RESUMO
Acute kidney injury (AKI) is considered an inflammatory disease in which toll-like receptors (TLRs) signaling pathways play an important role. The activation of TLRs results in production of several inflammatory cytokines leading to further renal damage. In contrast, TLRs are key players on autophagy induction, which is associated with a protective function on cisplatin-induced AKI. Hence, the present study aimed to evaluate the specific participation of TLR2 and TLR4 molecules on the development of cisplatin-induced AKI. Complementarily, we also investigated the link between TLRs and heme oxygenase-1 (HO-1), a promisor cytoprotective molecule. First, we observed that only the absence of TLR2 but not TLR4 in mice exacerbated the renal dysfunction, tissue injury and mortality rate, even under an immunologically privileged microenvironment. Second, we demonstrated that TLR2 knockout (KO) mice presented lower expression of autophagy-associated markers when compared with TLR4 KO animals. Similar parameter was confirmed in vitro, using tubular epithelial cells derived from both KO mice. To test the cross-talking between HO-1 and TLRs, hemin (an HO-1 internal inducer) was administrated in cisplatin-treated TLR2 and TLR4 KO mice and it was detected an improvement in the global renal tissue parameters. However, this protection was less evident at TLR2 KO mice. In summary, we documented that TLR2 plays a protective role in cisplatin-induced AKI progression, in part, by a mechanism associated with autophagy up-regulation, considering that its interplay with HO-1 can promote renal tissue recover.
Assuntos
Injúria Renal Aguda/genética , Autofagia/genética , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Injúria Renal Aguda/metabolismo , Animais , Células Cultivadas , Cisplatino , Citocinas/metabolismo , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Rim/metabolismo , Rim/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismoRESUMO
Mesenchymal stromal cells (MSCs) orchestrate tissue repair by releasing cell-derived microvesicles (MVs), which, presumably by small RNA species, modulate global gene expression. The knowledge of miRNA/mRNA signatures linked to a reparative status may elucidate some of the molecular events associated with MSC protection. Here, we used a model of cisplatin-induced kidney injury (acute kidney injury) to assess how MSCs or MVs could restore tissue function. MSCs and MVs presented similar protective effects, which were evidenced in vivo and in vitro by modulating apoptosis, inflammation, oxidative stress, and a set of prosurvival molecules. In addition, we observed that miRNAs (i.e., miR-880, miR-141, miR-377, and miR-21) were modulated, thereby showing active participation on regenerative process. Subsequently, we identified that MSC regulates a particular miRNA subset which mRNA targets are associated with Wnt/TGF-ß, fibrosis, and epithelial-mesenchymal transition signaling pathways. Our results suggest that MSCs release MVs that transcriptionally reprogram injured cells, thereby modulating a specific miRNA-mRNA network.
RESUMO
AIM: Experimental autoimmune encephalomyelitis (EAE) is a CD4(+)-mediated autoimmune pathology of the central nervous system (CNS) that is used as a model for the study of the human neuroinflammatory disease, multiple sclerosis. During the development of EAE, auto-reactive Th1 and Th17 CD4(+) T cells infiltrate the CNS promoting inflammatory cells recruitment, focal inflammation and tissue destruction. In this sense, statins, agents used to lower lipid levels, have recently shown to exert interesting immunomodulatory function. In fact, statins promote a bias towards a Th2 response, which ameliorates the clinical outcome of EAE. Additionally, simvastatin can inhibit Th17 differentiation. However, many other effects exerted on the immune system by statins have yet to be clarified, in particular during neuroinflammation. Thus, the aim of this study was to investigate the effects of simvastatin on the development of experimental autoimmune encephalomyelitis. METHODS: Mice were immunized with MOG(35-55) and EAE severity was assessed daily and scored using a clinical scale. Cytokine secretion by mononuclear cells infiltrating the CNS was evaluated by flow cytometry. RESULTS: Simvastatin (5 mg/kg/day) improved clinical outcome, induced an increase in TGF-ß mRNA expression and inhibited IL-6, IL-12p40, IL-12p70, RANTES and MIP-1ß secretion (p < 0.05). This was accompanied by a significant decrease in CNS inflammatory mononuclear cell infiltration, with reduced frequencies of both Th1 and Th17 cells. Simvastatin inhibited the proliferation of T lymphocytes co-cultured with primary microglial cells. CONCLUSIONS: Simvastatin treatment promotes EAE clinical amelioration by inhibiting T cell proliferation and CNS infiltration by pathogenic Th1 and Th17 cells.
Assuntos
Encefalomielite Autoimune Experimental/tratamento farmacológico , Sinvastatina/farmacologia , Células Th1/efeitos dos fármacos , Células Th17/efeitos dos fármacos , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Proliferação de Células/efeitos dos fármacos , Sistema Nervoso Central/efeitos dos fármacos , Sistema Nervoso Central/imunologia , Quimiocina CCL5/imunologia , Encefalomielite Autoimune Experimental/imunologia , Feminino , Inflamação/tratamento farmacológico , Inflamação/imunologia , Subunidade p40 da Interleucina-12/imunologia , Interleucina-6/imunologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Esclerose Múltipla/tratamento farmacológico , Esclerose Múltipla/imunologia , Sinvastatina/imunologia , Células Th1/imunologia , Células Th17/imunologia , Fator de Crescimento Transformador beta/imunologiaRESUMO
Renovascular hypertension induced by 2 Kidney-1 Clip (2K-1C) is a renin-angiotensin-system (RAS)-dependent model, leading to renal vascular rarefaction and renal failure. RAS inhibitors are not able to reduce arterial pressure (AP) and/or preserve the renal function, and thus, alternative therapies are needed. Three weeks after left renal artery occlusion, fluorescently tagged mesenchymal stem cells (MSC) (2×10(5) cells/animal) were injected weekly into the tail vein in 2K-1C hypertensive rats. Flow cytometry showed labeled MSC in the cortex and medulla of the clipped kidney. MSC prevented a further increase in the AP, significantly reduced proteinuria and decreased sympathetic hyperactivity in 2K-1C rats. Renal function parameters were unchanged, except for an increase in urinary volume observed in 2K-1C rats, which was not corrected by MSC. The treatment improved the morphology and decreased the fibrotic areas in the clipped kidney and also significantly reduced renal vascular rarefaction typical of 2K-1C model. Expression levels of IL-1ß, TNF-α angiotensinogen, ACE, and Ang II receptor AT1 were elevated, whereas AT2 levels were decreased in the medulla of the clipped kidney. MSC normalized these expression levels. In conclusion, MSC therapy in the 2K-1C model (i) prevented the progressive increase of AP, (ii) improved renal morphology and microvascular rarefaction, (iii) reduced fibrosis, proteinuria and inflammatory cytokines, (iv) suppressed the intrarenal RAS, iv) decreased sympathetic hyperactivity in anesthetized animals and v) MSC were detected at the CNS suggesting that the cells crossed the blood-brain barrier. This therapy may be a promising strategy to treat renovascular hypertension and its renal consequences in the near future.
Assuntos
Hipertensão Renovascular/terapia , Transplante de Células-Tronco Mesenquimais , Proteinúria/terapia , Animais , Pressão Sanguínea , Barreira Hematoencefálica/citologia , Barreira Hematoencefálica/metabolismo , Corantes Fluorescentes , Expressão Gênica , Hipertensão Renovascular/genética , Hipertensão Renovascular/metabolismo , Hipertensão Renovascular/patologia , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Rim/metabolismo , Rim/patologia , Masculino , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , Proteinúria/genética , Proteinúria/metabolismo , Proteinúria/patologia , Ratos , Ratos Wistar , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/metabolismo , Receptor Tipo 2 de Angiotensina/genética , Receptor Tipo 2 de Angiotensina/metabolismo , Artéria Renal/cirurgia , Sistema Renina-Angiotensina/genética , Resultado do Tratamento , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND/AIMS: Renal ischaemia-reperfusion injury (IRI) is a systemic inflammatory process in which Th1 responses predominate affecting other organs including the lungs. The present study explored the phagocytic and microbicidal capacity of macrophages in rats with lung inflammation that underwent IRI. METHODS: The alveolar macrophages of rats sensitised to OVA were evaluated for phagocytosis and bacterial killing 24h after antigen challenge in animals with or without prior submission to 60 min of renal ischaemia. RESULTS: Bronchoalveolar lavage had a high level of cellular infiltrate in immunised animals (420%) compared with control animals; IRI significantly reduced this infiltration (52%). Macrophages from animals immunised and challenged with OVA presented a 10x increase in phagocytic capacity compared to the control group, whereas immunised animals subjected to IRI showed a reduction in the phagocytic index of 68%. The killing of Klebsiella pneumoniae by macrophages from immunised animals was higher (56%) compared with the control group but reduced in animals submitted to IRI (45%). Immunised and challenged group showed an increase in gene expression levels of IL-10(450%), HO-1 (259%), INF-γ (460%) and MCP-1 (370%) compared to the immunised group subjected to IRI. CONCLUSIONS: Renal ischaemia and reperfusion injury apparently alters the phagocytic and microbicidal capacity of macrophages, reducing lung inflammation to OVA.
Assuntos
Injúria Renal Aguda/imunologia , Macrófagos Alveolares/fisiologia , Fagocitose/fisiologia , Injúria Renal Aguda/metabolismo , Injúria Renal Aguda/patologia , Animais , Líquido da Lavagem Broncoalveolar/citologia , Células Cultivadas , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Modelos Animais de Doenças , Expressão Gênica , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-10/genética , Interleucina-10/metabolismo , Klebsiella pneumoniae/fisiologia , Macrófagos Alveolares/citologia , Macrófagos Alveolares/imunologia , Masculino , Óxido Nítrico/metabolismo , Ovalbumina/imunologia , Ratos , Ratos Wistar , Traumatismo por Reperfusão/imunologia , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologiaRESUMO
OBJECTIVE: the purpose of this study was to investigate the effect of low-level laser therapy (LLLT) on chronic kidney disease (CKD) in a model of unilateral ureteral obstruction (UUO). BACKGROUND DATA: Regardless of the etiology, CKD involves progressive widespread tissue fibrosis, tubular atrophy, and loss of kidney function. This process also occurs in kidney allograft. At present, effective therapies for this condition are lacking. We investigated the effects of LLLT on the interstitial fibrosis that occurs after experimental UUO in rats. METHODS: The occluded kidney of half of the 32 Wistar rats that underwent UUO received a single intraoperative dose of LLLT (AlGaAs laser, 780 nm, 22.5 J/cm(2), 30 mW, 0.75 W/cm(2), 30 sec on each of nine points). After 14 days, renal fibrosis was assessed by Sirius red staining under polarized light. Immunohistochemical analyses quantitated the renal tissue cells that expressed fibroblast (FSP-1) and myofibroblast (α-SMA) markers. Reverse transcriptase polymerase chain reaction (RT-PCR) was performed to determine the mRNA expression of interleukin (IL)-6, monocyte chemotactic protein-1 (MCP-1), transforming growth factor (TGF)-ß1 and Smad3. RESULTS: The UUO and LLLT animals had less fibrosis than the UUO animals, as well having decreased expression inflammatory and pro-fibrotic markers. CONCLUSIONS: For the first time, we showed that LLLT had a protective effect regarding renal interstitial fibrosis. It is conceivable that by attenuating inflammation, LLLT can prevent tubular activation and transdifferentiation, which are the two processes that mainly drive the renal fibrosis of the UUO model.
Assuntos
Terapia com Luz de Baixa Intensidade/métodos , Nefrite Intersticial/patologia , Nefrite Intersticial/radioterapia , Animais , Biópsia por Agulha , Modelos Animais de Doenças , Imuno-Histoquímica , Nefropatias/patologia , Nefropatias/radioterapia , Masculino , Ratos , Ratos Wistar , Sensibilidade e Especificidade , Resultado do TratamentoRESUMO
IL-4 produced by Th2 cells can block cytokine production by Th1 cells, and Th1 IFN-γ is known to counterregulate Th2 immune response, inhibiting allergic eosinophilia. As intrauterine undernutrition can attenuate lung inflammation, we investigated the influence of intrauterine undernourishment on the Th1/Th2 cytokine balance and allergic lung inflammation. Intrauterine undernourished offspring were obtained from dams fed 50% of the nourished diet of their counterparts and were immunized at 9 weeks of age. We evaluated the cell counts and cytokine protein expression in the bronchoalveolar lavage, mucus production and collagen deposition, and cytokine gene expression and transcription factors in lung tissue 21 days after ovalbumin immunization. Intrauterine undernourishment significantly reduced inflammatory cell airway infiltration, mucus secretion and collagen deposition, in rats immunized and challenged. Intrauterine undernourished rats also exhibited an altered cytokine expression profile, including higher TNF-α and IL-1ß expression and lower IL-6 expression than well-nourished rats following immunization and challenge. Furthermore, the intrauterine undernourished group showed reduced ratios of the IL-4/IFN-γ and the transcription factors GATA-3/T-Bet after immunization and challenge. We suggest that the attenuated allergic lung inflammation observed in intrauterine undernourished rats is related to an altered Th1/Th2 cytokine balance resulting from a reduced GATA-3/T-bet ratio.
Assuntos
Hipersensibilidade/metabolismo , Desnutrição/imunologia , Pneumonia/metabolismo , Células Th1/metabolismo , Células Th2/metabolismo , Animais , Líquido da Lavagem Broncoalveolar/citologia , Feminino , Fator de Transcrição GATA3/metabolismo , Hipersensibilidade/imunologia , Hipersensibilidade/patologia , Interferon gama/metabolismo , Interleucina-1beta/metabolismo , Interleucina-4/metabolismo , Interleucina-6/metabolismo , Masculino , Desnutrição/fisiopatologia , Ovalbumina/imunologia , Ovalbumina/toxicidade , Pneumonia/imunologia , Pneumonia/patologia , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Fenômenos Fisiológicos da Nutrição Pré-Natal/imunologia , Ratos , Ratos Wistar , Proteínas com Domínio T/metabolismo , Células Th1/imunologia , Equilíbrio Th1-Th2/efeitos dos fármacos , Células Th2/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The present study investigated the early presence of inflammatory response in renal tissue of young offspring from diabetic mothers. The effect of L-arginine (L-arg) supplementation was also investigated. The offspring was divided into four groups: group CO (controls); group DO (diabetic offspring); group CA (CO receiving 2% L-arg solution) and group DA (DO receiving the 2% L-arg solution). Glycemia, arterial pressure and renal function were evaluated; gene and protein expression of pro-inflammatory cytokines were also measured. Blood pressure levels were significantly increased in 2 and 6 month-old DO rats, whereas L-arg administration caused a significant decrease in the DA group, at both ages. DO rats showed a significantly blunted glycemic response to exogenous insulin. In 2 month-old DO animals, renal protein expression of pro-inflammatory molecules was significantly increased. At six months of age, we also observed an increase in gene expression of pro-inflammatory molecules, whereas L-arg supplementation prevented this increase at both ages. Our data suggest that activation of inflammatory pathways is present early in the kidney of DO rats, and that L-arg can attenuate the expression of these markers of tissue inflammation. Our results also reinforce the concept that intrauterine environmental factors are a fundamental determinant in the development of metabolic and vascular diseases later in life.
Assuntos
Injúria Renal Aguda/patologia , Diabetes Mellitus Experimental/patologia , Mediadores da Inflamação/administração & dosagem , Complicações na Gravidez/patologia , Efeitos Tardios da Exposição Pré-Natal/patologia , Injúria Renal Aguda/diagnóstico , Injúria Renal Aguda/etiologia , Animais , Arginina/administração & dosagem , Arginina/toxicidade , Biomarcadores/metabolismo , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/diagnóstico , Diagnóstico Precoce , Feminino , Mediadores da Inflamação/toxicidade , Masculino , Gravidez , Complicações na Gravidez/diagnóstico , Complicações na Gravidez/etiologia , Efeitos Tardios da Exposição Pré-Natal/diagnóstico , Efeitos Tardios da Exposição Pré-Natal/etiologia , Distribuição Aleatória , Ratos , Ratos WistarRESUMO
A growing body of evidence demonstrates a correlation between Th2 cytokines and the development of focal and segmental glomerulosclerosis (FSGS). Therefore, we hypothesized that GSL-1, a monoglycosylceramide from Sphingomonas ssp. with pro-Th1 activity on invariant Natural Killer T (iNKT) lymphocytes, could counterbalance the Th2 profile and modulate glomerulosclerosis. Using an adriamycin(ADM)-based model of FSGS, we found that BALB/c mice presented albuminuria and glomerular degeneration in association with a Th2-like pro-fibrogenic profile; these mice also expressed a combination of inflammatory cytokines, such as IL-4, IL-1α, IL-1ß, IL-17, TNF-α, and chemokines, such as RANTES and eotaxin. In addition, we observed a decrease in the mRNA levels of GD3 synthase, the enzyme responsible for GD3 metabolism, a glycolipid associated with podocyte physiology. GSL-1 treatment inhibited ADM-induced renal dysfunction and preserved kidney architecture, a phenomenon associated with the induction of a Th1-like response, increased levels of GD3 synthase transcripts and inhibition of pro-fibrotic transcripts and inflammatory cytokines. TGF-ß analysis revealed increased levels of circulating protein and tissue transcripts in both ADM- and GSL-1-treated mice, suggesting that TGF-ß could be associated with both FSGS pathology and iNKT-mediated immunosuppression; therefore, we analyzed the kidney expression of phosphorylated SMAD2/3 and SMAD7 proteins, molecules associated with the deleterious and protective effects of TGF-ß, respectively. We found high levels of phosphoSMAD2/3 in ADM mice in contrast to the GSL-1 treated group in which SMAD7 expression increased. These data suggest that GSL-1 treatment modulates the downstream signaling of TGF-ß through a renoprotective pathway. Finally, GSL-1 treatment at day 4, a period when proteinuria was already established, was still able to improve renal function, preserve renal structure and inhibit fibrogenic transcripts. In conclusion, our work demonstrates that the iNKT agonist GSL-1 modulates the pathogenesis of ADM-induced glomerulosclerosis and may provide an alternative approach to disease management.
Assuntos
Ceramidas/farmacologia , Regulação da Expressão Gênica/fisiologia , Glomerulosclerose Segmentar e Focal/imunologia , Células T Matadoras Naturais/imunologia , Transdução de Sinais/fisiologia , Sphingomonas/química , Análise de Variância , Animais , Western Blotting , Quimiocinas/imunologia , Citocinas/imunologia , Primers do DNA/genética , Doxorrubicina/toxicidade , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Regulação da Expressão Gênica/efeitos dos fármacos , Glomerulosclerose Segmentar e Focal/induzido quimicamente , Glomerulosclerose Segmentar e Focal/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Células T Matadoras Naturais/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Sialiltransferases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Células Th2/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
Adipose tissue-derived stem cells (ASCs) are an attractive source of stem cells with regenerative properties that are similar to those of bone marrow stem cells. Here, we analyze the role of ASCs in reducing the progression of kidney fibrosis. Progressive renal fibrosis was achieved by unilateral clamping of the renal pedicle in mice for 1 h; after that, the kidney was reperfused immediately. Four hours after the surgery, 2 × 10(5) ASCs were intraperitoneally administered, and mice were followed for 24 h posttreatment and then at some other time interval for the next 6 weeks. Also, animals were treated with 2 × 10(5) ASCs at 6 weeks after reperfusion and sacrificed 4 weeks later to study their effect when interstitial fibrosis is already present. At 24 h after reperfusion, ASC-treated animals showed reduced renal dysfunction and enhanced regenerative tubular processes. Renal mRNA expression of IL-6 and TNF was decreased in ASC-treated animals, whereas IL-4, IL-10, and HO-1 expression increased despite a lack of ASCs in the kidneys as determined by SRY analysis. As expected, untreated kidneys shrank at 6 weeks, whereas the kidneys of ASC-treated animals remained normal in size, showed less collagen deposition, and decreased staining for FSP-1, type I collagen, and Hypoxyprobe. The renal protection seen in ASC-treated animals was followed by reduced serum levels of TNF-α, KC, RANTES, and IL-1α. Surprisingly, treatment with ASCs at 6 weeks, when animals already showed installed fibrosis, demonstrated amelioration of functional parameters, with less tissue fibrosis observed and reduced mRNA expression of type I collagen and vimentin. ASC therapy can improve functional parameters and reduce progression of renal fibrosis at early and later times after injury, mostly due to early modulation of the inflammatory response and to less hypoxia, thereby reducing the epithelial-mesenchymal transition.
Assuntos
Tecido Adiposo/citologia , Nefropatias/patologia , Transplante de Células-Tronco , Células-Tronco/citologia , Animais , Quimiocina CCL5/sangue , Quimiocinas/sangue , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Feminino , Heme Oxigenase-1/metabolismo , Interleucina-10/metabolismo , Interleucina-1alfa/sangue , Interleucina-4/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Isquemia/complicações , Isquemia/patologia , Isquemia/terapia , Nefropatias/complicações , Nefropatias/terapia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/metabolismo , Fator de Necrose Tumoral alfa/sangue , Vimentina/genética , Vimentina/metabolismoRESUMO
The pathophysiology of sepsis involves complex cytokine and inflammatory mediator networks, a mechanism to which NF-κB activation is central. Downregulation of endothelial nitric oxide synthase (eNOS) contributes to sepsis-induced endothelial dysfunction. Erythropoietin (EPO) has emerged as a major tissue-protective cytokine in the setting of stress. We investigated the role of EPO in sepsis-related acute kidney injury using a cecal ligation and puncture (CLP) model. Wistar rats were divided into three primary groups: control (sham-operated); CLP; and CLP+EPO. EPO (4,000 IU/kg body wt ip) was administered 24 and 1 h before CLP. Another group of rats received N-nitro-l-arginine methyl ester (l-NAME) simultaneously with EPO administration (CLP+EPO+l-NAME). A fifth group (CLP+EPOtreat) received EPO at 1 and 4 h after CLP. At 48 h postprocedure, CLP+EPO rats presented significantly higher inulin clearance than did CLP and CLP+EPO+l-NAME rats; hematocrit levels, mean arterial pressure, and metabolic balance remained unchanged in the CLP+EPO rats; and inulin clearance was significantly higher in CLP+EPOtreat rats than in CLP rats. At 48 h after CLP, creatinine clearance was significantly higher in the CLP+EPO rats than in the CLP rats. In renal tissue, pre-CLP EPO administration prevented the sepsis-induced increase in macrophage infiltration, as well as preserving eNOS expression, EPO receptor (EpoR) expression, IKK-α activation, NF-κB activation, and inflammatory cytokine levels, thereby increasing survival. We conclude that this protection, which appears to be dependent on EpoR activation and on eNOS expression, is attributable, in part, to inhibition of the inflammatory response via NF-κB downregulation.
Assuntos
Injúria Renal Aguda/prevenção & controle , Eritropoetina/antagonistas & inibidores , NF-kappa B/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Sepse/tratamento farmacológico , Injúria Renal Aguda/tratamento farmacológico , Injúria Renal Aguda/metabolismo , Animais , Creatinina/urina , Citocinas/análise , Regulação para Baixo , Quimioterapia Combinada , Inibidores Enzimáticos/farmacologia , Quinase I-kappa B/metabolismo , Inflamação/metabolismo , Inulina/urina , Masculino , NG-Nitroarginina Metil Éster/farmacologia , Ratos , Ratos Wistar , Receptores da Eritropoetina/biossíntese , Sepse/metabolismo , Regulação para CimaRESUMO
The pathogenesis of focal segmental glomerulosclerosis (FSGS) appears to be associated with type-2 cytokines and podocyte dysfunction. In this study, we tested the hypothesis that immunization with the polysaccharide fraction of Propionibacterium acnes (PS), a pro-Th1 agonist, may subvert the type-2 profile and protect podocytes from adriamycin-induced glomerulosclerosis. Adriamycin injection resulted in albuminuria and increased serum creatinine in association with loss of glomerular podocin and podoplanin expression, which is consistent with podocyte dysfunction. Renal tissue analysis revealed the expression of transcripts for GATA3 and fibrogenic-related proteins, such as TGF-ß, tissue inhibitor of metalloproteinase-1 (TIMP-1) and metalloproteinase 9 (MMP9). In association with the expression of fibrogenic transcripts, we observed peri-glomerular expression of α-smooth muscle actin (α-SMA), indicating epithelial-to-mesenchymal transition, and increased expression of proliferating cell nuclear antigen (PCNA) in tubular cells, suggesting intense proliferative activity. Previous immunization with PS inhibited albuminuria and serum creatinine in association with the preservation of podocyte proteins and inhibition of fibrogenic transcripts and the expression of α-SMA and PCNA proteins. Tissue analysis also revealed that PS treatment induced expression of mRNA for GD3 synthase, which is a glycosiltransferase related to the synthesis of GD3, a ganglioside associated with podocyte physiology. In addition, PS treatment inhibited the influx of inflammatory CD8(pos) and CD11b(pos) cells to kidney tissue. Finally, PS treatment on day 4 post-ADM, a period when proteinuria was already established, was able to improve renal function. Thus, we demonstrate that the PS fraction of P. acnes can inhibit FSGS pathogenesis, suggesting that immunomodulation can represent an alternative approach for disease management.
Assuntos
Glomerulosclerose Segmentar e Focal/etiologia , Polissacarídeos Bacterianos/farmacologia , Propionibacterium acnes/química , Substâncias Protetoras/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Doxorrubicina/efeitos adversos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Fibrose/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Glomerulosclerose Segmentar e Focal/tratamento farmacológico , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Polissacarídeos Bacterianos/administração & dosagem , Polissacarídeos Bacterianos/isolamento & purificação , Substâncias Protetoras/administração & dosagem , Substâncias Protetoras/isolamento & purificação , Proteinúria/tratamento farmacológico , Insuficiência Renal/induzido quimicamente , Insuficiência Renal/tratamento farmacológico , Linfócitos T/imunologia , Linfócitos T/metabolismo , Células Th2/imunologia , Células Th2/metabolismo , Transcrição GênicaRESUMO
Tubule-interstitial nephritis (TIN) results in decreased renal function and interstitial inflammation, which ultimately leads to fibrosis. Excessive adenine intake can cause TIN because xanthine dehydrogenase (XDH) can convert this purine into an insoluble compound, which precipitates in the tubuli. Innate immune sensors, such as Toll-like receptors (TLR) and inflammasome complex, play a crucial role in the initiation of inflammation. The aim of this study was to evaluate the roles of TLR-2 and -4, Myd88 and inflammasome complex in an experimental model of TIN. Here, we show that wild-type (WT) mice fed adenine-enriched food exhibited significant renal dysfunction and enhanced cellular infiltration accompanied by collagen deposition. They also presented higher gene and protein expression of pro-inflammatory cytokines. In contrast, TLR-2, -4, MyD88, ASC and Caspase-1 KO mice showed renoprotection associated with expression of inflammatory molecules at levels comparable to controls. Furthermore, treatment of WT animals with allopurinol, an XDH inhibitor, led to reduced levels of uric acid, oxidative stress, collagen deposition and a downregulation of the NF-kB signaling pathway. We concluded that MyD88 signaling and inflammasome participate in the development of TIN. Furthermore, inhibition of XDH seems to be a promising way to therapeutically target the developing inflammatory process.
Assuntos
Inflamassomos/metabolismo , Túbulos Renais/patologia , Fator 88 de Diferenciação Mieloide/metabolismo , Nefrite Intersticial/metabolismo , Nefrite Intersticial/patologia , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Adenina/administração & dosagem , Adenina/farmacologia , Alopurinol/farmacologia , Alopurinol/uso terapêutico , Animais , Dieta , Progressão da Doença , Inflamassomos/efeitos dos fármacos , Inflamação/patologia , Túbulos Renais/metabolismo , Camundongos , Camundongos Knockout , Nefrite Intersticial/tratamento farmacológico , Nefrite Intersticial/prevenção & controle , Transdução de Sinais/efeitos dos fármacos , Xantina Desidrogenase/antagonistas & inibidores , Xantina Desidrogenase/metabolismoRESUMO
Therapy with mesenchymal stem cells (MSCs) has showed to be promising due to its immunomodulatory function. Traumatic brain injury (TBI) triggers immune response and release of inflammatory mediators, mainly cytokines, by glial cells creating a hostile microenvironment for endogenous neural stem cells (NSCs). We investigated the effects of factors secreted by MSCs on NSC in vitro and analyzed cytokines expression in vitro in a TBI model. Our in vitro results show that MSC-secreted factors increase NSC proliferation and induce higher expression of GFAP, indicating a tendency toward differentiation into astrocytes. In vivo experiments showed that MSC injection at an acute model of brain injury diminishes a broad profile of cytokines in the tissue, suggesting that MSC-secreted factors may modulate the inflammation at the injury site, which may be of interest to the development of a favorable microenvironment for endogenous NSC and consequently to repair the injured tissue.
RESUMO
Focal and segmental glomerulosclerosis (FSGS) is one of the most important causes of end-stage renal failure. The bradykinin B1 receptor has been associated with tissue inflammation and renal fibrosis. To test for a role of the bradykinin B1 receptor in podocyte injury, we pharmacologically modulated its activity at different time points in an adriamycin-induced mouse model of FSGS. Estimated albuminuria and urinary protein to creatinine ratios correlated with podocytopathy. Adriamycin injection led to loss of body weight, proteinuria, and upregulation of B1 receptor mRNA. Early treatment with a B1 antagonist reduced albuminuria and glomerulosclerosis, and inhibited the adriamycin-induced downregulation of podocin, nephrin, and α-actinin-4 expression. Moreover, delayed treatment with antagonist also induced podocyte protection. Conversely, a B1 agonist aggravated renal dysfunction and even further suppressed the levels of podocyte-related molecules. Thus, we propose that kinin has a crucial role in the pathogenesis of FSGS operating through bradykinin B1 receptor signaling.
Assuntos
Bradicinina/análogos & derivados , Glomerulosclerose Segmentar e Focal/induzido quimicamente , Podócitos/efeitos dos fármacos , Receptor B1 da Bradicinina/agonistas , Transdução de Sinais/efeitos dos fármacos , Actinina/metabolismo , Albuminúria/induzido quimicamente , Albuminúria/metabolismo , Albuminúria/prevenção & controle , Animais , Bradicinina/farmacologia , Bradicinina/toxicidade , Antagonistas de Receptor B1 da Bradicinina , Modelos Animais de Doenças , Doxorrubicina , Regulação da Expressão Gênica/efeitos dos fármacos , Glomerulosclerose Segmentar e Focal/genética , Glomerulosclerose Segmentar e Focal/metabolismo , Glomerulosclerose Segmentar e Focal/patologia , Glomerulosclerose Segmentar e Focal/prevenção & controle , Heme Oxigenase-1/metabolismo , Interleucina-1beta/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Podócitos/metabolismo , Podócitos/patologia , RNA Mensageiro/metabolismo , Receptor B1 da Bradicinina/genética , Receptor B1 da Bradicinina/metabolismo , Fatores de Tempo , Inibidor Tecidual de Metaloproteinase-1/metabolismoRESUMO
Objective: To analyze the role of adipose tissue-derived stem cells in reducing the progression of renal fibrosis. Methods: adipose tissue-derived stem cells were isolated from C57Bl/6 mice and characterized by cytometry and differentiation. Renal fibrosis was established after unilateral clamping of the renal pedicle for 1 hour. Four hours after reperfusion, 2.105 adipose tissue-derived stem cells were administered intraperitoneally and the animals were followed for 24 hours during 6 weeks. In another experimental group, 2.105 adipose tissue-derived stem cells were administered only after 6 weeks of reperfusion, and they were euthanized and studied 4 weeks later. Twenty-four hours after reperfusion, the animals treated with adipose tissue-derived stem cells displayed reduced renal and tubular dysfunction and an increase of the regenerative process. Renal expression of IL-6 and TNF mRNA were decreased in the animals treated with adipose tissue-derived stem cells, while the levels of IL-4, IL-10, and HO-1 were increased, despite the fact that adipose tissue-derived stem cells were not observed in the kidneys via SRY analysis. Results: In 6 weeks, the kidneys of non-treated animals decreased in size, and the kidneys of the animals treated with adipose tissue-derived stem cells remained at normal size and display less deposition of type 1 collagen and FSP-1. The renal protection observed in animals treated with adipose tissue-derived stem cells was followed by a drop in serum levels of TNF-alpha, KC, RANTES, and IL-1a. Treatment with adipose tissue-derived stem cells after 6 weeks, when the animals already displayed established fibrosis, demonstrated an improvement in functional parameters and less fibrosis analyzed by Picrosirius stain, as well as a reduction of the expression of type 1 collagen and vimentin mRNA.
Objetivo: Analisar o papel das células-tronco derivadas do tecido adiposo na redução da progressão da fibrose renal. Métodos: células-tronco derivadas do tecido adiposo foram isoladas de camundongos C57Bl/6 e caracterizadas por citometria e diferenciação. Fibrose renal foi instaurada após clampeamento unilateral do pedículo renal por 1 hora. Após 4 horas de reperfusão, 2.105 células-tronco derivadas do tecido adiposo foram administradas por via intraperitoneal, e os animais foram acompanhados por 24 horas e 6 semanas. Em outro grupo de experimentos, 2.105 células-tronco derivadas do tecido adiposo foram administradas somente após 6 semanas de reperfusão, e os animais foram sacrificados e estudados 4 semanas mais tarde. Após 24 horas da reperfusão, animais tratados com células-tronco derivadas do tecido adiposo apresentaram reduzida disfunção renal e tubular, além de aumento do processo regenerativo. Expressão renal de RNAm de IL-6 e TNF foi diminuída nos animais tratados com células-tronco derivadas do tecido adiposo, enquanto IL-4, IL-10 e HO-1 foram aumentadas, apesar de células-tronco derivadas do tecido adiposo não serem observadas nos rins por meio da análise SRY. Resultados: Em 6 semanas, os rins dos animais não tratados diminuíram; no entanto, os rins dos animais tratados com células-tronco derivadas do tecido adiposo permaneceram com o tamanho normal e apresentaram menor deposição de colágeno tipo 1 e FSP-1. Proteção renal observada em animais tratados com células-tronco derivadas do tecido adiposo foi seguida por redução nos níveis séricos de TNF-alfa, KC, RANTES e IL-1a. O tratamento com células-tronco derivadas do tecido adiposo após 6 semanas, quando os animais já apresentavam fibrose instalada, demonstrou melhora em parâmetros funcionais e menos fibrose, analisada pela coloração de Picrosirius, e redução da expressão de RNAm de colágeno tipo I e vimentina.
Assuntos
Animais , Camundongos , Fibrose , Inflamação , Células-Tronco Mesenquimais , Insuficiência Renal , Traumatismo por ReperfusãoRESUMO
OBJECTIVE: To analyze the role of adipose tissue-derived stem cells in reducing the progression of renal fibrosis. METHODS: adipose tissue-derived stem cells were isolated from C57Bl/6 mice and characterized by cytometry and differentiation. Renal fibrosis was established after unilateral clamping of the renal pedicle for 1 hour. Four hours after reperfusion, 2.105 adipose tissue-derived stem cells were administered intraperitoneally and the animals were followed for 24 hours during 6 weeks. In another experimental group, 2.105adipose tissue-derived stem cells were administered only after 6 weeks of reperfusion, and they were euthanized and studied 4 weeks later. Twenty-four hours after reperfusion, the animals treated with adipose tissue-derived stem cells displayed reduced renal and tubular dysfunction and an increase of the regenerative process. Renal expression of IL-6 and TNF mRNA were decreased in the animals treated with adipose tissue-derived stem cells, while the levels of IL-4, IL-10, and HO-1 were increased, despite the fact that adipose tissue-derived stem cells were not observed in the kidneys via SRY analysis. RESULTS: In 6 weeks, the kidneys of non-treated animals decreased in size, and the kidneys of the animals treated with adipose tissue-derived stem cells remained at normal size and display less deposition of type 1 collagen and FSP-1. The renal protection observed in animals treated with adipose tissue-derived stem cells was followed by a drop in serum levels of TNF-α, KC, RANTES, and IL-1a. Treatment with adipose tissue-derived stem cells after 6 weeks, when the animals already displayed established fibrosis, demonstrated an improvement in functional parameters and less fibrosis analyzed by Picrosirius stain, as well as a reduction of the expression of type 1 collagen and vimentin mRNA. CONCLUSION: Treatment with adipose tissue-derived stem cells may deter the progression of renal fibrosis by modulation of the early inflammatory response, likely via reduction of the epithelial-mesenchymal transition.
RESUMO
BACKGROUND: The tubule-interstitial fibrosis is the hallmark of progressive renal disease and is strongly associated with inflammation of this compartment. Heme-oxygenase-1 (HO-1) is a cytoprotective molecule that has been shown to be beneficial in various models of renal injury. However, the role of HO-1 in reversing an established renal scar has not yet been addressed. AIM: We explored the ability of HO-1 to halt and reverse the establishment of fibrosis in an experimental model of chronic renal disease. METHODS: Sprague-Dawley male rats were subjected to unilateral ureteral obstruction (UUO) and divided into two groups: non-treated and Hemin-treated. To study the prevention of fibrosis, animals were pre-treated with Hemin at days -2 and -1 prior to UUO. To investigate whether HO-1 could reverse established fibrosis, Hemin therapy was given at days 6 and 7 post-surgery. After 7 and/or 14 days, animals were sacrificed and blood, urine and kidney tissue samples were collected for analyses. Renal function was determined by assessing the serum creatinine, inulin clearance, proteinuria/creatininuria ratio and extent of albuminuria. Arterial blood pressure was measured and fibrosis was quantified by Picrosirius staining. Gene and protein expression of pro-inflammatory and pro-fibrotic molecules, as well as HO-1 were performed. RESULTS: Pre-treatment with Hemin upregulated HO-1 expression and significantly reduced proteinuria, albuminuria, inflammation and pro-fibrotic protein and gene expressions in animals subjected to UUO. Interestingly, the delayed treatment with Hemin was also able to reduce renal dysfunction and to decrease the expression of pro-inflammatory molecules, all in association with significantly reduced levels of fibrosis-related molecules and collagen deposition. Finally, TGF-ß protein production was significantly lower in Hemin-treated animals. CONCLUSION: Treatment with Hemin was able both to prevent the progression of fibrosis and to reverse an established renal scar. Modulation of inflammation appears to be the major mechanism behind HO-1 cytoprotection.
Assuntos
Fibrose/metabolismo , Heme Oxigenase-1/biossíntese , Hemina/farmacologia , Túbulos Renais/metabolismo , Animais , Modelos Animais de Doenças , Progressão da Doença , Ensaio de Imunoadsorção Enzimática/métodos , Perfilação da Expressão Gênica , Imuno-Histoquímica/métodos , Inflamação , Nefropatias/patologia , Masculino , Modelos Biológicos , Ratos , Ratos Sprague-Dawley , Fator de Crescimento Transformador beta/metabolismoRESUMO
One of the early phases that lead to fibrosis progression is inflammation. Once this stage is resolved, fibrosis might be prevented. Bone marrow mononuclear cells (BMMCs) are emerging as a new therapy for several pathologies, including autoimmune diseases, because they enact immunosuppression. In this study we aimed to evaluate the role of BMMC administration in a model of kidney fibrosis induced by an acute injury. C57Bl6 mice were subjected to unilateral severe ischemia by clamping the left renal pedicle for 1h. BMMCs were isolated from femurs and tibia, and after 6h of reperfusion, 1 x 10(6) cells were administrated intraperitoneally. At 24h after surgery, treated animals showed a significant decrease in creatinine and urea levels when compared with untreated animals. Different administration routes were tested. Moreover, interferon (IFN) receptor knockout BMMCs were used, as this receptor is necessary for BMMC activation. Labeled BMMCs were found in ischemic kidney on FACS analysis. This improved outcome was associated with modulation of inflammation in the kidney and systemic modulation, as determined by cytokine expression profiling. Despite non-amelioration of functional parameters, kidney mRNA expression of interleukin (IL)-6 at 6 weeks was lower in BMMC-treated animals, as were levels of collagen 1, connective tissue growth factor (CTGF), transforming growth factor-beta (TGF-beta) and vimentin. Protective molecules, such as IL-10, heme oxygenase 1 (HO-1) and bone morphogenetic 7 (BMP-7), were increased in treated animals after 6 weeks. Moreover, Masson and Picrosirius red staining analyses showed less fibrotic areas in the kidneys of treated animals. Thus, early modulation of inflammation by BMMCs after an ischemic injury leads to reduced fibrosis through modulation of early inflammation.
Assuntos
Células da Medula Óssea/citologia , Nefropatias/cirurgia , Rim/patologia , Leucócitos Mononucleares/citologia , Doença Aguda , Animais , Antígenos CD34/análise , Células da Medula Óssea/metabolismo , Proteína Morfogenética Óssea 7/genética , Proteína Morfogenética Óssea 7/metabolismo , Transplante de Células/métodos , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Feminino , Fibrose/cirurgia , Expressão Gênica , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Imuno-Histoquímica , Imunofenotipagem , Isquemia/complicações , Rim/irrigação sanguínea , Nefropatias/etiologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/transplante , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-kit/análise , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Lipopolysaccharides from gram-negative bacteria are amongst the most common causative agents of acute lung injury, which is characterized by an inflammatory response, with cellular infiltration and the release of mediators/cytokines. There is evidence that bradykinin plays a role in lung inflammation in asthma but in other types of lung inflammation its role is less clear. In the present study we evaluated the role of the bradykinin B1 receptor in acute lung injury caused by lipopolysaccharide inhalation and the mechanisms behind bradykinin actions participating in the inflammatory response. We found that in C57Bl/6 mice, the bradykinin B1 receptor expression was up-regulated 24h after lipopolysaccharide inhalation. At this time, the number of cells and protein concentration were significantly increased in the bronchoalveolar lavage fluid and the mice developed airway hyperreactivity to methacholine. In addition, there was an increased expression of tumor necrosis factor-alpha, interleukin-1 beta and interferon-gamma and chemokines (monocytes chemotactic protein-1 and KC) in the bronchoalveolar lavage fluid and in the lung tissue. We then treated the mice with a bradykinin B1 receptor antagonist, R-954 (Ac-Orn-[Oic2, alpha-MePhe5, D-betaNal7, Ile8]desArg9-bradykinin), 30 min after lipopolysaccharide administration. We observed that this treatment prevented the airway hyperreactivity as well as the increased cellular infiltration and protein content in the bronchoalveolar lavage fluid. Moreover, R-954 inhibited the expression of cytokines/chemokines. These results implicate bradykinin, acting through B1 receptor, in the development of acute lung injury caused by lipopolysaccharide inhalation.