title: sh_MI_D1_2_51

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 384”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2784 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.292a)


title: NFF1_19

status: Public on Jul 25 2022

type: genomic

city: Grand Rapids

first: Peter

middle: W

last: Laird

state: MI

address: 333 Bostwick Ave NE

department: Epigenetics

country: USA

institute: Van Andel Institute

hyb_protocol: Bisulfite-converted DNA was amplified, fragmented, and hybridized to Illumina Infinium Human Methylation850k Beadchip using standard Illumina protocol.

scan_protocol: Arrays were imaged using BeadArray Reader (Illumina HiSeq2000) using standard recommended Illumina scanner setting

data_processing: Raw IDATs were processed with R (v4.1.1) package ‘SeSAMe’ with noob background correction, non-linear dye bias correction, and non-detection masking. P-value threshold was set at 0.1. Probe masking was performed using the standard mask list in SeSAMe, including probes that overlap with SNPs and repeat elements.

label: Cy3 and Cy5

molecule: genomic DNA

organism: Homo sapiens

source_name: Foreskin

label_protocol: Cy3 and Cy5

extract_protocol: Frozen cell pellets were thawed and lysed using QIAshredder spin columns (Qiagen).  Genomic DNA was extracted from each sample using the AllPrep DNA/RNA Mini Kit (Qiagen), then stored at -80°C before analysis.

tag: subexperiment

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

tag: subculture

value: b

tag: growth protocol

value: Fibroblasts were maintained in Ham’s F12/DMEM 1:1 media supplemented with 10% v/v fetal bovine serum. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: MI_D1_RZ_2_20

type: SRA

city: Tokyo

first: Shintaro

last: Yamada

address: 7-3-1, Hongo, Bunkyo-ku

department: Department of Cardiovascular Medicine

country: Japan

institute: The University of Tokyo

description:
Smart-seq2
ReadCounts.xlsx

library_source: transcriptomic

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.027 with the parameters “--cut_tail --cut_tail_window_size 1 --cut_tail_mean_quality 30 --length_required 30”
The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.2b)
The reads aligned to exon were counted using featureCounts
For nonCM RNA-seq, raw data was processed using Cell Ranger (version 4.0.0)
For spatial transcriptome, raw data was processed using Space Ranger (version 1.2.1)
Genome_build: For single-cardiomyocyte RNA-seq, the reference transcript data and the gene annotation file for mouse was downloaded from GENCODE (release 23, https://www.gencodegenes.org/)
Genome_build: For nonCM RNA-seq, the reference transcript data and the gene annotation file for mouse was downloaded from  https://cf.10xgenomics.com/supp/cell-exp/refdata-gex-mm10-2020-A.tar.gz.
Genome_build: For spatial transcrptome, the reference transcript data and the gene annotation file for mouse was downloaded from https://cf.10xgenomics.com/supp/spatial-exp/refdata-gex-mm10-2020-A.tar.gz.
Supplementary_files_format_and_content: ReadCounts file generated using featureCounts.
Supplementary_files_format_and_content: Each barcode.tsv.gz, features.tsv.gz and matrix.mtx.gz files genereted using Cell Ranger or Space Ranger.

molecule: total RNA

organism: Mus musculus

source_name: Cardiomyocyte

growth_protocol: All the animal experiments were approved by the University of Tokyo Ethics Committee for Animal Experiments and strictly adhered to the guidelines for animal experiments of the University of Tokyo (Approved Number P14-106). All of wild type C57BL/6 male mice were purchased from CLEA JAPAN. All mouse were housed in separate cages at a maximum of 6 mice per cage in a specific-pathogen–free, temperature-controlled vivarium under a 12-h light/dark cycle with ad libitum access to food and water.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described7. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2500 (Illumina) in rapid mode

treatment_protocol: Myocardial infarction (MI) was induced as previously described25. Mice were anaesthetized by inhalation of 2% isoflurane. MI was induced by ligation of the left anterior descending artery. Mice that failed to develop MI or died within one week after the operation were excluded from the analysis.

tag: strain

tag: operation

value: MI day1

tag: region

value: remote

tag: tissue

value: single-cardiomyocyte

tag: name in supplementary file


title: MI_D1_RZ_2_18

description:
Smart-seq2
ReadCounts.xlsx


title: MI_D1_RZ_2_17

description:
Smart-seq2
ReadCounts.xlsx


title: MI_D1_RZ_2_16

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_21


title: MI_D1_RZ_2_14

description:
Smart-seq2
ReadCounts.xlsx


title: sh_MI_D1_2_56

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 389”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2789 (Illumina) in rapid mode

treatment_protocol: Myocardial infarction (MI) was induced as previously described. Mice were anaesthetized by inhalation of 2% isoflurane. MI was induced by ligation of the left anterior descending artery. Mice that failed to develop MI or died within one week after the operation were excluded from the analysis.

value: MI day1_shCsrp3

value: ishchemic + border

tag: genome build

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.297a)


title: MI_D1_RZ_2_15

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_25


title: NFF1_15


title: MI_D1_RZ_2_11

description:
Smart-seq2
ReadCounts.xlsx


title: MI_D1_RZ_2_12

description:
Smart-seq2
ReadCounts.xlsx


title: sh_MI_D1_2_49

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 381”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2781 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.289a)


title: sh_MI_D1_2_47

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 379”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2779 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.287a)


title: MI_D1_RZ_2_10

description:
Smart-seq2
ReadCounts.xlsx


title: MI_D1_RZ_2_9

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_16


title: MI_D1_RZ_2_8

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_17


title: NFF1_13


title: sh_MI_D1_2_45

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 377”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2777 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.285a)


title: sh_MI_D1_2_48

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 380”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2780 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.288a)


title: MI_D1_RZ_2_6

description:
Smart-seq2
ReadCounts.xlsx


title: sh_MI_D1_2_44

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 376”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2776 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.284a)


title: MI_D1_RZ_2_4

description:
Smart-seq2
ReadCounts.xlsx


title: sh_MI_D1_2_5

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 382”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2782 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.290a)


title: MI_D1_RZ_2_3

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_12

value: Baseline profiling

value: 0 years

value: male


title: MI_D1_RZ_2_5

description:
Smart-seq2
ReadCounts.xlsx


title: MI_D1_RZ_2_2

description:
Smart-seq2
ReadCounts.xlsx


title: MI_D1_RZ_2_13

description:
Smart-seq2
ReadCounts.xlsx


title: sh_MI_D1_2_42

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 374”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2774 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.282a)


title: MI_D1_RZ_1_66

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_11

value: Ambient

value: Full

value: b

value: Fibroblasts were maintained in Ham’s F12/DMEM 1:1 media supplemented with 10% v/v fetal bovine serum. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: NFF1_10


title: MI_D1_RZ_1_65

description:
Smart-seq2
ReadCounts.xlsx

value: MI day1

value: remote

value: single-cardiomyocyte


title: sh_MI_D1_2_41

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 373”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2773 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: ishchemic + border

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.281a)


title: MI_D1_RZ_2_1

description:
Smart-seq2
ReadCounts.xlsx


title: sh_MI_D1_2_43

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 375”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2775 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.283a)


title: NFF1_9


title: NFF1_8


title: sh_MI_D1_2_40

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 372”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2772 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.280a)


title: MI_D1_RZ_1_64

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_4


title: RimRun SF 96

city: Lexington

first: Shavahn

middle: C

last: Loux

state: KY

address: 1400 Nicholasville

department: Veterinary Science

institute: University of Kentucky

data_processing:
mapping (STAR 2.7.2a - STAR --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonicalU  nannotated --alignEndsType Local --chimOutType WithinBAM --twopassMode Basic --t  wopass1readsN -1 --quantMode GeneCounts --outSAMunmapped Within --outSAMtype BAM   SortedByCoordinate )
quantification (STAR-2.7.2a - GeneCounts - annotation Ensembl v 104)
Normalization (GeTMM via EdgeR; rpk.norm <- DGEList(counts=rpk,group=group)  rpk.norm <- calcNormFactors(rpk.norm)  norm.counts.rpk_edger <- cpm(rpk.norm))
DEG (EdgeR; glmQLFTest)
Genome_build: EquCab3.0
Supplementary_files_format_and_content: GeTMM gene expression values - csv file

organism: Equus caballus

source_name: Bone marrow mononuclear cells

growth_protocol: Samples identified as SF(n,y,z, …i) were cultured in autologous normal equine synovial fluid for up to 10 days

extract_protocol:
Cultured cells were recovered in 10 mM EDTA, centrifuged (12,000g; 10 min; 4°C) and the cell pellet placed in 2 mL guanidinium chloride-phenol (Trizol®, Life Technologies). RNA was purified with on-column DNase digest (DirectZolTM RNA microprep kit, , Zymo Research, I), quantified (Qubit® 3.0 Fluorometer), and stored at -80°C. RNA quality was assessed using a Bioanalyzer 2100, Agilent Technologies, Santa Clara, CA).
cDNA libraries were prepared using TruSeq DNA Library Preparation kits (Illumina, Inc., San Diego, CA).
cDNA libraries were sequenced (NovaSeq 6000 S4, Illumina) to generate an average of 34.5 (range, 24-54) million stranded paired-end reads (2 x 150 nt) per sample

treatment_protocol: Samples identified as ISF(n,y,z, …i) were cultured in autologous inflamed equine synovial fluid for up to 10 days

tag: cell type

value: Bone marrow mononuclear cells

tag: breed

value: Thoroughbred

tag: age

value: Adult

tag: name

value: RimRun

tag: Sex

value: Female

tag: type

value: Mare

tag: treatment

tag: time

value: 96 hours


title: RimRun SF 48

value: cultured in autologous normal equine synovial fluid

value: 48 hours


title: NFF1_18

tag: subexperiment

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

value: b

value: Fibroblasts were maintained in Ham’s F12/DMEM 1:1 media supplemented with 10% v/v fetal bovine serum. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: sh_MI_D1_2_4

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 371”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2771 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: ishchemic + border

value: single-cardiomyocyte

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.279a)


title: sh_MI_D1_2_46

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 378”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2778 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.286a)


title: sh_MI_D1_2_39

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 370”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2770 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.278a)


title: MI_D1_RZ_1_62

description:
Smart-seq2
ReadCounts.xlsx

value: MI day1

value: remote


title: RimRun SF 10

value: Bone marrow mononuclear cells

value: Thoroughbred

value: Adult

value: RimRun

value: Female

value: Mare

value: cultured in autologous normal equine synovial fluid

value: 10 days


title: sh_MI_D1_2_38

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 369”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2769 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.277a)


title: MI_D1_RZ_1_61

description:
Smart-seq2
ReadCounts.xlsx

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.027 with the parameters “--cut_tail --cut_tail_window_size 1 --cut_tail_mean_quality 30 --length_required 30”
The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.2b)
The reads aligned to exon were counted using featureCounts
For nonCM RNA-seq, raw data was processed using Cell Ranger (version 4.0.0)
For spatial transcriptome, raw data was processed using Space Ranger (version 1.2.1)
Genome_build: For single-cardiomyocyte RNA-seq, the reference transcript data and the gene annotation file for mouse was downloaded from GENCODE (release 23, https://www.gencodegenes.org/)
Genome_build: For nonCM RNA-seq, the reference transcript data and the gene annotation file for mouse was downloaded from  https://cf.10xgenomics.com/supp/cell-exp/refdata-gex-mm10-2020-A.tar.gz.
Genome_build: For spatial transcrptome, the reference transcript data and the gene annotation file for mouse was downloaded from https://cf.10xgenomics.com/supp/spatial-exp/refdata-gex-mm10-2020-A.tar.gz.
Supplementary_files_format_and_content: ReadCounts file generated using featureCounts.
Supplementary_files_format_and_content: Each barcode.tsv.gz, features.tsv.gz and matrix.mtx.gz files genereted using Cell Ranger or Space Ranger.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described7. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2500 (Illumina) in rapid mode


title: NFF1_5

data_processing: Raw IDATs were processed with R (v4.1.1) package ‘SeSAMe’ with noob background correction, non-linear dye bias correction, and non-detection masking. P-value threshold was set at 0.1. Probe masking was performed using the standard mask list in SeSAMe, including probes that overlap with SNPs and repeat elements.

extract_protocol: Frozen cell pellets were thawed and lysed using QIAshredder spin columns (Qiagen).  Genomic DNA was extracted from each sample using the AllPrep DNA/RNA Mini Kit (Qiagen), then stored at -80°C before analysis.

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

tag: subculture

value: b

value: Fibroblasts were maintained in Ham’s F12/DMEM 1:1 media supplemented with 10% v/v fetal bovine serum. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: RimRun SF 6

value: 6 days


title: sh_MI_D1_2_37

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 368”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2768 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: ishchemic + border

value: single-cardiomyocyte

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.276a)


title: MI_D1_RZ_1_63

description:
Smart-seq2
ReadCounts.xlsx

value: MI day1

value: remote


title: RimRun ISF 96

value: Bone marrow mononuclear cells

value: Thoroughbred

value: Adult

value: RimRun

value: Female

value: Mare

value: cultured in inflammed equine synovial fluid

value: 96 hours


title: MI_D1_RZ_1_60

description:
Smart-seq2
ReadCounts.xlsx


title: NFF1_2

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

value: b

tag: growth protocol


title: sh_MI_D1_2_36

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 367”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2767 (Illumina) in rapid mode

tag: strain

value: MI day1_shCsrp3

value: ishchemic + border

value: single-cardiomyocyte

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.275a)


title: MI_D1_RZ_1_59

description:
Smart-seq2
ReadCounts.xlsx

tag: operation

value: MI day1

value: remote


title: RimRun ISF 48

value: Bone marrow mononuclear cells

value: Thoroughbred

value: Adult

value: RimRun

value: 48 hours


title: MI_D1_RZ_1_58

description:
Smart-seq2
ReadCounts.xlsx

tag: region


title: RimRun ISF 10

value: Female

value: Mare

value: cultured in inflammed equine synovial fluid

value: 10 days


title: MI_D1_RZ_1_57

description:
Smart-seq2
ReadCounts.xlsx

tag: tissue


title: RimRun ISF 6

value: 6 days


title: MI_D1_RZ_1_56

description:
Smart-seq2
ReadCounts.xlsx

tag: name in supplementary file


title: RimRun BMNC 0hr

value: 0 h - baseline


title: NFF1_3

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

value: b

value: Fibroblasts were maintained in Ham’s F12/DMEM 1:1 media supplemented with 10% v/v fetal bovine serum. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: sh_MI_D1_2_33

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 364”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2764 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: ishchemic + border

value: single-cardiomyocyte

tag: genome build

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.272a)


title: MI_D1_RZ_2_7

description:
Smart-seq2
ReadCounts.xlsx

value: MI day1

value: remote


title: sh_MI_D1_2_32

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 363”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2763 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.271a)


title: MI_D1_RZ_1_55

description:
Smart-seq2
ReadCounts.xlsx


title: Ollie SF 96

tag: cell type

value: Bone marrow mononuclear cells

value: Thoroughbred

value: Adult

value: Ollie

value: gelding

value: cultured in autologous normal equine synovial fluid

value: 96 hours


title: NFF1_7

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full


title: NFK1_19

value: a

value: Keratinocytes were maintained in serum-free human epidermal keratinocyte media on collagen IV-coated dishes. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: MI_D1_RZ_1_54

description:
Smart-seq2
ReadCounts.xlsx

value: MI day1

value: single-cardiomyocyte


title: Ollie SF 48

tag: breed

value: 48 hours


title: sh_MI_D1_2_31

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 362”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2762 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: ishchemic + border

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.270a)


title: sh_MI_D1_2_35

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 366”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2766 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.274a)


title: MI_D1_RZ_1_53

description:
Smart-seq2
ReadCounts.xlsx

value: remote


title: Ollie SF 10

value: Bone marrow mononuclear cells

value: Thoroughbred

tag: age

value: Adult

value: Ollie

value: gelding

value: cultured in autologous normal equine synovial fluid

value: 10 days


title: NFF1_1

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

value: Fibroblasts were maintained in Ham’s F12/DMEM 1:1 media supplemented with 10% v/v fetal bovine serum. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: sh_MI_D1_2_3

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 360”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2760 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: ishchemic + border

value: single-cardiomyocyte

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.268a)


title: NFK1_18

value: c

value: Keratinocytes were maintained in serum-free human epidermal keratinocyte media on collagen IV-coated dishes. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: Ollie SF 6

value: Bone marrow mononuclear cells

value: Thoroughbred

value: Adult

tag: name

value: 6 days


title: NFK1_16

value: a


title: MI_D1_RZ_1_51

description:
Smart-seq2
ReadCounts.xlsx

value: MI day1

value: remote


title: Ollie ISF 96

value: Ollie

tag: Sex

value: gelding

value: cultured in inflammed equine synovial fluid

value: 96 hours


title: sh_MI_D1_2_29

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 359”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2759 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.267a)


title: MI_D1_RZ_1_50

description:
Smart-seq2
ReadCounts.xlsx

value: single-cardiomyocyte


title: NFF1_14

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

value: b

value: Fibroblasts were maintained in Ham’s F12/DMEM 1:1 media supplemented with 10% v/v fetal bovine serum. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: MI_D1_RZ_1_52

description:
Smart-seq2
ReadCounts.xlsx


title: NFK1_17

value: Keratinocytes were maintained in serum-free human epidermal keratinocyte media on collagen IV-coated dishes. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: NFF1_6


title: Ollie ISF 48

value: Bone marrow mononuclear cells

value: Thoroughbred

value: Adult

value: 48 hours


title: sh_MI_D1_2_28

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 358”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2758 (Illumina) in rapid mode

value: ishchemic + border

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.266a)


title: sh_MI_D1_2_30

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 361”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2761 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.269a)


title: NFK1_11

value: a


title: MI_D1_RZ_1_49

description:
Smart-seq2
ReadCounts.xlsx

value: MI day1

value: remote

value: single-cardiomyocyte


title: sh_MI_D1_2_34

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 365”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2765 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.273a)


title: Ollie ISF 10

data_processing:
mapping (STAR 2.7.2a - STAR --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonicalU  nannotated --alignEndsType Local --chimOutType WithinBAM --twopassMode Basic --t  wopass1readsN -1 --quantMode GeneCounts --outSAMunmapped Within --outSAMtype BAM   SortedByCoordinate )
quantification (STAR-2.7.2a - GeneCounts - annotation Ensembl v 104)
Normalization (GeTMM via EdgeR; rpk.norm <- DGEList(counts=rpk,group=group)  rpk.norm <- calcNormFactors(rpk.norm)  norm.counts.rpk_edger <- cpm(rpk.norm))
DEG (EdgeR; glmQLFTest)
Genome_build: EquCab3.0
Supplementary_files_format_and_content: GeTMM gene expression values - csv file

extract_protocol:
Cultured cells were recovered in 10 mM EDTA, centrifuged (12,000g; 10 min; 4°C) and the cell pellet placed in 2 mL guanidinium chloride-phenol (Trizol®, Life Technologies). RNA was purified with on-column DNase digest (DirectZolTM RNA microprep kit, , Zymo Research, I), quantified (Qubit® 3.0 Fluorometer), and stored at -80°C. RNA quality was assessed using a Bioanalyzer 2100, Agilent Technologies, Santa Clara, CA).
cDNA libraries were prepared using TruSeq DNA Library Preparation kits (Illumina, Inc., San Diego, CA).
cDNA libraries were sequenced (NovaSeq 6000 S4, Illumina) to generate an average of 34.5 (range, 24-54) million stranded paired-end reads (2 x 150 nt) per sample

value: Ollie

value: gelding

value: cultured in inflammed equine synovial fluid

value: 10 days


title: sh_MI_D1_2_27

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 357”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2757 (Illumina) in rapid mode

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.265a)


title: MI_D1_RZ_1_48

description:
Smart-seq2
ReadCounts.xlsx


title: NFK1_13

value: Baseline profiling

value: 0 years

value: male

value: Ambient

value: Full

value: b

value: Keratinocytes were maintained in serum-free human epidermal keratinocyte media on collagen IV-coated dishes. Triplicate cultures derived from the same parent plate or vial obtained from Coriell were maintained in parallel through replicative senescence, which was defined in this study as drastically slowed growth (inability to reach near-confluence at 14 days after previous passage) or viable fraction of cells falling below 60%. Passaging occurred as cells became approximately 90% confluent.


title: sh_MI_D1_2_26

data_processing:
Raw sequencing reads from single-cardiomyocyte RNA sequencing libraries were trimmed to remove adapter sequences and low-quality bases using fastp-0.20.038 with the parameters “--cut_tail --cut_tail_window_size 10 --cut_tail_mean_quality 30 --length_required 356”.
ReadCounts file generated using featureCounts.

extract_protocol:
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricle as previously described. Enzymatic dissociation using Langendorff perfusion was performed with 35 mL enzyme solution (type 2 collagenase 1 mg/mL [Worthington], protease type XIV 0.05 mg/mL [Sigma-Aldrich], NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM; pH 7.4) pre-heated to 37 °C, at a rate of 3 mL/min. To neutralize enzymatic activity, fetal bovine serum (FBS) was added to the solution to a final concentration of 0.2% (v/v). After perfusion of digestion buffer, left ventricle was cut according to each sectional region (IR, RR) and cardiomyocytes were subsequently isolated. Cell suspensions were filtered through a 100-μm nylon mesh cell strainer and centrifuged at 100 g for 2 min, after which the supernatant was discarded. To prevent hypercontraction, the cardiomyocytes were resuspended in medium (NaCl 130 mM, KCl 5.4 mM, MgCl2 0.5 mM, NaH2PO4 0.33 mM, D-glucose 22 mM, HEPES 25 mM, FBS 0.2%; pH 7.4) containing a low concentration of calcium (0.1 mM). Rod-shaped live cardiomyocytes (viability of cardiomyocytes was ≥80% for all time points) were collected immediately with a 0.2–2-µL pipette (sample volume, 0.5 µL) and incubated in lysis buffer.
Single-cardiomyocyte cDNA libraries were generated using the Smart-seq2 protocol26 and the efﬁciency of reverse transcription was assessed by examining the cycle threshold (Ct) values of control genes (Tnnt2) from quantitative real-time polymerase chain reaction (qRT-PCR) using a CFX96 Real-Time PCR Detection System (Bio-Rad), and the distribution of cDNA fragment lengths was assessed using LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for qRT-PCR: Tnnt2 mRNA forward: TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse: TGCAGGTCGA ACTTCTCAGC. A Ct value of 25 was set as the threshold. The sequencing libraries were subjected to paired-end 51-bp RNA sequencing on a HiSeq 2756 (Illumina) in rapid mode

value: MI day1_shCsrp3

value: ishchemic + border

value: The clean reads were aligned to the mouse genome (mm10) using STAR (version 2.7.264a)


title: Ollie ISF 6

value: Bone marrow mononuclear cells

value: Thoroughbred

value: Adult

value: 6 days


title: NFK1_12

value: c


title: Ollie BMNC 0hr

value: Ollie

value: gelding

value: 0 h - baseline