RESUMO
In this study we evaluated whether administration of stem cells of neural origin (neural precursor cells, NPCs) could be protective against renal ischemia-reperfusion injury (IRI). We hypothesized that stem cell outcomes are not tissue-specific and that NPCs can improve tissue damage through paracrine mechanisms, especially due to immunomodulation. To this end, Wistar rats (200-250 g) were submitted to 1-hour ischemia and treated with NPCs (4 x 10(6) cells/animal) at 4 h of reperfusion. To serve as controls, ischemic animals were treated with cerebellum homogenate harvested from adult rat brain. All groups were sacrificed at 24 h of reperfusion. NPCs were isolated from rat fetus telencephalon and cultured until neurosphere formation (7 days). Before administration, NPCs were labeled with carboxyfluorescein diacetate succinimydylester (CFSE). Kidneys were harvested for analysis of cytokine profile and macrophage infiltration. At 24 h, NPC treatment resulted in a significant reduction in serum creatinine (IRI + NPC 1.21 + 0.18 vs. IRI 3.33 + 0.14 and IRI + cerebellum 2.95 + 0.78 mg/dl, p < 0.05) and acute tubular necrosis (IRI + NPC 46.0 + 2.4% vs. IRI 79.7 + 14.2%, p < 0.05). NPC-CFSE and glial fibrillary acidic protein (GFAP)-positive cells (astrocyte marker) were found exclusively in renal parenchyma, which also presented GFAP and SOX-2 (an embryonic neural stem cell marker) mRNA expression. NPC treatment resulted in lower renal proinflammatory IL1-beta and TNF-alpha expression and higher anti-inflammatory IL-4 and IL-10 transcription. NPC-treated animals also had less macrophage infiltration and decreased serum proinflammatory cytokines (IL-1beta, TNF-alpha and INF-gamma). Our data suggested that NPC therapy improved renal function by influencing immunological responses.
Assuntos
Rim/irrigação sanguínea , Neurônios , Traumatismo por Reperfusão/terapia , Transplante de Células-Tronco , Animais , Masculino , Ratos , Ratos WistarRESUMO
The Kallikrein-kinin system works through activation of two receptors. One constitutive, named B2 receptor (B2R) and another inducible, denominated B1 receptor (B1R). In renal fibrosis, B2R receptor activation appears to be protective, however B1R participation is unveiled. The aim of this study was to analyze how the deletion of the B1R would modify tissue responses after unilateral ureteral obstruction (UUO). For that, B1R knockout (B1KO) and wild-type mice (B1B2WT) were subjected to UUO and sacrificed at days 1, 5 and 14. Renal dysfunction was assayed by urine proteinuria/creatinine ratio and percentage of tubulointerstitial fibrosis. Kidneys were harvested at day 5 to analyze anti and pro-inflammatory molecules expression by real-time PCR. We demonstrated that at all time points, B1KO mice presented lower proteinuria/creatinine ratio from bladder urine. B1KO protection was reinforced by its lower tubular interstitial fibrosis percentage at day 14 (B1B2WT: 12.16+/-1.53% vs. B1KO: 6.73+/-1.07%, p<0.02). UUO was able to induce B1R expression and its highest transcription was achieved at day 5. At this day, B1KO had significant lower expression of pro-inflammatory molecules such as TGF-beta, MCP-1, OPN and IL-6 and higher anti-inflammatory components, as IL-10 and HO-1. Herein, we observed that B1R deletion may be an important component in renal fibrosis prevention.
Assuntos
Nefropatias/genética , Nefropatias/patologia , Rim/patologia , Receptor B1 da Bradicinina/genética , Animais , Quimiocina CCL2/imunologia , Quimiocina CCL2/metabolismo , Creatinina/sangue , Creatinina/urina , Fibrose , Deleção de Genes , Heme Oxigenase-1/imunologia , Heme Oxigenase-1/metabolismo , Interleucina-10/imunologia , Interleucina-10/metabolismo , Interleucina-6/imunologia , Interleucina-6/metabolismo , Sistema Calicreína-Cinina/genética , Nefropatias/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteinúria/urina , Fator de Crescimento Transformador beta/imunologia , Fator de Crescimento Transformador beta/metabolismo , Obstrução Ureteral/complicaçõesRESUMO
Ischemic-reperfusion injury (IRI) triggers an inflammatory response involving neutrophils/macrophages, lymphocytes and endothelial cells. Galectin-3 is a multi-functional lectin with a broad range of action such as promotion of neutrophil adhesion, induction of oxidative stress, mastocyte migration and degranulation, and production of pro-inflammatory cytokines. The aim of this study was evaluate the role of galectin-3 in the inflammation triggered by IRI. Galectin-3 knockout (KO) and wild type (wt) mice were subjected to 45 min of renal pedicle occlusion. Blood and kidney samples were collected at 6, 24, 48 and 120 h. Blood urea was analyzed enzymatically, while MCP-1, IL-6 and IL-1beta were studied by real-time PCR. Reactive oxygen species (ROS) was investigated by flow cytometry. Morphometric analyses were performed at 6, 24, 48 and 120 h after reperfusion. Urea peaked at 24 h, being significantly lower in knockout animals (wt = 264.4 +/- 85.21 mg/dl vs. gal-3 KO = 123.74 +/- 29.64 mg/dl, P = 0.001). Galectin-3 knockout animals presented less acute tubular necrosis and a more prominent tubular regeneration when compared with controls concurrently with lower expression of MCP-1, IL-6, IL-1beta, less macrophage infiltration and lower ROS production at early time points. Galectin-3 seems to play a role in renal IRI involving the secretion of macrophage-related chemokine, pro-inflammatory cytokines and ROS production.
Assuntos
DNA/genética , Galectina 3/genética , Expressão Gênica , Transplante de Rim/patologia , Rim/irrigação sanguínea , Traumatismo por Reperfusão/metabolismo , Animais , Autoantígenos , Biomarcadores , Quimiocina CCL2/biossíntese , Quimiocina CCL2/genética , Modelos Animais de Doenças , Citometria de Fluxo , Seguimentos , Galectina 3/biossíntese , Imuno-Histoquímica , Interleucina-1beta/biossíntese , Interleucina-1beta/genética , Interleucina-6/biossíntese , Interleucina-6/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Reação em Cadeia da Polimerase , Espécies Reativas de Oxigênio/metabolismo , Traumatismo por Reperfusão/patologiaRESUMO
Bradykinin B1 receptors are exclusively expressed in inflamed tissues. For this reason, they have been related with the outcomes of several pathologies. Ischemia-reperfusion injury is caused by the activation of inflammatory and cytoprotective genes, such as macrophage chemoattractant protein-1 and heme oxygenase-1, respectively. This study was aimed to analyze the involvement of bradykinin B1 and B2 receptors (B1R and B2R) in tissue response after renal ischemia-reperfusion injury. For that, B1R (B1-/-), B2R (B2-/-) knockout animals and its control (wild-type mice, B1B2+/+) were subjected to renal bilateral ischemia, followed by 24, 48 and 120 h of reperfusion. At these time points, blood serum samples were collected for creatinine and urea dosages. Kidneys were harvested for histology and molecular analyses by real-time PCR. At 24 and 48 h of reperfusion, B1-/- group resulted in the lowest serum creatinine and urea levels, indicating less renal damage, which was proved by renal histology. Renal protection associated with B1-/- mice was also related with higher expression of HO-1 and lower expression of MCP-1. In conclusion, the absence of B1R had a protective role against inflammatory responses developed after renal ischemia-reperfusion injury.