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1.
Int J Mol Sci ; 25(10)2024 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-38791159

RESUMO

Glomerulonephritis (GN) is characterized by podocyte injury or glomerular filtration dysfunction, which results in proteinuria and eventual loss of kidney function. Progress in studying the mechanism of GN, and developing an effective therapy, has been limited by the absence of suitable in vitro models that can closely recapitulate human physiological responses. We developed a microfluidic glomerulus-on-a-chip device that can recapitulate the physiological environment to construct a functional filtration barrier, with which we investigated biological changes in podocytes and dynamic alterations in the permeability of the glomerular filtration barrier (GFB) on a chip. We also evaluated the potential of GN-mimicking devices as a model for predicting responses to human GN. Glomerular endothelial cells and podocytes successfully formed intact monolayers on opposite sides of the membrane in our chip device. Permselectivity analysis confirmed that the chip was constituted by a functional GFB that could accurately perform differential clearance of albumin and dextran. Reduction in cell viability resulting from damage was observed in all serum-induced GN models. The expression of podocyte-specific marker WT1 was also decreased. Albumin permeability was increased in most models of serum-induced IgA nephropathy (IgAN) and membranous nephropathy (MN). However, sera from patients with minimal change disease (MCD) or lupus nephritis (LN) did not induce a loss of permeability. This glomerulus-on-a-chip system may provide a platform of glomerular cell culture for in vitro GFB in formation of a functional three-dimensional glomerular structure. Establishing a disease model of GN on a chip could accelerate our understanding of pathophysiological mechanisms of glomerulopathy.


Assuntos
Glomerulonefrite , Glomérulos Renais , Dispositivos Lab-On-A-Chip , Podócitos , Humanos , Podócitos/metabolismo , Podócitos/patologia , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Glomerulonefrite/metabolismo , Glomerulonefrite/fisiopatologia , Glomerulonefrite/patologia , Barreira de Filtração Glomerular/metabolismo , Glomerulonefrite Membranosa/metabolismo , Glomerulonefrite Membranosa/patologia , Glomerulonefrite Membranosa/fisiopatologia , Glomerulonefrite por IGA/metabolismo , Glomerulonefrite por IGA/patologia , Glomerulonefrite por IGA/fisiopatologia , Permeabilidade , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Nefrite Lúpica/metabolismo , Nefrite Lúpica/patologia , Nefrite Lúpica/fisiopatologia , Sobrevivência Celular , Nefrose Lipoide/metabolismo , Nefrose Lipoide/patologia , Nefrose Lipoide/fisiopatologia
2.
Int J Mol Sci ; 23(22)2022 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-36430531

RESUMO

Renal fibrosis, the final pathway of chronic kidney disease, is caused by genetic and epigenetic mechanisms. Although DNA methylation has drawn attention as a developing mechanism of renal fibrosis, its contribution to renal fibrosis has not been clarified. To address this issue, the effect of zebularine, a DNA methyltransferase inhibitor, on renal inflammation and fibrosis in the murine unilateral ureteral obstruction (UUO) model was analyzed. Zebularine significantly attenuated renal tubulointerstitial fibrosis and inflammation. Zebularine decreased trichrome, α-smooth muscle actin, collagen IV, and transforming growth factor-ß1 staining by 56.2%. 21.3%, 30.3%, and 29.9%, respectively, at 3 days, and by 54.6%, 41.9%, 45.9%, and 61.7%, respectively, at 7 days after UUO. Zebularine downregulated mRNA expression levels of matrix metalloproteinase (MMP)-2, MMP-9, fibronectin, and Snail1 by 48.6%. 71.4%, 31.8%, and 42.4%, respectively, at 7 days after UUO. Zebularine also suppressed the activation of nuclear factor-κB (NF-κB) and the expression of pro-inflammatory cytokines, including tumor necrosis factor-α, interleukin (IL)-1ß, and IL-6, by 69.8%, 74.9%, and 69.6%, respectively, in obstructed kidneys. Furthermore, inhibiting DNA methyltransferase buttressed the nuclear expression of nuclear factor (erythroid-derived 2)-like factor 2, which upregulated downstream effectors such as catalase (1.838-fold increase at 7 days, p < 0.01), superoxide dismutase 1 (1.494-fold increase at 7 days, p < 0.05), and NAD(P)H: quinone oxidoreduate-1 (1.376-fold increase at 7 days, p < 0.05) in obstructed kidneys. Collectively, these findings suggest that inhibiting DNA methylation restores the disrupted balance between pro-inflammatory and anti-inflammatory pathways to alleviate renal inflammation and fibrosis. Therefore, these results highlight the possibility of DNA methyltransferases as therapeutic targets for treating renal inflammation and fibrosis.


Assuntos
Nefrite , Insuficiência Renal Crônica , Obstrução Ureteral , Camundongos , Animais , Fibrose , Nefrite/patologia , Obstrução Ureteral/complicações , Obstrução Ureteral/tratamento farmacológico , Obstrução Ureteral/genética , Inflamação/patologia , Insuficiência Renal Crônica/complicações , Metilases de Modificação do DNA , DNA/uso terapêutico
4.
Sci Rep ; 9(1): 3021, 2019 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-30816234

RESUMO

Prolonged hypokalemia induces a decrease of urinary concentrating ability via down-regulation of aquaporin 2 (AQP2); however, the precise mechanisms remain unknown. To investigate the role of autophagy in the degradation of AQP2, we generated the principal cell-specific Atg7 deletion (Atg7Δpc) mice. In hypokalemic Atg7-floxed (Atg7f/f) mice, huge irregular shaped LC3-positive autophagic vacuoles accumulated mainly in inner medullary collecting duct (IMCD) cells. Total- and pS261-AQP2 were redistributed from apical and subapical domains into these vacuoles, which were not co-localized with RAB9. However, in the IMCD cells of hypokalemic Atg7Δpc mice, these canonical autophagic vacuoles were markedly reduced, whereas numerous small regular shaped LC3-negative/RAB9-positive non-canonical autophagic vacuoles were observed along with diffusely distributed total- and pS261-AQP2 in the cytoplasm. The immunoreactivity of pS256-AQP2 in the apical membrane of IMCD cells was markedly decreased, and no redistribution was observed in both hypokalemic Atg7f/f and Atg7Δpc mice. These findings suggest that AQP2 down regulation in hypokalemia was induced by reduced phosphorylation of AQP2, resulting in a reduction of apical plasma labeling of pS256-AQP2 and degradation of total- and pS261-AQP2 via an LC3/ATG7-dependent canonical autophagy pathway.


Assuntos
Aquaporina 2/metabolismo , Proteína 7 Relacionada à Autofagia/metabolismo , Autofagia/fisiologia , Hipopotassemia/metabolismo , Animais , Regulação para Baixo/fisiologia , Rim/metabolismo , Masculino , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Fosforilação/fisiologia , Proteínas rab de Ligação ao GTP/metabolismo
5.
Cell Death Dis ; 10(2): 78, 2019 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-30692509

RESUMO

Renal fibrosis is the final common pathway of various renal injuries and it leads to chronic kidney disease. Autophagy is a cellular process of degradation of damaged cytoplasmic components and regulates cell death and proliferation. Cellular response during unilateral ureteral obstruction (UUO) is tubular segment specific. Thus the role of autophagy on renal tubulointerstitial fibrosis (TIF) after UUO may be different according to segment of nephron. The role of autophagy during UUO remains unclear especially in distal tubules. In this study, we investigated the role of autophagy in distal tubules on renal TIF using conditional knockout mice in which Atg7 was genetically ablated specifically in distal tubular epithelial cell (TEC). In green fluorescent protein (GFP)-LC3 transgenic mice, GFP-LC3 puncta was highly expressed in distal tubular cells of the obstructed kidneys after UUO. Genetic deletion of Atg7 specifically in distal TEC increased renal tubulointerstial fibrosis and epithelial-mesenchymal transition-like phenotype change after UUO through Smad4-dependent transforming growth factor (TGF)-ß pathway. Distal tubule-specific autophagy-deficient mice increased the accumulation of damaged mitochondria and SQSTM1/p62-positive aggregates in the obstructed kidney and resulted in increased expression of NLRP3 inflammasome, interleukin (IL) 1-ß and caspase-1. Distal TEC-specific Atg7 deletion enhanced apoptosis of TECs after UUO. In summary, our data showed that autophagy in distal TEC plays a protective role in development of renal tubulointerstial fibrosis through regulating the expression of TGF-ß an IL1-ß after UUO.


Assuntos
Fibrose/genética , Inflamassomos/metabolismo , Rim/patologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Autofagia , Camundongos , Camundongos Knockout , Transdução de Sinais
6.
Biochem Biophys Res Commun ; 508(3): 965-972, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30545632

RESUMO

Renal fibrosis is the final common pathway of various renal injuries and it leads to chronic kidney disease. Recent studies reported that FOXD1-lineage pericyte plays a critical role in tubulointerstitial fibrosis (TIF). However the regulatory mechanisms remain unclear. Autophagy is a cellular process of degradation of damaged cytoplasmic components that regulates cell death and proliferation. To investigate the role of autophagy in FOXD1-lineage pericytes on renal TIF, we generated the FOXD1-lineage stromal cell-specific Atg7 deletion (Atg7△FOXD1) mice. FOXD1-lineage stromal cell-specific Atg7 deletion enhanced renal TIF through Smad-dependent transforming growth factor (TGF)-ß signaling after unilateral ureteral obstruction (UUO). FOXD1-lineage stromal cell-specific Atg7 deletion increased the accumulation of interstitial myofibroblasts and enhanced the differentiation of pericytes into myofibroblasts after UUO. Peritubular capillary rarefaction was accelerated in Atg7△FOXD1 mice after UUO. Atg7△FOXD1 mice increased the accumulation of SQSTM1/p62-positive aggregates in the obstructed kidney and resulted in increased expression of NLRP3 inflammasome, interleukin (IL) 1-ß and caspase-1 signaling pathway, which enhanced apoptosis of interstitial cells after UUO. In summary, our data showed that autophagy in FOXD1-lineage stromal cells plays a protective role in renal TIF through regulating the Smad4 dependent TGF-ß an NLRP3 inflammasome signaling pathway.


Assuntos
Autofagia , Fatores de Transcrição Forkhead/análise , Inflamassomos/metabolismo , Rim/patologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Apoptose , Proteína 7 Relacionada à Autofagia/genética , Diferenciação Celular , Linhagem da Célula , Fibrose , Fatores de Transcrição Forkhead/genética , Rim/citologia , Rim/metabolismo , Masculino , Camundongos , Camundongos Knockout , Miofibroblastos/citologia , Pericitos/citologia , Receptores do Fator de Crescimento Derivado de Plaquetas/análise , Transdução de Sinais , Proteínas Smad/metabolismo , Células Estromais/química , Obstrução Ureteral/complicações
7.
Korean J Intern Med ; 33(4): 774-782, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-28602064

RESUMO

Background/Aims: Mind bomb-1 (Mib1) encodes an E3 ubiquitin ligase, which is required for the initiation of Notch signaling. Recently, it was demonstrated that the renal collecting duct plays an important role in renal fibrosis. Here, we investigated the role of Notch signaling in renal fibrosis using conditional knockout mice with the specific ablation of Mib1 in renal collecting duct principal cells. METHODS: Mib1-floxed mice (Mib1f/f) were crossed with aquaporin 2 (AQP2)-Cre mice in order to generate principal cell-specific Mib1 knockout mice (Mib1f/f :AQP2-Cre+). Unilateral ureteral obstruction (UUO) was performed, and mice were sacrificed 7 days after UUO. RESULTS: After performing the UUO, renal tubulointerstitial fibrosis and the expression of transforming growth factor ß were markedly enhanced in the obstructed kidneys of Mib1f/f mice compared with the sham-operated kidney of Mib1f/f mice. These changes were shown to be even more pronounced in the obstructed kidneys of Mib1f/f :AQP2-Cre+ mice than in those of the Mib1f/f mice . Furthermore, the number of TUNNEL-positive cells in renal collecting duct was higher in the obstructed kidneys of Mib1f/f :AQP2-Cre+ mice than in the kidneys of Mib1f/f mice. Conclusions: Notch signaling in the renal collecting duct plays an important role in the regulation of renal tubulointerstitial fibrosis and apoptosis after UUO.


Assuntos
Aquaporina 2 , Receptores Notch , Obstrução Ureteral , Animais , Aquaporina 2/metabolismo , Fibrose , Rim , Nefropatias/metabolismo , Masculino , Camundongos , Receptores Notch/metabolismo , Transdução de Sinais , Obstrução Ureteral/metabolismo
8.
Korean J Intern Med ; 33(4): 763-773, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-28192889

RESUMO

Background/Aims: This study was designed to investigate the roles of aristolochic acid I (AA-I) and hypokalemia in acute aristolochic acid nephropathy (AAN). METHODS: After an adaptation period (1 week), a total of 40 C57BL/6 mice (male, 8 weeks old) were divided into four groups: I (control group), II (low potassium [K] diet), III (normal K diet with administration of AA-I [10 mg/kg weight]), and IV (low K diet with AA-I). After collecting 24 hours of urine at 2 weeks, the mice were sacrificed, and their blood and kidneys were obtained to perform immunochemical staining and/or Western blot analysis. RESULTS: Proteinuria, glycosuria, and increased fractional excretion of sodium and K were prominent in groups III and IV (p < 0.05). Diffuse swelling and poor staining of collecting duct epithelial cells were evident in the medullas of group II. Typical lesions of toxic acute tubular injury were prominent in the cortices of groups III and IV. Α-Smooth muscle actin (α-SMA) was higher in the cortices of the mice in groups III and IV versus group II (p < 0.05), and higher in the medullas of group IV than groups I and III (p < 0.05). E-cadherin was higher in the cortices of groups III and IV compared to group I (p < 0.05). The F4/80 value was higher in the cortices and medullas of groups II, III, and IV compared to group I (p < 0.05), particularly in the case of group II. Conclusions: AA-I can induce acquired Fanconi syndrome in the acute stage of AAN. Macrophages appear to play a key role in the pathogenesis of AAN and hypokalemic nephropathy. It remains uncertain whether hypokalemia plays any role in AAN and hypokalemia.


Assuntos
Ácidos Aristolóquicos , Hipopotassemia , Nefropatias , Túbulos Renais , Animais , Ácidos Aristolóquicos/farmacologia , Modelos Animais de Doenças , Túbulos Renais/efeitos dos fármacos , Túbulos Renais/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , República da Coreia
9.
Sci Rep ; 6: 31931, 2016 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-27550469

RESUMO

Renal tubulointerstitial fibrosis (TIF) is the final pathway of various renal injuries that result in chronic kidney disease. The mammalian Hippo-Salvador signaling pathway has been implicated in the regulation of cell proliferation, cell death, tissue regeneration, and tumorigenesis. Here, we report that the Hippo-Salvador pathway plays a role in disease development in patients with TIF and in a mouse model of TIF. Mice with tubular epithelial cell (TEC)-specific deletions of Sav1 (Salvador homolog 1) exhibited aggravated renal TIF, enhanced epithelial-mesenchymal transition-like phenotypic changes, apoptosis, and proliferation after unilateral ureteral obstruction (UUO). Moreover, Sav1 depletion in TECs increased transforming growth factor (TGF)-ß and activated ß-catenin expression after UUO, which likely accounts for the abovementioned enhanced TEC fibrotic phenotype. In addition, TAZ (transcriptional coactivator with PDZ-binding motif), a major downstream effector of the Hippo pathway, was significantly activated in Sav1-knockout mice in vivo. An in vitro study showed that TAZ directly regulates TGF-ß and TGF-ß receptor II expression. Collectively, our data indicate that the Hippo-Salvador pathway plays a role in the pathogenesis of TIF and that regulating this pathway may be a therapeutic strategy for reducing TIF.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Nefropatias/metabolismo , Túbulos Renais/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Aciltransferases , Animais , Proteínas de Ciclo Celular/genética , Modelos Animais de Doenças , Transição Epitelial-Mesenquimal , Fibrose , Deleção de Genes , Regulação da Expressão Gênica , Via de Sinalização Hippo , Humanos , Nefropatias/genética , Nefropatias/patologia , Túbulos Renais/patologia , Camundongos , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/metabolismo , beta Catenina/metabolismo
10.
Histochem Cell Biol ; 146(1): 1-12, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27091563

RESUMO

A new intermediate type of Henle's loop has been reported that it extends into the inner medulla and turns within the first millimeter beyond the outer medulla. This study aimed to identify the descending thin limb (DTL) of the intermediate loop in the adult C57Bl/6 mouse kidney using aquaporin 1 (AQP1) and urea transporter A2 (UT-A2) antibodies. In the upper part of the inner stripe of the outer medulla (ISOM), AQP1 was expressed strongly in the DTL with type II epithelium of the long loop, but not in type I epithelium of the short loop. The DTL of the intermediate loop exhibited weak AQP1 immunoreactivity. UT-A2 immunoreactivity was not observed in the upper part of any DTL type. AQP1 expression was similar in the upper and middle parts of the ISOM. UT-A2 expression was variable, being expressed strongly in the DTL with type I epithelium of the short loop, but not in type II epithelium of the long loop. In the innermost part of the ISOM, AQP1 was expressed only in type III epithelium of the long loop. UT-A2-positive and UT-A2-negative cells were intermingled in type I epithelium of the intermediate loop, but were not observed in type III epithelium of the long loop. UT-A2-positive DTLs of the intermediate loop extended into the UT-A2/AQP1-negative type I epithelium in the initial part of the inner medulla. These results demonstrate that the DTL of the intermediate loop is composed of type I epithelium and expresses both AQP1 and UT-A2. The functional role of the DTL of the intermediate loop may be distinct from the short or long loops.


Assuntos
Aquaporina 1/metabolismo , Medula Renal/metabolismo , Rim/metabolismo , Alça do Néfron/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Animais , Aquaporina 1/análise , Rim/química , Medula Renal/química , Alça do Néfron/química , Masculino , Proteínas de Membrana Transportadoras/análise , Camundongos , Camundongos Endogâmicos C57BL , Transportadores de Ureia
11.
Histochem Cell Biol ; 145(1): 17-24, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26496924

RESUMO

The mammalian renal collecting duct consists of principal cells (PCs) and intercalated cells (ICs). Both PCs and ICs are involved in potassium (K(+)) homeostasis, PCs through their role in K(+) secretion and ICs through their ability to facilitate K(+) resorption. We previously hypothesized that PCs may differentiate into ICs upon K(+) depletion. However, no direct evidence has yet been obtained to conclusively demonstrate that PCs differentiate into ICs in response to K(+) depletion. Here, we present direct evidence for the differentiation of PCs into ICs by cell lineage tracing using aquaporin 2 (AQP2)-Cre mice and R26R-EYFP transgenic mice. In control mice, AQP2-EYFP(+) cells exhibited mainly a PC phenotype (AQP2-positive/H(+)-ATPase-negative). Interestingly, some AQP2-EYFP(+) cells exhibited an IC phenotype (H(+)-ATPase-positive/AQP2-negative); these cells accounted for 1.7 %. After K(+) depletion, the proportion of AQP2-EYFP(+) cells with an IC phenotype was increased to 4.1 %. Furthermore, some AQP2-EYFP(+) cells exhibited a "null cell" phenotype (AQP2-negative/H(+)-ATPase-negative) after K(+) depletion. Collectively, our data demonstrate that AQP2-labeled cells can differentiate into ICs, as well as null cells, in response to K(+) depletion. This finding indicates that some of AQP2-labeled cells possess properties of progenitor cells and that they can differentiate into ICs in the adult mouse kidney.


Assuntos
Aquaporina 2/genética , Células Epiteliais/citologia , Túbulos Renais Coletores/citologia , Potássio/metabolismo , Animais , Proteínas de Bactérias/genética , Diferenciação Celular , Linhagem da Célula , Células Cultivadas , Proteínas Luminescentes/genética , Camundongos , Camundongos Knockout
13.
PLoS One ; 10(5): e0127429, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25993027

RESUMO

The homeobox transcription factor Prox1 is critical to the development of many embryonic organs and tissues, although current understanding of its expression in the developing renal medulla is limited. We examined the functional role of Prox1 during mouse kidney development with particular emphasis on the developing loop of Henle. Our data show that Prox1 is expressed in the transdifferentiating region from the NKCC2-positive thick ascending limb, into the CLC-K1-positive ascending thin limb of Henle's loop beginning at embryonic day 18. From 1 to 14 days of age, Prox1-positive cells gradually disappeared from the papillary tip, and remained in the initial part of inner medulla after 21 days. In this transforming area, no Prox1 was observed in cells undergoing apoptosis but was expressed strongly in the remaining cells, which differentiated into ascending thin limb epithelial cells. In vitro and in vivo approaches showed that Prox1 expression increases where the osmolality is near optimal range, but decreases at below- or above-optimal ranges. Renal hypoosmolality induced by furosemide (NKCC2 inhibitor) inhibited Prox1 expression and delayed maturation of the ascending limb of Henle's loop. Together, these studies suggest that Prox1 appears to be a critical stage specific regulator of specifying ascending thin limb cell fate and that its expression is regulated by osmolality.


Assuntos
Proteínas de Homeodomínio/fisiologia , Medula Renal/embriologia , Alça do Néfron/embriologia , Proteínas Supressoras de Tumor/fisiologia , Animais , Apoptose , Proliferação de Células , Camundongos , Concentração Osmolar
14.
Histol Histopathol ; 30(6): 751-62, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25594189

RESUMO

The anion exchanger pendrin is exclusively expressed by non-type A intercalated cells (ICs), type B ICs and non A-non B ICs. Pendrin-positive ICs are mainly localized in the cortical collecting duct (CCD) and connecting tubule (CNT) rather than the outer medullary collecting duct (OMCD). Our previous study reported that Notch signaling is required for the specification of ureteric bud cells to the principal cells (PCs) and ICs in the medullary collecting duct. The purpose of this study was to determine whether the deletion of Mind bomb-1 (Mib1), an E3 ubiquitin ligase required for the initiation of Notch signaling, would affect the differentiation of pendrin-positive type B and non A-non B ICs in Hoxb7-Cre;Mib1f/f mice. In Hoxb7-Cre;Mib1f/f mice, there was a significant increase in the fraction of pendrin-negative/AE1-positive type A ICs not only in the OMCD (67.02±2.04% vs. 33.78±0.71%; Hoxb7-Cre;Mib1f/f vs. Mib1f/f) but also in the CCD (23.70±2.68% vs. 19.71±0.43%; Hoxb7-Cre;Mib1f/f vs. Mib1f/f) and CNT (23.70±2.68% vs. 19.71±0.43%; Hoxb7-Cre;Mib1f/f vs. Mib1f/f) as compared with Mib1f/f. In contrast, there were no significant differences in the fraction of pendrin-positive type B ICs (7.11±3.84% vs. 7.61±4.45%; Hoxb7-Cre;Mib1f/f vs. Mib1f/f) between the two groups in the cortex including CCD and CNT. Furthermore, there was a significant decrease in the fraction of non A-non B ICs (8.95±2.28% vs. 13.06±4.81%; Hoxb7-Cre;Mib1f/f vs. Mib1f/f) in these tubules in the Hoxb7-Cre;Mib1f/f mice. These results suggest that the degree of differentiation of subtypes of ICs may vary depending on the Notch signaling pathway.


Assuntos
Proteínas de Transporte de Ânions/metabolismo , Túbulos Renais Coletores/metabolismo , Rim/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Rim/citologia , Túbulos Renais Coletores/citologia , Camundongos , Camundongos Transgênicos , Transdução de Sinais/genética , Transportadores de Sulfato , Ubiquitina-Proteína Ligases/genética
15.
J Anim Sci Technol ; 56: 11, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-26290700

RESUMO

This study was conducted to evaluate the effects of dietary conjugated linoleic acid (CLA) on the performance of lactating sows and piglets as well as the immunity of piglets suckling from sows fed CLA. Eighteen multiparous Duroc sows with an average body weight (BW) of 232.0 ± 6.38 kg were randomly selected and assigned to two dietary treatments (n = 9 for each treatment), control (no CLA addition) and 1% CLA supplementation. For the control diet, CLA was replaced with soybean oil. Experimental diets were fed to sows during a 28-day lactation period. Litter size for each sow was standardized to nine piglets by cross-fostering within 24 hours after birth. Sow milk and blood samples were taken from sows and piglets after 21 and 27 days of lactation, respectively. Loss of BW was significantly (p < 0.05) higher in sows fed control diet compared to sows fed CLA diet. Piglet weights at weaning and weight gain during suckling were significantly (p < 0.05) higher in sows fed CLA compared to sows fed control diet. Serum non-esterified fatty acid (NEFA) and urea nitrogen concentrations were significantly (p < 0.05) lower in sows fed CLA than in sows fed soybean oil. IgG concentrations of the groups supplemented with CLA increased by 49% in sow serum (p < 0.0001), 23% in milk (p < 0.05), and 35% in piglet serum (p < 0.05) compared with the control group. Sows fed CLA showed an increase of 10% in milk yield compared with sows fed soybean oil (p < 0.05), even though there was no difference in daily feed intake between the treatments. Milk fat content was significantly (p < 0.05) lower in sows fed CLA than in sows fed soybean oil. Solid-not-fat yield was significantly (p < 0.05) higher in sows supplemented with CLA than in sows fed control diet and also protein-to-fat ratio in milk was significantly (p < 0.05) higher in sows fed CLA compared with the control group. The results show that CLA supplementation to sows increased milk yield without losing BW during lactation, whereas soybean oil supplementation resulted in severe BW loss.

16.
Lab Invest ; 92(11): 1583-96, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22964853

RESUMO

The accumulation of extracellular matrix proteins in the interstitial area is the final common feature of chronic kidney diseases. Accumulating evidence suggests that transforming growth factor (TGF)-ß1 promotes the development of renal fibrosis. Heat shock protein (Hsp) 90 inhibitors have been shown to repress TGF-ß1 signaling, but whether they inhibit renal fibrosis is unknown. The purpose of this study is to determine the therapeutic efficacy of Hsp90 inhibitor on renal fibrosis. In TGF-ß1-treated HK2 cells and unilateral ureteral obstruction (UUO) kidneys, we found that 17-allylamino-17-demethoxygeldanamycin (17AAG), an Hsp90 inhibitor, decreased the expression of α-smooth muscle actin, fibronectin, and collagen I and largely restored the expression of E-cadherin. 17AAG inhibited TGF-ß1-mediated phosphorylation of Smad2, Akt, glycogen synthase kinase-3ß, and extracellular signal-regulated kinase in HK2 cells. Inhibition of Hsp90 also blocked TGF-ß1-mediated induction of snail1. This 17AAG-induced reduction was completely restored by simultaneous treatment with proteasome inhibitor MG132. Furthermore, 17AAG blocked the interaction between Hsp90 and TGF-ß type II receptor (TßRII) and promoted ubiquitination of TßRII, leading to the decreased availability of TßRII. Smurf2-specific siRNA reversed the ability of 17AAG to inhibit TGF-ß1 signaling. The effect of 17AAG on TßRII expression and renal fibrosis was confirmed in UUO kidneys. These findings suggest that Hsp90 inhibitor prevents the development of renal fibrosis via a mechanism dependent on Smurf2-mediated degradation of TßRII.


Assuntos
Benzoquinonas/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Nefropatias/prevenção & controle , Lactamas Macrocíclicas/farmacologia , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Animais , Benzoquinonas/uso terapêutico , Linhagem Celular , Modelos Animais de Doenças , Fibrose , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Nefropatias/metabolismo , Nefropatias/patologia , Lactamas Macrocíclicas/uso terapêutico , Masculino , Camundongos , Fosforilação , Ratos , Receptor do Fator de Crescimento Transformador beta Tipo II , Proteínas Smad/metabolismo , Ubiquitina-Proteína Ligases/metabolismo
17.
Nephrology (Carlton) ; 17(2): 148-59, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22085202

RESUMO

AIM: Autophagy is a cellular process of degradation of damaged cytoplasmic components and regulates cell death or proliferation. Unilateral ureteral obstruction (UUO) is a model of progressive renal fibrosis in the obstructed kidney. And UUO is followed by compensatory cellular proliferation in the contralateral kidney. We investigate the role of autophagy in the obstructed kidney and contralateral kidney after UUO. METHODS: To obtain the evidence and the patterns of autophagy during UUO, the rats were sacrificed 3, 7 and 14 days after UUO. To examine the efficacy of the autophagy inhibitors, 3-methyladenine (3-MA), the rats were treated daily with intraperitoneal injection of 3-MA (30 mg/kg per day) for 7 days. RESULTS: After UUO, autophagy was induced in the obstructed kidney in a time-dependent manner. Inhibition of autophagy by 3-MA enhanced tubular cell apoptosis and tubulointerstitial fibrosis in the obstructed kidney after UUO. In the contralateral kidney, autophagy was also induced and prolonged during UUO. Inhibition of autophagy by 3-MA increased the protein expression of proliferating cell nuclear antigen significantly in the contralateral kidney after UUO. The Akt-mammalian target of rapamycin (mTOR) signalling pathway was involved in the induction of autophagy after UUO in both kidneys. CONCLUSION: Our present results support that autophagy induced by UUO has a renoprotective role in the obstructed kidney and regulatory role of compensatory cellular proliferation in the contralateral kidney through Akt-mTOR signalling pathway.


Assuntos
Autofagia , Rim/patologia , Obstrução Ureteral/patologia , Adenina/administração & dosagem , Adenina/análogos & derivados , Animais , Apoptose , Autofagia/efeitos dos fármacos , Proliferação de Células , Citoproteção , Modelos Animais de Doenças , Fibrose , Injeções Intraperitoneais , Rim/efeitos dos fármacos , Rim/metabolismo , Túbulos Renais/patologia , Masculino , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Fatores de Tempo , Obstrução Ureteral/metabolismo
18.
Nephrol Dial Transplant ; 26(11): 3474-84, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21478303

RESUMO

BACKGROUND: Despite the beneficial effects of alagebrium (ALA), a putative advanced glycation end-product (AGE) breaker, on diabetic nephropathy, its renoprotective mechanisms are incompletely understood. Since oxidative stress exacerbates diabetic renal injury through interaction with AGE, the present study examined the antioxidative property of ALA in db/db mice, mesangial cells cultured under high glucose or H(2)O(2) and a test tube. METHODS: ALA (2 mg/kg/day) was administered intraperitoneally for 12 weeks to 8-week-old db/m and db/db (D(ALA)E) mice or for 4 weeks to 16-week-old db/db mice (D(ALA)L). Oxidative stress markers (nitrotyrosine accumulation, expression and translocation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits, cellular DCF-DA fluorescence) together with urinary albumin excretion and histological changes including mesangial expansion were measured. The concentration of H(2)O(2) in the presence and absence of ALA was measured by iodometric analysis in a test tube. RESULTS: ALA significantly reduced not only urinary albumin excretion and renal pathological changes but also accumulation of pentosidine and nitrotyrosine and expression of NADPH oxidase subunits in db/db mice regardless of treatment protocol. In mesangial cells, ALA effectively prevented not only high glucose- but also H(2)O(2)-induced membrane translocation of NADPH oxidase subunit (p47 phox, p67 phox and rac1) and protein kinase C isoform (α, ßI and ßII) and Nox4 messenger RNA expression concomitant with cellular reactive oxygen species. Furthermore, ALA directly decreased H(2)O(2) in a test tube. CONCLUSION: ALA has both direct and indirect antioxidant effects that may play important roles in ALA's renoprotective effect in diabetic kidneys.


Assuntos
Injúria Renal Aguda/prevenção & controle , Antioxidantes/uso terapêutico , Diabetes Mellitus Experimental/fisiopatologia , Nefropatias Diabéticas/prevenção & controle , Células Mesangiais/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Tiazóis/uso terapêutico , Injúria Renal Aguda/metabolismo , Injúria Renal Aguda/patologia , Albuminúria/etiologia , Albuminúria/prevenção & controle , Animais , Western Blotting , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Peróxido de Hidrogênio/farmacologia , Técnicas Imunoenzimáticas , Masculino , Células Mesangiais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Proteína Quinase C/genética , Proteína Quinase C/metabolismo , RNA Mensageiro/genética , Espécies Reativas de Oxigênio/metabolismo , Reação em Cadeia da Polimerase em Tempo Real
19.
Cell Tissue Res ; 343(2): 429-44, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21181199

RESUMO

Our knowledge of the embryonic development of the lymphatic vessels within the kidney is limited. The aim of this study was to establish the time of appearance and the distribution of intra-renal lymphatic vessels in the developing mouse kidney by using the lymphatic marker, LYVE-1. Kidneys from embryonic day 12 (E12) to E18, from neonates at post-natal day 1 (P1) to P21, and from adults were studied. In the adult mouse kidney, LYVE-1 was expressed mainly in the lymphatic endothelial cells (LECs) and in a subset of endothelial cells in the glomerular capillaries. However, in the developing mouse kidney, LYVE-1 was also expressed transiently in F4/80(+)/CD11b(-) immature macrophages/dendritic cells and in the developing renal vein. LYVE-1(+) lymphatic vessels connected with extra-renal lymphatics were detected in the kidney at E13. F4/80(+)/CD11b(-)/LYVE-1(+) immature macrophages/dendritic cells appeared prior to the appearance of LYVE-1(+) renal lymphatic vessels and were closely intermingled or even formed part of the lymphatic vascular wall. Prox1 was expressed only in the LYVE-1(+) LECs from fetus to adult-hood, but not in LYVE-1(+) endothelial cells of the developing renal vein and macrophages/dendritic cells. Thus, lymphatic vessels of the kidney might originate by extension of extra-renal lymphatics through an active branching process possibly associated with F4/80(+)/CD11b(-)/LYVE-1(+) macrophages/dendritic cells.


Assuntos
Endotélio Linfático/metabolismo , Glicoproteínas/metabolismo , Receptores de Hialuronatos/metabolismo , Rim/embriologia , Animais , Células Endoteliais/metabolismo , Endotélio Linfático/embriologia , Feminino , Rim/metabolismo , Proteínas de Membrana Transportadoras , Camundongos , Camundongos Endogâmicos C57BL
20.
Am J Nephrol ; 33(1): 7-16, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21124021

RESUMO

BACKGROUND/AIMS: The increased permeability of chloride in the distal cortical nephron in cyclosporine nephrotoxicity may involve the transcellular pathway mediated by the thiazide-sensitive Na(+)-Cl⁻ cotransporter and/or the paracellular pathway mediated by the tight junctions (TJs). METHODS: Cyclosporine was subcutaneously administered to Sprague-Dawley rats for 6 (7.5 mg/kg body weight) and 2 (25 mg/kg body weight) weeks, and immunoblot analysis and immunohistochemistry were carried out from the kidneys. Electrically tight epithelial Madin-Darby canine kidney (MDCK) I cells were exposed to cyclosporine for 72 h to measure changes in transepithelial electrical resistance (ΔTER). RESULTS: Cyclosporine treatment induced a decrease in Na(+)-Cl⁻ cotransporter in rat renal cortex. WNK4 protein was increased in both rat kidneys and MDCK I cells. Occludin was also increased in rat kidneys and MDCK I cells exposed to 100 ng/ml cyclosporine. In contrast, cyclosporine treatment induced a decrease in zonula occludens 1 protein abundance and no changes in claudin-1 and claudin-4 in both rat kidneys and MDCK I cells. As a measure of the barrier to small ions, ΔTER of MDCK monolayers was decreased by 100 ng/ml cyclosporine. CONCLUSION: Renal TJ proteins are affected by cyclosporine treatment. Changes in TJ protein assembly induced by altered expression of WNK4, occludin, and zonula occludens 1 may affect paracellular permeability.


Assuntos
Ciclosporina/efeitos adversos , Regulação da Expressão Gênica , Imunossupressores/efeitos adversos , Rim/efeitos dos fármacos , Proteínas de Membrana/biossíntese , Néfrons/metabolismo , Fosfoproteínas/biossíntese , Junções Íntimas/metabolismo , Animais , Claudina-1 , Claudina-4 , Cães , Rim/metabolismo , Masculino , Ocludina , Ratos , Ratos Sprague-Dawley , Proteína da Zônula de Oclusão-1
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