Formaldehyde induces toxic effects and regulates the expression of damage response genes in BM-MSCs.
In this study, we assessed the toxic effects of formaldehyde (FA) on mouse bone marrow mesenchymal stem cells (BM-MSCs). Cytotoxicity was measured by using MTT assay. DNA strand breakage was detected by standard alkaline comet assay and comet assay modified with proteinase K (PK). DNA-protein crosslinks (DPCs) were detected by KCl-SDS precipitation assay. We found that FA at a concentration from 75 to 200 μM inhibited cell survival and induced DPCs over 125 μM. The PK-modified comet assay showed that FA-induced DNA strand breakage was increased in a dose-dependent manner from 75 to 200 μM. On the other hand, standard alkaline comet assay showed that DNA strand breakage was decreased with FA concentration over 125 μM. We confirmed by using Pearson correlation that there was a negative linear correlation between DPCs and survival rate (r = -0.987, P < 0.01) and positive linear relationships between DPCs and (i) sister chromatid exchange and (ii) micronucleus (r = 0.995, P < 0.01; r = 0.968, P < 0.01). DNA damage RT(2) profiler polymerase chain reaction array was used to investigate the changes in the expression of damage response genes. Xpa and Xpc of the nucleotide excision repair pathway and Brca2, Rad51, and Xrcc2 of the homologous recombination pathway were all up-regulated in both 75 and 125 μM FA. However, the same genes were down-regulated with 175 μM FA. The expressions of Chek1 and Hus1, which are involved in cell cycle regulation, were altered in the same manner with 75, 125, and 175 μM FA. These results indicated that Xpa, Xpc, Brca2, Rad51, Xrcc2, Chek1, and Hus1 were essential for the BM-MSCs to counteract the effects of FA.
College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China.
Yali
Y
She
Yi
Y
Li
Yongqi
Y
Liu
Guli
G
Asai
Shaobo
S
Sun
Jianxin
J
He
Zheng
Z
Pan
Yan
Y
Cui
D000818:Animals; D024682:BRCA2 Protein / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D000071877:Checkpoint Kinase 1; D020552:Comet Assay; D053960:DNA Breaks / Q000187:drug effects; D004249:DNA Damage / Q000235:genetics*; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D005557:Formaldehyde / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D059630:Mesenchymal Stem Cells / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D048629:Micronuclei, Chromosome-Defective / Q000187:drug effects; D015162:Micronucleus Tests; D011494:Protein Kinases / Q000235:genetics; D051135:Rad51 Recombinase / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012854:Sister Chromatid Exchange / Q000187:drug effects; D051760:Xeroderma Pigmentosum Group A Protein / Q000235:genetics
D024682:BRCA2 Protein; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C549605:HUS1 protein, human; C497314:XPA protein, human; C113187:XRCC2 protein, human; D051760:Xeroderma Pigmentosum Group A Protein; C086984:XPC protein, human; D005557:Formaldehyde; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D051135:Rad51 Recombinase
DNA–protein crosslinks; PCR array; comet assay; micronuclei; sister chromatid exchange