Normal Kidney Tissue Research Articles

Elevated Expression of ADAP2 is Associated With Aggressive Behavior of Human Clear-Cell Renal Cell Carcinoma and Poor Patient Survival.

Clear cell renal cell carcinoma (ccRCC) is the most common and lethal cancer of the adult kidney. ADAP2 is a GTPase-activating protein was upregulated in clear cell renal cell carcinoma. The role of ADAP2 in ccRCC progression is unknown. ADAP2 expression in ccRCC cell lines and tissues was examined via real-time PCR, Western blot and IHC. MTS, colony formation and transwell assay to explore the role of ADAP2 in ccRCC. ADAP2 in growth and metastasis of ccRCC were evaluated in vivo through ccRCC xenograft tumor growth, lung metastatic mice model. The prognostic role of ADAP2 was evaluated by survival analysis. ADAP2 mRNA was expressed at significantly higher levels in 23 pairs of ccRCC tissues than in normal kidney tissues (P < 0.01). Immunohistochemical analysis of 298 ccRCC tissues revealed elevated ADAP2 expression as an independent unfavorable prognostic factor for the overall survival (P = 0.0042) and progression-free survival (P = 0.0232) of patients. The KaplanMeier survival curve showed that patients with a higher expression of ADAP2 showed a significantly lower overall survival rate and disease-free survival rate. Moreover, high expression of ADAP2 at the mRNA level was associated with a worse prognosis for overall survival (P = 0.0083) in The Cancer Genome Atlas (TCGA) cohort. In vivo and in vitro functional study showed that overexpression of ADAP2 promotes ccRCC cell proliferation and metastasis ability, whereas knockdown of ADAP2 inhibited cell proliferation, colony formation, migration and invasion. ADAP2 is a novel prognostic marker and could promotes tumor progression in ccRCC.

PD17-02 UNIVERSAL LOSS OF COMPLEX I IN ONCOCYTOMA IS ASSOCIATED WITH DEFECTIVE MITOPHAGY VIA ALTERED GLUTATHIONE METABOLISM

You have accessJournal of UrologyCME1 Apr 2023PD17-02 UNIVERSAL LOSS OF COMPLEX I IN ONCOCYTOMA IS ASSOCIATED WITH DEFECTIVE MITOPHAGY VIA ALTERED GLUTATHIONE METABOLISM Lin Lin, Neal Patel, Blake Wilde, Lucia Fernandez-del-Rio, Eirini Christodoulou, Shinji Ohtake, Randy Caliliw, Orian Shirihai, Xiaowu Gai, Heather Christofk, David Shackelford, and Brian Shuch Lin LinLin Lin More articles by this author , Neal PatelNeal Patel More articles by this author , Blake WildeBlake Wilde More articles by this author , Lucia Fernandez-del-RioLucia Fernandez-del-Rio More articles by this author , Eirini ChristodoulouEirini Christodoulou More articles by this author , Shinji OhtakeShinji Ohtake More articles by this author , Randy CaliliwRandy Caliliw More articles by this author , Orian ShirihaiOrian Shirihai More articles by this author , Xiaowu GaiXiaowu Gai More articles by this author , Heather ChristofkHeather Christofk More articles by this author , David ShackelfordDavid Shackelford More articles by this author , and Brian ShuchBrian Shuch More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003272.02AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Renal oncocytoma (RO) is the most commonly resected benign renal neoplasm. ROs are characterized by a dense accumulation of dysfunctional mitochondria possibly resulting by increased mitochondrial biogenesis and defective mitophagy. Although it is well known that ROs harbor recurrent inactivating mutations in mitochondrial genes encoding Complex I (NADH dehydrogenase) in the electron transport chain, the mechanism of defective mitophagy remains unclear. Here, we hypothesize that mitophagy inhibition in RO is regulated by Complex I loss, leading to increased glutathione that oxidizes key players in the mitophagy pathway. METHODS: RO and normal kidney (NK) tissues were obtained from patients undergoing surgical resection. DNA was isolated from frozen RO samples (n=18) and matched whole blood. Long-range PCR was performed to sequence mitochondrial DNA (mtDNA). Protein expression was analyzed using Western Blot. Complex I-IV activity, as measured by oxygen consumption rate, was analyzed by the Seahorse XF96 analyzer. Gene expression changes were evaluated using the Nanostring nCounter human metabolic pathway panel and analyzed by Rosalind. Metabolic profiles were measured using liquid chromatography-tandem mass spectrometry (LC-MS). Statistical calculations were performed using Graphpad Prism. RESULTS: A total of 13 ROs (72.2%) had mtDNA Complex I loss-of-function (LoF) mutations with 8 (44.4%) having high allele frequency (>50%). Compared to NK, nearly all ROs had reduced Complex I activity with a compensatory increase of Complex IV activity, irrespective of identified mtDNA mutation. On the protein level, there was a universal loss of Complex I with resultant compensatory increase of other complexes regardless of mtDNA status. Gene expression profiling revealed increased activation of AMPK pathway in ROs as well as altered glutathione metabolism, including increased biosynthesis and decreased breakdown. Metabolic profiling confirmed that both reduced and oxidized forms of glutathione were significantly enriched in ROs compared to NKs. The mechanism of glutathione inhibition of mitophagy is currently being investigated with possible candidates having dysregulated oxidation. CONCLUSIONS: The universal loss of Complex I activity in ROs is independent of mtDNA mutations. Complex I loss in RO is associated with increased glutathione, which in turn may result in impaired mitophagy. We continue to develop models to interrogate key drivers in this process. Source of Funding: AUA Urology Care Foundation Residency Research Award AUA Research Scholar Award © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e495 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Lin Lin More articles by this author Neal Patel More articles by this author Blake Wilde More articles by this author Lucia Fernandez-del-Rio More articles by this author Eirini Christodoulou More articles by this author Shinji Ohtake More articles by this author Randy Caliliw More articles by this author Orian Shirihai More articles by this author Xiaowu Gai More articles by this author Heather Christofk More articles by this author David Shackelford More articles by this author Brian Shuch More articles by this author Expand All Advertisement PDF downloadLoading ...

Immune landscape and the key role of APOE+ monocytes of lupus nephritis under the single-cell and spatial transcriptional vista.

Lupus nephritis (LN) is among the most common complication of systemic lupus erythematosus (SLE) with high mortality and morbidity. The analysis of LN kidney's local immune response through single-cell and spatial transcriptome enables the study of potential therapeutic targets. By single cell sequencing and spatial transcriptome, we profile cells from LN kidney and normal kidney tissues to characterize cellular composition and elucidate the potential upstream monocyte/macrophage (Mono/MΦ) initiating the auto-immune response. After the high-throughput synergy screening, we performed the immunofluorescence to identify the specific cells in LN patients. The function experiments were finished by flow cytometry and Elisa. By immunofluorescence and spatial transcriptome, we identified differential subsets of Mono/MΦ and demonstrated that they exhibit temporal expression of TIMP1, IL1B, SPP1 and APOE. With the function experiments, we found that the APOE+ Mono may be compensatorily increased in LN, and the capacity of antigen presenting was decreased with the overexpression of APOE. Furthermore, how do the LN-specific Mono/MΦ transport in and out the glomerulus to active the local immune response remains unclear. Our results showed that lymphangiogenesis occurred in LN kidneys but not in normal kidneys, suggesting the presence of a new lymphatic vessel may serve as a 'green channel' for LN-specific Mono/MΦ. In LN, APOE+ Mono are compensatorily elevated, with decreased antigen presenting ability and reduced secretion of interferons. The lymphangiogenesis in LN prompts the trafficking of Mono/MΦ in LN kidney.

Integrated bioinformatic analysis and cell line experiments reveal the significant role of the novel immune checkpoint TIGIT in kidney renal clear cell carcinoma.

T cell immunoglobulin and ITIM domain (TIGIT) is a widely concerned immune checkpoint, which plays an essential role in immunosuppression and immune evasion. However, the role of TIGIT in normal organ tissues and renal clear cell carcinoma is unclear. We aim to identify the critical role of TIGIT in renal clear cell carcinoma and find potential targeted TIGIT drugs. Data retrieved from the GTEX database and TCGA database was used to investigate the expression of TIGIT in normal whole-body tissues and abnormal pan-cancer, then the transcriptome atlas of patients with kidney renal clear cell carcinoma (KIRC) in the TCGA database were applied to distinguish the TIGIT related features, including differential expression status, prognostic value, immune infiltration, co-expression, and drug response of sunitinib an anti-PD1/CTLA4 immunotherapy in KIRC. Furthermore, we constructed a gene-drug network to discover a potential drug targeting TIGIT and verified it by performing molecular docking. Finally, we conducted real-time polymerase chain reaction (PCR) and assays for Transwell migration and CCK-8 to explore the potential roles of TIGIT. TIGIT showed a moderate expression in normal kidney tissues and was confirmed as an essential prognostic factor that was significantly higher expressed in KIRC tissues, and high expression of TIGIT is associated with poor OS, PFS, and DSS in KIRC. Also, the expression of TIGIT was closely associated with the pathological characteristics of the tumor, high expression of TIGIT in KIRC was observed with several critical functions or pathways such as apoptosis, BCR signaling, TCR signaling etal. Moreover, the expression of TIGIT showed a strong positive correlation with infiltration of CD8+ T cells and Tregs and a positive correlation with the drug sensitivity of sunitinib simultaneously. Further Tide ips score analysis and submap analysis reveal that patients with high TIGIT expression significantly show a better response to anti-PD1 immunotherapy. Following this, we discovered Selumetinib and PD0325901 as potential drugs targeting TIGIT and verified the interaction between these two drugs and TIGIT protein by molecular docking. Finally, we verified the essential role of TIGIT in the proliferation and migration functions by using KIRC cell lines. TIGIT plays an essential role in tumorigenesis and progression in KIRC. High expression of TIGIT results in poor survival of KIRC and high drug sensitivity to sunitinib. Besides, Selumetinib and PD0325901 may be potential drugs targeting TIGIT, and combined therapy of anti-TIGIT and other treatments show great potential in treating KIRC.

A chemokine network of T cell exhaustion and metabolic reprogramming in renal cell carcinoma.

Renal cell carcinoma (RCC) is frequently infiltrated by immune cells, a process which is governed by chemokines. CD8+T cells in the RCC tumor microenvironment (TME) may be exhausted which most likely influence therapy response and survival. The aim of this study was to evaluate chemokine-driven T cell recruitment, T cell exhaustion in the RCC TME, as well as metabolic processes leading to their functional anergy in RCC. Eight publicly available bulk RCC transcriptome collectives (n=1819) and a single cell RNAseq dataset (n=12) were analyzed. Immunodeconvolution, semi-supervised clustering, gene set variation analysis and Monte Carlo-based modeling of metabolic reaction activity were employed. Among 28 chemokine genes available, CXCL9/10/11/CXCR3, CXCL13/CXCR5 and XCL1/XCR1 mRNA expression were significantly increased in RCC compared to normal kidney tissue and also strongly associated with tumor-infiltrating effector memory and central memory CD8+ T cells in all investigated collectives. M1 TAMs, T cells, NK cells as well as tumor cells were identified as the major sources of these chemokines, whereas T cells, B cells and dendritic cells were found to predominantly express the cognate receptors. The cluster of RCCs characterized by high chemokine expression and high CD8+ T cell infiltration displayed a strong activation of IFN/JAK/STAT signaling with elevated expression of multiple T cell exhaustion-associated transcripts. Chemokinehigh RCCs were characterized by metabolic reprogramming, in particular by downregulated OXPHOS and increased IDO1-mediated tryptophan degradation. None of the investigated chemokine genes was significantly associated with survival or response to immunotherapy. We propose a chemokine network that mediates CD8+ T cell recruitment and identify T cell exhaustion, altered energy metabolism and high IDO1 activity as key mechanisms of their suppression. Concomitant targeting of exhaustion pathways and metabolism may pose an effective approach to RCC therapy.

Establishment of a new prognostic risk model of MAPK pathway-related molecules in kidney renal clear cell carcinoma based on genomes and transcriptomes analysis.

The mitogen-activated protein kinase (MAPK) signaling pathway is often studied in oncology as the most easily mentioned signaling pathway. This study aims to establish a new prognostic risk model of MAPK pathway related molecules in kidney renal clear cell carcinoma (KIRC) based on genome and transcriptome analysis. In our study, RNA-seq data were acquired from the KIRC dataset of The Cancer Genome Atlas (TCGA) database. MAPK signaling pathway-related genes were obtained from the gene enrichment analysis (GSEA) database. We used "glmnet" and the "survival" extension package for LASSO (Least absolute shrinkage and selection operator) regression curve analysis and constructed a prognosis-related risk model. The survival curve and the COX regression analysis were used the "survival" expansion packages. The ROC curve was plotted using the "survival ROC" extension package. We then used the "rms" expansion package to construct a nomogram plot. We performed a pan-cancer analysis of CNV (copy number variation), SNV (single nucleotide variant), drug sensitivity, immune infiltration, and overall survival (OS) of 14 MAPK signaling pathway-related genes using several analysis websites, such as GEPIA website and TIMER database. Besides, the immunohistochemistry and pathway enrichment analysis used The Human Protein Atlas (THPA) database and the GSEA method. Finally, the mRNA expression of risk model genes in clinical renal cancer tissues versus adjacent normal tissues was further verified by real-time quantitative reverse transcription (qRT-PCR). We performed Lasso regression analysis using 14 genes and created a new KIRC prognosis-related risk model. High-risk scores suggested that KIRC patients with lower-risk scores had a significantly worse prognosis. Based on the multivariate Cox analysis, we found that the risk score of this model could serve as an independent risk factor for KIRC patients. In addition, we used the THPA database to verify the differential expression of proteins between normal kidney tissues and KIRC tumor tissues. Finally, the results of qRT-PCR experiments suggested large differences in the mRNA expression of risk model genes. This study constructs a KIRC prognosis prediction model involving 14 MAPK signaling pathway-related genes, which is essential for exploring potential biomarkers for KIRC diagnosis.

COL28 promotes proliferation, migration, and EMT of renal tubular epithelial cells

Type XXVIII collagen (COL28) is involved in cancer and lung fibrosis. COL28 polymorphisms and mutations might be involved in kidney fibrosis, but the exact role of COL28 in renal fibrosis is unknown. This study explored the function of COL28 in renal tubular cells by examining the expression of COL28 mRNA and the effects of COL28 overexpression in human tubular cells. COL28 mRNA expression and localization were observed in normal and fibrotic kidney tissues from humans and mice using real-time PCR, western blot, immunofluorescence, and immunohistochemistry. The consequences of COL28 overexpression on cell proliferation, migration, cell polarity, and epithelial-to-mesenchymal transition (EMT) induced by TGF-β1 were examined in human tubular HK-2 cells. COL28 expression was low in human normal renal tissues, mainly observed in the renal tubular epithelial cells and especially in proximal renal tubules. COL28 protein expression in human and mouse obstructive kidney disease was higher than in normal tissues (p < 0.05) and more significant in the UUO2-Week than the UUO1-Week group. The overexpression of COL28 promoted HK-2 cell proliferation and enhanced their migration ability (all p < 0.05). TGF-β1 (10 ng/ml) induced COL28 mRNA expression in HK-2 cells, decreased E-cadherin and increased α-SMA in the COL28-overexpression group compared with controls (p < 0.05). ZO-1 expression decreased while COL6 increased in the COL28-overexpression group compared with controls (p < 0.05). In conclusion, COL28 overexpression promotes the migration and proliferation of renal tubular epithelial cells. The EMT could also be involved. COL28 could be a therapeutic target against renal- fibrotic diseases.

The relationship of apparent diffusion coefficient values of renal cell carcinoma before and after cryotherapy ablation

IntroductionThe diagnosis of renal cell carcinoma (RCC) is increasing due to incidental findings with more frequent use of cross-sectional imaging. Therefore improvements to diagnostic and follow up imaging techniques is necessary. MRI diffusion weighted imaging (DWI) is a recognised method of measuring the diffusion of water within lesions using the apparent diffusion coefficient (ADC), and may have a role in monitoring the efficacy of cryotherapy ablation of RCC. MethodsA retrospective cohort study of 50 patients was approved to investigate if the ADC value can determine the success of cryotherapy ablation treatment for RCC. DWI was performed at a single centre using 1.5 T MRI before and after cryotherapy ablation to the RCC. The control group was considered as the unaffected kidney. The ADC value of the RCC tumour and normal kidney tissue prior to and after cryotherapy ablation was measured, and compared to the result of the MRI. ResultsA statistically significant change in the ADC values was observed, pre ablation (1.562 × 10¯mm2/sec) to the post ablation (1.126 × 10¯³mm2/sec), p < 0.0005. There was no statistical significance in any of the other outcomes measured. ConclusionAlthough a change of ADC value occurred this is likely due to cryotherapy ablation causing coagulative necrosis at the site, and does not determine the success of the cryotherapy ablation. This can be considered a feasibility study for future research. Implications for practiceDWI is a quick addition to routine protocols, does not require intravenous gadolinium based contrast agent, and provides qualitative and quantitative data. Further research is required to establish the role of ADC for treatment monitoring.

Research on automatic classification technology of kidney tumor and normal kidney tissue based on computed tomography radiomics.

By using a radiomics-based approach, multiple radiomics features can be extracted from regions of interest in computed tomography (CT) images, which may be applied to automatically classify kidney tumors and normal kidney tissues. The study proposes a method based on CT radiomics and aims to use extracted radiomics features to automatically classify of kidney tumors and normal kidney tissues and to establish an automatic classification model. CT data were retrieved from the 2019 Kidney and Kidney Tumor Segmentation Challenge (KiTS19) in The Cancer Imaging Archive (TCIA) open access database. Arterial phase-enhanced CT images from 210 cases were used to establish an automatic classification model. These CT images of patients were randomly divided into training (168 cases) and test (42 cases) sets. Furthermore, the radiomics features of gross tumor volume (GTV) and normal kidney tissues in the training set were extracted and screened, and a binary logistic regression model was established. For the test set, the radiomic features and cutoff value of P were consistent with the training set. Three radiomics features were selected to establish the binary logistic regression model. The accuracy (ACC), sensitivity (SENS), specificity (SPEC), area under the curve (AUC), and Youden index of the training and test sets based on the CT radiomics classification model were all higher than 0.85. The automatic classification model of kidney tumors and normal kidney tissues based on CT radiomics exhibited good classification ability. Kidney tumors could be distinguished from normal kidney tissues. This study may complement automated tumor delineation techniques and warrants further research.

Proteotranscriptomic Discrimination of Tumor and Normal Tissues in Renal Cell Carcinoma.

Clear cell renal carcinoma is the most frequent type of kidney cancer, with an increasing incidence rate worldwide. In this research, we used a proteotranscriptomic approach to differentiate normal and tumor tissues in clear cell renal cell carcinoma (ccRCC). Using transcriptomic data of patients with malignant and paired normal tissue samples from gene array cohorts, we identified the top genes over-expressed in ccRCC. We collected surgically resected ccRCC specimens to further investigate the transcriptomic results on the proteome level. The differential protein abundance was evaluated using targeted mass spectrometry (MS). We assembled a database of 558 renal tissue samples from NCBI GEO and used these to uncover the top genes with higher expression in ccRCC. For protein level analysis 162 malignant and normal kidney tissue samples were acquired. The most consistently upregulated genes were IGFBP3, PLIN2, PLOD2, PFKP, VEGFA, and CCND1 (p < 10-5 for each gene). Mass spectrometry further validated the differential protein abundance of these genes (IGFBP3, p = 7.53 × 10-18; PLIN2, p = 3.9 × 10-39; PLOD2, p = 6.51 × 10-36; PFKP, p = 1.01 × 10-47; VEGFA, p = 1.40 × 10-22; CCND1, p = 1.04 × 10-24). We also identified those proteins which correlate with overall survival. Finally, a support vector machine-based classification algorithm using the protein-level data was set up. We used transcriptomic and proteomic data to identify a minimal panel of proteins highly specific for clear cell renal carcinoma tissues. The introduced gene panel could be used as a promising tool in the clinical setting.

ANO4 Expression Is a Potential Prognostic Biomarker in Non-Metastasized Clear Cell Renal Cell Carcinoma.

Background: Over the past decade, transcriptome profiling has elucidated many pivotal pathways involved in oncogenesis. However, a detailed comprehensive map of tumorigenesis remains an enigma to solve. Propelled research has been devoted to investigating the molecular drivers of clear cell renal cell carcinoma (ccRCC). To add another piece to the puzzle, we evaluated the role of anoctamin 4 (ANO4) expression as a potential prognostic biomarker in non-metastasized ccRCC. Methods: A total of 422 ccRCC patients with the corresponding ANO4 expression and clinicopathological data were obtained from The Cancer Genome Atlas Program (TCGA). Differential expression across several clinicopathological variables was performed. The Kaplan-Meier method was used to assess the impact of ANO4 expression on the overall survival (OS), progression-free interval (PFI), disease-free interval (DFI), and disease-specific survival (DSS). Univariate and multivariate Cox logistic regression analyses were conducted to identify independent factors modulating the aforementioned outcomes. Gene set enrichment analysis (GSEA) was used to discern a set of molecular mechanisms involved in the prognostic signature. Tumor immune microenvironment was estimated using xCell. Results: ANO4 expression was upregulated in tumor samples compared to normal kidney tissue. Albeit the latter finding, low ANO4 expression is associated with advanced clinicopathological variables such as tumor grade, stage, and pT. In addition, low ANO4 expression is linked to shorter OS, PFI, and DSS. Multivariate Cox logistic regression analysis identified ANO4 expression as an independent prognostic variable in OS (HR: 1.686, 95% CI: 1.120-2.540, p = 0.012), PFI (HR: 1.727, 95% CI: 1.103-2.704, p = 0.017), and DSS (HR: 2.688, 95% CI: 1.465-4.934, p = 0.001). GSEA identified the following pathways to be enriched within the low ANO4 expression group: epithelial-mesenchymal transition, G2-M checkpoint, E2F targets, estrogen response, apical junction, glycolysis, hypoxia, coagulation, KRAS, complement, p53, myogenesis, and TNF-α signaling via NF-κB pathways. ANO4 expression correlates significantly with monocyte (ρ = -0.1429, p = 0.0033) and mast cell (ρ = 0.1598, p = 0.001) infiltration. Conclusions: In the presented work, low ANO4 expression is portrayed as a potential poor prognostic factor in non-metastasized ccRCC. Further experimental studies should be directed to shed new light on the exact molecular mechanisms involved.

Ex vivo tumor dissection followed by kidney autotransplantation in bilateral wilms tumor.

Successful management of bilateral Wilm's tumor (BWT) involves a radical resection while preserving enough normal kidney tissue. Nephron-sparing surgery often results in an R1/R2 resection with a high recurrence rate in children with huge or multiple tumors, or tumors proximity to the renal hilum. In contrast, kidney autotransplantation can completely resect the tumor while maintaining homeostasis and preserving the patient's healthy kidney tissues. We summarized the clinical data of 8 synchronous BWT patients who underwent kidney autotransplantation at the First Affiliated Hospital of Sun Yat-sen University from 2018 to 2020. Ex vivo tumor resection and kidney autotransplantions were performed on 11 kidneys. The baseline characteristics, perioperative management, and survival status were reported. Nephron-sparing surgeries were performed on 5 kidneys in vivo. Among all the 8 patients, six of them (75%) received staged operation and the other 2 patients (25%) received single-stage operation. No residual tumors were found on the postoperative imaging in all the 8 patients. In total, 6 (75%) patients occurred complications after the autotransplantation, among which, 2 (33.3%) patients had complication of Clavien-Dindo grade IIIa, and 4 (66.7%) patients had complication of grade < 3. During the 38 months of follow-up, 87.5% (7/8) of patients were tumor-free survival with normal renal function. One patient died from renal failure without tumor recurrence. Therefore, our study indicated that autologous kidney transplantation can be an option for patients with complex BWT if the hospital's surgical technique and perioperative management conditions are feasible.

Identify AGAP2 as prognostic biomarker in clear cell renal cell carcinoma based on bioinformatics and IHC staining

BackgroundArf GTPase-activating proteins are aberrantly expressed in a variety of tumors, but their role in clear cell renal cell carcinoma (ccRCC) was unclear. Exploring the biological role of Arf GAP with GTP binding protein like domain, Ankyrin repeat and PH domain 2 (AGAP2) in ccRCC could improve our understanding on the aggressiveness and immune relevance of ccRCC. MethodsThe expression of AGAP2 was analyzed based on the Cancer Genome Atlas (TCGA) database and verified in ccRCC samples using immunohistochemistry. The association between AGAP2 and clinical cancer stages was explored by TCGA dataset and UALCAN. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed to analyze the biological functions of AGAP2-related genes. Moreover, the relationship between AGAP2 and immune cell infiltration was investigated with TIME and TCGA dataset. ResultsCompared to normal tissues, AGAP2 was upregulated in ccRCC tissues. Higher expression of AGAP2 was associated with clinical cancer stages, TNM stages, pathologic stages, and status. Prognostic analysis on AGAP2 showed that AGAP2 overexpression was associated with KIRC overall survival (OS) reduction (P = 0.019). However, higher expression of AGAP2 may improve the OS of CESC (P = 0.002), THYM (P = 0.006) and UCEC (P = 0.049). GO and KEGG analysis showed that AGAP2-related genes was related to T cell activation, immune activity and PD-L1 expression and PD-1 checkpoint pathway. Furthermore, our study showed that AGAP2 were significantly associated with T cells, Cytotoxic cells, Treg, Th1 cells, CD8 T cells, T helper cells. And AGAP2 expression level affected the abundance of immune cells infiltration. The infiltrating level of immune cells was different between the AGAP2 high-expression and low-expression groups. ConclusionThe expression of AGAP2 in ccRCC was higher than that in normal kidney tissues. It was significantly associated with clinical stage, poor prognosis, and immune cell infiltration. Therefore, AGAP2 may become an important component for ccRCC patients who receive precision cancer therapy and may be a promising prognostic biomarker.

Magnetic resonance imaging diagnosis of renal infection in children

OBJECTIVE: To analyze the magnetic resonance imaging (MRI) manifestations of renal infection in children, evaluate the reliability of diffusion-weighted imaging (DWI) in detecting renal infections, and the difference in apparent diffusion coefficient (ADC) values between acute renal pelvis nephritis (APN), acute focal bacterial nephritis (AFBN) and renal abscess (RA). METHODS: Data for 513 children with a clinical orpathological diagnosis of acute kidney infection. Both MRI and voiding cystourethrography were performed in 28 cases. Group A comprised 17 cases of retrograde infection; Group B comprised 11 cases of blood–borne infection. The Kappa test was used to analyze differences in lesion DWI distribution between the groups, and the t-test was used to compare the ADC values. RESULTS: The MRI manifestations of pediatric renal infection were characteristic. The lesions had high-signal intensity on DWI and were located mainly in the renal cortex blood–borne infections versus the renal cortex and medulla in retrograde infections. The sensitivity and specificity of DWI for detecting lesions were 100% and 96%. The average ADC values of APN, AFBN, and RA lesions were lower than that of normal kidney tissue, with statistically significant differences between the three conditions; the ADC value of RA was the lowest, and that of APN was the highest. CONCLUSION: There were differences in the lesion distributions between renal retrograde and blood–borne infections. The quantitative decrease in ADC value may be related to the duration of the disease and the degree of renal damage.

Deregulated expression of the 14q32 miRNA cluster in clear cell renal cancer cells.

Clear cell renal cell carcinomas (ccRCC) are characterized by arm-wide chromosomal alterations. Loss at 14q is associated with disease aggressiveness in ccRCC, which responds poorly to chemotherapeutics. The 14q locus contains one of the largest miRNA clusters in the human genome; however, little is known about the contribution of these miRNAs to ccRCC pathogenesis. In this regard, we investigated the expression pattern of selected miRNAs at the 14q32 locus in TCGA kidney tumors and in ccRCC cell lines. We demonstrated that the miRNA cluster is downregulated in ccRCC (and cell lines) as well as in papillary kidney tumors relative to normal kidney tissues (and primary renal proximal tubule epithelial (RPTEC) cells). We demonstrated that agents modulating expression of DNMT1 (e.g., 5-Aza-deoxycytidine) could modulate 14q32 miRNA expression in ccRCC cell lines. Lysophosphatidic acid (LPA, a lysophospholipid mediator elevated in ccRCC) not only increased labile iron content but also modulated expression of a 14q32 miRNA. Through an overexpression approach targeting a subset of 14q32 miRNAs (specifically at subcluster A: miR-431-5p, miR-432-5p, miR-127-3p, and miR-433-3p) in 769-P cells, we uncovered changes in cellular viability and claudin-1, a tight junction marker. A global proteomic approach was implemented using these miRNA overexpressing cell lines which uncovered ATXN2 as a highly downregulated target. Collectively, these findings support a contribution of miRNAs at 14q32 in ccRCC pathogenesis.

Pathological Significance of Kidney and Brain Expressed Protein (KIBRA) in Clear Cell Renal Cell Carcinoma.

Kidney and brain expressed protein (KIBRA), a member of the WW domain-containing protein family, has an important role in tumour growth and progression in various cancers. However, the pathological significance of KIBRA expression in clear cell renal cell carcinoma (ccRCC) tissues is not fully understood. The aim of this study was to clarify the pathological significance and prognostic roles of KIBRA expression in patients with ccRCC. KIBRA immunoreactivity, proliferation index (PI; with anti-Ki-67 antibody), apoptotic index (AI; using anti-cleaved caspase-3), and large tumour suppressor kinases (LATS-2) were evaluated in 157 ccRCC specimens by immunohistochemistry. Fifty normal kidney tissues were also evaluated as controls. The relationships between KIBRA expression and these cancer-related variables as well as clinicopathological features and survival were analysed. Moderate to strong immunoreactivity of KIBRA was identified in all normal kidney tissues; however, ccRCC cells with strong KIBRA expression was rare. The immunoreactivity score (IRS) of KIBRA was negatively associated with grade, T stage, tumour diameter, and metastasis. Kaplan-Meier survival curves showed that high KIBRA expression was a favourite predictor for overall survival. KIBRA IRS was negatively associated with PI and positively associated with the IRS of LATS-2 by univariate analysis. In addition, multivariate analysis showed that KIBRA was significantly associated with PI. KIBRA demonstrated important roles as a tumour suppressor in ccRCC. In addition, its expression was significantly associated with survival in these patients. Several such KIBRA-related functions were speculated to be modulated by cancer cell proliferation and LATS-2.

Metabolic Activation of Benzo[a]pyrene by Human Tissue Organoid Cultures.

Organoids are 3D cultures that to some extent reproduce the structure, composition and function of the mammalian tissues from which they derive, thereby creating in vitro systems with more in vivo-like characteristics than 2D monocultures. Here, the ability of human organoids derived from normal gastric, pancreas, liver, colon and kidney tissues to metabolise the environmental carcinogen benzo[a]pyrene (BaP) was investigated. While organoids from the different tissues showed varied cytotoxic responses to BaP, with gastric and colon organoids being the most susceptible, the xenobiotic-metabolising enzyme (XME) genes, CYP1A1 and NQO1, were highly upregulated in all organoid types, with kidney organoids having the highest levels. Furthermore, the presence of two key metabolites, BaP-t-7,8-dihydrodiol and BaP-tetrol-l-1, was detected in all organoid types, confirming their ability to metabolise BaP. BaP bioactivation was confirmed both by the activation of the DNA damage response pathway (induction of p-p53, pCHK2, p21 and γ-H2AX) and by DNA adduct formation. Overall, pancreatic and undifferentiated liver organoids formed the highest levels of DNA adducts. Colon organoids had the lowest responses in DNA adduct and metabolite formation, as well as XME expression. Additionally, high-throughput RT-qPCR explored differences in gene expression between organoid types after BaP treatment. The results demonstrate the potential usefulness of organoids for studying environmental carcinogenesis and genetic toxicology.

Highlights from Recent Cancer Literature.

Highlights from Recent Cancer Literature.

HNF1β-associated cyst development and electrolyte disturbances are not explained by BAIAP2L2 expression.

Mutations or deletions in transcription factor hepatocyte nuclear factor 1 homeobox β (HNF1β) cause renal cysts and/or malformation, maturity-onset diabetes of the young and electrolyte disturbances. Here, we applied a comprehensive bioinformatic approach on ChIP-seq, RNA-seq, and gene expression array studies to identify novel transcriptional targets of HNF1β explaining the kidney phenotype of HNF1β patients. We identified BAR/IMD Domain Containing Adaptor Protein 2 Like 2 (BAIAP2L2), as a novel transcriptional target of HNF1β and validated direct transcriptional activation of the BAIAP2L2 promoter by a reporter luciferase assay. Using mass spectrometry analysis, we show that BAIAP2L2 binds to other members of the I-BAR domain-containing family: BAIAP2 and BAIAP2L1. Subsequently, the role of BAIAP2L2 in maintaining epithelial cell integrity in the kidney was assessed using Baiap2l2 knockout cell and mouse models. Kidney epithelial cells lacking functional BAIAP2L2 displayed normal F-actin distribution at cell-cell contacts and formed polarized three-dimensional spheroids with a lumen. In vivo, Baiap2l2 knockout mice displayed normal kidney and colon tissue morphology and serum and urine electrolyte concentrations were not affected. Altogether, our study is the first to characterize the function of BAIAP2L2 in the kidney in vivo and we report that mice lacking BAIAP2L2 exhibit normal electrolyte homeostasis and tissue morphology under physiological conditions.

Speckle-type POZ protein could play a potential inhibitory role in human renal cell carcinoma

BackgroundSpeckle-type POZ protein(SPOP), a substrate adaptor of Cul3 ubiquitin ligase, plays crucial roles in solid neoplasms by promoting the ubiquitination and degradation of substrates. Limited studies have shown that SPOP is overexpressed in human renal cell carcinoma (RCC) tissue. However, the exact role of SPOP in RCC remains unclear and needs to be further elucidated. The present study showed that SPOP was expressed at different levels in different RCC cell lines. The purpose of this study was to explore the roles of SPOP in the biological features of RCC cells and the expression levels of SPOP in human tissue microarray (TMA) and kidney tissues.MethodsHere, SPOP was overexpressed by lentiviral vector transfection in ACHN and Caki-1 cells, and SPOP was knocked down in Caki-2 cells with similar transfection methods. The transfection efficiency was evaluated by quantitative PCR and western blotting analyses. The role of SPOP in the proliferation, migration, invasion and apoptosis of cell lines was determined by the MTT, wound-healing, transwell and flow cytometry assays. Moreover, the cells were treated with different drug concentrations in proliferation and apoptosis assays to investigate the effect of sunitinib and IFN-α2b on the proliferation and apoptosis of SPOP-overexpressing cells and SPOP-knockdown RCC cells. Finally, immunohistochemical staining of SPOP was performed in kidney tissues and TMAs, which included RCC tissues and corresponding adjacent normal tissues.ResultsOverexpression of SPOP inhibited cell proliferation, migration and invasion and increased cell apoptosis. Interestingly, sunitinib and IFN-α2b at several concentrations increased the proliferation inhibitory rate and total apoptosis rate of cells overexpressing SPOP. The findings of the present study showed that the SPOP protein was significantly expressed at low levels in most clear cell RCC (ccRCC) tissues and at relatively high levels in the majority of adjacent normal tissues and kidney tissues. Kaplan–Meier survival analysis showed that there was no statistically significant difference in cumulative survival based on the data of different SPOP expression levels in TMA and patients.ConclusionsIn contrast to previous studies, our findings demonstrated that overexpression of SPOP might suppress the progression of RCC cells, which was supported by cell experiments and immunohistochemical staining. SPOP could be a potential tumour inhibitor in RCC.