+ Site Statistics
+ Search Articles
+ Subscribe to Site Feeds
EurekaMag Most Shared ContentMost Shared
EurekaMag PDF Full Text ContentPDF Full Text
+ PDF Full Text
Request PDF Full TextRequest PDF Full Text
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter
Follow on Google+Follow on Google+
Follow on LinkedInFollow on LinkedIn

+ Translate

Regulation of gene expression in adult rat hepatocytes cultured on a basement membrane matrix

Journal of Cellular Physiology 134(3): 309-323
Regulation of gene expression in adult rat hepatocytes cultured on a basement membrane matrix
Freshly isolated adult rat hepatocytes, when cultured on type I collagen (commercially available as Vitrogen), assume a polygonal shape, form a stable monolayer within 24 hours, but lose the capacity to express some liver-specific functions over time in culture. We incubated hepatocytes in a serum-free medium on a reconstituted basement membrane gel, "matrigel" (prepared from an extract of extracellular matrix of the murine Engelbreth-Holm-Swarm sarcoma), and observed that the cells adhered firmly, remained rounded as single cells or clusters, and maintained liver-specific gene expression for more than 1 week in vitro. Hepatocytes on matrigel secreted substantially higher amounts of albumin, transferrin, haptoglobin, and hemopexin, contained higher amounts of glutathione peroxidase, and, as judged by Northern blot analyses of extracted cellular RNA, expressed increased amounts of mRNA for the liver-specific protein albumin (as compared with cells on vitrogen). In cultures treated with phenobarbital, cytochrome P-450b, and cytochrome P-450e, mRNAs and proteins were barely detectable in cells on Vitrogen but were induced to levels similar to those in the liver in vivo in matrigel cultures. Likewise, the use of matrigel greatly enhanced the induction of mRNA and protein for P-450c by 3-methylcholanthrene and for P-450p by steroidal and nonsteroidal inducers. However, neither substratum permitted induction of P-450d by 3-methylcholanthrene, suggesting that the effects of matrigel are selective even for expression in liver of members of the superfamily of cytochrome P-450 genes. Within 5 days in cultures on Vitrogen, hepatocytes expressed detectable amounts of fetal liver aldolase activity and also mRNA for vimentin and type I collagen, each considered a phenotypic change reflecting hepatocyte "dedifferentiation". None of these was present in cells on matrigel. Responsiveness to mitogenic stimuli, as judged by incorporation of 3H-thymidine into DNA, was also decreased in hepatocytes cultured on matrigel. Finally, there was a remarkable increase in the levels of mRNAs for the cytoskeleton proteins actin and tubulin in hepatocytes on both matrices during the first 2 days in culture. However, the continuously flattening hepatocytes on Vitrogen maintained substantially higher levels of cytoskeleton mRNA over time in culture than did the rounded cells on matrigel. We conclude that hepatocytes cultured on matrigel, as opposed to the standard collagen, exhibit remarkably enhanced expression of many liver-specific functions.

Accession: 006288002

PMID: 3350857

DOI: 10.1002/jcp.1041340302

Download PDF Full Text: Regulation of gene expression in adult rat hepatocytes cultured on a basement membrane matrix

Related references

Regulation of cytochrome P-450b/e and P-450p gene expression by growth hormone in adult rat hepatocytes cultured on a reconstituted basement membrane. Journal of Biological Chemistry 265(2): 1188-1192, 1990

Transcriptional regulation of the albumin gene in cultured rat hepatocytes. Role of basement-membrane matrix. Molecular Biology & Medicine 7(2): 187-197, 1990

Growth hormone regulation of sex specific and phenobarbital inducible cytochromes p 450 in adult rat hepatocytes cultured on the basement membrane matrix matrigel. Hepatology 8(5): 1227, 1988

Regulation of gene expression in adult rat hepatocytes cultured on extracellular matrix. Hepatology: 1188, 1986

Regulation of milk protein and basement membrane gene expression: the influence of the extracellular matrix. Journal of Dairy Science 71(10): 2830-2842, 1988

Regulation of glutathione S-transferase gene expression by phenobarbital in cultured adult rat hepatocytes. Febs Letters 284(1): 103-108, 1991

The role of fetal and adult hepatocyte extracellular matrix in the regulation of tissue-specific gene expression in fetal and adult hepatocytes. European Journal of Cell Biology 81(1): 43-50, 2002

Influence of isolation procedure, extracellular matrix and dexamethasone on the regulation of membrane transporters gene expression in rat hepatocytes. Biochemical Pharmacology 64(11): 1637-1650, 1 December, 2002

Transient expression and regulation of inducible nitric oxide synthase gene in primary cultured adult murine hepatocytes. Japanese Journal of Pharmacology 75(SUPPL 1): 84P, 1997

Differential expression of basement membrane genes in fetal and adult hepatocytes. Journal of Cell Biology 111(5 PART 2): 399A, 1990