Spermatogenesis is the process by which a single spermatogonium develops into 256 spermatozoa, one of which will fertilize the ovum. Since the 1950s when the stages of the epithelial cycle were first described, reproductive biologists have been in pursuit of one question: How can a spermatogonium traverse the epithelium, while at the same time differentiating into elongate spermatids that remain attached to the Sertoli cell throughout their development? Although it was generally agreed upon that junction restructuring was involved, at that time the types of junctions present in the testis were not even discerned. Today, it is known that tight, anchoring, and gap junctions are found in the testis. The testis also has two unique anchoring junction types, the ectoplasmic specialization and tubulobulbar complex. However, attention has recently shifted on identifying the regulatory molecules that "open" and "close" junctions, because this information will be useful in elucidating the mechanism of germ cell movement. For instance, cytokines have been shown to induce Sertoli cell tight junction disassembly by shutting down the production of tight junction proteins. Other factors such as proteases, protease inhibitors, GTPases, kinases, and phosphatases also come into play. In this review, we focus on this cellular phenomenon, recapping recent developments in the field.

Sertoli-Sertoli and Sertoli-Germ Cell Interactions and Their Significance in Germ Cell Movement in the Seminiferous Epithelium during Spermatogenesis

Although it has been known for many years that estrogen administration has deleterious effects on male fertility, data from transgenic mice deficient in estrogen receptors or aromatase point to an essential physiological role for estrogen in male fertility. This review summarizes the current knowledge on the localization of estrogen receptors and aromatase in the testis in an effort to understand the likely sites of estrogen action. The review also discusses the many studies that have used models employing the administration of estrogenic substances to show that male fertility is responsive to estrogen, thus providing a mechanism by which inappropriate exposure to estrogenic substances may cause adverse effects on spermatogenesis and male fertility. The reproductive phenotypes of mice deficient in estrogen receptors alpha and/or beta and aromatase are also compared to evaluate the physiological role of estrogen in male fertility. The review focuses on the effects of estrogen administration or deprivation, primarily in rodents, on the hypothalamo-pituitary-testis axis, testicular function (including Leydig cell, Sertoli cell, and germ cell development and function), and in the development and function of the efferent ductules and epididymis. The requirement for estrogen in normal male sexual behavior is also reviewed, along with the somewhat limited data on the fertility of men who lack either the capacity to produce or respond to estrogen. This review highlights the ability of exogenous estrogen exposure to perturb spermatogenesis and male fertility, as well as the emerging physiological role of estrogens in male fertility, suggesting that, in this local context, estrogenic substances should also be considered "male hormones."

Estrogen and spermatogenesis.

THE ability of cells to interact and communicate is a biological phenomenon that developed with the evolution of multicellular organisms. The cell-cell interactions that have evolved are essential for the survival of both simple organisms such as dictyostelium and complex organisms such as mammals. These cellular associations allow an organism to develop capacities that are greater than the simple sum of their individual parts. As early as 1878, Claude Bernard proposed that the “milieu interieur” (i.e. internally produced fluid environment) and a cybernetic-like control system (i.e. cell-cell interactions and communication) in multicellular organisms are needed to adaptively regulate the growth, development, and maintenance of normal tissue function (1, 2). The experimental analysis of cellular interactions was initiated with the investigation of cell aggregation in simple organisms such as sponges (3, 4) and progressed into the areas of embryology (5) and cell biology (6, 7). Analysis of cell biology on ...

Cell-Cell Interactions in the Testis

Widely varied chemicals--including certain herbicides, plasticizers, drugs, and natural products--induce peroxisome proliferation in rodent liver and other tissues. This phenomenon is characterized by increases in the volume density and fatty acid oxidation of these organelles, which contain hydrogen peroxide and fatty acid oxidation systems important in lipid metabolism. Research showing that some peroxisome proliferating chemicals are nongenotoxic animal carcinogens stimulated interest in developing mode of action (MOA) information to understand and explain the human relevance of animal tumors associated with these chemicals. Studies have demonstrated that a nuclear hormone receptor implicated in energy homeostasis, designated peroxisome proliferator-activated receptor alpha (PPARalpha), is an obligatory factor in peroxisome proliferation in rodent hepatocytes. This report provides an in-depth analysis of the state of the science on several topics critical to evaluating the relationship between the MOA for PPARalpha agonists and the human relevance of related animal tumors. Topics include a review of existing tumor bioassay data, data from animal and human sources relating to the MOA for PPARalpha agonists in several different tissues, and case studies on the potential human relevance of the animal MOA data. The summary of existing bioassay data discloses substantial species differences in response to peroxisome proliferators in vivo, with rodents more responsive than primates. Among the rat and mouse strains tested, both males and females develop tumors in response to exposure to a wide range of chemicals including DEHP and other phthalates, chlorinated paraffins, chlorinated solvents such as trichloroethylene and perchloroethylene, and certain pesticides and hypolipidemic pharmaceuticals. MOA data from three different rodent tissues--rat and mouse liver, rat pancreas, and rat testis--lead to several different postulated MOAs, some beginning with PPARalpha activation as a causal first step. For example, studies in rodent liver identified seven "key events," including three "causal events"--activation of PPARalpha, perturbation of cell proliferation and apoptosis, and selective clonal expansion--and a series of associative events involving peroxisome proliferation, hepatocyte oxidative stress, and Kupffer-cell-mediated events. Similar in-depth analysis for rat Leydig-cell tumors (LCTs) posits one MOA that begins with PPARalpha activation in the liver, but two possible pathways, one secondary to liver induction and the other direct inhibition of testicular testosterone biosynthesis. For this tumor, both proposed pathways involve changes in the metabolism and quantity of related hormones and hormone precursors. Key events in the postulated MOA for the third tumor type, pancreatic acinar-cell tumors (PACTs) in rats, also begin with PPARalpha activation in the liver, followed by changes in bile synthesis and composition. Using the new human relevance framework (HRF) (see companion article), case studies involving PPARalpha-related tumors in each of these three tissues produced a range of outcomes, depending partly on the quality and quantity of MOA data available from laboratory animals and related information from human data sources.

PPARα Agonist-Induced Rodent Tumors: Modes of Action and Human Relevance

Steroid hormone biosynthesis in steroidogenic cells is regulated through trophic hormone activation of protein kinase A (PKA) signaling pathways. However, many examples of the regulation of steroid synthesis via pathways other than the PKA pathway have been documented. In some cases these pathways act independently of PKA activation whereas in other cases, they act synergistically with it. The current understanding of additional signaling pathways and factors, such as the protein kinase C pathway, arachidonic acid metabolites, growth factors, chloride ion, the calcium messenger system, and others capable of regulating/modulating steroid hormone biosynthesis, and in many cases steroidogenic acute regulatory protein expression, are discussed in this review.

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Multiple Signaling Pathways Regulating Steroidogenesis and Steroidogenic Acute Regulatory Protein Expression: More Complicated than We Thought

Homogenates of whole testis, isolated seminiferous tubules, testicular cytosol, conditioned media from seminiferous tubules obtained from intact or cryptorchid rats, as well as seminiferous tubules devoid of peritubular cells, showed high concentrations of interleukin-1 (IL-1). Cytosol from spleen showed low IL-1 activity, while no activity was found in cytosol from heart, kidney, prostate, ovary or liver. Interleukin-1 activity was not detected in spent medium from cultures of immature Sertoli cells (10-day-old rats) or from peritubular cells or in homogenates of interstitial cells from adult rats. Ultrogel AcA 44 gel chromatography and HPLC size exclusion chromatography exhibited a single peak of IL-1 activity corresponding to a relative molecular mass of 17 000–20 000 (Mr= 17–20 K). Similarly, chromatofocusing revealed only one peak of activity with an apparent isoelectric point of 5–6. It is concluded that the rat testis contains large amounts of an IL-1α-like factor. The adult Sertoli cell or possibly germ cells are suggested as its primary source. Testicular IL-1-like activity is of particular interest in view of the intense cell proliferation during spermatogenesis, and the tendency to testicular relapse of acute lymphoblastic leukaemia.

The rat testis produces large amounts of an interleukin-1-like factor.

Puberty in the male is dependent upon the elevated production of testosterone by the Leydig cells. LH affects this increase in testosterone output by increasing the total number of Leydig cells in the testis and by stimulating the steroidogenic pathway in these cells. Since Leydig cell proliferation is a prerequisite for the onset of puberty, we have examined the ability of LH and growth factors known to be present in the testis to promote DNA synthesis. Leydig cells were isolated from 21 -day-old rats, cultured in serum-free medium for 48 h to become quiescent; and then treated with UI and growth factors for 18 Is. l5HIThymidine incorporation into DNA was assessed over the subsequent 4-h incubation period. Cells in control cultures incorporated low levels

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Growth factor requirements for DNA synthesis by Leydig cells from the immature rat.

The number of interstitial macrophages in the testis fluctuates according to age, increasing gradually during prepubertal development to reach 15-20% in the interstitial compartment in the adult rat. These macrophages are in close morphological association with Leydig cells. Macrophage products, interleukin-1 (IL-1) and tumor necrosis factor alpha stimulate and/or inhibit steroid production in cultures of Leydig cells. We have studied the effects of macrophage products on DNA synthesis in rat Leydig cells to investigate a possible paracrine role of testicular macrophage products on the proliferation of Leydig cells. Leydig cells isolated from 10-, 20-, and 70-day-old rats were cultured for 48 h in serum-free medium, washed, and treated with different cytokines for 18 h. The medium was then removed, fresh medium containing 0.5 microCi [3H]thymidine was added, and cells were incubated for 4 h prior to determining the incorporation of [3H]thymidine into DNA. Human recombinant IL-1 beta caused a dose-dependen...

Shafiq A. Khan

Papers

The rat testis produces large amounts of an interleukin-1-like factor.

Growth factor requirements for DNA synthesis by Leydig cells from the immature rat.

Interleukin-1 stimulates deoxyribonucleic acid synthesis in immature rat Leydig cells in vitro.

Interactions between FSH, estradiol-17β and transforming growth factor-β regulate growth and differentiation in the rat gonad

Stage-specific inhibition of interstitial cell testosterone secretion by rat seminiferous tubules in vitro