Mouse Kidney Tissue Research Articles

Spatial Segmentation of Mass Spectrometry Imaging Data by Combining Multivariate Clustering and Univariate Thresholding.

Spatial segmentation partitions mass spectrometry imaging (MSI) data into distinct regions, providing a concise visualization of the vast amount of data and identifying regions of interest (ROIs) for downstream statistical analysis. Unsupervised approaches are particularly attractive, as they may be used to discover the underlying subpopulations present in the high-dimensional MSI data without prior knowledge of the properties of the sample. Herein, we introduce an unsupervised spatial segmentation approach, which combines multivariate clustering and univariate thresholding to generate comprehensive spatial segmentation maps of the MSI data. This approach combines matrix factorization and manifold learning to enable high-quality image segmentation without an extensive hyperparameter search. In parallel, some ion images inadequately represented in the multivariate analysis were treated using univariate thresholding to generate complementary spatial segments. The final spatial segmentation map was assembled from segment candidates that were generated using both techniques. We demonstrate the performance and robustness of this approach for two MSI data sets of mouse uterine and kidney tissue sections that were acquired with different spatial resolutions. The resulting segmentation maps are easy to interpret and project onto the known anatomical regions of the tissue.

Suppressed Hepatic Production of Indoxyl Sulfate Attenuates Cisplatin-Induced Acute Kidney Injury in Sulfotransferase 1a1-Deficient Mice.

Endogenous factors involved in the progression of cisplatin nephropathy remain undetermined. Here, we demonstrate the toxico-pathological roles of indoxyl sulfate (IS), a sulfate-conjugated uremic toxin, and sulfotransferase 1A1 (SULT1A1), an enzyme involved in its synthesis, in cisplatin-induced acute kidney injury using Sult1a1-deficient (Sult1a1-/- KO) mice. With cisplatin administration, severe kidney dysfunction, tissue damage, and apoptosis were attenuated in Sult1a1-/- (KO) mice. Aryl hydrocarbon receptor (AhR) expression was increased by treatment with cisplatin in mouse kidney tissue. Moreover, the downregulation of antioxidant stress enzymes in wild-type (WT) mice was not observed in Sult1a1-/- (KO) mice. To investigate the effect of IS on the reactive oxygen species (ROS) levels, HK-2 cells were treated with cisplatin and IS. The ROS levels were significantly increased compared to cisplatin or IS treatment alone. IS-induced increases in ROS were reversed by downregulation of AhR, xanthine oxidase (XO), and NADPH oxidase 4 (NOX4). These findings suggest that SULT1A1 plays toxico-pathological roles in the progression of cisplatin-induced acute kidney injury, while the IS/AhR/ROS axis brings about oxidative stress.

Experimental evidence of pathogenic role of IgG autoantibodies in IgA nephropathy.

IgA nephropathy is thought to be an autoimmune disease wherein galactose-deficient IgA1 (Gd-IgA1) is recognized by IgG autoantibodies, resulting in formation and renal accumulation of nephritogenic immune complexes. Although this hypothesis is supported by recent findings that, in renal immunodeposits of IgA nephropathy patients, IgG is enriched for Gd-IgA1-specific autoantibodies, experimental proof is still lacking. IgG isolated from sera of IgA nephropathy patients or produced as a recombinant IgG (rIgG) was mixed with human Gd-IgA1 to form immune complexes. IgG from healthy individuals served as a control. Nude and SCID mice were injected with human IgG and Gd-IgA1, in immune complexes or individually, and their presence in kidneys was ascertained by immunofluorescence. Pathologic changes in the glomeruli were evaluated by quantitative morphometry and exploratory transcriptomic profiling was performed by RNA-Seq. Immunodeficient mice injected with Gd-IgA1 mixed with IgG autoantibodies from patients with IgA nephropathy, but not Gd-IgA1 mixed with IgG from healthy individuals, displayed IgA, IgG, and mouse complement C3 glomerular deposits and mesangioproliferative glomerular injury with hematuria and proteinuria. Un-complexed Gd-IgA1 or IgG did not induce pathological changes. Moreover, Gd-IgA1-rIgG immune complexes injected into immunodeficient mice induced histopathological changes characteristic of human disease. Exploratory transcriptome profiling of mouse kidney tissues indicated that these immune complexes altered gene expression of multiple pathways, in concordance with the changes observed in kidney biopsies of patients with IgA nephropathy. This study provides the first in vivo evidence for a pathogenic role of IgG autoantibodies specific for Gd-IgA1 in the pathogenesis of IgA nephropathy.

Evaluation of dietary selenium, vitamin C and E as the multi-antioxidants on the methylmercury intoxicated mice based on mercury bioaccumulation, antioxidant enzyme activity, lipid peroxidation and mitochondrial oxidative stress

Mercury (Hg) in high exposures can be a potent life threatening heavy metal that bioaccumulate in aquatic food-chain mainly as organic methylmercury (MeHg). In this regard, fish and seafood consumptions could be the primary sources of MeHg exposure for human and fish-eating animals. The objective of the present study was to elucidate the effects of dietary supplementation of some antioxidants on induced mercury toxicity in mice model. In this study, a 30-day long investigation has been conducted to evaluate the dietary effect of selenium (Se) in combination with vitamin C and vitamin E on methylmercury induced toxicity in mice. Total 54 mice fed the diets with three levels of Hg (0, 50 or 500 μg kg−1) and two levels of Se in combination with vitamin C and E (Se: 0, 2 mg kg−1; vitamin C: 0, 400 mg kg−1; vitamin E: 0, 200 mg kg−1) in triplicates. The results show that Hg accumulated in blood and different tissues such as muscle, liver and kidney tissues of mice on dose dependent manner. The bioaccumulation pattern of dietary Hg, in decreasing order, kidney > liver > muscle > blood. Superoxide dismutase levels in blood serum showed no significant differences in mice fed the diets. However, dietary antioxidants significantly reduced the levels of thiobarbituric acid reactive substances in mice fed the mercury containing diets. Cytochrome c oxidase enzyme activities showed no significant differences as the mercury level increases in liver and kidney tissues of mice. Kaplan-Meier curve showed a dose- and time-dependent survivability of mice. Cumulative survival rate of Hg intoxicated mice fed the antioxidant supplemented diets were increased during the experimental period. Overall, the results showed that dietary Se, vitamin C and vitamin E had no effect on reducing the mercury bioaccumulation in tissues but reduced the serum lipid peroxidation as well as prolonged the cumulative survival rate in terms of high Hg exposures in mice.

Deep Learning Approach for Dynamic Sparse Sampling for High-Throughput Mass Spectrometry Imaging.

A Supervised Learning Approach for Dynamic Sampling (SLADS) addresses traditional issues with the incorporation of stochastic processes into a compressed sensing method. Statistical features, extracted from a sample reconstruction, estimate entropy reduction with regression models, in order to dynamically determine optimal sampling locations. This work introduces an enhanced SLADS method, in the form of a Deep Learning Approach for Dynamic Sampling (DLADS), showing reductions in sample acquisition times for high-fidelity reconstructions between ~ 70-80% over traditional rectilinear scanning. These improvements are demonstrated for dimensionally asymmetric, high-resolution molecular images of mouse uterine and kidney tissues, as obtained using Nanospray Desorption ElectroSpray Ionization (nano-DESI) Mass Spectrometry Imaging (MSI). The methodology for training set creation is adjusted to mitigate stretching artifacts generated when using prior SLADS approaches. Transitioning to DLADS removes the need for feature extraction, further advanced with the employment of convolutional layers to leverage inter-pixel spatial relationships. Additionally, DLADS demonstrates effective generalization, despite dissimilar training and testing data. Overall, DLADS is shown to maximize potential experimental throughput for nano-DESI MSI.

A quantitative LC–MS/MS method for the determination of tissue brincidofovir and cidofovir diphosphate in a MuPyV‐infected mouse model

Brincidofovir (BCV) is an investigational lipid conjugate of the nucleotide analog cidofovir (CDV), which is being developed as a medical countermeasure for the treatment of smallpox. BCV is active against double-stranded DNA viruses including BK and JC viruses. Here, we validated procedures for quantifying BCV and its pharmacologically active moiety cidofovir diphosphate (CDV-PP) in mouse kidney, brain and spleen tissue homogenates. Following homogenization, BCV and CDV-PP were extracted from the tissues by protein precipitation with their stable, isotopically labeled internal standards, BCV-d6 and 13 C3 15 N2 -CDV-PP. Then, samples were analyzed for BCV by reverse-phase chromatography on a Waters Xterra MS C18 (50 × 2.1 mm, 3.5 μm particle size) column while CDV-PP was analyzed on a Thermo BioBasic AX (50 × 2.1 mm, 5 μm particle size) column using anion exchange chromatography. Detection was achieved by electrospray ionization in positive ion mode on an AB Sciex API-5000 triple quadrupole mass spectrometer. The calibration curves were linear over a range of 1.00-1,000 ng/ml homogenate and 0.050-50.0 ng/ml homogenate for BCV and CDV-PP, respectively. These methods were validated according to US Food and Drug Administration guidance for industry and may be used to characterize the tissue pharmacology of both analytes to advance its preclinical development.

Downregulation of Salusin-β protects renal tubular epithelial cells against high glucose-induced inflammation, oxidative stress, apoptosis and lipid accumulation via suppressing miR-155-5p

ABSTRACT Diabetic nephropathy (DN) is the main contributor to the excess mortality for patients suffering from diabetes. Here, C57BL/6 mice received 4 weeks of high-fat diet and intraperitoneal injection of STZ (100 mg/kg). Mice with random blood glucose level ≥16.7 mmol/L and positive urine protein were recognized as successful DN model. To construct an in vitro model, HK-2 cells were incubated with 30 mM glucose. RT-qPCR and western blot were employed to measure Salusin-β levels in kidney tissues of DN mice and HG-induced HK-2 cells. Meanwhile, RT-qPCR was performed to detect miR-155-5p level in kidney tissues of DN mice and HG-induced HK-2 cells. TNF-α, IL-6, IL-1β, ROS, SOD and CAT levels were assessed using commercial assay kits. Furthermore, apoptosis of HK-2 cells was assessed via flow cytometric analysis and TUNEL staining. In addition, intracellular lipid accumulation and total cholesterol levels were detected using Oil red O staining and TC ELISA kit. Herein, Salusin-β and miR-155-5p levels were distinctly upregulated in kidney tissues of DN mice and HG-induced HK-2 cells. Downregulation of Salusin-β reduced miR-155-5p expression. Salusin-β silencing dramatically relieved inflammatory and oxidative injury, suppressed apoptosis as well as lipid accumulation induced by HG in HK-2 cells. Besides, miR-155-5p elevation partially abrogated the alleviating effects Salusin-β silencing on HG-induced RTEC injury. In summary, downregulation of Salusin-β protected HK-2 cells against HG-induced inflammation, oxidative stress, apoptosis and ameliorated lipid accumulation through suppressing miR-155-5p, which indicated that Salusin-β could be a potential therapeutic drug for DN.

Aristolochic Acid Induces Chronic Kidney Disease in ACE Knockout Mice.

Background:Aristolochic acid I (AAI) is an extract from Chinese herbs that causes progressive interstitial nephritis. The aim of this research is to know whether chymases play the crucial role in AAI-induced nephropathy.Methods:The mice were treated with AAI via intraperitoneal injection and the accumulated AAI dosages are 30 mg/kg of body weight for two, four, six, and eight weeks. The animals were sacrificed after another two or four weeks for nephropathy development. Collection of blood, urine, and kidney samples for the further biochemical analysis, hematoxylin–eosin (H and E) and Masson's trichrome stained to detected pathologic, and MMP2 and MMP9 activity assays.Results:After the treatment of AAI, of the mice, their body weights were decreased (P < 0.01), and concentration of creatinine and blood urea nitrogen (BUN) in serum (P < 0.01) and urine collection were increased (P < 0.01). In the renal tissue sections, high amount of inflammatory cells were found by H and E stain, and increased fibrosis in renal interstitial tissue were observed by Masson's trichrome stain. In mice kidney tissue, significantly increased chymase activity after treatment of AAI was found (P < 0.01), but ACE activity did not show significant changes. In ACE KO mice, increased MMP2 and decreased MMP9 activity were found in the AAI-treated mice compared with AAI-untreated control (P < 0.01).Conclusions:Moreover, it was also observed that the deficiency of ACE would accelerate the disease development of AAI-induced nephropathy. These results may help to know more information about the role of AAI-induced chronic kidney disease and can be applied in developing new drug targets for nephropathy.

General Control of Amino Acid Synthesis 5-Like 1-Mediated Acetylation of Manganese Superoxide Dismutase Regulates Oxidative Stress in Diabetic Kidney Disease.

Diabetic kidney disease (DKD) is the major cause of end-stage renal disease (ESRD). In the past few decades, there has been a large amount of evidence to highlight the pivotal role of oxidative stress in the development and progression of DKD. However, the detailed molecular mechanisms are not fully elucidated. A new sight has been established that the mitochondrial acetyltransferase GCN5L1 participates in cellular redox homeostasis maintenance in DKD. Firstly, we found that the expression of GCN5L1 is significantly elevated both in human and mouse kidney tissues with DKD and in hyperglycemic renal tubular epithelial cells (TECs), while deletion of GCN5L1 could effectively ameliorate oxidative stress-induced renal injury in DKD. Furthermore, deletion of GCN5L1 could reduce MnSOD acetylation on lysine 68 and activate its activity, thereby scavenging excessive ROS and relieving oxidative stress-induced renal inflammation and fibrosis. In general, GCN5L1-mediated acetylation of MnSOD exacerbated oxidative stress-induced renal injury, suggesting that GCN5L1 might be a potential intervention target in DKD.

The role of discoidin domain receptor 2 in the renal dysfunction of alport syndrome mouse model

Alport syndrome (AS) is a hereditary glomerular nephritis caused by mutation in one of the type IV collagen genes α3/α4/α5 that encode the heterotrimer COL4A3/4/5. Failure to form a heterotrimer due to mutation leads to the dysfunction of the glomerular basement membrane, and end-stage renal disease. Previous reports have suggested the involvement of the receptor tyrosine kinase discoidin domain receptor (DDR) 1 in the progression of AS pathology. However, due to the similarity between DDR1 and DDR2, the role of DDR2 in AS pathology is unclear. Here, we investigated the involvement of DDR2 in AS using the X-linked AS mouse model. Mice were treated subcutaneously with saline or antisense oligonucleotide (ASO; 5 mg/kg or 15 mg/kg per week) for 8 weeks. Renal function parameters and renal histology were analyzed, and the gene expressions of inflammatory cytokines were determined in renal tissues. The expression level of DDR2 was highly elevated in kidney tissues of AS mice. Knockdown of Ddr2 using Ddr2-specific ASO decreased the Ddr2 expression. However, the DDR2 ASO treatment did not improve the proteinuria or decrease the BUN level. DDR2 ASO also did not significantly ameliorate the renal injury, inflammation and fibrosis in AS mice. These results showed that Ddr2 knockdown by ASO had no notable effect on the progression of AS indicating that DDR2 may not be critically involved in AS pathology. This finding may provide useful information and further understanding of the role of DDRs in AS.

Deficiency of the X-inactivation escaping gene KDM5C in clear cell renal cell carcinoma promotes tumorigenicity by reprogramming glycogen metabolism and inhibiting ferroptosis.

Background: Clear cell renal cell carcinoma (ccRCC) is characterized by glycogen-laden, unexplained male predominance, and frequent mutations in the Von Hippel-Lindau (VHL) gene and histone modifier genes. Besides, poor survival rates of ccRCC patients seem to be associated with up-regulation of the pentose phosphate pathway (PPP). However, the mechanism underlying these features remains unclear.Methods: Whole exome sequencing was used to identify the gene mutation that implicated in the rewired glucose metabolism. RNA-seq analyses were performed to evaluate the function of KDM5C in ccRCC. Furthermore, heavy isotope tracer analysis and metabolites quantification assays were used to study how KDM5C affects intracellular metabolic flux. To provide more in vivo evidence, we generated the Kdm5c-/- mice by CRISPR-Cas9 mediated gene knockout and performed the xenografts with KDM5C overexpressing or depleted cell lines.Results: A histone demethylase gene KDM5C, which can escape from X-inactivation and is predominantly mutated in male ccRCC patients, was identified to harbor the frameshift mutation in the ccRCC cell line with the highest glycogen level, while the restoration of KDM5C significantly reduced the glycogen level. Transcriptome and metabolomic analysis linked KDM5C to metabolism-related biological processes. KDM5C specifically regulated the expression of several hypoxia-inducible factor (HIF)-related genes and Glucose-6-phosphate dehydrogenase (G6PD) that were involved in glycogenesis/glycogenolysis and PPP, respectively, mainly through the histone demethylase activity of KDM5C. Depletion of KDM5C increased the production of glycogen, which was then directed to glycogenolysis to generate glucose-6-phosphate (G6P) and subsequently PPP to produce nicotinamide adenine dinucleotide phosphate hydride (NADPH) and glutathione (GSH), thus conferring cells resistance to reactive oxygen species (ROS) and ferroptosis. KDM5C re-expression suppressed the glucose flux through PPP and re-sensitized cancer cells to ferroptosis. Notably, Kdm5c-knockout mice kidney tissues exhibited elevated glycogen level, reduced lipid peroxidation and displayed a transformation of renal cysts into hyperplastic lesions, implying a cancer-protective benefit of ferroptosis. Furthermore, KDM5C deficiency predicted the poor prognosis, and clinically relevant KDM5C mutants failed to suppress glycogen accumulation and promoted ferroptosis as wild type.Conclusion: This work revealed that a histone modifier gene inactive mutation reprogramed glycogen metabolism and helped to explain the long-standing puzzle of male predominance in human cancer. In addition, our findings may suggest the therapeutic value of targeting glycogen metabolism in ccRCC.

MicroRNA-302b mitigates renal fibrosis via inhibiting TGF-β/Smad pathway activation.

Renal fibrosis is one of the most significant pathological changes after ureteral obstruction. Transforming growth factor-β (TGF-β) signaling pathway plays essential roles in kidney fibrosis regulation. The aims of the present study were to investigate effects of microRNA-302b (miR-302b) on renal fibrosis, and interaction between miR-302b and TGF-β signaling pathway in murine unilateral ureteral obstruction (UUO) model. Microarray dataset GSE42716 was downloaded by retrieving Gene Expression Omnibus database. In accordance with bioinformatics analysis results, miR-302b was significantly down-regulated in UUO mouse kidney tissue and TGF-β1-treated HK-2 cells. Masson's trichrome staining showed that miR-302b mimics decreased renal fibrosis induced by UUO. The increased mRNA expression of collagen I and α-smooth muscle actin (α-SMA) and decreased expression of E-cadherin were reversed by miR-302b mimics. In addition, miR-302b up-regulation also inhibited TGF-β1-induced epithelial mesenchymal transition (EMT) of HK-2 cells by restoring E-cadherin expression and decreasing α-SMA expression. miR-302b mimics suppressed both luciferase activity and protein expression of TGF-βR2. However, miR-302b inhibitor increased TGF-βR2 luciferase activity and protein expression. Meanwhile, miR-302b mimics inhibited TGF-βR2 mRNA expression and decreased Smad2 and Smad3 phosphorylation in vivo and in vitro. Furthermore, over-expression of TGF-βR2 restored the miR-302b-induced decrease of collagen I and α-SMA expression. In conclusion, this study demonstrated that miR-302b attenuated renal fibrosis by targeting TGF-βR2 to suppress TGF-β/Smad signaling activation. Our findings showed that elevating renal miR-302b levels may be a novel therapeutic strategy for preventing renal fibrosis.

Two-photon multifocal structured light microscopy based on high-speed phase-type spatial light modulator

Multifocal structured illumination microscopy (MSIM) can achieve a doubled improvement in the resolution of the diffraction limit within an imaging depth of 50 μm. But when imaging thick samples, scattered light and defocused light limit its optical sectioning capability and image contrast. Two-photon MSIM (2P-MSIM) overcomes the influence of sample tissue scattering and further improves the imaging depth and imaging characteristics. However, the existing 2P-MSIM usually adopts galvanometer based scanning mirrors for precisely scanning imaging, which is a complicated and poor flexibility system. Here we propose a simpler 2P-MSIM. Two-photon multifocal scanning imaging can be realized by a spatial light modulator (SLM) with a high frame rate (&lt; 845 Hz). The phase map of generating multi-focus array and linear phase grating loaded on the SLM simultaneously, high-precision parallel digital random address scanning and excitation imaging on the sample surface can be realized. The mechanical inertia problem of the galvanometer scanner in multifocal imaging can be solved by the proposed method while reducing the complexity of the system and improving flexibility. We finally realize two-photon multifocal imaging of mouse kidney tissue slices and lily of the valley rhizome by this system, which verifies the three-dimensional super-resolution imaging capability of this method. It is of great significance in developing the 2P-MSIM.

Effects of exposure to prothioconazole and its metabolite prothioconazole-desthio on oxidative stress and metabolic profiles of liver and kidney tissues in male mice

Prothioconazole (PTC), a popular agricultural fungicide, and its main metabolite prothioconazole-desthio (PTCd) are receiving great attention due to their toxicological effects in the non-target organisms. This study investigated their dosage-dependent (1 and 5mg/kg BW/day) toxicological effects on oxidative stress and metabolic profiles of liver and kidney tissues using male mice. PTC and PTCd significantly inhibited the growth phenotype including body weights gain, liver and kidney indices. Furthermore, these effects were deeply investigated using the biomarkers of oxidative stress, and metabolomics. Notably, these effects were dose and tissue-dependent. Specifically, the more serious impacts involving oxidative stress and metabolic disorders were observed in the high concentration treatment groups. Also, the liver tissue was more severely affected than the kidney tissue. Lastly, the change in oxidative stress biomarkers and metabolomics profile revealed that PTCd induced more severe toxic effects than the parent compound PTC. In brief, these results indicate that exposure to PTC and PTCd could cause potential health risks in mammals.

Kadmiyum Uygulanan Farelerde Oluşturulan Oksidatif Strese Karşı Resveratrolün Koruyucu Etkisinin Araştırılması*

Kadmiyum ve diğer ağır metaller hem çevresel hem de mesleki maruziyetlerle insan sağlığını tehdit etmekte ve sanayide yaygın olarak kullanılmaktadır. Resveratrol anti-inflamatuvar, antioksidan, antitümör ve immunomodulatör özellikleri olan doğal bir polifenolik bileşiktir. Bu nedenle yapılan çalışmada farelerin plazma, karaciğer, böbrek ve kalp dokusunda kadmiyum toksisitesine karşı resveratrolün koruyucu etkisi araştırılmıştır. Çalışmada 35 fare eşit olarak 5 gruba ayrılmıştır. Kontrol grubu, alkol grubu, kadmiyum grubu (CdCl2, 1 mg/kg/gün), resveratrol grubu (10 mg/kg/gün), kadmiyum+resveratrol grubu (1 mg/kg/gün+10 mg/kg/gün) olarak belirlenmiştir. Tüm uygulamalar 21 gün süreyle oral gavaj yoluyla uygulanmıştır. Uygulama süresi sonunda karaciğer, böbrek ve kalp dokusunda oksidatif stresin gösterge-si olarak malondialdehit (MDA) ve nitrik oksit (NO) miktarları, antioksidan savunmanın biyobelirteci olarak redükte glu-tatyon (GSH) miktarı, akut faz yanıtın bir göstergesi olan total siyalik asit (TSA) ve plazmada aspartat aminotransferaz (AST), alanin aminotransferaz (ALT), gamma glutamil transpeptidaz (GGT) aktiviteleri spektrofotometrik yöntemlerle belirlenmiştir. Kontrole göre, kadmiyum verilen grupta doku (karaciğer, böbrek ve kalp) MDA, NO, TSA düzeyleri ile plazma AST, ALT, GGT aktiviteleri önemli derecede artarken, doku GSH düzeyi ise önemli derecede azalmıştır. Kad-miyum+resveratrol verilen grupta ise doku MDA, NO, TSA düzeyleri ile plazma AST, ALT, GGT aktivitelerinin kadmi-yum verilen gruba göre istatistiksel olarak önemli derecede azalırken, doku GSH düzeyi ise önemli derecede artmıştır. Sonuç olarak kadmiyumun toksik etkisine karşı resveratrolün karaciğer, böbrek ve kalpte koruyucu etkisinin olabileceği kanısına varılmıştır.

Piperine Inhibits Cell Proliferation and Induces Apoptosis of Human Gastric Cancer Cells by Downregulating Phosphatidylinositol 3-Kinase (PI3K)/Akt Pathway.

BackgroundPiperine has been reported to inhibit proliferation and induce apoptosis in various cancer cells. This study aimed to explore the efficacy and underlying mechanism of piperine in human gastric cancer.Material/MethodsMTT assay was performed to examine the effect of piperine (concentrations of 0–300 μM) on the proliferation of human gastric cancer SNU-16 cells and normal human gastric epithelial GES-1 cells. Flow cytometry and Western blot were used to determine cell apoptosis and the expression level of protein (Cyto C, cleaved PARP, cleaved caspase-3, Bax, Bcl-2, Bad, Bcl-xl, PI3K, pPI3K, Akt, and pAkt), respectively. To further investigate the anti-tumor mechanism of piperine in SNU-16 cells, we used a small-molecule Akt activator SC79 in this study. The in vivo mechanism of piperine against gastric cancer was evaluated using a xenograft tumor model.ResultsThe results showed that piperine inhibited proliferation and induced apoptosis of SNU-16 cells. Piperine upregulated the protein expression of Bax, Bad, Cyto C, cleaved PARP, and cleaved caspase-3, but downregulated the protein expression of Bcl-2, Bcl-xl, pPI3k, and pAkt. However, SC79 reversed the function of piperine on the apoptosis-related proteins. An in vivo study revealed that, compared with the control group, the tumor volume of mice treated with piperine was significantly reduced. Piperine enhanced cleaved caspase-3 expression but decreased Ki-67 expression in a dose-dependent manner. Moreover, the nontoxicity effect of piperine was confirmed by H&E staining analysis in kidney and heart tissues of mice.ConclusionsOur findings suggest that piperine inhibits proliferation and induces apoptosis of human gastric cancer cells through inhibition of the PI3K/Akt signaling pathway.

CdSe/ZnS quantum dots exhibited nephrotoxicity through mediating oxidative damage and inflammatory response

Objective: This study aimed to the evaluate the nephrotoxicity of CdSe/ZnS QDs in vitro and vivo, as well as investigate the underlying toxicity mechanisms.Results: In vitro experiments showed that compared with control cells, CdSe/ZnS QDs treatment significantly inhibited cell viability and promoted cell apoptosis in dose-dependent manner in NRK cells. Notably, CdSe/ZnS QDs treatment increased the contents of MDA and ROS, and decreased the activities of SOD, CAT and GSH-Px; however, the co-treatment of NAC and QDs relieved the oxidative damage of NRK cells. Moreover, in vivo experiments also revealed that CdSe/ZnS QDs treatment obviously increased kidney weight coefficient, damaged the kidney function, as well as induced inflammatory response and inhibited the activation of NRF2/Keap1 pathway in kidney tissues of mice.Conclusions: CdSe/ZnS QDs exhibited obvious nephrotoxicity by mediating oxidative damage and inflammatory response in vitro and in vivo via NRF2/Keap1 pathway.Methods: The characterization of CdSe/ZnS QDs was analyzed by transmission electron microscope, emission spectrum scanning, and dynamic light scattering. Rat kidney cells (NRK) were exposed to different doses of CdSe/ZnS QDs with or without N-acetylcysteine (NAC, antioxidant). Then, cellular uptake of CdSe/ZnS QDs was detected, and in vitro cytotoxicity was evaluated by MTT assay and TUNEL assay.

Involvement of Klotho, TNF‑&alpha; and ADAMs in radiation‑induced senescence of renal epithelial cells.

While radiation nephropathy is a major problem associated with radiotherapy, the exact mechanisms underlying its pathogenesis and the mediators involved in kidney deterioration remain to be elucidated. In view of the finding that senescence is typically increased post-irradiation, the present study examined whether ionizing radiation may cause kidney injury by enhancing premature senescence. The present study explored the relevance of the aging suppressor, Klotho, which has anti-aging activity and is highly expressed in murine renal cells/kidney tissues, under irradiation conditions. Firstly, the effects of radiation on mouse inner medullary collecting duct-3 (mIMCD-3) cells and kidney tissues of mice were assessed. Subsequently, the mRNA expression levels of Klotho, TNF-α and ADAM metallopeptidase domain (ADAM)9/10/17 were analyzed by reverse transcription-quantitative PCR following exposure to radiation. In addition, the levels of these proteins were measured by western blotting or ELISA. The results revealed that irradiation of mIMCD-3 cells clearly triggered cellular senescence. Notably, Klotho gene expression was considerably decreased in radiation-exposed mIMCD-3 cells and in the kidney tissues of irradiated BALB/c mice, and the corresponding translated protein was consistently expressed following radiation exposure. Moreover, expression of TNF-α, a negative regulator of Klotho, was significantly increased, whereas ADAM9/10/17, an ectodomain shedding enzyme of Klotho, was decreased in irradiated mIMCD-3 cells and in the kidney tissues of BALB/c mice. Collectively, these data suggested that TNF-α-mediated inhibition of Klotho expression and blockage of soluble Klotho formation via decreased ADAM expression following irradiation may contribute to the development of renal dysfunction through acceleration of radiation-induced cellular senescence.

Dexmedetomidine Alleviates Lipopolysaccharide-Induced Acute Kidney Injury by Inhibiting p75NTR-Mediated Oxidative Stress and Apoptosis.

Oxidative stress and apoptosis play a key role in the pathogenesis of sepsis-associated acute kidney injury (AKI). Dexmedetomidine (DEX) may present renal protective effects in sepsis. Therefore, we studied antioxidant effects and the mechanism of DEX in an inflammatory proximal tubular epithelial cell model and lipopolysaccharide- (LPS-) induced AKI in mice. Methods. We assessed renal function (creatinine, urea nitrogen), histopathology, oxidative stress (malondialdehyde (MDA) and superoxide dismutase (SOD)), and apoptosis (TUNEL staining and Cleaved caspase-3) in mice. In vitro experiments including Cleaved caspase-3 and p75NTR/p38MAPK/JNK signaling pathways were evaluated using western blot. Reactive oxidative species (ROS) production and apoptosis were determined using flow cytometry. Results. DEX significantly improved renal function and kidney injury and also revert the substantially increased level of MDA concentrations as well as the reduction of the SOD enzyme activity found in LPS-induced AKI mice. In parallel, DEX treatment also reduced the apoptosis and Cleaved caspase-3 expression evoked by LPS. The expression of p75NTR was increased in kidney tissues of mice with AKI but decreased after treatment with DEX. In cultured human renal tubular epithelial cell line (HK-2 cells), DEX inhibited LPS-induced apoptosis and generation of ROS, but this was reversed by overexpression of p75NTR. Furthermore, pretreatment with DEX significantly downregulated phosphorylation of JNK and p38MAPK in LPS-stimulated HK-2 cells, and this effect was abolished by overexpression of p75NTR. Conclusion. DEX ameliorated AKI in mice with sepsis by partially reducing oxidative stress and apoptosis through regulation of p75NTR/p38MAPK/JNK signaling pathways.

Micromechanical poroelastic and viscoelastic properties of ex-vivo soft tissues

Soft biological tissues demonstrate strong time-dependent mechanical behavior, arising from their intrinsic viscoelasticity and fluid flow-induced poroelasticity. It is increasingly recognized that time-dependent mechanical properties of soft tissues influence their physiological functions and are linked to several pathological processes. Nevertheless, soft tissue time-dependent characteristics, especially their micromechanical variation with tissue composition and location, remain poorly understood. Nanoindentation is a well-established technique to measure local elastic properties but has not been fully explored to determine micro-scale time-dependent properties of soft tissues. Here, a nanoindentation-based experimental strategy is implemented to characterize the micro-scale poroelastic and viscoelastic behavior of mouse heart, kidney, and liver tissues. It is demonstrated that heart tissue exhibits substantial mechanical heterogeneity where the elastic modulus varies spatially from 1 to 30 kPa. In contrast, both kidney and liver tissues show relatively homogeneous response with elastic modulus 0.5–3 kPa. All three tissues demonstrate marked load relaxation under constant indentation, where the relaxation behavior is observed to be largely dominated by tissue viscoelasticity. Intrinsic permeability varies among different tissues, where heart tissue is found to be less permeable compared to kidney and liver tissues. Overall, the results presented herein provide key insights into the time-dependent micromechanical behavior of different tissues and can therefore contribute to studies of tissue pathology and tissue engineering applications.