Testis Development and Ultrastructural Features During Spermatogenesis in Cultured Small Yellow Croaker,Larimichthys polyactis

The small yellow croaker Larimichthys polyactis is an economically important marine fish in Northeast Asia.Currently,its natural resources are threatened by overfishing and environmental pollution.Therefore,research on the reproductive system of the fish is crucial.Here,we studied the testis development and ultrastructural features of spermatogenesis in cultured L.polyactis using anatomical,histological,and ultrastructural techniques.A pair of testes,consisting of a central sperm duct and radial seminiferous lobules,were observed.The reproduction cycle of testes can be divided into stages I–VI.March to May was confirmed as the breeding season for male L.polyactis,while April is the ideal period for artificial breeding.The male L.polyactis can attain sexual maturity within 1 year.The spermatogenesis of L.polyactis comprised spermatogonium,spermatocyte,spermatid,and mature spermatozoon.The morphology of spermatogenic cells changed obviously during spermiogenesis,including nuclear shaping,midpiece and flagellum formation.The mature sperms consist of an ellipsoidal head,a short midpiece,and a long flagellum.The anterior of the head with a kidney-shaped nucleus can be distinguished.The midpiece is located posterior to the head and includes four to six spherical mitochondria.The flagellum has irregular lateral fins.The testis of L.polyactis is an unrestricted lobular type,with cystic spermatogenesis,type II spermiogenesis,and type II spermatozoa.These features are highly similar to those of other Sciaenid species.Our findings provide useful insights into the mechanism underlying testis development and spermatogenesis of L.polyactis,which can facilitate the artificial breeding of this species.

Long-Term Impact of Acute Retinoic Acid Supplementation at the Young Age on Testicular Architecture of Wistar Albino Rats

Introduction: Inappropriate and excess vitamin supplementation, particularly for vitamin A, is increasingly recognized as a public health problem in developed countries. On the other hand, blind supplementation of vitamin A, for children in developing countries is a subject of controversy in the literature. The crucial role of vitamin A in the process of spermatogenesis in adult rodents is well established, but only a few publications are consecrated to the long-term effect of vitamin A intake at a young age on testicular development and differentiation. Objectives: Our study aimed to evaluate the long-term effects of acute supplementation at an early age, in the post-natal period, on spermatogenesis and testicular trophicity at adult age. Material and Methods: Young Wistar Albinos rats of 22 days received an acute high dose of supplementation of vitamin A (retinyl palmitate). The control group, group 1, received only extra virgin olive oil, Group 2 a dose of 7000 IU/kg of retinyl palmitate, group 3, 14,000 IU/kg, and Group 4 a dose of 28,000 IU/kg. At 10 weeks of age, the testes’ testosterone levels were measured by ELISA. For histological assessment, sections were stained with Hematoxylin eosin, and the Johnsen score was used to evaluate spermatogenesis in the seminiferous tubules. Results: The average testicular weights of rats were significantly lower in group 4 (p < 0.05), and so was the testosterone level in the testis compared to the control group (p .01). Most of the seminiferous tubules were concerned by an arrest of spermatogenesis and the Johnsen score was decreased with a mean score of 5.96 ± 1.60 (p .001) in that Group. In Group 3, Johnsen’s score was significantly better than the one obtained with the control. Conclusion: We observed a negative effect in the long term with a high acute dose of supplementation of retinyl palmitate at a young age, on testicular development and differentiation. Despite a return to normal diet after that supplementation, during childhood, impaired spermatogenesis was identified at the adult age with an arrest of spermatogenesis. The reversibility of that lack of differentiation by a return to a normal diet is questionable and would need more investigation.

The expression and significance of CATSPER1 in human testis and ejaculated spermatozoa

Aim： To investigate the distribution of cation channel of sperm 1 （CATSPER1） protein and the presence of CATSPER1 mRNA in human testis and ejaculated spermatozoa. The influence of anti-human CATSPER1 antibody upon human sperm motility was used to evaluate the function of human CATSPER1 and to estimate its possible use as a target for immunocontraception. Methods： Human ejaculated sperm from normozoospermic donors （n = 12） and liquid nitrogen frozen human testis were used for the study of mRNA and protein expression of CATSPER1 by reverse transcription polymerase chain reaction （RT-PCR） and immunohistochemistry, respectively. Spermatozoa from normozoospermic donors （n = 12） were individually processed using a swim-up procedure and were then incubated with CATSPER1 antibody at final concentrations of 20, 4 and 0.8 μg/mL. After 1, 2 and 6 h incubation, progressive motility and fast progressive motility were measured by means of computer-assisted semen analysis. Results： CATSPER1 transcript was detected in both human testis and each human ejaculated semen sample. CATSPER1 protein expressed in the membrane of spermatid and was localized in the principal piece of the sperm tail. The application of CATSPER1 antibody at all concentrations significantly inhibited both progressive motility and fast progressive motility after 1, 2 and 6 h incubation, and significant dose-dependent changes were observed. Conclusion： CATSPER1 is meiotically and post-meiotically expressed in human testis tissue. CATSPER1 mRNA in human ejaculated spermatozoa could be a more feasible target for study and infertility screening than testis biopsy. In addition, our results suggest that human CATSPER1 could be a possible target for immunocontraception.

Effects of melatonin on lipid peroxidation and antioxidant enzymes in streptozotocin-induced diabetic rat testis

Aim： To examine the effects of melatonin treatment on lipid peroxidation （LPO） and the activities of antioxidant enzymes in the testicular tissue of streptozotocin （STZ）-induced diabetic rats. Methods： Twenty-six male rats were randomly divided into three groups as follows： group Ⅰ, control, non-diabetic rats （n = 9）; group Ⅱ, STZ-induced, untreated diabetic rats （n = 8）; group Ⅲ, STZ-induced, melatonin-treated （dose of 10 mg/kg·day） diabetic rats （n = 9）. Following 8-week melatonin treatment, all rats were anaesthetized and then were killed to remove testes from the scrotum. Results： As compared to group Ⅰ, in rat testicular tissues of grouap Ⅱ, increased levels of malondialdehyde （MDA） （P 〈 0.01） and superoxide dismutase （SOD） （P 〈 0.01） as well as, decreased levels of catalase （CAT） （P 〈 0.01） and glutathione peroxidase （GSH-Px） （P 〉 0.05） were found. In contrast, as compared to group Ⅱ, in rat testicular tissues of group Ⅲ, levels of MDA decreased （but this decrease was not significant, P 〉 0.05） and SOD （P 〈 0.01） as well as CAT （P 〈 0.05） increased. GSH-Px was not influenced by any of the treatment. Melatonin did not significantly affect the elevated glucose concentration of diabetic group. At the end of the study, there was no significant difference between the melatonin-treated group and the untreated group by means of body and testicular weight. Conclusion： Diabetes mellitus increases oxidative stress and melatonin inhibits lipid peroxidation and might regulate the activities of antioxidant enzymes of diabetic rat testes.

Expression and localization of Smadl, Smad2 and Smad4 proteins in rat testis during postnatal development

<abstract>Aim: To study the expression and regulation of Smadl, Smad2 and Smad4 proteins (intracellular signaling molecules of transforming growth factor-β family) in rat testis during postnatal development. Methods: The whole testes were collected from SD rats aged 3, 7, 14, 28 and 90 (adult) days. The cellular localization and developmental changes were examined by immunohistochemistry ABC method with the glucose oxidase-DAB-nickel enhancement technique. Quantitative analysis of the immunostaining was made by the image analysis system. The Smads proteins coexistence in the adult rat testis was tested by the double immune staining for CD14-Smad4 and Smad2-Smad4. The protein expression of Smad during rat testicular development was examined by means of Western blots. Results: Smadl, Smad2 and Smad4 were present throughout testicular development. The immunostaining of Smadl and Smad2 were present in spermatogenic cells. A positive immunoreactivity was located at the cytoplasm, but the nucleus was negative. Smadl was immunolocalized at the d14, d28 and adult testes, while Smad2, at the d7, d14, d28 and adult testis. There was positive irnmunoreaction in the Sertoli cells and Leydig cells as well. The immunolocalization of Smad4 was exclusively at the cytoplasm of Leydig cells and the nuclei were negative throughout the testicular development. No expression was detected in the germ cells. The results of image and statistical analysis showed that generally the expression of Smadl, Smad2 and Smad4 in the testis tended to increase gradually with the growth of the rat. Conclusion: The present data provide direct evidences for the molecular mechanism of TGF-βaction in rat testes during postnatal development and spermatogenesis.

Histological changes of the testis and epididymis in adult rats as a result of Leydig cell destruction after ethane dimethane sulfonate treatment： a morphometric study

Aim： To quantitatively study the histological changes of the testis and epididymis as a result of a drastic reduction of testosterone secretion. Methods： Fourteen adult Sprague-Dawley rats were injected intraperitoneally with ethane dimethane sulfonate （EDS, 75 mg/kg） and the same number of animals were injected with normal saline as a control. At days 7 and 12 （after treatment）, respectively, half of the animals from each group were killed. The testes and epididymides were removed and tissue blocks embedded in methacrylate resin. The cell number per testis was estimated using the stereological optical disector and some other parameters were obtained using other morphometric methods. Results： The EDS treatment resulted in an almost complete elimination of Leydig cells but had no effect on the numbers of Sertoli cells per testis. At day 7 after EDS treatment, many elongated spermatids were retained in the seminiferous epithelium and many round spermatids could be seen in the epididymal ducts. At day 12, a looser arrangement of spermatids and spermatocytes became evident, with apparent narrow empty spaces being formed between germ cells in an approximately radial direction towards the tubule lumen; the numbers （per testis） of non-type B spermatogonia and spermatocytes were similar to controls, whereas that of type B spermatogonia increased by 59%, and that of early round, elongating and late elongated spermatids decreased by 37%, 72% and 52%, respectively. Conclusion： The primary spermatogenic lesions following EDS administration were （i） spermiation failure and （ii） detachment of spermatids and spermatocytes associated with impairment in spermiogenesis and meiosis.

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.

Targeting cancer testis antigens for biomarkers and immunotherapy in colorectal cancer: Current status and challenges

Colorectal cancer ranks third among the estimatedcancer cases and cancer related mortalities in United States in 2014. Early detection and efficient therapy remains a significant clinical challenge for this disease. Therefore, there is a need to identify novel tumor asso-ciated molecules to target for biomarker development and immunotherapy. In this regard, cancer testis antigens have emerged as a potential targets for developing novel clinical biomarkers and immunotherapy for various malignancies. These germ cell specific proteins exhibit aberrant expression in cancer cells and contribute in tumorigenesis. Owing to their unique expression profile and immunogenicity in cancer patients, cancer testis antigens are clinically referred as the most promising tumor associated antigens. Several cancer testis antigens have been studied in colorectal cancer but none of them could be used in clinical practice. This review is an attempt to address the promising cancer testis antigens in colorectal cancer and their possible clinical implications as biomarkers and immunotherapeutic targets with particular focus on challenges and future interventions.

Identification of a novel testis-specific gene and its potential roles in testis development/spermatogenesis

Aim:To identify and characterize a novel gene with potential roles in testis development and spermatogenesis. Methods:A cDNA microarray was constructed from a human testis large insert cDNA library and hybridized with probes of human or mouse adult and fetal testes.Differentially expressed genes were isolated and sequenced.RT-PCR was used to test the tissue distribution of the genes of interest and in situ hybridization was performed to localize the gene expression in the mouse testis.A range of bioinformatical programs including Gene Runner,SMART,NCBI Blast and Emboss CpGPlot were used to characterize the new gene's feature.Results:A novel testis-specific gene, NYD-SP5,was differentially expressed in fetal and adult testes.The deduced protein structure of NYD-SP5 was found to contain an IQ motif (a short calmodulin-binding motif containing conserved lie and Gin residues),a Carbam- ate kinase-like domain,a Zn-dependent exopeptidase domain and a lactate dehydrogenase (LDH) C-terminal-like domain.RT-PCR analysis revealed that NYD-SP5 was predominantly expressed in the testis but not in other 15 tissues examined.In situ hybridization and RT-PCR examinations revealed that the expression of NYD-SP5 was confined in the male germ cell but not present in the somatic cell in the testes.Conclusion:NYD-SP5 is a newly found testis- specific gene with potential roles in testis development and spermatogenesis through a calmodulin-activated enzyme.

Influence of selenium induced oxidative stress on spermatogenesis and lactate dehydrogenase-X in mice testis

Aim: To evaluate the effect of oxidative stress on the spermatogenesis and lactate dehydrogenase-X (LDH-X) activity in mouse testis. Methods: For creating different levels of oxidative stress in mice, three selenium (Se) level diets were fed in separate groups for 8 weeks. Group 1 animals were fed yeast-based Se-deficient (0.02 ppm) diet. Group 2 and Group 3 animals were fed with the same diet supplemented with 0.2 ppm and 1 ppm Se as sodium selenite, respectively. After 8 weeks, biochemical and histopathological observations of the testis were carried out. LDH-X levels in the testis were analyzed by western immunoblot and ELISA. Results: A significant decrease in testis Se level was observed in Group 1 animals, whereas it was enhanced in Group 3 as compared to Group 2. The glutathione peroxidase (GSH-Px) activity was significantly reduced in both the liver and testis in Group 1, but not in Group 2 and 3. A significant increase in the testis glutathione-S-transferase (GST) activity was observed in Group 1, whereas no significant change was seen in Groups 2 and 3. Histological analysis of testis revealed a normal structure in Group 2. A significant decrease in the germ cell population in Group 1 was observed as compared to Group 2 with the spermatids and mature sperm affected the most. Decrease in the lumen size was also observed. In the Se-excess group (Group 3), displacement of germ cell population was observed. Further, a decrease in the LDH-X level in testis was observed in Group 1. Conclusion: Excessive oxidative stress in the Se deficient group, as indicated by changes in the GSH-Px/GST activity, affects the spermatogenic process with a reduction in mature sperm and in turn the LDH-X level.

Apparent diffusion coefficient values of normal testis and variations with age

The usefulness of diffusion-weighted magnetic resonance imaging （DWI） in the evaluation of scrotal pathology has recently been reported. A standard reference of normal testicular apparent diffusion coefficient （ADC） values and their variations with age is necessary when interpreting normal testicular anatomy and pathology. We evaluated 147 normal testes using DWI, including 71 testes from 53 men aged 20-39years （group 1）, 67 testes from 42 men aged 40-69 years （group 2） and nine testes from six men older than 70years （group 3）. DWI was performed along the axial plane, using a single shot, multislice spin-echo planar diffusion pulse sequence and b-values of 0 and 900 s mm-2. The mean and standard deviation of the ADC values of normal testicular parenchyma were calculated for each age group separately. Analysis of variance （ANOVA） followed by post hoc analysis （Dunnett T3） was used for statistical purposes. The ADC values （x 10-3 mm2s-1） of normal testicular tissue were different among age groups （group 1：1.08 ± 0.13; group 2：1.15 ±0.15 and group 3：1.31± 0.22）. ANOVA revealed differences in mean ADC among age groups （F= 11.391, P〈 0.001）. Post hoc analysis showed differences between groups 1 and 2 （P= 0.008） and between groups 1 and 3 （P= 0.043）, but not between groups 2 and 3 （P= 0.197）. Our findings suggest that ADC values of normal testicular tissue increase with advancing age.

Studies on relationship between testicular capsule and sperm transport in rat testis

Aim: In SD rats, histological changes in the testis were observed after bilateral capsulotomy (of the tunica albug-inea) in order to investigate the physiological role of the testicular capsule on sperm transport. Methods: Bilaterallongitudinal capsulotomy was devised to disrupt the capsular contractile function. With this technique, only the tunicavaginalis and tunica albuginea were slit open, leaving the tunica vasculosa intact to embrace the underlying testicularparenchyma. After capsulotomy, the structural changes in the seminiferous tubules, the transitional distal seminiferoussegment, and the rete testis were observed. Results: In the capsulotomized testis, there was sperm retention at thetransitional seminiferous segment and progressive degenerative changes in seminiferous tubules. Conclusion: Theresults clearly indicated that an intact testicular capsule was required for normal sperm transport from the seminiferoustubules into the rete testis. This is the first attempt to study the physiological role of the testicular capsule in intact ani-mals.Aim: In SD rats, histological changes in the testis were observed after bilateral capsulotomy (of the tunica albug-inea) in order to investigate the physiological role of the testicular capsule on sperm transport. Methods: Bilaterallongitudinal capsulotomy was devised to disrupt the capsular contractile function. With this technique, only the tunicavaginalis and tunica albuginea were slit open, leaving the tunica vasculosa intact to embrace the underlying testicularparenchyma. After capsulotomy, the structural changes in the seminiferous tubules, the transitional distal seminiferoussegment, and the rete testis were observed. Results: In the capsulotomized testis, there was sperm retention at thetransitional seminiferous segment and progressive degenerative changes in seminiferous tubules. Conclusion: Theresults clearly indicated that an intact testicular capsule was required for normal sperm transport from the seminiferoustubules into the rete testis. This is the first attempt to study the physiological role of the testicular capsule in intact ani-mals.Aim: In SD rats, histological changes in the testis were observed after bilateral capsulotomy (of the tunica albug-inea) in order to investigate the physiological role of the testicular capsule on sperm transport. Methods: Bilaterallongitudinal capsulotomy was devised to disrupt the capsular contractile function. With this technique, only the tunicavaginalis and tunica albuginea were slit open, leaving the tunica vasculosa intact to embrace the underlying testicularparenchyma. After capsulotomy, the structural changes in the seminiferous tubules, the transitional distal seminiferoussegment, and the rete testis were observed. Results: In the capsulotomized testis, there was sperm retention at thetransitional seminiferous segment and progressive degenerative changes in seminiferous tubules. Conclusion: Theresults clearly indicated that an intact testicular capsule was required for normal sperm transport from the seminiferoustubules into the rete testis. This is the first attempt to study the physiological role of the testicular capsule in intact ani-mals.

Expression of a novel bHLH-Zip gene in human testis

<abstract>Aim: To identify specifically expressed genes in the adult and fetal testes. Methods: A human testis cDNA microarray was established. Then the mRNA of adult and fetal testis was purified and probes were prepared by a reverse transcription reaction with the testis mRNA as template. The microarray was hybridized with probes of adult and fetal testes. The nucleic sequences of differentially expressed genes were determined and homologies were searched in the databases of the GenBank. Results: When hybridized with adult or fetal testis probes, the positive clones were 96.8 % and 95.4 %, respectively. Among these genes, one was a new testis-specific gene, which was named TSP1. TSP1 was highly expressed in human adult testis. The cDNA of TSP1 was 1,484 bp in length. The cDNA sequence of this clone was deposited in the Genbank (AF333098). TSP1 was also determined as Interim Gen Symbol (Unigene, No. Hs.98266). Protein analysis showed that TSP1 contained two functional domains: an N-terminal basic helix-loop-helix (bHLH) and a C-terminal leucine zipper (Zip). Homologous analysis showed that the 430 amino acid sequences deduced from the 1,293 bp open reading frame (ORF) had a homology with the human gene FLJ2509 (AK098575). TSP1 had also a sequence homology with Spz 1 protein of mouse. Expression profiles showed that TSP1 was specifically and strongly expressed in the testis. Conclusion: TSP1 is a gene highly expressed in adult testis. It may play an important role in spermatogenesis in the humans.

Cytokines in the BALB/c mouse testis in various conditions

Aim: To investigate whether testosterone, estrogens, vasectomy, experimental cryptorchidism, varicocele or agingwould induce changes in the cytokine environment of the mouse testis. Methods: In adult male BALB/c mice,testosterone implants, estradiol benzoate, vasectomy, unilateral cryptorchidism, unilateral varicocele were adminis-tered/performed. The mice were followed up for different periods of time and were then sacrificed with testes incisedfor examination. The control mice received the vehicle or sham-operation. Results: IL-10 was present in Leydigcells of nearly every testis and IL-10 + macrophages in 39% of testes. IL-6 was found in the testes of intact adultmice, mice treated with testosterone for 70 days, cryptorchid testes and sham-operated testes. Conclusion: Resultssuggest that IL-10 might be involved in the generation of the immunologically privileged microenvironment in the testis.(Asian J Androl 2001 Mar; 3: 9-19)

Protective effects of estrogens and caloric restriction during aging on various rat testis parameters

Aim： To investigate the effects of 17β-estradiol （E2）, Peganum harmala extract （PHE） and caloric restriction （CR） on various testis parameters during aging. Methods： Twelve-month-old male rats were treated for 6 months with either E2 or PHE, or submitted to CR （40%）. Results： Our results show that estrogens and CR are able to protect the male gonad by preventing the decrease of testosterone and E2 levels as well as the decrease of aromatase and estrogen receptor gene expressions. Indeed, E2, PHE and CR treatments induced an increase in the superoxide dismutase activities and decreased the activity of testicular enzymes： gamma-glutamyl transferase, alkaline phosphatase, lactate deshydrogenase as well as the aspartate and lactate transaminases in aged animals. In addition, the testicular catalase and gluthatione peroxidase activities were enhanced in E2, PHE and CR-treated rats compared to untreated animals at 18 months of age. Moreover, the positive effects of estradiol, PHE and CR were further supported by a lower level of lipid peroxidation. Recovery of spermatogenesis was recorded in treated rats. Conclusion： Besides a low caloric diet which is beneficial for spermatogenesis, a protective antioxydant role of estrogens is suggested. Estrogens delay testicular cell damage, which leads to functional senescence and, therefore, estrogens are helpful in protecting the reproductive functions from the adverse effects exerted by reactive oxygen species （ROS） produced in large quanti- ties in the aged testis.

Normal and varicocele testis in adolescents

The authors reviewed the results of their research on the structure and composition of normal and varicocele semi-niferous tubules in adolescents. They give new evidences of normal structure of adolescent testis and demonstrate, forthe first time, the ultrastructural and immunohistochemical modifications of the lamina propria and basal lamina in theadolescent varicocele patients, which are similar to those observed in adults, but less severe, and of the adherence junc-tions in seminiferous tubules. They also report the presence of oxidative stress in adolescents limited to testis and notgeneralised as in the adults. These data are well correlated to different clinical studies that support the hypothesis of aprogressive course of varicocele and the need for surgical treatment in adolescent varicocele patients. (Asian J Androl2001 Dec; 3: 259-262)

Interstitial tissue-specific gene expression in mouse testis by intra-tunica albuguineal injection of recombinant baculovirus

The purpose of this study is to establish a gene delivery system for interstitial tissue-specific protein expression in mice testes using modified recombinant baculovirus. Green fluorescent protein （GFP）-expressing recombinant baculovirus （GFP-baculovirus）, in which the insect cell-specific polyhedron promoter was replaced by the cytomegalovirus （CMV）-IE promoter, was used to transfect testicular cells in vitro, and for intra-tunica albuguineal injection of the interstitial tissue of the testis. GFP expression was monitored in frozen testes sections by fluorescence microscopy. Expression of GFP in testicular tissues was also assessed by reverse transcription polymerase chain reaction （RT-PCR）, and protein expression was assessed by Western blot. Testicular cells in vitro were infected efficiently by modified recombinant GFP-baculovirus. lntra-tunica albuguineal injection of GFP- baculovirus into the mouse testis resulted in a high level of GFP expression in the interstitial tissues. RT-PCR analysis clearly showed GFP gene expression in the testis, particularly interstitial tissues. Intra-tunica albuguineal injection of a modified baculovirus that encoded recombinant rat insulin-like growth factor binding protein （IGFBP）-5 resulted in an increase in IGFBP-5 in testis and semen. In conclusion, we have developed an efficient delivery system for gene expression in vivo in testicular cells, particularly cells of the interstitial tissue using intratunica albuguineal injection of a modified recombinant baculovirus. This method will be particularly relevant for application that requires gene delivery and protein expression in the testicular cells of the outer seminiferous tubule of the testis.

Vitamin E ameliorates aflatoxin-induced biochemical changes in the testis of mice

Aim: To assess the effect of aflatoxin on biochemical changes in the testis of mice and the possibility of ameliorationby vitamin E treatment. Methods: Adult male albino mice were orally administered with 25 or 50μg of aflatoxin/animal/day (750 or 1500 μg/kg body weight) for 45 days. The testis was isolated and processed for biochemical anal-ysis. Results; There was a significant, dose-dependent reduction in DNA, RNA, protein, sialic acid contents andthe activities of succinic dehydrogenase, adenosine triphosphatase and alkaline phosphatase in the testis of aflatoxin-treated mice as compared to the vehicle control. However, the acid phosphatase activity was significantly increased inthe aflatoxin-treated mice. Vitamin E (2 mg/animal/day) treatment significantly ameliorated the aflatoxin-inducedchanges, except the acid and alkaline phosphatase activities in the high dose group. Conclusion; Vitamin E treat-ment ameliorates the aflatoxin-induced changes in the testis of mice. (Asian J Androl 2001 Dec; 3: 305 - 309)

Ameliorative effect of vitamin E on aflatoxin-induced lipid peroxidation in the testis of mice

Aim: To evaluate the ameliorative effect of vitamin E on aflatoxin-induced lipid peroxidation in the testis. Meth-ods: Adult male albino mice were orally administered 25 or 50 μg of aflatoxin in 0.2 mL olive oil per d for 45 d.The testis was isolated, blotted free of blood and processed for biochemical analysis. Results: There was a dose-de-pendent significantly higher lipid peroxidation in the testis of aflatoxin treated mice than in the controls. The levels ofnon-enzymatic antioxidants such as glutathione, total and reduced ascorbic acid, as well as the activities of enzymaticantioxidants, such as superoxide dismutase, glutathione peroxidase and catalase were significantly lower in the testis ofaflatoxin treated mice. Vitamin E (2 mg/d per animal; orally) pretreatment significantly ameliorates the aflatoxin-in-duced lipid peroxidation which could be due to higher enzymatic and non-enzymatic antioxidants in the testis of mice ascompared with those given aflatoxin alone. Conclusion: Vitamin E pretreatment significantly ameliorates aflatoxin-induced lipid peroxidation in the testis of mice. (Asian J Androl 2001 Sep; 3: 217 - 221)

Expression of the retinoic acid-metabolizing enzymes RALDH2 and CYP26b1 during mouse postnatal testis development

Aim： To study the expression pattern of the retinoic acid metabolizing enzymes RALDH2 and CYP26bl during mouse postnatal testis development at both mRNA and protein levels. Methods： Real-time polymerase chain reaction and Western blot analysis were performed to determine the relative quantity of RALDH2 and CYP26bl at both mRNA and protein levels at postnatal day 1, 5, 10, 20, and in adult mice （70 days testes）. Testicular localization of RALDH2 and CYP26b 1 during mouse postnatal development was examined using immunohistochemistry assay. Results： Aldhla2 transcripts and its protein RALDH2 began to increase at postnatal day 10, and remained at a high level through postnatal day 20 to adulthood. Cyp2661 transcripts and CYP26bl protein did not change significantly during mouse postnatal testis development. RALDH2 was undetectable in the postnatal 1, 5 and 10 day testes using immunohis- tochemistry assay. At postnatal day 20 it was detected in pachytene spermatocytes. Robust expression of RALDH2 was restricted in round spermatids in the adult mouse testis. In the developing and adult testis, CYP26bl protein was confined to the peritubular myoepithelial cells. Conclusion： Our results indicate that following birth, the level of retinoic acid in the seminiferous tubules might begin to increase at postnatal day 10, and maintain a high level through postnatal day 20 to adulthood.