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1.
Methodist Debakey Cardiovasc J ; 18(4): 27-33, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36132587

RESUMO

Amyloidosis encompasses a collection of disorders of pathological protein folding. The extracellular location where these "amyloid fibril" proteins are deposited determines the clinical presentation of the disease. The abnormal architecture of these fibrils makes them insoluble and not easily removed, leading to disruption of normal tissue structure and interference with normal physiology. Amyloidosis of the heart and kidney can be inherited, secondary to unrelated diseases, or due to a plasma cell disorder. This review will focus on immunoglobulin light chain amyloidosis, which is life-threatening and must be diagnosed as early as possible by employing precise and accurate typing to ensure timely and frequently curative therapy.


Assuntos
Amiloidose , Amiloidose de Cadeia Leve de Imunoglobulina , Amiloide/química , Amiloide/metabolismo , Amiloide/uso terapêutico , Amiloidose/complicações , Amiloidose/diagnóstico , Amiloidose/terapia , Coração , Humanos , Amiloidose de Cadeia Leve de Imunoglobulina/diagnóstico , Amiloidose de Cadeia Leve de Imunoglobulina/terapia , Rim/metabolismo
2.
Front Endocrinol (Lausanne) ; 13: 932286, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36133305

RESUMO

2-Hydroxyglutarate (2HG) overproducing tumors arise in a number of tissues, including the kidney. The tumorigenesis resulting from overproduced 2HG has been attributed to the ability of 2HG alter gene expression by inhibiting α-ketoglutarate (αKG)-dependent dioxygenases, including Ten-eleven-Translocation (TET) enzymes. Genes that regulate cellular differentiation are reportedly repressed, blocking differentiation of mesenchymal cells into myocytes, and adipocytes. In this report, the expression of the enzyme responsible for L2HG degradation, L-2HG dehydrogenase (L2HGDH), is knocked down, using lentiviral shRNA, as well as siRNA, in primary cultures of normal Renal Proximal Tubule (RPT) cells. The knockdown (KD) results in increased L-2HG levels, decreased demethylation of 5mC in genomic DNA, and increased methylation of H3 Histones. Consequences include reduced tubulogenesis by RPT cells in matrigel, and reduced expression of molecular markers of differentiation, including membrane transporters as well as HNF1α and HNF1ß, which regulate their transcription. These results are consistent with the hypothesis that oncometabolite 2HG blocks RPT differentiation by altering the methylation status of chromatin in a manner that impedes the transcriptional events required for normal differentiation. Presumably, similar alterations are responsible for promoting the expansion of renal cancer stem-cells, increasing their propensity for malignant transformation.


Assuntos
Dioxigenases , Histonas , Diferenciação Celular/genética , Cromatina , Dioxigenases/metabolismo , Epigênese Genética , Glutaratos , Histonas/metabolismo , Ácidos Cetoglutáricos/metabolismo , Rim/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Oxirredutases/metabolismo , RNA Interferente Pequeno
3.
Cytokine ; 159: 156000, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36058192

RESUMO

BACKGROUND: Renal interstitial fibrosis (RIF) is the main pathological change of a variety of chronic kidney diseases (CKD). Epigenetic modifications of fibrosis-prone genes regulate RIF progression. This study aimed to investigate long non-coding RNA (lncRNA) N6-methyladenosine (m6A) modification and its role in regulating RIF progression. METHODS: Unilateral ureteral occlusion (UUO) was employed to construct the RIF in vivo model; and TGF-ß1-treated HK-2 and HKC-8 cells were used for in vitro experiments. The mRNA and protein expressions were assessed using qRT-PCR and western blot. The proliferation and migration were evaluated by EdU assay and transwell assay, respectively. In addition, levels of inflammatory cytokines were determined by ELISA assay and qRT-PCR. Moreover, lncRNA GAS5 m6A level was detected using Me-RIP assay. HE and Masson staining were employed to evaluate fibrotic lesions of the kidney. RESULTS: FTO expression was elevated in HK-2 and HKC-8 cells after TGF-ß1 treatment and mouse kidney tissue following UUO, and lncRNA GAS5 was downregulated. LncRNA GAS5 overexpression or FTO silencing suppressed TGF-ß1-induced the increase of EMT-related proteins (Vimentin, Snail and N-cadherin) and inflammatory cytokines (IL-6, IL-1ß and TNF-α) levels in HK-2 cells. FTO suppressed lncRNA GAS5 expression by reducing the m6A modification of lncRNA GAS5. Additionally, FTO knockdown could suppress EMT process and inflammation response induced by TGF-ß1 and UUO in vitro and in vivo. As expected, FTO knockdown abrogated the promotion effects of lncRNA GAS5 silencing on TGF-ß1-induced EMT process and inflammation response in HK-2 and HKC-8 cells. CONCLUSION: FTO promoted EMT process and inflammation response through reducing the m6A modification of lncRNA GAS5.


Assuntos
RNA Longo não Codificante , Insuficiência Renal Crônica , Obstrução Ureteral , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Animais , Caderinas , Transição Epitelial-Mesenquimal/genética , Fibrose , Inflamação/genética , Inflamação/patologia , Interleucina-6/farmacologia , Rim/metabolismo , Camundongos , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Mensageiro , Insuficiência Renal Crônica/genética , Insuficiência Renal Crônica/patologia , Fator de Crescimento Transformador beta1/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Obstrução Ureteral/genética , Obstrução Ureteral/metabolismo , Vimentina
4.
BMC Urol ; 22(1): 152, 2022 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-36104680

RESUMO

BACKGROUND: Kidney renal papillary cell carcinoma (KIRP) is a dangerous cancer, which accounts for 15-20% of all kidney malignancies. Ferroptosis is a rare kind of cell death that overcomes medication resistance. Ferroptosis-related long non-coding RNAs (LNCRNAs) in KIRP, remain unknown. METHOD: We wanted to express how ferroptosis-related LNCRNAs interact with immune cell infiltration in KIRP. Gene set enrichment analysis in the GO and KEGG databases were used to explore gene expression enrichment. The prognostic model was constructed using Lasso regression. In addition, we also analyzed the modifications in the tumor microenvironment (TME) and immunological association. RESULT: The expression of LNCRNA was closely connected to the ferroptosis, according to co-expression analyses. CASC19, AC090197.1, AC099850.3, AL033397.2, LINC00462, and B3GALT1-AS1 were found to be significantly increased in the high-risk group, indicating that all of these markers implicates the malignancy processes for KIRP patients and may be cancer-promoting variables. LNCTAM34A and AC024022.1 were shown to be significantly elevated in the low-risk group; these might represent as the KIRP tumor suppressor genes. According to the TCGA, CCR, and inflammation-promoting genes were considered to be significantly different between the low-risk and high-risk groups. The expression of CD160, TNFSF4, CD80, BTLA, and TNFRSF9 was different in the two risk groups. CONCLUSION: LNCRNAs associated with ferroptosis were linked to the occurrence and progression of KIRP. Ferroptosis-related LNCRNAs and immune cell infiltration in the TME may be potential biomarkers in KIRP that should be further investigated.


Assuntos
Carcinoma de Células Renais , Ferroptose , Neoplasias Renais , RNA Longo não Codificante , Biomarcadores , Carcinoma de Células Renais/genética , Ferroptose/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Rim/metabolismo , Neoplasias Renais/genética , Ligante OX40/genética , Ligante OX40/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Microambiente Tumoral
5.
Physiol Rep ; 10(18): e15470, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-36117297

RESUMO

Excessive fructose consumption has been associated with hypertension and metabolic disorders and can alter physiological adaptations during pregnancy, with long-term detrimental consequences. This study evaluated in post-weaning mothers the effects of increased fructose consumption during pregnancy and lactation on blood pressure and renal function. Female Wistar rats were assigned to one of four experimental groups: non-pregnant control (NPC); pregnant control (PC); non-pregnant fructose (NPF), and pregnant fructose (PF). Control rats had free access to food and water, while the fructose groups had free access to food and to a 20% fructose solution, over the time period of the experiment. The systolic BP and renal function parameters were measured at the end of the experimental period, one week after weaning (28 days after delivery). The results were presented as means ± standard error. Higher values of BP were observed in both pregnant and non-pregnant rats treated with fructose compared to control. Creatinine clearance was reduced only in the PF group; however, both the PF and NPF groups had reduced Na+ and K+ excretions. In the PF group, there was also glomerular enlargement and changes in the media/lumen (M/L) ratio of interlobular arteries. Additionally, the PF group showed increased macrophage infiltration and expression of alpha-SM-actin and reduced expression of nitric-oxide-synthase endothelial in renal tissue. These findings suggest that the association of high fructose intake with pregnancy aggravated kidney changes that persisted for up to four weeks after delivery, which may represent a risk factor for maternal health.


Assuntos
Actinas , Efeitos Tardios da Exposição Pré-Natal , Animais , Creatinina , Feminino , Frutose , Humanos , Rim/metabolismo , Lactação , Óxidos/farmacologia , Gravidez , Ratos , Ratos Wistar , Água
6.
Biomed Res Int ; 2022: 3002353, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36119929

RESUMO

lncRNAs and mRNA are closely associated with hypertensive renal damage, and Astragalus membranaceus and Salvia miltiorrhiza (AS) have a therapeutic effect; however, the mechanism of AS to ameliorate hypertensive renal damage through the co-expression network of lncRNA-mRNA was unclear. In this study, we investigated the role of AS regulated the coexpression network of lncRNA-mRNA in improving hypertensive renal damage. Sixteen 24-week old spontaneous hypertensive rats (SHRs) were randomly divided into model group (M) and drug intervention group (AS, 5.9 g/kg), 8 Wistar Kyoto rats (WKY) of the same age as normal group (N). The treatment of rats was 4 weeks. Detecting the change of blood pressure, renal pathology and renal function related indicators, and lncRNA and mRNA sequencing and joint analysis was performed on the kidney. AS reduced blood pressure; decreased urine NAG, urine mALB, serum CysC, and IL-6; and improved renal pathology compared with group M. Simultaneously, AS reversed the disordered expression of 178 differential expression (DE) mRNAs and 237 DE-lncRNAs in SHRs, and their joint analysis showed that 13 DE-mRNAs and 32 DE-lncRNAs were coexpressed. Further analysis of 13 coexpressed DE-mRNAs showed negative regulation of blood pressure and fatty acid beta-oxidation was highly enriched in GO pathways, PPAR signaling pathway was highly enriched in KEGG pathways, and the verification related to these pathways was also highly consistent with the sequence. AS can alleviate hypertensive renal damage through the coexpression network of lncRNA-mRNA, of which coexpressed 13 DE-mRNAs and 32 DE-lncRNAs were the important targets, and the pathway negative regulation of blood pressure, fatty acid beta-oxidation, and PPAR signaling pathway play a major regulatory role.


Assuntos
Hipertensão , RNA Longo não Codificante , Salvia miltiorrhiza , Animais , Astragalus propinquus , Ácidos Graxos , Hipertensão/tratamento farmacológico , Hipertensão/genética , Interleucina-6 , Rim/metabolismo , Receptores Ativados por Proliferador de Peroxissomo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos SHR
7.
J Transl Med ; 20(1): 420, 2022 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-36104729

RESUMO

Diabetic kidney disease (DKD) is the leading cause of end-stage renal disease (ESRD) worldwide. SGLT2 inhibitors are clinically effective in halting DKD progression. However, the underlying mechanisms remain unclear. The serum and kidneys of mice with DKD were analyzed using liquid chromatography with tandem mass spectrometry (LC-MS/MS)-based metabolomic and proteomic analyses. Three groups were established: placebo-treated littermate db/m mice, placebo-treated db/db mice and EMPA-treated db/db mice. Empagliflozin (EMPA) and placebo (10 mg/kg/d) were administered for 12 weeks. EMPA treatment decreased Cys-C and urinary albumin excretion compared with placebo by 78.60% and 57.12%, respectively (p < 0.001 in all cases). Renal glomerular area, interstitial fibrosis and glomerulosclerosis were decreased by 16.47%, 68.50% and 62.82%, respectively (p < 0.05 in all cases). Multi-omic analysis revealed that EMPA treatment altered the protein and metabolic profiles in the db/db group, including 32 renal proteins, 51 serum proteins, 94 renal metabolites and 37 serum metabolites. Five EMPA-related metabolic pathways were identified by integrating proteomic and metabolomic analyses, which are involved in renal purine metabolism; pyrimidine metabolism; tryptophan metabolism; nicotinate and nicotinamide metabolism, and glycine, serine and threonine metabolism in serum. In conclusion, this study demonstrated metabolic reprogramming in mice with DKD. EMPA treatment improved kidney function and morphology by regulating metabolic reprogramming, including regulation of renal reductive stress, alleviation of mitochondrial dysfunction and reduction in renal oxidative stress reaction.


Assuntos
Diabetes Mellitus , Nefropatias Diabéticas , Inibidores do Transportador 2 de Sódio-Glicose , Animais , Compostos Benzidrílicos , Cromatografia Líquida , Diabetes Mellitus/tratamento farmacológico , Nefropatias Diabéticas/tratamento farmacológico , Nefropatias Diabéticas/metabolismo , Glucosídeos , Rim/metabolismo , Camundongos , Proteômica , Inibidores do Transportador 2 de Sódio-Glicose/metabolismo , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Espectrometria de Massas em Tandem
8.
Physiol Rep ; 10(18): e15468, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-36117389

RESUMO

Meprin metalloproteinases have been implicated in the pathophysiology of ischemia/reperfusion (IR)-induced kidney injury. Previous in vitro data showed that meprin ß proteolytically processes interleukin-6 (IL-6) resulting in its inactivation. Recently, meprin-ß was also shown to cleave the IL-6 receptor. The goal of this study was to determine how meprin ß expression impacts IL-6 and downstream modulators of the JAK2-STAT3-mediated signaling pathway in IR-induced kidney injury. IR was induced in 12-week-old male wild-type (WT) and meprin ß knockout (ßKO) mice and kidneys obtained at 24 h post-IR. Real-time PCR, western blot, and immunostaining/microscopy approaches were used to quantify mRNA and protein levels respectively, and immunofluorescence counterstaining with proximal tubule (PT) markers to determine protein localization. The mRNA levels for IL-6, CASP3 and BCL-2 increased significantly in both genotypes. Interestingly, western blot data showed increases in protein levels for IL-6, CASP3, and BCL-2 in the ßKO but not in WT kidneys. However, immunohistochemical data showed increases in IL-6, CASP3, and BCL-2 proteins in select kidney tubules in both genotypes, shown to be PTs by immunofluorescence counterstaining. IR-induced increases in p-STAT-3 and p-JAK-2 in ßKO at a global level but immunoflourescence counterstaining demonstrated p-JAK2 and p-STAT3 increases in select PT for both genotypes. BCL-2 increased only in the renal corpuscle of WT kidneys, suggesting a role for meprins expressed in leukocytes. Immunohistochemical analysis confirmed higher levels of leukocyte infiltration in WT kidneys when compared to ßKO kidneys. The present data demonstrate that meprin ß modulates IR-induced kidney injury in part via IL-6/JAK2/STAT3-mediated signaling.


Assuntos
Interleucina-6 , Traumatismo por Reperfusão , Animais , Caspase 3/metabolismo , Interleucina-6/metabolismo , Isquemia/metabolismo , Rim/metabolismo , Masculino , Metaloendopeptidases , Metaloproteases/metabolismo , Camundongos , Camundongos Knockout , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/metabolismo , Receptores de Interleucina-6/metabolismo , Reperfusão , Traumatismo por Reperfusão/metabolismo , Transdução de Sinais
9.
Lipids Health Dis ; 21(1): 90, 2022 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-36123608

RESUMO

OBJECTIVE: We wanted to explore how angiopoietin-like 3 (ANGPTL3) impact hyperlipidemia-induced renal injury. METHODS: ANGPTL3 knockout mice and wild-type C57 mice were set up in four groups (N = 5) depending on a normal or 60% high-fat diet: wild-type with normal diet (WT), angptl3-/- with normal diet (KO), wild-type + high-fat diet (WT + HF) and angptl3-/- + high-fat diet (KO + HF). The detection time points were the 9th, 13th, 17th and 21st weeks after modeling. Serum lipid and urinary protein levels of mice in each group were detected, and pathological changes in the kidney were analyzed. Moreover, the expression of ANGPTL3, α-actinin-4 (ACTN4), CD2-associated protein (CD2AP) and podocin was tested in the glomerulus by immunohistochemistry (IHC). RESULTS: In the WT + HF group, hyperlipidemia and proteinuria could be observed at the 9th week and were gradually aggravated with time. Compared with WT + HF mice, the levels of serum lipids and proteinuria in KO + HF mice were significantly reduced, and the width of podocyte foot processes (FPs) fusion was also markedly improved. The IHC results suggested that in WT + HF mice, the expression of ANGPTL3 was significantly enhanced. After modeling, ACTN4 expression was markedly weakened in the glomeruli of WT + HF mice. Different to WT mice, ACTN4 expression was not observed obviously change in KO + HF mice. Compared with the normal diet group, the expression of podocin showed a decline in WT mice treated with high-fat diet and showed a significant difference from the 17th week. In addition, podocin expression in KO + HF glomeruli was also found to be weak but not significantly different from that in WT + HF glomeruli at the four time points. The expression of CD2AP showed similar results among the four groups. CONCLUSION: ANGPTL3 could play a role in the mechanism of hyperlipidemia-associated podocyte injury via ACTN4.


Assuntos
Dieta Hiperlipídica , Hiperlipidemias , Actinina/genética , Proteína 3 Semelhante a Angiopoietina , Proteínas Semelhantes a Angiopoietina/genética , Proteínas Semelhantes a Angiopoietina/metabolismo , Angiopoietinas , Animais , Dieta Hiperlipídica/efeitos adversos , Hiperlipidemias/genética , Hiperlipidemias/metabolismo , Rim/metabolismo , Lipídeos , Camundongos , Camundongos Knockout , Proteinúria
10.
FASEB J ; 36(10): e22545, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36094323

RESUMO

The kidneys are radiosensitive and dose-limiting organs for radiotherapy (RT) targeting abdominal and paraspinal tumors. Excessive radiation doses to the kidneys ultimately lead to radiation nephropathy. Our prior work unmasked a novel role for the lipid-modifying enzyme, sphingomyelin phosphodiesterase acid-like 3b (SMPDL3b), in regulating the response of renal podocytes to radiation injury. In this study, we investigated the role of SMPDL3b in DNA double-strand breaks (DSBs) repair in vitro and in vivo. We assessed the kinetics of DSBs recognition and repair along with the ATM pathway and nuclear sphingolipid metabolism in wild-type (WT) and SMPDL3b overexpressing (OE) human podocytes. We also assessed the extent of DNA damage repair in SMPDL3b knock-down (KD) human podocytes, and C57BL6 WT and podocyte-specific SMPDL3b-knock out (KO) mice after radiation injury. We found that SMPDL3b overexpression enhanced DSBs recognition and repair through modulating ATM nuclear shuttling. OE podocytes were protected against radiation-induced apoptosis by increasing the phosphorylation of p53 at serine 15 and attenuating subsequent caspase-3 cleavage. SMPDL3b overexpression prevented radiation-induced alterations in nuclear ceramide-1-phosphate (C1P) and ceramide levels. Interestingly, exogenous C1P pretreatment radiosensitized OE podocytes by delaying ATM nuclear foci formation and DSBs repair. On the other hand, SMPDL3b knock-down, in vitro and in vivo, induced a significant delay in DSBs repair. Additionally, increased activation of apoptosis was induced in podocytes of SMPDL3b-KO mice compared to WT mice at 24 h post-irradiation. Together, our results unravel a novel role for SMPDL3b in radiation-induced DNA damage response. The current work suggests that SMPDL3b modulates nuclear sphingolipid metabolism, ATM nuclear shuttling, and DSBs repair.


Assuntos
Podócitos , Lesões por Radiação , Animais , Ceramidas/metabolismo , Quebras de DNA de Cadeia Dupla , Humanos , Rim/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Podócitos/metabolismo , Lesões por Radiação/genética , Lesões por Radiação/metabolismo , Esfingomielina Fosfodiesterase/genética , Esfingomielina Fosfodiesterase/metabolismo
11.
Anal Chem ; 94(37): 12604-12613, 2022 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-36067026

RESUMO

Core histones including H2A, H2B, H3, and H4 are key modulators of cellular repair, transcription, and replication within eukaryotic cells, playing vital roles in the pathogenesis of disease and cellular responses to environmental stimuli. Traditional mass spectrometry (MS)-based bottom-up and top-down proteomics allows for the comprehensive identification of proteins and of post-translational modification (PTM) harboring proteoforms. However, these methodologies have difficulties preserving near-cellular spatial distributions because they typically require laser capture microdissection (LCM) and advanced sample preparation techniques. Herein, we coupled a matrix-assisted laser desorption/ionization (MALDI) source with a Thermo Scientific Q Exactive HF Orbitrap MS upgraded with ultrahigh mass range (UHMR) boards for the first demonstration of complementary high-resolution accurate mass (HR/AM) measurements of proteoforms up to 16.5 kDa directly from tissues using this benchtop mass spectrometer. The platform achieved isotopic resolution throughout the detected mass range, providing confident assignments of proteoforms with low ppm mass error and a considerable increase in duty cycle over other Fourier transform mass analyzers. Proteoform mapping of core histones was demonstrated on sections of human kidney at near-cellular spatial resolution, with several key distributions of histone and other proteoforms noted within both healthy biopsy and a section from a renal cell carcinoma (RCC) containing nephrectomy. The use of MALDI-MS imaging (MSI) for proteoform mapping demonstrates several steps toward high-throughput accurate identification of proteoforms and provides a new tool for mapping biomolecule distributions throughout tissue sections in extended mass ranges.


Assuntos
Histonas , Proteômica , Análise de Fourier , Histonas/metabolismo , Humanos , Rim/metabolismo , Proteômica/métodos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
12.
Cell Physiol Biochem ; 56: 484-499, 2022 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-36126285

RESUMO

BACKGROUND/AIMS: In kidney, extracellular [Ca2+] can modulate intracellular [Ca2+] to control key cellular processes. Hence, extracellular [Ca2+] is normally maintained within narrow range. We tested effect of extracellular ATP on viability of human proximal (HK-2) cells at high calcium. Modulation of intracellular calcium was assessed by imaging cytosolic [Ca2+], and expression of calcium-binding proteins (CaBPs). We present an artificial intelligence enabled deep learning model for prediction of injury and protection against extracellular [Ca2+] in HK-2 cells. METHODS: HK-2 cells were cultured in calcium-free DMEM supplemented with CaCl2. Morphological changes were detected using light microscopy. Cell viability was determined using MTT Assay. Intracellular [Ca2+] was detected using fluorescence microscopy. For easy detection of HK-2 cells injury, we performed light microscopy image classification based on Convolutional Neural Network. Expression of CaBPs, p21, and Mcl-1 was measured using real-time PCR. RESULTS: We show decreased viability of HK-2 cells cultured in elevated calcium levels, which was prevented by adenosine triphosphate (ATP). Exposure of cells to elevated extracellular [Ca2+] correlated with increasing fluorescence of intracellular calcium indicator, which was attenuated in presence of ATP. Since features cannot be detected easily by human eyes, we propose a customized deep learning-based CNN model for classification of HK-2 cells injury by extracellular calcium with high accuracy of 98%. Our data demonstrated significant increase in mRNA levels of calmodulin, S100A8, S100A14 and CaBP28k, with elevated extracellular [Ca2+]. Expression of these genes was enhanced with ATP. CONCLUSION: The results suggest that ATP protects human proximal (HK-2) cells against elevated extracellular calcium levels. We present a CNN model as user friendly tool to study calcium dependent injury in (HK-2) cells. Finally, we show that ATP-mediated protection is correlated with enhanced expression of calcium-binding proteins.


Assuntos
Cálcio , Aprendizado Profundo , Trifosfato de Adenosina/metabolismo , Inteligência Artificial , Cálcio/metabolismo , Cloreto de Cálcio/metabolismo , Calmodulina/metabolismo , Humanos , Rim/metabolismo , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , RNA Mensageiro
13.
Int J Mol Sci ; 23(17)2022 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-36077583

RESUMO

The activity of drug transporters is central to the secretory function of the kidneys and a defining feature of renal proximal tubule epithelial cells (RPTECs). The expression, regulation, and function of these membrane-bound proteins is well understood under normal renal physiological conditions. However, the impact of drug transporters on the pathophysiology of kidney cancer is still elusive. In the present study, we employed different renal cell carcinoma (RCC) cell lines and a prototypical non-malignant RPTEC cell line to characterize the activity, expression, and potential regulatory mechanisms of relevant renal drug transporters in RCC in vitro. An analysis of the uptake and efflux activity, the expression of drug transporters, and the evaluation of cisplatin cytotoxicity under the effects of methylation or epidermal growth factor receptor (EGFR) inhibition showed that the RCC cells retained substantial drug transport activity. In RCC cells, P-glycoprotein was localized in the nucleus and its pharmacological inhibition enhanced cisplatin toxicity in non-malignant RPTECs. On the other hand, methylation inhibition enhanced cisplatin toxicity by upregulating the organic cation uptake activity in RCC cells. Differential effects of methylation and EGFR were observed in transporter expression, showing regulatory heterogeneity in these cells. Interestingly, the non-malignant RPTEC cell line that was used lacked the machinery responsible for organic cation transport, which reiterates the functional losses that renal cells undergo in vitro.


Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Carcinoma de Células Renais/metabolismo , Cátions/metabolismo , Cisplatino/metabolismo , Cisplatino/farmacologia , Receptores ErbB/metabolismo , Humanos , Rim/metabolismo , Neoplasias Renais/metabolismo , Túbulos Renais Proximais/metabolismo , Proteínas de Membrana Transportadoras/metabolismo
14.
Int J Mol Sci ; 23(17)2022 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-36077596

RESUMO

Acute kidney injury (AKI) is a common renal injury leading to relevant morbidity and mortality worldwide. Most of the clinical cases of AKI are caused by ischemia reperfusion (I/R) injury with renal ischemia injury followed by reperfusion injury and activation of the innate immune response converging to NF-ĸB pathway induction. Despite the clear role of NF-ĸB in inflammation, it has recently been acknowledged that NF-ĸB may impact other cell functions. To identify NF-ĸB function with respect to metabolism, vascular function and oxidative stress after I/R injury and to decipher in detail the underlying mechanism, we generated a transgenic mouse model with targeted deletion of IKKß along the tubule and applied I/R injury followed by its analysis after 2 and 14 days after I/R injury. Tubular IKKß deletion ameliorated renal function and reduced tissue damage. RNAseq data together with immunohistochemical, biochemical and morphometric analysis demonstrated an ameliorated vascular organization and mRNA expression profile for increased angiogenesis in mice with tubular IKKß deletion at 2 days after I/R injury. RNAseq and protein analysis indicate an ameliorated metabolism, oxidative species handling and timely-adapted cell proliferation and apoptosis as well as reduced fibrosis in mice with tubular IKKß deletion at 14 days after I/R injury. In conclusion, mice with tubular IKKß deletion upon I/R injury display improved renal function and reduced tissue damage and fibrosis in association with improved vascularization, metabolism, reactive species disposal and fine-tuned cell proliferation.


Assuntos
Injúria Renal Aguda , Traumatismo por Reperfusão , Injúria Renal Aguda/genética , Injúria Renal Aguda/metabolismo , Animais , Apoptose/genética , Fibrose , Quinase I-kappa B/genética , Isquemia , Rim/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , NF-kappa B/metabolismo , Traumatismo por Reperfusão/genética
15.
J Clin Pharmacol ; 62 Suppl 1: S129-S139, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-36106785

RESUMO

Physiologically based pharmacokinetic modeling (PBPK) could be used to predict changes in exposure during pregnancy and possibly inform medicine use in pregnancy in situations where there are currently no available clinical data. The Medicines and Healthcare Product Regulatory Agency has been evaluating the available models for a number of medicines cleared by the kidney. Models were evaluated for ceftazidime, cefuroxime, metformin, oseltamivir, and amoxicillin. Because the passive renal process contributes significantly to the renal elimination of these drugs and changes of the process during pregnancy have been implemented in existing pregnancy physiology models, simulations using these models can reasonably describe the pharmacokinetics of ceftazidime changes during pregnancy and appears to generally capture the changes in the other medicines; however, there are insufficient data on drugs solely passively cleared to fully qualify the models. In addition, in many cases, active transport processes are involved in a drug's renal clearance. Knowledge of changes in renal transport functions during pregnancy is emerging, and incorporation of such changes in current physiologically based pharmacokinetic modeling software is a work in progress. Filling this gap is expected to further enhance predictive performance of the models and increase the confidence in predicting pharmacokinetic changes in pregnant women for other renally cleared drugs.


Assuntos
Ceftazidima , Modelos Biológicos , Feminino , Humanos , Rim/metabolismo , Testes de Função Renal , Gravidez , Eliminação Renal
16.
Front Immunol ; 13: 929244, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36059489

RESUMO

Background: Monocytes are involved in the pathogenesis of ANCA-associated vasculitis (AAV). Monocyte/macrophages are the dominant infiltrating cells in the glomeruli of patients with myeloperoxidase-AAV (MPO-AAV). However, how human monocyte subsets extravasate to the kidney in MPO-AAV with renal damage is unclear. Methods: 30 MPO-AAV patients with renal damage and 22 healthy controls were enrolled in this study. Monocyte subsets and monocyte-related chemokines in the blood and kidneys of MPO-AAV patients were detected. The chemoattractant activity of the CX3CL1-CX3CR1 axis on CD16+ monocytes was observed. The effect of MPO-ANCA on the migration of CD16+ monocytes to human glomerular endothelial cells (HGECs) was detected by flow cytometry and transwell migration assay. Results: Compared with controls, CD16+ monocytes were significantly decreased in the blood and increased in the glomeruli of MPO-AAV patients with renal damage. The level of CX3CL1, but not CCL2, was significantly increased in the plasma of MPO-AAV patients. CX3CL1 co-localized with glomerular endothelial cells in MPO-AAV patients with renal damage. Moreover, we initially found that MPO-ANCA promotes an increase of the chemokine CX3CL1 on HGECs, imposing recruitment on CD16+ monocytes. Finally, the percentage of CD16+ monocytes in the blood was found to be positively correlated with estimated glomerular filtration rate (eGFR) and negatively correlated with urinary protein creatinine ratio in MPO-AAV patients with renal damage. Furthermore, the urinary protein creatinine ratio was positively correlated with the infiltrating of CD14+ and CD16+ cells in the kidneys. Conclusion: Enhanced extravasation of CD16+ monocytes to the kidney via the CX3CL1-CX3CR1 axis may be involved in renal damage in MPO-AAV.


Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos , Quimiocina CX3CL1 , Monócitos , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/genética , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/metabolismo , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/patologia , Anticorpos Anticitoplasma de Neutrófilos/metabolismo , Quimiocina CX3CL1/genética , Quimiocina CX3CL1/metabolismo , Creatinina , Células Endoteliais/metabolismo , Humanos , Rim/metabolismo , Rim/patologia , Monócitos/metabolismo , Peroxidase/efeitos adversos , Peroxidase/metabolismo
17.
Cell Death Dis ; 13(9): 770, 2022 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-36068197

RESUMO

Mixed lineage leukemia 1 (MLL1) is a histone H3 lysine 4 (H3K4) methyltransferase that interacts with WD repeat domain 5 (WDR5) to regulate cell survival, proliferation, and senescence. The role of MLL1 in the pathogenesis of acute kidney injury (AKI) is unknown. In this study, we demonstrate that MLL1, WDR5, and trimethylated H3K4 (H3K4me3) were upregulated in renal tubular cells of cisplatin-induced AKI in mice, along with increased phosphorylation of p53 and decreased expression of E-cadherin. Administration of MM102, a selective MLL1/WDR5 complex inhibitor, improved renal function and attenuated tubular injury and apoptosis, while repressing MLL1, WDR5, and H3K4me3, dephosphorylating p53 and preserving E-cadherin. In cultured mouse renal proximal tubular cells (RPTCs) exposed to cisplatin, treatment with MM102 or transfection with siRNAs for either MLL1 or WDR5 also inhibited apoptosis and p53 phosphorylation while preserving E-cadherin expression; p53 inhibition with Pifithrin-α lowered cisplatin-induced apoptosis without affecting expression of MLL1, WDR5, and H3K4me3. Interestingly, silencing of E-cadherin offset MM102's cytoprotective effects, but had no effect on p53 phosphorylation. These findings suggest that MLL1/WDR5 activates p53, which, in turn, represses E-cadherin, leading to apoptosis during cisplatin-induced AKI. Further studies showed that MM102 effectively inhibited cisplatin-triggered DNA damage response (DDR), as indicated by dephosphorylation of ataxia telangiectasia mutated (ATM) and ATM and Rad-3 related (ATR) proteins, dephosphorylation of checkpoint kinase 1 and 2 (Chk1 and Chk2); depression of γ-H2AX; and restrained cell cycle arrest, as evidenced by decreased expression of p21 and phospho-histone H3 at serine 10 in vitro and in vivo. Overall, we identify MLL1 as a novel DDR regulator that drives cisplatin-induced RPTC apoptosis and AKI by modulating the MLL1/WDR5-/ATR/ATM-Chk-p53-E-cadherin axis. Targeting the MLL1/WDR5 complex may have a therapeutic potential for the treatment of AKI.


Assuntos
Injúria Renal Aguda , Leucemia , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/genética , Injúria Renal Aguda/metabolismo , Animais , Apoptose , Caderinas/genética , Caderinas/metabolismo , Cisplatino/farmacologia , Histona Metiltransferases/metabolismo , Histonas/metabolismo , Rim/metabolismo , Leucemia/tratamento farmacológico , Camundongos , Proteína de Leucina Linfoide-Mieloide/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
18.
Virol J ; 19(1): 147, 2022 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-36096838

RESUMO

BACKGROUND: Human cytomegalovirus (CMV) can establish a latent infection with periodic or sporadic reactivation after the first infection happens. Primary and recurrent infection, results in different problems in patients with impaired or immature immune systems, such as kidney transplant recipients (KTRs). MicroRNAs (miRNAs, miRs) are important regulatory molecules in the outcome of CMV-infected KTRs. Therefore, in this study the expression level of CMV miRNAs were evaluated in active vs. latent CMV infected KTRs. METHODS: Expression of viral miRNAs were studied in 61 KTRs which were divided into 30 active CMV and 31 latent CMV infected individuals. In order to study the expression level of selected miRNAs, SYBR Green Real-time PCR technique was exploited. Also, mature miRNAs expression level that were produced from one precursor, studied both in active and latent situations. RESULTS: Among studied miRNAs' expression level, CMV miR-UL112-3p/5p, -UL22A-3p/5p, -US25-1-5p, -US25-2-3p/5p, -UL36-3p/5p and -UL70-3p showed significant increase in active CMV infected KTRs in comparison to latent ones. The ROC curve analysis results for miR-UL112-3p, -UL22A-3p, -US25-2-3p, -UL36-3p and -UL70-3p showed significant difference between two studied patient groups. CONCLUSION: This study revealed an extremely high expression level in CMV miR-UL112-3p/5p, -UL22A-3p/5p, -US25-1-5p, -US25-2-3p/5p, -UL36-3p/5p and -UL70-3p in active CMV infected KTRs in comparison to latent ones. Further studies might help in finding the capability of miRNAs to differentiate active from latent stage of CMV infection in KTRs.


Assuntos
Infecções por Citomegalovirus , MicroRNAs , Citomegalovirus/genética , Humanos , Rim/metabolismo , MicroRNAs/genética , Reação em Cadeia da Polimerase em Tempo Real
19.
J Clin Invest ; 132(16)2022 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-35968780

RESUMO

Understanding the loss of kidney function resulting from kidney aging has become an emerging research focus that will facilitate the future development of antisenolytic treatments. In this issue of the JCI, Pippin et al. first identified PD-1 upregulation in the aged mouse podocyte via unbiased RNA-seq analysis. Overexpression of PD-1 in immortalized mouse podocytes induced cell death and a senescence-associated secretory phenotype, suggesting the pathological role of PD-1 upregulation in aged podocytes. Blocking PD-1 signaling via a neutralizing anti-PD-1 antibody reversed the aged phenotype in the aged mice and ameliorated proteinuria in an experimental focal segmental glomerulosclerosis (FSGS) mouse model. These findings highlight the role of PD-1 signaling in kidney aging and its therapeutic potential for human clinical trials.


Assuntos
Glomerulosclerose Segmentar e Focal , Podócitos , Idoso , Animais , Glomerulosclerose Segmentar e Focal/genética , Glomerulosclerose Segmentar e Focal/patologia , Humanos , Rim/metabolismo , Glomérulos Renais/patologia , Camundongos , Podócitos/patologia , Proteinúria/patologia
20.
Fish Shellfish Immunol ; 128: 228-237, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35940536

RESUMO

2,2,4,4-tetra-brominated diphenyl ether (PBDE-47)-the dominant homologue of polybrominated diphenyl ethers-is a toxic environmental pollutant in the aquatic environment that continuously exists and bioaccumulates in the aquatic food chain. In experimental disease models, melatonin (MEL) has been reported to attenuate necroptosis and inflammatory responses. To further explore the mechanism underlying PBDE-47 toxicity and the mitigative impact of MEL detoxification, in this study, fish kidney cell models of PBDE-47 poisoning and/or MEL treatment were developed. The Ctenopharyngodon idellus kidney (CIK) cell line was treated with PBDE-47 (100 µM) and/or MEL (60 µM) for 24 h. Experimental data suggest that PBDE-47 exposure resulted in the enhancement of cytoplasmic Ca2+ concentration, induction of calcium dysmetabolism, decrease in the miR-140-5p miRNA level, upregulation of Toll-like Receptor 4 (TLR4) and nuclear factor-kappaB (NF-κB), triggering of receptor interacting serine/threonine kinase-induced necroptosis, and NF-κB pathway mediated secretion of inflammatory factors in CIK cells. PBDE-47-induced CIK cell damage could be mitigated by MEL through the regulation of calcium channels and the restoration of disorders of the miR-140-5p/TLR4/NF-κB axis. Overall, MEL relieved PBDE-47-induced necroptosis and the secretion of inflammatory factors through the miR-140-5p/TLR4/NF-κB axis. These findings enrich the current understanding of the toxicological molecular mechanisms of the PBDE-47 as well as the detoxification mechanisms of the MEL.


Assuntos
Poluentes Ambientais , Melatonina , MicroRNAs , Bifenil Polibromatos , Animais , Cálcio/metabolismo , Canais de Cálcio , Éter , Éteres Difenil Halogenados/toxicidade , Rim/metabolismo , Melatonina/farmacologia , MicroRNAs/genética , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Necroptose , Bifenil Polibromatos/toxicidade , Proteínas Serina-Treonina Quinases , Serina , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo
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