Expression of tumor necrosis factor-alpha and its receptors type I and type II in human oocytes

Abstract

The present study was conducted to examine the expression of tumor necrosis factor-alpha (TNF-alpha) and its receptors (types I and II, designated TNFR-I and TNFR-II, respectively) in human oocytes and cumulus cells at the mRNA and protein levels. mRNA expression was investigated using a reverse transcriptase-polymerase chain reaction (PCR)/Southern hybridization procedure. DNA-free RNA was isolated from the oocytes/cumulus cells, reverse-transcribed, and PCR-amplified using specific oligonucleotide primers based upon genomic/cDNA sequences. The expected bands of 303 bp and 513 bp were observed in oocytes and cumulus cells using primers based on genomic/cDNA sequences of TNF-alpha and TNFR-II, respectively, that hybridized with specific cDNA probes in Southern blot hybridization procedure. The expected band of 368 bp was not observed in oocytes and cumulus cells using primers based on the TNFR-I cDNA sequence. Similar results were observed for expression at the protein level, as seen by the immunoreactivity of the specific antibodies with the paraformaldehyde-fixed oocytes and cumulus cells in the indirect immunofluorescence technique (IFT). These results indicate that human oocytes and cumulus cells express TNF-alpha and its receptor type II (TNFR-II), and not type I (TNFR-I), both at the mRNA and protein levels. These findings provide further evidence and substantiate the proposed physiologic role of TNF-alpha in ovarian function, and may lead to clinical applications in in vitro fertilization programs and in diagnosis and treatment of infertility in women, especially in cases attributed to ovarian dysfunction.