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1.
Kidney Int ; 97(1): 130-142, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31685310

RESUMO

Innate lymphoid cells are a recently recognized group of immune cells with critical roles in tissue homeostasis and inflammation. Regulatory innate lymphoid cells are a newly identified subset of innate lymphoid cells, which play a suppressive role in the innate immune response, favoring the resolution of intestinal inflammation. However, the expression and role of regulatory innate lymphoid cells in kidney has not been reported. Here, we show that regulatory innate lymphoid cells are present in both human and mouse kidney, express similar surface markers and form a similar proportion of total kidney innate lymphoid cells. Regulatory innate lymphoid cells from kidney were expanded in vitro with a combination of IL-2, IL-7 and transforming growth factor-ß. These cells exhibited immunosuppressive effects on innate immune cells via secretion of IL-10 and transforming growth factor-ß. Moreover, treatment with IL-2/IL-2 antibody complexes (IL-2C) promoted expansion of regulatory innate lymphoid cells in vivo, and prevent renal ischemia/reperfusion injury in Rag-/- mice that lack adaptive immune cells including Tregs. Depletion of regulatory innate lymphoid cells with anti-CD25 antibody abolished the beneficial effects of IL-2C in the Rag-/- mice. Adoptive transfer of ex vivo expanded regulatory innate lymphoid cells improved renal function and attenuated histologic damage when given before or after induction of ischemia/reperfusion injury in association with reduction of neutrophil infiltration and induction of reparative M2 macrophages in kidney. Thus, our study shows that regulatory innate lymphoid cells suppress innate renal inflammation and ischemia/reperfusion injury.


Assuntos
Imunidade Inata , Rim/citologia , Subpopulações de Linfócitos/imunologia , Nefrite/imunologia , Traumatismo por Reperfusão/complicações , Transferência Adotiva , Animais , Separação Celular , Células Cultivadas , Técnicas de Cocultura , Modelos Animais de Doenças , Citometria de Fluxo , Proteínas de Homeodomínio/genética , Humanos , Interleucina-10/metabolismo , Interleucina-2/antagonistas & inibidores , Interleucina-2/metabolismo , Rim/irrigação sanguínea , Rim/imunologia , Rim/patologia , Subpopulações de Linfócitos/metabolismo , Subpopulações de Linfócitos/transplante , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Knockout , Nefrite/patologia , Cultura Primária de Células , Traumatismo por Reperfusão/imunologia , Traumatismo por Reperfusão/patologia , Fator de Crescimento Transformador beta/metabolismo
2.
Nephrol Dial Transplant ; 34(11): 1853-1863, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-30590794

RESUMO

BACKGROUND: Chronic kidney disease (CKD) is a global public health problem, which lacks effective treatment. Previously, we have shown that CD103+ dendritic cells (DCs) are pathogenic in adriamycin nephropathy (AN), a model of human focal segmental glomerulosclerosis (FSGS). Fms-like tyrosine kinase 3 (Flt3) is a receptor that is expressed with high specificity on tissue resident CD103+ DCs. METHODS: To test the effect on CD103+ DCs and kidney injury of inhibition of Flt3, we used a selective Flt3 inhibitor (AC220) to treat mice with AN. RESULTS: Human CD141+ DCs, homologous to murine CD103+ DCs, were significantly increased in patients with FSGS. The number of kidney CD103+ DCs, but not CD103- DCs or plasmacytoid DCs, was significantly decreased in AN mice after AC220 administration. Treatment with AC220 significantly improved kidney function and reduced kidney injury and fibrosis in AN mice. AC220-treated AN mice had decreased levels of inflammatory cytokines and chemokines, tumor necrosis factor-α, interleukin (IL)-1ß, IL-6, CCL2 and CCL5 and reduced kidney infiltration of CD4 T cells and CD8 T cells. The protective effect of AC220 was associated with its suppression of CD103+ DCs-mediated CD8 T cell proliferation and activation in AN mice. CONCLUSION: Flt3 inhibitor AC220 effectively reduced kidney injury in AN mice, suggesting that this inhibitor might be a useful pharmaceutical agent to treat CKD.


Assuntos
Antígenos CD/metabolismo , Benzotiazóis/farmacologia , Células Dendríticas/imunologia , Cadeias alfa de Integrinas/metabolismo , Rim/efeitos dos fármacos , Ativação Linfocitária/imunologia , Compostos de Fenilureia/farmacologia , Insuficiência Renal Crônica/prevenção & controle , Tirosina Quinase 3 Semelhante a fms/antagonistas & inibidores , Animais , Citocinas/metabolismo , Células Dendríticas/efeitos dos fármacos , Humanos , Rim/imunologia , Rim/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Insuficiência Renal Crônica/imunologia , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/patologia
3.
J Am Soc Nephrol ; 29(3): 961-976, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29295873

RESUMO

The IL-33-type 2 innate lymphoid cell (ILC2) axis has an important role in tissue homeostasis, inflammation, and wound healing. However, the relative importance of this innate immune pathway for immunotherapy against inflammation and tissue damage remains unclear. Here, we show that treatment with recombinant mouse IL-33 prevented renal structural and functional injury and reduced mortality in mice subjected to ischemia-reperfusion injury (IRI). Compared with control-treated IRI mice, IL-33-treated IRI mice had increased levels of IL-4 and IL-13 in serum and kidney and more ILC2, regulatory T cells (Tregs), and anti-inflammatory (M2) macrophages. Depletion of ILC2, but not Tregs, substantially abolished the protective effect of IL-33 on renal IRI. Adoptive transfer of ex vivo-expanded ILC2 prevented renal injury in mice subjected to IRI. This protective effect associated with induction of M2 macrophages in kidney and required ILC2 production of amphiregulin. Treatment of mice with IL-33 or ILC2 after IRI was also renoprotective. Furthermore, in a humanized mouse model of renal IRI, treatment with human IL-33 or transfer of ex vivo-expanded human ILC2 ameliorated renal IRI. This study has uncovered a major protective role of the IL-33-ILC2 axis in renal IRI that could be potentiated as a therapeutic strategy.


Assuntos
Interleucina-33/uso terapêutico , Nefropatias/prevenção & controle , Linfócitos/imunologia , Linfócitos/metabolismo , Traumatismo por Reperfusão/prevenção & controle , Anfirregulina/metabolismo , Animais , Feminino , Humanos , Imunidade Inata , Interleucina-13/metabolismo , Interleucina-4/metabolismo , Nefropatias/imunologia , Nefropatias/patologia , Contagem de Linfócitos , Macrófagos/imunologia , Masculino , Camundongos , Proteínas Recombinantes/uso terapêutico , Traumatismo por Reperfusão/imunologia , Traumatismo por Reperfusão/patologia , Linfócitos T Reguladores/imunologia , Células Th2/imunologia
4.
Am J Physiol Renal Physiol ; 314(4): F561-F571, 2018 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-29357438

RESUMO

Cell therapy using macrophages requires large amounts of cells, which are difficult to collect from patients. Patients undergoing peritoneal dialysis (PD) discard huge numbers of peritoneal macrophages in dialysate daily. Macrophages can be modulated to become regulatory macrophages, which have shown great promise as a therapeutic strategy in experimental kidney disease and human kidney transplantation. This study aimed to examine the potential of using peritoneal macrophages (PMs) from peritoneal dialysate to treat kidney disease. Monocytes/macrophages accounted for >40% of total peritoneal leukocytes in both patients and mice undergoing PD. PMs from patients and mice undergoing PD were more mature than peripheral monocytes/macrophages, as shown by low expression of C-C motif chemokine receptor 2 (CCR2) and morphological changes during in vitro culture. PMs from patients and mice undergoing PD displayed normal macrophage function and could be modulated into a regulatory (M2) phenotype. In vivo, adoptive transfer of peritoneal M2 macrophages derived from PD mice effectively protected against kidney injury in mice with adriamycin nephropathy (AN). Importantly, the transfused peritoneal M2 macrophages maintained their M2 phenotype in kidney of AN mice. In conclusion, PMs derived from patients and mice undergoing PD exhibited conventional macrophage features. Peritoneal M2 macrophages derived from PD mice are able to reduce kidney injury in AN, suggesting that peritoneal macrophages from patients undergoing PD may have the potential for clinical therapeutic application.


Assuntos
Transferência Adotiva , Soluções para Diálise , Doxorrubicina , Nefropatias/prevenção & controle , Rim , Macrófagos Peritoneais/transplante , Diálise Peritoneal , Animais , Plasticidade Celular , Separação Celular/métodos , Células Cultivadas , Citocinas/metabolismo , Modelos Animais de Doenças , Humanos , Mediadores da Inflamação/metabolismo , Rim/imunologia , Rim/metabolismo , Rim/patologia , Nefropatias/induzido quimicamente , Nefropatias/imunologia , Nefropatias/metabolismo , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos Endogâmicos BALB C , Fenótipo , Fatores de Tempo
5.
Am J Pathol ; 186(7): 1847-1860, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27182643

RESUMO

Loss of E-cadherin marks a defect in epithelial integrity and polarity during tissue injury and fibrosis. Whether loss of E-cadherin plays a causal role in fibrosis is uncertain. α3ß1 Integrin has been identified to complex with E-cadherin in cell-cell adhesion, but little is known about the details of their cross talk. Herein, E-cadherin gene (Cdh1) was selectively deleted from proximal tubules of murine kidney by Sglt2Cre. Ablation of E-cadherin up-regulated α3ß1 integrin at cell-cell adhesion. E-cadherin-deficient proximal tubular epithelial cell displayed enhanced transforming growth factor-ß1-induced α-smooth muscle actin (α-SMA) and vimentin expression, which was suppressed by siRNA silencing of α3 integrin, but not ß1 integrin. Up-regulation of transforming growth factor-ß1-induced α-SMA was mediated by an α3 integrin-dependent increase in integrin-linked kinase (ILK). Src phosphorylation of ß-catenin and consequent p-ß-catenin-Y654/p-Smad2 transcriptional complex underlies the transcriptional up-regulation of ILK. Kidney fibrosis after unilateral ureteric obstruction or ischemia reperfusion was increased in proximal tubule E-cadherin-deficient mice in comparison to that of E-cadherin intact control mice. The exacerbation of fibrosis was explained by the α3 integrin-dependent increase of ILK, ß-catenin nuclear translocation, and α-SMA/proximal tubular-specific Cre double positive staining in proximal tubular epithelial cell. These studies delineate a nonconventional integrin/ILK signaling by α3 integrin-dependent Src/p-ß-catenin-Y654/p-Smad2-mediated up-regulation of ILK through which loss of E-cadherin leads to kidney fibrosis.


Assuntos
Caderinas/deficiência , Integrina alfa3beta1/metabolismo , Nefropatias/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Western Blotting , Adesão Celular , Imunoprecipitação da Cromatina , Modelos Animais de Doenças , Fibrose/metabolismo , Fibrose/patologia , Imuno-Histoquímica , Imunoprecipitação , Nefropatias/metabolismo , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , Camundongos , Camundongos Knockout , Microscopia Eletrônica de Transmissão , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/fisiologia
6.
Nephrol Dial Transplant ; 32(5): 781-791, 2017 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-27566305

RESUMO

BACKGROUND: Endothelial cells are known to contribute to kidney fibrosis via endothelial-mesenchymal transition (EndoMT). Matrix metalloproteinase 9 (MMP-9) is known to be profibrotic. However, whether MMP-9 contributes to kidney fibrosis via EndoMT is unknown. METHODS: Primary mouse renal peritubular endothelial cells (MRPECs) were isolated and treated by recombinant human transforming growth factor beta 1 (rhTGF-ß1) with or without MMP-9 inhibitor or by recombinant human MMP-9 (rhMMP-9) alone. Kidney fibrosis was induced by unilateral ureteral obstruction (UUO) in MMP-9 knockout (KO) and wide-type (WT) control mice. The effects of MMP-9 on EndoMT of MRPECs and kidney fibrosis were examined. RESULTS: We showed that MRPECs underwent EndoMT after rhTGF-ß1 treatment or in UUO kidney as evidenced by decreased expression of endothelial markers, vascular endothelial cadherin (VE-cadherin) and CD31, and increased levels of mesenchymal markers, α-smooth muscle actin (α-SMA) and vimentin. The expression of fibrosis markers was also up-regulated significantly after rhTGF-ß1 treatment in MRPECs. The EndoMT and fibrosis markers were significantly less in rhTGF-ß1-treated MMP-9 KO MRPECs, whereas MMP-9 alone was sufficient to induce EndoMT in MRPECs. UUO kidney of MMP-9 KO mice showed significantly less interstitial fibrosis and EndoMT in MRPECs. Notch signaling shown by Notch intracellular domain (NICD) was increased, while Notch-1 was decreased in rhTGF-ß1-treated MRPECs of MMP-9 WT but not MMP-9 KO mice. Inhibition of MMP-9 or Notch signaling prevented rhTGF-ß1- or rhMMP-9-induced α-SMA and NICD upregulation in MRPECs. UUO kidney of MMP-9 KO mice had less staining of Notch signaling transcription factor Hey-1 in VE-cadherin-positive MRPECs than WT controls. CONCLUSIONS: Our results demonstrate that MMP-9-dependent Notch signaling plays an important role in kidney fibrosis through EndoMT of MRPECs.


Assuntos
Endotélio/patologia , Fibrose/patologia , Nefropatias/patologia , Metaloproteinase 9 da Matriz/metabolismo , Mesoderma/patologia , Receptores Notch/metabolismo , Animais , Endotélio/metabolismo , Fibrose/metabolismo , Humanos , Nefropatias/metabolismo , Masculino , Mesoderma/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Transdução de Sinais , Fator de Crescimento Transformador beta1/metabolismo , Obstrução Ureteral/metabolismo , Obstrução Ureteral/patologia
7.
J Am Soc Nephrol ; 27(5): 1344-60, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-26376858

RESUMO

CD103(+) dendritic cells (DCs) in nonlymphoid organs exhibit two main functions: maintaining tolerance by induction of regulatory T cells and protecting against tissue infection through cross-presentation of foreign antigens to CD8(+) T cells. However, the role of CD103(+) DCs in kidney disease is unknown. In this study, we show that CD103(+) DCs are one of four subpopulations of renal mononuclear phagocytes in normal kidneys. CD103(+) DCs expressed DC-specific surface markers, transcription factors, and growth factor receptors and were found in the kidney cortex but not in the medulla. The number of kidney CD103(+) DCs was significantly higher in mice with adriamycin nephropathy (AN) than in normal mice, and depletion of CD103(+) DCs attenuated kidney injury in AN mice. In vitro, kidney CD103(+) DCs preferentially primed CD8(+) T cells and did not directly induce tubular epithelial cell apoptosis. Adoptive transfer of CD8(+) T cells significantly exacerbated kidney injury in AN SCID mice, whereas depletion of CD103(+) DCs in these mice impaired activation and proliferation of transfused CD8(+) T cells and prevented the exacerbation of kidney injury associated with this transfusion. In conclusion, kidney CD103(+) DCs display a pathogenic role in murine CKD via activation of CD8(+) T cells.


Assuntos
Antibióticos Antineoplásicos/efeitos adversos , Antígenos CD/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Doxorrubicina/efeitos adversos , Cadeias alfa de Integrinas/imunologia , Nefropatias/induzido quimicamente , Nefropatias/imunologia , Animais , Camundongos
8.
BMC Cell Biol ; 17(1): 21, 2016 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-27130612

RESUMO

BACKGROUND: Endothelial-mesenchymal transition (EndoMT) is a major source of myofibroblast formation in kidney fibrosis. Our previous study showed a profibrotic role for matrix metalloproteinase 9 (MMP-9) in kidney fibrosis via induction of epithelial-mesenchymal transition (EMT). Inhibition of MMP-9 activity reduced kidney fibrosis in murine unilateral ureteral obstruction. This study investigated whether MMP-9 also plays a role in EndoMT in human glomerular endothelial cells. RESULTS: TGF-ß1 (10 or 20 ng/ml) induced EndoMT in HKGECs as shown by morphological changes. In addition, VE-cadherin and CD31 were significantly downregulated, whereas α-SMA, vimentin, and N-cadherin were upregulated. RT-PCR revealed that Snail, a known inducer of EMT, was upregulated. The MMP inhibitor GM6001 abrogated TGF-ß1-induced EndoMT. Zymography indicated that MMP-9 was also upregulated in TGF-ß1-treated HKGECs. Recombinant MMP-9 (2 µg/ml) induced EndoMT in HKGECs via Notch signaling, as evidenced by increased formation of the Notch intracellular domain (NICD) and decreased Notch 1. Inhibition of MMP-9 activity by its inhibitor showed a dose-dependent response in preventing TGF-ß1-induced α-SMA and NICD in HKGECs, whereas inhibition of Notch signaling by γ-secretase inhibitor (GSI) blocked rMMP-9-induced EndoMT. CONCLUSIONS: Taken together, our results demonstrate that MMP-9 plays an important role in TGF-ß1-induced EndoMT via upregulation of Notch signaling in HKGECs.


Assuntos
Células Endoteliais/metabolismo , Glomérulos Renais/citologia , Metaloproteinase 9 da Matriz/metabolismo , Mesoderma/citologia , Receptores Notch/metabolismo , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Secretases da Proteína Precursora do Amiloide/metabolismo , Dipeptídeos/farmacologia , Células Endoteliais/efeitos dos fármacos , Humanos , Inibidores de Metaloproteinases de Matriz/farmacologia , Mesoderma/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/farmacologia
9.
J Am Soc Nephrol ; 26(9): 2199-211, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25556172

RESUMO

IL-25 is an important immune regulator that can promote Th2 immune response-dependent immunity, inflammation, and tissue repair in asthma, intestinal infection, and autoimmune diseases. In this study, we examined the effects of IL-25 in renal ischemic/reperfusion injury (IRI). Treating IRI mice with IL-25 significantly improved renal function and reduced renal injury. Furthermore, IL-25 treatment increased the levels of IL-4, IL-5, and IL-13 in serum and kidney and promoted induction of alternatively activated (M2) macrophages in kidney. Notably, IL-25 treatment also increased the frequency of type 2 innate lymphoid cells (ILC2s) and multipotent progenitor type 2 (MPP(type2)) cells in kidney. IL-25-responsive ILC2 and MPP(type2) cells produced greater amounts of Th2 cytokines that associated with the induction of M2 macrophages and suppression of classically activated (M1) macrophages in vitro. Finally, adoptive transfer of ILC2s or MPP(type2) cells not only reduced renal functional and histologic injury in IRI mice but also induced M2 macrophages in kidney. In conclusion, our data identify a mechanism whereby IL-25-elicited ILC2 and MPP(type2) cells regulate macrophage phenotype in kidney and prevent renal IRI.


Assuntos
Injúria Renal Aguda/imunologia , Injúria Renal Aguda/prevenção & controle , Fatores Imunológicos/uso terapêutico , Interleucina-17/uso terapêutico , Linfócitos/efeitos dos fármacos , Células-Tronco Multipotentes/efeitos dos fármacos , Traumatismo por Reperfusão/imunologia , Traumatismo por Reperfusão/prevenção & controle , Injúria Renal Aguda/etiologia , Transferência Adotiva , Animais , Sobrevivência Celular , Células Cultivadas , Imunidade Inata/efeitos dos fármacos , Imunidade Inata/imunologia , Fatores Imunológicos/farmacologia , Interleucina-13/metabolismo , Interleucina-17/farmacologia , Interleucina-4/metabolismo , Interleucina-5/metabolismo , Rim/irrigação sanguínea , Rim/metabolismo , Linfócitos/citologia , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Células-Tronco Multipotentes/citologia , Traumatismo por Reperfusão/complicações , Células Th2/imunologia
10.
J Am Soc Nephrol ; 26(2): 349-63, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25012165

RESUMO

Conventional markers of macrophages (Mфs) and dendritic cells (DCs) lack specificity and often overlap, leading to confusion and controversy regarding the precise function of these cells in kidney and other diseases. This study aimed to identify the phenotype and function of renal mononuclear phagocytes (rMPs) expressing key markers of both Mфs and DCs. F4/80(+)CD11c(+) cells accounted for 45% of total rMPs in normal kidneys and in those from mice with Adriamycin nephropathy (AN). Despite expression of the DC marker CD11c, these double-positive rMPs displayed the features of Mфs, including Mф-like morphology, high expression of CD68, CD204, and CD206, and high phagocytic ability but low antigen-presenting ability. F4/80(+)CD11c(+) cells were found in the cortex but not in the medulla of the kidney. In AN, F4/80(+)CD11c(+) cells displayed an M1 Mф phenotype with high expression of inflammatory mediators and costimulatory factors. Adoptive transfer of F4/80(+)CD11c(+) cells separated from diseased kidney aggravated renal injury in AN mice. Furthermore, adoptive transfer of common progenitors revealed that kidney F4/80(+)CD11c(+) cells were derived predominantly from monocytes, but not from pre-DCs. In conclusion, renal F4/80(+)CD11c(+) cells are a major subset of rMPs and display Mф-like phenotypic and functional characteristics in health and in AN.


Assuntos
Antígenos de Diferenciação/metabolismo , Antígenos CD11/metabolismo , Nefropatias/patologia , Rim/patologia , Macrófagos/patologia , Fagócitos/patologia , Fenótipo , Transferência Adotiva , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Biomarcadores/metabolismo , Modelos Animais de Doenças , Doxorrubicina/efeitos adversos , Técnicas In Vitro , Rim/fisiologia , Nefropatias/induzido quimicamente , Nefropatias/fisiopatologia , Lectinas Tipo C/metabolismo , Macrófagos/imunologia , Macrófagos/fisiologia , Receptor de Manose , Lectinas de Ligação a Manose/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fagócitos/imunologia , Fagócitos/fisiologia , Receptores de Superfície Celular/metabolismo , Receptores Depuradores Classe A/metabolismo
11.
J Cell Sci ; 126(Pt 1): 67-76, 2013 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-23203799

RESUMO

Transforming growth factor ß1 (TGF-ß1) is known to be both anti-inflammatory and profibrotic. Cross-talk between TGF-ß/Smad and Wnt/ß-catenin pathways in epithelial-mesenchymal transition (EMT) suggests a specific role for ß-catenin in profibrotic effects of TGF-ß1. However, no such mechanistic role has been demonstrated for ß-catenin in the anti-inflammatory effects of TGF-ß1. In the present study, we explored the role of ß-catenin in the profibrotic and anti-inflammatory effects of TGF-ß1 by using a cytosolic, but not membrane, ß-catenin knockdown chimera (F-TrCP-Ecad) and the ß-catenin/CBP inhibitor ICG-001. TGF-ß1 induced nuclear Smad3/ß-catenin complex, but not ß-catenin/LEF-1 complex or TOP-flash activity, during EMT of C1.1 (renal tubular epithelial) cells. F-TrCP-Ecad selectively degraded TGF-ß1-induced cytoplasmic ß-catenin and blocked EMT of C1.1 cells. Both F-TrCP-Ecad and ICG-001 blocked TGF-ß1-induced Smad3/ß-catenin and Smad reporter activity in C1.1 cells, suggesting that TGF-ß1-induced EMT depends on ß-catenin binding to Smad3, but not LEF-1 downstream of Smad3, through canonical Wnt. In contrast, in J774 macrophages, the ß-catenin level was low and was not changed by interferon-γ (IFN-γ) or lipopolysaccharide (LPS) with or without TGF-ß1. TGF-ß1 inhibition of LPS-induced TNF-α and IFN-γ-stimulated inducible NO synthase (iNOS) expression was not affected by F-TrCP-Ecad, ICG-001 or by overexpression of wild-type ß-catenin in J774 cells. Inhibition of ß-catenin by either F-TrCP-Ecad or ICG-001 abolished LiCl-induced TOP-flash, but not TGF-ß1-induced Smad reporter, activity in J774 cells. These results demonstrate for the first time that ß-catenin is required as a co-factor of Smad in TGF-ß1-induced EMT of C1.1 epithelial cells, but not in TGF-ß1 inhibition of macrophage activation. Targeting ß-catenin may dissociate the TGF-ß1 profibrotic and anti-inflammatory effects.


Assuntos
Anti-Inflamatórios/farmacologia , Fator 1 de Ligação ao Facilitador Linfoide/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , beta Catenina/metabolismo , Animais , Western Blotting , Linhagem Celular , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Imunoprecipitação , Fator 1 de Ligação ao Facilitador Linfoide/genética , Camundongos , Microscopia de Fluorescência , Ligação Proteica/efeitos dos fármacos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína Smad3/genética , beta Catenina/genética
12.
Kidney Int ; 85(4): 794-806, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24048378

RESUMO

Alternatively activated macrophages (M2) regulate immune responses and ex vivo polarized splenic M2 are able to ameliorate renal injury including models of renal disease, such as adriamycin nephropathy. Whether M2 derived from other organs have similar protective efficacy is unknown. Here, we report adoptively transferred bone marrow M2 macrophages did not improve renal function or reduce renal injury in adriamycin nephropathy, whereas splenic M2 macrophages were protective. Bone marrow and splenic M2 macrophages showed similar regulatory phenotypes and suppressive functions in vitro. Within the inflamed kidney, suppressive phenotypes in bone marrow but not in splenic M2 macrophages, were dramatically reduced. Loss of the suppressive phenotype in bone marrow M2 was related to strong proliferation of bone marrow M2. Bone marrow M2 proliferation in vivo correlated with M-CSF expression by tubular cells in the inflamed kidney. Inhibition of M-CSF in vitro limited bone marrow M2 proliferation and prevented switch of phenotype. Proliferating cells derived from transfused bone marrow M2 were inflammatory rather than regulatory in their phenotype and function. Thus bone marrow in contrast to splenic M2 macrophages do not protect against renal structural and functional injury in murine adriamycin nephropathy. The failed renoprotection of bone marrow M2 is due to the switch of transfused M2 macrophages from a regulatory to an inflammatory phenotype.


Assuntos
Transferência Adotiva , Nefropatias/terapia , Macrófagos/transplante , Baço/imunologia , Animais , Células da Medula Óssea/citologia , Proliferação de Células , Doxorrubicina , Nefropatias/induzido quimicamente , Túbulos Renais/metabolismo , Fator Estimulador de Colônias de Macrófagos/metabolismo , Macrófagos/citologia , Macrófagos/fisiologia , Masculino , Camundongos Endogâmicos BALB C , Fenótipo
13.
Lab Invest ; 93(4): 434-49, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23358111

RESUMO

A pro-fibrotic role of matrix metalloproteinase-9 (MMP-9) in tubular cell epithelial-mesenchymal transition (EMT) is well established in renal fibrosis; however studies from our group and others have demonstrated some previously unrecognized complexity of MMP-9 that has been overlooked in renal fibrosis. Therefore, the aim of this study was to determine the expression pattern, origin and the exact mechanism underlying the contribution of MMP-9 to unilateral ureteral obstruction (UUO), a well-established model of renal fibrosis via MMP-9 inhibition. Renal MMP-9 expression in BALB/c mice with UUO was examined on day 1, 3, 5, 7, 9, 11 and 14. To inhibit MMP-9 activity, MMP-2/9 inhibitor or MMP-9-neutralizing antibody was administered daily for 4 consecutive days from day 0-3, 6-9 or 10-13 and tissues harvested at day 14. In UUO, there was a bi-phasic early- and late-stage upregulation of MMP-9 activity. Interestingly, tubular epithelial cells (TECs) were the predominant source of MMP-9 during early stage, whereas TECs, macrophages and myofibroblasts produced MMP-9 during late-stage UUO. Early- and late-stage inhibition of MMP-9 in UUO mice significantly reduced tubular cell EMT and renal fibrosis. Moreover, MMP-9 inhibition caused a significant reduction in MMP-9-cleaved osteopontin and macrophage infiltration in UUO kidney. Our in vitro study showed MMP-9-cleaved osteopontin enhanced macrophage transwell migration and MMP-9 of both primary TEC and macrophage induced tubular cell EMT. In summary, our result suggests that MMP-9 of both TEC and macrophage origin may directly or indirectly contribute to the pathogenesis of renal fibrosis via osteopontin cleavage, which, in turn further recruit macrophage and induce tubular cell EMT. Our study also highlights the time dependency of its expression and the potential of stage-specific inhibition strategy against renal fibrosis.


Assuntos
Nefropatias/imunologia , Rim/patologia , Metaloproteinase 9 da Matriz/metabolismo , Osteopontina/metabolismo , Obstrução Ureteral/metabolismo , Animais , Movimento Celular , Células Cultivadas , Células Epiteliais/fisiologia , Transição Epitelial-Mesenquimal , Fibrose , Rim/metabolismo , Nefropatias/metabolismo , Nefropatias/patologia , Macrófagos/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Fatores de Transcrição da Família Snail , Fatores de Transcrição/metabolismo , beta Catenina/metabolismo
14.
Kidney Int ; 84(4): 745-55, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23636175

RESUMO

Two types of alternatively activated macrophages, M(2a) induced by IL-4/IL-13 and M(2c) by IL-10/TGF-ß, exhibit anti-inflammatory functions in vitro and protect against renal injury in vivo. Since their relative therapeutic efficacy is unclear, we compared the effects of these two macrophage subsets in murine adriamycin nephrosis. Both subsets significantly reduced renal inflammation and renal injury; however, M(2c) macrophages more effectively reduced glomerulosclerosis, tubular atrophy, interstitial expansion, and proteinuria than M(2a) macrophages. The M(2c) macrophages were also more effective than M(2a) in reduction of macrophage and CD4(+) T-cell infiltration in kidney. Moreover, nephrotic mice treated with M(2c) had a greater reduction in renal fibrosis than those treated with M(2a). M(2c) but not M(2a) macrophages induced regulatory T cells (Tregs) from CD4(+)CD25(-) T cells in vitro, and increased Treg numbers in local draining lymph nodes of nephrotic mice. To determine whether the greater protection with M(2c) was due to their capability to induce Tregs, the Tregs were depleted by PC61 antibody in nephrotic mice treated with M(2a) or M(2c). Treg depletion diminished the superior effects of M(2c) compared to M(2a) in protection against renal injury, inflammatory infiltrates, and renal fibrosis. Thus, M(2c) are more potent than M(2a) macrophages in protecting against renal injury due to their ability to induce Tregs.


Assuntos
Transferência Adotiva/métodos , Terapia Baseada em Transplante de Células e Tecidos/métodos , Macrófagos/classificação , Macrófagos/fisiologia , Insuficiência Renal Crônica/terapia , Animais , Diferenciação Celular/fisiologia , Células Cultivadas , Gerenciamento Clínico , Modelos Animais de Doenças , Doxorrubicina/efeitos adversos , Técnicas In Vitro , Macrófagos/citologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Nefrose/induzido quimicamente , Nefrose/fisiopatologia , Nefrose/prevenção & controle , Fenótipo , Insuficiência Renal Crônica/patologia , Insuficiência Renal Crônica/fisiopatologia , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/fisiologia
15.
Eur J Immunol ; 42(9): 2441-51, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22684996

RESUMO

CD39 is an ecto-enzyme that degrades extracellular nucleotides, such as ATP, and is highly expressed on by the vasculature and circulating cells including Foxp3+ regulatory T (Treg) cells. To study the role of purinergic regulation in renal disease, we used the adriamycin nephropathy (AN) mouse model of chronic renal injury, using human CD39-transgenic (hCD39Tg) and wild-type (WT) BALB/c mice. Effects of CD39 expression by Treg cells were assessed in AN by adoptive transfer of CD4(+) CD25(+) and CD4(+) CD25(-) T cells isolated from hCD39Tg and WT mice. hCD39Tg mice were protected from renal injury in AN with decreased urinary protein and serum creatinine, and significantly less renal injury compared with WT mice. While WT CD25(+) and hCD39Tg CD25(-) T cells conferred some protection against AN, hCD39Tg CD25(+) Treg cells offered greater protection. In vitro studies showed direct pro-apoptotic effects of ATP on renal tubular cells. In conclusion, hCD39 expressed by circulating leukocytes and intrinsic renal cells limits innate AN injury. Specifically, CD39 expression by Treg cells contributes to its protective role in renal injury. These findings suggest that extracellular nucleotides mediate AN kidney injury and that CD39, expressed by Treg cells and other cells, is protective in this model.


Assuntos
Antígenos CD/imunologia , Antígenos CD/metabolismo , Apirase/imunologia , Apirase/metabolismo , Nefropatias/imunologia , Linfócitos T Reguladores/imunologia , Trifosfato de Adenosina/imunologia , Trifosfato de Adenosina/metabolismo , Animais , Apoptose/imunologia , Creatinina/imunologia , Creatinina/metabolismo , Modelos Animais de Doenças , Doxorrubicina , Humanos , Nefropatias/induzido quimicamente , Nefropatias/metabolismo , Túbulos Renais/imunologia , Túbulos Renais/metabolismo , Leucócitos/imunologia , Leucócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Reguladores/metabolismo
16.
JHEP Rep ; 5(10): 100837, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37691688

RESUMO

Background and Aims: Although type 2 innate lymphoid cells (ILC2s) were originally found to be liver-resident lymphocytes, the role and importance of ILC2 in liver injury remains poorly understood. In the current study, we sought to determine whether ILC2 is an important regulator of hepatic ischaemia/reperfusion injury (IRI). Methods: ILC2-deficient mice (ICOS-T or NSG) and genetically modified ILC2s were used to investigate the role of ILC2s in murine hepatic IRI. Interactions between ILC2s and eosinophils or macrophages were studied in coculture. The role of human ILC2s was assessed in an immunocompromised mouse model of hepatic IRI. Results: Administration of IL-33 prevented hepatic IRI in association with reduction of neutrophil infiltration and inflammatory mediators in the liver. IL-33-treated mice had elevated numbers of ILC2s, eosinophils, and regulatory T cells. Eosinophils, but not regulatory T cells, were required for IL-33-mediated hepatoprotection in IRI mice. Depletion of ILC2s substantially abolished the protective effect of IL-33 in hepatic IRI, indicating that ILC2s play critical roles in IL-33-mediated liver protection. Adoptive transfer of ex vivo-expanded ILC2s improved liver function and attenuated histologic damage in mice subjected to IRI. Mechanistic studies combining genetic and adoptive transfer approaches identified a protective role of ILC2s through promoting IL-13-dependent induction of anti-inflammatory macrophages and IL-5-dependent elevation of eosinophils in IRI. Furthermore, in vivo expansion of human ILC2s by IL-33 or transfer of ex vivo-expanded human ILC2s ameliorated hepatic IRI in an immunocompromised mouse model of hepatic IRI. Conclusions: This study provides insight into the mechanisms of ILC2-mediated liver protection that could serve as therapeutic targets to treat acute liver injury. Impact and Implications: We report that type 2 innate lymphoid cells (ILC2s) are important regulators in a mouse model of liver ischaemia/reperfusion injury (IRI). Through manipulation of macrophage and eosinophil phenotypes, ILC2s mitigate liver inflammation and injury during liver IRI. We propose that ILC2s have the potential to serve as a therapeutic tool for protecting against acute liver injury and lay the foundation for translation of ILC2 therapy to human liver disease.

17.
Kidney Int ; 81(9): 892-902, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22318423

RESUMO

Plasmacytoid dendritic cells play important roles in inducing immune tolerance, preventing allograft rejection, and regulating immune responses in both autoimmune disease and graft-versus-host disease. In order to evaluate a possible protective effect of plasmacytoid dendritic cells against renal inflammation and injury, we purified these cells from mouse spleens and adoptively transferred lipopolysaccharide (LPS)-treated cells, modified ex vivo, into mice with adriamycin nephropathy. These LPS-treated cells localized to the kidney cortex and the lymph nodes draining the kidney, and protected the kidney from injury during adriamycin nephropathy. Glomerulosclerosis, tubular atrophy, interstitial expansion, proteinuria, and creatinine clearance were significantly reduced in mice with adriamycin nephropathy subsequently treated with LPS-activated plasmacytoid dendritic cells as compared to the kidney injury in mice given naive plasmacytoid dendritic cells. In addition, LPS-pretreated cells, but not naive plasmacytoid dendritic cells, convert CD4+CD25- T cells into Foxp3+ regulatory T cells and suppress the proinflammatory cytokine production of endogenous renal macrophages. This may explain their ability to protect against renal injury in adriamycin nephropathy.


Assuntos
Transferência Adotiva , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/transplante , Nefropatias/prevenção & controle , Rim/imunologia , Lipopolissacarídeos/farmacologia , Animais , Atrofia , Biomarcadores/sangue , Linfócitos T CD4-Positivos/imunologia , Células Cultivadas , Doença Crônica , Técnicas de Cocultura , Creatinina/sangue , Citocinas/metabolismo , Células Dendríticas/imunologia , Modelos Animais de Doenças , Doxorrubicina , Fatores de Transcrição Forkhead/metabolismo , Glomerulonefrite/imunologia , Glomerulonefrite/prevenção & controle , Mediadores da Inflamação/metabolismo , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Rim/metabolismo , Rim/patologia , Rim/fisiopatologia , Nefropatias/sangue , Nefropatias/induzido quimicamente , Nefropatias/imunologia , Nefropatias/patologia , Nefropatias/fisiopatologia , Linfonodos/imunologia , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Nefrite Intersticial/imunologia , Nefrite Intersticial/prevenção & controle , Fenótipo , Proteinúria/imunologia , Proteinúria/prevenção & controle , Linfócitos T Reguladores/imunologia , Receptor 4 Toll-Like/metabolismo
18.
J Am Soc Nephrol ; 22(7): 1229-39, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21719780

RESUMO

The kidney contains receptors for the cytokine IL-25, but the effects of IL-25 in CKD are unknown. Here, we induced adriamycin nephropathy in both BALB/c mice and severe combined immunodeficient (SCID) mice, and we injected IL-25 for 7 consecutive days starting at day 5 after adriamycin administration. BALB/c mice treated with IL-25 had less glomerulosclerosis, tubular atrophy, interstitial expansion, and proteinuria than control mice at day 28. IL-25 increased the levels of IL-4 and IL-13 in serum, kidney, renal draining lymph nodes, and CD4+ lymphocytes. IL-25 also directly suppressed effector macrophages in vitro and in vivo and induced alternatively activated (M2) macrophages in vivo. However, in SCID mice and in BALB/c mice treated with IL-4/13-neutralizing antibody, IL-25 failed to protect against renal injury and did not induce M2. In conclusion, IL-25 protects against renal injury in adriamycin nephropathy in mice by, at least in part, inducing Th2 immune responses.


Assuntos
Glomerulosclerose Segmentar e Focal/metabolismo , Interleucinas/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Proteinúria/metabolismo , Células Th2/efeitos dos fármacos , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/prevenção & controle , Animais , Antibióticos Antineoplásicos , Doxorrubicina , Glomerulosclerose Segmentar e Focal/induzido quimicamente , Glomerulosclerose Segmentar e Focal/tratamento farmacológico , Glomerulosclerose Segmentar e Focal/imunologia , Interleucina-13/metabolismo , Interleucina-4/metabolismo , Interleucinas/farmacologia , Interleucinas/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos SCID , Proteinúria/induzido quimicamente , Proteinúria/tratamento farmacológico , Proteinúria/imunologia
19.
Am J Pathol ; 176(3): 1256-70, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20075196

RESUMO

As a rich source of pro-fibrogenic growth factors and matrix metalloproteinases (MMPs), macrophages are well-placed to play an important role in renal fibrosis. However, the exact underlying mechanisms and the extent of macrophage involvement are unclear. Tubular cell epithelial-mesenchymal transition (EMT) is an important contributor to renal fibrosis and MMPs to induction of tubular cell EMT. The aim of this study was to investigate the contribution of macrophages and MMPs to induction of tubular cell EMT. The murine C1.1 tubular epithelial cell line and primary tubular epithelial cells were cultured in activated macrophage-conditioned medium (AMCM) derived from lipopolysaccharide-activated J774 macrophages. MMP-9, but not MMP-2 activity was detected in AMCM. AMCM-induced tubular cell EMT in C1.1 cells was inhibited by broad-spectrum MMP inhibitor (GM6001), MMP-2/9 inhibitor, and in AMCM after MMP-9 removal by monoclonal Ab against MMP-9. AMCM-induced EMT in primary tubular epithelial cells was inhibited by MMP-2/9 inhibitor. MMP-9 induced tubular cell EMT in both C1.1 cells and primary tubular epithelial cells. Furthermore, MMP-9 induced tubular cell EMT in C1.1 cells to an extent similar to transforming growth factor-beta. Transforming growth factor-beta-induced tubular cell EMT in C1.1 cells was inhibited by MMP-2/9 inhibitor. Our in vitro study provides evidence that MMPs, specifically MMP-9, secreted by effector macrophages can induce tubular cell EMT and thereby contribute to renal fibrosis.


Assuntos
Células Epiteliais/enzimologia , Células Epiteliais/patologia , Epitélio/enzimologia , Túbulos Renais/patologia , Macrófagos/enzimologia , Metaloproteinase 9 da Matriz/metabolismo , Mesoderma/enzimologia , Actinas/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Meios de Cultivo Condicionados/farmacologia , Dipeptídeos/farmacologia , Células Epiteliais/efeitos dos fármacos , Epitélio/patologia , Fibrose/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Lipopolissacarídeos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Metaloproteinase 2 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz , Mesoderma/efeitos dos fármacos , Mesoderma/patologia , Camundongos , Camundongos Endogâmicos BALB C , Fenótipo , Transporte Proteico/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Obstrução Ureteral/enzimologia , Obstrução Ureteral/patologia
20.
Nephron Exp Nephrol ; 118(4): e87-99, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21311199

RESUMO

BACKGROUND: Alternatively activated macrophages (M2 macrophages) are able to reduce renal injury in murine adriamycin nephropathy. However, the effect of M2 macrophages in other renal diseases such as diabetic nephropathy remains unknown. METHODS: Macrophages were separated from splenocytes and polarized with IL-4 and IL-13 into a protective phenotype. Mice underwent adoptive transfer with M2 macrophages, and then diabetes was induced by tail vein injection with streptozotocin (STZ). Blood glucose levels were monitored daily. Mice were sacrificed at week 10 after STZ. Renal function and histopathological injury were assessed quantitatively. RESULTS: Transfused M2 macrophages accumulated progressively in kidneys for up to 10 weeks after STZ. Kidneys from diabetic mice transfused with M2 macrophages had less tubular atrophy, glomerular hypertrophy and interstitial expansion than did control diabetic mice. M2 macrophages suppressed the development of interstitial fibrosis. In addition, the degree of pancreatic islet injury, as assessed by insulin staining, haemoglobin A1c and blood glucose was reduced after transfusion of M2 macrophages. In vivo, activation of kidney endogenous macrophage cytokine expression was inhibited by M2 macrophages. CONCLUSION: Our findings show that M2 macrophages can protect against islet and renal injury in streptozotocin-induced diabetes, providing a potential therapeutic strategy for diabetes and diabetic nephropathy.


Assuntos
Diabetes Mellitus Experimental/terapia , Nefropatias/terapia , Transfusão de Leucócitos , Macrófagos/transplante , Pancreatopatias/terapia , Animais , Glicemia/metabolismo , Células Cultivadas , Diabetes Mellitus Experimental/sangue , Nefropatias/sangue , Nefropatias/patologia , Transfusão de Leucócitos/métodos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pancreatopatias/sangue , Pancreatopatias/patologia , Baço/citologia , Baço/transplante
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