Lipopolysaccharide stimulation of 70 kilo Dalton heat shock protein messenger ribonucleic acid production in cultured human fetal membranes.
OBJECTIVE
The 70 kilo Dalton heat shock protein is up-regulated when cells are under physiological stress. It prevents protein denaturation and incorrect polypeptide assembly, and inhibits apoptosis as well as the transcription of genes coding for pro-inflammatory cytokines. To evaluate if up-regulation of heat shock protein 70 can occur during pregnancy, we examined whether addition of bacterial lipopolysaccharide to human amniochorion membranes in vitro stimulated heat shock protein 70 gene transcription.


METHODS
Amniochorionic membranes (n = 5), collected at the time of elective repeat cesarean section prior to labor from normal term gestations, were placed in an organ explant system. After 48 hour in culture, the membranes were stimulated with lipopolysaccharide for 24 hours. Total RNA was extracted and subjected to an oligo dT primed reverse transcriptase reaction followed by polymerase chain reaction (PCR) using heat shock protein 70 specific primers. PCR products were hybridized with biotinylated internal probes and identified by enzyme-linked immunosorbent assay (ELISA). Results were analyzed by Mann-Whitney U test. A p < 0.05 was significant.


RESULTS
Heat shock protein 70 messenger RNA was expressed by all fetal membrane preparations both prior to and following in vitro culture. Addition of lipopolysaccharide increased the concentrations of heat shock protein 70 messenger RNA in each sample tested from a mean of 35.5 +/- 29.6 ng/milliliter (12.1-80.1 ng/milliliter) to 169.6 +/- 69.9 ng/ml (51.7-218.2 ng/milliliter) (p = 0.03).


CONCLUSIONS
Human fetal membranes constitutively express heat shock protein 70 messenger ribonucleic acid. Bacterial lipopolysaccharide markedly stimulated heat shock protein 70 messenger RNA gene transcription in human fetal membranes. Thus, heat shock protein 70 is inducible in fetal membranes and may facilitate fetal survival under adverse conditions.
Menon
R
R
Perinatal Research Center, Women's Health Research and Education Foundation and Maternal-Fetal Group, Centennial Women's Hospital, Nashville, TN, USA.
Gerber
S
S
Fortunato
S J
SJ
Witkin
S S
SS
D002585:Cesarean Section; D046508:Culture Techniques; D005321:Extraembryonic Membranes / Q000187:drug effects / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018840:HSP70 Heat-Shock Proteins; D008070:Lipopolysaccharides; D012333:RNA, Messenger