S. Vaithinathan

Lindane induces testicular apoptosis in adult Wistar rats through the involvement of Fas–FasL and mitochondria-dependent pathways

Lindane, an organochlorine pesticide, is known to impair testicular functions and fertility. To elucidate the mechanism(s) underpinning the gonadal effects of lindane, we sought to investigate the levels of apoptosis-related proteins, namely cytochrome c, caspase-3 and-9, Fas and FasL in the testis of adult rats. Furthermore, the study aims to delineate whether nuclear factor kappa B (NF-kappaB) is involved in meditating the testicular effects of lindane. Animals were administered with a single dose of lindane (5mg/kg body weight) and sacrificed at specific post-treatment intervals (0, 3, 6, 12, 24 and 72h). Significant elevations in the levels of cytosolic cytochrome c with a parallel increase in pro-caspase-9 were observed as early as 6h following exposure. Time-dependent elevations in the levels of Fas, FasL and caspase-3 were observed. Immunofluorescence studies revealed increased colocalization of Fas and caspase-3 in peritubular germ cells. FasL levels were increased in Sertoli and peritubular germ cells. The cytoplasmic levels of NF-kappaB p65 decreased from 3h following exposure with a maximal decline at 12 and 24h. Changes in the localization of NF-kappaB were observed with maximal nuclear translocation in germ cells at 12 and 24h. Terminal deoxynucleotidyl transferase-mediated dUTP nickend-labeling (TUNEL) assay revealed a time-dependent increase in the number of apoptotic cells. Taken together, the data illustrate induction of testicular apoptosis in adult rats following exposure to a single dose of lindane. Early activation of NF-kappaB in contrast to late increase in Fas expression suggests a pro-apoptotic role of NF-kappaB in testicular response to lindane.

Effects of plants and plant products on the testis

For centuries, plants and plant-based products have been used as a valuable and safe natural source of medicines for treating various ailments. The therapeutic potential of most of these plants could be ascribed to their anticancer, antidiabetic, hepatoprotective, cardioprotective, antispasmodic, analgesic and various other pharmacological properties. However, several commonly used plants have been reported to adversely affect male reproductive functions in wildlife and humans. The effects observed with most of the plant and plant-based products have been attributed to the antispermatogenic and/or antisteroidogenic properties of one or more active ingredients. This review discusses the detrimental effects of some of the commonly used plants on various target cells in the testis. A deeper insight into the molecular mechanisms of action of these natural compounds could pave the way for developing therapeutic strategies against their toxicity.

Methoxychlor induces apoptosis via mitochondria- and FasL-mediated pathways in adult rat testis.

In the past few years, there has been much concern about the adverse health effects of environmental contaminants in general and organochlorine in particular. Studies have shown the repro-toxic effects of long-term exposure to methoxychlor, a member of the organochlorine family. However, the insight into the mechanisms of gonadal toxicity induced by methoxychlor is not well known. In the present study we sought to elucidate the mechanism(s) underpinning the gonadal effects within hours of exposure to methoxychlor. Experimental rats were divided into six groups of four each. Animals were orally administered with a single dose of methoxychlor (50mg/kg body weight) and killed at 0, 3, 6, 12, 24, and 72h post-treatment. The levels and time-course of induction of apoptosis-related proteins like cytochorome C, caspase 3 and procaspase 9, Fas-FasL and NF-kappaB were determined to assess sequential induction of apoptosis in the rat testis. DNA damage was assessed by TUNEL assay and flowcytometry. Administration of methoxychlor resulted in a significant increase in the levels of cytosolic cytochrome c and procaspase 9 as early as 6h following exposure. Time-dependent elevations in the levels of Fas, FasL, pro- and cleaved caspase 3 were observed. The DNA damage was measured and showed time-dependent increase in the TUNEL positive cells, and also by flowcytometry of testicular cells. The study demonstrates induction of testicular apoptosis in adult rats following exposure to a single dose of methoxychlor.

Kolaviron prevents carbendazim-induced steroidogenic dysfunction and apoptosis in testes of rats

The study evaluated the protective role of kolaviron (an isolated biflavonoid from the seed of Garcinia kola) and vitamin E in carbendazim-induced reproductive dysfunction in male rats. Adult male Wistar rats were orally exposed to carbendazim (200mg/kg) singly or in combination with kolaviron (100 and 200mg/kg). Exposure to carbendazim significantly decreased the activities of superoxide dismutase and catalase but markedly increased sialic acid concentration and lipid peroxidation in the testes of rats. Western blot analysis revealed that carbendazim treatment decreased the expression of steroid acute regulatory (StAR) protein and androgen binding protein (ABP) with concomitant decrease in activities of steroidogenic enzymes. Germ cell apoptosis in carbendazim-treated rats was confirmed by TUNEL assay. However, pretreatment with kolaviron and vitamin E restored the testicular antioxidant status and steroidogenesis and decreased apoptotic nuclei to near control level in carbendazim-treated rats. Kolaviron may prove useful in combating carbendazim-induced reproductive toxicity.

Single exposure to low dose of lindane causes transient decrease in testicular steroidogenesis in adult male Wistar rats

Substantial evidence has piled up portending the adverse effects of environmental toxicants on male reproduction. Lindane, an organochlorine pesticide, has been reported to perturb testicular functions and hence fertility. To get more insight into the mechanism(s) involved in gonadal effect(s) of lindane, we sought to investigate whether treatment with lindane at a low dose would alter the levels of steroidogenic acute regulatory protein (StAR), androgen binding protein (ABP) and activities of steroidogenic enzymes (3beta-hydroxysteroid dehydrogenase and 17beta-hydroxysteroid dehydrogenase) in adult rat. Furthermore, the levels of H2O2 were monitored to delineate the possible role of H2O2 in mediating the testicular effects of lindane. Animals used for experimentation were divided into six groups and four animals were maintained in each group. Animals were administered with a single dose of lindane (5mg/kg body weight) and terminated at specific post-treatment intervals (0h, 3h, 6h, 12h, 24h and 72h) to analyze the early testicular response. Administration of lindane resulted in a sequential reduction in the levels of StAR and the activities of steroidogenic enzymes with a parallel increase in the levels of H2O2. These changes elicited by lindane were significant at 12h and 24h post-treatment. In case of ABP, a significant decline in the level was found at 12h after treatment. These findings demonstrate transient inhibitory effects of lindane on testicular steroidogenesis and the possible role of H2O2 in mediating these effects.

Methoxychlor‐induced alteration in the levels of HSP70 and clusterin is accompanied with oxidative stress in adult rat testis

Methoxychlor, an organochlorine pesticide, has been reported to induce abnormalities in male reproductive tract. However, the insight into the mechanisms of gonadal toxicity induced by methoxychlor is not well known. We investigated whether treatment with methoxychlor would alter the levels of stress proteins, heat shock proteins (HSP), and clusterin (CLU), and oxidative stress‐related parameters in the testis of adult male rats. Animals were exposed to a single dose of methoxychlor (50 mg/kg body weight) orally and were terminated at various time points (0, 3, 6, 12, 24, and 72 h) using anesthetic ether. The levels of HSP70, CLU, and the activities of superoxide dismutase (SOD), catalase, and lipid peroxidation levels were evaluated in a 10% testis homogenate. A sequential reduction in the activities of catalase and SOD with concomitant increase in the levels of thiobarbituric acid reactive substance (TBARS) was observed. These changes elicited by methoxychlor were very significant between 6–12 h of posttreatment. Immunoblot analysis of HSP revealed the expression of HSP72, an inducible form of HSP, at certain time points (3–24 h) following exposure to methoxychlor. Similarly, the levels of secretory CLU (sCLU) were also found to be elevated between 3–24 h of treatment. The present data demonstrate methoxychlor‐elicited increase in the levels of inducible HSP72 and sCLU, which could be a part of protective mechanism mounted to reduce cellular oxidative damage.

Environmental Toxicants and Testicular Apoptosis

When apoptosis is improperly activated or regulated in the testis, infertility or even cancer can result. Studies have implicated elevated rates of apoptosis in infertile male patients. Pinpointing how environmental toxicity affects apoptosis is important for the advancement of preventative medicine and behavior, especially as potentially harmful compounds continue to proliferate in households and workplaces. Moreover, familiarity with testicular processes, particularly the induction of apoptosis, is essential for promoting male fertility. This review examines environmental toxicants that have been implicated in testicular apoptosis. We elucidate the mechanistic pathways through which specific xenobiotic compounds trigger cell death in the testis. This review highlights the role of oxidative stress in mediating these apoptotic actions.

Piperine activates testicular apoptosis in adult rats

Piperine, an alkaloid present in the fruits of commonly used spice pepper, is known to impair reproductive functions. In the present study, piperine was administered to adult male rats at the dose levels of 1, 10, and 100 mg/kg body weight for 30 days to evaluate its effects on the testis. A significant decrease in the activities of antioxidant enzymes superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase in the testis was observed at 10 and 100 mg of piperine administration when compared with the controls. A dose‐dependent increase in lipid peroxidation and hydrogen peroxide generation was also observed. Sialic acid levels in the testis were also found to be decreased when piperine was administered at 10 and 100 mg dose levels. Immunofluorescence studies demonstrated a dose‐dependent increase in caspase 3 and Fas protein in testicular germ cells after piperine treatment. These observations indicate that piperine induces oxidative stress and thereby triggers apoptosis in the testis, contributing to hampered reproductive functions.

Apoptosis and Male Infertility

The pathogenesis of male infertility can be reflected by defective spermatogenesis due to pituitary disorders, testicular cancer, germ cell aplasia, varicocele, and environmental factors or due to defective sperm transport resulting from congenital abnormalities, immunological or neurological factors. Recent findings show that male infertility could be increased incidence of genetic disorders and apoptosis. Of these, apoptosis has been identified as a major factor contributing to male infertility and has been studied extensively in recent years. Apoptosis, also known as programmed cell death (PCD), is required for normal spermatogenesis in mammals and is believed to ensure cellular homeostasis, and an adequate number of germ cells are eliminated via the process of apoptosis in order to maintain a precise germ cell population in compliance with the supportive capacity of the Sertoli cells. This chapter briefs both physiological and pathological events that can trigger apoptosis and their effects on the male reproductive system.

NF-κB in Male Reproduction: A Boon or a Bane?

The apoptotic process involved in the male reproductive system regulates the ability of the male to fertilize and pass on his genes through the process of spermatogenesis. Numerous factors are involved in mediating this essential and intricate process. It is known that without programmed cell death there would be an overwhelming amount of chaos within the seminiferous tubules of the testis, which would lead to dysfunctional spermatogenesis, and problems within the male reproductive system. Of the factors effecting testicular apoptosis, we are interested in studying the effects of Nuclear Factor kappa B (NF-κB) as well as select cytokines as they appear to have a significant role in germ cell death. Other mediators will also be briefly discussed in this paper.