Spermatogenic Cells In Testis Research Articles

Ursonic acid attenuates spermatogenesis in oligozoospermia mice through inhibiting ferroptosis

Ursonic acid (UNA) is a natural pentacyclic triterpene found in some medicinal plants and foods. The reproductive protective effect of UNA was evaluated in a mouse model of oligozoospermia induced by busulfan (BUS) at 30 mg/kg b.w.. The mice were initially divided into groups with UNA concentrations of 10, 30, 50, 100 mg/kg. Subsequently, based on sperm parameters, the optimal concentration of 50 mg/kg was identified. As a control, an additional group was supplemented with ursolic acid at a concentration of 50 mg/kg. The results indicated that BUS caused the loss of spermatogenic cells in testis, the decrease of sperm in epididymis, the disorder of testicular cytoskeleton, the decrease of serum sex hormones such as testosterone which induced an increase in feedback of androgen receptor and other testosterone-related proteins, the increase of malondialdehyde and reactive oxygen species levels and the increase of ferroptosis in testis while UNA successfully reversed these injuries. High-throughput sequencing revealed that UNA administration significantly upregulated the expression of genes associated with spermatogenesis, such as Tnp1, Tnp2, Prm1, among others. These proteins are crucial in the histone to protamine transition during sperm chromatin remodeling. Network pharmacology analysis reveals a close association between UNA and proteins related to the transformation of histones to protamine. Molecular docking studies reveal that UNA can interact with the ferroptosis-inhibiting gene SLC7A11, thereby modulating ferroptosis. Taken together, UNA alleviated BUS-induced oligozoospermia by regulating hormone secretion, mitigating oxidative stress and promoting recovery of spermatogenesis by inhibiting the ferroptosis.

Time-resolved ICP-MS analysis of mineral element contents and distribution patterns in spermatogenic cells of different types

Mineral elements play an important role in the spermatogenesis, maturation, and fertilization of sperm. It is of great scientific significance to study the role of mineral elements in spermatogenesis by accurately measuring the content of elements in different spermatogenic cells and analyzing the distribution pattern of elements in spermatogenesis. Here, time-resolved inductively coupled plasma mass spectrometry (ICP-MS) was used to analyze the content and distribution patterns of mineral elements in spermatogenic cells of different types at the single cell level. Firstly, spermatogonia, spermatocytes, round spermatids and elongating spermatids were successfully isolated from testis of mice of different weeks of age by differential adherent method and discontinuous bovine serum albumin (BSA) density gradient method. Then, signal profiles and elemental distributions of 24Mg, 31P, 52Cr, 55Mn, 56Fe and 66Zn in spermatogenic cells were obtained with dwell time at 0.1 ms. Based on the results of acid digestion, we derived a formula to calculate element content in single cell from peak area for each element, and the feasibility and universality of the formula in the quantitative detection of single cell elements were verified by sperm samples to a certain extent. The detection results of element content in single cell showed that the content of 31P in elongating spermatids was significantly higher than that in spermatogonia, spermatocytes and round spermatids (P < 0.01), and the distribution range was wider. However, the 52Cr and 56Fe content of elongating spermatids was lower than that of spermatogonia, spermatocytes and round spermatids (P < 0.05). When spermatogonia developed into round spermatids, the contents of 55Mn and 66Zn in single cell increased significantly (P < 0.05), then decreased to the lowest in elongating spermatids. In addition, the significant decrease of 52Cr, 55Mn, 56Fe and 66Zn content in elongating spermatids also be visually observed from the center of the fitting curve of the element signal intensity distribution moving to the left. This study provides an elemental view of the changes in elemental content at various stages of spermatogenesis at the single-cell level. Time-resolved ICP-MS is used to detect mineral elements content and distribution patterns in spermatogenic cells of testis, which is helpful to better explore the stages and modes of action of various elements in spermatogenesis, and provide direct evidence for revealing the effects of element content changes on spermatogenesis and semen quality regulation.

Temperature-induced testicular germ cell loss and recovery in Nile tilapia Oreochromis niloticus

Water temperature is a critical external factor influencing gonadal development in fish. This research aimed to study the impact of elevated temperature on testicular germ cell survival and reproductive capacity of Nile tilapia. Male Nile tilapia were exposed to high temperatures of either 36 (HT1) or 37 °C (HT2) for 3000 degree-days (DD) and thereafter returned to the control temperature of 27 °C (CT) for 2200 DD. The deleterious effects on testicular germ and somatic cells were observed histologically, characterised by vacuolisation, atrophy and the loss of spermatogenic cells in testes with a more severe impact of HT2 compared to HT1. Interestingly, serum 11-ketotestosterone (11-KT) and testosterone (T) levels tended to be higher during the heat treatments than CT. Expression levels of germline-specific genes piwil1, piwil2 and nanos2 and Bcl-2 family genes, bcl-xLb and baxa were significantly reduced during the heat treatment compared to CT, more so in the HT2, while the levels of nanos3 and gfra1 transcripts were only significantly reduced in HT2, implying a significant loss of spermatogonial stem cell (SSC) and spermatogonia in HT2. The effect of HT2 is further evidenced by the significantly reduced sperm density and fertilisation rate compared to CT and HT1 at the end of the recovery period but complete sterility was not induced by HT2. Overall, the present study showed significant effects of HT2 on germ cell survival with histological changes in testes, reduced milt quality, increased 11-KT, and decreased expression of germline-specific genes, SSC marker genes and Bcl-2 family genes in testes which could therefore be potential target genes for sterilisation by genome editing.

Electron microscopic changes of the testis germinal epithelium after an experimental thermal trauma in the application of cryo-lyophilized xenograft skin substrate

Deep, large area of thermal trauma of the skin leads to significant morphofunctional changes in organs in the composition of the burned organism. However, condition of the central organ of the male reproductive system - testicle after burns and, especially, in the treatment of thermally damaged areas of the skin remain poorly understood. The purpose of this study was to establish an ultrastructural re-organization of testis germinal epithelium cells in the stages of toxemia and septicotoxemia following an experimental thermal trauma when using xenograft skin substrate. The studies were carried out on 35 sexually mature white male rats, which were divided into three groups: 1 - intact animals (5 rats), 2 - animals with burn injury (15 rats), 3 - animals with burn injury, for which were used xenograft skin substrate (15 rats). The burn was applied under ketamine anesthesia with copper plates, heated in boiling water, on 1820% shaved body surface of animals. Early necrectomy of the damaged areas of skin was carried out 1 day after the thermal burn. The formed wounds were covered with cryo-lyophilized xenograft skin substrate. Experimental animals were decapitated at 7, 14 and 21 days of the experiment (early, late toxemia and septicotoxemia). The material sampling for electron microscopy was carried out in accordance with a generally accepted methodology. It is established that in the stage of early toxemia (7 days of experiment) there are adaptive-compensatory changes and signs of destruction of spermatogenic cells of testis. In the stages of late toxemia and septicotoxemia (14 and 21 days of the trial), significant destructive changes occur in all components of the seminiferous tubules of the organ. Electron microscopically found that the closure of the burn wound after the early necrectomy of cryo-lyophilized xenograft skin substrate in severe experimental burn injury in the early stages of the experiment significantly reduces the damage of testis germinal epithelium of experimental animals and activates regenerative processes. This contributes to a significant improvement in the ultrastructure of the components of the convoluted tubules of the organ in the late stages of the experiment.

Overexpression of TDRP1 Gene in Swine Testis Cell and Its Global Transcriptome Analysis

The testis development-related protein-1 (TDRP1) gene was first isolated in spermatogenic cells of testis, and was related to the spermatogenesis and reproduction traits in mammals. This study was performed to further analyze the function of the pig TDRP1 gene by microarray screening of the TDRP1-overexpressed swine testis (ST) cell. After transfection of the recombinant pcDNA3.1-TDRP1 vector into the ST cell, the expression of the TDRP1 gene continued to increase at 12, 24, and 36 h post-transfection, and then decreased at 48 h, as indicated by both the fluorescence signal and quantitative PCR. Microarray screening revealed 529 upregulated and 1086 downregulated genes in TDRP1-overexpressed ST cells at 24 h post-transfection (TD-24) versus untransfected control (TD-blank), and 764 upregulated and 858 downregulated genes at 36 h post-transfection (TD-36) versus TD-blank, as well as 720 upregulated and 375 downregulated genes in TD-36 versus TD-24. As far as three contrasts were totally considered, 29 common genes were identified, including 17 upregulated genes and 12 downregulated ones. Further, Kyoto encyclopedia of genes and genomes prediction analysis indicated that some of these 29 differently expressed genes were involved in a number of biological pathways. Among the 17 upregulated genes, the PMAIP1 gene was significantly enriched in the p53 signaling pathway, and the others were the DDIT3 gene in the mitogen-activated protein kinases (MAPK) signaling pathway, the PPP1R3C gene in the insulin signaling pathway, as well as the GADD45A gene in the p53 signaling pathway, MAPK signaling pathway, and the cell cycle. Among the 12 downregulated genes, the SFRP4 gene was significantly enriched in the Wnt signaling pathway. This study indicated that the TDRP1 gene regulated PMAIP1, GADD45A, DDIT3, and PPP1R3C expression, and these genes involving p53, MAPK, insulin, and Wnt signaling pathways are related to spermatogenesis or reproduction in pig.

Expression and tissue localization of renalase, a novel soluble FAD-dependent protein, in reproductive/steroidogenic systems

Renalase was initially identified in human kidney as a soluble monoamine oxidase. Here we show that renalase is predominantly expressed in reproductive/steroidogenic systems, with particularly substantial expression in oocytes, granulosa, interstitial and luteal cells of ovary, spermatogenic cells of testis, and cortex of adrenal gland, suggesting its function(s) in maturation of germ cells and steroid hormone regulation. Renalase expression increases in testes and ovaries as mice develop and its expression is further enhanced in the ovaries of pregnant mice, indicating an activity of renalase in reproduction. Gonadotropin-releasing hormone (GnRH) antagonist, cetrorelix, repressed renalase expression in mice ovaries and testes, suggesting that steroids regulate renalase expression. Leptin is an effector and modulator of steroid hormones and reproduction. Surprisingly, knockout of leptin causes a dramatic increase of renalase expression in mice testes. Taken together, our results suggest that reproductive/steroidogenic systems are also the sources for renalase secretion and renalase may play a critical role in reproduction and hormone regulation. This provides a novel insight into understanding the function of renalase.

Maternal cypermethrin exposure during lactation impairs testicular development and spermatogenesis in male mouse offspring

Within the last decade, numerous epidemiological studies have demonstrated that endocrine disruptors are a possible cause for a decline in semen quality. Cypermethrin is a widely used pyrethroid insecticide, but little is known about its potentially adverse effects on male reproduction. In the present study, we investigated the effects of maternal cypermethrin exposure during lactation on testicular development and spermatogenesis in male offspring. Maternal mice were administered with cypermethrin (25 mg/kg) by gavage daily from postnatal day 0 (PND0) to PND21. Results showed that the weight of testes at PND21 was significantly decreased in pups whose mothers were exposed to cypermethrin during lactation. Maternal cypermethrin exposure during lactation markedly decreased the layers of spermatogenic cells, increased the inside diameter of seminiferous tubules, and disturbed the array of spermatogenic cells in testes of pups at PND21. In addition, maternal cypermethrin exposure during lactation markedly reduced mRNA and protein levels of testicular P450scc, a testosterone (T) synthetic enzyme. Correspondingly, the level of serum and testicular T at weaning was significantly decreased in pups whose mothers were exposed to cypermethrin during lactation. Although the expression of testicular T synthetic enzymes and serum and testicular T in adulthood had restored to control level, the decreased testicular weight and histological changes were irreversible. Importantly, the number of spermatozoa was significantly decreased in adult male offspring whose mothers were exposed to cypermethrin during lactation. In conclusion, maternal cypermethrin exposure during lactation permanently impairs testicular development and spermatogenesis in male offspring, whereas cypermethrin-induced endocrine disruption is reversible.

Molecular cloning of a novel nuclear factor, TDRP1, in spermatogenic cells of testis and its relationship with spermatogenesis

We reported the identification of a novel gene termed TDRP (encoding testis development-related protein) that might be involved in spermatogenesis. The human TDRP gene had two distinct transcripts, TDRP1 and TDRP2, which encoded proteins of 183 aa and 198 aa respectively. Tdrp mRNA was predominantly expressed in testis tissue. We generated rabbit polyclonal antibodies specific against human TDRP1. Immunohistochemistry analysis showed TDRP1 was expressed in spermatogenic cells, especially with high expression in spermatocytes. We provided evidence that TDRP1 distributed in both cytoplasm and nuclei of spermatogenic cells. Expression patterns of Tdrp1 mRNA and its protein were investigated in the rat testis tissues of different developmental stages. Both Tdrp1 mRNA and its protein were barely detected in the testis of neonatal rats, increased remarkably at 3weeks postpartum, and peaked at 2months postpartum. We also investigated TDRP1 expressions in testis tissues of azoospermic men with defective spermatogenesis. Western blot analysis showed that TDRP1 expressions were significantly lower in the testis tissues of azoospermic men compared with normal controls. These current data demonstrated that as a nuclear factor, TDRP1 might play an important role in spermatogenesis.

Heat Shock Protein and Heat Shock Factor Expression in Sperm: Relation to Oligozoospermia and Varicocele

Varicocele may be associated with normozoospermia or oligozoospermia. Much controversy still exists regarding the diagnosis, management and pathophysiology of spermatogenesis alterations associated with varicocele. The increased temperature induced by varicocele and stress in general may activate heat shock proteins and heat shock factors with a protective function in cells. We analyzed the expression of 5 heat shock proteins and heat shock factors in the sperm of men with normozoospermia and oligozoospermia with or without varicocele. We performed a prospective study between June 2008 and February 2009 at an academic clinic in 117 consecutive patients with varicocele and 68 controls without varicocele. Four groups were based on the presence/absence of varicocele and normozoospermia/oligozoospermia. Subjects were studied by history, physical examination, scrotal Doppler ultrasound, semen analysis, reproductive hormone plasma levels and quantitative real-time polymerase chain reaction in RNA extracted from ejaculated sperm to analyze HSP90, HSPA4, HSF1, HSF2 and HSFY expression. Increased HSPA4, HSF1 and HSF2 were observed in the sperm of men with varicocele and in those with oligozoospermia. Levels were maximum when the 2 conditions were present. Increased HSP90 was observed in oligozoospermia cases independent of varicocele. HSFY was up-regulated only in patients with varicocele, especially those with normozoospermia. To our knowledge we describe for the first time the expression of different heat shock proteins and heat shock factors in ejaculated sperm. While some of these proteins are up-regulated in men with oligozoospermia and varicocele, HSFY is up-regulated only in the presence of varicocele and especially in men with normozoospermia. This suggests that it may be a molecular marker of an adequate or inadequate response to the damaging effect of varicocele on spermatogenesis.

Localization of EFA6A, a guanine nucleotide exchange factor for ARF6, in spermatogenic cells of testes of adult mice

ADP ribosylation factors (ARFs) of small GTPase are molecular switches regulating various membrane dynamics. Among them, ARF6 has recently been highlighted because of its function to facilitate the interaction between the cytoskeleton and the plasma membrane. Each ARFs has its preferable or even specific guanine nucleotide exchange factors (GEFs) as its activators. According to our previous RT-PCR analysis, EFA6A, a guanine nucleotide exchange factor for ARF6, was restrictedly expressed in the brain, retina and testis. Different from previous studies on neurons, EFA6A, a guanine nucleotide exchange factor for ARF6, was first shown to be localized intensely in nuclei of spermatocytes of adult mouse testes in the present immunohistochemical study. This suggests a possible involvement of EFA6A-ARF6 signaling in the karyokinesis and cytokinesis.

Cypermethrin induced histological changes in gonadotrophic cells, liver, gonads, plasma levels of estradiol-17β and 11-ketotestosterone, and sperm motility in Heteropneustes fossilis (Bloch)

The aim of the present investigation is to assess the impact of cypermethrin on reproductive physiology in catfish, Heteropneustes fossilis during prespawning phase. Results indicate that there is a decrease in the size of gonadotrophic cells with less granulation, pycnosis in the liver, presence of immature oocytes and atretic follicles in the ovaries and gross condensation of spermatogenic cells in testes after cypermethrin exposure at sublethal concentration. The gonado-somatic index (GSI), plasma levels of estradiol-17β (E2) and 11-ketotestosterone (11-KT) also decreases. The motility of sperm cells is dependent on the dilution (2000 times) and duration of motility is recorded 2 min maximally at 90 s after post-activation. The dose 0.1 and 0.01 ppm is sublethal, while 1 ppm is lethal on sperm motility. Results indicate that cypermethrin causes inhibition of reproduction by acting at the hypothalamo-hypophyseal-gonadal axis as is manifest from the histological observations of gonadotrophs along with disruption of follicular wall and spermatogenic cells. Obviously such changes are responsible for decreasing the steroid hormone levels which result in decreasing scale and duration of sperm motility after 45 d exposure of cypermethrin in this species.

Distribution of α-MSH-like immunoreactive cells in the nervous system, Hatschek’s pit and other tissues of amphioxus, Branchiostoma belcheri

Immunohistochemical localization of α-melanocyte-stimulating hormone (α-MSH) in the nervous system, Hatschek’s pit and other tissues of amphioxus ( Branchiostoma belcheri) was performed using the antibody against synthetic α-MSH. The results revealed that α-MSH-like immunoreactive cells were distributed at the dorsal side and ventral side of brain vesicle, the dorsal side and the surrounding of nerve tube, and in the epithelial cells of Hatschek’s pit, the zone 1, 3, and 6 of endostyle and gut. The immunoreactive substance was also found in the primary oocytes of the small and large growth stage of ovary and early stage spermatogenic cells in testis. These findings indicate that α-MSH is an ancient and highly conserved hormone and it is extensively distributed in amphioxus. Although Hatschek’s pit in amphioxus does not have a structure of the intermediate lobe of vertebrate adenohypophysis, it has already hosted α-MSH-like endocrine cells, implying that the functional differentiation of α-MSH-like cells occurred earlier than the differentiation of the tissue structure. The results of the present study provided a new evidence for the endocrinology of Hatschek’s pit and for the origin and evolution of vertebrate adenohypophysis.

Identification of a heat‐shock protein Hsp40, DjB1, as an acrosome‐ and a tail‐associated component in rodent spermatozoa

Iba1 is a 17-kDa EF-hand protein highly expressed in the cytoplasm of elongating spermatids in testis. Using Iba1 as a bait, we performed yeast Two-hybrid screening and isolated a heat-shock protein Hsp40, DjB1, from cDNA library of mouse testis. To characterize DjB1 that is encoded by Dnajb1 gene, we carried out immunoblot analyses, in situ hybridization, and immunohistochemistry. Immunoblot analyses showed that DjB1was constitutively expressed in mouse testis and that its expression level was not changed by heat shock. Dnajb1 mRNA was exclusively expressed in spermatocytes and round spermatids in mouse testis, and Dnajb1 protein DjB1 was predominantly expressed in the cytoplasm of spermatocytes, round spermatids, and elongating spermatids. In mature mouse spermatozoa, DjB1 was localized in the middle and the end pieces of flagella as well as in association with the head (acrosomal region). Association of DjB1 with the acrosomal region in sperm head was also observed in rat spermatozoa. These data suggested that DjB1, which was constitutively expressed in postmeiotic spermatogenic cells in testis, was integrated into spermatozoa as at least two components, that is, sperm head and tail of rodent spermatozoa.

Altered expression pattern of heat shock transcription factor, Y chromosome (HSFY) may be related to altered differentiation of spermatogenic cells in testes with deteriorated spermatogenesis

To evaluate the expression patterns of heat shock transcription factor, Y chromosome (HSFY), in the testes showing deteriorated spermatogenesis. Prospective study. University hospital, its branch hospital, and academic laboratory. Men undergoing testicular biopsy for the investigation of infertility and men undergoing orchiectomy for testicular cancer. After pathologic evaluation, specimens were subdivided into three groups: normal spermatogenesis (n = 8), maturation arrest (n = 5), and Sertoli cell-only syndrome (n = 4). Immunostaining and Western blotting techniques determined the expression of HSFY. Expression of HSFY in testes. Western blotting data revealed HSFY in the testicular tissues with normal spermatogenesis, maturation arrest, and Sertoli cell-only syndrome, but the amount of the protein in the maturation arrest and Sertoli cell-only syndrome samples was altered. The immunohistochemical data demonstrated that HSFY was expressed in spermatogenic cells and Sertoli cells in all specimens. However, the expression of HSFY was low or absent in spermatogenic cells of maturation arrest specimens, and the ratio of HSFY expressed in Sertoli cells was different in the specimens with maturation arrest and with Sertoli cell-only syndrome. Altered expression of the HSFY in the testis showing deteriorated spermatogenesis may be associated with alteration of spermatogenic cell differentiation.

Concerted changes in the YB2/RYB-a protein and protamine 2 messenger RNA in the mouse testis under heat stress.

Translation of a number of mRNAs is under strict regulation via RNA-binding proteins in the spermatogenic cells of testes. A family of Y-box binding proteins represents promising candidates for these presently uncharacterized RNA-binding proteins. The effects of heat stress on the expression of a Y-box binding protein, YB2/RYB-a, and mouse protamine 2 (mP2) were investigated in cultured spermatogenic cells and mouse testes by immunoblot and Northern blot analyses. Localization and alterations in the expression of the YB2/RYB-a protein and the mP2 mRNA in heat-stressed testes were examined by immunohistochemistry and in situ hybridization, respectively. Levels of the YB2/RYB-a protein in spermatogenic cells decreased rapidly as the result of exposure to higher temperature, 37 degrees C or 43 degrees C, compared with the scrotal temperature, 32.5 degrees C, under the culture conditions used. In experimental cryptorchidism, levels of the YB2/RYB-a protein were decreased after Day 10, while the mRNA levels were affected only slightly. The levels of the mP2 mRNA were also decreased and about comparable with those of the YB2/RYB-a protein. Exposure of the lower abdomen to a high temperature, 43 degrees C for 15 min, also damaged the testis and led to a decrease in YB2/RYB-a protein and the mP2 mRNA levels in a coordinated manner. Because YB2/RYB-a is proposed to function as a stabilizer of mP2 mRNA, the perturbation of YB2/RYB-a by heat stress could account for the decline of the mP2 mRNA in elongated spermatids.

Quantitative (stereological) study of the effects of vasectomy on spermatogenesis in rhesus monkeys (Macaca mulatta).

Vasectomy reversal by vasovasostomy after long-term vasectomy in men results in lower sperm counts and pregnancy rates compared with controls, and severe damage to spermatogenesis has been observed in some animal models such as mice. The primary aim of this study was to evaluate, using sophisticated stereological methods, whether vasectomy of 6 and 12 months in a non-human primate would lead to, among other morphometric changes, reduced numbers of germ cells in testes and spermatozoa in epididymides. Five normal adult male rhesus monkeys (Macaca mulatta) underwent bilateral vasectomy, with another three aged-matched normal monkeys not undergoing vasectomy. One testis together with the ipsilateral epididymis was removed from each animal at 6 months, and the other testis and epididymis, the prostate gland and seminal vesicles were removed at 12 months. Various morphometric data were obtained using stereological methods and an unbiased and efficient stereological tool, the optical disector, was used to estimate nuclear numbers of all types of spermatogenic cells in testes and spermatozoa in epididymides using methacrylate-embedded sections 25 microm in thickness. As shown by a two-way repeated measures analysis of variance, vasectomy or hemicastration (removal of the organs at 6 months) had no significant effects on all quantitative parameters of stereology obtained from the testis, epididymis, prostate gland and seminal vesicle, except that (i) sperm granuloma was observed from three of five vasectomized animals both at 6 and 12 months, and (ii) hemicastration significantly reduced the diameter of the seminiferous tubules and increased the number of type A spermatogonia per testis. In conclusion, vasectomy in the non-human primate is a safe procedure in terms of effects on the structures of the reproductive organs.

Expression of MAGE-antigens in normal tissues and cancer

The human MAGE gene family encodes products that can be recognized by autologous cytotoxic T cells. Because MAGE genes are silent in most normal tissues except testis but are activated in a variety of neoplastic lesions, MAGE antigens represent ideal targets for immunotherapy. Current knowledge of MAGE gene expression is based primarily on mRNA typing and relatively little is known about MAGE protein expression. Monoclonal antibody (MAb) 57B, originally thought to be specific for MAGE-3, but now known to be reactive with other MAGE components, was used in the present study to analyze MAGE expression in a panel of normal and malignant tissues. In tests with a wide range of normal tissues, only spermatogenic cells of testis were reactive with 57B. In tumor tissues, significant immunoreactivity was observed in malignant melanomas and carcinomas of the lung, head and neck as well as urinary bladder. No 57B reactivity was seen with colorectal, prostatic or renal cell carcinomas. Lipo- and myosarcomas, as well as malignant fibrous histiocytoma (MFH), were negative, but synovial sarcomas showed intense immunoreactivity. A subset of seminomas was also strongly reactive with 57B. Tumor specimens showed great variability in the number of tumor cells showing 57B reactivity, with some tumors showing only small isolated clusters of positive cells to other tumors with uniform staining throughout the tumor. Int. J. Cancer 85:460–465, 2000. © 2000 Wiley-Liss, Inc.

Expression of MAGE-antigens in normal tissues and cancer

The human MAGE gene family encodes products that can be recognized by autologous cytotoxic T cells. Because MAGE genes are silent in most normal tissues except testis but are activated in a variety of neoplastic lesions, MAGE antigens represent ideal targets for immunotherapy. Current knowledge of MAGE gene expression is based primarily on mRNA typing and relatively little is known about MAGE protein expression. Monoclonal antibody (MAb) 57B, originally thought to be specific for MAGE-3, but now known to be reactive with other MAGE components, was used in the present study to analyze MAGE expression in a panel of normal and malignant tissues. In tests with a wide range of normal tissues, only spermatogenic cells of testis were reactive with 57B. In tumor tissues, significant immunoreactivity was observed in malignant melanomas and carcinomas of the lung, head and neck as well as urinary bladder. No 57B reactivity was seen with colorectal, prostatic or renal cell carcinomas. Lipo- and myosarcomas, as well as malignant fibrous histiocytoma (MFH), were negative, but synovial sarcomas showed intense immunoreactivity. A subset of seminomas was also strongly reactive with 57B. Tumor specimens showed great variability in the number of tumor cells showing 57B reactivity, with some tumors showing only small isolated clusters of positive cells to other tumors with uniform staining throughout the tumor.

Developmental studies of sperm surface antigens using sperm-specific monoclonal antibodies

Sperm-specific monoclonal antibodies generated against mouse sperm isoantigens were used to analyze the developmental expressions of sperm surface antigens during different stages of spermatogenesis. Indirect immunofluorescent assays using the freshly isolated testicular cells from mature and immature mice revealed that a number of monoclonal antibodies did not stain the surface of spermatogenic cells in testis. Instead these antibodies reacted with the sperm surface antigens in testis and/or epididymis. Further analysis was performed using frozen testicular sections from mice of day 14 to day 30 after birth. It was generally observed that a significant number of these antibodies reacted with the cytoplasmic components of spermatogenic cells in testis at the postmeiotic stages (e.g. day 22 after birth). After meiosis, the percentage of seminiferous tubules that were stained by immunofluorescence was found to increase with the ages of the mice. The results of this study suggest that some cytoplasmic components (especially acrosomal) of spermatogenic cells are expressed postmeiotically in testis, but later translocated to the sperm surface during very late stages of spermatogenesis.