Molecular mechanisms of Fas-mediated cell death in oligodendrocytes

Oligodendrocyte cell death is a significant component of the secondary damage following spinal cord injury (SCI) and other neurodegenerative disorders. However, the mechanisms underlying oligodendroglial apoptotic cell death and the potential relationship to Fas receptor (FasR) activation require further clarification. Here, using MO3.13, a human oligodendroglial cell line, we show clear evidence of apoptosis upon exposure to soluble Fas ligand (sFasL). Apoptosis was linked to caspase-8, -9, and -3 activity and resulted in DNA fragmentation detected by deoxynucleotide transferase dUTP nick end-labeling (TUNEL). Dissipation of mitochondrial membrane potential (DeltaPsim) was an early event and temporally coincided with mitochondrial outer membrane permeability (MOMP), demonstrated by the presence of cytochrome c and apoptosis inducing factor (AIF) in cytosolic fractions. Pretreatment with 100 microM of the caspase inhibitor zVAD-fmk prior to sFasL exposure reduced caspase activation, the dissipation of DeltaPsim, MOMP, and apoptotic cell death. These data provide clear evidence that Fas activation induces apoptosis in oligodendrocytes signaling through intrinsic and extrinsic events. Moreover, we provide evidence for the first time that AIF may play a role in caspase-independent apoptotic execution following Fas activation of oligodendrocytes. These data also add to an emerging body of evidence, which strongly implicates Fas-mediated apoptosis of oligodendrocytes as a potential mediator in the pathobiology of a variety of neurological disorders, including SCI.     

Cell proliferation/cell death balance in renal cell cultures after exposure to a static magnetic field

The effect of a static magnetic field (MF) of 0.5 mT of intensity on the cell proliferation/cell death balance was investigated in renal cells (VERO) and cortical astrocyte cultures from rats. Magnetic stimulation was delivered by magnetic disks at known intensities. The percentage of apoptotic and necrotic cells was evaluated using flow cytometry and morphological analysis following Hoechst chromatin staining. An index of cell proliferation was determined using sulfonated tetrazolium (WST-1). Control cultures were prepared without exposure to MFs. After 2, 4 and 6 days of exposure to a MF, we observed a gradual decrease in apoptosis and proliferation and a gradual increase in cells with a necrotic morphology with respect to the control group. In astrocyte cultures, over a 6-day exposure period. A gradual increase was observed in apoptotic, proliferating, and necrotic cells. Our findings suggest that the effect of exposure to MFs varies, depending on the cell type; MFs may also have a nephropathogenic effect.     

Intracellular mediators of programmed cell death initiated at the cell surface receptor Fas

Abstract Apoptosis is a programmed cell death process, which plays a pivotal role in development, in tissue homeostasis and in several human diseases. Fas (CD95/Apo‐1) is a member of the “death receptors” family, a group of cell surface proteins that trigger apoptosis upon binding with their natural ligands. In the immune system, intracellular signal transduction triggered from Fas splits into two different pathways. The proteolytic pathway is mediated by a family of cysteine proteases, the caspases, responsible for the morphological changes occurring in the apoptotic process. To complete this death program, another series of events, involving a lipid pathway, is necessary. Upon Fas stimulation, a sequential activation of specific enzymes results in the accumulation of ceramides and GD3 ganglioside. GD3 directly induces mitochondrial damage and triggers the release of apoptogenic factors, allowing efficient execution of Fas‐mediated apoptosis.     

Many mechanisms for hsp70 protection from cerebral ischemia

Overexpression of inducible Hsp70 has been shown to provide protection from cerebral ischemia both in animal models of stroke and in cell culture models. New work suggests that there are multiple routes of cell death, including apoptotic and necrotic cell death. Hsp70 is known to protect from both necrotic and apoptotic cell death. In addition to the well-studied role of Hsp70 as a molecular chaperone assisting in correct protein folding, several new mechanisms by which Hsp70 can prevent cell death have been described. Hsp70 is now known to regulate apoptotic cell death both directly by interfering with the function of several proteins that induce apoptotic cell death as well as indirectly by increasing levels of the anti-death protein bcl-2. Despite these new insights into the ways in which Hsp70 functions as an anti-death protein, further surprises are likely as we continue to gain insight into the functioning of this multifaceted protein.     

Mitochondrial participation in ischemic and traumatic neural cell death

Mitochondria play critical roles in cerebral energy metabolism and in the regulation of cellular Ca2+ homeostasis. They are also the primary intracellular source of reactive oxygen species, due to the tremendous number of oxidation-reduction reactions and the massive utilization of O2 that occur there. Metabolic trafficking among cells is also highly dependent upon normal, well-controlled mitochondrial activities. Alterations of any of these functions can cause cell death directly or precipitate death indirectly by compromising the ability of cells to withstand stressful stimuli. Abnormal accumulation of Ca2+ by mitochondria in response to exposure of neurons to excitotoxic levels of excitatory neurotransmitters, for example, glutamate, is a primary mediator of mitochondrial dysfunction and delayed cell death. Excitoxicity, along with inflammatory reactions, mechanical stress, and altered trophic signal transduction, all likely contribute to mitochondrial damage observed during the evolution of traumatic brain injury. The release of apoptogenic proteins from mitochondria into the cytosol serves as a primary mechanism responsible for inducing apoptosis, a form of cell death that contributes significantly to neurologic impairment following neurotrauma. Although several signals for the release of mitochondrial cell death proteins have been identified, the mechanisms by which these signals increase the permeability of the mitochondrial outer membrane to apoptogenic proteins is controversial. Elucidation of the precise biochemical mechanisms responsible for mitochondrial dysfunction during neurotrauma and the roles that mitochondria play in both necrotic and apoptotic cell death should provide new molecular targets for neuroprotective interventions.     

Proteasome inhibition blocks caspase-8 degradation and sensitizes prostate cancer cells to death receptor-mediated apoptosis

BACKGROUND: Proteasome inhibition through the administration of Velcade is a viable chemotherapeutic strategy that is approved to treat multiple myeloma and is being evaluated for efficacy against prostate cancer. Currently, the apoptotic pathways that contribute to this anticancer response are poorly understood. Our goal is to test the extent to which proteasome inhibition modulates apoptosis through death receptor pathways. METHODS: Several prostate cancer cell lines and primary prostate epithelial cells (PrECs) were used as models. The death receptor pathway was activated by the expression of Fas ligand (FasL) or addition of TNF-related apoptosis-inducing ligand (TRAIL) in the presence or absence of proteasome inhibitors. The apoptotic response was quantified by annexin V, TUNEL and nuclear condensation assays. Western blot analysis was conducted to quantify protein levels and enzyme assays were used to measure caspase activity. RESULTS: Proteasome inhibition markedly sensitized prostate cancer cells to apoptosis initiated by Fas ligand (FasL) or TRAIL. In the presence of either death ligand, procaspase-8 processing occurred, but led to minimal amounts of active caspase-8. The addition of Velcade, however, led to robust active caspase-8 protein abundance and activity. In the presence of Velcade the caspase-8 p18 subunit half-life increased from 22 min to over 2 hr. CONCLUSIONS: These findings demonstrate that proteasome inhibition can sensitize cells to apoptosis elicited by tumor necrosis factor ligands and retarding caspase-8 degradation provides one explanation for this activity. This study suggests that the clinical efficacy of Velcade may result, at least in part, from the activity of death ligands.     

Role of endonucleases in renal tubular epithelial cell injury

The study of cell death has emerged as an important and exciting area of research in cell biology. Although two kinds of cell death, apoptosis and necrosis, are recognized, one of the major advances in our understanding of cell death has been the recognition that the pathways traditionally associated with apoptosis may be very critical in the form of cell injury associated with necrosis. Renal tubular epithelial cell injury from ischemia or toxins has generally been regarded as a result of a necrotic form of cell death. We briefly describe recent evidence indicating apoptotic mechanisms including endonuclease activation in renal tubular injury and some mediators (oxidants, caspases and ceramide) which regulate this process. The pathway that is followed by the cell is dependent on both the nature and severity of insults, and it is likely that the cascades that lead to the apoptotic or necrotic mode of cell death are activated almost simultaneously and may share some common pathways.     

Roles of NF-kappaB and bcl-2 in two differential modes of cell death of mouse cortical collecting duct cells

Recent data have implicated nuclear factor-kappaB (NF-kappaB) and Bcl-2 in the regulation of apoptotic and necrotic cell death in various cells. However, mechanisms of their effects on cell death of renal epithelial cells are not clear. First, we investigated the effect of specific inhibition of NF-kappaB and overexpression of Bcl-2 on necrotic cell death induced by hydrogen peroxide or cisplatin in renal collecting duct cells. M-1 cells, which were derived from outer cortical collecting duct, were stably transfected with the non-phosphorylatable mutant of inhibitory-kappaBalpha (I-kappaBalpha) and Bcl-2. Overexpression of I-kappaBalpha and Bcl-2 did not affect cisplatin-induced necrotic cell death, but overexpression of I-kappaBalpha significantly decreased H2O2-induced cell death. Regarding apoptotic cell death induced by cisplatin, serum deprivation and contact inhibition was increased by overexpression of I-kappaBalpha, whereas overexpression of bcl-2 inhibited the apoptotic cell death. I-kappaBalpha overexpression increased Bax expression and decreased cIAP-1 and -2 expression compared to vector-transfected cells, but did not alter SAPK/JNK activity in the presence or absence of cisplatin. NF-kappaB activity was significantly higher in bcl-2-overexpressing cells than in control cells. These data show that activation of NF-kappaB mediates H2O2-induced necrotic injury, but inhibits apoptotic cell death in renal collecting duct cells, and that Bcl-2 selectively protects apoptotic cell death in M-1 cells.     

Programmed cell death and survival pathways in prostate cancer cells

Programmed cell death, or apoptosis, is a series of morphologically and biochemically related processes. The extrinsic (death receptor mediated) and intrinsic (mitochondrial-mediated) apoptotic pathways can be triggered by physiological and pharmacological substances. However, other molecular events influence the sensitivity of prostate cancer cells to apoptotic stimuli, leading to marked variations in the responsiveness of prostate cancer cell lines to individual stimuli. Modulation of apoptotic responses by over expression of anti-apoptotic proteins (NF-kappaB, IAPs and Bcl-2), or attenuation of pro-apoptotic proteins (PTEN and Bax) may be responsible for the variations in sensitivity of these cell lines to hormone and chemotherapy. The expression of anti- and pro-apoptotic proteins in some of the widely used in vitro models of prostate cancer is reviewed.     

Tumor necrosis factor-alpha and lipopolysaccharide induce apoptotic cell death in bovine glomerular endothelial cells.

BACKGROUND: The glomerular endothelial cell is a specialized microvascular cell type involved in the regulation of glomerular ultrafiltration. During gram-negative sepsis, glomerulonephritis, and acute renal failure, bacterial lipopolysaccharide (LPS) and tumor necrosis factor-alpha (TNF-alpha) may cause severe cell damage. Our aim was to study and compare the direct effects of TNF-alpha and LPS on the induction of apoptosis in bovine glomerular endothelial cells. METHODS: Primary bovine glomerular endothelial cells were stimulated with TNF-alpha or LPS, and apoptotic cell death was investigated by DNA fragmentation analysis, morphological studies, measurement of cytochrome c efflux and mitochondrial permeability transition, Bak, Bad, Bax, Bcl-2, Bcl-xL protein expression, and caspase-3-like protease activity. RESULTS: TNF-alpha, as well as LPS, elicited apoptotic cell death both time and concentration dependently. Along with DNA ladder formation, we detected the formation of 50 kbp high molecular weight DNA fragments, nuclear condensation, and mitochondrial permeability transition. Concerning all parameters, LPS signaling proved to be more rapid than TNF-alpha. Mechanistically, TNF-alpha-induced cell death was preceded by an efflux of mitochondrial cytochrome c into the cytosol and, subsequently, by a marked increase in the proapoptotic protein Bak and a decrease in the anti-apoptotic Bcl-xL protein content. Comparable but more pronounced effects were seen with LPS. Later, caspase-3-like protease activity was first detectable after 10 hours and was continuously increased up to 24 hours in both TNF-alpha- and LPS-stimulated cells. Correspondingly, we detected an extended cleavage of the nuclear enzyme poly(ADP-ribose) polymerase. Caspase inhibitors Z-Asp-CH2-DCB and Z-VAD-fmk blocked both TNF-alpha- and LPS-induced apoptosis in a comparable manner. Only Z-Asp-CH2-DCB was able to block apoptotic cell death completely. CONCLUSION: Both bacterial LPS and TNF-alpha potently induced apoptotic cell death in glomerular endothelial cells. Direct endotoxin-induced apoptosis may therefore be relevant in the progression of acute renal failure, which is a frequent complication of gram-negative sepsis.     

Influence of p53 and bcl-2 on chemosensitivity in benign and malignant prostatic cell lines

The administration of cancer chemotherapeutic agents results in an increase in the apoptotic cells in the tumor: therefore, it has been assumed that anticancer drugs exhibit their cytotoxic effects via apoptotic signaling pathways. Characteristics that confer sensitivity to drug-induced apoptosis are, a functional p53 protein and expression of the apoptosis-promoting protein, bax. The role of p53 and bax/bcl-2 in drug-induced apoptosis was assessed in six prostate cell lines, 1532T, 1535T, 1542T, 1542N, BPH-1 and LNCaP using TD(50) concentrations of etoposide, vinblastine and estramustine. Cell death was monitored morphologically by fluorescent microscopy, and by flow cytometry (Annexin-V assay). Apoptotic morphology was rather low and ranged from 0.1% to 12.1%, 3.0% to 6.0% and 0.1% to 8.5% for etoposide, estramustine and vinblastine, respectively. Annexin-V binding and flow cytometry indicated apoptotic propensities of 0% to 4%, 0% to 3% and 0% to 5%, respectively. The percentage of cells responding to drug-induced apoptosis was, on average, higher in the tumor cell lines than in the normal cell lines, but showed no correlation with p53 status. The percentage of cells showing necrosis, assessed by Annexin binding and Propidium Iodide permeability in aqueous medium, tended to be much higher, and was found to be at the level of 5% to 30%. Immunoblotting demonstrated that bax and bcl-2 proteins were expressed at a basal level in all cell lines, but did not increase after exposure to TD(50) doses of the three drugs. The ratio of bax and bcl-2, measured by laser scanning densitometry, was not altered by the drug-induced DNA damage. The results suggest that apoptosis is not a major mechanism of drug-induced cell death in prostate cell lines and appears to be independent of p53 status and bax/bcl-2 expression.     

Association of HIF- expression and cell apoptosis after traumatic brain injury in the rat

Objective: To explore the expression of hypoxia inducible factor-1α (HIF-1~) and the correlation between HIF-1α and apoptosis after traumatic brain injury.Methods: Using experimental traumatic brain injury in the rats, the expression of HIF-1α was studied by immunohisto-chemistry in cerebral tissue, apoptotic cell death was evaluated with TUNEL (transferase-mediated XdUTP nick end labeling ), and double-labeled immunohistochemistry and TUNEL methods were used to investigate the relationship between HIF-1α and apoptosis.Results: There was remarkable difference in the expression of HIF-1α between the experimental groups and the control groups (P ＜ 0.01), in the experimental groups,the expression of HIF-1α at 48 hours was highest; the evidence of apoptotic cell death after experimental traumatic brain injury was found by TUNEL; the apoptotic percentage increased or decreased according to the changes of the positive expression of HIF-1α (r = 0.99).Conclusions: The results suggest that secondary brain ischemia plays a crucial role in apoptotic cell death after traumatic brain injury; HIF-1α can prompt apoptotic cell death after experimental traumatic brain injury.*e expres     

Association of HIF—1α expression and cell apoptosis after traumatic brain injury in the rat

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Renal tubular epithelial cell death and cyclosporin A.

BACKGROUND: The pathogenesis of chronic cyclosporin A (CsA) nephrotoxicity is largely unknown. In this study we examined whether CsA produces cell death through necrosis or apoptosis of either cultured human proximal tubular epithelial cells (PTEC) or the porcine tubular cell line LLC-PK(1). METHODS: Primary isolates of human PTEC and LLC-PK(1) cells were treated for various time periods with CsA at concentrations of 0.01-100 microg/ml. Apoptosis was studied by the assessment of annexin binding and propidium iodide uptake, the measurement of cellular DNA content and cell cycle analysis, and by the evaluation of nuclear morphology. Cell death was studied by the trypan blue exclusion method. Hypoxic conditions were simulated through chemical ATP depletion. RESULTS: In human PTEC, cell death was observed at CsA concentrations higher than 10 microg/ml; at these concentrations PTEC died as a result of necrosis and the toxicity of its vehicle Cremophore EL, and not as a result of CsA inducing apoptosis. The addition of cycloheximide to relieve a possible block in the apoptotic process had no effect on human PTEC, but did result in apoptosis of LLC-PK(1). In human PTEC, CsA did not augment cell death induced by chemical ATP depletion. CONCLUSIONS: The results of this in vitro study do not support the hypothesis that CsA directly induces cell death of proximal tubular epithelial cells.     

Complement-mediated killing of mesangial cells in experimental glomerulonephritis: cell death by a combination of apoptosis and necrosis

Immune system mediated, particularly antibody- and complement-mediated, glomerular injury triggers glomerulonephritis (GN). To characterize complement-mediated cytotoxicity in GN, we assessed the process of mesangial cell death induced by C5b-9 attack in Thy-1 GN. Cell injury was recognized morphologically, and nuclear DNA breaks were confirmed by the DNA nick end labeling (TUNEL) method as well as DNA gel electrophoresis. Thy-1 GN was induced in rats with anti-Thy-1.1 antibody injection. Mouse IgG (administered antibody) and rat C3 were detected in all glomeruli within 5 min after antibody injection. Damaged mesangial cells with condensed as well as TUNEL-positive nuclei could be observed at 20 min and became prominent at 40-60 min. Ultrastructurally, damaged mesangial cells contained condensed apoptotic nuclei from 40 to 60 min, whereas the cytoplasm showed necrotic degeneration. This was followed by progressive lysis of both nuclei and cytoplasm. The DNA 'ladder' pattern was observed by gel electrophoresis of extracted DNA between 40 and 60 min and correlated with the increased number of TUNEL-positive damaged mesangial cells. To examine the role of complement in this form of cell death, complement depletion was induced in rats by cobra venom factor. Complement-depleted rats showed no rat C3 deposition, rare TUNEL-positive mesangial cells, rare ultrastructural degenerated mesangial cells with apoptotic nuclei and necrotic cytoplasm, and no DNA 'ladder' pattern on gel electrophoresis at 40 min, although prominent mouse IgG was seen in glomeruli. To analyze milder forms of complement injury, a low dose of the antibody was administered to rats with a normal complement level. A few TUNEL-positive mesangial cells were detected in the glomeruli which contained apoptotic nuclei and necrotic cytoplasm. Our results indicate that an apoptotic death mechanism accompanies cell necrosis in complement-mediated mesangial cell destruction in GN and that this unusual form of cell death may represent a combination of apoptosis-necrosis within the same cell. Complement injury activates a 'death program' which in turn leads to irreversible damage of mesangial cells and which may contribute to initiation and development of GN.     

Cellular effects of imatinib on medullary thyroid cancer cells harboring multiple endocrine neoplasia Type 2A and 2B associated RET mutations

BACKGROUND: Activating mutations in the RET gene, which encodes a tyrosine kinase receptor, often cause medullary thyroid carcinoma (MTC). Surgical resection is the only curative treatment; no effective systemic treatment is available. We evaluated imatinib, a tyrosine kinase inhibitor currently used to treat chronic myelogenous leukemia and gastrointestinal stromal tumors, as a potential drug for systemic treatment of MTC, in 2 MTC-derived cell lines expressing multiple endocrine neoplasia-associated mutant RET receptors. METHODS: We determined RET expression and Y1062 phosphorylation using Western blot analysis and quantitative polymerase chain reaction. We determined the effects on cell proliferation by a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, and we used fluorescence-activated cell sorter analysis with annexin V/propidium iodide staining to study imatinib-induced cell-cycle arrest, apoptosis, and cell death. RESULTS: Imatinib inhibited RET Y1062 phosphorylation in a dose-dependent manner after 1.5 hours of exposure. After 16 hours both RET Y1062 phosphorylation and protein expression levels were affected. Dose-dependent decreases in cell proliferation of both cell lines after exposure to imatinib with inhibitory concentration of 50% levels of 23 +/- 2 micromol/L and 25 +/- 4 micromol/L were seen. These values are high, compared with those for chronic myelogenous leukemia and gastrointestinal stromal tumors. We further could show that imatinib induced cell-cycle arrest, and apoptotic and nonapoptotic cell death. CONCLUSIONS: Imatinib inhibits RET-mediated MTC cell growth affecting RET protein levels in vitro in a dose-dependent manner. The concentration of imatinib necessary to inhibit RET in vitro, however, makes it impossible to conclude that imatinib monotherapy will be a good option for systemic therapy of MTC.     

Relationship between DNA fragmentation and apoptosis in the programmed cell death in the rat prostate following castration

Previous studies have demonstrated that the rapid involution of the rat ventral prostate following castration involves the death of the androgen-dependent epithelial cells present within the gland and that this death is the result of a series of discrete biochemical steps. The degradation of genomic DNA into nucleosomal-sized fragments is an early event in this process and is catalyzed by calcium magnesium-dependent endonuclease activity. The morphologic correlation of the involution process involves a series of structural changes which are collectively referred to as apoptosis. The apoptotic process describes the earliest apparent signs of morphologic change exhibited by the dying cells through their eventual complete destruction and deletion from the tissue. The temporal relationship between these recently described biochemical events and the morphologic changes of the apoptotic process were compared in the present study, in order to test the cause versus effect nature of DNA fragmentation in the programmed death of androgen dependent prostatic cells following castration. These studies demonstrated that the early elevation of the Ca+2 Mg+2-dependent endonuclease activity and the fragmentation of DNA into nucleosomal oligomers occurs within prostatic glandular epithelial cells and probably does not involve the direct participation of extraprostatic cells which may subsequently migrate into the gland. Once the DNA is initially cleaved into the nucleosomal oligomers, the subsequent participation of lysosomal enzymes act in a less restricted fashion to degrade both the nucleosomal DNA as well as the cytoplasmic elements and the cell becomes morphologically apoptotic. As the elevations in Ca+2 Mg+2-dependent endonuclease activity and DNA fragmentation are initiated at a time well before the cell is morphologically dead, as defined by apoptosis, these changes in DNA metabolism must not be the consequences of cell death but instead are early causal events in an active process of programmed cell death.     

Syndecan-2 overexpression induces osteosarcoma cell apoptosis: Implication of syndecan-2 cytoplasmic domain and JNK signaling

Syndecans are cell surface heparan sulfate proteoglycans that serve as co-receptors and modulate the actions of a number of extracellular ligands. Syndecans thereby regulate cell adhesion, proliferation, and differentiation. Studies in cancer cells suggest that syndecans may also modulate cell viability. We previously showed that syndecan-2 controls the growth of normal human osteoblastic cells. In this study, we examined the role of syndecan-2 in osteosarcoma cell proliferation and apoptosis. To this goal, MG63 osteosarcoma cells which express low syndecan-2 levels were transfected to overexpress full-length syndecan-2 or truncated syndecan-2 lacking its cytoplasmic domain. Determination of cell growth by cell counting and 3H-thymidine incorporation showed that truncated syndecan-2 inhibited MG63 cell proliferation. Flow cytometry analysis of DNA content and colony forming test revealed that syndecan-2, but not truncated syndecan-2, induced MG63 cell death. We show that characteristic features of apoptosis such as caspase activation, PARP cleavage, cytochrome c release, increased Bax expression, and DNA fragmentation were associated with syndecan-2-induced cell death. We further show that expression of full-length or truncated syndecan-2 induced differential activation of MAPK phosphorylation in MG63 cells. Notably, syndecan-2 but not truncated syndecan-2 overexpression increased JNK phosphorylation. Moreover, SP600125, a specific inhibitor of JNK, suppressed Bax expression induced by syndecan-2 overexpression, indicating that JNK activation mediates syndecan-2-induced apoptosis. The results show that syndecan-2 induces osteoblastic cell apoptosis through the JNK/Bax apoptotic pathway and that the cytoplasmic domain of syndecan-2 is required for this action. This supports a role for syndecan-2 in the regulation of osteosarcoma cell fate and identifies one signaling pathway by which syndecan-2 induces apoptosis in osteosarcoma cells.     

Apoptotic Sertoli cell death in hypogonadic （hgn/hgn） rat testes during early postnatal development

Aim:To determine the involvement of apoptotic cell death in postnatal pathogenesis in mutant strain of hypogonadic(hgn/hgn)rats testes.We evaluated the numbers and types of cells undergoing apoptotic cell death.Methods:Tissuesections were stained by the TUNEL method for in situ detection of apoptotic cells,with specific antibodies used asmarkers of testicular somatic and germ cells.Results:We found that apoptosis in the hgn/hgn testes during the earlypostnatal period occurred primarily in Sertoli cells,which should actively proliferate during this stage of differentiation.These findings strongly suggest that the normal allele of hgn is involved in the direct or indirect control of differentia-tion and proliferation of Sertoli cells.Conclusion:To our knowledge,this is the first report demonstrating earlypostnatal apoptosis of Sertoli cells,suggesting that the hgn/hgn rat is a unique model for the study of Sertoli celldeficiency.(Asian J Androl 2006 Sep;8:535-541)