title: Human primary cultured hepatic stellate cells can be cryopreserved.
abstract: We compared the morphological and functional characteristics of cultured unfrozen hepatic stellate cells (HSCs) and cryopreserved HSCs obtained from human livers. We used liver tissues obtained by surgical resection from patients with metastatic liver cancer or with hepatocellular carcinoma. HSCs were isolated and allowed to spread in culture. Comparison of morphological and functional features between the unfrozen HSCs and cryopreserved HSCs was performed at each passage using the following techniques: light microscopy, immunohistochemistry, cell growth curve, metallothionein (MTT) assay, and PI staining, Western blot, real-time polymerase chain reaction (PCR), and gene expression analysis using microarrays. The purity of HSCs was more than 90% in all passages. alpha-Smooth muscle actin (SMA-)positive HSCs gradually increased in successive passages, and the positive cell rate and rate of increase in cell number were similar in both groups. Expression of platelet-derived growth factor (PDGF) receptor, transforming growth factor (TGF)-beta receptor, and alpha-SMA mRNAs and protein was similar during each passage in the two groups. Gene expression was nearly identical at each passage in unfrozen and frozen/thawed samples obtained from the same patient. In conclusion, an adequate protocol for the cryopreservation of human primary cultured HSCs could be established.
authors.affiliation: Research Center for Innovative Cancer Therapy, Kurume University, 67 Asahi-machi, Kurume, 830-0011, Japan.
authors.forename: Anna
authors.initials: A
authors.lastname: Nakamura
authors.forename: Takato
authors.initials: T
authors.lastname: Ueno
authors.forename: Yumihiko
authors.initials: Y
authors.lastname: Yagi
authors.forename: Koji
authors.initials: K
authors.lastname: Okuda
authors.forename: Toshiro
authors.initials: T
authors.lastname: Ogata
authors.forename: Toru
authors.initials: T
authors.lastname: Nakamura
authors.forename: Takuji
authors.initials: T
authors.lastname: Torimura
authors.forename: Hideki
authors.initials: H
authors.lastname: Iwamoto
authors.forename: Sivakumar
authors.initials: S
authors.lastname: Ramadoss
authors.forename: Michio
authors.initials: M
authors.lastname: Sata
authors.forename: Victor
authors.initials: V
authors.lastname: Tsutsumi
authors.forename: Kaori
authors.initials: K
authors.lastname: Yasuda
authors.forename: Yumi
authors.initials: Y
authors.lastname: Tomiyasu
authors.forename: Kenichi
authors.initials: K
authors.lastname: Obayashi
authors.forename: Kosuke
authors.initials: K
authors.lastname: Tashiro
authors.forename: Satoru
authors.initials: S
authors.lastname: Kuhara
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D015153:Blotting, Western; D002478:Cells, Cultured; D016000:Cluster Analysis; D015925:Cryopreservation / Q000379:methods*; D005260:Female; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation; D055166:Hepatic Stellate Cells / Q000166:cytology* / Q000648:ultrastructure; D006801:Humans; D008099:Liver / Q000166:cytology / Q000378:metabolism; D008297:Male; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics
chemical_list: D012333:RNA, Messenger