Testicular macrophages (TM) comprise the largest immune cell population in the mammalian testis. They are characterized by a subdued proinflammatory response upon adequate stimulation, and a ...polarization toward the immunoregulatory and immunotolerant M2 phenotype. This enables them to play a relevant role in supporting the archetypical functions of the testis, namely spermatogenesis and steroidogenesis. During infection, the characteristic blunted immune response of TM reflects the need for a delicate balance between a sufficiently strong reaction to counteract invading pathogens, and the prevention of excessive proinflammatory cytokine levels with the potential to disturb or destroy spermatogenesis. Local microenvironmental factors that determine the special phenotype of TM have just begun to be unraveled, and are discussed in this review.
Thousands of long noncoding RNAs (lncRNAs) have been identified in mouse, rat, and human testes, some of which play important roles in testis development and spermatogenesis. However, systematic ...analysis of lncRNAs expressed in postnatal pig testes has not been reported. Thus, in this study, we present the expression and characterization of lncRNAs in immature (30-day-old D30) and mature (180-day-old D180) pig testes. A total of 90 440 168 (85.75%) and 97 001 700 (95.35%) 150-base-pair paired-end clean reads were generated in D30 and D180 cDNA libraries, respectively, using the Illumina HiSeq 4000 platform; 36 727 transcripts were assembled in those two libraries, 777 lncRNA transcripts from 752 lncRNA gene loci were identified using the highly stringent pipeline, and 101 of those lncRNA transcripts were significantly differentially expressed. Those lncRNAs shared some characteristics with other mammals, including fewer exons, shorter length and exon length, and lower expression level compared with those of protein-coding genes; 402 protein-coding genes (<10 kb) were found as nearest neighbors of 294 out of 752 lncRNA genes, and gene ontology enrichment showed that they were enriched in transcription- and development-related processes. Fifteen differentially expressed and 10 novel lncRNAs were randomly selected and validated by quantitative polymerase chain reaction (PCR) and reverse transcriptase PCR. In addition, one of the 10 novel lncRNAs was further confirmed using RACE clone technology. This study provides a catalog of porcine testes lncRNAs for further understanding their regulation roles in pig testis development and spermatogenesis.
A healthy man typically produces between 50 × 10(6) and 200 × 10(6) spermatozoa per day by spermatogenesis; in the absence of Sertoli cells in the male gonad, this individual would be infertile. In ...the adult testis, Sertoli cells are sustentacular cells that support germ cell development by secreting proteins and other important biomolecules that are essential for germ cell survival and maturation, establishing the blood-testis barrier, and facilitating spermatozoa detachment at spermiation. In the fetal testis, on the other hand, pre-Sertoli cells form the testis cords, the future seminiferous tubules. However, the role of pre-Sertoli cells in this process is much less clear than the function of Sertoli cells in the adult testis. Within this framework, we provide an overview of the biology of the fetal, pubertal, and adult Sertoli cell, highlighting relevant cell biology studies that have expanded our understanding of mammalian spermatogenesis.
STUDY QUESTION
Is it possible to derive a scaffold from human testis for the purpose of tissue engineering and regenerative medicine?
SUMMARY ANSWER
We developed a method to produce a cytocompatible ...decellularized testicular matrix (DTM) while maintaining the native tissue-specific characteristics and components.
WHAT IS KNOWN ALREADY
The potential benefits of tissue-specific scaffolds consisting of naturally-derived extracellular matrix (ECM) have been demonstrated using a wide variety of animal and human tissue sources. However, so far, testis scaffolds have never been considered for constructive remodelling purposes.
STUDY DESIGN, SIZE, DURATION
Human cadaveric testicular tissue was exposed for 24 or 48 h to 1% Triton X-100 and/or 1% sodium dodecyl sulphate (SDS). Acellular samples were used for further scaffold characterization purposes.
PARTICIPANTS/MATERIALS, SETTING, METHODS
The extent of decellularization was evaluated by histology. Confirmation of cell removal in DTM was done by a DNA quantification technique. Retention of testicular tissue-specific characteristics was evaluated by mass spectrometry, immunohistochemistry, Alcian blue staining and scanning electron microscopy. Soluble toxicity and testicular cell attachment was assessed to check the cytocompatibility of DTM scaffolds.
MAIN RESULTS AND THE ROLE OF CHANCE
Histological analysis showed that DTM could be obtained by mechanical agitation in 1% SDS for 24 h. The resulting DTM was found to be clear of cells while retaining the typical three-dimensional structure and the major components of the native tissue scaffold, including collagen type I and IV, fibronectin, laminin and glycosaminoglycans. In addition, using proteomic analysis, we revealed numerous additional ECM proteins in DTM, indicating its complex nature. The mass spectrometry data were deposited to the ProteomeXchange with identifier PXD001524. Importantly, we demonstrated that DTM scaffolds are not cytotoxic, as evidenced by MTT assay not showing an aberrant fibroblast proliferation activity after indirect exposure, and support testicular cell attachment and infiltration.
LIMITATIONS, REASONS FOR CAUTION
The functionality of human testicular cells in DTM needs to be investigated.
WIDER IMPLICATIONS OF THE FINDINGS
Our results suggest that the insights into the molecular composition of the testicular ECM provide new clues for the unravelling of its important yet poorly understood role in regulating testicular function, and DTM-based bioscaffolds are promising components for the development of human in vitro spermatogenesis as a treatment for various types of male fertility disorders.
STUDY FUNDING/COMPETING INTEREST(S)
This study is supported by a Ph.D. grant from the Agency for Innovation by Science and Technology (IWT) and research grants from the Flemish League Against Cancer-Public Utility Foundation (VLK), the Scientific Research Foundation Flanders (FWO), the Vrije Universiteit Brussel, the Swedish Research Council/ Finnish Academy of Science, Emil och Wera Cornells Stiftelse, the Swedish Childhood Cancer Foundation as well as through the regional agreement on medical training and clinical research (ALF) between Stockholm County Council and Karolinska Institutet. J.-B.S. was supported by the German Research Foundation (DFG-Grant No.: STU 506/3-1). E.G. and J.D.K. are postdoctoral fellows of the Scientific Research Foundation Flanders. The authors declare that no competing interests exist.
TRIAL REGISTRATION NUMBER
Not applicable.
Fluoride is known to affect the pro-inflammatory cytokines in the testis. Most of the recent literatures cited that cytokines regulate the blood-testis-barrier (BTB). However, the involvement of ...cytokines in the fluoride induced toxicity in BTB remains unclear. In order to study this, 60 male Sprague-Dawley (SD) rats were taken and randomly divided into 5 groups which included four fluoride groups exposed to 0, 25, 50, and 100 mg/L NaF in distilled water and one positive control group. On the 29th day of fluoride exposure, the positive control group rats were administered 0.1% CaCl2 solution. Biotin tracer technology and transmission electron microscopy (TEM) analysis were applied to evaluate the function and ultra-structure of BTB. The expression levels of the BTB associated proteins, actin relative protein 3 (Arp3), interleukin-1 alpha (IL-1α), and transforming growth factor beta-3 (TGF-β3) were determined using Western blotting and Enzyme Linked Immunosorbent Assay (ELISA) respectively, meanwhile the actin filament (F-actin) was detected by fluorescent phalloidin conjugates. Our results revealed that the function and the ultra-structure of BTB in all the fluoride treated groups were damaged with a concomitant significant decreases in basal ectoplasmic specialization (basal ES), associated protein β-catenin, and F-actin. Moreover, Arp3 levels were significantly increased in 50 and 100 mg/L NaF groups. Meanwhile, IL-1α significantly increased in all the fluoride treated groups. In summary, we concluded that an increase in IL-1α induced by NaF significantly decreased the expression of F-actin and the organization of F-actin highly branched, which might facilitate the BTB's functional and ultra-structural variations.
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•Fluoride affected the function and ultra-structure of BTB in the testis.•Fluoride induced F-actin bundles into highly branched arrangement.•Fluoride markedly increased the expression level of actin related protein 3(Arp3).•IL-1α induced the damage of BTB in the rat exposed to fluoride for 30 days.
Cisplatin exposure represents a significant fertility problem for childhood cancer. In this study we examined the possible therapeutic role of Zinc oxide nanoparticles (ZnO-NPs) on Cisplatin (Cis) ...induced impairment in the spermatogenesis initiation during puberty. Seventy-two male rats aged 30 days were distributed into four equal groups: Control group; ZnO-NPs group (intraperitoneal i.p. injected with 5 mg/kg ZnO-NPs once a week for eight weeks); Cis group (i.p. injected with a single dose of 5 mg/kg); ZnO-NPs + Cis group (ZnO-NPs injection 2 hrs before Cis). Each group was subdivided into three groups and was sacrificed 7, 30 and, 60 days after cisplatin induction, which considered prepubertal at 37-day-old, productive at 60-day-old, and completely adult at 90-day-old. Biochemical, molecular, immunohistochemical, and ultrastructural examinations were studied on the testicular tissues and sperm samples. Group treated with Cis showed a decrease in the antioxidant activity and an increase in the reactive oxygen species (ROS), which in turn caused disruption in blood-testis barrier (BTB) proteins in the three different rat ages, and sperm DNA damage in the adult rats compared to control group (p < 0.05). Moreover, alterations in the structural and the ultrastructural morphology of the testis were observed compared with the control at 37, 60 and 90 days old rats. ZnO-NPs administration to Cis group manifested a significant decrease in the ROS that restored the BTB proteins, enhanced the architecture of the testis in the three different rat ages, and increased sperm DNA integrity in the adult rats. Zinc oxide nanoparticles could restore the male reproductive capacity in the adult rats after induction of Cis in the prepubertal period by promoting spermatogenesis.
To facilitate temperature adjustments, the testicles are located outside the body cavity. In most mammals, the temperature of the testes is lower than the body temperature to ensure the normal ...progression of spermatogenesis. Rising temperatures affect spermatogenesis and eventually lead to a decline in male fertility or even infertility. However, the testes are composed of different cell types, including spermatogonial stem cells (SSCs), spermatocytes, spermatozoa, Leydig cells, and Sertoli cells, which have different cellular responses to heat stress. Recent studies have shown that using different drugs can relieve heat stress-induced reproductive damage by regulating different signaling pathways. Here, we review the mechanisms by which heat stress damages different cells in testes and possible treatments.
Background: Several studies have described an increasing frequency of male reproductive disorders, which may have a common origin in fetal life and which are hypothesized to be caused by endocrine ...disruptors. Phthalate esters represent a class of environmental endocrine-active chemicals known to disrupt development of the male reproductive tract by decreasing testosterone production in the fetal rat. Objectives: Using the organ culture system we developed previously, we investigated the effects on the development of human fetal testis of one phthalate--mono-2-ethylhexyl phthalate (MEHP)--an industrial chemical found in many products, which has been incriminated as a disruptor of male reproductive function. Methods: Human fetal testes were recovered during the first trimester (7-12 weeks) of gestation, a critical period for testicular differentiation, and cultured for 3 days with or without MEHP in basal conditions or stimulated with luteinizing hormone (LH). Results: Whatever the dose, MEHP treatment had no effect on basal or LH-stimulated testosterone produced by the human fetal testis in vitro, although testosterone production can be modulated in our culture system. MEHP (10⁻⁴ M) did not affect proliferation or apoptosis of Sertoli cells, but it reduced the mRNA expression of anti-Müllerian hormone. MEHP (10⁻⁴ M) reduced the number of germ cells by increasing their apoptosis, measured by the detection of caspase-3-positive germ cells, without modification of their proliferation. Conclusions: This is the first experimental demonstration that phthalates alter the development of the germ cell lineage in humans. However, in contrast to results observed in the rat, phthalates did not affect steroidogenesis.
In rat testes, the ectoplasmic specialization (ES) at the Sertoli-Sertoli and Sertoli-spermatid interface known as the basal ES at the blood-testis barrier and the apical ES in the adluminal ...compartment, respectively, is a testis-specific adherens junction. The remarkable ultrastructural feature of the ES is the actin filament bundles that sandwiched in between the cisternae of endoplasmic reticulum and apposing plasma membranes. Although these actin filament bundles undergo extensive reorganization to switch between their bundled and debundled state to facilitate blood-testis barrier restructuring and spermatid adhesion/transport, the regulatory molecules underlying these events remain unknown. Herein we report findings of an actin filament cross-linking/bundling protein palladin, which displayed restrictive spatiotemporal expression at the apical and the basal ES during the epithelial cycle. Palladin structurally interacted and colocalized with Eps8 (epidermal growth factor receptor pathway substrate 8, an actin barbed end capping and bundling protein) and Arp3 (actin related protein 3, which together with Arp2 form the Arp2/3 complex to induce branched actin nucleation, converting bundled actin filaments to an unbundled/branched network), illustrating its role in regulating actin filament bundle dynamics at the ES. A knockdown of palladin in Sertoli cells in vitro with an established tight junction (TJ)-permeability barrier was found to disrupt the TJ function, which was associated with a disorganization of actin filaments that affected protein distribution at the TJ. Its knockdown in vivo also perturbed F-actin organization that led to a loss of spermatid polarity and adhesion, causing defects in spermatid transport and spermiation. In summary, palladin is an actin filament regulator at the ES.
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•Glyphosate (GLY) causes blood-testis barrier (BTB) damage and low-quality sperm.•GLY-induced BTB injury contributes to sperm quality decrease.•NOX1-derived oxidative stress results ...in the disruption of BTB integrity.•NOX1 upregulation may be due to the interaction of GLY with estrogen receptor-α.
Blood-testis barrier (BTB) creates a privileged niche indispensable for spermatogenesis. Glyphosate (GLY), the most commonly used herbicide worldwide, has been reported to decrease sperm quality. However, whether and how GLY destroys the BTB to affect sperm quality remains to be elucidated. Herein, this study was designed to investigate the influence of GLY on the BTB in vivo and in vitro experiments. The results showed that male rats exposed to GLY for 4 months exhibited a decrease in sperm quality and quantity, accompanied by BTB integrity disruption and testicular oxidative stress. Additionally, GLY-induced reactive oxygen species (ROS) contributed to the downregulation of BTB-related proteins in primary Sertoli cells (SCs). Intriguingly, we identified a marked upregulation of oxidative stress-related gene NOX1 in GLY-exposed testis based on transcriptome analysis. NOX1 knockdown blocked the GLY-induced oxidative stress, as well as prevented BTB-related protein decrease in SCs. Furthermore, the estrogen receptor (ER)-α was significantly upregulated in vivo and in vitro models. An ER-α inhibitor decreased the expression levels of both ER-α and NOX1. Mechanistically, GLY directly interacted with ER-α at the site of Pro39 and Lys401 to promote ER-α activation, which boosted NOX1 expression to trigger ROS accumulation. Collectively, these results demonstrate that long-term GLY exposure adversely affects BTB integrity, which disrupts spermatogenesis via activation of ER-α/NOX1 axis. This study presents a better understanding of the risk of long-term GLY exposure to male fertility.
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