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Frontiers in Cell and Developmental... 2023Recessive mutation of the X-linked gene, (), causes familial ciliopathy. PIH1D3 deficiency is associated with the defects of dynein arms in cilia, but how PIH1D3...
Recessive mutation of the X-linked gene, (), causes familial ciliopathy. PIH1D3 deficiency is associated with the defects of dynein arms in cilia, but how PIH1D3 specifically affects the structure and function of dynein arms is not understood yet. To gain insights into the underlying mechanisms of the disease, it is crucial to create a reliable animal model. In humans, rats, and mice, one copy of the gene is located on the X chromosome. Interestingly, mice have an additional, intronless copy of the gene on chromosome 1. To develop an accurate disease model, it is best to manipulate the X-linked gene, which contains essential regulatory sequences within the introns for precise gene expression. This study aimed to develop a tailored rat model for PIH1D3-associated ciliopathy with the ultimate goal of uncovering the intricate molecular mechanisms responsible for ciliary defects in the disease. Novel Pih1d3-knockout (KO) rats were created by using TALEN-mediated non-homologous DNA recombination within fertilized rat eggs and, subsequently, underwent a comprehensive characterization through a battery of behavioral and pathological assays. A series of biochemical and histological analyses were conducted to elucidate the identity of protein partners that interact with PIH1D3, thus shedding light on the intricate molecular mechanisms involved in this context. PIH1D3-KO rats reproduced the cardinal features of ciliopathy including situs inversus, defects in spermatocyte survival and mucociliary clearance, and perinatal hydrocephalus. We revealed the novel function of PIH1D3 in cerebrospinal fluid circulation and elucidated the mechanism by which PIH1D3 deficiency caused communicating hydrocephalus. PIH1D3 interacted with the proteins required for the pre-assembly and uploading of outer (ODA) and inner dynein arms (IDA), regulating the integrity of dynein arm structure and function in cilia. PIH1D3-KO rats faithfully reproduced the cardinal features of ciliopathy associated with PIH1D3 deficiency. PIH1D3 interacted with the proteins responsible for the pre-assembly and uploading of dynein arms in cilia, and its deficiency led to dysfunctional cilia and, thus, to ciliopathy by affecting the pre-assembly and uploading of dynein arms. The resultant rat model is a valuable tool for the mechanistic study of PIH1D3-caused diseases.
PubMed: 37900281
DOI: 10.3389/fcell.2023.1282787 -
Ecotoxicology and Environmental Safety Sep 2023China has the world's largest reserves of rare earth elements (REEs), but widespread mining and application of REEs has led to an increased risk of potential pollution....
China has the world's largest reserves of rare earth elements (REEs), but widespread mining and application of REEs has led to an increased risk of potential pollution. Yttrium (Y), the first heavy REEs to be discovered, poses a substantial threat to human health. Unfortunately, little attention has been given to the impact of Y on human reproductive health. In this study, we investigated the toxic effects of YCl on mouse testes and four types of testicular cells, including Sertoli, Leydig, spermatogonial and spermatocyte cells. The results showed that YCl exposure causes substantial damage to mouse testes and induces apoptosis and autophagy, but not pyroptosis or necrosis, in testicular cells. Genome-wide gene expression analysis revealed that YCl induced significant changes in gene expression, with Ca and mitochondria-related genes being the most significantly altered. Mechanistically, YCl exposure induced mitochondrial dysfunction in testicular cells, triggering the overproduction of reactive oxygen species (ROS) by impairing the Nrf2 pathway, regulating downstream Ho-1 target protein expression, and increasing Ca levels to activate the CamkII/Ampk signaling pathway. Blocking ROS production or Ca signaling significantly attenuates apoptosis and autophagy, while supplementation with Ca reverses the suppression of apoptosis and autophagy by ROS blockade in testicular cells. Notably, apoptosis and autophagy induced by YCl treatment are independent of each other. Thus, our study suggests that YCl may impair the antioxidant stress signaling pathway and activate the calcium pathway through the ROS-Ca axis, which promotes testicular cell apoptosis and autophagy independently, thus inducing testicular damage and impairing male reproductive function.
Topics: Humans; Animals; Mice; Male; Yttrium; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Reactive Oxygen Species; Metals, Rare Earth; Apoptosis; Autophagy; DNA, Mitochondrial; Genitalia, Male
PubMed: 37480693
DOI: 10.1016/j.ecoenv.2023.115262 -
Animals : An Open Access Journal From... May 2024We identified Wdr17 as a highly expressed gene in pachytene spermatocytes by transcriptomic analysis of mouse testis. Germ cell-deficient infertile mouse models had...
We identified Wdr17 as a highly expressed gene in pachytene spermatocytes by transcriptomic analysis of mouse testis. Germ cell-deficient infertile mouse models had significantly reduced Wdr17 expression. We performed gene interference and overexpression in the mouse spermatocyte cell line GC-2spd(ts) and investigated how Wdr17 affects spermatocyte growth and development. Our results showed that Wdr17 suppression significantly decreased cell growth rate and increased cell apoptosis in GC-2spd(ts) cells. Wdr17 suppression also arrested the cell cycle at the G1 phase. On the contrary, Wdr17 overexpression significantly promoted cell proliferation and inhibited cell apoptosis in GC-2spd(ts) cells. More cells were enriched at the S stage with a concomitant reduction of cells at the G1 stage. Wdr17 promotes mouse spermatocyte proliferation by advancing cell cycle progression and inhibiting cell apoptosis, indicating its potential role in regulating spermatogenesis in the mouse.
PubMed: 38791636
DOI: 10.3390/ani14101418 -
Frontiers in Bioscience (Landmark... Jul 2023Hydroxychloroquine (HCQ) toxicity can adversely affect vital organs, cause pathologic ocular damage, and can have direct cardiovascular effects. This study aims to...
BACKGROUND
Hydroxychloroquine (HCQ) toxicity can adversely affect vital organs, cause pathologic ocular damage, and can have direct cardiovascular effects. This study aims to identify the biochemical, hematological, and histological alterations of the vital organs associated with the effects of HCQ.
METHODS
Male albino rats were exposed to the equivalent of HCQ therapeutic doses given to human patients being affected by malaria, lupus erythematosus, and COVID-19. The animal blood samples were subjected to hematological analysis, biochemical analysis, liver function tests, kidney function tests, and cardiac biomarkers. Liver, kidney, heart, spleen, and testis biopsies were subjected to histological examination.
RESULTS
HCQ significantly lowered the values of erythrocytes, hemoglobin, hematocrit, platelets, leucocytes, and lymphocytes but significantly increased the values of aspartate aminotransferase (AST), alanine aminotransferase (ALT), amylase, alkaline phosphatase, lactate dehydrogenase, cholesterol, and chlorine ions. The renal tissues of HCQ-treated animals demonstrated glomerular fragmentation, partial atrophy degeneration, renal tubules hydropic degeneration, hyaline cast formation, and interstitial edema formation. Additionally, the heart exhibited myofiber necrosis, myolysis, wavy appearance, disorganization, and disarray. The testicular tissues also demonstrated spermatocyte degeneration, spermatogenic cell sloughing, testicular interstitial edema, and occasional spermatogenic arrest. Additionally, the spleen showed a decrease in the number and size of the white pulp follicles, a decrease in the number of apoptotic activity, and a decline in the number of T-rich cells. However, the red pulp demonstrated a diffuse decline in B rich-lymphocytes and macrophages. The liver was also the least affected but showed Kupffer cell hyperplasia and occasional hepatocyte dysplasia.
CONCLUSIONS
The results indicate that chronic exposure to HCQ could alter the structures and functions of the vital organs.
Topics: Rats; Animals; Humans; Male; Hydroxychloroquine; COVID-19; COVID-19 Drug Treatment; Liver; Necrosis
PubMed: 37525906
DOI: 10.31083/j.fbl2807137 -
Journal of Clinical Medicine Feb 2024Gender-affirming hormone therapy (GAHT) is an important component in the process of transitioning for many transgender and gender-diverse (TGD) individuals. Multiple... (Review)
Review
Gender-affirming hormone therapy (GAHT) is an important component in the process of transitioning for many transgender and gender-diverse (TGD) individuals. Multiple medical organizations recommend fertility preservation counseling prior to initiation of GAHT; however, there remains little high-quality data regarding the impact of GAHT on fertility and reproductive function. A PubMed literature review was performed using Boolean search operators linking keywords or phrases such as "mouse", "rat", "primate", "animal model", "transgender", "gender", "estrogen", "testosterone", "fertility", and "fertility preservation". Recent research has produced a number of animal models of GAHT that utilize similar hormonal regimens and produce similar phenotypic results to those used and observed in human patients. Specific to testosterone(T)-containing GAHT, animals demonstrate loss of menstrual cyclicity with therapy, resumption of menses on cessation of therapy, suppression of gonadotropin levels, and physical changes such as clitoromegaly. Models mimicking GAHT for transmasculine individuals in the peripubertal period demonstrate that pretreatment with GnRHa therapy does not modify the effects of subsequent T administration, which were similar to those described in adult models. Both models suggest promising potential for future fertility with cessation of T. With estradiol (E)-containing GAHT, animals exhibit decreased size of testicles, epididymis, and seminal vesicles, as well as ongoing production of spermatocytes, and seminiferous tubule vacuolization. Given the ethical challenges of conducting human studies in this area, high-fidelity animal models represent a promising opportunity for investigation and could eventually transform clinical counseling about the necessity of fertility preservation. Future studies should better delineate the interactions (if any exist) between treatment attributes such as dosing and duration with the extent of reversibility of reproductive perturbations. The development of models of peripubertal feminizing GAHT is an additional area for future work.
PubMed: 38398495
DOI: 10.3390/jcm13041183 -
BioRxiv : the Preprint Server For... Nov 2023Meiotic sex chromosome inactivation (MSCI) is a critical feature of meiotic prophase I progression in males. While the ATR kinase and its activator TOPBP1 are key...
Meiotic sex chromosome inactivation (MSCI) is a critical feature of meiotic prophase I progression in males. While the ATR kinase and its activator TOPBP1 are key drivers of MSCI within the specialized sex body (SB) domain of the nucleus, how they promote silencing remains unclear given their multifaceted meiotic functions that also include DNA repair, chromosome synapsis and SB formation. Here we report a novel mutant mouse harboring mutations in the TOPBP1-BRCT5 domain. males are infertile, with impaired MSCI despite displaying grossly normal events of early prophase I, including synapsis and SB formation. Specific ATR-dependent events are disrupted including phosphorylation and localization of the RNA:DNA helicase Senataxin. spermatocytes initiate, but cannot maintain ongoing, MSCI. These findings reveal a non-canonical role for the ATR-TOPBP1 signaling axis in MSCI dynamics at advanced stages in pachynema and establish the first mouse mutant that separates ATR signaling and MSCI from SB formation.
PubMed: 37398453
DOI: 10.1101/2023.05.31.543071 -
Cell Proliferation May 2024The mammalian Pre-B cell leukaemia transcription factors 1-4 (PBX1-4) constitutes the PBC class of the homeodomain (HD)-containing proteins, which play important roles...
The mammalian Pre-B cell leukaemia transcription factors 1-4 (PBX1-4) constitutes the PBC class of the homeodomain (HD)-containing proteins, which play important roles in diverse developmental processes. The functions and the underlying molecular mechanisms of PBX1-3 but not PBX4 have been extensively studied, and they have been reported to direct essential morphogenetic processes and organogenesis. In the present study, we generated knockin mice of FLAG-tagged PBX4 and the Pbx4 knockout (KO) mice and carried out in-depth characterisation of PBX4 expression and function. PBX4 was initially detected only in the testis among several organs of the adult mice and was expressed in spermatocytes and spermatids. However, no abnormality in spermatogenesis, but growth retardation and premature death after birth were observed in most adult Pbx4 KO mice. These animals were inactive and had shorter hindlimbs and lower numbers of reticulocytes and lymphocytes, probably caused by abnormalities at earlier developmental stages. Pbx4 mRNAs were indeed detected in several embryonic cell types related to limb development by in situ hybridisation and single-cell RNA-sequencing analysis. Pbx4 protein was also detected in the bone marrow of adult mice with a lower level compared with that in the testis. PBX4 preferentially binds to the promoters of a large number of genes including those for other HD-containing proteins and ribosomal proteins whose mutations are related to anaemia. PBX4-binding sites are enriched in motifs similar to those of other HD-containing proteins such as PKNOX1 indicating that PBX4 may also act as a co-transcription factor like other PBC proteins. Together, these results show that PBX4 participates in limb development and haematopoiesis while its function in spermatogenesis has not been revealed by gene KO probably due to the complementary effects of other genes.
Topics: Animals; Male; Mice; Extremities; Gene Expression Regulation, Developmental; Hematopoiesis; Homeodomain Proteins; Mice, Inbred C57BL; Mice, Knockout; Promoter Regions, Genetic; Testis; Transcription Factors; DNA-Binding Proteins
PubMed: 38230761
DOI: 10.1111/cpr.13580 -
Frontiers in Veterinary Science 2024This study aimed to explore the effects of neonatal vitamin A (VA) supplementation on testis development and spermatogenesis. A total of 32 newborn lambs were...
This study aimed to explore the effects of neonatal vitamin A (VA) supplementation on testis development and spermatogenesis. A total of 32 newborn lambs were intramuscularly injected with corn oil (control group) or corn oil + 2500 IU/kg BW VA (VA group). They were slaughtered and sampled at 3 weeks and 8 months of age to analyze spermatogenesis, cell proliferation, hormone secretion, antioxidant status of the testis, and adult sheep sperm parameters. Compared with the control group, the expression of spermatogonial differentiation-related genes in VA group was up-regulated ( < 0.05). Testis weight, seminiferous tubule diameter, number of spermatogonium and spermatocyte, and sperm density increased significantly in VA group at 8 months of age ( < 0.05). Neonatal VA injection upregulated the expression of the cell proliferation marker and cell cycle-related genes in the testis ( < 0.05). VA increased the concentrations of testosterone (T), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) in the serum and upregulated steroidogenesis-related genes in the testis ( < 0.05). The antioxidant levels in the VA group were maintained at high levels. The total antioxidant capacity (T-AOC), antioxidant enzyme content and antioxidant-related genes were increased in the testis ( < 0.05). Furthermore, neonatal VA injection activated retinoic acid (RA) signaling to maintain the blood-testosterone barrier (BTB) in the testis of 3-week-old sheep. AMP-activated protein kinase (AMPK) and protein kinase B (AKT) signaling were also modulated in the sheep testis ( < 0.05). Taken together, VA supplementation in newborn rams promotes testis development and spermatogenesis to improve fertility.
PubMed: 38756517
DOI: 10.3389/fvets.2024.1370576 -
American Journal of Human Genetics Jun 2024Sperm production and function require the correct establishment of DNA methylation patterns in the germline. Here, we examined the genome-wide DNA methylation changes...
Sperm production and function require the correct establishment of DNA methylation patterns in the germline. Here, we examined the genome-wide DNA methylation changes during human spermatogenesis and its alterations in disturbed spermatogenesis. We found that spermatogenesis is associated with remodeling of the methylome, comprising a global decline in DNA methylation in primary spermatocytes followed by selective remethylation, resulting in a spermatids/sperm-specific methylome. Hypomethylated regions in spermatids/sperm were enriched in specific transcription factor binding sites for DMRT and SOX family members and spermatid-specific genes. Intriguingly, while SINEs displayed differential methylation throughout spermatogenesis, LINEs appeared to be protected from changes in DNA methylation. In disturbed spermatogenesis, germ cells exhibited considerable DNA methylation changes, which were significantly enriched at transposable elements and genes involved in spermatogenesis. We detected hypomethylation in SVA and L1HS in disturbed spermatogenesis, suggesting an association between the abnormal programming of these regions and failure of germ cells progressing beyond meiosis.
Topics: Humans; Spermatogenesis; Male; DNA Methylation; Genome, Human; Spermatids; Spermatocytes; DNA Transposable Elements; Spermatozoa; Meiosis; Transcription Factors
PubMed: 38759652
DOI: 10.1016/j.ajhg.2024.04.017 -
PLoS Genetics Aug 2023The double stranded RNA binding protein Adad1 (adenosine deaminase domain containing 1) is a member of the adenosine deaminase acting on RNAs (Adar) protein family with...
The double stranded RNA binding protein Adad1 (adenosine deaminase domain containing 1) is a member of the adenosine deaminase acting on RNAs (Adar) protein family with germ cell-specific expression. In mice, Adad1 is necessary for sperm differentiation, however its function outside of mammals has not been investigated. Here, through an N-ethyl-N-nitrosourea (ENU) based forward genetic screen, we identified an adad1 mutant zebrafish line that develops as sterile males. Further histological examination revealed complete lack of germ cells in adult mutant fish, however germ cells populated the gonad, proliferated, and entered meiosis in larval and juvenile fish. Although meiosis was initiated in adad1 mutant testes, the spermatocytes failed to progress beyond the zygotene stage. Thus, Adad1 is essential for meiosis and germline maintenance in zebrafish. We tested if spermatogonial stem cells were affected using nanos2 RNA FISH and a label retaining cell (LRC) assay, and found that the mutant testes had fewer LRCs and nanos2-expressing cells compared to wild-type siblings, suggesting that failure to maintain the spermatogonial stem cells resulted in germ cell loss by adulthood. To identify potential molecular processes regulated by Adad1, we sequenced bulk mRNA from mutants and wild-type testes and found mis-regulation of genes involved in RNA stability and modification, pointing to a potential broader role in post-transcriptional regulation. Our findings suggest that the RNA regulatory protein Adad1 is required for fertility through regulation of spermatogonial stem cell maintenance in zebrafish.
Topics: Animals; Male; Mice; Adenosine Deaminase; Germ Cells; Mammals; Meiosis; RNA; RNA-Binding Proteins; Semen; Testis; Zebrafish; Zebrafish Proteins
PubMed: 37552671
DOI: 10.1371/journal.pgen.1010589