Germ Cell Maturation Research Articles

"Second-look" Micro Testicular Sperm Extraction (MicroTESE) in Patients with Non-obstructive Azoospermia Following Histopathological Analysis.

Introduction:Microdissection testicular sperm extraction (microTESE) is considered the gold standard method for surgical sperm retrieval among patients with non-obstructive azoospermia (NOA).Aim:This study aimed to evaluate the correlation between histopathological findings after failed microTESE procedure and outcomes of the „second-look“ procedure and to provide insight into the most common histopathological patterns after testicular biopsy within our population.Methods:The retrospective study included 33 selected patients with NOA, who had undergone unsuccessful sperm retrieval. The diagnosis of NOA was made after the assessment of the patient’s history data, a physical examination, semen analysis, the hormonal profile, and genetic studies. After negative sperm retrieval, histopathological report has been analyzed for „second-look“ microTESE attempt.Results:Five testicular histopathological patterns were found: hypospermatogenesis (9,1%), Sertoli cell-only syndrome (43%), germ cell maturation arrest (15%), seminiferous tubule hyalinization (15%), mixed pattern (21%). Y-microdeletions were detected in 5 patients, of which 3 patients showed AZFc region deletions. Only 3 patients (9,1%) underwent a „second-look“ procedure after the evaluation of histopathological reports. After the stimulation therapy and „second-look“ procedure, we had a positive outcome in a single patient (33,3%). Mean FSH value in patients with confirmed spermatogenesis was 17.26±3.11IU/l, while mean FSH value in patients without presence or germ cell statistically significantly exceeded and was 24.28±4.71IU/L (p=0.038).Conclusion:Histopathological reports following the microTESE procedure are obligatory for the proper selection of patients who are candidates for the „second-look“ microTESE attempt. Patients with Sertoli cell-only syndrome and hypospermatogenesis particularly can benefit from the “second-look” procedure.

The efficiency of use lentiviral vectors for the transformation of rooster spermatogenic cells in vivo

Male gonad cells are considered as promising target cells for the introduction of recombinant DNA within obtaining genetically modified individuals with given characteristics. The use of testicular spermatogonial stem cells is of the greatest interest. In the process of differentiation, this type of cell gives rise to a significant population of mature male germ cells. In the case of their genetic transformation, differentiated cells can be used to inseminate females in order to produce transgenic progeny. The aim of the research was to study the efficiency of using lentiviral vectors for the local transformation of roosters’ testicular spermatogenic cells. We used a lentiviral vector containing the ZsGreen reporter gene under the control of the CMV promoter. In vitro transformation of rooster spermatogenic cells was carried out by infection with a viral preparation, in vivo through multiple injections of the viral preparation into the testicular parenchyma of roosters ( n = 5). The efficiency of transformation was assessed by expression of the reporter ZsGreen gene in transfected spermatogenic cells. The success of using lentiviral vectors for the genetic transformation of rooster spermatogenic cells was shown in experiments in vitro and in vivo . The transformation efficiency of this cells types in an in vitro culture varied from 45 to 57% and averaged 48 ± 4%. The expression of the ZsGreen reporter gene in the cells of the spermatogenic epithelium of the testes was established in almost all experimental roosters in the in vivo experiments. The number of seminiferous tubules with transformed spermatogenic cells varied in the studied experimental roosters from 10 to 22%. The effectiveness of genetic transformation of the testes spermatogenic cells was 1.8 ± 0.2%. The obtained results indicate to the success of using lentiviral vectors for the genetic transformation of spermatogenic cells of rooster testes in vivo in order to create individuals with genetically transformed germ cells for the further production of transgenic offspring with given characteristics.

PSXIII-5 The age dynamics of the spermatogenic cells development in the roosters’ testes

Abstract The cells of the male gonads are considered as a valuable genetic material for the conservation of the gene pool of breeds and lines of agricultural birds, as well as the directed modification of the poultry genome. Mature germ cells – spermatozoa and their predecessors – spermatogonia, spermatocytes and spermatids can be used for these purposes. To obtain these types of cells, it is necessary to know the characteristics of their development (spermatogenesis). The dynamics of the development of certain spermatogenic cell types in the testicular tubules of different-aged roosters has been studied. Histological studies were performed on testes of roosters aged from 1 week to 6 months with an interval of 2 weeks. Samples of testis tissue were fixed in Bouin’s solution. Histological sections were stained with hematoxylin-eosin. Identification of different cell types (Sertoli, spermatogonia, spermatocytes, spermatids, sperm cells) was carried out according to their morphology. At the age of 1–6 weeks in the seminiferous tubule of roosters, the mainly presence of two cell types was noted: Sertoli cells and spermatogonia. From 7 weeks of age, spermatocytes were detected in the seminiferous tubules, in the 4 months - spermatids, in the 5.5 months - sperm cells. The number of Sertoli cells remained almost unchanged with age and was 21 ± 2. The percentage of these cells decreased with age from 71 ± 3 % to 5 ± 1 %. The percentage of spermatogonia also decreased with age from 75 ± 2 % to 7 ± 1 %. The number of spermatids and spermatozoa, on the contrary, increased to puberty (6 months) and reached 54 %. The study was supported by the RFBR within Project no.18-29-07079.

137 Recovery of quail spermatogenesis by donor spermatogonia transplantation

Spermatogonia, the stem cell precursors of male germ cells, are used as convenient biological material for the preservation of genetic resources (cryobanks) and the introduction of recombinant DNA (transgenesis). Donor spermatogonia subsequently differentiate into mature germ cells (spermatozoa), which are used to produce offspring. Our laboratory is investigating methods to improve the efficiency of spermatogonial germ cell transplantation in quail. The objectives of this study were to (1) determine the optimal age for spermatogonia isolation from the testes of donor male quail and (2) identify the most appropriate concentration of busulfan for treatment of recipient quail testes. Statistical analysis was performed using SPSS ver. 15.0 (IBM Corp.; analysis of variance test). In order to determine the optimal age for spermatogonia isolation, testes from male quail at 1-4 weeks of age were isolated and histological studies were performed on a population of spermatogenic cells in the seminiferous tubules. Histological studies of quail testes isolated at different ages showed that the optimal age for obtaining a culture of spermatogonia is a period from 1-2 weeks of age. During this period, spermatogenic cells were represented mainly by spermatogonia (P<0.01). Therefore, testes of 1-week-old quails were used to obtain a culture of spermatogonia. The resulting cell culture consisted mainly of spermatogonia (85%) with a small number of Sertoli cells. Next, a series of experiments introducing busulfan into quail testes was carried out using concentrations from 10-150mg kg−1 of liveweight. Experiments showed that an effective dose to remove the recipient male's own spermatogenic cells was a concentration of 100mg kg−1 of liveweight (P<0.05). Finally, using the optimal parameters described above, spermatogonia cultures were obtained and introduced into the testes of quail recipients (n=6), following administration of busulfan for 2-3 weeks before donor spermatogonia were introduced. The effectiveness of spermatogenesis recovery was assessed based on the analysis of sperm from quail recipients at 3 months after the injection of donor cells. The presence of donor germ cells in the testes of quail recipient drakes was confirmed by microsatellite analysis of DNA isolated from the blood and sperm of recipients as well as the donor cells (spermatogonia). The microsatellite profiles of the blood and sperm DNA in quail recipient males were different, which confirms the restoration of spermatogenesis in the studied recipients due to the development of donor germ cells. The reported study was funded by RFBR, project number 18-29-07079.

Long non-coding RNA LOC105611671 modulates fibroblast growth factor 9 (FGF9) expression by targeting oar-miR-26a to promote testosterone biosynthesis in Hu sheep.

Fibroblast growth factors (FGFs) play crucial roles in early gonadal development and germ cell maturation of mammals; FGF9 is involved in mammalian testis steroidogenesis. However, the upstream regulators of FGF9 in ovine testosterone biosynthesis remain unknown. Long non-coding RNAs (lncRNAs) are crucial regulators of multiple biological functions that act by altering gene expression. In the present study, we analysed the role of LOC105611671, a lncRNA upstream of FGF9, in Hu sheep steroidogenesis. We found that LOC105611671 expression increased significantly in Hu sheep testes during sexual maturation (P<0.05). Moreover, levels of FGF9 and testosterone were decreased by LOC105611671 knockdown in Hu sheep Leydig cells (LCs). Results of transient transfection and luciferase assays revealed that FGF9 is a functional target gene of oar-miR-26a in ovine LCs. Further functional validation experiments revealed that LOC105611671 regulates testosterone biosynthesis by targeting oar-miR-26a. Overall, the present study describes the expression profile of LOC105611671 during sexual maturation and demonstrates that LOC105611671 modulates FGF9 expression by targeting oar-miR-26a to promote testis steroidogenesis in Hu sheep. Our research provides a new theoretical basis for genetic and molecular research on testosterone biosynthesis in sheep.

The oncogene Gankyrin is expressed in testicular cancer and contributes to cisplatin sensitivity in embryonal carcinoma cells

BackgroundTesticular germ cell cancer (TGCC) develops from pre-malignant germ neoplasia in situ (GCNIS) cells. GCNIS originates from fetal gonocytes (POU5F1+/MAGE-A4−), which fail to differentiate to pre-spermatogonia (POU5F1−/MAGE-A4+) and undergo malignant transformation. Gankyrin is an oncogene which has been shown to prevent POU5F1 degradation and specifically interact with MAGE-A4 in hepatocellular carcinoma (HCC) cells. We aimed to investigate the role of Gankyrin in progression from gonocyte to pre-invasive GCNIS and subsequent invasive TGCC.MethodsWe determined Gankyrin expression in human fetal testicular tissue (gestational weeks 9–20; n = 38), human adult testicular tissue with active spermatogenesis (n = 9), human testicular tissue with germ cell maturation delay (n = 4), testicular tissue from patients with pre-invasive GCNIS (n = 6), and invasive TGCC including seminoma (n = 6) and teratoma (n = 7). Functional analysis was performed in-vitro by siRNA knock-down of Gankyrin in the NTera2 cells (derived from embryonal carcinoma).ResultsGerm cell expression of Gankyrin was restricted to a sub-population of prespermatogonia in human fetal testes. Nuclear Gankyrin was also expressed in GCNIS cells of childhood and adult pre-invasive TGCC patients, and in GCNIS from seminoma and non-seminoma patients. Cytoplasmic expression was observed in seminoma tumour cells and NTera2 cells. Gankyrin knock-down in NTera2 cells resulted in an increase in apoptosis mediated via the TP53 pathway, whilst POU5F1 expression was unaffected. Furthermore, Gankyrin knock-down in NTera2 cells increased cisplatin sensitivity with an increase in cell death (13%, p < 0.05) following Gankyrin knock-down, when compared to cisplatin treatment alone, likely via BAX and FAS. Our results demonstrate that Gankyrin expression changes in germ cells during normal transition from gonocyte to prespermatogonia. In addition, changes in Gankyrin localisation are associated with progression of pre-invasive GCNIS to invasive TGCC. Furthermore, we found that Gankyrin is involved in the regulation of NTera2 cell survival and that a reduction in Gankyrin expression can modulate cisplatin sensitivity.ConclusionsThese results suggest that manipulation of Gankyrin expression may reduce the cisplatin dose required for the treatment of TGCC, with benefits in reducing dose-dependent side effects of chemotherapy. Further studies are required in order to assess the effects of modulating Gankyrin on GCNIS/TGCC using in vivo models.

Blood Testis Barrier and Somatic Cells Impairment in a Series of 35 Adult Klinefelter Syndrome Patients.

Klinefelter Syndrome (KS) is the most common genetic cause of infertility in men. Degeneration of the testicular tissue starts in utero and accelerates at puberty with hyalinisation of seminiferous tubules, spermatogonia apoptosis and germ cell maturation arrest. Therefore, fertility preservation in young KS boys has been proposed, although this measure is still debated due to insufficient knowledge of the pathophysiology of the disease. To better understand the underlying mechanisms of testicular failure and germ cell loss, we analysed functional and morphological alterations in the somatic compartment of KS testis, i.e., Sertoli cells, including the blood–testis barrier (BTB) and Leydig cells (LC). We compared three populations: 35 KS 47,XXY non-mosaic patients, 28 Sertoli-cell-only (SCO) syndrome patients and 9 patients with normal spermatogenesis. In KS patients the expression of BTB proteins connexin-43 and claudin-11 assessed with a semi-quantitative scoring system appeared significantly reduced with a disorganised pattern. A significant reduction in seminiferous tubules expressing androgen receptors (AR) was observed in KS compared to normal spermatogenesis controls. INSL3 expression, a marker of LC maturation, was also significantly reduced in KS compared to patients with normal spermatogenesis or SCO. Hence, the somatic compartment impairment in KS could be involved in degeneration of seminiferous tubules.

Central Precocious Puberty in two Boys with Prader-Willi Syndrome on Growth Hormone Treatment

Prader-Willi syndrome (PWS) is a rare genetic neuroendocrine disorder characterized by hypotonia, obesity, short stature, and mental retardation. Incomplete or delayed pubertal development as well as premature adrenarche are usually found in PWS, whereas central precocious puberty is rarely seen. This study reports the clinical, biochemical, and histologic findings in 2 boys with PWS who developed central precocious puberty. Both boys were started on growth hormone therapy during the first years of life according to the PWS indication. They had both bilateral cryptorchidism at birth and had orchidopexy in early childhood. Retrospective histologic analysis of testicular biopsies demonstrated largely normal tissue architecture and germ cell maturation, but severely decreased number of prespermatogonia in one of the patients. Both boys had premature adrenarche around the age of 6. Precocious puberty was diagnosed in both boys with enlargement of testicular volume (>3 mL), signs of virilization and a pubertal response to a gonadotropin-releasing hormone (GnRH) test and they were both treated with GnRH analog. The cases described here displayed typical characteristics for PWS, a considerable heterogeneity of the hypothalamic-pituitary function, as well as testicular histology. Central precocious puberty is extremely rare in PWS boys, but growth hormone treatment may play a role in the pubertal timing.

Dazl determines primordial follicle formation through the translational regulation of Tex14.

Deleted in azoospermia-like (DAZL) is a germ cell RNA-binding protein that is essential for entry and progression through meiosis. The phenotype of the Dazl knockout mouse has extensive germ cell loss because of incomplete meiosis. We have created a Dazl hypomorph model using short interfering RNA knockdown in mouse fetal ovary cultures, allowing investigation of Dazl function in germ cell maturation. Dazl hypomorph ovaries had a phenotype of impaired germ cell nest breakdown with a 66% reduction in total follicle number and an increase in the proportion of primordial follicles (PMFs), with smaller oocytes within these follicles. There was no significant early germ cell loss or meiotic delay. Immunostaining of intercellular bridge component testis-expressed protein (Tex)14 showed ∼59% reduction in foci number and size, without any change in Tex14 mRNA levels. TEX14 expression was also confirmed in the human fetal ovary across gestation. Using 3'UTR-luciferase reporter assays, translational regulation of TEX14 was demonstrated to be DAZL-dependant. Dazl is therefore essential for normal intercellular bridges within germ cell nests and their timely breakdown, with a major impact on subsequent assembly of PMFs.-Rosario, R., Crichton, J. H., Stewart, H. L., Childs, A. J., Adams, I. R., Anderson, R. A. Dazl determines primordial follicle formation through the translational regulation of Tex14.

Characterization of RNA binding proteins required for receptor-mediated endocytosis in C. elegans

Background and Hypothesis: Recently, a study conducted by the Hundley lab revealed that mutation of a specific RNA editing protein, ADR2, in the model organism C. elegans increased receptor-mediated endocytosis of yolk protein vit-2 and oocyte maturation. Of interest to us was what additional RNA binding proteins were involved in the expression of vit-2. To this end, RNA interference (RNAi) was used to knock out relevant RNA binding proteins. We hypothesized we would find candidates that both enhanced and reduced the expression of vit-2.&#x0D; Project Methods: Previously, adr-2(-) C. elegans strain expressing a yolk protein (vit-2) fused to GFP and a library of E. coli strains expressing RNAi directed to specific RNA binding proteins were engineered. vit-2:GFP reporter worms were grown on each RNAi bacterial strain. The COPAS Biosort, a large particle sorter which detects fluorescent intensity, was used to visualize and quantify vit-2 endocytosis.&#x0D; Results: Our screen yielded ten candidate proteins. These proteins had various functions and were expressed in different tissues, most commonly the nervous and reproductive systems. Remarkably, proteins gld-1 and puf-8 are negative regulators of oocyte fate determination and spermatogenesis respectively. Unlike the rest of our candidates, mutation of these two proteins did not have an observed effect on developmental progression but still caused significant increase in vit-2 expression.&#x0D; Conclusion and Potential Impact: This screen successfully gave direction to future studies of ADR-2 involvement in yolk protein uptake, oocyte maturation, and embryonic development. Two of our identified targets seem to be involved in reproductive processes. The remainder may be affecting vit-2 expression by altering development or receptor-mediated endocytosis. Our goal in the immediate future will be to compare loss of these target proteins in wildtype worms to loss in adr-2(-) worms. This would add to what is known about ADARs and life span and possibly demonstrate a role for ADARs in germ cell maturation.&#x0D;

Tissue-specific roles of p73 in development and homeostasis.

p73 (TP73) belongs to the p53 family of transcription factors. Its gene locus encodes two opposing types of isoforms, the transcriptionally active TAp73 class and the dominant-negative DNp73 class, which both play critical roles in development and homeostasis in an astonishingly diverse array of biological systems within specific tissues. While p73 has functions in cancer, this Review focuses on the non-oncogenic activities of p73. In the central and peripheral nervous system, both isoforms cooperate in complex ways to regulate neural stem cell survival, self-renewal and terminal differentiation. In airways, oviduct and to a lesser extent in brain ependyma, TAp73 is the master transcriptional regulator of multiciliogenesis, enabling fluid and germ cell transport across tissue surfaces. In male and female reproduction, TAp73 regulates gene networks that control cell-cell adhesion programs within germinal epithelium to enable germ cell maturation. Finally, p73 participates in the control of angiogenesis in development and cancer. While many open questions remain, we discuss here key findings that provide insight into the complex functions of this gene at the organismal, cellular and molecular level.

Dynamic paraspeckle component localisation during spermatogenesis.

Expression profiles and subcellular localisations of core Drosophila behaviour/human splicing (DBHS) proteins (PSPC1, SFPQ and NONO) and NEAT1, a long noncoding RNA (lncRNA), are investigated in developing and adult mouse testes. Core DBHS proteins are markers for the distinct subnuclear domain termed paraspeckles, while a long NEAT1 isoform scaffold facilitates paraspeckle nucleation. Paraspeckles contain many proteins (>40) and are broadly involved in RNA metabolism, including transcriptional regulation by protein sequestration, nuclear retention of A-to-I edited RNA transcripts to regulate translation and promoting survival during cellular stress. Immunohistochemistry reveals cell-specific profiles for core DBHS paraspeckle protein expression, indicating their functional diversity. PSPC1 is an androgen receptor co-activator, and it is detected in differentiating Sertoli cell nuclei from day 15 onwards, as they develop androgen responsiveness. PSPC1 is nuclear in the most mature male germ cell type present at each age, from foetal to adult life. In adult mouse testes, PSPC1 and SFPQ are present in Sertoli cells, spermatocytes and round spermatids, while the NEAT1 lncRNA appears in the punctate nuclear foci delineating paraspeckles only within Leydig cells. Identification of NEAT1 in the cytoplasm of spermatogonia and spermatocytes must reflect non-paraspeckle-related functions. NONO was absent from germ cells but nuclear in Sertoli cells. Reciprocal nuclear profiles of PSPC1 and γ-H2AX in spermatogenic cells suggest that each performs developmentally regulated roles in stress responses. These findings demonstrate paraspeckles and paraspeckle-related proteins contribute to diverse functions during testis development and spermatogenesis.

A Novel Dual Immunostain to Characterize Sloughed Cells in Testicular Biopsies for Infertility.

Evaluation of testicular biopsies from azoospermic men requires recognition of phases of germ cell maturation as organized architecturally within the seminiferous tubule, as well as distinguishing the inability to generate mature spermatozoa (germ cell aplasia or maturation arrest) from normal spermatogenesis, which may be associated with a reversible obstruction. While traditional fixatives (eg, Bouin solution) provide exquisite nuclear detail and preserve the architectural integrity of the seminiferous tubule, formalin fixation yields biopsies with relatively poor nuclear detail and frequent luminal sloughing of cells, making it difficult to assess sperm maturation. One clone of the anti-DOG1 antibody was recently found to be expressed in late (postspermatogonial) germ cells. We developed a dual stain including DOG1 and SF-1 to mark late germ cells and Sertoli cells, respectively, in both sloughed and intact cells. Consecutive testicular biopsies (N=28) from men with azoospermia were classified by hematoxylin and eosin morphology and stained with a dual SF-1 (Perseus)/DOG1 (Cell Marque) immunohistochemical stain. Histologic patterns included normal spermatogenesis (5 cases), hypospermatogenesis (5 cases), late maturation arrest (2 cases), Sertoli cell only pattern (15 cases), and extensive tubular hyalinization (1 case). Architectural disruption of seminiferous tubules with sloughing of cells into the lumens was noted in all biopsies, at least focally. SF-1 (nuclear) was expressed in sloughed Sertoli cells; DOG1 (cytoplasmic) in sloughed postspermatogonial germ cells (spermatocytes and spermatids). This resulted in two distinct immunophenotypes: SF-1(+)/DOG1(-) sloughed cells in cases with the Sertoli cell only pattern and SF-1(+)/DOG1(+) sloughed cells in all other histologic patterns (normal spermatogenesis, hypospermatogenesis, and maturation arrest). Because the rate of sperm retrieval is lower in men with the Sertoli cell only pattern, this immunohistochemical stain may assist pathologists in the proper interpretation of testicular biopsies, allowing better-informed decision making by patients and clinicians regarding the subsequent use of assisted reproductive technologies.

DNA cross-link repair safeguards genomic stability during premeiotic germ cell development.

Germline de novo mutations are the basis of evolutionary diversity but also of genetic disease. However, the molecular origin, mechanisms and timing of germline mutagenesis are not fully understood. Here, we define a fundamental role for DNA interstrand crosslink repair in the germline. This repair process is essential for primordial germ cell (PGC) maturation during embryonic development. Inactivation of crosslink repair leads to genetic instability that is restricted to PGCs within the genital ridge during a narrow temporal window. Having successfully activated the PGC transcriptional program, a potent quality control mechanism detects and drives damaged PGCs into apoptosis. These findings therefore define a source of DNA damage and the nature of the subsequent DNA repair response in germ cells, which ensures faithful transmission of the genome between generations.

Postnatal germ cell development in cryptorchid boys.

Cryptorchidism is associated with infertility in adulthood. Early orchiopexy is suggested to reduce the risk. Information is lacking on the potential link between infant germ cell maturation and the risk of future infertility. The objective of the study was to evaluate age-related germ cell development in cryptorchidism. Immunostaining for markers of germ cell development (octamer-binding transcription factor 3/4 [OCT3/4], placental alkaline phosphatase [PLAP], KIT proto-oncogene [C-KIT], podoplanin [D2-40], Lin-28 homolog A [LIN28], and G antigen 7 [GAGE-7]) was performed in testicular biopsies from 40 cryptorchid boys aged 4–35 months. Germ cell numbers and distributions were evaluated in cross sections of seminiferous tubules, with and without immunostaining. OCT3/4, D2-40, and LIN28 were generally expressed in the early stages of germ cell development, as shown by positive expression in germ cells in the central region of seminiferous tubules. In contrast, PLAP and GAGE-7 were expressed in both central and peripheral parts of the tubules in the early stages of development and expressed mainly in a peripheral position with advancing age. Germ cell maturation was delayed in this study population as compared with that observed in our previous study on germ cell markers in a healthy population. The number of GAGE-7-positive germ cells per tubular cross section obtained by immunostaining was significantly higher than that obtained by standard hematoxylin and eosin staining. Double immunostaining revealed heterogeneity in germ cell development in cryptorchid testes. These results shed light on the pathophysiology of germ cell development in boys with cryptorchidism.

Expression analysis and characterization of dmrt2 in Chinese tongue sole (Cynoglossus semilaevis)

Dmrt2 is a member of the dmrt gene family with a conserved zinc finger-like DNA-binding motif (DM domain). In the present study, CS-dmrt2 was cloned from the gonads of Chinese tongue sole (Cynoglossus semilaevis). The full-length cDNA of CS-dmrt2 is 2834 bp in length, with a 251 bp 5′-untranslated region (UTR), a 1086 bp 3′-UTR and a 1503 bp open reading frame (ORF) that encodes a 501-amino-acid peptide. qPCR revealed that CS-dmrt2 was mainly expressed in C. semilaevis testes. In situ hybridization (ISH) showed CS-dmrt2 expression throughout early gonadal development (36 days after hatching (dah) and 86 dah), but the expression was higher in male gonads than in female gonads. CS-dmrt2 mRNA was highly expressed in male germ cells. Comparison of methylation levels between females and males demonstrated hypo-methylated levels of the CS-dmrt2 promoter in the male gonads, which is consistent with the high mRNA expression. These results suggest that the CS-dmrt2 gene may play a functional role in gonadal differentiation/development and germ cell maturation in the testis.

Fertility Preservation in Pubertal and Pre-Pubertal Boys with Cancer.

Children diagnosed with cancer continue to have improved survival due to advances in effective treatment options. Increased attention is therefore now focused on quality of life issues once they are cured. Fertility preservation is of paramount concern since gonadotoxic treatments, especially radiation and chemotherapy, often impair future fertility. The importance of family counseling and having an informed discussion about the potential for treatment to impair fertility and the options available for fertility preservation is crucial. However, fertility preservation in prepubertal boys is challenging, but not impossible. Experimental methods are being investigated including cryopreservation of immature testicular tissue, xenografting, and in vitro germ cell maturation. Despite the success and relative ease of sperm banking, barriers exist and affect the number of patients offered treatment. Education and awareness of the possibilities can overcome these barriers. In this way we will continue to preserve young patients' future fertility and quality of life.

Combined Antioxidant Recovery Properties of Fruit Juice Extracts of Cucumber and Water Melon on Lipid Profile and Gonadal Steroid Levels of Cadmium Induced Testicular Damage on Male Albino Rats

The antioxidant protective effects of fruit juice of cucumber and watermelon on lipid profile of cadmium induced toxicity on male albino rats was investigated. Forty male rats were divided into eight groups. Group NC served as normal control group while group PC was positive control that was not treated but induced with cadmium. Groups I to VI received high dose and low dose of juice of Cucumber and Watermelon respectively. Excluding the normal control group, other groups were fed with lard 14 days before treatment commenced. Doses of 0.8 mg/kg-high dose and 0.4 mg/kg-low dose for cucumber and watermelon respectively. At the 4th and 6th week, biochemical parameters were assayed. Results revealed that the levels of total cholesterol, LDL, VLDL and triglyceride significantly (P˂0.05) were decreased compared to positive control but HDL was increased in treatment groups compared to positive control. Pretreatment with cucumber and watermelon juice indicated that total cholesterol, LDL, VLDL and triglyceride significantly (P˂0.05) were decreased compared to positive control but HDL was increased in treatment groups compared to positive control. The result also revealed an increase in testosterone levels in treated groups after 4 weeks of administration of whole extract of cucumber and watermelon when compared to their week 2 values. Testosterone level in positive control was also reduced significantly from 1.5±0.14 ng/ml to 0.46±0.31 ng/ml. Histological evaluation of the testes of normal control group revealed that the interstitium was intact with leydig cells present and maturing germ cells embedded in normal seminiferous tubules while the other groups that were induced with cadmium only showed morphology of testes with empty seminiferous tubules and consolidated interstitial spaces.

Genetic causes of nonsyndromic forms of azoospermia and severe oligozoospermia in infertility men

Spermatogenesis is a complex biological process of male germ cells differentiation and maturation involving more than 2300 genes in the human. Until recently, only a small part of them was investigated in severe pathozoospermia and male infertility. During the last years clinical utilisation of genomic data obtained by technologies, such as whole exome sequencing and array comparative genomic hybridization, has significantly expanded the opportunities of genomic and cytogenomic analysis, the detecting of multiple pathogenic mutations/variants and copy number variations related to different genetic syndromes and inherited disorders, including male and female infertility. This review provides an update on the genetics of male infertility, presenting a number of genes which related to non-syndromal male infertility as-sotiated with azoospermia and severe oligozoospermia.

Fertility concerns of the transgender patient.

Transgender individuals who undergo gender-affirming medical or surgical therapies are at risk for infertility. Suppression of puberty with gonadotropin-releasing hormone agonist analogs (GnRHa) in the pediatric transgender patient can pause the maturation of germ cells, and thus, affect fertility potential. Testosterone therapy in transgender men can suppress ovulation and alter ovarian histology, while estrogen therapy in transgender women can lead to impaired spermatogenesis and testicular atrophy. The effect of hormone therapy on fertility is potentially reversible, but the extent is unclear. Gender-affirming surgery (GAS) that includes hysterectomy and oophorectomy in transmen or orchiectomy in transwomen results in permanent sterility. It is recommended that clinicians counsel transgender patients on fertility preservation (FP) options prior to initiation of gender-affirming therapy. Transmen can choose to undergo cryopreservation of oocytes or embryos, which requires hormonal stimulation for egg retrieval. Uterus preservation allows transmen to gestate if desired. For transwomen, the option for FP is cryopreservation of sperm either through masturbation or testicular sperm extraction. Experimental and future options may include cryopreservation and in vitro maturation of ovarian or testicular tissue, which could provide prepubertal transgender youth an option for FP since they lack mature gametes. Successful uterus transplantation with subsequent live birth is a new medical breakthrough for cisgender women with uterus factor infertility. Although it has not yet been performed in transgender women, uterus transplantation is a potential solution for those who wish to get pregnant. The transgender population faces many barriers to care, such as provider discrimination, lack of information, legal barriers, scarcity of fertility centers, financial burden, and emotional cost. Further research is necessary to investigate the feasibility of experimental FP options, provide better evidence-based information to clinicians and transgender patients alike, and to improve access to and quality of reproductive services for the transgender population.