RESUMO
The Renin-Angiotensin System (RAS) possesses a counter-regulatory axis composed of angiotensin converting enzyme (ACE)2, angiotensin-(1-7) [Ang-(1-7)] and the Mas receptor, which opposes many AT1-receptor-mediated effects of ligand angiotensin II. Ang-(1-7), as a ligand of the Mas receptor, has inhibitory effects on renal inflammation and fibrosis in experimental diabetes. However, Ang-(1-7) has a short half-life in plasma, which may render it unsuitable for use in clinics. Here, we investigated the effects of the lanthionine-stabilized Ang-(1-7), cyclic (c)Ang-(1-7), a lanthipeptide that is more peptidase-resistant than the linear peptide, in BTBR ob/ob mice with type 2 diabetic nephropathy. BTBR ob/ob mice received vehicle, cAng-(1-7), or the ACE inhibitor lisinopril. The treatment started at ten weeks of age, when the animals had already developed albuminuria, and ended at 19-20 weeks of age. cAng-(1-7) limited albuminuria progression, and limited podocyte dysfunction similarly to lisinopril. cAng-(1-7), unlike lisinopril, reduced glomerular fibrosis and inflammation, and counteracted glomerular capillary rarefaction. Furthermore, when cAng-(1-7) was combined with lisinopril, a superior antiproteinuric effect than with lisinopril alone was found, in association with better preservation of podocyte proteins and amelioration of capillary density. Thus, adding cAng-(1-7) to ACE-inhibitor therapy could benefit those diabetic patients who do not respond completely to ACE-inhibitor therapy.
Assuntos
Angiotensina I/administração & dosagem , Inibidores da Enzima Conversora de Angiotensina/administração & dosagem , Diabetes Mellitus Tipo 2/complicações , Nefropatias Diabéticas/tratamento farmacológico , Fragmentos de Peptídeos/administração & dosagem , Proteinúria/tratamento farmacológico , Alanina/administração & dosagem , Alanina/análogos & derivados , Alanina/química , Alanina/farmacocinética , Angiotensina I/química , Angiotensina I/farmacocinética , Animais , Diabetes Mellitus Tipo 2/genética , Nefropatias Diabéticas/diagnóstico , Nefropatias Diabéticas/etiologia , Nefropatias Diabéticas/patologia , Modelos Animais de Doenças , Quimioterapia Combinada/métodos , Meia-Vida , Humanos , Glomérulos Renais/irrigação sanguínea , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/patologia , Glomérulos Renais/ultraestrutura , Lisinopril/administração & dosagem , Masculino , Camundongos , Camundongos Transgênicos , Microscopia Eletrônica de Transmissão , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/farmacocinética , Peptídeos Cíclicos/administração & dosagem , Peptídeos Cíclicos/química , Peptídeos Cíclicos/farmacocinética , Proteinúria/diagnóstico , Proteinúria/etiologia , Proteinúria/patologia , Sulfetos/administração & dosagem , Sulfetos/química , Sulfetos/farmacocinéticaRESUMO
BACKGROUND: Focal segmental glomerulosclerosis is a kidney disease that is manifested as the nephrotic syndrome. It is often resistant to glucocorticoid therapy and progresses to end-stage renal disease in 50 to 70% of patients. Genetic studies have shown that familial focal segmental glomerulosclerosis is a disease of the podocytes, which are major components of the glomerular filtration barrier. However, the molecular cause in over half the cases of primary focal segmental glomerulosclerosis is unknown, and effective treatments have been elusive. METHODS: We performed whole-genome linkage analysis followed by high-throughput sequencing of the positive-linkage area in a family with autosomal recessive focal segmental glomerulosclerosis (index family) and sequenced a newly discovered gene in 52 unrelated patients with focal segmental glomerulosclerosis. Immunohistochemical studies were performed on human kidney-biopsy specimens and cultured podocytes. Expression studies in vitro were performed to characterize the functional consequences of the mutations identified. RESULTS: We identified two mutations (A159P and Y695X) in MYO1E, which encodes a nonmuscle class I myosin, myosin 1E (Myo1E). The mutations in MYO1E segregated with focal segmental glomerulosclerosis in two independent pedigrees (the index family and Family 2). Patients were homozygous for the mutations and did not have a response to glucocorticoid therapy. Electron microscopy showed thickening and disorganization of the glomerular basement membrane. Normal expression of Myo1E was documented in control human kidney-biopsy specimens in vivo and in glomerular podocytes in vitro. Transfection studies revealed abnormal subcellular localization and function of the A159P-Myo1E mutant. The Y695X mutation causes loss of calmodulin binding and of the tail domains of Myo1E. CONCLUSIONS: MYO1E mutations are associated with childhood-onset, glucocorticoid-resistant focal segmental glomerulosclerosis. Our data provide evidence of a role of Myo1E in podocyte function and the consequent integrity of the glomerular filtration barrier.
Assuntos
Glomerulosclerose Segmentar e Focal/genética , Mutação , Miosina Tipo I/genética , Idade de Início , Animais , Criança , Pré-Escolar , Resistência a Medicamentos , Feminino , Genes Recessivos , Ligação Genética , Estudo de Associação Genômica Ampla , Glomerulosclerose Segmentar e Focal/patologia , Glucocorticoides/uso terapêutico , Humanos , Lactente , Glomérulos Renais/irrigação sanguínea , Glomérulos Renais/ultraestrutura , Masculino , Camundongos , Microscopia de Fluorescência , Mutação de Sentido Incorreto , Miosina Tipo I/química , Miosina Tipo I/metabolismo , Linhagem , Podócitos/metabolismo , Podócitos/ultraestrutura , Alinhamento de SequênciaRESUMO
Bone marrow-derived mesenchymal stromal cells (MSC) have emerged as useful cell population for immunomodulation therapy in transplantation. Moving this concept towards clinical application, however, should be critically assessed by a tailor-made step-wise approach. Here, we report results of the second step of the multistep MSC-based clinical protocol in kidney transplantation. We examined in two living-related kidney transplant recipients whether: (i) pre-transplant (DAY-1) infusion of autologous MSC protected from the development of acute graft dysfunction previously reported in patients given MSC post-transplant, (ii) avoiding basiliximab in the induction regimen improved the MSC-induced Treg expansion previously reported with therapy including this anti-CD25-antibody. In patient 3, MSC treatment was uneventful and graft function remained normal during 1 year follow-up. In patient 4, acute cellular rejection occurred 2 weeks post-transplant. Both patients had excellent graft function at the last observation. Circulating memory CD8(+) T cells and donor-specific CD8(+) T-cell cytolytic response were reduced in MSC-treated patients, not in transplant controls not given MSC. CD4(+) FoxP3(+) Treg expansion was comparable in MSC-treated patients with or without basiliximab induction. Thus, pre-transplant MSC no longer negatively affect kidney graft at least to the point of impairing graft function, and maintained MSC-immunomodulatory properties. Induction therapy without basiliximab does not offer any advantage on CD4(+) FoxP3(+) Treg expansion (ClinicalTrials.gov number: NCT 00752479).
Assuntos
Rejeição de Enxerto/prevenção & controle , Transplante de Rim/métodos , Transplante de Células-Tronco Mesenquimais/métodos , Imunologia de Transplantes , Adulto , Anticorpos Monoclonais/efeitos adversos , Soro Antilinfocitário/uso terapêutico , Basiliximab , Protocolos Clínicos , Feminino , Humanos , Imunossupressores/efeitos adversos , Falência Renal Crônica/cirurgia , Masculino , Proteínas Recombinantes de Fusão/efeitos adversos , Linfócitos T Reguladores/imunologiaRESUMO
Tolerance induction toward allogeneic organ grafts represents one of the major aims of transplantation medicine. Stem cells are promising candidates for promoting donor-specific tolerance. In this study, we investigated the immunomodulatory properties of murine embryonic stem cells (ESCs), obtained either by in vitro fertilization (IVF-ESCs) or by nuclear transfer (NT-ESCs), in heart transplant mouse models. IVF-ESCs did not prolong the survival of fully allogeneic cardiac transplants but significantly prolonged the survival of semiallogeneic hearts from the same ESC donor strain for >100 d in 44% of the animals. However, 28% of transplanted animals infused with IVF-ESCs experienced development of a teratoma. NT-ESCs similarly prolonged semiallogeneic heart graft survival (>100 d in 40% of the animals) but were less teratogenic. By in vitro studies, IVF-ESC and NT-ESC immunoregulation was mediated both by cell contact-dependent mechanisms and by the release of soluble factors. By adding specific inhibitors, we identified PGE(2) as a soluble mediator of ESC immunoregulation. Expansion of regulatory T cells was found in lymphoid organs and in the grafts of IVF-ESC- and NT-ESC-tolerized mice. Our study demonstrates that both IVF-ESCs and NT-ESCs modulate recipient immune response toward tolerance to solid organ transplantation, and that NT-ESCs exhibit a lower tendency for teratoma formation. Because NT-ESCs are obtained by NT of a somatic cell from living individuals into an enucleated oocyte, they could represent a source of donor-derived stem cells to induce tolerance to solid organ allograft.
Assuntos
Células-Tronco Embrionárias/imunologia , Células-Tronco Embrionárias/transplante , Fertilização in vitro , Sobrevivência de Enxerto/imunologia , Transplante de Coração/imunologia , Proteínas Nucleares/administração & dosagem , Transferência Adotiva , Animais , Linhagem Celular , Feminino , Rejeição de Enxerto/prevenção & controle , Transplante de Coração/métodos , Transplante de Coração/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Transgênicos , Distribuição Aleatória , Transplante Homólogo/imunologia , Transplante Homólogo/patologiaRESUMO
Anemia can contribute to chronic allograft injury by limiting oxygen delivery to tissues, particularly in the tubulointerstitium. To determine mechanisms by which erythropoietin (EPO) prevents chronic allograft injury we utilized a rat model of full MHC-mismatched kidney transplantation (Wistar Furth donor and Lewis recipients) with removal of the native kidneys. EPO treatment entirely corrected post-transplant anemia. Control rats developed progressive proteinuria and graft dysfunction, tubulointerstitial damage, inflammatory cell infiltration, and glomerulosclerosis, all prevented by EPO. Normalization of post-transplant hemoglobin levels by blood transfusions, however, had no impact on chronic allograft injury, indicating that EPO-mediated graft protection went beyond the correction of anemia. Compared to syngeneic grafts, control allografts had loss of peritubular capillaries, higher tubular apoptosis, tubular and glomerular oxidative injury, and reduced expression of podocyte nephrin; all prevented by EPO treatment. The effects of EPO were associated with preservation of intragraft expression of angiogenic factors, upregulation of the anti-apoptotic factor p-Akt in tubuli, and increased expression of Bcl-2. Inhibition of p-Akt by Wortmannin partially antagonized the effect of EPO on allograft injury and tubular apoptosis, and prevented EPO-induced Bcl-2 upregulation. Thus non-erythropoietic derivatives of EPO may be useful to prevent chronic renal allograft injury.
Assuntos
Anemia/tratamento farmacológico , Eritropoetina/farmacologia , Hematínicos/farmacologia , Nefropatias/prevenção & controle , Transplante de Rim/efeitos adversos , Rim/efeitos dos fármacos , Anemia/sangue , Anemia/etiologia , Animais , Apoptose/efeitos dos fármacos , Biomarcadores/sangue , Transfusão de Sangue , Doença Crônica , Glomerulonefrite/imunologia , Glomerulonefrite/prevenção & controle , Hemoglobinas/metabolismo , Histocompatibilidade , Rim/imunologia , Rim/metabolismo , Rim/patologia , Nefropatias/imunologia , Nefropatias/metabolismo , Nefropatias/patologia , Transplante de Rim/imunologia , Camundongos , Disfunção Primária do Enxerto/imunologia , Disfunção Primária do Enxerto/prevenção & controle , Proteinúria/imunologia , Proteinúria/prevenção & controle , Ratos , Ratos Endogâmicos Lew , Ratos Endogâmicos WF , Fatores de TempoRESUMO
One of the leading causes of long-term kidney graft loss is chronic allograft injury (CAI), a pathological process triggered by alloantigen-dependent and alloantigen-independent factors. Alloantigen-independent factors, such as cold ischemia (CI) may amplify the recipient immune response against the graft. We investigated the impact of prolonged cold ischemia and the subsequent delayed graft function on CAI in a fully MHC-mismatched rat model of kidney allotransplantation. Prolonged CI was associated with anticipation of proteinuria onset and graft function deterioration (ischemia: 90d; no ischemia: 150d), more severe tubular atrophy, interstitial fibrosis, and glomerulosclerosis, and increased mortality rate (180d survival, ischemia: 0%; no ischemia: 67%). In ischemic allografts, T and B cells were detected very early and were organized in inflammatory clusters. Higher expression of BAFF-R and TACI within the ischemic allografts indicates that B cells are mature and activated. As a consequence of B cell activity, anti-donor antibodies, glomerular C4d and IgG deposition, important features of chronic humoral rejection, appeared earlier in ischemic than in non-ischemic allograft recipients. Thus, prolonged CI time plays a main role in CAI development by triggering acceleration of cellular and humoral reactions of chronic rejection. Limiting CI time should be considered as a main target in kidney transplantation.
Assuntos
Isquemia Fria/efeitos adversos , Função Retardada do Enxerto/etiologia , Rejeição de Enxerto/etiologia , Transplante de Rim/imunologia , Animais , Apoptose , Função Retardada do Enxerto/imunologia , Função Retardada do Enxerto/patologia , Citometria de Fluxo , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/patologia , Marcação In Situ das Extremidades Cortadas , Rim/imunologia , Rim/patologia , Transplante de Rim/métodos , Transplante de Rim/mortalidade , Masculino , Ratos , Ratos Endogâmicos WF , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Tempo , Transplante Homólogo/imunologia , Transplante Homólogo/métodos , Transplante Homólogo/mortalidadeRESUMO
In search for new sources of mesenchymal stem cells (MSCs) for renal repair in acute kidney injury (AKI), we investigated the potential of human cord blood (CB)-MSCs to cure mice with AKI. Infusion of CB-MSCs in immunodeficient mice with cisplatin-induced AKI ameliorated both renal function and tubular cell injury, and prolonged survival. Transplanted CB-MSCs localized in peritubular areas, limited capillary alterations and neutrophil infiltration. Apoptosis reduced and tubular cell proliferation increased by virtue of stem cell capacity to produce growth factors. The reno-protective effect of CB-MSCs was further confirmed by their ability to inhibit oxidative damage and to induce the prosurvival factor Akt in tubular cells. The evidence that CB-MSCs in vitro increased the production of growth factors and inhibited IL-1 beta and TNFalpha synthesis when cocultured with damaged proximal tubular cells indicates a regenerative and anti-inflammatory action of stem cell treatment. Altogether these results highlight the potential of human CB-MSCs as future cell therapy for testing in human AKI.
Assuntos
Diferenciação Celular/fisiologia , Transplante de Células-Tronco de Sangue do Cordão Umbilical/métodos , Sobrevivência de Enxerto/fisiologia , Nefropatias/cirurgia , Rim/cirurgia , Transplante de Células-Tronco Mesenquimais/métodos , Doença Aguda , Animais , Anti-Inflamatórios/metabolismo , Apoptose/fisiologia , Técnicas de Cultura de Células , Proliferação de Células , Técnicas de Cocultura , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Rim/patologia , Rim/fisiopatologia , Nefropatias/fisiopatologia , Túbulos Renais/citologia , Túbulos Renais/metabolismo , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos SCID , Estresse Oxidativo/fisiologia , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Ischemia/reperfusion (I/R) injury is an important cause of renal graft dysfunction in humans. Increases in cold and warm ischemia times lead to a higher risk of early post-transplant complications including delayed graft function and acute rejection. Moreover, prolonged cold ischemia is a predictor of long-term kidney graft loss. The protective effect of rabbit anti-rat thymocyte immunoglobulin (rATG) was evaluated in a rat model of I/R injury following syngeneic kidney transplantation. Serum creatinine concentration was evaluated at 16 h and 24 h post-transplant. Animals were sacrificed 24 h post-transplant for evaluation of histology, infiltrating leukocytes, nitrotyrosine staining, and apoptosis. rATG was effective in preventing renal function impairment, tissue damage and tubular apoptosis associated with I/R only when was given 2 h before transplantation but not at the time of reperfusion. Pretransplant rATG treatment of recipient animals effectively reduced the amount of macrophages, CD4(+), CD8(+) T cells and LFA-1(+) cells infiltrating renal graft subjected to cold ischemia as well as granzyme-B expression within ischemic kidney. On the other hand, granulocyte infiltration and oxidative stress were not modified by rATG. If these results will be translated into the clinical setting, pretransplant administration of Thymoglobuline(®) could offer the additional advantage over peri-transplant administration of limiting I/R-mediated kidney graft damage.
Assuntos
Imunoglobulinas/metabolismo , Transplante de Rim/métodos , Rim/metabolismo , Traumatismo por Reperfusão/metabolismo , Timócitos/citologia , Animais , Apoptose , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Granzimas/farmacologia , Isquemia/patologia , Masculino , Microscopia de Fluorescência/métodos , Coelhos , Ratos , Ratos Endogâmicos LewRESUMO
Members of the TLR/IL-1R superfamily mediate ischemia/reperfusion injury and initiate immune response in transplanted organs. In this study, we tested the hypothesis that Toll-IL-1R8 (TIR8), a negative regulator of TLR/IL-1R highly expressed in the kidney, modulates immune cell activation underlying kidney rejection. In a mouse model of fully mismatched kidney allotransplantation in which the graft is spontaneously accepted, intragraft Tir8 expression was enhanced compared with naive kidneys. Targeted deletion of Tir8 in the graft exerted a powerful antitolerogenic action leading to acute rejection. Similarly, in a mouse model of kidney graft acceptance induced by costimulation blockade, most Tir8(-/-) grafts were acutely rejected. Despite similar levels of TLR4, IL-1R, and their ligands, the posttransplant ischemia/reperfusion-induced inflammatory response was more severe in Tir8(-/-) than in Tir8(+/+) grafts and was followed by expansion and maturation of resident dendritic cell precursors. In vitro, Tir8(-/-) dendritic cell precursors acquired higher allostimulatory activity and released more IL-6 upon stimulation with a TLR4 ligand and TNF-alpha than Tir8(+/+) cells, which may explain the increased frequency of antidonor-reactive T cells and the block of regulatory T cell formation in recipients of a Tir8(-/-) kidney. Thus, TIR8 acts locally as a key regulator of allogeneic immune response in the kidney. Tir8 expression and/or signaling in donor tissue are envisaged as a novel target for control of innate immunity and amelioration of graft survival.
Assuntos
Regulação para Baixo/imunologia , Rejeição de Enxerto/imunologia , Transplante de Rim/imunologia , Glicoproteínas de Membrana/fisiologia , Receptores de Interleucina-1/fisiologia , Doença Aguda , Animais , Modelos Animais de Doenças , Regulação para Baixo/genética , Rejeição de Enxerto/genética , Rejeição de Enxerto/prevenção & controle , Tolerância Imunológica/genética , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Masculino , Glicoproteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Endogâmicos DBA , Camundongos Knockout , Receptores de Interleucina-1/antagonistas & inibidores , Receptores de Interleucina-1/deficiência , Receptores de Interleucina-1/genética , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/imunologiaRESUMO
BACKGROUND: Vulvovestibular syndrome (VVS) or vulvodynia is a chronic, heterogeneous and multifactorial disease that dramatically affects women's health and quality of life. Despite important advancements in understanding VVS etiology have been achieved in the past decades, VVS still remains an elusive and complex condition without identifiable causes and effective treatments. In the present observational, retrospective, case-control study, we sought to investigate whether gut dysbiosis developed in patients with VVS. METHODS: To this aim, we compared both bacterial and fungal composition in VVS patients (n = 74; 34.3 ± 10.9 years old) with those of women without gynecological symptoms (n = 13 healthy control; 38.3 ± 10.4 years old). Furthermore, to assess whether gut ecology may have an impact on gut function, the degree of intestinal inflammation (calprotectin levels) and gut permeability (zonulin levels) were also evaluated. RESULTS: VVS patient developed gut dysbiosis, mainly characterized by a significant increase of Escherichia coli along with increased colonization of mold/yeast compared to healthy controls. Furthermore, fecal levels of zonulin indicated that in VVS patients gut dysbiosis translated into increased gut permeability. CONCLUSION: Our preliminary study, by demonstrating that alterations in gut microbiota and intestinal permeability are present in patients with VVS, highlights the novel notion that gut dysbiosis may be considered an important associated factor for VVS. These findings, if confirmed, may be clinically relevant and may help in choosing further diagnostic methods and more effective therapies for these patients.
RESUMO
Angiotensin II (Ang II), the central product of renin-angiotensin system, has a role in the etiology of hypertension and in pathophysiology of cardiac and renal diseases in humans. Other functions of Ang II include effects on immune response, inflammation, cell growth and proliferation, which are largely mediated by Ang II type 1 receptor (AT(1)). Several experimental studies have demonstrated that Ang II acts through AT(1) as a mediator of normal aging processes by increasing oxidant damage to mitochondria and in consequences by affecting mitochondrial function. Recently, our group has demonstrated that the inhibition of Ang II activity by targeted disruption of the Agtr1a gene encoding Ang II type 1A receptor (AT(1A)) in mice translates into marked prolongation of life span. The absence of AT(1A) protected multiple organs from oxidative damage and the alleviation of aging-like phenotype was associated with increased number of mitochondria and upregulation of the prosurvival gene sirtuin 3. AT(1) receptor antagonists have been proven safe and well-tolerated for chronic use and are used as a key component of the modern therapy for hypertension and cardiac failure, therefore Ang II/AT(1) pathway represents a feasible therapeutic strategy to prolong life span in humans.
Assuntos
Envelhecimento/fisiologia , Angiotensina II/fisiologia , Longevidade/fisiologia , Receptores de Angiotensina/fisiologia , Sirtuínas/fisiologia , Envelhecimento/efeitos dos fármacos , Antagonistas de Receptores de Angiotensina , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Animais , Humanos , Mitocôndrias/fisiologia , Estresse Oxidativo/fisiologia , Sistema Renina-Angiotensina/fisiologia , Sirtuína 3/fisiologiaRESUMO
In this study, we investigated whether mesenchymal stem cells (MSC) had immunomodulatory properties in solid organ allotransplantation, using a semiallogeneic heart transplant mouse model, and studied the mechanism(s) underlying MSC tolerogenic effects. Either single (portal vein, day -7) or double (portal vein, day -7 and tail vein, day -1) pretransplant infusions of donor-derived B6C3 MSC in B6 recipients induced a profound T cell hyporesponsiveness and prolonged B6C3 cardiac allograft survival. The protolerogenic effect was abrogated when donor-derived MSC were injected together with B6C3 hematopoietic stem cells (HSC), suggesting that HSC negatively impact MSC immunomodulatory properties. Both the induction (pretransplant) and the maintenance phase (>100 days posttransplant) of donor-derived MSC-induced tolerance were associated with CD4(+)CD25(+)Foxp3(+) Treg expansion and impaired anti-donor Th1 activity. MSC-induced regulatory T cells (Treg) were donor-specific since adoptive transfer of splenocytes from tolerant mice prevented the rejection of fully MHC-mismatched donor-specific secondary allografts but not of third-party grafts. In addition, infusion of recipient-derived B6 MSC tolerized a semiallogeneic B6C3 cardiac allograft, but not a fully MHC-mismatched BALB/c graft, and expanded Treg. A double i.v. pretransplant infusion of recipient-derived MSC had the same tolerogenic effect as the combined intraportal/i.v. MSC infusions, which makes the tolerogenic protocol applicable in a clinical setting. In contrast, single MSC infusions given either peritransplant or 1 day after transplant were less effective. Altogether these findings indicate that MSC immunomodulatory properties require HSC removal, partial sharing of MHC Ags between the donor and the recipient and pretransplant infusion, and are associated with expansion of donor-specific Treg.
Assuntos
Diferenciação Celular/imunologia , Sobrevivência de Enxerto/imunologia , Transplante de Coração/imunologia , Transplante de Células-Tronco Mesenquimais , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologia , Condicionamento Pré-Transplante , Tolerância ao Transplante/imunologia , Animais , Células da Medula Óssea/imunologia , Feminino , Transplante de Coração/métodos , Masculino , Transplante de Células-Tronco Mesenquimais/métodos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Condicionamento Pré-Transplante/métodos , Transplante Heterotópico , Transplante Homólogo , Transplante IsogênicoRESUMO
In childhood, almost all swallowed objects that successfully navigate the esophagus pass through the gut without complications. In a 15-year-old male adolescent with the initial working diagnosis of acalculous cholecystitis, computed tomography revealed a thickened wall of the second duodenal portion, some infiltration of the periduodenal tissue, and a hyperdense needle-shape structure probably passing through the duodenal wall. Endoscopy revealed a wooden toothpick perforating the duodenum that was carefully retracted. An uneventful recovery followed the endoscopic removal of the foreign body. A computer-based search of the literature to examine the injuries caused by ingested toothpicks since 1960 found only 4 reports in 5 children.
Assuntos
Duodeno , Corpos Estranhos/complicações , Perfuração Intestinal/etiologia , Dor Abdominal/etiologia , Adolescente , Duodeno/diagnóstico por imagem , Endoscopia , Corpos Estranhos/diagnóstico por imagem , Corpos Estranhos/cirurgia , Humanos , Perfuração Intestinal/diagnóstico por imagem , Masculino , Recidiva , Tomografia Computadorizada por Raios XRESUMO
More effective treatments for diabetic nephropathy remain a major unmet clinical need. Increased oxidative stress is one of the most important pathological mechanisms that lead to kidney damage and functional impairment induced by diabetes. Sirtuin 3 (SIRT3) is the main mitochondrial deacetylase and critically regulates cellular reactive oxygen species (ROS) production and detoxification. Honokiol is a natural biphenolic compound that, by activating mitochondrial SIRT3, can carry out anti-oxidant, anti-inflammatory and anti-fibrotic activities. Here, we sought to investigate the renoprotective effects of honokiol in BTBR ob/ob mice with type 2 diabetes. Diabetic mice were treated with vehicle or honokiol between the ages of 8 and 14 weeks. Wild-type mice served as controls. Renal Sirt3 expression was significantly reduced in BTBR ob/ob mice, and this was associated with a reduction in its activity and increased ROS levels. Selective activation of SIRT3 through honokiol administration translated into the attenuation of albuminuria, amelioration of glomerular damage, and a reduction in podocyte injury. SIRT3 activation preserved mitochondrial wellness through the activation of SOD2 and the restoration of PGC-1α expression in glomerular cells. Additionally, the protective role of SIRT3 in glomerular changes was associated with enhanced tubular Sirt3 expression and upregulated renal Nampt levels, indicating a possible tubule-glomerulus retrograde interplay, which resulted in improved glomerular SIRT3 activity. Our results demonstrate the hitherto unknown renoprotective effect of SIRT3 against diabetic glomerular disease and suggest that the pharmacological modulation of SIRT3 activity is a possible novel approach to treating diabetic nephropathy.
Assuntos
Anti-Inflamatórios/uso terapêutico , Antioxidantes/uso terapêutico , Compostos de Bifenilo/uso terapêutico , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Nefropatias Diabéticas/tratamento farmacológico , Glomérulos Renais/patologia , Lignanas/uso terapêutico , Albuminúria/prevenção & controle , Animais , Citocinas/metabolismo , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/patologia , Masculino , Camundongos , Camundongos Obesos , Mitocôndrias/metabolismo , Nicotinamida Fosforribosiltransferase/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Podócitos/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Sirtuína 3/metabolismo , Superóxido Dismutase/metabolismoRESUMO
Renal activation of the complement system has been described in patients with diabetic nephropathy (DN), although its pathological relevance is still ill-defined. Here, we studied whether glomerular C3a, generated by uncontrolled complement activation, promotes podocyte damage, leading to proteinuria and renal injury in mice with type 2 diabetes. BTBR ob/ob mice exhibited podocyte loss, albuminuria, and glomerular injury accompanied by C3 deposits and increased C3a and C3a receptor (C3aR) levels. Decreased glomerular nephrin and α-actinin4 expression, coupled with integrin-linked kinase induction, were also observed. Treatment of DN mice with a C3aR antagonist enhanced podocyte density and preserved their phenotype, limiting proteinuria and glomerular injury. Mechanistically, ultrastructural and functional mitochondrial alterations, accompanied by downregulation of antioxidant superoxide dismutase 2 (SOD2) and increased protein oxidation, occurred in podocytes and were normalized by C3aR blockade. In cultured podocytes, C3a induced cAMP-dependent mitochondrial fragmentation. Alterations of mitochondrial membrane potential, SOD2 expression, and energetic metabolism were also found in response to C3a. Notably, C3a-induced podocyte motility was inhibited by SS-31, a peptide with mitochondrial protective effects. These data indicate that C3a blockade represents a potentially novel therapeutic strategy in DN for preserving podocyte integrity through the maintenance of mitochondrial functions.
Assuntos
Complemento C3a/metabolismo , Nefropatias Diabéticas/patologia , Podócitos/patologia , Receptores de Complemento/antagonistas & inibidores , Animais , Ativação do Complemento , AMP Cíclico/metabolismo , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/metabolismo , Nefropatias Diabéticas/metabolismo , Modelos Animais de Doenças , Glomérulos Renais/patologia , Camundongos , Mitocôndrias/metabolismo , Estresse Oxidativo , Podócitos/metabolismo , Receptores de Complemento/metabolismo , Superóxido Dismutase/metabolismoRESUMO
Introduction: Animal models have provided significant insights into the mechanisms responsible for the development of glomerular lesions and proteinuria; they have also helped to identify molecules that control the podocyte function as suitable target-specific therapeutics. Areas covered: We discuss putative therapeutic targets for proteinuric glomerular diseases. An exhaustive search for eligible studies was performed in PubMed/MEDLINE. Most of the selected reports were published in the last decade, but we did not exclude older relevant milestone publications. We consider the molecules that regulate podocyte cytoskeletal dynamics and the transcription factors that regulate the expression of slit-diaphragm proteins. There is a focus on SGLT2 and sirtuins which have recently emerged as mediators of podocyte injury and repair. We also examine paracrine signallings involved in the cross-talk of injured podocytes with the neighbouring glomerular endothelial cells and parietal epithelial cells. Expert opinion: There is a need to discover novel therapeutic moleecules with renoprotective effects for those patients with glomerular diseases who do not respond completely to standard therapy. Emerging strategies targeting components of the podocyte cytoskeleton or signallings that regulate cellular communication within the glomerulus are promising avenues for treating glomerular diseases.
Assuntos
Nefropatias/tratamento farmacológico , Glomérulos Renais/fisiopatologia , Proteinúria/tratamento farmacológico , Animais , Citoesqueleto/metabolismo , Modelos Animais de Doenças , Descoberta de Drogas , Células Endoteliais/metabolismo , Humanos , Nefropatias/fisiopatologia , Podócitos/metabolismo , Substâncias Protetoras/farmacologia , Proteinúria/patologiaRESUMO
Aims: Sirtuins, a family of NAD+-dependent deacetylases, are recognized as nondispensable regulators of aging processes. Sirtuin 3 (SIRT3) is the main mitochondrial deacetylase that maintains mitochondrial bioenergetics, an essential prerequisite for healthy aging. In this study, using Sirt3 knockout (Sirt3-/-) mice, we sought to establish whether Sirt3 deficiency affected life span, an endpoint that has never been tested formally in mammals, and uncover the mechanisms involved in organ damage associated with aging. Results:Sirt3-/- mice experienced a shorter life span than wild-type mice and severe cardiac damage, characterized by hypertrophy and fibrosis, as they aged. No alterations were found in organs other than the heart. Sirt3 deficiency altered cardiac mitochondrial bioenergetics and caused hyperacetylation of optic atrophy 1 (OPA1), a SIRT3 target. These changes were associated with aberrant alignment of trans-mitochondrial cristae in cardiomyocytes, and cardiac dysfunction. Gene transfer of deacetylated Opa1 restored cristae alignment in Sirt3-/- mice, ameliorated cardiac reserve capacity, and protected the heart against hypertrophy and fibrosis. The translational relevance of these findings is in the data showing that SIRT3 silencing in human-induced pluripotent stem cell-derived cardiomyocytes led to mitochondrial dysfunction and altered contractile phenotype, both rescued by Opa1 gene transfer. Innovation: Our findings indicate that future approaches to heart failure could include SIRT3 as a plausible therapeutic target. Conclusion: SIRT3 has a major role in regulating mammalian life span. Sirt3 deficiency leads to cardiac abnormalities, due to defective trans-mitochondrial cristae alignment and impaired mitochondrial bioenergetics. Correcting cardiac OPA1 hyperacetylation through gene transfer diminished heart failure in Sirt3-/- mice during aging. Antioxid. Redox Signal. 31, 1255-1271.
Assuntos
GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Longevidade , Mitocôndrias Cardíacas/metabolismo , Sirtuína 3/deficiência , Sirtuína 3/metabolismo , Acetilação , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
BACKGROUND: Mesenchymal stromal cells (MSCs) have protolerogenic effects in renal transplantation, but they induce long-term regulatory T cells (Treg)-dependent graft acceptance only when infused before transplantation. When given posttransplant, MSCs home to the graft where they promote engraftment syndrome and do not induce Treg. Unfortunately, pretransplant MSC administration is unfeasible in deceased-donor kidney transplantation. METHODS: To make MSCs a therapeutic option also for deceased organ recipients, we tested whether MSC infusion at the time of transplant (day 0) or posttransplant (day 2) together with inhibition of complement receptors prevents engraftment syndrome and allows their homing to secondary lymphoid organs for promoting tolerance. We analyzed intragraft and splenic MSC localization, graft survival, and alloimmune response in mice recipients of kidney allografts and syngeneic MSCs given on day 0 or on posttransplant day 2. C3a receptor (C3aR) or C5a receptor (C5aR) antagonists were administered to mice in combination with the cells or were used together to treat MSCs before infusion. RESULTS: Syngeneic MSCs given at day 0 homed to the spleen increased Treg numbers and induced long-term graft acceptance. Posttransplant MSC infusion, combined with a short course of C3aR or C5aR antagonist or administration of MSCs pretreated with C3aR and C5aR antagonists, prevented intragraft recruitment of MSCs and graft inflammation, inhibited antidonor T-cell reactivity, but failed to induce Treg, resulting in mild prolongation of graft survival. CONCLUSIONS: These data support testing the safety/efficacy profile of administering MSCs on the day of transplant in deceased-donor transplant recipients and indicate that complement is crucial for MSC recruitment into the kidney allograft.
Assuntos
Inativadores do Complemento/administração & dosagem , Rejeição de Enxerto/prevenção & controle , Sobrevivência de Enxerto/efeitos dos fármacos , Transplante de Rim , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/efeitos dos fármacos , Receptores de Complemento/antagonistas & inibidores , Tolerância ao Transplante/efeitos dos fármacos , Animais , Esquema de Medicação , Feminino , Rejeição de Enxerto/imunologia , Células-Tronco Mesenquimais/imunologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Receptores de Complemento/imunologia , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Acoplados a Proteínas G/imunologia , Baço/efeitos dos fármacos , Baço/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Fatores de Tempo , Transplante Homólogo , Transplante IsogênicoRESUMO
Ischemia-reperfusion injury is an important cause of graft failure. Because carnitine regulates substrate flux and energy balance across membranes which may be deranged in ischemia we determined whether its use was effective in preventing kidney injury in an allogeneic transplant model. Brown Norway rats received a Lewis rat kidney transplant and were then treated with cyclosporine A to avoid rejection. The grafts were stored in Belzer solution supplemented with propionyl-L-carnitine during the cold ischemia period. Compared to rats receiving untreated kidneys but with equal cold ischemia times, the post-transplant serum creatinine values of the carnitine-treated transplants were significantly lower. Histological evaluation 16 h after transplant showed that propionyl-L-carnitine significantly inhibited tubular necrosis and neutrophil infiltration of the allografts and improved the 3 month graft survival. Treated transplants also had decreased lipid peroxidation, inducible nitric oxide synthase expression and protein nitration compared to the untreated grafts. Post-transplant serum creatinine levels were significantly reduced and graft survival was slightly prolonged in rats not receiving cyclosporine A treatment and transplanted with a kidney treated with propionyl-L-carnitine. The efficacy of propionyl-L-carnitine to modulate ischemia-reperfusion injury during transplantation suggests that its use in human transplantation is worth testing.
Assuntos
Carnitina/análogos & derivados , Sobrevivência de Enxerto/efeitos dos fármacos , Transplante de Rim/métodos , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Carnitina/farmacologia , Ciclosporina/farmacologia , Necrose do Córtex Renal/prevenção & controle , Transplante de Rim/efeitos adversos , Infiltração de Neutrófilos/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Ratos , Traumatismo por Reperfusão , Transplante HomólogoRESUMO
CONTEXT: Expansion and culture of beta cell progenitors in vitro may represent an alternative to the use of differentiated beta cells from donor pancreata. OBJECTIVE: The aim of our study was to investigate to what extent exocrine or endocrine pancreatic cells can be differentiated in insulin-producing cells in vitro. SETTING: Bovine exocrine tissue (n=4) and islets (n=4) were cultured in DMEM with serum. INTERVENTIONS: After 7 days, the cells were trypsinized and cultured in the same medium for cell proliferation, or in DMEM/F-12 containing growth factors to induce cell differentiation. MAIN OUTCOME MEASURE: Proliferating capacity after 4 weeks in culture. In addition, insulin expression was evaluated by RT-PCR and by immunohistochemical staining. RESULTS: After 4 weeks of culture, cells from exocrine tissue showed a 69.5+/-10.0 fold increase, while cells from islets showed a 31.2+/-11.4 fold increase (P=0.059). In differentiating medium, monolayers from exocrine and islet tissue were organized into islet-like structures containing cells which stained positively for insulin. Morphometrical analysis and RT-PCR confirmed the presence of insulin in the cells at the protein and the mRNA level. CONCLUSIONS: In our experimental conditions, cells from pancreatic tissue proliferated and differentiated in insulin-containing cells. However, the level of insulin as well as mRNA expression is only a small fraction of that shown by fresh islets. Only selective identification of cell precursors may allow efficient generation of insulin-producing cells in vitro.