Involvement of Bcl-2 family genes and Fas signaling system in primary and secondary male germ cell apoptosis induced by 2-bromopropane in rat.
Epidemiological surveys and animal experimental studies suggest that exposure to 2-bromopropane (2-BP) could result in reproductive and hematopoietic disorders. The objectives of this study were to investigate the role of apoptosis in 2-BP-induced testicular toxicity and whether this process involves Bcl-2 family genes and the Fas signaling system. Rats were injected percutaneously with 1350 mg/kg 2-BP for 1 to 5 days and then were euthanized at 6 or 12 h after one dose, 6 h after two, three, or five doses, and 2 or 9 days after the final treatment. Light and electron microscopic analyses, TUNEL staining of DNA fragments, agarose gel electrophoresis of low-molecular-weight DNA, and Western blotting analysis of Bcl-2 family proteins and Fas receptor and ligand were conducted. Two-day treatment resulted in selective degeneration of spermatogonia with marked nuclear chromatin condensation. DNA ladder formation on the agarose gel further validated the findings of TUNEL-stained apoptotic cells. The percentage of apoptotic-positive tubules and apoptotic cell index increased time dependently. 2-BP treatment resulted in two distinct morphological changes: an immediate effect on spermatogonia and secondary apoptosis of spermatocytes 9 days after treatment. Downregulation of Bcl-2 after the first or second injection of 2-BP and upregulation of Bax after the first treatment contributed to the initiation of primary apoptosis of spermatogonia. Expression of FasL was inhibited while expression of Fas increased after the 2-BP treatment and remained at levels about two times of the control. However, it increased about sixfold of the control by day 9 after final injection, which contributed to the induction of secondary apoptosis of spermatocytes. Our results indicate that 2-BP resulted in apoptotic death of testicular germ cells and that this process involves the Bcl-2 family genes and the Fas signaling system.
Yu
X
X
National Institute of Industrial Health, Kawasaki, Japan.
Kubota
H
H
Wang
R
R
Saegusa
J
J
Ogawa
Y
Y
Ichihara
G
G
Takeuchi
Y
Y
Hisanaga
N
N
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D004247:DNA / Q000032:analysis; D053938:DNA Fragmentation; D004587:Electrophoresis, Agar Gel; D053222:Fas Ligand Protein; D015870:Gene Expression; D006842:Hydrocarbons, Brominated / Q000633:toxicity*; D020287:In Situ Nick-End Labeling; D008297:Male; D008562:Membrane Glycoproteins / Q000032:analysis / Q000378:metabolism; D008854:Microscopy, Electron; D009153:Mutagens / Q000633:toxicity*; D011518:Proto-Oncogene Proteins / Q000032:analysis; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000032:analysis / Q000235:genetics*; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction; D013090:Spermatocytes / Q000187:drug effects; D013093:Spermatogonia / Q000187:drug effects; D013094:Spermatozoa / Q000166:cytology / Q000187:drug effects*; D013737:Testis / Q000737:chemistry; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D019014:fas Receptor / Q000378:metabolism*
C494828:Bax protein, rat; C494813:Bcl2l1 protein, rat; D053222:Fas Ligand Protein; C505662:Faslg protein, rat; D006842:Hydrocarbons, Brominated; D008562:Membrane Glycoproteins; D009153:Mutagens; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D019014:fas Receptor; D004247:DNA; C103736:2-bromopropane