Hepatitis E virus infects human testicular tissue and Sertoli cells.

Globally, hepatitis E virus (HEV) infections are prevalent. The finding of high viral loads and persistent viral shedding in ejaculate suggests that HEV replicates within the human male genital tract, but its target organ is unknown and appropriate models are lacking. We aimed to determine the HEV tropism in the human testis and its potential influence on male reproductive health. We conducted an ex vivo culture of human testis explants and in vitro culture of primary human Sertoli cells. Clinically derived HEV genotype 1 (HEV1) and HEV3 virions, as well as rat-derived HEV-C1, were used for inoculation. Transcriptomic analysis was performed on testis tissues collected from tacrolimus-treated rabbits with chronic HEV3 infection. Our findings reveal that HEV3, but not HEV1 or HEV-C1, can replicate in human testis explants and primary human Sertoli cells. Tacrolimus treatment significantly enhanced the replication efficiency of HEV3 in testis explants and enabled successful HEV1 infection in Sertoli cells. HEV3 infection disrupted the secretion of several soluble factors and altered the cytokine microenvironment within primary human Sertoli cells. Finally, intratesticular transcriptomic analysis of immunocompromised rabbits with chronic HEV infection indicated downregulation of genes associated with spermatogenesis. HEV can infect the human testicular tissues and Sertoli cells, with increased replication efficiency when exposed to tacrolimus treatment. These findings shed light on how HEV may persist in the ejaculate of patients with chronic hepatitis E and provide valuable ex vivo tools for studying countermeasures.

The gut metabolite 3-hydroxyphenylacetic acid rejuvenates spermatogenic dysfunction in aged mice through GPX4-mediated ferroptosis.

BACKGROUND: Aging-related fertility decline is a prevalent concern globally. Male reproductive system aging is mainly characterized by a decrease in sperm quality and fertility. While it is known that intestinal physiology changes with age and that microbiota is shaped by physiology, the underlying mechanism of how the microbiota affects male reproductive aging is still largely unexplored. RESULTS: Here, we utilized fecal microbiota transplantation (FMT) to exchange the fecal microbiota between young and old mice. Cecal shotgun metagenomics and metabolomics were used to identify differences in gut microbiota composition and metabolic regulation during aging. Our results demonstrated that FMT from young to old mice alleviated aging-associated spermatogenic dysfunction through an unexpected mechanism mediated by a gut bacteria-derived metabolite, 3-hydroxyphenylacetic acid (3-HPAA). 3-HPAA treatment resulted in an improvement of spermatogenesis in old mice. RNA sequencing analysis, qRT-PCR and Western blot revealed that 3-HPAA induced an upregulation of GPX4, thereby restraining ferroptosis and restoring spermatogenesis. These findings were further confirmed by in vitro induction of ferroptosis and inhibition of GPX4 expression. CONCLUSIONS: Our results demonstrate that the microbiome-derived metabolite, 3-HPAA, facilitates spermatogenesis of old mice through a ferroptosis-mediated mechanism. Overall, these findings provide a novel mechanism of dysregulated spermatogenesis of old mice, and suggest that 3-HPAA could be a potential therapy for fertility decline of aging males in clinical practice. Video Abstract.

IL-6 translation is a therapeutic target of human cytokine release syndrome.

Chimeric antigen receptor (CAR) T therapies have achieved remarkable success for treating hematologic malignancies, yet are often accompanied by severe cytokine release syndrome (CRS). Here, an accidental clinical observation raised the possibility that metoprolol, an FDA-approved ß1 adrenergic receptor blocker widely used for cardiovascular conditions, may alleviate CAR T-induced CRS. Metoprolol effectively blocked IL-6 production in human monocytes through unexpected mechanisms of action of targeting IL-6 protein translation but not IL6 mRNA expression. Mechanistically, metoprolol diminished IL-6 protein synthesis via attenuating eEF2K-eEF2 axis-regulated translation elongation. Furthermore, an investigator-initiated phase I/II clinical trial demonstrated a favorable safety profile of metoprolol in CRS management and showed that metoprolol significantly alleviated CAR T-induced CRS without compromising CAR T efficacy. These results repurposed metoprolol, a WHO essential drug, as a potential therapeutic for CRS and implicated IL-6 translation as a mechanistic target of metoprolol, opening venues for protein translation-oriented drug developments for human inflammatory diseases.

Testicular sperm aspiration has a poor effect in predicting micro-TESE outcomes in NOA patients with AZFc deletion.

BACKGROUND: Testicular sperm aspiration (TESA) is widely used in the diagnosis and management of nonobstructive azoospermia. However, its ability for predicting microdissection testicular sperm extraction in nonobstructive azoospermia (NOA) patients with AZFc deletion remains uncertain. To investigate whether TESA affected the sperm retrieval rate (SRR) in NOA patients with AZFc deletion, a retrospective analysis of the clinical data of NOA patients with AZFc deletion who underwent microdissection testicular sperm extraction (micro-TESE) was conducted. The effects of age, testicular volume, follicle-stimulating hormone (FSH) levels, luteinizing hormone (LH) levels, testosterone (T) levels and TESA on the SRR were analyzed in this group of patients. RESULTS: A total of 181 individuals had their sperm successfully collected and underwent micro-TESE, with an SRR of 67.4%. The patients were separated into two groups based on their micro-TESE results (sperm acquisition and nonsperm acquisition), with no significant variations in age, testicular volume, FSH levels, LH levels, or T levels between the two groups. There was no significant difference in the SRR between any of the groups into which patients were classified based on reproductive hormone reference value ranges. Binary logistic regression was used to explore the absence of significant effects of age, testicular volume, FSH levels, LH levels, and T levels on sperm acquisition in patients undergoing micro-TESE. In the preoperative testicular diagnostic biopsy group, the sperm acquisition and nonsperm acquisition groups had SRRs of 90.1% and 65.1%, respectively. More significantly, there was no significant difference in the SRR between the negative preoperative testicular diagnostic biopsy group and the nonpreoperative testicular diagnostic biopsy group (65.1 vs. 63.8%, p = 0.855). CONCLUSION: There is a high probability of successful sperm acquisition in the testis of men undergoing micro-TESE. In this group of patients, age, testicular volume, FSH levels, LH levels, and T levels may have little bearing on the micro-TESE outcome. In patients whose preoperative TESA revealed the absence of sperm, the probability of obtaining sperm by micro-TESE remained high (65.1%); negative TESA results appeared to not influence the SRR (63.8%) in patients undergoing micro-TESE.

RéSUMé: CONTEXTE: L'aspiration testiculaire de spermatozoïdes (TESA) est largement utilisée dans le diagnostic et la prise en charge de l'azoospermie non obstructive. Cependant, sa capacité à prédire la présence de spermatozoïdes testiculaires lors de l'extraction par microdissection chez les patients atteints d'azoospermie non obstructive (NOA) et porteurs de la délétion AZFc reste incertaine. Pour déterminer si la TESA affectait le taux de récupération de spermatozoïdes (SRR) chez les patients atteints d'ANO avec délétion AZFc, nous avons mené une analyse rétrospective des données cliniques des patients atteints de NOA et d'une délétion AZFc ayant subi une extraction testiculaire de spermatozoïdes (micro-TESE) par microdissection. Les effets de l'âge, du volume testiculaire, des taux d'hormone folliculostimulante (FSH), d'hormone lutéinisante (LH), de testostérone (T) et de TESA sur le SRR ont été analysés chez ces patients. RéSULTATS: Au total, 181 personnes ont eu leur spermatozoïdes collectés avec succès par micro-TESE, avec un SRR de 67,4%. Les patients ont été répartis en 2 groupes en fonction de leurs résultats à la micro-TESE (obtention de spermatozoïdes et non obtention de spermatozoïdes), sans variations significatives de l'âge, du volume testiculaire, des taux de FSH, LH ou de T entre les 2 groupes. Aucune différence significative du SRR n'a été retrouvée entre les groupes dans lesquels les patients ont été classés en fonction des plages de valeurs de référence des hormones reproductives. La régression logistique binaire a été utilisée pour explorer l'absence d'effets significatifs de l'âge, du volume testiculaire, des taux de FSH, de LH et de T sur la récupération de spermatozoïdes chez les patients bénéficiant d'une micro-TESE. Dans le groupe de biopsie diagnostique testiculaire préopératoire, les groupes avec récupération de spermatozoïdes et sans récupération de spermatozoïdes avaient respectivement des SRR de 90,1% et 65,1%. Plus important encore, il n'y avait pas de différence significative du SRR entre le groupe de biopsie diagnostique testiculaire préopératoire négatif et le groupe sans biopsie diagnostique testiculaire préopératoire (65,1 vs 63,8%, p = 0,855). CONCLUSIONS: Il existe une forte probabilité de récupération réussie de spermatozoïdes testiculaires chez les hommes bénéficiant d'une micro-TESE. Dans ce groupe de patients, l'âge, le volume testiculaire, les taux de FSH, de LH et de T ont peu d'incidence sur le résultat de la micro-TESE. Chez les patients dont la TESA préopératoire a révélé l'absence de spermatozoïdes, la probabilité d'obtenir des spermatozoïdes par micro-TESE est restée élevée (65,1%); les résultats négatifs d'une TESA ne semblaient pas influencer le SRR (63,8%) chez les patients bénéficiant d'une micro-TESE.

Development of a predictive model for increasing sperm retrieval success by microdissection testicular sperm extraction in patients with nonobstructive azoospermia.

Microdissection testicular sperm extraction (micro-TESE) is widely used to treat nonobstructive azoospermia. However, a good prediction model is required to anticipate a successful sperm retrieval rate before performing micro-TESE. This retrospective study analyzed the clinical records of 200 nonobstructive azoospermia patients between January 2021 and December 2021. The backward method was used to perform binary logistic regression analysis and identify factors that predicted a successful micro-TESE sperm retrieval. The prediction model was constructed using acquired regression coefficients, and its predictive performance was assessed using the receiver operating characteristic curve. In all, 67 patients (sperm retrieval rate: 33.5%) underwent successful micro-TESE. Follicle-stimulating hormone, anti-Müllerian hormone, and inhibin B levels varied significantly between patients who underwent successful and unsuccessful micro-TESE. Binary logistic regression analysis yielded the following six predictors: anti-Müllerian hormone (odds ratio [OR] = 0.902, 95% confidence interval [CI]: 0.821-0.990), inhibin B (OR = 1.012, 95% CI: 1.001-1.024), Klinefelter's syndrome (OR = 0.022, 95% CI: 0.002-0.243), Y chromosome microdeletion (OR = 0.050, 95% CI: 0.005-0.504), cryptorchidism with orchiopexy (OR = 0.085, 95% CI: 0.008-0.929), and idiopathic nonobstructive azoospermia (OR = 0.031, 95% CI: 0.003-0.277). The prediction model had an area under the curve of 0.720 (95% CI: 0.645-0.794), sensitivity of 65.7%, specificity of 72.2%, Youden index of 0.379, and cut-off value of 0.305 overall, indicating good predictive value and accuracy. This model can assist clinicians and nonobstructive azoospermia patients in decision-making and avoiding negative micro-TESE results.

Psychological status of infertile men during the Coronavirus Disease 2019 Pandemic in China: a cross-sectional investigation.

BACKGROUND: The coronavirus disease 2019 (COVID-19) outbreak has had a widespread and profound impact on people's mental health. The factors associated with mental symptoms among men diagnosed with infertility, a disease closely related to psychological conditions, remain unclear. The aim of this study is to investigate the risk factors associated with mental symptoms among infertile Chinese men during the pandemic. RESULTS: A total of 4,098 eligible participants were recruited in this cross-sectional, nationwide study, including 2,034 (49.6%) with primary infertility and 2,064 (50.4%) with secondary infertility. The prevalence of mental health conditions was 36.3%, 39.6%, and 6.7% for anxiety, depression, and post-pandemic stress, respectively. Sexual dysfunction is associated with a higher risk with adjusted odds ratios (ORs) of 1.40 for anxiety, 1.38 for depression, and 2.32 for stress. Men receiving infertility drug therapy displayed a higher risk for anxiety (adjusted OR, 1.31) and depression (adjusted OR, 1.28) symptoms, while those receiving intrauterine insemination had a lower risk of anxiety (adjusted OR, 0.56) and depression (adjusted OR, 0.55) symptoms. CONCLUSION: The COVID-19 pandemic has had a significant psychological impact on infertile men. Several psychologically vulnerable populations were identified, including individuals with sexual dysfunction, respondents receiving infertility drug therapy, and those experiencing control measures for COVID-19. The findings provide a comprehensive profile of the mental health status of infertile Chinese men during the COVID-19 outbreak and provide potential psychological intervention strategies.

RéSUMé: CONTEXTE: L'épidémie de maladie à coronavirus 2019 (COVID-19) a eu un impact étendu et profond sur la santé mentale des gens. Les facteurs associés aux symptômes mentaux chez les hommes diagnostiqués comme infertiles, une maladie étroitement liée aux conditions psychologiques, restent flous. L'objectif de cette étude est d'étudier les facteurs de risque associés aux symptômes mentaux chez les hommes chinois infertiles pendant la pandémie. RéSULTATS: Au total, 4 098 participants admissibles ont été recrutés dans cette étude transversale à l'échelle nationale, dont 2 034 (49,6%) présentaient une infertilité primaire et 2 064 (50,4%) une infertilité secondaire. La prévalence des problèmes de santé mentale était respectivement de 36,3 %, 39,6 % et 6,7 % pour l'anxiété, la dépression, et le stress postpandémique. La dysfonction sexuelle est associée à un risque plus élevé avec des odds ratios ajustés (OR) de 1,40 pour l'anxiété, 1,38 pour la dépression et 2,32 pour le stress. Les hommes recevant un traitement médicamenteux contre l'infertilité présentaient un risque plus élevé de symptômes d'anxiété (OR ajusté, 1,31) et de dépression (OR ajusté, 1,28), alors que ceux dont le traitement consistait à faire des inséminations intra-utérines présentaient un risque plus faible de symptômes d'anxiété (OR ajusté, 0,56) et de dépression (OR ajusté, 0,55). CONCLUSIONS: La pandémie de COVID-19 a eu un impact psychologique important sur les hommes infertiles. Plusieurs populations psychologiquement vulnérables ont été identifiées, notamment les personnes souffrant de dysfonction sexuelle, les hommes recevant un traitement médicamenteux contre l'infertilité, et ceux subissant des mesures de contrôle de la COVID-19. Les résultats fournissent un profil complet de l'état de santé mentale des hommes Chinois infertiles pendant l'épidémie de COVID-19 et fournissent des stratégies potentielles d'intervention psychologique.

Synthesis and anti-HIV activity of non-nucleoside reverse-transcriptase inhibitor DB02 phosphate derivatives based on water-soluble optimization.

To improve the water solubility of anti-human immunodeficiency virus (HIV) agent DB02, an excellent non-nucleoside reverse-transcriptase inhibitor (NNRTI) obtained in our previous efforts, we designed and synthesized four phosphate derivatives of DB02 based on the molecular model of DB02 with RT. Here, the antiviral activity of these four derivatives was detected, leading to the discovery of compound P-2, which possessed a superior potency to the lead compound DB02 against wild-type HIV-1 and a variety of HIV-resistant mutant viruses significantly. Furthermore, the water solubility of P-2 was nearly 17 times higher than that of DB02, and the pharmacokinetic test in rats showed that P-2 demonstrate significantly improved oral bioavailablity of 14.6%. Our study showed that the introduction of a phosphate ester group at the end of the C-2 side chain of DB02 was beneficial to the improvement of its antiviral activity and pharmacokinetic properties, which provided a promising lead for the further development of S-DACOs type of NNRTIs.

Preclinical evaluation of the fluid dynamics and hemocompatibility of the Corheart 6 left ventricular assist device.

BACKGROUND: Corheart 6 (Corheart) is a newly developed magnetically levitated continuous-flow left ventricular assist device currently undergoing multicenter clinical trials in China. Featuring a small size, minimal weight, and low power consumption, the Corheart aims to improve pump hemocompatibility, reduce adverse events, and enhance the quality of life of heart failure patients. METHODS: Computational simulations assessed flow field, shear stress, and washout, while in vitro and in vivo experiments were performed to further demonstrate hemocompatibility. RESULTS: Numerical results show that the flow path in the Corheart blood pump is well designed. There is no significantly high shear stress in the majority of the flow domain. Short secondary flow paths and small pump size (small priming volume) provide good washing (0.049 and 0.165 s to remove 55% and 95% old blood, respectively), allowing low hemolysis levels both in computational and in vitro hemolysis tests (in vitro hemolysis index ranges from 0.00092 ± 0.00006 g/100 L to 0.00134 ± 0.00019 g/100 L). Good hemocompatibility was further evidenced by ten 60-day sheep implants tested with relatively low flow rates of 2.0 ± 0.2 L/min; the results showed no hemolysis or thrombosis. CONCLUSIONS: Numerical and experimental results shed light on the fluid dynamics characteristics and hemocompatibility of the Corheart. It is believed that the Corheart will provide more promising possibilities for minimally invasive implantation techniques and for those patients with a small body surface area.

Inhibin B-to-Anti-Mullerian Hormone Ratio as Noninvasive Predictors of Positive Sperm Retrieval in Idiopathic Non-Obstructive Azoospermia.

Background: The lack of clinically useful biomarkers for predicting micro-TESE outcomes in males with idiopathic NOA. To find clinically reliable serum reproductive hormone markers to predict the outcome of sperm retrieval in men with idiopathic NOA undergoing micro-TESE. Methods: We analyzed the clinical data of 168 men with idiopathic NOA treated with micro-TESE. The clinical predictors of a successful sperm retrieval were put to the test using logistic regression analysis. The clinical net benefit was calculated using a decision-curve analysis, and the predictive power of each variable was assessed using the ROC-derived area under the curve. Result: Between positive group and negative group for sperm retrieval, there was a statistically significant difference in INHB, AMH, and INHB/AMH. AMH, INHB, and INHB/AMH were each independent predictors of successful sperm retrieval, with INHB (OR1.02, p = 0.03), AMH (OR0.85, p = 0.01), INHB/AMH (OR1.08, p < 0.01). The ROC curve determined the optimal cut-off values for serum INHB and INHB/AMH in positive sperm retrieval patients undergoing micro-TESE. 21.51 pg/mL was the cut-off value for INHB. The cut-off value for INHB/AMH was 3.19, which had a 86.3% sensitivity and a 53.8% specificity. Using INHB and INHB/AMH prior to micro-TESE sperm retrieval in idiopathic NOA patients improved the net benefit of positive sperm retrieval, and the net benefit score of INHB/AMH was better than that of INHB, according to decision analysis curves. Conclusion: Serum INHB have predictive value for sperm retrieval outcomes in idiopathic NOA patients undergoing micro-TESE. Combining INHB and AMH, INHB/AMH seems to be a better predictor.

Micro-TESE strategy in patients with NOA caused by AZFc deletion: synchronous or asynchronous?

In the treatment of infertile patients with non-obstructive azoospermia (NOA) caused by the deletion of the azoospermia factor c region (AZFc) on the Y chromosome, synchronous and asynchronous surgical strategies are discussed. Clinical data from NOA patients with the AZFc deletion who underwent micro-TESE were analyzed retrospectively. The sperm retrieval rate (SRR) and sperm utilization rate of synchronous and asynchronous operation groups were followed up and compared. The fertilization rate, high-quality embryo rate, clinical pregnancy rate, abortion rate, and cumulative live birth rate of ICSI in patients with successful sperm retrieval were compared between the two groups. The two groups had sperm utilization rates of 98.9% (93/94) and 50.0% (14/28), respectively. The asynchronous group's sperm consumption rates were much lower than those of the synchronous operation group. Fertilization rate, high-quality embryo rate, clinical pregnancy rate of fresh transfer cycle, abortion rate, and cumulative live birth rate of patients in the synchronous operation group with fresh sperm, and the asynchronous operation group with thawed sperm, respectively, were 30.6% vs 33.8%, 33.8% vs 40.7%, 40.0% vs 12.5%, 30.4% vs 7.1%. Between the two groups, there was no significant difference. This suggests that individuals with NOA caused by the AZFc deletion have a high possibility of successfully acquiring sperm using micro-TESE and ICSI to conceive their own offspring. Synchronous micro-TESE is recommended to improve sperm utilization rate and the cumulative live birth rate.

Identification of 6ω-cyclohexyl-2-(phenylamino carbonylmethylthio)pyrimidin-4(3H)-ones targeting the ZIKV NS5 RNA dependent RNA polymerase.

Zika virus (ZIKV), a mosquito-borne flavivirus, is a global health concern because of its association with severe neurological disorders such as neonatal microcephaly and adult Guillain-Barre syndrome. Although many efforts have been made to combat ZIKV infection, there is currently no approved vaccines or antiviral drugs available and there is an urgent need to develop effective anti-ZIKV agents. In this study, 26 acetylarylamine-S-DACOs derivatives were prepared, and eight of them were found to have inhibitory activity against Zika virus. Among these substances, 2-[(4-cyclohexyl-5-ethyl-6-oxo-1,6-dihydropyrimidin-2-yl)thio]-N-(3,5-difluorophenyl)acetamide (4w) with the best anti-ZIKV activity was selected for in-depth study of antiviral activity and mechanism of action. Here, we discovered 4w targeted on the ZIKV NS5 RNA -dependent RNA polymerase (RdRp), which exhibited good in vitro antiviral activity without cell species specificity, both at the protein level and at the RNA level can significantly inhibit ZIKV replication. Preliminary molecular docking studies showed that 4w preferentially binds to the palm region of NS5A RdRp through hydrogen bonding with residues such as LYS468, PHE466, GLU465, and GLY467. ZIKV NS5 RdRp enzyme activity experiment showed that 4w could directly inhibit ZIKV RdRp activity with EC50 = 11.38 ± 0.51 µM. In antiviral activity studies, 4w was found to inhibit ZIKV RNA replication with EC50 = 6.87 ± 1.21 µM. ZIKV-induced plaque formation was inhibited with EC50 = 7.65 ± 0.31 µM. In conclusion, our study disclosed that acetylarylamine-S-DACOs is a new active scaffolds against ZIKV, among which compound 4w was proved to be a potent novel anti-ZIKV compound target ZIKV RdRp protein. These promising results provide a future prospective for the development of ZIKV RdRp inhibitors.

Theoretical insights into the gaseous and heterogeneous reactions of halogenated phenols with ˙OH radicals: mechanism, kinetics and role of (TiO2)n clusters in degradation processes.

Halogenated phenols are highly toxic chemicals with serious health risks, and the removal of these persistent environmental pollutants remains a challenge. Based on quantum chemistry calculations, the homogeneous/heterogeneous degradation mechanism and kinetics of C6X5OH (X = F, Cl, and Br) initiated by ˙OH radicals in the gas phase and TiO2 cluster surfaces are investigated in this work. Four ˙OH-addition and one proton-coupled electron-transfer (PCET) reaction channels for each halogenated phenol were found and the ˙OH-addition channels were more favorable than the PCET pathway without TiO2 clusters. At 296 K, the calculated total rate constant for ˙OH with C6F5OH in the atmosphere well agreed with the limited experimental data of (6.88 ± 1.37) × 10-12 cm3 molecule-1 s-1. The lifetimes of C6F5OH, C6Cl5OH, and C6Br5OH were about 12.04-12.86 h at 296 K, which favored their medium-range transport in the atmosphere. In the presence of (TiO2)n clusters (n = 4, 6, 8, 12, and 16), the PCET mechanism for hydrogen transfer reaction of C6F5OH with ˙OH radicals was changed from the previous four-electron/three-center into four-electron/two-center, which results in the PCET pathway becoming more favorable than the ˙OH-addition channels. Meanwhile, the heterogeneous degradation rate constants of C6F5OH were accelerated by more than 10 orders of magnitude within 200-430 K compared with those of the naked reaction. The effects of (TiO2)n cluster (n = 4, 6, 8, 12, and 16) size on the degradation rates were analyzed at 200-430 K, and the reaction on the (TiO2)8 cluster had a faster rate. The subsequent reactions including the bond cleavage of the benzene ring and O2 addition or abstraction were studied. This work provides new insights into halogenated aromatic atmospheric chemistry and nanoscale TiO2 photocatalysis in air or wastewater management.

Identification and characterization of circular RNAs in the testicular tissue of patients with non-obstructive azoospermia.

Circular RNAs (circRNAs) are highly conserved and ubiquitously expressed noncoding RNAs that participate in multiple reproduction-related diseases. However, the expression pattern and potential functions of circRNAs in the testes of patients with non-obstructive azoospermia (NOA) remain elusive. In this study, according to a circRNA array, a total of 37 881 circRNAs were identified that were differentially expressed in the testes of NOA patients compared with normal controls, including 19 874 upregulated circRNAs and 18 007 downregulated circRNAs. Using quantitative real-time polymerase chain reaction (qRT-PCR) analysis, we confirmed that the change tendency of some specific circRNAs, including hsa_circ_0137890, hsa_circ_0136298, and hsa_circ_0007273, was consistent with the microarray data in another larger sample. The structures and characteristics of these circRNAs were confirmed by Sanger sequencing, and fluorescence in situ hybridization revealed that these circRNAs were primarily expressed in the cytoplasm. Bioinformatics analysis was used to construct the competing endogenous RNA (ceRNA) network, and numerous miRNAs that could be paired with circRNAs validated in this study were reported to be vital for spermatogenesis regulation. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses indicated that genes involved in axoneme assembly, microtubule-based processes, and cell proliferation were significantly enriched. Our data suggest that there are aberrantly expressed circRNA profiles in patients with NOA and that these circRNAs may help identify key diagnostic and therapeutic molecular biomarkers for NOA patients.

Traditional fermented soybean products: processing, flavor formation, nutritional and biological activities.

Traditional fermented soybean food has emerged as an important part of people's dietary structure because of the unique flavors and improved health benefit. During fermentation, the nutrients in soybean undergo a series of biochemical reactions catalyzed naturally by microorganism secreted enzymes. Thereafter, many functional and bioactive substances such as bioactive peptides, unsaturated fatty acids, free soy isoflavones, vitamins and minerals are produced, making fermented soy products more advantageous in nutrition and health. This review comprehensively discusses the historical evolution, distribution, traditional fermentation processing, main sources and characteristics of fermented strains, flavor components, nutritional properties, and biological activities of four traditional fermented soybean foods including douchi, sufu, dajiang, and soy sauce. In the end, we introduce four major challenges encountered by traditional fermented soybean foods including high salt content, formation of biogenic amine, the presence of pathogenic microorganisms and mycotoxins, and quality inconsistency. We conclude that the establishment of scientific quality standard and innovated fermentation processing is the potential solutions to combat the issues and improve the safety of traditional fermented soybean products.

Nonspecific interactions between SpCas9 and dsDNA sites located downstream of the PAM mediate facilitated diffusion to accelerate target search.

RNA-guided Streptococcus pyogenes Cas9 (SpCas9) is a sequence-specific DNA endonuclease that works as one of the most powerful genetic editing tools. However, how Cas9 locates its target among huge amounts of dsDNAs remains elusive. Here, combining biochemical and single-molecule fluorescence assays, we revealed that Cas9 uses both three-dimensional and one-dimensional diffusion to find its target with high efficiency. We further observed surprising apparent asymmetric target search regions flanking PAM sites on dsDNA under physiological salt conditions, which accelerates the target search efficiency of Cas9 by ∼10-fold. Illustrated by a cryo-EM structure of the Cas9/sgRNA/dsDNA dimer, non-specific interactions between DNA ∼8 bp downstream of the PAM site and lysines within residues 1151-1156 of Cas9, especially lys1153, are the key elements to mediate the one-dimensional diffusion of Cas9 and cause asymmetric target search regions flanking the PAM. Disrupting these non-specific interactions, such as mutating these lysines to alanines, diminishes the contribution of one-dimensional diffusion and reduces the target search rate by several times. In addition, low ionic concentrations or mutations on PAM recognition residues that modulate interactions between Cas9 and dsDNA alter apparent asymmetric target search behaviors. Together, our results reveal a unique searching mechanism of Cas9 under physiological salt conditions, and provide important guidance for both in vitro and in vivo applications of Cas9.

Mitocytosis, a migrasome-mediated mitochondrial quality-control process.

Damaged mitochondria need to be cleared to maintain the quality of the mitochondrial pool. Here, we report mitocytosis, a migrasome-mediated mitochondrial quality-control process. We found that, upon exposure to mild mitochondrial stresses, damaged mitochondria are transported into migrasomes and subsequently disposed of from migrating cells. Mechanistically, mitocytosis requires positioning of damaged mitochondria at the cell periphery, which occurs because damaged mitochondria avoid binding to inward motor proteins. Functionally, mitocytosis plays an important role in maintaining mitochondrial quality. Enhanced mitocytosis protects cells from mitochondrial stressor-induced loss of mitochondrial membrane potential (MMP) and mitochondrial respiration; conversely, blocking mitocytosis causes loss of MMP and mitochondrial respiration under normal conditions. Physiologically, we demonstrate that mitocytosis is required for maintaining MMP and viability in neutrophils in vivo. We propose that mitocytosis is an important mitochondrial quality-control process in migrating cells, which couples mitochondrial homeostasis with cell migration.

Altered gene expression profiles of testicular tissues from azoospermic patients with maturation arrest.

Maturation arrest is a common cause of male infertility which has caused worldwide concern, and its pathophysiological process remains further elucidation. Our study aimed to identify genetic characteristics of maturation arrest by comparing gene expression between maturation arrest and normal samples using microarray technology. A total of 6,373 genes were identified differentially expressed (p < .05, fold change > 2.0 or <-2.0) and 1,594 genes were selected as statistically significant after Bonferroni correction, including 419 up-regulated and 1,175 down-regulated genes. Microarray data were validated by quantitative reverse transcriptase-polymerase chain reaction. Bioinformation analysis was performed to explore genetic function of statistically significant genes. Gene Ontology results showed the statistically significant genes enriched in sexual reproduction, spermatogenesis and male gamete generation. Reactome pathway analysis highlighted the olfactory signalling pathway, fertilisation, developmental biology, etc. One module and eight hub genes were found to be involved in ubiquitin-mediated proteolysis and may affect as indicators of spermatogenic process through protein-protein interaction analysis. Our study provided a comprehensive genetic characteristic of differential expressed genes in testicular tissues from maturation arrest patients and speculated several genes as potential indicators of disease.

Altered microRNA profiles of testicular biopsies from patients with nonobstructive azoospermia.

Many studies have shown that microRNAs (miRNAs) play vital roles during the spermatogenesis. However, little is known about the altered miRNA profiles of testicular tissues in nonobstructive azoospermia (NOA). Using microarray technology, the miRNA expression profiles of testicular biopsies from patients with NOA and of normal testicular tissues were determined. Bioinformatics analyses were conducted to predict the enriched biological processes and functions of identified miRNAs. The microarray data were validated by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), the results of which were then validated with a larger sample size. Correlations between the miRNA expression levels and clinical characteristics were analyzed. Receiver operating characteristic (ROC) curve analysis was used to evaluate the diagnostic ability of miRNAs for azoospermia. Hierarchical clustering showed that 129 miRNAs were significantly differentially expressed between the NOA and control groups. Bioinformatics analysis indicated that the differentially expressed miRNAs were involved in spermatogenesis, cell cycle, and mitotic prometaphase. In the subsequent qRT-PCR assays, the selected miRNA expression levels were consistent with the microarray results, and similar validated results were obtained with a larger sample size. Some clinical characteristics were significantly associated with the expression of certain miRNAs. In particular, we identified a combination of two miRNAs (miR-10b-3p and miR-34b-5p) that could serve as a predictive biomarker of azoospermia. This study provides altered miRNA profiles of testicular biopsies from NOA patients and examines the roles of miRNAs in spermatogenesis. These profiles may be useful for predicting and diagnosing the presence of testicular sperm in individuals with azoospermia.

Nanotechnology-assisted adipose-derived stem cell (ADSC) therapy for erectile dysfunction of cavernous nerve injury: In vivo cell tracking, optimized injection dosage, and functional evaluation.

Stem cell therapy is a potentially promising option for erectile dysfunction; however, its risk of tumorigenicity is a clinical hurdle and the risk is positively related to the number of injected cells. Our previous study showed that nanotechnology improved adipose-derived stem cell (ADSC) therapy for erectile dysfunction of cavernous nerve injury (CNI) by attracting cells in the corpus cavernosum. These results indicated the possibility of using a reduced dosage of ADSCs for intracavernous injection. In this exploratory study, we used lower dosage (2 × 105 cells) of ADSCs for intracavernous injection (ICI) and the nanotechnology approach. Intracavernous pressure and mean arterial pressure were measured at day 28 to assess erectile function. The low-dose ADSC therapy group showed favorable treatment effects, and nanotechnology further improved these effects. In vivo imaging of ICI cells revealed that the fluorescein signals of NanoShuttle-bound ADSCs (NanoADSCs) were much stronger than those of ADSCs at days 0, 1, and 3. Both immunofluorescence and Western blot analysis showed a significant increase in smooth muscle, endothelium, and nerve tissue in the ADSC group compared to that in the CNI group; further improvement was achieved with assisted nanotechnology. These findings demonstrate that nanotechnology can be used to further improve the effect of small dosage of ADSCs to improve erectile function. Abundant NanoADSCs remain in the corpus cavernosum in vivo for at least 3 days. The mechanism of erectile function improvement may be related to the regeneration of the smooth muscle, endothelium, and nerve tissues.