Clinical Utility of Sperm Function Tests in Predicting Male Fertility: A Systematic Review.

Routine semen analysis provides considerable information regarding sperm parameters; however, it is not solely adequate to predict male fertility potential. In the past two decades, several advance sperm function tests have been developed. The present systematic review intends to assess the clinical utility of available advance sperm function tests in predicting the male fertility potential. A systematic literature search was conducted as per PRISMA guidelines using PubMed, MEDLINE, Google Scholar, and Cochrane Library. Different keywords either singly or in combination were used to retrieve the relevant articles related to sperm function tests, male fertility, and pregnancy outcomes. A total of 5169 articles were obtained, out of which 110 meeting the selection criteria were included in this review. The majorly investigated sperm function tests are hypo-osmotic swelling test, acrosome reaction test, sperm capacitation test, hemizona binding assay, sperm DNA fragmentation test, seminal reactive oxygen species test, mitochondrial dysfunction tests, antisperm antibody test, nuclear chromatin de-condensation (NCD) test, etc. The different advance sperm function tests analyse different aspects of sperm function. Hence, any one test may not be helpful to appropriately predict the male fertility potential. Currently, the unavailability of high-quality clinical data, robust thresholds, complex protocols, high cost, etc., are the limiting factors and prohibiting current sperm function tests to reach the clinics. Further multi-centric research efforts are required to fulfil the existing lacunas and pave the way for these tests to be introduced into the clinics.

Dopamine receptor D2 regulates genes involved in germ cell movement and sperm motility in rat testes†.

The function of dopamine receptor D2 (D2R) is well associated with sperm motility; however, the physiological role of D2R present on testicular cells remains elusive. The aim of the present study is to delineate the function of testicular D2R. Serum dopamine levels were found to decrease with age, whereas testicular D2R expression increased. In rat testicular sections, D2R immunolabeling was observed in interstitial cells, spermatogonia, spermatocytes and mature elongated spermatids, whereas tyrosine hydroxylase immunolabeling was selectively detected in Leydig cells. In vitro seminiferous tubule culture following bromocriptine (D2R agonist) treatment resulted in decreased cAMP levels. Microarray identified 1077 differentially expressed genes (511 up-regulated, 566 down-regulated). The majority of differentially expressed genes were present in post-meiotic cells including early and late spermatids, and sperm. Gene ontology elucidated processes related to extra-cellular matrix to be enriched and was supported by differential expression of various collagens and laminins, thereby indicating a role of dopamine in extra-cellular matrix integrity and transport of spermatids across the seminiferous epithelium. Gene ontology and enrichment map also highlighted cell/sperm motility to be significantly enriched. Therefore, genes involved in sperm motility functions were further validated by RT-qPCR. Seven genes (Akap4, Ccnyl1, Iqcf1, Klc3, Prss55, Tbc1d21, Tl18) were significantly up-regulated, whereas four genes (Dnah1, Dnah5, Clxn, Fsip2) were significantly down-regulated by bromocriptine treatment. The bromocriptine-stimulated reduction in seminiferous tubule cyclic AMP and associated changes in spermatid gene expression suggests that dopamine regulates both spermatogenesis and spermiogenesis within the seminiferous epithelium, and spermatozoa motility following spermiation, as essential processes for fertility.

DNA methylation biomarkers to identify epigenetically abnormal spermatozoa in male partners from couples experiencing recurrent pregnancy loss.

Previously, we showed that DNA methylation defects in spermatozoa from male partners of couples undergoing recurrent pregnancy loss (RPL) could be a contributing paternal factor. In the present study, we aimed to determine whether the methylation levels of selected imprinted genes can be used as diagnostic markers to identify epigenetically abnormal spermatozoa sample in these cases. The methylation levels of selected imprinted genes in spermatozoa, which were previously found to be differentially methylated, were combined into a probability score (between 0-1) using multiple logistic regression. Different combinations of these genes were investigated using Receiver Operating Characteristic analysis, and the threshold values were experimentally validated in an independent cohort of 38 control and 45 RPL spermatozoa samples. Among the different combinations investigated, a combination of five imprinted genes comprising IGF2-H19 DMR, IG-DMR, ZAC, KvDMR, and PEG3 (AUC = 0.88) with a threshold value of 0.61 was selected with a specificity of 90.41% and sensitivity of 70%. The results from the validation study indicated that 97% of the control samples had probability scores below this threshold, whereas 40% of the RPL samples were above this threshold with a post-hoc power of 97.8%. Thus, this combination can correctly classify control samples and potentially identify epigenetically abnormal spermatozoa samples in the male partners of couples undergoing RPL. We propose that the combined DNA methylation levels of these imprinted genes can be used as a diagnostic tool to identify spermatozoa samples with epigenetic defects which could contribute to the pathophysiology of RPL and the couple could be counselled appropriately.

Preclinical safety assessment of red emissive gold nanocluster conjugated crumpled MXene nanosheets: a dynamic duo for image-guided photothermal therapy.

Red emissive gold nanoclusters have potential as biological fluorescent probes, but lack sufficient light-to-heat conversion efficiency for photothermal therapy (PTT). MXene nanomaterials, on the other hand, have shown promise in PTT due to their strong near-infrared absorption abilities, but their instability caused by restacking of the sheets can decrease their available surface area. One approach to address this issue is to design sheets with wrinkles or folds. However, the crumpled or 3D MXene materials reported in the literature are actually aggregates of multiple nanosheets rather than a single sheet that is folded. In this study, a modified method for crumpling a single MXene sheet and further conjugating it with red emissive gold nanoclusters and folic acid was developed. A detailed in vitro toxicity study was performed in various cell lines and cellular uptake in cancer cells was studied using AFM to understand its interaction at the nano-bio interface. The material also demonstrated excellent utility as a bioimaging and PTT agent in vitro, with its high fluorescence allowing bioimaging at a lower concentration of 12 µg mL-1 and a photothermal conversion efficiency of 43.51%. In vitro analyses of the cell death mechanisms induced by PTT were conducted through studies of apoptosis, cell proliferation, and ROS production. In vivo acute toxicity tests were conducted on male and female Wistar rats through oral and intravenous administration (20 mg kg-1 dose), and toxicity was evaluated using various measures including body weight, hematology, serum biochemistry, and H&E staining. The findings from these studies suggest that the MXene gold nanoconjugate could be useful in a range of biomedical applications, with no observed toxicity following either oral or intravenous administration.

Whole genome bisulfite sequencing of sperm reveals differentially methylated regions in male partners of idiopathic recurrent pregnancy loss cases.

OBJECTIVE: To study the genome wide alterations in sperm DNA methylation in male partners of idiopathic recurrent pregnancy loss (iRPL) cases and note regions as potential diagnostic markers. DESIGN: Case-control study and methylome analysis of human sperm. SETTING: Obstetrics and Gynaecology clinics. PATIENT(S): Control group consists of apparently healthy fertile men having fathered a child within the last 2 years (n = 39); and case group consists of male partners of iRPL cases having ≥2 consecutive 1st trimester pregnancy losses (n = 47). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Sperm DNA samples of controls and cases were selected for whole genome bisulfite sequencing analysis based on the previously set thresholds of global methylation levels and methylation levels of imprinted genes (KvDMR and ZAC). Whole genome bisulfite sequencing of selected sperm genomic DNA was performed to identify differentially methylated CpG sites of iRPL cases compared with fertile controls. Pathway analysis of all the differentially methylated genes was done by Database for Annotation, Visualization, and Integrated Discovery annotation tool and Kyoto Encyclopedia of Genes and Genomes tool. Differentially methylated CpGs within genes relevant to embryo and placenta development were selected to further validate their methylation levels in study population by pyrosequencing. RESULT(S): A total of 9497 differentially methylated CpGs with highest enrichment in intronic regions were obtained. In addition, 5352 differentially methylated regions and 2087 differentially methylated genes were noted. Signaling pathways involved in development were enriched on pathway analysis. Select CpGs within genes PPARG, KCNQ1, SETD2, and MAP3K4 showed distinct hypomethylated subpopulations within iRPL study population. CONCLUSION(S): Our study highlights the altered methylation landscape of iRPL sperm, and their possible implications in pathways of embryo and placental development. The CpG sites that are hypomethylated specifically in sperm of iRPL subpopulation can be further assessed as predictive biomarkers.

Varicocoele-associated male infertility: Cellular and molecular perspectives of pathophysiology.

BACKGROUND: Varicocoele is a common risk factor associated with reduced male fertility potential. The current understanding of varicocoele pathophysiology does not completely explain the clinical manifestation of infertility. The present treatment options such as antioxidant supplementation and varicocoelectomy only help ≈35% of men to achieve spontaneous pregnancy. OBJECTIVE: This review aims to summarize the available knowledge on cellular and molecular alterations implicated to varicocoele-associated male infertility and also highlights the new knowledge generated by "omics" technologies. MATERIALS AND METHODS: PubMed, MEDLINE, Cochrane and Google Scholar databases are searched using different combinations of keywords (varicocoele, infertile/fertile men with varicocoele, cellular changes, molecular mechanisms, proteome, epigenome, transcriptome and metabolome). A total of 229 relevant human and animal studies published till 2021 were included in this review. RESULTS: Current understanding advocates oxidative stress (OS) as a major contributory factor to varicocoele-associated male infertility. Excessive OS causes alteration in testicular microenvironment and sperm DNA fragmentation, which further contributes to infertility. Molecular and omics studies have identified several promising biomarkers such as AAMP, SPINT1, MKI67 (genetic markers), sperm quality and function related protein markers, global sperm DNA methylation level (epigenetic marker), Hspa2, Protamine, Gadd7, Dynlt1 and Beclin1 (mRNA markers), PRDX2, HSPA, APOA2, YKL40 (seminal protein markers), total choline and PHGDH (metabolic markers). DISCUSSION: Mature spermatozoa harbours a plethora of molecular information in form of proteome, epigenome and transcriptome, which could provide very important clues regarding pathophysiology of varicocoele-associated infertility. Recent molecular and omics studies in infertile men with varicocoele have identified several promising biomarkers. Upon further validation with larger and well-defined studies, some of these biomarkers could aid in varicocoele management. CONCLUSION: The present evidences suggest that inclusion of OS and sperm DNA fragmentation tests could be useful to the diagnostic workup for men with varicocoele. Furthermore, including precise molecular markers may assist in diagnostics and prognostics of varicocoele-associated male infertility.

Maternal cypermethrin exposure during perinatal period dysregulates gonadal steroidogenesis, gametogenesis and sperm epigenome in F1 rat offspring.

Evidences suggest that maternal exposure to endocrine disruptor insecticide cypermethrin (CYP) affects the reproductive functions of offspring. However, its molecular effects are not well researched. This study elucidates the effects of CYP on gonadal steroidogenesis, gametogenesis and sperm epigenome of perinatally exposed F1 rat offspring. CYP (1, 10, 25 mg/kg bw/day) and Diethylestilbestrol (10 µg/kg bw/day; positive control) were gavaged once daily to pregnant dams from gestational day 6 to postnatal day 21 and their effects were assessed in F1 adults. Dysregulated steroidogenesis was observed in F1 testis; expression of Star, Cyp11a1 and Cyp19a1 were upregulated and Hsd3b1, Cyp17a1, Hsd17b3 downregulated. Expression of spermatogenesis cell-type markers Pcna, Plzf, Sohlh2 were upregulated and Ccna1, Sycp1, Ccnb1, Acrv, Amh downregulated. Upon epigenetic investigations of F1 spermatozoa; global hypermethylation, H19 DMR hypomethylation and increased expression of testicular Dnmts were observed. In F1 ovaries, expression of all studied steroidogenesis genes and oogenesis markers such as Amh, Gdf9, Ccnb1 and Pcna were altered. Amh was downregulated and Gdf9 was found to be upregulated. Overall, perinatal CYP exposure hampers molecular control of steroidogenesis and gametogenesis processes along with sperm epigenome in CYP exposed F1 offspring.

pH Stress Mediated Alteration in Protein Composition and Reduction in Cytotoxic Potential of Gardnerella vaginalis Membrane Vesicles.

The vagina of healthy women is predominantly colonized by lactobacilli but it also harbors a limited proportion of certain anaerobes such as Gardnerella vaginalis. An increase in G. vaginalis along with other anaerobes on account of perturbation in the vaginal microbiota is associated with bacterial vaginosis (BV). Although strategies adopted by G. vaginalis for survival and pathogenesis in a conducive environment (i.e., high vaginal pH, characteristic of BV) have been previously studied, the approaches potentially employed for adaptation to the low pH of the healthy vagina are unknown. In the present study, we investigated the effect of acidic stress on the modulation of the production and function of membrane vesicles (MVs) of G. vaginalis. pH stress led to a distortion of the bacterial cell morphology as well as an altered biogenesis of MVs, as revealed by transmission electron microscopy (TEM). Both qualitative and quantitative differences in protein content of MVs produced in response to pH stress were observed by flow cytometry. A significant change in the protein composition characterized by presence of chaperones despite a reduction in number of proteins was also noted in the stress induced MVs. Further, these changes were also reflected in the reduced cytotoxic potential toward vaginal epithelial cells. Although, these findings need to be validated in the in vivo settings, the modulation of G. vaginalis MV biogenesis, composition and function appears to reflect the exposure to acidic conditions prevailing in the host vaginal mileu in the absence of vaginal infection.

Collapsed mitochondrial cristae in goat spermatozoa due to mercury result in lethality and compromised motility along with altered kinematic patterns.

Earlier we have reported mercury-induced alterations in functional dynamics of buck spermatozoa through free radicals-mediated oxidative stress and spontaneous acrosome reaction. Based on our earlier findings, we aimed to investigate the effect of mercury exposure on motility, kinematic patterns, DNA damage, apoptosis and ultra-structural alterations in goat spermatozoa following in vitro exposure to different concentrations (0.031-1.25 µg/ml) of mercuric chloride for 15 min and 3 h. Following exposure of sperm cells to 0.031 µg/ml of mercuric chloride for 3 h, livability and motility of sperms was significantly reduced along with altered kinematic patterns, significant increase in per cent necrotic sperm cells and number of cells showing DNA damage; and this effect was dose- and time-dependent. Contrary to up-regulation of Bax gene after 3 h in control group, there was significant increase in expression of Bcl-2 in mercury-treated groups. Transmission electron microscopy studies revealed rifts and nicks in plasma and acrosomal membrane, mitochondrial sheath, and collapsed mitochondria with loss of helical organization of mitochondria in the middle piece of spermatozoa. Our findings evidently suggest that mercury induces necrosis instead of apoptosis and targets the membrane, acrosome, mid piece of sperms; and the damage to mitochondria seems to be responsible for alterations in functional and kinematic attributes of spermatozoa.

Causes and consequences of sperm mitochondrial dysfunction.

Mitochondria have multiple functions, including synthesis of adenine triphosphate, production of reactive oxygen species, calcium signalling, thermogenesis and apoptosis. Mitochondria have a significant contribution in regulating the various physiological aspects of reproductive function, from spermatogenesis up to fertilisation. Mitochondrial functionality and intact mitochondrial membrane potential are a pre-requisite for sperm motility, hyperactivation, capacitation, acrosin activity, acrosome reaction and DNA integrity. Optimal mitochondrial activity is therefore crucial for human sperm function and semen quality. However, the precise role of mitochondria in spermatozoa remains to be fully explored. Defects in sperm mitochondrial function severely impair the maintenance of energy production required for sperm motility and may be an underlying cause of asthenozoospermia. Sperm mtDNA is susceptible to oxidative damage and mutations that could compromise sperm function leading to infertility. Males with abnormal semen parameters have increased mtDNA copy number and reduced mtDNA integrity. This review discusses the role of mitochondria in sperm function, along with the causes and impact of its dysfunction on male fertility. Greater understanding of sperm mitochondrial function and its correlation with sperm quality could provide further insights into their contribution in the assessment of the infertile male.

Characterization of Gardnerella vaginalis membrane vesicles reveals a role in inducing cytotoxicity in vaginal epithelial cells.

Bacterial vaginosis (BV) is a common polymicrobial infection affecting women in the reproductive age and is associated with adverse obstetric and gynaecological outcomes. Gardnerella vaginalis is the most virulent anaerobic bacterial species predominantly associated with BV. However, a clear understanding of the mechanisms by which it contributes to the pathogenesis and persistence of BV is lacking. In this report, we demonstrate for the first time, the isolation of membrane vesicles (MVs) from G. vaginalis ATCC 14019. These MVs are approximately 120-260 nm in diameter. Proteomic characterization of the MVs by LC-MS/MS led to the identification of 417 proteins, including proteins involved in cellular metabolism as well as molecular chaperones and certain virulence factors. Immunoblot analysis of the MVs confirmed the presence of vaginolysin, the most well-characterized virulence factor of G. vaginalis. The exposure of the vaginal epithelial cells, VK2/E6E7 to the G. vaginalis MVs resulted in the internalization of the MVs. The MVs induced cytotoxicity and an increase in the levels of the pro-inflammatory cytokine, IL-8 in VK2 cells as well lysis of erythrocytes. The results of the study indicate that G. vaginalis MVs may be involved in the delivery of cytotoxic proteins and other virulence factors to the host cells and could thereby contribute towards enhancing the cellular damage associated with pathogenesis of BV.

Cypermethrin exposure during perinatal period affects fetal development and impairs reproductive functions of F1 female rats.

Cypermethrin (CYP) is a ubiquitously present synthetic pyrethroid insecticide. It has endocrine disrupting activities which may adversely affect reproductive development and functions of offspring if exposed during critical developmental period. The present study was undertaken to delineate the effects of CYP exposure in pregnant female rats during perinatal period on the sexual maturation, hormonal regulation, reproductive development and fertility of F1 female offspring and its molecular mechanism of action. Pregnant rats (F0) were gavaged daily with 0, 1, 10, 25 mg/kg bw/day CYP and 10 µg/kg bw/day Diethylstilbestrol (DES; positive control) from gestation day (GD) 6 to postnatal day (PND) 21. The reproductive development and function parameters were evaluated at PND 45 and 75. Reduced body weight, delayed vaginal opening, and disrupted estrous cyclicity were observed at 25 mg/kg CYP dose. CYP exposure significantly affected the reproductive organ development and their functions at all doses. Significant alterations in ovarian and uterine histology such as luteinization, reduction of primordial follicular reserves, presence of multi-oocyte follicles and thin degenerative luminal and glandular uterine epithelium were observed at adulthood. Altered circulatory steroid hormone levels and expression of ovarian and uterine steroid hormone receptors were observed at PND 75 in the F1 female offspring. Expression of HOXA10 and α-SMA which are important for uterine integrity and functions, were found to be altered at PND 75. Increased pre-implantation loss (PIL%), post-implantation loss (POL%), and reduced litter size in F1 females when cohabitated with unexposed fertile male rats were observed. Overall, perinatal exposure of pregnant rats to CYP led to significant long lasting effects on the reproductive functions of F1 female offspring. The adverse effects were passed on to F2 generation via female germ line and posed developmental anomalies. The present finding necessitates additional molecular studies to understand its trans-generational mechanism of action via female germline.

Is there plagiarism in the most influential publications in the field of andrology?

Plagiarism is a common form of academic misconduct that extensively jeopardises the quality of scientific publication. The purpose of this study is to determine the extent of plagiarism in the most influential andrology articles. A total of 77 highly cited andrology articles were analysed for their similarity index using iThenticate and Turnitin. The articles were categorised based on the year (before and on/after 2000) and type of publication (review and research articles), and the similarity indices were compared. Furthermore, the analysed articles were categorised based on the level of similarity using an arbitrary similarity index range (low: ≤10, moderate: 11-20, high: 21-50 and very high: >50) and average incidence rate (%) was determined. Our analysis revealed a higher percentage of the similarity indices for reviews than research articles. We noticed a higher similarity index for articles published on/after 2000 than those published before. The majority of the influential articles in the field of andrology showed a low similarity index, while some articles exhibited moderate to high levels of similarity. These findings support the need for the development of similarity index guidelines as a major pre-requisite for establishing a more transparent and efficient system to address plagiarism in scientific publications.

Heterogeneity in pneumolysin expression governs the fate of Streptococcus pneumoniae during blood-brain barrier trafficking.

Outcome of host-pathogen encounter is determined by the complex interplay between protective bacterial and host defense strategies. This complexity further amplifies with the existence of cell-to-cell phenotypic heterogeneity in pathogens which remains largely unexplored. In this study, we illustrated that heterogeneous expression of pneumolysin (Ply), a pore-forming toxin of the meningeal pathogen, S. pneumoniae (SPN) gives rise to stochastically different bacterial subpopulations with variable fate during passage across blood-brain barrier (BBB). We demonstrate that Ply mediated damage to pneumococcus containing vacuolar (PCV) membrane leads to recruitment of cytosolic "eat-me" signals, galectin-8 and ubiquitin, targeting SPN for autophagic clearance. However, a majority of high Ply producing subset extensively damages autophagosomes leading to pneumococcal escape into cytosol and efficient clearance by host ubiquitination machinery. Interestingly, a low Ply producing subset halts autophagosomal maturation and evades all intracellular defense mechanisms, promoting its prolonged survival and successful transcytosis across BBB, both in vitro and in vivo. Ply therefore acts as both, sword and shield implying that its smart regulation ensures optimal disease manifestation. Our elucidation of heterogeneity in Ply expression leading to disparate infection outcomes attempts to resolve the dubious role of Ply in pneumococcal pathogenesis.

Estradiol: A Steroid with Multiple Facets.

Seventy-five glorious years have passed since estradiol was discovered by Edward Doisy. From discovery in the ovaries to delineation of diverse physiological effects, research on estrogens has covered a lot of ground. Estrogen receptors that mediate estrogenic effects, have been detected not only in reproductive organs, but also in other body organs. Estrogen receptors function either as conventional transcription factors or as rapid signal transducers. These different modes of action are opted by estrogens to elicit an array of reproductive and non-reproductive functions. It is well established that estrogens promote cell proliferation in various tissues and hence are also linked to carcinogenesis. Anti-estrogens are being used as adjunct therapies for cancers since several years. On the other hand, estrogen-based strategies are used to alleviate adverse effects of menopause. Apart from estrogens synthesized in various organs, exposure to environmental estrogens can also impact physiology. Thus, too much or too less of estrogens can tip the balance and lead to unfavorable consequences. Multiple estrogen receptors with their tissue- or cell type-specific expression eliciting dose-dependent effects make it perplexing to 'unify' estrogenic actions in diverse tissues/organs. This warrants more research on estrogen-mediated effects and their regulation in somatic and reproductive tissues. This review presents physiological and pathological aspects of estrogens thus highlighting the good, bad, and ugly facets of estrogens.

Bisphenol A-induced ultrastructural changes in the testes of common marmoset.

BACKGROUND & OBJECTIVES: Bisphenol A (BPA) is an endocrine disruptor that is widely used in the manufacture of polycarbonate plastics, epoxy resins and dental sealants. It is known to have adverse effects on spermatogenesis in rodents. This study was aimed to evaluate the effects of BPA in adult common marmoset owing to its similarities with human spermatogenesis. METHODS: Sixteen marmosets were divided into four groups (n=4 per group) and given oral doses of BPA (2.5, 12.5 and 25 µg/kg BW/day) for 70 days to cover two spermatogenic cycles, and the control group received only vehicle (honey). Testes were processed for histological and transmission electron microscopy studies. RESULTS: Histology of the testis showed sloughing of germ cells into the lumen, increase in interstitial space and vacuolation of Sertoli cell cytoplasm. Ultrastructural analysis of the testis revealed several degenerative effects on the basement membrane, Sertoli cells, Leydig cells and other developing germ cells in the 12.5 and 25 µg/kg BW/day groups as compared to control. INTERPRETATION & CONCLUSIONS: The observed ultrastructural changes caused by BPA in testicular morphology might be indicative of a perturbed sperm production. Considering the genetic and spermatogenic similarities of common marmoset (Callithrix jacchus) and humans, the study findings are of significance. Further studies are, however, needed to elucidate the mechanism of action.

Perinatal exposure of pregnant rats to cypermethrin delays testicular descent, impairs fertility in F1 male progeny leading to developmental defects in F2 generation.

Cypermethrin (CYP) is a widely used synthetic pyrethroid insecticide and regarded as a potential endocrine disruptor. CYP exposure may pose a great risk to human health including adverse effect on their reproductive functions. This study aimed to delineate the effects of perinatal exposure of rats to CYP on the sexual maturation and fertility of F1 male progeny. Pregnant rats (F0) were gavaged daily with CYP (0, 1, 10, 25 mg/kg BW/day) and Diethylestilbestrol (DES, 10 µg/kg BW/day), as positive control from gestation day 6 to postnatal day 21. The effects of CYP on body weight gain and reproductive functions were evaluated at the Juvenile (PND 22), peri-pubertal (PND 45) and adult (PND 75) stages of development. A significant delay in the age of testicular descent and prepuce separation was observed at 1 and 25 mg/kg doses of CYP. At the same dose level, reproductive organ development and their functions were also affected. A significant alteration in testicular histology, expression of steroid hormone receptors, and circulatory steroid hormones was observed throughout development. Reduced sperm count and motility were observed at PND 75 leading to subfertility and reduced litter size. These adult male rats when cohabitated with unexposed normal cycling females, the F2 fetuses exhibited developmental defects. Taken together, CYP perinatal exposure caused significant long lasting effects of the reproductive functions of F1 generation male rats, which were vertically transmitted to F2 generation leading to developmental defects. The mechanism of transgenerational effects needs to be explored in details.

Embryonic Zebrafish Model - A Well-Established Method for Rapidly Assessing the Toxicity of Homeopathic Drugs: - Toxicity Evaluation of Homeopathic Drugs Using Zebrafish Embryo Model.

OBJECTIVES: Advancements in nanotechnology have led to nanoparticle (NP) use in various fields of medicine. Although the potential of NPs is promising, the lack of documented evidence on the toxicological effects of NPs is concerning. A few studies have documented that homeopathy uses NPs. Unfortunately, very few sound scientific studies have explored the toxic effects of homeopathic drugs. Citing this lack of high-quality scientific evidence, regulatory agencies have been reluctant to endorse homeopathic treatment as an alternative or adjunct treatment. This study aimed to enhance our insight into the impact of commercially-available homeopathic drugs, to study the presence of NPs in those drugs and any deleterious effects they might have, and to determine the distribution pattern of NPs in zebrafish embryos (Danio rerio). METHODS: Homeopathic dilutions were studied using high-resolution transmission electron microscopy with selected area electron diffraction (SAED). For the toxicity assessment on Zebrafish, embryos were exposed to a test solution from 4 - 6 hours post-fertilization, and embryos/larvae were assessed up to 5 days post-fertilization (dpf) for viability and morphology. Toxicity was recorded in terms of mortality, hatching delay, phenotypic defects and metal accumulation. Around 5 dpf was found to be the optimum developmental stage for evaluation. RESULTS: The present study aimed to conclusively prove the presence of NPs in all high dilutions of homeopathic drugs. Embryonic zebrafish were exposed to three homeopathic drugs with two potencies (30CH, 200CH) during early embryogenesis. The resulting morphological and cellular responses were observed. Exposure to these potencies produced no visibly significant malformations, pericardial edema, and mortality and no necrotic and apoptotic cellular death. CONCLUSION: Our findings clearly demonstrate that no toxic effects were observed for these three homeopathic drugs at the potencies and exposure times used in this study. The embryonic zebrafish model is recommended as a well-established method for rapidly assessing the toxicity of homeopathic drugs.

ZapC promotes assembly and stability of FtsZ filaments by binding at a different site on FtsZ than ZipA.

ZapC, a component of the divisome in Escherichia coli, is known to co-localize with FtsZ at the mid-cell position. A deletion or an overexpression of ZapC has been found to induce elongation of bacterial cells implying a role of ZapC in the cell division. ZapC has also been shown to enhance the assembly of purified FtsZ. In this study, ZapC was found to prevent the dilution-induced disassembly of preformed FtsZ polymers and to decorate FtsZ protofilaments along the length. ZapC interacted with FtsZ with a dissociation constant of 30±7nM. Salt had no discernable effect on the binding of ZapC to FtsZ; however, bis-ANS inhibited the binding of ZapC to FtsZ suggesting that the interaction was predominantly hydrophobic in nature. Several of the positive regulators of FtsZ assembly including ZipA are shown to bind FtsZ at the C-terminal tail of FtsZ. Using a 12-residue C-terminal tail peptide (LDIPAFLRKQAD) of FtsZ and a C-terminal tail truncated FtsZ construct, we provided data suggesting that ZapC does not bind at the C-terminal tail of FtsZ. The results indicated that ZapC and ZipA, two functionally similar proteins of the divisome complex, regulate FtsZ assembly through different sites of action on FtsZ.

Histopathological and ultra structural effects of nanoparticles on rat testis following 90 days (Chronic study) of repeated oral administration.

BACKGROUND: Nanoparticles (Ag NPs) have recently received much attention for their possible applications in biotechnology and biomedical. However, little is known about the toxicity in reproductive organs of animal model following exposure to nanoparticles. OBJECTIVE: This study therefore, tried to examine the effects of nanoparticles with a diameter range of 5-20 nm on the histology of the testis of wistar rats and correlate it with Transmission Electron Microscopy results. MATERIALS AND METHODS: Sixteen wistar rats were randomly divided into two groups of 8 rats each. Each group received the following via gavage technique for 90 days: Control Group (Group-1)-tap water; Experimental group (Group 2) - nanoparticles (20ug/kg/day). After ninety days (chronic study), rats were sacrificed and testis tissues was processed for histology and transmission electron microscopic study. RESULTS: There was significant difference between the observations of group-1 and group 2. The changes observed in the testis were disarray of the spermatogenic cells and disorientation of the testis. These changes were observed to have been disappearing from normal histological features. Detailed structural damages were observed with TEM analysis, such as depletion of germ cells, germinal cells necrosis, especially in spermatogonia and Leydig cells had an abnormal fibroblast-like appearance, abnormal space between neighboring sertoli cells, mitochondria, lost cristae and vacuolated (none energized) with those animals exposed to nanoparticles. CONCLUSION: It seems that nanoparticles have acute and significant effects on spermatogenesis and number of spermatogenic cells. More experimental investigations are necessary to elucidate better conclusion regarding the safety of nanoparticles on male reproduction system.