Document Detail


Attenuated expression of 70-kDa heat shock protein in WI-38 human fibroblasts during aging in vitro.
MedLine Citation:
PMID:  10502396     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
We examined the effects of cellular aging on the expression of the heat shock-inducible HSP70 gene in WI-38 diploid human fibroblasts serially passaged in vitro. The senescence of the cells was established by evaluating population doubling level, cell density at confluency, and cell morphology along with the detection of senescence-associated beta-galactosidase activity (histochemically detectable at pH 6), a reliable marker of aging in low-density cultures. A marked decrease in the synthesis and accumulation of the inducible HSP70 protein was observed in serum-fed late passage cells exposed to a severe heat shock (30 min at 45 degrees C) in comparison to early passage cells. However, the degree of HSF-DNA binding, monitored by gel retardation assay was similar in both early and late passage cells. Similarly, Northern blotting analysis indicated that comparable amounts of inducible HSP70 mRNA were present in the total RNA fraction, in the total polyadenylated RNA fraction, or in the nuclear polyadenylated RNA fraction extracted from both early and late passage cells. In contrast, much less inducible HSP70 mRNA was detected in the total cytoplasmic RNA fraction or in the polyadenylated cytoplasmic RNA fraction of late passage cells. Thus age-related differences in heat-induced HSP70 synthesis and accumulation observed in serum-fed WI-38 cells appeared to result from an impairment in the posttranscriptional processing of the HSP70 mRNA at a level following the polyadenylation step and preceding translocation from the nucleus to the cytoplasm. When HF were serum deprived for 20 h before heat shock, the induction of HSP70 mRNA was less than 30% reduced in early passage cells in comparison to serum-fed cells; however, the level of HSP70 mRNA was markedly (over 80%) decreased in serum-deprived late passage cells. This result indicated that the presence of serum has a strong influence on heat shock-induced HSP70 gene expression in human fibroblasts aging in vitro.
Authors:
M A Bonelli; R R Alfieri; P G Petronini; M Brigotti; C Campanini; A F Borghetti
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Publication Detail:
Type:  In Vitro; Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Experimental cell research     Volume:  252     ISSN:  0014-4827     ISO Abbreviation:  Exp. Cell Res.     Publication Date:  1999 Oct 
Date Detail:
Created Date:  1999-11-01     Completed Date:  1999-11-01     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  0373226     Medline TA:  Exp Cell Res     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  20-32     Citation Subset:  IM    
Copyright Information:
Copyright 1999 Academic Press.
Affiliation:
Istituto di Patologia Generale, Università degli Studi di Parma, Parma, Italy.
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MeSH Terms
Descriptor/Qualifier:
Base Sequence
Biological Markers
Cell Aging / genetics*,  physiology
Cell Line
Cell Nucleus / metabolism
Culture Media, Serum-Free
Cytoplasm / metabolism
DNA Primers / genetics
Fibroblasts / cytology,  metabolism
Gene Expression
HSP70 Heat-Shock Proteins / genetics*
Heat-Shock Response
Humans
RNA, Messenger / genetics,  metabolism
beta-Galactosidase / metabolism
Chemical
Reg. No./Substance:
0/Biological Markers; 0/Culture Media, Serum-Free; 0/DNA Primers; 0/HSP70 Heat-Shock Proteins; 0/RNA, Messenger; EC 3.2.1.23/beta-Galactosidase

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