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Effects of acid sphingomyelinase deficiency on male germ cell development and programmed cell death
Marjut Otala1, Markku O Pentikäinen, Tiina Matikainen
1Program for Developmental and Reproductive Biology, Biomedicum Helsinki and Hospital for Children and Adolescents, University of Helsinki, FIN-00029 HUS, Helsinki, Finland. marjut.otala@hus.fi
Abstract:
Deficiency of acid sphingomyelinase (ASM), an enzyme responsible for producing a pro-apoptotic second messenger ceramide, has previously been shown to promote the survival of fetal mouse oocytes in vivo and to protect oocytes from chemotherapy-induced apoptosis in vitro. Here we investigated the effects of ASM deficiency on testicular germ cell development and on the ability of germ cells to undergo apoptosis. At the age of 20 weeks, ASM knock-out (ASMKO) sperm concentrations were comparable with wild-type (WT) sperm concentrations, whereas sperm motility was seriously affected. ASMKO testes contained significantly elevated levels of sphingomyelin at the age of 8 weeks as detected by high-performance, thin-layer chromatography. Electron microscopy revealed that the testes started to accumulate pathological vesicles in Sertoli cells and in the interstitium at the age of 21 days. Irradiation of WT and ASMKO mice did not elevate intratesticular ceramide levels at 16 h after irradiation. In situ end labeling of apoptotic cells also showed a similar degree of cell death in both groups. After a 21-day recovery period, the numbers of primary spermatocytes and spermatogonia at G2 as well as spermatids were essentially the same in the WT and ASMKO testes, as detected by flow cytometry. In serum-free cultures both ASMKO and WT germ cells showed a significant increase in the level of ceramide, as well as massive apoptosis. In conclusion, ASM is required for maintenance of normal sphingomyelin levels in the testis and for normal sperm motility, but not for testicular ceramide production or for the ability of the germ cells to undergo apoptosis.
Insights
Acid sphingomyelinase (ASM) deficiency impacts sperm motility and testicular sphingomyelin levels but does not affect germ cell apoptosis or ceramide production. ASM is crucial for maintaining normal testicular function.
Area of Science:
- Reproductive biology
- Cellular biology
- Biochemistry
Background:
- Acid sphingomyelinase (ASM) deficiency protects oocytes from apoptosis.
- ASM produces ceramide, a pro-apoptotic second messenger.
- The role of ASM in testicular germ cell development and apoptosis is not well understood.
Purpose of the Study:
- To investigate the effects of ASM deficiency on testicular germ cell development.
- To assess the impact of ASM deficiency on germ cell apoptosis.
- To determine ASM's role in testicular ceramide production and sphingomyelin levels.
Main Methods:
- Comparison of sperm concentration and motility in ASM knock-out (ASMKO) and wild-type (WT) mice.
- High-performance, thin-layer chromatography to measure sphingomyelin levels.
- Electron microscopy to observe testicular pathology.
- Irradiation and in situ end labeling to assess apoptosis.
- Flow cytometry to analyze germ cell populations.
Main Results:
- ASMKO mice showed significantly reduced sperm motility but comparable sperm concentrations to WT mice at 20 weeks.
- Elevated sphingomyelin levels were detected in ASMKO testes at 8 weeks.
- Pathological vesicles accumulated in Sertoli cells and interstitium of ASMKO testes from 21 days.
- Irradiation did not alter intratesticular ceramide levels or apoptosis rates in either group.
- Germ cell populations (spermatocytes, spermatogonia, spermatids) were similar in WT and ASMKO testes after recovery.
- Both ASMKO and WT germ cells underwent massive apoptosis with increased ceramide levels in vitro.
Conclusions:
- ASM is essential for maintaining normal sphingomyelin levels in the testis.
- ASM plays a critical role in ensuring normal sperm motility.
- ASM is not required for testicular ceramide production or the intrinsic apoptotic capacity of germ cells.
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