Immortalized differentiated hepatocyte lines derived from transgenic mice harboring SV40 T-antigen genes*1

Paul, Dieter

doi:10.1016/0014-4827(88)90199-1

Cited by 95 publications

(49 citation statements)

References 18 publications

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“…The connexins are a family of gap-junction proteins which are differentially expressed in tissue-specific patterns. Cx 26 and Cx 32 are the only types known to exist in hepatocytes, whereas Cx 43 is found in bile ductules and dedifferentiated liver cell lines (6,26,27,33). At 4 months in culture, both AML lines synthesized the 2.4-kb Cx 26 and 1.8-kb Cx 32 transcripts at levels comparable to that of 1-day cultured hepatocytes (Fig.…”

Section: Resultsmentioning

confidence: 87%

“…More recently, well-differentiated lines have been derived by introducing viral oncogenes into hepatocytes. While useful, these lines have the drawback of either being fully transformed or undergoing transformation with repeated passaging (3)(4)(5)(6).Transforming growth factor a (TGFa) is a polypeptide that regulates normal growth in epithelial tissues, and its overproduction in cells possessing epidermal growth factor receptors (EGFRs) is often correlated with malignant transformation (7-10). TGFa is expressed in the liver during embryogenesis…”

mentioning

confidence: 99%

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confidence: 99%

“…Northern blot analysis was performed on total RNA extracted from cultured cells at 70% confluence (12,21). Filters were hybridized at 50°C [for the connexin (Cx) 32 probe] or 42°C (all other probes) with the following 32P-labeled probes: a 240-bp Pst I cDNA fragment recognizing human and mouse TGFa mRNA; a 2.4-kb human EGFR cDNA insert from pE7 (22); the mouse albumin cDNA insert from pmalb2 (23); a 0.5-kb Bgl I-BamHI a-fetoprotein (AFP) cDNA fragment from pBAF700 (24); the mouse a1-antitrypsin (AlAT) insert in pLivS3 and transferrin insert in pLivS6 (from H. Isom, Pennsylvania State University College of Medicine) (25); the rat Cx 26 and 32 cDNAs in pSP42T and pGEM-3, respectively (from D. Paul, Harvard Medical School) (6,26); the rat Cx 43 insert in pBS (from E. Beyer, Washington University School of Medicine) (27); and a 1.2-kb rat glyceraldehyde-3-phosphate dehydrogenase cDNA from pRLC-GAP (from J. Tso, X.-H. Sun, and R. Wu, Cornell University). For multiple probings, filters were stripped in 1% (vol/vol) glycerol at 80°C for 3 min before rehybridization.…”

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confidence: 99%

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Establishment and characterization of differentiated, nontransformed hepatocyte cell lines derived from mice transgenic for transforming growth factor alpha.

Merlino

Fausto

1994

Proc. Natl. Acad. Sci. U.S.A.

268

202

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Hepatocytes are extensively used in studies of gene regulation but cannot be maintained in long-term culture as replicating, differentiated cells while remaining nontumorigenic. We have derived two hepatocyte lines from livers of transgenic mice overexpressing transforming growth factor a, a potent hepatocyte mitogen, which overcome these limitations. The transgenic hepatocytes were maintained for .2 months in serum-supplemented primary culture and gave rise to cell lines, of which two (AML12 and AML14) have been cultured for >1.5 years (>80 passages). Both lines have typical hepatocyte features such as peroxisomes and bile canalicular-like structures, do not grow in soft agar, and are nontumorigenic in nude mice. Like normal hepatocytes, AML cells express high levels of mRNA for serum (albumin, al-antitrypsin, and transferrin) and gap junction (connexins 26 and 32) proteins, secrete albumin, and contain solely isozyme 5 of lactate dehydrogenase. After extensive passaging, AML12 cells continue to strongly coexpress hepatocyte connexin mRNAs but do not display nonparenchymal cell markers. Although mRNA levels for some serum proteins progressively fall, high expression in late AML12 cultures may be regained by passage in serum-free medium. The AML14 line loses expression of both differentiated markers and transgene mRNA with extended passaging, and hepatocytic traits are only partially restored by passage in serum-free medium. These differentiated, nontumorigenic cell lines should serve as models in which to study hepatocyte growth and differentiation.Mammalian hepatocytes have long been used to investigate mechanisms of cell growth, differentiation, and tumorigenesis and as vectors for gene therapy. These studies have been hampered because dissociated hepatocytes rapidly lose their cell-specific functions, are short-lived, and have a limited ability to replicate in culture. Longer-term differentiated cultures can be maintained with specialized media and substrata or by coculture with nonparenchymal cells, but under these conditions the cells remain nonproliferative (1-3).Hepatocyte lines which express some tissue-specific markers have been propagated in vitro from hepatocellular tumors. More recently, well-differentiated lines have been derived by introducing viral oncogenes into hepatocytes. While useful, these lines have the drawback of either being fully transformed or undergoing transformation with repeated passaging (3)(4)(5)(6).Transforming growth factor a (TGFa) is a polypeptide that regulates normal growth in epithelial tissues, and its overproduction in cells possessing epidermal growth factor receptors (EGFRs) is often correlated with malignant transformation (7-10). TGFa is expressed in the liver during embryogenesisThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. and transiently increases during liver regeneration after partial hepa...

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Section: Resultsmentioning

confidence: 87%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Establishment and characterization of differentiated, nontransformed hepatocyte cell lines derived from mice transgenic for transforming growth factor alpha.

Merlino

Fausto

1994

Proc. Natl. Acad. Sci. U.S.A.

268

202

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“…Previously, it has been shown that cells extracted from various tissues of TG rats were immortalized and retained temperature sensitivity (AutilloTouati et al 1986;Paul et al 1988;Yanai et al 1991;Kameoka et al 1995;Okuyama et al 1995). Cell lines established from TG rats typically proliferate strongly when grown at 33 C and differentiate when the temperature is shifted to 39 C (Yanai et al 1991;Jat and Sharp 1989).…”

Section: Introductionmentioning

confidence: 99%

Establishment of Periodontal Ligament Cell Lines from Temperature-Sensitive Simian Virus 40 Large T-antigen Transgenic Rats

Kubota¹,

Chiba²,

Obinata³

et al. 2004

Cytotechnology

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Orthodontic tooth movement is controlled by various cell types in the periodontal ligament (PDL). Mechanical stresses, such as orthodontic force, are thought to induce differentiation of the mesenchymal cells in the PDL into osteoblasts and cementoblasts. The details of the process of differentiation, however, are not known, in part because adequate in vitro systems for their study do not yet exist. The purpose of this study was to establish and characterize immortalized PDL cell lines derived from the PDL of transgenic rats harboring the temperaturesensitive simian virus 40 T-antigen gene (TG rats). The PDL was removed from the molar roots of TG rats and incubated in tissue culture. Outgrowth cells from the PDL explant were passaged and cloned, depending on the shape of the colonies formed. The cell lines thus established were analyzed by reverse transcription-polymerase chain reaction for expression of type-I collagen, osteopontin, fibronectin, alkaline phosphatase (bone type), bone sialoprotein, the receptor activator of NF-B ligand, and osteoprotegerin. In addition, the capacity for formation of mineralized nodules was assessed by incubating cells in calcification-promoting medium at 37 C. A total of 15 stable cell lines were successfully established and characterized. These cell lines were classified into six groups based on their pattern of gene expression at 33 C. Moreover, three of these clones were capable of forming calcified nodules. In conclusion, differential gene expression was demonstrated in 15 established PDL cell lines. Some cells had the potential to differentiate into cell types found in mineralized tissues, such as osteoblasts and cementoblasts, as well as cells expressing molecules that regulate osteoclast differentiation.Abbreviations: ALP -Alkaline phosphatase; BSP -Bone sialoprotein; COLL1 -Type-I collagen; DEX -Dexamethasone; FN -Fibronectin; OPG -Osteoprotegerin; OPN -Osteopontin; PDLPeriodontal ligament; RANKL -Receptor activator of NF-B ligand; TG rats -Transgenic rats harboring the temperature-sensitive simian virus 40 T-antigen gene

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Flow cytometric detection of the association between cell surface receptors and the cytoskeleton

et al. 1997

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Immortalized differentiated hepatocyte lines derived from transgenic mice harboring SV40 T-antigen genes*1

Cited by 95 publications

References 18 publications

Establishment and characterization of differentiated, nontransformed hepatocyte cell lines derived from mice transgenic for transforming growth factor alpha.

Establishment and characterization of differentiated, nontransformed hepatocyte cell lines derived from mice transgenic for transforming growth factor alpha.

Establishment of Periodontal Ligament Cell Lines from Temperature-Sensitive Simian Virus 40 Large T-antigen Transgenic Rats

Flow cytometric detection of the association between cell surface receptors and the cytoskeleton

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