Expression of tumor necrosis factor-alpha in the rat ovary.
Tumor necrosis factor-alpha (TNF-alpha) messenger RNA (mRNA) in rat ovaries was detected by Northern blot analysis and by reverse transcription-polymerase chain reaction (RT-PCR). Northern blot analysis of total cellular RNA (tcRNA) isolated from adult rat ovaries showed a single distinct band at approximately 2 kilobases, while rat white blood cells, used as control, also showed a second lower band at approximately 0.5 kilobases. TNF-alpha expression in ovaries was also detected by RT-PCR, using RNA-specific primers. Southern blot analysis of the RT-PCR products showed a single band of the expected 500 base pairs, from the ovary and white blood cells, using either tcRNA or polyA+ RNA. In order to validate the RT-PCR product, it was digested with restriction enzymes, HhaI, HindIII, and XhoI. The results indicate that the ovarian TNF-alpha mRNA does not contain major alterations with respect to the known structure of rat TNF-alpha mRNA. Ovarian RNA was also subjected to PCR without previous RT in order to amplify, if any, contaminating genomic DNA. A single band of approximately 900 base pairs was observed, which contained introns 1 and 2 (determined by restriction enzyme digestion). In summary, these results indicate that rat ovaries contain TNF-alpha mRNA which makes it a likely source of local TNF-alpha secretion. The possibility exists that ovarian cells and/or macrophages are the source of the ovarian TNF-alpha mRNA.
Sancho-Tello
M
M
Department of Physiology, University of Kansas Medical Center, Kansas 66103.
Pérez-Roger
I
I
Imakawa
K
K
Tilzer
L
L
Terranova
P F
PF
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D004247:DNA / Q000235:genetics; D005260:Female; D015870:Gene Expression / Q000235:genetics*; D008969:Molecular Sequence Data; D010053:Ovary / Q000737:chemistry* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D051381:Rats; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis / Q000235:genetics* / Q000378:metabolism
D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D004247:DNA