RESUMO
Mesangioproliferative glomerulonephritis is the most common nephritis worldwide. We examined the effects of low- and high-dose telmisartan, an angiotensin II receptor blocker, in rats with progressive anti-Thy1.1 mesangioproliferative glomerulonephritis in a clinically relevant situation of established renal damage. Uninephrectomized nephritic rats were randomized on day 28 to remain untreated (control treatment; CT), or to receive low- (0.1 mg/kg/day, LT) or high-dose telmisartan (10 mg/kg/day, HT), hydrochlorothiazide + hydralazine (8 + 32 mg/kg/day, HCT + H), or atenolol (100 mg/kg/day, AT). CT and LT rats were hypertensive, whereas HT, HCT + H and AT treatment normalized blood pressures. On day 131, despite similar blood lowering effects, only HT, but not AT or HCT + H, prevented loss of renal function and reduced proteinuria compared to CT. Only HT potently ameliorated glomerulosclerosis, tubulointerstitial damage, cortical matrix deposition, podocyte damage and macrophage infiltration. HT reduced cortical expression of platelet derived growth factor receptor-α and -ß as well as transforming growth factor-ß1. LT exhibited minor but significant efficacy even in the absence of antihypertensive effects. Transcript array analyses revealed a four-fold down-regulation of renal cortical chemokine (C-C motif) receptor 6 (CCR6) mRNA by HT, which was confirmed at the protein level. Silencing of CCR6 did not alter podocyte function in vitro, thus indicating a predominant role in the tubulo-interstitium. In human kidney biopsies, CCR6 mRNA and mRNA of its ligand chemokine (C-C motif) ligand 20 was up-regulated in patients with progressive IgA nephropathy compared to stable disease. Thus, delayed treatment with high-dose telmisartan exerted a pronounced benefit in progressive mesangioproliferative glomerulonephritis, which extended beyond that of equivalent blood pressure lowering. We identified down-regulation of platelet-derived growth factor receptors and CCR6 as potential mediators of telmisartan-related renoprotection. CCR6 may also regulate the renal outcome in human mesangioprolfierative glomerulonephritis.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Anti-Hipertensivos/farmacologia , Benzimidazóis/farmacologia , Benzoatos/farmacologia , Mesângio Glomerular/efeitos dos fármacos , Glomerulonefrite Membranoproliferativa/tratamento farmacológico , Bloqueadores do Receptor Tipo 1 de Angiotensina II/administração & dosagem , Animais , Anti-Hipertensivos/administração & dosagem , Atenolol/farmacologia , Benzimidazóis/administração & dosagem , Benzoatos/administração & dosagem , Pressão Sanguínea/efeitos dos fármacos , Desdiferenciação Celular/efeitos dos fármacos , Linhagem Celular , Quimiocina CCL20/genética , Citoproteção , Modelos Animais de Doenças , Fibrose , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Mesângio Glomerular/metabolismo , Mesângio Glomerular/patologia , Mesângio Glomerular/fisiopatologia , Glomerulonefrite Membranoproliferativa/etiologia , Glomerulonefrite Membranoproliferativa/genética , Glomerulonefrite Membranoproliferativa/metabolismo , Glomerulonefrite Membranoproliferativa/patologia , Glomerulonefrite Membranoproliferativa/fisiopatologia , Humanos , Hidralazina/farmacologia , Hidroclorotiazida/farmacologia , Hipertensão/tratamento farmacológico , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/fisiopatologia , Isoanticorpos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Nefrectomia , Podócitos/efeitos dos fármacos , Podócitos/metabolismo , Podócitos/patologia , Proteinúria/tratamento farmacológico , Proteinúria/metabolismo , Proteinúria/fisiopatologia , Interferência de RNA , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Receptores CCR6/genéticaRESUMO
Matricellular proteins participate in the pathogenesis of chronic kidney diseases. We analyzed glomerular gene expression profiles from patients with proteinuric diseases to identify matricellular proteins contributing to the progression of human nephropathies. Several genes encoding matricellular proteins, such as SPARC, THBS1, and CTGF, were induced in progressive nephropathies, but not in nonprogressive minimal-change disease. Periostin showed the highest induction, and its transcript levels correlated negatively with glomerular filtration rate in both glomerular and tubulointerstitial specimen. In well-preserved renal tissue, periostin localized to the glomerular tuft, the vascular pole, and along Bowman's capsule; no signal was detected in the tubulointerstitial compartment. Biopsies from patients with glomerulopathies and renal dysfunction showed enhanced periostin expression in the mesangium, tubular interstitium, and sites of fibrosis. Periostin staining correlated negatively with renal function. α-smooth muscle actin-positive mesangial and interstitial cells localized close to periostin-positive sites, as indicated by co-immunofluorescence. In vitro stimulation of mesangial cells by external addition of TGF-ß1 resulted in robust induction of periostin. Addition of periostin to mesangial cells induced cell proliferation and decreased the number of cells expressing activated caspase-3, a marker of apoptosis. These human data indicate for the first time a role of periostin in glomerular and interstitial injury in acquired nephropathies.
Assuntos
Moléculas de Adesão Celular/metabolismo , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose , Biópsia , Moléculas de Adesão Celular/genética , Proliferação de Células , Feminino , Fibrose , Humanos , Imuno-Histoquímica , Falência Renal Crônica/genética , Masculino , Células Mesangiais/metabolismo , Células Mesangiais/patologia , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Transporte Proteico , Proteinúria/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Preservação de Tecido , Transcrição Gênica , Adulto JovemRESUMO
The B-cell survival factors APRIL and BLyS are important for B-cell maturation and activation and contribute to human autoimmune diseases. Interference with B-cell function by targeting these molecules is currently being investigated in large clinical trials for systemic lupus erythematosus. The local expression patterns of APRIL and BLyS have not been investigated in detail in kidneys with lupus nephritis. We studied the mRNA expression of APRIL, BLyS, and the corresponding receptors BCMA, TACI, and BAFF-R in microdissected human biopsies with proliferative lupus nephritis (n=25) and compared it with pretransplant biopsies of living donors (n=9). APRIL and BLyS mRNA levels were significantly higher in glomeruli of patients with proliferative lupus nephritis (12- and 30-fold, respectively). Tubulointerstitial expression of APRIL, BLyS, BCMA, and TACI was also significantly elevated. To localize the respective proteins in the kidney, APRIL, BLyS, and BAFF-R were studied by immunohistochemistry in renal biopsies with proliferative (n=21) or membranous (n=8) lupus nephritis. APRIL was prominently expressed in glomeruli with proliferative, but not membranous, lupus nephritis. The staining pattern was consistent with mesangial cells. A prominent accumulation of CD68-positive cells was present in glomeruli in association with APRIL expression. APRIL, BLyS, and BAFF-R were also expressed in interstitial inflammatory cell accumulation. This is the first study, which details local expression of APRIL and BLyS in glomeruli and tubulointerstitium of human proliferative lupus nephritis. This information might help define intrarenal effects of APRIL and BLyS inhibition in human lupus nephritis.
Assuntos
Fator Ativador de Células B/metabolismo , Receptor do Fator Ativador de Células B/genética , Nefrite Lúpica/genética , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/genética , Adolescente , Adulto , Idoso , Fator Ativador de Células B/genética , Linfócitos B , Sobrevivência Celular , Feminino , Expressão Gênica , Glomerulonefrite Membranoproliferativa/genética , Glomerulonefrite Membranoproliferativa/patologia , Glomerulonefrite Membranosa/genética , Glomerulonefrite Membranosa/patologia , Humanos , Imuno-Histoquímica , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Nefrite Lúpica/patologia , Masculino , Microdissecção , Pessoa de Meia-Idade , RNA Mensageiro/genética , Adulto JovemRESUMO
In the kidney, hypoxia contributes to tubulointerstitial fibrosis, but little is known about its implications for glomerular damage and glomerulosclerosis. Chronic hypoxia was hypothesized to be involved in nephrosclerosis (NSC) or "hypertensive nephropathy." In the present study genome-wide expression data from microdissected glomeruli were studied to examine the role of hypoxia in glomerulosclerosis of human NSC. Functional annotation analysis revealed prominent regulation of hypoxia-associated biological processes in NSC, including angiogenesis, fibrosis, and inflammation. Glomerular expression levels of a majority of genes regulated by the hypoxia-inducible factors (HIFs) were significantly altered in NSC. Among these HIF targets, chemokine C-X-C motif receptor 4 (CXCR4) was prominently induced. Glomerular CXCR4 mRNA induction was confirmed by quantitative RT-PCR in an independent cohort with NSC but not in those with other glomerulopathies. By immunohistological analysis, CXCR4 showed enhanced positivity in podocytes in NSC biopsy specimens. This CXCR4 positivity was associated with nuclear localization of HIF1alpha only in podocytes of NSC, indicating transcriptional activity of HIF. As the CXCR4 ligand CXCL12/SDF-1 is constitutively expressed in podocytes, autocrine signaling may contribute to NSC. In addition, a blocking CXCR4 antibody caused significant inhibition of wound closure by podocytes in an in vitro scratch assay. These data support a role for CXCR4/CXCL12 in human NSC and indicate that hypoxia not only is involved in tubulointerstitial fibrosis but also contributes to glomerular damage in NSC.
Assuntos
Hipóxia/complicações , Nefropatias/etiologia , Nefroesclerose/complicações , Adolescente , Adulto , Idoso , Animais , Estudos de Casos e Controles , Células Cultivadas , Feminino , Perfilação da Expressão Gênica , Humanos , Hipóxia/genética , Nefropatias/genética , Nefropatias/patologia , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Nefroesclerose/genética , Análise de Sequência com Séries de Oligonucleotídeos , Oxigênio/farmacologia , Adulto JovemRESUMO
CXCR7 is an atypical receptor for the chemokines CXCL11 and CXCL12, which were found to be involved in animal models of allograft injury. We studied the expression of CXCR7 and its ligands in human kidneys by first quantifying the mRNA in 53 renal allograft biopsies. Receptor and ligand mRNAs were expressed in renal allografts, with a significant induction of CXCL11 and CXCL12 in biopsies showing borderline lesions and acute rejection. Immunohistochemical analysis for CXCR7 was performed in a series of 64 indication and 24 protocol biopsies. The indication biopsies included 46 acute rejections, 6 with interstitial fibrosis and tubular atrophy, and 12 pretransplant biopsies as controls. In control biopsies, CXCR7 protein was found on smooth muscle and on endothelial cells of a small number of peritubular vessels. The number of CXCR7-positive vessels was increased in acute rejection and, using double immunofluorescence labeling, a subset of these CXCR7-positive endothelial cells were identified as lymphatic vessels. Both CXCR7-positive blood and lymphatic vessels increased during allograft rejection. We found that CXCR7 is present in both blood and lymphatic endothelial cells in human renal allografts. Whether its presence modulates the formation of chemokine gradients and the recruitment of inflammatory cells will require further experimental studies.
Assuntos
Quimiocinas/metabolismo , Rim/metabolismo , Receptores de Quimiocinas/metabolismo , Biópsia , Proteínas de Transporte/genética , Proteínas de Transporte/imunologia , Proteínas de Transporte/metabolismo , Quimiocina CXCL11/genética , Quimiocina CXCL11/metabolismo , Quimiocina CXCL12/genética , Quimiocina CXCL12/metabolismo , Quimiocinas/genética , Quimiocinas/imunologia , Quimiocinas CXC/genética , Quimiocinas CXC/imunologia , Quimiocinas CXC/metabolismo , Células Endoteliais/imunologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Humanos , Rim/imunologia , Rim/patologia , Ligantes , Nefrectomia , RNA Mensageiro/imunologia , RNA Mensageiro/metabolismo , Receptores de Quimiocinas/genética , Receptores de Quimiocinas/imunologiaRESUMO
Renal proximal tubular epithelial cells, a target of infiltrating T cells during renal allograft rejection, may be protected from this injury by the cell surface protein CD274 (also termed PD-L1 for programmed death ligand 1). The co-inhibitory molecules PD-L1 (CD274) and PD-L2 (CD273) are ligands of PD-1 (programmed death 1; CD279). Here we determine the functional role of PD-1/PD-L pathways in human renal allograft rejection. Treatment of human primary tubular epithelial cells with interferon-beta and -gamma caused a dose-dependent and synergistic increase of PD-L1 and PD-L2 expression. Blockade of surface PD-L1, but not PD-L2, on interferon-treated tubular epithelial cells resulted in a significant increase in CD4+ T-cell proliferation and cytokine production by CD4+ and CD8+ T cells. The expression of PD-L1, PD-L2, and PD-1 mRNA and protein was upregulated in biopsies of patients with renal allograft rejection compared to the respective levels found in the pre-transplant biopsies. Induction of PD-L1 was significantly associated with acute vascular rejection. Our study suggests that the renal epithelial PD-1/PD-L1 pathway exerts an inhibitory effect of on alloreactive T-cell responses. The upregulation of PD-L1 on proximal tubular epithelial cells in patients with acute allograft rejection may reduce T-cell-mediated injury.
Assuntos
Antígenos CD/fisiologia , Túbulos Renais Proximais/metabolismo , Linfócitos T/imunologia , Antígeno B7-H1 , Proliferação de Células , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Rejeição de Enxerto/genética , Rejeição de Enxerto/imunologia , Humanos , Interferon beta/genética , Interferon beta/imunologia , Interferon beta/metabolismo , Ligantes , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Proteínas/genética , Proteínas/imunologia , Proteínas/metabolismo , Linfócitos T/metabolismo , Regulação para CimaRESUMO
The endoplasmic reticulum (ER) is an important site for protein folding and becomes "stressed" when its capacity to fold proteins is overwhelmed. In response, "unfolded protein response" (UPR) genes are induced, increasing the capacity to fold proteins; if the response is insufficient, then apoptosis ensues. For investigation of whether proteinuria and hyperglycemia induce ER stress in renal epithelial cells, microarray data from biopsies of established diabetic nephropathy (DN) were analyzed. Expression of UPR genes was significantly different in these biopsies than in control kidneys or biopsies of patients with mild DN, suggesting an association between the degree of DN and UPR gene expression. Expression of the transcription factor XBP1 and the ER chaperones HSPA5 and HYOU1 were increased, but the proapoptotic gene DDIT3 was unchanged. These findings were replicated in an independent cohort of patients with established DN by real-time reverse transcriptase-PCR. Immunofluorescence of renal biopsies from patients with DN confirmed the upregulation for HSPA5 and HYOU1 proteins in tubular epithelia. In biopsies of minimal-change disease, the mRNA levels of some ER stress molecules were also induced, but protein expression of HSPA5 and HYOU1 remained significantly lower than that observed in DN. Exposure of renal tubular epithelial cells to albumin and high glucose in vitro enhanced expression of genes involved in ER stress. These observations suggest that in proteinuric diseases, tubular epithelial cells undergo ER stress, which induces an adaptive, protective UPR. Although this may protect the cells from ER stress, persistence of hyperglycemia and proteinuria may eventually lead to apoptosis.
Assuntos
Retículo Endoplasmático/metabolismo , Hiperglicemia/metabolismo , Proteinúria/metabolismo , Albuminas/farmacologia , Linhagem Celular , Proteínas de Ligação a DNA/genética , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Chaperona BiP do Retículo Endoplasmático , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Glucose/farmacologia , Proteínas de Choque Térmico HSP70 , Proteínas de Choque Térmico/genética , Humanos , Hiperglicemia/genética , Túbulos Renais/citologia , Túbulos Renais/efeitos dos fármacos , Túbulos Renais/metabolismo , Chaperonas Moleculares/genética , Análise de Sequência com Séries de Oligonucleotídeos , Dobramento de Proteína , Proteínas/genética , Proteinúria/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Transcrição de Fator Regulador X , Tapsigargina/farmacologia , Fatores de Transcrição/genética , Tunicamicina/farmacologia , Proteína 1 de Ligação a X-BoxRESUMO
With the advent of transcriptome profiling techniques, an enormous amount of data has been generated in the field of molecular nephrology. We will review analysis tools and challenges for genomic approaches and present their application in gene-expression studies on kidney biopsies. The findings in this rapidly evolving field may ultimately complement histopathological analysis, the current diagnostic and prognostic gold standard. Altogether, genomics may bring nephrology one-step closer to a systematic understanding of biological processes involved in renal disease.
Assuntos
Genômica , Nefropatias/genética , Biópsia , Biologia Computacional , Expressão Gênica , Perfilação da Expressão Gênica , Humanos , Rim/anatomia & histologia , Rim/fisiologia , Nefropatias/patologia , Transplante de Rim/efeitos adversos , Nefrologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Falha de TratamentoRESUMO
OBJECTIVE: Glomerular mesangial expansion and podocyte loss are important early features of diabetic nephropathy, whereas tubulointerstitial injury and fibrosis are critical for progression of diabetic nephropathy to kidney failure. Therefore, we analyzed the expression of genes in glomeruli and tubulointerstitium in kidney biopsies from diabetic nephropathy patients to identify pathways that may be activated in humans but not in murine models of diabetic nephropathy that fail to progress to glomerulosclerosis, tubulointerstitial fibrosis, and kidney failure. RESEARCH DESIGN AND METHODS: Kidney biopsies were obtained from 74 patients (control subjects, early and progressive type 2 diabetic nephropathy). Glomerular and tubulointerstitial mRNAs were microarrayed, followed by bioinformatics analyses. Gene expression changes were confirmed by real-time RT-PCR and immunohistological staining. Samples from db/db C57BLKS and streptozotocin-induced DBA/2J mice, commonly studied murine models of diabetic nephropathy, were analyzed. RESULTS: In human glomeruli and tubulointerstitial samples, the Janus kinase (Jak)-signal transducer and activator of transcription (Stat) pathway was highly and significantly regulated. Jak-1, -2, and -3 as well as Stat-1 and -3 were expressed at higher levels in patients with diabetic nephropathy than in control subjects. The estimated glomerular filtration rate significantly correlated with tubulointerstitial Jak-1, -2, and -3 and Stat-1 expression (R(2) = 0.30-0.44). Immunohistochemistry found strong Jak-2 staining in glomerular and tubulointerstitial compartments in diabetic nephropathy compared with control subjects. In contrast, there was little or no increase in expression of Jak/Stat genes in the db/db C57BLKS or diabetic DBA/2J mice. CONCLUSIONS: These data suggest a direct relationship between tubulointerstitial Jak/Stat expression and progression of kidney failure in patients with type 2 diabetic nephropathy and distinguish progressive human diabetic nephropathy from nonprogressive murine diabetic nephropathy.