Role of Natural Killer Cells in Antibody-mediated Rejection of Kidney Transplantation.

Objective: This study explicitly demonstrates the roles of natural killer (NK) cells in different types of kidney transplantation. Methods: We'd done the whole study from October 2022 to October 2023. To further explore the significance of NK cells during renal transplantation, we provide a theoretical basis for clinically overcoming immune rejection after renal transplantation by developing new anti-rejection drugs. We selected twelve male mice and divided them into three groups (Syngeneic transplant group allograft transplant group allograft transplant (priming) group) by random. Initially, the morphological and histopathological changes in the kidney transplantation graft model of mice in different groups are observed. Further, the DSA-IgG levels in peripheral blood and C3d and IgG deposition in mice are detected by ELISA and immunohistochemical staining. Then, the Banff 2015 score is recorded to screen a suitable AMR mouse model. Finally, the expression of NK cells in different rejection modes is detected by flow cytometry, and the expressions of various cytokines (INF-γ, perforin, granzyme B, TNF-α) in peripheral blood are detected by enzyme-linked immunosorbent assay (ELISA). Results: In the allogeneic transplantation (priming) group, peritubular capillary inflammatory cell infiltration, moderate endarteritis, and small arterial fibrinoid necrosis are evident. The Banff score showed that the allogeneic transplantation (pre-sensitized) group is significantly higher than the syngeneic and allogeneic transplantation groups. The C3H→C57BL/6 mice are pre-sensitized by skin transplantation, and then kidney transplantation is performed to establish the antibody-mediated rejection (AMR) model. After kidney transplantation, the expression levels of NK cells in the peripheral blood, spleen, and transplanted kidney tissue of mice in the pre-sensitized group are significantly higher than in the allogeneic transplantation and control groups. In the C3H→C57BL/6 mouse model of AMR, NK cells and the related cytokines in the peripheral blood are highly expressed after kidney transplantation, proving that NK cells play an essential role in the occurrence of AMR. Conclusion: Our study proved the significance of NK cells in the occurrence of AMR by systematically monitoring the expression of NK cell-related cytokines in different types, which provided some ideas for the clinical treatment of AMR.

RAB3D, upregulated by aryl hydrocarbon receptor (AhR), promotes the progression of prostate cancer by activating the PI3K/AKT signaling pathway.

Many patients with prostate cancer (PCa) cannot be diagnosed until an advanced stage, which make PCa become a large threat to human health. It is an urgent need to explore novel biomarkers for accurate diagnosis and targets for the effective treatment of PCa. This study aimed to investigate the effects of RAB3D (which belongs to the secretory Rab GTPases) on the progression of PCa. The results showed that RAB3D was highly expressed in PCa tissues compared to normal tissues according to the gene expression omnibus dataset. Consistent with the bioinformatics results, RAB3D exhibited a higher expression in PCa cells. Overexpression of RAB3D promoted the proliferation, migration, and invasion of PCa cells, whereas the knockdown of RAB3D led to the opposite results. The procancer effects of RAB3D were further confirmed by the in vivo growth of xenograft model. Subsequently, RAB3D upregulated the PI3K/AKT signaling pathway both in vivo and in vitro. LY294002 (a PI3K inhibitor) rescued the RAB3D upregulation-induced promotion of malignant phenotypes of PCa cells. Furthermore, the transcription activity of RAB3D was found to be enhanced by aryl hydrocarbon receptor (AhR; a transcription factor). The AhR silencing-induced inhibition of the proliferation and migration of PCa cells was reversed by the overexpression of RAB3D. Taken together, RAB3D, upregulated by AhR, promotes the PCa progression by activating the PI3K/AKT signaling pathway.

Construction and Analysis of lncRNA-miRNA-mRNA ceRNA Network Identify an Eight-Gene Signature as a Potential Prognostic Factor in Kidney Renal Papillary Cell Carcinoma (KIRP).

Aims: This study was conducted to establish the potential competing endogeneous RNA (ceRNA) network for predicting prognoses in kidney papillary renal cell carcinoma (KIRP) and explore novel therapeutic targets. Methods: The edgeR package in R was used to determine differentially expressed messenger RNAs (mRNAs), long non-coding RNAs (lncRNAs) and microRNAs (miRNAs), based on data from The Cancer Gene Atlas Program (TCGA) and the Genotype Expression (GTEx) databases. Weighted gene co-expression network analysis (WGCNA) was performed to filter out the mRNAs or lncRNAs that were strongly related to KIRP. The miRNAs that possibly sponged by differentially expressed RNAs lncRNAs (DElncRNAs) were screened using miRcode. Starbase, miRDB, and TargetScan sets were utilized to predict target mRNAs to corresponding miRNAs. LASSO and multivariate Cox regression analyses were applied for the determination of potential prognostic significance. Finally, the lncRNA-miRNA-mRNA ceRNA network was constructed. Results: A total of 1739 DEmRNAs and 1599 DElncRNAs were identified in KIRP. WGCNA analysis suggested that DEmRNAs in the blue module and DElncRNAs in the turquoise module were closely correlated with KIRP. An 8-gene signature was constructed, which had prognostic significance and predictive value in KIRP. Of note, a lncRNA-miRNA-mRNA ceRNA network (including 18 lncRNAs, 5 miRNAs, and 7 mRNAs) was established. Conclusion: This investigation constructed a new lncRNA-miRNA-mRNA ceRNA network, and proposed some genes that may be novel targets, as well as a theoretical basis for the treatment of patients with KIRP.

CREB1 protects against the renal injury in a rat model of kidney stone disease and calcium oxalate monohydrate crystals-induced injury in NRK-52E cells.

Kidney stone disease (KSD) is a common urinary disease with increasing prevalence worldwide. In this study, we investigated the effect of cyclic AMP responsive element binding protein (CREB) 1 in a KSD model of rat and calcium oxalate monohydrate (COM) crystals-treated NRK-52E cells. Rats were pretreated with lentivirus (LV)-CREB1 vector or LV-control vector and administrated with ethylene glycol + ammonium chloride to induce KSD. It was found that CREB1 was activated in the renal tissue of non-treated KSD rats. Pretreating with LV-CREB1 vector significantly enhanced CREB1 expression in KSD rats. Biochemical analysis for serum and urine showed that upregulation of CREB1 could improve the renal function of KSD rats. Histological analysis confirmed that upregulation of CREB1 alleviated the renal injury in KSD rats. Moreover, the upregulation of CREB1 suppressed the apoptosis in renal tissue of KSD rats through regulating apoptosis-associated proteins. Further study showed that the upregulation of CREB1 could attenuate the oxidative stress in KSD rats as well. More interestingly, the upregulation of CREB1 enhanced the activity of complex I and complex III and the expression of mitochondrial cytochrome c, implicating the effect of CREB1 on improving mitochondrial function in KSD rats. In vitro study confirmed that upregulation of CREB1 inhibited the apoptosis and oxidative stress, while improved the mitochondrial function of NRK-52E cells treated with COM crystals, demonstrating the protective effect of CREB1 on COM crystals-induced renal epithelial cell injury. Therefore, CREB1 might be served as a promising target in the prophylaxis and treatment of KSD.

Identification of potential crucial genes associated with carcinogenesis of clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is a common genitourinary malignancy with high mortality. However, the molecular pathogenesis of ccRCC remains unclear and effective biomarkers for daily practice are still limited. Thus, we aimed to identify the potential crucial genes and pathways associated with carcinogenesis of ccRCC and further analyze the molecular mechanisms implicated in tumorigenesis. In the present study, expression profiles GSE 66270, GSE 53757, GSE 36895, and GSE 76351 were downloaded from GEO database, including 244 matched primary and adjacent normal tissues, furthermore, the level 3 RNAseq dataset (RNAseqV2 RSEM) of KIRC was also downloaded from The Cancer Genome Atlas (TCGA), which consist of 529 ccRCC tumors and 72 normal tissues. Then, differentially expressed genes (DEGs) and pathway enrichment were analyzed by using R software. A total of 129 up- and 123 down-regulated genes were identified, which were aberrantly expressed both in GEO and TCGA data. Second, Gene ontology (GO) analyses revealed that most of the DEGs were significantly enriched in integral component of membrane, extracellular exosome, plasma membrane, cell adhesion, and receptor binding. Signaling pathway analyses indicated that DEGs had common pathways in signal transduction, metabolism, and immune system. Third, hub genes were identified with protein-protein interaction (PPI) network, including PTPRC, TGFB1, EGF, MYC, ITGB2, CTSS, FN1, CCL5, KNG1, and CD86. Additionally, sub-networks analyse was also performed by using MCODE plugin. In conclusion, the novel DEGs and pathways in ccRCC identified in this study may provide new insight into the underlying molecular mechanisms that facilitates RCC carcinogenesis.

Aryl hydrocarbon receptor enhances the expression of miR-150-5p to suppress in prostate cancer progression by regulating MAP3K12.

It has been reported that mircoRNAs (miRNAs) can act as tumor inhibitors in multiple malignant tumors. As a tumor suppressor, miR-150-5p has been reported in some cancers. However, the biological impacts of miR-150-5p in prostate cancer is not fully elaborated. This study aims to explore the biological role and mechanism of miR-150-5p in prostate cancer. The expression level of miR-150-5p was examined with Quantitative real time polymerase chain reaction (qRT-PCR). Moreover, Kaplan Meier analysis revealed that downregulation of miR-150-5p predicted unfavorable prognosis for patients with prostate cancer. To identify the inhibitory effects of miR-150-5p on the cellular processes of prostate cancer, gain-of function assay was conducted. Next, the inhibitory effects of Tetrachlorodibenzo-p-dioxin (TCDD) and 3,3'-Diindolylmethane (DIM) on the proliferation and invasion of prostate cancer cells were demonstrated. Knockdown of Ahr reversed the TCDD/DIM-mediated proliferation and invasion. The expression level of CYP1A1 also was measured to confirm that Ahr was activated by TCDD or DIM in prostate cancer cells. Mechanism experiments revealed that MAP3K12 is a target mRNA of miR-150-5p in prostate cancer cells. In conclusion, Aryl hydrocarbon receptor enhances the expression of miR-150-5p to suppress cell proliferation and invasion in prostate cancer by regulating MAP3K12.

Calcium oxalate crystals induces tight junction disruption in distal renal tubular epithelial cells by activating ROS/Akt/p38 MAPK signaling pathway.

Tight junction plays important roles in regulating paracellular transports and maintaining cell polarity. Calcium oxalate monohydrate (COM) crystals, the major crystalline composition of kidney stones, have been demonstrated to be able to cause tight junction disruption to accelerate renal cell injury. However, the cellular signaling involved in COM crystal-induced tight junction disruption remains largely to be investigated. In the present study, we proved that COM crystals induced tight junction disruption by activating ROS/Akt/p38 MAPK pathway. Treating Madin-Darby canine kidney (MDCK) cells with COM crystals induced a substantial increasing of ROS generation and activation of Akt that triggered subsequential activation of ASK1 and p38 mitogen-activated protein kinase (MAPK). Western blot revealed a significantly decreased expression of ZO-1 and occludin, two important structural proteins of tight junction. Besides, redistribution and dissociation of ZO-1 were observed by COM crystals treatment. Inhibition of ROS by N-acetyl-l-cysteine (NAC) attenuated the activation of Akt, ASK1, p38 MAPK, and down-regulation of ZO-1 and occludin. The redistribution and dissociation of ZO-1 were also alleviated by NAC treatment. These results indicated that ROS were involved in the regulation of tight junction disruption induced by COM crystals. In addition, the down-regulation of ZO-1 and occludin, the phosphorylation of ASK1 and p38 MAPK were also attenuated by MK-2206, an inhibitor of Akt kinase, implying Akt was involved in the disruption of tight junction upstream of p38 MAPK. Thus, these results suggested that ROS-Akt-p38 MAPK signaling pathway was activated in COM crystal-induced disruption of tight junction in MDCK cells.

Identification of biological targets of therapeutic intervention for clear cell renal cell carcinoma based on bioinformatics approach.

BACKGROUND: We aimed to discover the potential microRNA (miRNA) targets and to explore the underlying molecular mechanisms of clear cell renal cell carcinoma (ccRCC). METHODS: Microarray data of GSE16441 was downloaded from Gene Expression Omnibus database. Differentially expressed genes (DEGs) and differentially expressed miRNAs between ccRCC tumors and matched non-tumor samples were analyzed. Target genes of differentially expressed miRNAs were screened. Besides, functional enrichment analysis of DEGs was performed, followed by protein-protein interaction (PPI) network construction and sub-module analysis. Finally, the integrated miRNA-DEGs network was constructed. RESULTS: A total of 1758 up- and 2465 down-regulated DEGs were identified. Moreover, 15 up- and 12 down-regulated differentially expressed miRNAs were screened. The up-regulated DEGs were significantly enriched in pathways such as cell adhesion molecules and focal adhesion. Besides, the down-regulated DEGs were enriched in oxidative phosphorylation, and citrate cycle (TCA cycle). Moreover, eight sub-modules of PPI network were obtained. Totally, eight down-regulated miRNAs were identified to significantly regulate the DEGs and miRNA-200c that could regulate collagen, type V, alpha 2 (COL5A2) as well as COL5A3 was found to be the most significant. Additionally, 10 up-regulated miRNAs were identified to be significantly associated with the DEGs. Thereinto, miRNA-15a that could regulate ATPase, H(+) transporting, lysosomal 21 kDa, V0 subunit b (ATP6V0B) and miRNA-155 were found to be the most significant. CONCLUSIONS: miRNA-200c that could regulate COL5A2 and COL5A3, miRNA-15a that could regulate ATP6V0B and miRNA-155 may play key roles in ccRCC progression. These miRNAs may be potential targets for ccRCC treatment.

Reduced ING4 Expression Is Associated with the Malignancy of Human Bladder.

INTRODUCTION: Inhibitor of growth 4 (ING4) is a tumor suppressor. However the role of ING4 in human bladder malignancy is unknown. In this study, ING4 expression in human bladder cancer and its potential effects were studied. MATERIALS AND METHODS: ING4 expression in 47 human bladder cancer tissues and paired adjacent normal tissues was detected by Western blotting, quantitative reverse transcription-polymerase chain reaction, and immunohistochemistry. The migration and cell cycle progression of SV-HUC-1 and T24 cells with aberrant ING4 expression were examined. RESULTS: ING4 protein and mRNA were significantly decreased in bladder cancer tissues. ING4 protein level was significantly lower in the group of patients over 50 years of age. ING4 knockdown caused more rapid cell migration and increased the population of SV-HUC-1 and T24 cells in the G2-M phase. CONCLUSION: Our data suggest a close connection between aberrant ING4 expression and the carcinogenesis of human bladder cells. ING4 may be a potential target for bladder cancer chemotherapy.

USP28 is a potential prognostic marker for bladder cancer.

This study was conducted to analyze the expression of the ubiquitin-specific protease Usp28 and assess its clinical significance in human bladder cancer. mRNA and protein expression levels of Usp28 were determined by real-time polymerase chain reaction (PCR) and Western blot in 24 paired bladder cancers and the adjacent non-cancerous tissues. In addition, the expression of Usp28 protein in 186 bladder cancers was also determined by immunohistochemistry. The relationship between expression of Usp28 and clinico-pathologic features and prognosis was finally evaluated. Usp28 was expressed at a higher level in bladder cancers compared to adjacent non-cancerous tissues at both the mRNA and protein levels in 24 paired samples (all P < 0.01). In immunohistochemical examination, 78 (41.9%) of 186 cases displayed low Usp28 expression in cancerous tissues, whereas 108 (58.1%) cases displayed high Usp28 expression. In the universal analysis, Usp28 correlated strongly with histopathological grade, clinical stage, tumor number and recurrence rate (P = 0.0001, 0.0001, 0.0001 and 0.0051, respectively), but did not correlate with gender or age (P = 0.5588 and 0.6574). After multiple analysis of the above factors and consideration of confounding factors, tumor number, histological grade, clinical stage, and recurrence were related to Usp28 expression (P = 0.001, 0.001, 0.001 and 0.001, respectively). Finally, Usp28 expression was indentified as a independent predictors of survival (P = 0.001). Usp28 protein expression is potentially valuable in prognostic evaluation of bladder cancer.