Transcriptomics-based identification of TYROBP and TLR8 as novel macrophage-related biomarkers for the diagnosis of acute rejection after kidney transplantation.

Macrophages play an important role in the development and progression of acute rejection after kidney transplantation. The study aims to investigate the biological role and significance of macrophage-associated genes (MAG) in acute rejection after kidney transplantation. We utilized transcriptome sequencing results from public databases related to acute rejection of kidney transplantation for comprehensive analysis and validation in animal experiments. We found that a large number of immune-related signaling pathways are activated in acute rejection. PPI protein interaction networks and machine learning were used to establish a Hub gene consisting of TYROBP and TLR8 for the diagnosis of acute rejection. The single-gene GSEA enrichment analysis and immune cell correlation analysis revealed a close correlation between the expression of Hub genes and immune-related biological pathways as well as the expression of multiple immune cells. In addition, the study of TF, miRNAs, and drugs provided a theoretical basis for regulating and treating the Hub genes in acute rejection. Finally, the animal experiments demonstrated once again that acute rejection can aggravate kidney tissue damage, apoptosis level, and increase the release of inflammatory factors. We established and validated a macrophage-associated diagnostic model for acute rejection after kidney transplantation, which can accurately diagnose the biological alterations in acute rejection after kidney transplantation.

Maternal Diabetes and Risk of Hypospadias: A Systemic Review and Meta-Analysis.

INTRODUCTION: This study aimed to investigate the association between maternal diabetes and the risk of hypospadias in male infants, as the relationship between them remains uncertain. METHODS: To comprehensively evaluate the association between pregestational diabetes mellitus and gestational diabetes mellitus with hypospadias, we conducted a systematic review and meta-analysis. A thorough literature search was conducted, encompassing relevant publications published prior to January 2023. Crude odds ratios (ORs) and their corresponding 95% confidence intervals (CIs) were calculated using a random-effects model. RESULTS: Our meta-analysis comprised a total of 13 studies, 11 of which investigated the relationship between pregestational diabetes mellitus and hypospadias, while 9 studies explored the association between gestational diabetes mellitus and hypospadias. Notably, these investigations yielded compelling evidence of significant positive associations between pregestational diabetes mellitus and hypospadias (OR = 1.51, 95% CI = 1.13-2.03), as well as between gestational diabetes mellitus and hypospadias (OR = 1.18, 95% CI = 1.04-1.35). CONCLUSION: Our findings suggest that both pregestational diabetes mellitus and gestational diabetes mellitus are associated with an increased risk of hypospadias in offspring. Further investigations are needed to explore the optimal range of blood glucose during pregnancy that minimizes the risk of congenital malformation in the fetus, as well as to develop more effective measures for glycemic control in pregnant women.

Differentiation Potential of Hypodifferentiated Subsets of Nephrogenic Rests and Its Relationship to Prognosis in Wilms Tumor.

Background Wilms tumor (WT) is highly curable, although anaplastic histology or relapse imparts a worse prognosis. Nephrogenic rests (NR) associated with a high risk of developing WT are abnormally retained embryonic kidney precursor cells. Methods After pseudo-time analysis using single-cell RNA sequencing (scRNA-seq) data, we generated and validated a WT differentiation-related gene (WTDRG) signature to predict overall survival (OS) in children with a poor OS. Results A differentiation trajectory from NR to WT was identified and showed that hypodifferentiated subsets of NR could differentiate into WT. Classification of WT children with anaplastic histology or relapse based on the expression patterns of WTDRGs suggested that patients with relatively high levels of hypodifferentiated NR presented a poorer prognosis. A WTDRG-based risk model and a clinically applicable nomogram was developed. Conclusions These findings may inform oncogenesis of WT and interventions directed toward poor prognosis in WT children of anaplastic histology or relapse.

Transcriptome-based exploration of potential molecular targets and mechanisms of selenomethionine in alleviating renal ischemia-reperfusion injury.

Renal ischemia-reperfusion injuries (IRIs) are one of the leading causes of acute kidney injuries (AKIs). Selenium, as an essential trace element, is able to antioxidant stress and reduces inflammatory responses. The regulation mechanism of selenomethionine, one of the major forms of selenium intake by humans, is not yet clear in renal IRIs. Therefore, we aimed to explore the key targets and related mechanisms of selenomethionine regulation in renal IRIs and provide new ideas for the treatment of selenomethionine with renal IRIs. We used transcriptome sequencing data from public databases as well as animal experiments to explore the key target genes and related mechanisms regulated by selenomethionine in renal IRI. We found that selenomethionine can effectively alleviate renal IRI by a mechanism that may be achieved by inhibiting the MAPK signaling pathway. Meanwhile, we also found that the key target of selenomethionine regulation in renal IRI might be selenoprotein GPX3 based on the PPI protein interaction network and machine learning. Through a comprehensive analysis of bioinformatic techniques and animal experiments, we found that Gpx3 might serve as a key gene for the regulation of selenomethionine in renal IRIs. Selenomethionine may exert a protective effect against renal IRI by up-regulating GPX3, inhibiting the MAPK signaling pathway, increased production of antioxidants, decreasing inflammation levels, mitigation of apoptosis in renal tubular epithelial cells, this reduces renal histopathological damage and protects renal function. Providing a theoretical basis for the mechanism of selenomethionine actions in renal IRIs.

Human umbilical cord mesenchymal stem cell exosomes alleviate acute kidney injury by inhibiting pyroptosis in rats and NRK-52E cells.

Human umbilical cord mesenchymal stem cells (hucMSCs) have been shown to improve kidney injury. Exosomes have been indicated to be important mediators of renal protection in MSC therapy. In spite of this, the mechanism remains unclear. Our study investigated how exosomes derived from hucMSCs (hucMSC-Ex) improve acute kidney injury (AKI). Exosomes were extracted by using an ultracentrifugation technique and identified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blot. Twenty-four male SD rats were randomly divided into four groups: sham group, sham + hucMSC-Ex group, ischemia-reperfusion injury (IRI) group, and IRI + hucMSC-Ex group. In vitro, we treated rat proximal renal tubular epithelial cell line (NRK-52E) with cisplatin to mimic in vivo models of AKI. The NRK-52E cells were treated with or without 160 µg/mL hucMSC-Ex, and 1 µg/mL cisplatin was added after 9 h. Cells were harvested after 24 h. In the IRI group, the levels of serum creatinine (Scr) and blood urea nitrogen (BUN) were increased; renal tubules were dilated, epithelial cells were vacuolated, and collagen fibers were deposited in the renal interstitium. After treatment with cisplatin, the NRK-52E cells displayed pyroptotic morphology characterized by pyroptotic bodies. The protein expression levels of fibronectin, α-smooth muscle actin (α-SMA), vimentin, gasdermin D (GSDMD), caspase-1, interleukin-1 (IL-1ß) and NLRP3 in IRI tissues and in cisplatin treatment NRK-52E cells were significantly upregulated. However, after the hucMSC-Ex intervention, kidney injury was effectively improved in vivo and in vitro. The current study shows that pyroptosis is involved in AKI and that hucMSC-Ex improves AKI by inhibiting pyroptosis.

Exploring the role of Hmox1 in ferroptosis and immune infiltration during renal ischemia-reperfusion injury.

Ischemia-reperfusion injury (IRI) is inevitable in kidney transplantations and, as a complex pathophysiological process, it can be greatly impacted by ferroptosis and immune inflammation. Our study aimed to identify the biomarkers of renal IRI (RIRI) and elucidate their relationship with immune infiltration. In this study, the GSE148420 database was used as a training set to analyze differential genes and overlap them with ferroptosis-related genes to identify hub genes using a protein-protein interaction (PPI) network, the least absolute shrinkage and selection operator (LASSO), and random forest algorithm (RFA). We verified the hub gene and ferroptosis-related phenotypes in a verification set and animal experiments involving unilateral IRI with contralateral nephrectomy in rats. Gene set enrichment analysis (GSEA) of single genes was conducted according to the hub gene to predict related endogenous RNAs (ceRNAs) and drugs to establish a network. Finally, we used the Cibersort to analyze immunological infiltration and conducted Spearman's correlation analysis. We identified 5456 differential genes and obtained 26 ferroptosis-related differentially expressed genes. Through PPI, LASSO, and RFA, Hmox1 was identified as the only hub gene and its expression levels were verified using verification sets. In animal experiments, Hmox1 was verified as a key biomarker. GSEA of single genes revealed the seven most related pathways, and the ceRNAs network included 138 mRNAs and miRNAs. We predicted 11 related drugs and their three-dimensional structural maps. Thus, Hmox1 was identified as a key biomarker and regulator of ferroptosis in RIRI and its regulation of ferroptosis was closely related to immune infiltration.

The efficacy of robotic-assisted laparoscopic pyeloplasty for pediatric ureteropelvic junction obstruction: a systematic review and meta-analysis.

To evaluate the safety and effectiveness of robotic-assisted laparoscopic pyeloplasty (RALP) for treating pediatric ureteropelvic junction obstruction through an extensive comparison of RALP, open pyeloplasty (OP) and laparoscopic pyeloplasty (LP). We conducted a comprehensive search of the following databases: PubMed, Excerpta Medica Database, Cochrane Library, Web of Science database, China National Knowledge Infrastructure, WanFang Data, and China Biology Medical Disc. Baseline data were compared, the sources of heterogeneity were assessed, and publication biases were detected. This study was registered with PROSPERO (CRD42023415667). 26 studies with 6074 cases performing pyeloplasty were included, and the overall data are comparable. Our analysis showed no significant difference in success rate and postoperative complications between RALP and OP, and RALP is associated with a shorter length of stay (LOS) (MD - 1.00 95%CI - 1.45 to - 0.55, p < 0.0001). In addition, compared to LP, RALP was associated with a shorter anastomosis time (MD - 18.35 95%CI - 29.88 to - 6.82, p = 0.002) and fewer postoperative analgesics (MD - 0.09 95% CI - 0.18 to - 0.01, p = 0.03); however, RALP has a longer operative time (OT) (MD 52.39, 95% CI 39.75-65.03, p < 0.00001) and higher cost. The heterogeneity of OT may be influenced by factors, such as age and region, while the heterogeneity of LOS primarily stems from regional differences. No significant publication bias was detected. Our meta-analysis shows that RALP can be an alternative to OP and LP with a high success rate, minimal postoperative complications, and shorter LOS. In addition, RALP contributes to reduce anastomosis time and postoperative analgesic drugs. However, further well-designed, large-scale, randomized controlled trials with additional parameters are needed to conduct a more comprehensive analysis of heterogeneity.

Inhibition of the NF-κB Signaling Pathway Alleviates Pyroptosis in Bladder Epithelial Cells and Neurogenic Bladder Fibrosis.

Most children with a neurogenic bladder (NB) have bladder fibrosis, which causes irreversible bladder dysfunction and damage to the upper urinary tract. However, the mechanism of bladder fibrosis remains unclear. This study aimed to investigate the underlying causes of bladder fibrosis. Here, the lumbar 6 (L6) and sacral 1 (S1) spinal nerves of Sprague Dawley rats were severed bilaterally to establish NB models. Using RNA-seq, we discovered that the NF-κB signaling pathway and inflammation were upregulated in spinal cord injury (SCI)-induced bladder fibrosis. Subsequent Western blotting, enzyme-linked immunosorbent assays, immunohistochemical staining, and immunofluorescence staining verified the RNA-seq findings. To further clarify whether the NF-κB signaling pathway and pyroptosis were involved in bladder fibrosis, a TGF-ß1-treated urinary epithelial cell line (SV-HUC-1 cells) was used as an in vitro model. Based on the results of RNA-seq, we consistently found that the NF-κB signaling pathway and pyroptosis might play important roles in TGF-ß1-treated cells. Further experiments also confirmed the RNA-seq findings in vitro. Moreover, using the NLRP3 inhibitor MCC950 rescued TGF-ß1-induced fibrosis, and the NF-κB signaling pathway inhibitor BAY 11-7082 effectively rescued TGF-ß1-induced pyroptosis and the deposition of extracellular matrix by SV-HUC-1 cells. In summary, our research demonstrated for the first time that the NF-κB signaling pathway inhibition rescued bladder epithelial cells pyroptosis and fibrosis in neurogenic bladders.

Laparoscopic versus open inguinal hernia repair in children: A systematic review.

PURPOSE: Considerable debates exist regarding the preferable technique to repair a paediatric inguinal hernia (PIH). This systematic review aims to compare the efficacy and safety of laparoscopic herniorrhaphy (LH) and open herniorrhaphy (OH) in PIH. METHODS: The randomised controlled trials (RCTs) that compared the outcomes of LH and OH in PIH without region and language restrictions searched from the following databases: PubMed, Web of Science Database, Cochrane Library, SciELO Citation Index, Russian Science Citation Index, China National Knowledge Infrastructure, WanFang Data and China Science and Technology Journal Database. RESULTS: A total of 13 RCTs that involving 1207 patients included in the review. The LH displayed a shorter operative time for bilateral hernia repair (weighted mean difference = -8.23, 95% confidence interval [CI]: -11.22~-5.23, P < 0.00001), a lower complication rate (odds ratio [OR] = 0.32, 95% CI: 013-0.83, P = 0.02) along with a lower wound infection (OR = 0.14, 95% CI: 0.04-0.55, P = 0.005) and major male-specific post-operative complications (OR = 0.10, 95% CI: 0.04-0.24, P < 0.00001) and a less contralateral metachronous inguinal hernia (CMIH) incidence rate (OR = 0.09, 95% CI: 0.02-0.42, P = 0.002). No significant difference was found for unilateral operative time, time to full recovery, length of hospital stay, recurrence and hydrocele rates between the two techniques. CONCLUSION: The present review reiterates that both the LH and OH techniques for the PIH repair are comparable. However, in some aspects, the LH is superior to the OH in terms of operative time for bilateral hernias, post-operative complications rate and CMIH incidence rate. Rigorously designed RCTs are anticipated to confirm the clinical effects of both LH and OH.

Demographic, clinical, and socioeconomic factors associated with delayed diagnosis and management of pediatric testicular torsion in West China: a retrospective study of 301 cases in a single tertiary children's hospital.

BACKGROUND: To investigate the association between geographic, clinical, socioeconomic factors and delayed management of pediatric testicular torsion (TT) in West China. METHODS: A retrospective study was conducted on TT at Children's Hospital of Chongqing Medical University in West China from November 2004 to December 2020. Univariate analysis and logistic regression analysis were conducted to determine the association between these factors and delayed management of TT. RESULTS: A total of 301 cases were included in this study. The misdiagnosis rate of TT in primary, secondary healthcare units and tertiary hospitals was 93.8, 71.1, and 8.9%, respectively. Approximately 26.9% of TT boys received timely surgical management (within 12 h from symptoms inset to surgery). Logistic regression analyses suggested the following factors were associated with delayed repair of TT: age less than 6 years (P = 0.001), with a history of symptoms progress (P = 0.001) or former treatment (P <0.001), absence of other diagnosis (P = 0.011) and those boys living far away from the main city zones (P <0.001). CONCLUSIONS: Delayed surgical management for TT was more likely for boys with age less than 6 years, the absence of other diagnosis, with a history of former treatment or symptoms progress, and those living far away from the main city zone. To maximize the possibility of timely surgical management for TT, it is vital to strengthen the public awareness of TT and conduct continuously re-education and update physicians working at primary and secondary healthcare units.

Maternal smoking during pregnancy and risk of cryptorchidism: a systematic review and meta-analysis.

The risk factors for undescended testes in male infants and the underlying pathogenesis still remain unclear. The aim of this study is to identify the relationship between maternal smoking during pregnancy and risk of cryptorchidism. A systematic review was conducted using appropriate search terms to identify articles pertaining to maternal smoking during pregnancy and risk of cryptorchidism. Entries up to December 23, 2017 were taken into consideration, without any language or regional restriction. The crude ORs and their 95% CIs were computed by using the fixed-effect model. Twenty studies involving 111,712 infants were included in our meta-analysis. The risk of having a male infant with cryptorchidism was significantly different between mothers who smoked during pregnancy and those who did not (pooled crude OR 1.18, 95% confidence interval [CI] 1.12-1.24, p < 0.00001).Conclusion: Our findings suggest that smoking during pregnancy increased the risk of cryptorchidism by 1.18 times. Further investigations that are well-designed, multicentric studies measuring variables, such as the number of cigarettes smoked in a day and the stage of pregnancy during which the mothers smoked, are necessary to precisely determine the relationship between maternal smoking and risk of cryptorchidism. What is Known: • Preterm and low birth weight have been definitively shown to be risk factors for cryptorchidism. • The relationship between with maternal smoking during pregnancy and risk of cryptorchidism remains controversial all the time. What is New: • Mothers who smoked during pregnancy had a 1.18 times higher risk of having a child with cryptorchidism as compared to those who did not smoke. • Evidence has been found that maternal smoking during pregnancy is a definitive risk factor for cryptorchidism.

Correction to: Maternal smoking during pregnancy and risk of cryptorchidism: a systematic review and meta-analysis.

The publisher regrets that the original version of this article contained an error. Table 1 was shown in the wrong version, thus corrected table is shown in this article. The original article has been corrected.

CCR7 and CD48 as Predicted Targets in Acute Rejection Related to M1 Macrophage after Pediatric Kidney Transplantation.

Background: Kidney transplantation (KT) is the best treatment for end-stage renal disease. Although long and short-term survival rates for the graft have improved significantly with the development of immunosuppressants, acute rejection (AR) remains a major risk factor attacking the graft and patients. The innate immune response plays an important role in rejection. Therefore, our objective is to determine the biomarkers of congenital immunity associated with AR after KT and provide support for future research. Materials and Methods: A differential expression genes (DEGs) analysis was performed based on the dataset GSE174020 from the NCBI gene Expression Synthesis Database (GEO) and then combined with the GSE5099 M1 macrophage-related gene identified in the Molecular Signatures Database. We then identified genes in DEGs associated with M1 macrophages defined as DEM1Gs and performed gene ontology (GO) and Kyoto Encyclopedia of Genomes (KEGG) enrichment analysis. Cibersort was used to analyze the immune cell infiltration during AR. At the same time, we used the protein-protein interaction (PPI) network and Cytoscape software to determine the key genes. Dataset, GSE14328 derived from pediatric patients, GSE138043 and GSE9493 derived from adult patients, were used to verify Hub genes. Additional verification was the rat KT model, which was used to perform HE staining, immunohistochemical staining, and Western Blot. Hub genes were searched in the HPA database to confirm their expression. Finally, we construct the interaction network of transcription factor (TF)-Hub genes and miRNA-Hub genes. Results: Compared to the normal group, 366 genes were upregulated, and 423 genes were downregulated in the AR group. Then, 106 genes related to M1 macrophages were found among these genes. GO and KEGG enrichment analysis showed that these genes are mainly involved in cytokine binding, antigen binding, NK cell-mediated cytotoxicity, activation of immune receptors and immune response, and activation of the inflammatory NF-κB signaling pathway. Two Hub genes, namely CCR7 and CD48, were identified by PPI and Cytoscape analysis. They have been verified in external validation sets, originated from both pediatric patients and adult patients, and animal experiments. In the HPA database, CCR7 and CD48 are mainly expressed in T cells, B cells, macrophages, and tissues where these immune cells are distributed. In addition to immunoinfiltration, CD4+T, CD8+T, NK cells, NKT cells, and monocytes increased significantly in the AR group, which was highly consistent with the results of Hub gene screening. Finally, we predicted that 19 TFs and 32 miRNAs might interact with the Hub gene. Conclusions: Through a comprehensive bioinformatic analysis, our findings may provide predictive and therapeutic targets for AR after KT.

Transcriptomics-based exploration of shared M1-type macrophage-related biomarker in acute kidney injury after kidney transplantation and acute rejection after kidney transplantation.

BACKGROUND: Macrophage type 1 (M1) cells are associated with both acute kidney injury (AKI) during kidney transplantation and acute rejection (AR) after kidney transplantation. Our study explored M1-related biomarkers involved in both AKI and AR and their potential biological functions. METHODS: Based on the Gene Expression Omnibus (GEO) database, the immune cell infiltration levels and differentially expressed genes were examined in AKI and AR in the kidney transplantation; M1-related genes shared in AKI and AR were identified using weighted gene co-expression analysis (WGCNA) system. Subsequently, protein-protein interaction (PPI) networks and machine learning methods to identify Hub genes and construct diagnostic models. Both AKI model and AR rat models were built to validate the expressions of Hub genes and test the injury phenotype, oxidative stress markers, and inflammatory factors. Finally, the transcription factor (TF)-Hub gene and micro-RNA (miRNA)-Hub gene regulatory networks were constructed based on identified Hub genes. RESULTS: Out of 2167 differential expression genes (DEGs) in AKI and 2100 DEGs in AR, four M1-related Hub genes were obtained by PPI networks and machine learning methods, namely GBP2, TYROBP, CCR5, and TLR8. The calibration curves in the nomogram diagnostic model for these four Hub genes suggested the same predictive probability as an ideal model for AKI and AR after kidney transplantation (AUC values of the area under the ROC curve were all >0.7). The same observations were confirmed in ischemia reperfusion injury (IRI) and AR rat models by identifying common four Hub genes (GBP2, TYROBP, TLR8, and CCR5). Western blots showed that these four Hub genes were significantly different in rat models of IRI and AR (all p<0.05). Compared with the control group, IRI and AR groups showed aggravated histopathological damage and increased secretion of oxidative stress markers and inflammatory factors in rat kidneys (all p<0.05). Finally, TF-Hub and miRNA-Hub gene regulatory networks were constructed to provide a theoretical basis for the regulation of Hub genes. CONCLUSION: We identified four macrophage M1-related Hub genes shared among AKI and AR after kidney transplantation. These genes may be considered for diagnosis of AKI and AR after kidney transplantation.

IL-13 alleviates acute kidney injury and promotes regeneration via activating the JAK-STAT signaling pathway in a rat kidney transplantation model.

AIMS: To identify whether and how a younger systemic internal milieu alleviates acute kidney injury (AKI) in grafts after kidney transplantation. MATERIALS AND METHODS: We conducted an allogenic heterotopic rat kidney transplantation model with young and adult recipients receiving similar donor kidneys. We evaluated the renal function, histological damage, apoptosis, dedifferentiation, proliferation, hub regulating cytokines, and signaling pathways involved in young and adult recipients based on transcriptomics, proteomics, and experimental validation. We also validated the protective effect and mechanism of interleukin-13 (IL-13) on tubular epithelial cell injury induced by transplantation in vivo and by cisplatin in vitro. KEY FINDINGS: Compared with adult recipients, the young recipients had lower levels of renal histological damage and apoptosis, while had higher levels of dedifferentiation and proliferation. Serum IL-13 levels were higher in young recipients both before and after surgery. Pretreating with IL-13 decreased apoptosis and promoted regeneration in injured rat tubular epithelial cells induced by cisplatin, while this effect can be counteracted by a JAK2 and STAT3 specific inhibitor, AG490. Recipients pretreated with IL-13 also had lower levels of histological damage and improved renal function. SIGNIFICANCE: Higher levels of IL-13 in young recipients ameliorates tubular epithelial cell apoptosis and promotes regeneration via activating the JAK-STAT signaling pathway both in vivo and in vitro. Our results suggest that IL-13 is a promising therapeutic strategy for alleviating AKI. The therapeutic potential of IL-13 in injury repair and immune regulation deserves further evaluation and clinical consideration.

GPX3 and GSTT1 as biomarkers related to oxidative stress during renal ischemia reperfusion injuries and their relationship with immune infiltration.

Background: Renal ischemia reperfusion injuries (IRIs) are very common in clinical diagnoses and treatments, which are a common cause of impaired renal functions, worsening pathological damage, affecting disease progression and hindering recovery. Renal IRIs are an inflammatory disease mediated by the adaptive and innate immune system. There is a complex interaction between oxidative stress and immune cell infiltration. Therefore, we aimed to determine biomarkers associated with oxidative stress during renal IRIs and their relationship with immune cell infiltration. Method: A differential gene expression analysis was made based on the GSE148420 dataset from the NCBI Gene Expression Comprehensive Database (GEO) combined with 92 oxidative-stress (OS)-related genes identified in the Molecular Signatures Database. Then we identified differentially-expressed genes (DEOSGs) associated with oxidative stress, which were used for gene ontology (GO) and a Kyoto Encyclopedia of Genomes (KEGG) enrichment analysis. At the same time, we used PPI protein interaction networks and Lasso regression analysis to identify key genes, which were verified by the validation sets GSE58438 and GSE71647, as well as Western Blot detection on rat renal IRI models. At the same time, PAS staining, HE staining and immunohistochemistry were used to detect tissue damage and expression of markers related to oxidative stress during renal ischemia-reperfusion. Single-gene enrichment analysis (GSEA) was used to further clarify the underlying biological functions of key genes. Cibersort was used to analyze the immune cell infiltration during renal IRI and the correlation of key genes with immune cells. At the same time, we constructed a network of transcription-factor (TF)-Hub genes and miRNA-Hub genes. DGIDB was used to predict drugs and molecular compounds that might interact with the Hub genes. Results: Compared with the control group, a total of 5456 differential genes (DEGs) were measured in the renal IRI group, 2486 of which were upregulated and 2970 were down-regulated. Among them, we found 30 DEGs (DEOSGs) associated with oxidative stress. The results of GO and KEGG enrichment analysis showed that these DEOSGs were mainly enriched in glutathione metabolism, the response to oxidative stress stimulation, the regulation of T cell activation and apoptosis signaling pathways. Through a protein interaction network (PPI) and a LASSO regression analysis, a total of two Hub genes were identified, namely GPX3 and GSTT1, which were validated through external validation sets and animal experiments. Through pathological methods, we found that the pathological damage of renal tissue and the expression of oxidative stress markers increased after renal ischemia-reperfusion. The results of GSEA showed that the Hub genes were related to oxidative stress pathways, apoptosis signaling pathways and immune-response-related signaling pathways. An immunoinfiltration correlation analysis showed that genes GPX3 and GSTT1 were significantly positively correlated with plasma cells and macrophage M0, while were negatively correlated with monocytes and macrophages M1 and M2. Using the Strust, Starbase and DGIDB database, we predicted that 81 transcription factors, 49 miRNAs and 13 drug or molecular compounds might interact with the Hub genes. Conclusion: Through a comprehensive analysis of gene expression, our findings may provide new potential biomarkers for the pathogenesis of renal IRIs and a reliable basis for its early diagnosis as well as treatment.

Difenoconazole Exposure Induces Retinoic Acid Signaling Dysregulation and Testicular Injury in Mice Testes.

Difenoconazole (DFZ) is a broad-spectrum triazole fungicide that is widely utilized in agriculture. Although DFZ has been demonstrated to induce reproductive toxicity in aquatic species, its toxic effects on the mammalian reproductive system have yet to be fully elucidated. In vivo, male mice were administered 0, 20 or 40 mg/kg/d of DFZ via oral gavage for 35 days. Consequently, DFZ significantly decreased testicular organ coefficient, sperm count and testosterone levels, augmented sperm malformation rates, and elicited histopathological alterations in testes. TUNEL assay showed increased apoptosis in testis. Western blotting results suggested abnormally high expression of the sperm meiosis-associated proteins STRA8 and SCP3. The concentrations of retinoic acid (RA), retinaldehyde (RE), and retinol (ROL) were increased in the testicular tissues of DFZ-treated groups. The mRNA expression level of genes implicated in RA synthesis significantly increased while genes involved in RA catabolism significantly decreased. In vitro, DFZ reduced cell viability and increased RA, RE, and ROL levels in GC-2 cells. Transcriptome analysis revealed a significant enrichment of numerous terms associated with the RA pathway and apoptosis. The qPCR experiment verified the transcriptome results. In conclusion, our results indicate that DFZ exposure can disrupt RA signaling pathway homeostasis, and induce testicular injury in mice testes.

Wnt10a downregulation contributes to MEHP-induced disruption of self-renewal and differentiation balance and proliferation inhibition in GC-1 cells: Insights from multiple transcriptomic profiling.

Di (2-ethylhexyl) phthalate (DEHP), one of phthalic acid esters, has been widely used in daily products. Its main metabolite, mono (2-ethylhexyl) phthalate (MEHP) was reported to possess higher testicular toxicity than DEHP. To explore the precise mechanism in MEHP-induced testis damage, multiple transcriptomic sequencing was employed in spermatogonia cell line GC-1 cells treated with MEHP (0, 100, and 200 µM) for 24 h. Integrative omics analysis and empirical validation revealed that Wnt signaling pathway was downregulated and wnt10a, one of hub genes, may be the key player in this process. Similar results were observed in DEHP-exposed rats. MEHP-induced disturbance of self-renewal and differentiation was dose-dependent. Moreover, self-renewal proteins were downregulated; the differentiation level was stimulated. Meanwhile, GC-1 proliferation was decreased. Stable transformation strain of wnt10a overexpression GC-1 cell line constructed from lentivirus was utilized in this study. The upregulation of Wnt10a significantly reversed the dysfunction of self-renewal and differentiation and promoted the cell proliferation. Finally, retinol, predicted to be useful in CONNECTIVITY MAP (cMAP), failed to rescue the damage caused by MEHP. Cumulatively, our findings revealed that the downregulation of Wnt10a induced the imbalance of self-renew and differentiation, and inhibition of cell proliferation in GC-1 cells after MEHP exposure.

Inhibition of inflammatory factor TNF-α by ferrostatin-1 in microglia regulates necroptosis of oligodendrocyte precursor cells.

OBJECTIVE: Inflammation of the surrounding environment is a major reason causing loss or injury of oligodendrocyte precursor cells (OPCs) in myelin-associated diseases. Lipopolysaccharide-activated microglia can release various inflammatory factors such as tumor necrosis factor-α (TNF-α). One of the ways of OPC death is necroptosis, which can be triggered by TNF-α, a death receptor ligand, by activating receptor-interacting protein kinase 1 (RIPK1)/RIPK3/mixed lineage kinase domain-like protein (MLKL) signaling pathway. This study investigated whether inhibiting microglia ferroptosis can decrease TNF-α release to alleviate OPC necroptosis. METHODS: Lipopolysaccharide and Fer-1 stimulate BV2 cells. The expressions of GPX4 and TNF-α were detected by western blot and quantitative real-time PCR; malondialdehyde, glutathione, iron, and reactive oxygen species were measured by the assay kits. After lipopolysaccharide stimulation of BV2 cells, the supernatant was taken to culture OPC. The protein expression levels of RIPK1, p-RIPK1, RIPK3, p-RIPK3, MLKL, and p-MLKL were detected by western blot. RESULTS: Lipopolysaccharide administration could induce ferroptosis in microglia by decreasing ferroptosis marker GPX4, while ferroptosis inhibitor Fer-1 could significantly increase GPX4 level. Fer-1 prevented oxidative stress and iron concentration elevation and alleviated mitochondrial damage in lipopolysaccharide-induced BV2 cells. The results revealed that Fer-1 downregulated the release of lipopolysaccharide-induced TNF-α in microglia and attenuated OPC necroptosis by significantly decreasing the expression levels of RIPK1, p-RIPK1, MLKL, p-MLKL, RIPK3, and p-RIPK3. CONCLUSION: Fer-1 may be a potential agent for inhibiting inflammation and treating myelin-related diseases.