Search

Testosterone, the primary sex hormone in male lizards, is closely linked to Leydig cell activity (the cells where steroidogenesis occurs) throughout the reproductive cycle, but testosterone action is related to androgen receptors (ARs) distribution in the seminiferous epithelium. In temperate zones, environmental factors detected through the hypothalamic-pituitary-gonadal axis, downregulate plasma testosterone, resulting in a seasonal reproductive cycle. The aim of this work is to study plasma testosterone in adult male lizards of Liolaemus cuyanus, an oviparous species, throughout its reproductive cycle and its relationship with Leydig cell histology, TotalLeydigCell/ActiveLeydigCell (TLC/ALC) ratio, environmental factors (temperature, relative humidity and solar irradiation) and ARs distribution in seminiferous epithelium. Specimens (N = 27) were captured (October to March) in a semi-arid zone (Valle de Matagusanos, San Juan, Argentina) and grouped into three relevant reproductive periods: pre-reproductive (PrR), reproductive (R), and post-reproductive (PsR). Significant differences in plasma testosterone were found among these periods, highest during R than in PsR. A significant positive correlation between plasma testosterone and TLC/ALC ratio was also observed. Plasma testosterone has a significant positive correlation only with solar irradiation, but not with the other variables. In PrR and R, ARs distribution was cytoplasmic and nuclear, shifting to only cytoplasmic in PsR. These results highlight the close correspondence between plasma testosterone, Leydig cell histology and activity, environmental factors, and ARs distribution, resulting in a synchronization that allows males of L. cuyanus to coordinate their reproductive cycle with the most favorable environmental conditions, probably for mating and birth of offspring., (© 2024 Wiley Periodicals LLC.)

Recent technological and algorithmic advances enable single-cell transcriptomic analysis with remarkable depth and breadth. Nonetheless, a persistent challenge is the compromise between the ability to profile high numbers of cells and the achievement of full-length transcript coverage. Currently, the field is progressing and developing new and creative solutions that improve cellular throughput, gene detection sensitivity and full-length transcript capture. Furthermore, long-read sequencing approaches for single-cell transcripts are breaking frontiers that have previously blocked full transcriptome characterization. We here present a comprehensive overview of available options for single-cell transcriptome profiling, highlighting the key advantages and disadvantages of each approach., Competing Interests: Declaration of interests The authors have declared no competing interest., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Rabbits are sensitive to dietary cholesterol and rapidly develop hypercholesterolemia, leading to prominent subfertility. Sterol regulatory element-binding protein isoform 2 drives the intracellular cholesterol pathway in many tissues, including the testicles. Its abnormal regulation could be the mainly responsible for the failure of suppressing cholesterol synthesis in a cholesterol-enriched environment, ultimately leading to testicular and seminal alterations. However, extra-virgin olive oil consumption has beneficial properties that promote lowering of cholesterol levels, including the recovery of seminal parameters altered under a high-fat diet., Objectives: Our goal was to investigate the effects of high-fat diet supplementation with extra-virgin olive oil at the molecular level on rabbit testes, by analyzing sterol regulatory element-binding protein isoform 2 protein and its corresponding downstream effectors., Materials and Methods: During 12 months, male rabbits were fed a control diet, high-fat diet, or 6-month high-fat diet followed by 6-month high-fat diet plus extra-virgin olive oil. Serum lipids, testosterone levels, bodyweight, and seminal parameters were tested. The mRNA and protein levels of sterol regulatory element-binding protein isoform 2, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase, and low-density lipoprotein receptor were determined by semi-quantitative polymerase chain reaction and Western blotting techniques. The expression pattern of sterol regulatory element-binding protein isoform 2 protein in the rabbit testicles was studied by indirect immunofluorescence. In addition, testicular cholesterol was detected and quantified by filipin staining and gas chromatography., Results: The data showed that the addition of extra-virgin olive oil to high-fat diet reduced testicular cholesterol levels and recovered the expression of sterol regulatory element-binding protein isoform 2, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase, and low-density lipoprotein receptor initially altered by the high-fat diet., Discussion and Conclusions: The combination of high-fat diet with extra-virgin olive oil encourages testicular recovery by modifying the expression of the enzymes related to intracellular cholesterol management., (© 2023 American Society of Andrology and European Academy of Andrology.)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the RNA virus responsible for the coronavirus disease 2019 (COVID-19) pandemic. Although SARS-CoV-2 was reported to alter several cellular pathways, its impact on DNA integrity and the mechanisms involved remain unknown. Here we show that SARS-CoV-2 causes DNA damage and elicits an altered DNA damage response. Mechanistically, SARS-CoV-2 proteins ORF6 and NSP13 cause degradation of the DNA damage response kinase CHK1 through proteasome and autophagy, respectively. CHK1 loss leads to deoxynucleoside triphosphate (dNTP) shortage, causing impaired S-phase progression, DNA damage, pro-inflammatory pathways activation and cellular senescence. Supplementation of deoxynucleosides reduces that. Furthermore, SARS-CoV-2 N-protein impairs 53BP1 focal recruitment by interfering with damage-induced long non-coding RNAs, thus reducing DNA repair. Key observations are recapitulated in SARS-CoV-2-infected mice and patients with COVID-19. We propose that SARS-CoV-2, by boosting ribonucleoside triphosphate levels to promote its replication at the expense of dNTPs and by hijacking damage-induced long non-coding RNAs' biology, threatens genome integrity and causes altered DNA damage response activation, induction of inflammation and cellular senescence., (© 2023. The Author(s).)

NK cell deficiencies (NKD) are a type of primary immune deficiency in which the major immunologic abnormality affects NK cell number, maturity, or function. Since NK cells contribute to immune defense against virally infected cells, patients with NKD experience higher susceptibility to chronic, recurrent, and fatal viral infections. An individual with recurrent viral infections and mild hypogammaglobulinemia was identified to have an X-linked damaging variant in the transcription factor gene ELF4. The variant does not decrease expression but disrupts ELF4 protein interactions and DNA binding, reducing transcriptional activation of target genes and selectively impairing ELF4 function. Corroborating previous murine models of ELF4 deficiency (Elf4-/-) and using a knockdown human NK cell line, we determined that ELF4 is necessary for normal NK cell development, terminal maturation, and function. Through characterization of the NK cells of the proband, expression of the proband's variant in Elf4-/- mouse hematopoietic precursor cells, and a human in vitro NK cell maturation model, we established this ELF4 variant as a potentially novel cause of NKD.

Human NK cell deficiency (NKD) is a primary immunodeficiency in which the main clinically relevant immunological defect involves missing or dysfunctional NK cells. Here, we describe a familial NKD case in which 2 siblings had a substantive NKD and neutropenia in the absence of other immune system abnormalities. Exome sequencing identified compound heterozygous variants in Go-Ichi-Ni-San (GINS) complex subunit 4 (GINS4, also known as SLD5), an essential component of the human replicative helicase, which we demonstrate to have a damaging impact upon the expression and assembly of the GINS complex. Cells derived from affected individuals and a GINS4-knockdown cell line demonstrate delayed cell cycle progression, without signs of improper DNA synthesis or increased replication stress. By modeling partial GINS4 depletion in differentiating NK cells in vitro, we demonstrate the causal relationship between the genotype and the NK cell phenotype, as well as a cell-intrinsic defect in NK cell development. Thus, biallelic partial loss-of-function mutations in GINS4 define a potentially novel disease-causing gene underlying NKD with neutropenia. Together with the previously described mutations in other helicase genes causing NKD, and with the mild defects observed in other human cells, these variants underscore the importance of this pathway in NK cell biology.

Male fertility has been shown to be dependent on cholesterol homeostasis. This lipid is essential for testosterone synthesis and spermatogenesis, but its levels must be maintained in an optimal range for proper testicular function. In particular, sperm cells' development is very sensitive to high cholesterol levels, noticeably during acrosomal formation. The aim of this work was to study whether the molecular pathway that regulates intracellular cholesterol, the sterol regulatory element-binding protein (SREBP) pathway, is affected in the testicles of animals under a fat diet. To investigate this, we took advantage of the non-obese hypercholesterolemia (HC) model in New Zealand rabbits that displays poor sperm and seminal quality. The testicular expression of SREBP isoform 2 (SREBP2) and its target molecules 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGCR) and low-density lipoprotein receptor (LDLR) were studied under acute (6 months) and chronic (more than 12 months) fat intake by RT-PCR, western blot and immunofluorescence. Our findings showed that fat consumption promoted down-regulation of the SREBP2 pathway in the testicle at 6 months, but upregulation after a chronic period. This was consistent with load of testicular cholesterol, assessed by filipin staining. In conclusion, the intracellular pathway that regulates cholesterol levels in the testicle is sensitive to dietary fats, and behaves differently depending on the duration of consumption: it has a short-term protective effect, but became deregulated in the long term, ultimately leading to a detrimental situation. These results will contribute to the understanding of the basic mechanisms of the effect of fat consumption in humans with idiopathic infertility., (© The Author(s) 2021. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Human natural killer cell deficiency (NKD) arises from inborn errors of immunity that lead to impaired NK cell development, function, or both. Through the understanding of the biological perturbations in individuals with NKD, requirements for the generation of terminally mature functional innate effector cells can be elucidated. Here, we report a cause of NKD resulting from compound heterozygous mutations in minichromosomal maintenance complex member 10 (MCM10) that impaired NK cell maturation in a child with fatal susceptibility to CMV. MCM10 has not been previously associated with monogenic disease and plays a critical role in the activation and function of the eukaryotic DNA replisome. Through evaluation of patient primary fibroblasts, modeling patient mutations in fibroblast cell lines, and MCM10 knockdown in human NK cell lines, we have shown that loss of MCM10 function leads to impaired cell cycle progression and induction of DNA damage-response pathways. By modeling MCM10 deficiency in primary NK cell precursors, including patient-derived induced pluripotent stem cells, we further demonstrated that MCM10 is required for NK cell terminal maturation and acquisition of immunological system function. Together, these data define MCM10 as an NKD gene and provide biological insight into the requirement for the DNA replisome in human NK cell maturation and function.

Pigment epithelium derived factor (PEDF) expression has been described in many organs as showing neurotrophic, anti-angiogenic, anti-apoptotic, anti-inflammatory, anti-oxidant and pro-cell survival properties. However, references to its activity in the male reproductive system are scarce. We aimed to characterize the expression of PEDF in the male reproductive tract of Wistar rats by using RT-PCR, western blot and immunostaining and also evaluate the effect of flutamide in PEDF expression. We found that PEDF is expressed in the epididymis, prostate and seminal vesicles in Wistar rats, but notably not in the testes. Under the effect of flutamide PEDF expression decreased, recovering by suppressing the antiandrogen. The epididymis is an essential organ in sperm maturation-storages. The role of PEDF in this physiological process has not been fully elucidated yet, but considering that in other systems PEDF has anti-apoptotic, anti-oxidants and pro-cell survival properties, its expression along the epididymis could play a role in the protection of spermatozoa while they are stored., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Incontinentia pigmenti (IP; Online Mendelian Inheritance in Man catalog #308300) is an X-linked dominant ectodermal disorder caused by mutations of the inhibitor of κ polypeptide gene enchancer in B cells, kinase γ ( IKBKG )/ nuclear factor κB, essential modulator ( NEMO ) gene. Hemizygous IKBKG/NEMO loss-of-function (LoF) mutations are lethal in males, thus patients are female, and the disease is always transmitted from an IP-affected mother to her daughter. We present 2 families with father-to-daughter transmission of IP and provide for the first time molecular evidence that the combination of somatic and germ-line mosaicism for IKBKG/NEMO loss of function mutations in IP males resulted in the transmission of the disease to a female child. We searched for the IKBKG/NEMO mutant allele in blood, urine, skin, and sperm DNA and found that the 2 fathers were somatic and germ-line mosaics for the p.Gln132×mutation or the exon 4-10 deletion of IKBKG/NEMO , respectively. The highest level of IKBKG/NEMO mutant cells was detected in the sperm, which might explain the recurrence of the disease. We therefore recommend careful clinical evaluation in IP male cases and the genetic investigation in sperm DNA to ensure correct genetic counseling and prevent the risk of paternal transmission of IP., Competing Interests: POTENTIAL CONFLICT OF INTEREST: The authors have indicated they have no potential conflicts of interest to disclose., (Copyright © 2017 by the American Academy of Pediatrics.)

Hypohidrotic ectodermal dysplasia (HED) consists of disorders resulting from molecular alterations of ectodysplasin-A (EDA) pathway. Hypomorphic mutations in NF-kB essential modulator, downstream EDA, result in HED with immunodeficiency (HED-ID), characterized by susceptibility to encapsulated pyogenic bacteria infections. Increased susceptibility to pneumococcal infections and poor response to polysaccharide antigens are associated with defect in T-independent B-cell immunity. We investigated B-cell differentiation and immunoglobulin secretion induced by the TLR9 ligand CpG in two HED-ID and in a HED patient caused by EDA mutations (XLHED). In HED-ID, only few B cells differentiated into plasma cells upon TLR9 stimulation and memory B cells did not produce IgG and IgA, but small amounts of IgM. Unexpectedly, memory B cells from XLHED patient failed to produce normal IgA or IgG amount upon TLR9 stimulation. Our findings expand the knowledge about the pathogenesis of humoral alterations in HED patients and help explain the susceptibility to pneumococcal infections., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Mendelian primary immunodeficiency diseases (MPIDs) are rare disorders affecting distinct constituents of the innate and adaptive immune system. Although they are genetically heterogeneous, a substantial group of MPIDs is due to mutations in genes affecting the nuclear factor-κB (NF-κB) transcription pathway, essential for cell proliferation and cell survival and involved in innate immunity and inflammation. Many of these genes encode for crucial regulatory components of the NF-κB pathway and their mutations are associated with immunological and developmental signs somehow overlapping in patients with MPIDs. At present, nine different MPIDs listed in the online mendelian inheritance in man (OMIM) are caused by mutations in at least nine different genes strictly involved in the NF-κB pathway that result in defects in immune responses. Here we report on the distinct function of each causative gene, on the impaired NF-κB step and more in general on the molecular mechanisms underlining the pathogenesis of the disease. Overall, the MPIDs affecting the NF-κB signalosome require a careful integrated diagnosis and appropriate genetic tests to be molecularly identified. Their discovery at an ever-increasing rate will help establish a common therapeutic strategy for a subclass of immunodeficient patients.

Anhidrotic Ectodermal Dysplasia with ImmunoDeficiency (EDA-ID, OMIM 300291) and Incontinentia Pigmenti (IP, OMIM 308300) are two rare diseases, caused by mutations of the IKBKG/NEMO gene. The protein NEMO/IKKγ is essential for the NF-κB activation pathway, involved in a variety of physiological and cellular processes, such as immunity, inflammation, cell proliferation, and survival. A wide spectrum of IKBKG/NEMO mutations have been identified so far, and, on the basis of their effect on NF-κB activation, they are considered hypomorphic or amorphic (loss of function) mutations. IKBKG/NEMO hypomorphic mutations, reducing but not abolishing NF-κB activation, have been identified in EDA-ID and IP patients. Instead, the amorphic mutations, abolishing NF-κB activation by complete IKBKG/NEMO gene silencing, cause only IP. Here, we present an overview of IKBKG/NEMO mutations in EDA-ID and IP patients and describe similarities and differences between the clinical/immunophenotypic and genetic aspects, highlighting any T and B lymphocyte defect, and paying particular attention to the cellular and molecular defects that underlie the pathogenesis of both diseases.