Preliminary assessment of a portable Raman spectroscopy system for post-operative urinary stone analysis.

PURPOSE: We aimed to evaluate the reliability of a portable device that applies Raman spectroscopy at an excitation wavelength of 1064 nm for the post-operative analysis of urinary stone composition. MATERIALS AND METHODS: Urinary stone samples were obtained post-operatively from 300 patients. All samples were analyzed by the portable Raman spectroscopy system at an excitation wavelength of 1064 nm as well as by infrared spectroscopy (IR), and the results were compared. RESULTS: Both Raman spectroscopy and IR could detect multiple stone components, including calcium oxalate monohydrate, calcium oxalate dihydrate, calcium phosphate, uric acid, cystine, and magnesium ammonium phosphate hexahydrate. The results from 1064-nm Raman analysis matched those from IR analysis for 96.0% (288/300) of cases. Although IR detected multiple components within samples more often than Raman analysis (239 vs 131), the Raman analysis required less time to complete than IR data acquisition (5 min vs 30 min). CONCLUSIONS: These preliminary results indicate that 1064-nm Raman spectroscopy can be applied in a portable and automated analytical system for rapid detection of urinary stone composition in the post-operative clinical setting. TRIAL REGISTRATION: Chinese Clinical Trail Register ID: ChiCTR2000039810 (approved WHO primary register) http://www.chictr.org.cn/showproj.aspx?proj=63662 .

SOX2 Promotes Invasion in Human Bladder Cancers through MMP2 Upregulation and FOXO1 Downregulation.

SOX2, a member of the SRY-related HMG-box (SOX) family, is abnormally expressed in many tumors and associated with cancer stem cell-like properties. Previous reports have shown that SOX2 is a biomarker for cancer stem cells in human bladder cancer (BC), and our most recent study has indicated that the inhibition of SOX2 by anticancer compound ChlA-F attenuates human BC cell invasion. We now investigated the mechanisms through which SOX2 promotes the invasive ability of BC cells. Our studies revealed that SOX2 promoted SKP2 transcription and increased SKP2-accelerated Sp1 protein degradation. As Sp1 is a transcriptionally regulated gene, HUR transcription was thereby attenuated, and, in the absence of HUR, FOXO1 mRNA was degraded fast, which promoted BC cell invasion. In addition, SOX2 promoted BC invasion through the upregulation of nucleolin transcription, which resulted in increased MMP2 mRNA stability and expression. Collectively, our findings show that SOX2 promotes BC invasion through both SKP2-Sp1-HUR-FOXO1 and nucleolin-MMP2 dual axes.

CircPTPRA blocks the recognition of RNA N6-methyladenosine through interacting with IGF2BP1 to suppress bladder cancer progression.

BACKGROUND: Circular RNAs (circRNAs) have been found to have significant impacts on bladder cancer (BC) progression through various mechanisms. In this study, we aimed to identify novel circRNAs that regulate the function of IGF2BP1, a key m6A reader, and explore the regulatory mechanisms and clinical significances in BC. METHODS: Firstly, the clinical role of IGF2BP1 in BC was studied. Then, RNA immunoprecipitation sequencing (RIP-seq) analysis was performed to identify the circRNAs interacted with IGF2BP1 in BC cells. The overall biological roles of IGF2BP1 and the candidate circPTPRA were investigated in both BC cell lines and animal xenograft studies. Subsequently, we evaluated the regulation effects of circPTPRA on IGF2BP1 and screened out its target genes through RNA sequencing. Finally, we explored the underlying molecular mechanisms that circPTPRA might act as a blocker in recognition of m6A. RESULTS: We demonstrated that IGF2BP1 was predominantly binded with circPTPRA in the cytoplasm in BC cells. Ectopic expression of circPTPRA abolished the promotion of cell proliferation, migration and invasion of BC cells induced by IGF2BP1. Importantly, circPTPRA downregulated IGF2BP1-regulation of MYC and FSCN1 expression via interacting with IGF2BP1. Moreover, the recognition of m6A-modified RNAs mediated by IGF2BP1 was partly disturbed by circPTPRA through its interaction with KH domains of IGF2BP1. CONCLUSIONS: This study identifies exonic circular circPTPRA as a new tumor suppressor that inhibits cancer progression through endogenous blocking the recognition of IGF2BP1 to m6A-modified RNAs, indicating that circPTPRA may serve as an exploitable therapeutic target for patients with BC.

CircLIFR synergizes with MSH2 to attenuate chemoresistance via MutSα/ATM-p73 axis in bladder cancer.

BACKGROUND: Cisplatin (CDDP) has become a standard-of-care treatment for muscle-invasive bladder cancer (MIBC), while chemoresistance remains a major challenge. Accumulating evidence indicates that circular RNAs (circRNAs) are discrete functional entities. However, the regulatory functions as well as complexities of circRNAs in modulating CDDP-based chemotherapy in bladder cancer are yet to be well revealed. METHODS: Through analyzing the expression profile of circRNAs in bladder cancer tissues, RNA FISH, circRNA pull-down assay, mass spectrometry analysis and RIP, circLIFR was identified and its interaction with MSH2 was confirmed. The effects of circLIFR and MSH2 on CDDP-based chemotherapy were explored by flow cytometry and rescue experiments. Co-IP and Western blot were used to investigate the molecular mechanisms underlying the functions of circLIFR and MSH2. Biological implications of circLIFR and MSH2 in bladder cancer were implemented in tumor xenograft models and PDX models. RESULTS: CircLIFR was downregulated in bladder cancer and expression was positively correlated with favorable prognosis. Moreover, circLIFR synergizing with MSH2, which was a mediator of CDDP sensitivity in bladder cancer cells, positively modulated sensitivity to CDDP in vitro and in vivo. Mechanistically, circLIFR augmented the interaction between MutSα and ATM, ultimately contributing to stabilize p73, which triggered to apoptosis. Importantly, MIBC with high expression of circLIFR and MSH2 was more sensitive to CDDP-based chemotherapy in tumor xenograft models and PDX models. CONCLUSIONS: CircLIFR could interact with MSH2 to positively modulate CDDP-sensitivity through MutSα/ATM-p73 axis in bladder cancer. CircLIFR and MSH2 might be act as promising therapeutic targets for CDDP-resistant bladder cancer.

The long non-coding RNA SNHG1 promotes bladder cancer progression by interacting with miR-143-3p and EZH2.

The long non-coding RNA (lncRNA) SNHG1 has been shown to be implicated in the progression of multiple human carcinomas. Nevertheless, the biological functions and potential mechanism of SNHG1 in bladder cancer (BC) are uncharacterized. In the present study, SNHG1 was found to be substantially up-regulated in BC tissues and cells and was intimately correlated with the TNM stage, lymphatic invasion, metastasis and recurrence-free survival in BC patients. Down-regulation of SNHG1 dramatically attenuated the proliferation, migration and invasion of BC cells, whereas the ectopic overexpression of SNHG1 had the opposite effects in vitro. The in vivo experimental results also indicated that SNHG1 down-regulation hampered the tumour growth and metastasis of BC cells. Mechanistic investigations revealed that SNHG1 enhances HK2 expression by serving as an endogenous sponge to regulate miR-143-3p in the cytoplasm of BC cells. In the nucleus, SNHG1 could interact with EZH2 and regulate the histone methylation of the CDH1 promoter, altering the biological behaviours of BC cells. Overall, these findings elucidate an oncologic role of SNHG1 in BC and provide a new therapeutic strategy against BC.

The RING domain in the anti-apoptotic protein XIAP stabilizes c-Myc protein and preserves anchorage-independent growth of bladder cancer cells.

X-linked inhibitor of apoptosis protein (XIAP) suppresses apoptosis and plays key roles in the development, growth, migration, and invasion of cancer cells. Therefore, XIAP has recently attracted much attention as a potential antineoplastic therapeutic target, requiring elucidation of the molecular mechanisms underlying its biological activities. Here, using shRNA-mediated gene silencing, immunoblotting, quantitative RT-PCR, anchorage-independent growth assay, and invasive assay, we found that XIAP's RING domain, but not its BIR domain, is crucial for XIAP-mediated up-regulation of c-Myc protein expression in human bladder cancer (BC) cells. Mechanistically, we observed that the RING domain stabilizes c-Myc by inhibiting its phosphorylation at Thr-58 and that this inhibition is due to activated ERK1/2-mediated phosphorylation of glycogen synthase kinase-3ß (GSK-3ß) at Ser-9. Functional studies further revealed that c-Myc protein promotes anchorage-independent growth and invasion stimulated by the XIAP RING domain in human BC cells. Collectively, the findings in our study uncover that the RING domain of XIAP supports c-Myc protein stability, providing insight into the molecular mechanism and role of c-Myc overexpression in cancer progression. Our observations support the notion of targeting XIAP's RING domain and c-Myc in cancer therapy.

Selenium-binding protein 1 transcriptionally activates p21 expression via p53-independent mechanism and its frequent reduction associates with poor prognosis in bladder cancer.

BACKGROUND: Recent studies have shown that selenium-binding protein 1 (SELENBP1) is significantly down-regulated in a variety of solid tumors. Nevertheless, the clinical relevance of SELENBP1 in human bladder cancer has not been described in any detail, and the molecular mechanism underlying its inhibitory role in cancer cell growth is largely unknown. METHODS: SELENBP1 expression levels in tumor tissues and adjacent normal tissues were evaluated using immunoblotting assay. The association of SELENBP1 expression, clinicopathological features, and clinical outcome was determined using publicly available dataset from The Cancer Genome Atlas bladder cancer (TCGA-BLCA) cohort. DNA methylation in SELENBP1 gene was assessed using online MEXPRESS tool. We generated stable SELENBP1-overexpression and their corresponding control cell lines to determine its potential effect on cell cycle and transcriptional activity of p21 by using flow cytometry and luciferase reporter assay, respectively. The dominant-negative mutant constructs, TAM67 and STAT1 Y701F, were employed to define the roles of c-Jun and STAT1 in the regulation of p21 protein. RESULTS: Here, we report that the reduction of SELENBP1 is a frequent event and significantly correlates with tumor progression as well as unfavorable prognosis in human bladder cancer. By utilizing TCGA-BLCA cohort, DNA hypermethylation, especially in gene body, is shown to be likely to account for the reduction of SELENBP1 expression. However, an apparent paradox is observed in its 3'-UTR region, in which DNA methylation is positively related to SELENBP1 expression. More importantly, we verify the growth inhibitory role for SELENBP1 in human bladder cancer, and further report a novel function for SELENBP1 in transcriptionally modulating p21 expression through a p53-independent mechanism. Instead, ectopic expression of SELENBP1 pronouncedly attenuates the phosphorylation of c-Jun and STAT1, both of which are indispensable for SELENBP1-mediated transcriptional induction of p21, thereby resulting in the G0/G1 phase cell cycle arrest in bladder cancer cell. CONCLUSIONS: Taken together, our findings provide clinical and molecular insights into improved understanding of the tumor suppressive role for SELENBP1 in human bladder cancer, suggesting that SELENBP1 could potentially be utilized as a prognostic biomarker as well as a therapeutic target in future cancer therapy.

Accurate prediction of secondary structure of tRNAs.

RNA structure plays a key role in understanding its biological function. Currently, the most successful de novo methods for RNA secondary structure prediction are based on free energy minimization. Existing free energy minimization methods focus more on building energy calculation models and optimizing energy parameters while paying less effort on developing structure space search algorithm. This study proposes a de novo method, namely DPARSS, for RNA secondary structure prediction. DPARSS proposes a novel dynamic programming algorithm for structure space search and intends to generate predictions with minimal free energy. An evaluation on a benchmark dataset consisting of tRNAs indicates that the DPARSS method achieves better predictive performance than existing free energy minimization methods. Additionally, case studies suggest that the improvement achieved by the DPARSS algorithm is likely due to the fact that the predictions generated by DPARSS algorithm form more base pairs, contain less mismatches and form longer base pair segments.

CircHIPK3 sponges miR-558 to suppress heparanase expression in bladder cancer cells.

Increasing evidences suggest that circular RNAs (circRNAs) exert crucial functions in regulating gene expression. In this study, we perform RNA-seq and identify 6,154 distinct circRNAs from human bladder cancer and normal bladder tissues. We find that hundreds of circRNAs are significantly dysregulated in human bladder cancer tissues. We further show that circHIPK3, also named bladder cancer-related circular RNA-2 (BCRC-2), is significantly down-regulated in bladder cancer tissues and cell lines, and negatively correlates with bladder cancer grade, invasion as well as lymph node metastasis, respectively. Over-expression of circHIPK3 effectively inhibits migration, invasion, and angiogenesis of bladder cancer cells in vitro and suppresses bladder cancer growth and metastasis in vivo Mechanistic studies reveal that circHIPK3 contains two critical binding sites for the microRNA miR-558 and can abundantly sponge miR-558 to suppress the expression of heparanase (HPSE). Taken together, our findings provide evidence that circRNAs act as "microRNA sponges", and suggest a new therapeutic target for the treatment of bladder cancer.

Transcriptional and post-transcriptional upregulation of p27 mediates growth inhibition of isorhapontigenin (ISO) on human bladder cancer cells.

There are few approved drugs available for the treatment of muscle-invasive bladder cancer (MIBC). Recently, we have demonstrated that isorhapontigenin (ISO), a new derivative isolated from the Chinese herb Gnetum cleistostachyum, effectively induces cell-cycle arrest at the G0/G1 phase and inhibits anchorage-independent cell growth through the miR-137/Sp1/cyclin D1 axis in human MIBC cells. Herein, we found that treatment of bladder cancer (BC) cells with ISO resulted in a significant upregulation of p27, which was also observed in ISO-treated mouse BCs that were induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). Importantly, knockdown of p27 caused a decline in the ISO-induced G0-G1 growth arrest and reversed ISO suppression of anchorage-independent growth in BC cells. Mechanistic studies revealed that ISO promoted p27 expression at mRNA transcription level through increasing direct binding of forkhead box class O1 (FOXO1) to its promoter, while knockdown of FOXO1 attenuated ISO inhibition of BC cell growth. On the other hand, ISO upregulated the 3'-untranslated region (3'-UTR) activity of p27, which was accompanied by a reduction of miR-182 expression. In line with these observations, ectopic expression of miR-182 significantly blocked p27 3'-UTR activity, whereas mutation of the miR-182-binding site at p27 mRNA 3'-UTR effectively reversed this inhibition. Accordingly, ectopic expression of miR-182 also attenuated ISO upregulation of p27 expression and impaired ISO inhibition of BC cell growth. Our results not only provide novel insight into understanding of the underlying mechanism related to regulation of MIBC cell growth but also identify new roles and mechanisms underlying ISO inhibition of BC cell growth.

Circular RNA BCRC-3 suppresses bladder cancer proliferation through miR-182-5p/p27 axis.

BACKGROUND: Circular RNAs (circRNAs) are a new member of noncoding RNAs (ncRNAs) that have recently been described as key regulators of gene expression. Our previous study had identified the negative correlation between circHIPK3 and bladder cancer grade, invasion, as well as lymph node metastasis. However, the roles of circRNAs in cellular proliferation in bladder cancer remain largely unknown. METHODS: We had analyzed circRNA high-throughout sequencing from human tissues and determined bladder cancer related circRNA-3 (BCRC-3, GenBank: KU921434.1) as a new candidate circRNA derived from PSMD1 gene. The expression levels of circRNAs, mRNAs and miRNAs in human tissues and cells were detected by quantitative real-time PCR (qRT-PCR). The effects of BCRC-3 on cancer cells were explored by transfecting with plasmids in vitro and in vivo. RNA pull down assay, luciferase reporter assay and fluorescence in situ hybridization were applied to verify the interaction between BCRC-3 and microRNAs. Anticancer effects of methyl jasmonate (MJ) were measured by flow cytometry assay, western blot and qRT-PCR. RESULTS: BCRC-3 was lowly expressed in bladder cancer tissues and cell lines. Proliferation of BC cells was suppressed by ectopic expression of BCRC-3 in vitro and in vivo. Mechanistically, overexpression of BCRC-3 induced the expression of cyclin-dependent kinase inhibitor 1B (p27). Importantly, BCRC-3 could directly interact with miR-182-5p, and subsequently act as a miRNA sponge to promote the miR-182-5p-targeted 3'UTR activity of p27. Furthermore, MJ significantly increased the expression of BCRC-3, resulting in an obvious up-regulation of p27. CONCLUSIONS: BCRC-3 functions as a tumor inhibitor to suppress BC cell proliferation through miR-182-5p/p27 axis, which would be a novel target for BC therapy.

XIAP overexpression promotes bladder cancer invasion in vitro and lung metastasis in vivo via enhancing nucleolin-mediated Rho-GDIß mRNA stability.

Our recent studies demonstrate that X-linked inhibitor of apoptosis protein (XIAP) is essential for regulating colorectal cancer invasion. Here, we discovered that RhoGDIß was a key XIAP downstream effector mediating bladder cancer (BC) invasion in vitro and in vivo. We found that both XIAP and RhoGDIß expressions were consistently elevated in BCs of N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-treated mice in comparison to bladder tissues from vehicle-treated mice and human BCs in comparison to the paired adjacent normal bladder tissues. Knockdown of XIAP attenuated RhoGDIß expression and reduced cancer cell invasion, whereas RhoGDIß expression was attenuated in BBN-treated urothelium of RING-deletion knockin mice. Mechanistically, XIAP stabilized RhoGDIß mRNA by its positively regulating nucleolin mRNA stability via Erks-dependent manner. Moreover, ectopic expression of GFP-RhoGDIß in T24T(shXIAP) cells restored its lung metastasis in nude mice. Our results demonstrate that XIAP-regulated Erks/nucleolin/RhoGDIß axis promoted BC invasion and lung metastasis.

NR4A1 is Involved in Fibrogenesis in Ovarian Endometriosis.

BACKGROUND/AIMS: Excess fibrosis may lead to chronic pain, scarring, and infertility as endometriosis develops and progresses. The pathogenesis of endometriosis has been linked to transforming growth factor-ß (TGF-ß), the most potent promoter of fibrosis. METHODS: Levels of NR4A1 and P-NR4A1 protein in human endometrial and endometriotic tissue were assessed by western blotting and immunohistochemistry. The expression levels of fibrotic markers in stromal cells were evaluated by real-time PCR. The degree of fibrosis in mouse endometriotic lesions was detected by Masson trichrome and Sirius red staining. RESULTS: The level of phosphorylated-NR4A1 was higher in ovarian endometriotic tissue than in normal endometrium, and long-term TGF-ß1 stimulation phosphorylated NR4A1 in an AKT-dependent manner and then promoted the expression of fibrotic markers. Furthermore, inhibition of NR4A1 in stromal cells increased the TGF-ß1-dependent elevated expression of fibrotic markers, and loss of NR4A1 stimulated fibrogenesis in mice with endometriosis. Additionally, Cytosporone B (Csn-B), an NR4A1 agonist, effectively decreased the TGF-ß1-dependent elevated expression of fibrotic markers in vitro and significantly inhibited fibrogenesis in vivo. CONCLUSION: NR4A1 can regulate fibrosis in endometriosis and may serve as a new target for the treatment of endometriosis.

Inhibition of BRD4 Suppresses Cell Proliferation and Induces Apoptosis in Renal Cell Carcinoma.

BACKGROUND/AIMS: Renal cell carcinoma (RCC) remains an intractable genitourinary malignancy. Resistance to chemotherapy or targeted therapies in RCC is presumably due to the complicated underlying molecular mechanisms and insufficient understanding. The aim of this research was to assess the expression and role of bromodomain-4 protein (BRD4) in RCC and evaluate the effects of BRD4 inhibitor JQ1 for RCC treatment. METHODS: BRD4 expressionlevels were assessed by qRT-PCR and western blot in RCC tissues and cells. The effects of BRD4 knockdown or JQ1 on RCC cells were assessed by MTT assay and flow cytometry. The effects of in vivo treatment were evaluated through xenograft experiments. RESULTS: BRD4 is significantly overexpressed in RCC, and is related to tumor stage and lymph node metastasis. Inhibition of BRD4 suppressed RCC cell proliferation, induced cell apoptosis in vitro and repressed tumor growth in vivo. Inhibition of BRD4 decreased BCL2 and C-MYC expression while increased BAX and cleaved caspase3 expression, and strikingly diminished the recruitment of BRD4 to BCL2 promoter. CONCLUSIONS: Our research reveals that BRD4 probably play a critical role in RCC progression, and is a new promising target for pharmacological treatment directed against this intractable disease.

Long Non-Coding RNA MEG3 Inhibits Cell Proliferation and Induces Apoptosis in Prostate Cancer.

BACKGROUND/AIMS: Long non-coding RNAs (lncRNAs) play important roles in diverse biological processes, such as cell growth, apoptosis and migration. Although downregulation of lncRNA maternally expressed gene 3 (MEG3) has been identified in several cancers, little is known about its role in prostate cancer progression. The aim of this study was to detect MEG3 expression in clinical prostate cancer tissues, investigate its biological functions in the development of prostate cancer and the underlying mechanism. METHODS: MEG3 expression levels were detected by qRT-PCR in both tumor tissues and adjacent non-tumor tissues from 21 prostate cancer patients. The effects of MEG3 on PC3 and DU145 cells were assessed by MTT assay, colony formation assay, western blot and flow cytometry. Transfected PC3 cells were transplanted into nude mice, and the tumor growth curves were determined. RESULTS: MEG3 decreased significantly in prostate cancer tissues relative to adjacent normal tissues. MEG3 inhibited intrinsic cell survival pathway in vitro and in vivo by reducing the protein expression of Bcl-2, enhancing Bax and activating caspase 3. We further demonstrated that MEG3 inhibited the expression of cell cycle regulatory protein Cyclin D1 and induced cell cycle arrest in G0/G1 phase. CONCLUSIONS: Our study presents an important role of MEG3 in the molecular etiology of prostate cancer and implicates the potential application of MEG3 in prostate cancer therapy.

Stabilization of Snail by HIF-1α and TNF-α is required for hypoxia-induced invasion in prostate cancer PC3 cells.

Hypoxia has been involved in the development of tumor by regulating the expression of invasiveness-associated genes. However, the specific function of hypoxia in cancer cell invasion is still unclear. The aim of the present study was to determine the role of hypoxia in invasion of prostate cancer PC3 cells and to investigate the underlying mechanisms. We found that hypoxia significantly increased the invasive activity of PC3 cells, via up-regulation of the expression of hypoxia inducible factor 1α (HIF-1α) and the autocrine production of tumor necrosis factor α (TNF-α). More important, TNF-α cooperated with HIF-1α in promoting stabilization of Snail, a transcriptional repressor of E-cadherin expression, which lead to the up-regulation of invasiveness-associated genes MMP-9, fibronectin and vimentin. Snail silencing by specific siRNA significantly inhibited hypoxia-induced invasion of PC3 cells, indicating an essential role of Snail in conferring the malignant phenotype to cancer cells under hypoxic conditions. In conclusion, our data demonstrate that hypoxia promoted the invasiveness of prostate cancer PC3 cells via HIF-1α- and TNF-α-induced stabilization of Snail, suggesting a signaling mechanism involving HIF-1α/TNF-α/Snail that mediates invasiveness hypoxic tumor cells in the absence of neoangiogenesis.

CLIC3 interacts with NAT10 to inhibit N4-acetylcytidine modification of p21 mRNA and promote bladder cancer progression.

Chromatin accessibility plays important roles in revealing the regulatory networks of gene expression, while its application in bladder cancer is yet to be fully elucidated. Chloride intracellular channel 3 (CLIC3) protein has been reported to be associated with the progression of some tumors, whereas the specific mechanism of CLIC3 in tumor remains unclear. Here, we screened for key genes in bladder cancer through the identification of transcription factor binding site clustered region (TFCR) on the basis of chromatin accessibility and TF motif. CLIC3 was identified by joint profiling of chromatin accessibility data with TCGA database. Clinically, CLIC3 expression was significantly elevated in bladder cancer and was negatively correlated with patient survival. CLIC3 promoted the proliferation of bladder cancer cells by reducing p21 expression in vitro and in vivo. Mechanistically, CLIC3 interacted with NAT10 and inhibited the function of NAT10, resulting in the downregulation of ac4C modification and stability of p21 mRNA. Overall, these findings uncover an novel mechanism of mRNA ac4C modification and CLIC3 may act as a potential therapeutic target for bladder cancer.

Exendin-4 ameliorates renal ischemia-reperfusion injury in the rat.

BACKGROUND: Glucagon-like peptide-1 receptor (GLP-1R) activation exerts protective effects against reactive oxygen species by inducing the oxidative defense gene heme oxygenase-1 (HO-1), and provides protection in mice against transient focal cerebral ischemia and ischemia-reperfusion injury in the rat heart. GLP-1R is also expressed in the kidney, but it is unknown whether GLP-1R activation is able to protect against ischemia-reperfusion injury in the rat kidney. MATERIALS AND METHODS: We used a rat model of renal ischemia-reperfusion injury. The rats were pretreated with the GLP-1R agonist, exendin-4 before reperfusion. We used real-time polymerase chain reaction to evaluate expression of the oxidative defense gene HO-1 and Western blot analysis for HO-1 and GLP-1R. Renal function was assessed at baseline and 24 and 72 h after reperfusion. The kidneys were processed for histologic and morphometric analysis, caspase-3, and ED1 immunohistochemistry at 72 h. The degree of apoptosis of the renal tubular cells was determined using terminal deoxynucleotidyl transferase deoxyuridine triphosphate-biotin nick end labeling assays. RESULTS: Exendin-4 pretreatment resulted in GLP-1R activation and upregulation of HO-1. Preconditional activation of GLP-1R significantly improved the serum creatinine levels compared with vehicle (P < 0.05). Furthermore, tissue injury, caspase-3 and ED1 expression, and apoptosis were less severe, as quantified by application of a standardized histologic scoring system in a blinded manner. CONCLUSIONS: These results have demonstrated that preconditional activation of the GLP-1R with exendin-4 in the kidney significantly protected against ischemia-reperfusion injury in rats by increasing HO-1 expression.