RESUMO
Plant-derived compounds can be useful for the management of liver disease. Traditionally, hepatic disorders have been treated with herbal extracts. Although many herbal extracts in Eastern medicine have been shown to possess hepatoprotective activities, single-origin herbal extracts primarily demonstrate either antioxidant or anti-inflammatory activities. The current study investigated the effects of combinatorial herbal extracts on alcohol-induced hepatic disorders in an ethanol-fed mouse model. Sixteen herbal combinations were evaluated as hepatoprotective formulations; the active constituents in these herbal extracts were daidzin, peonidin-3-glucoside, hesperidin, glycyrrhizin, and phosphatidylcholine. RNA sequencing analysis showed that exposure to ethanol altered hepatic gene expression profiles (compared to those of the non-alcohol-fed group), resulting in 79 differentially expressed genes. A majority of the differentially expressed genes in alcohol-induced hepatic disorders were associated with dysfunction of the normal cellular homeostasis in the liver; however, these genes were repressed by treatment with herbal extracts. Moreover, following treatment with herbal extracts, there were neither acute inflammatory responses in the liver tissue nor abnormalities in the cholesterol profile. These results suggest that combinatorial herbal extracts may alleviate alcohol-induced hepatic disorders by modulating the inflammatory response and lipid metabolism in the liver.
Assuntos
Fígado , Extratos Vegetais , Camundongos , Animais , Extratos Vegetais/farmacologia , Antioxidantes/farmacologia , Etanol/efeitos adversos , Anti-Inflamatórios/farmacologiaRESUMO
Sigumjang is a traditional Korean fermented barley bran consumed in the Gyungsang-do area. In this study, pork loin was marinated with sigumjang, and its physicochemical and textural properties were investigated. Sigumjang-marinated pork loin (SMPL) displayed an attractive yellowish-brown overall color. SMPL's pH marginally increased during storage and finally equilibrated to the pH of sigumjang, 5.5, on day 14. The amino-type nitrogen content of SMPL increased linearly during the marination period. Due to the lower extractability and higher pH compared to the control, the volume of myofibrillar fragments in SMPL decreased during the marination period. SMPL showed an increase in free amino acids related to umami (Asp and Glu) and sweet (Ser, Thr, Ala and Gly) tastes compared to the control. These may indicate improvement of sensory qualities of SMPL. The results also provide valuable information for the use of sigumjang in the development of novel meat products.
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Although the pharmacological action of red ginseng is mostly generated by ginsenosides, contents of ginsenosides in manufactured extracts are still varying according to processing and storage conditions. Rg1, Rb1, and Rh1 levels significantly decreased overtime under pH adjustment and thermal treatment during storage, and these changes were exacerbated at lower pH and higher storage temperature. However, Rg3 level showed the opposite pattern compared to other ginsenosides. Rg3 level linearly increased at higher temperature and pH while significantly decreased at pH 2. Furthermore, Rg3 level constantly increased during storage. This is the first combined study on the effects of manufacturing and storage conditions on ginsenoside contents of red-ginseng-extract products. To minimize loss of major marker and bioactive compounds of red ginseng products during manufacturing processes and storage, it is recommended that red-ginseng-extracts be maintained at pH 6-8, sterilized at below 105 °C, and stored at below 25 °C.
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Context Ligularia fischeri (Ledebour) Turcz. (Compositae) has been used as a leafy vegetable and in traditional medicine to treat hepatic disorder in East Asia. Objective The present study explores the antioxidant activity of LF aqueous extract on EtOH-induced oxidative stress accompanied by hepatotoxicity both in vitro and in vivo. Materials and methods In vitro study using the mouse liver NCTC-1469 cell line was conducted to estimate the cytotoxicity as well as the inhibitory effect of LF extract against alcohol-treated cell damage. In vivo study used an alcohol-fed Wister rat model orally administered EtOH (3.95 g/kg of body weight/d) with or without LF extract (100 or 200 mg/kg body weight) for 6 weeks. Serum and liver tissue were collected to evaluate hepatic injury and antioxidant-related enzyme activity. Results The EC50 value for the DPPH radical scavenging capacity of LF extract was 451.5 µg/mL, whereas the IC50 value of LF extract in terms of EtOH-induced reactive oxygen species (ROS) generation was 98.3 µg/mL without cell cytotoxicity. LF extract (200 mg/kg body weight) significantly reduced the triglyceride content of serum (33%) as well as hepatic lipid peroxidation (36%), whereas SOD activity was elevated three-fold. LF extract suppressed expression of CYP2E1 and TNF-α, and attenuated alcohol-induced abnormal morphological changes. Discussion and conclusion LF extract attenuated liver damage induced by alcoholic oxidative stress through inhibition of ROS generation, down-regulation of CYP2E1, and activation of hepatic antioxidative enzymes. Homeostasis of the antioxidative defence system in the liver by LF extract mitigated hepatic disorder following chronic alcohol intake.
Assuntos
Antioxidantes/farmacologia , Hepatopatias Alcoólicas/prevenção & controle , Fígado/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Antioxidantes/isolamento & purificação , Asteraceae/química , Compostos de Bifenilo/química , Linhagem Celular , Citocromo P-450 CYP2E1/metabolismo , Citoproteção , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Peroxidação de Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Hepatopatias Alcoólicas/metabolismo , Hepatopatias Alcoólicas/patologia , Masculino , Camundongos , Fitoterapia , Picratos/química , Extratos Vegetais/isolamento & purificação , Folhas de Planta , Plantas Medicinais , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Sigumjang prepared from fermented barley bran is a traditional fermented food found only in the Gyeongsang-do area of South Korea. There have been no studies reported to date despite the potential bioactivities of sigumjang. In this study, the anti-obesity activities of sigumjang extracts (SEs) during 3T3-L1 differentiation into adipocytes were investigated. SEs inhibited adipocyte differentiation by suppressing the CCAAT/enhancer binding protein-ß and sterol regulatory element binding protein-1c expression in the early stage of differentiation, followed by the suppression of the peroxisome proliferator-activated receptor-γ, CCAAT/enhancer binding protein-α, and adiponectin. These changes in adipogenic markers induced inhibition of lipogenesis via down-regulation of mainly fatty acid synthase, acetyl-CoA carboxylase, fatty acid binding protein 4, and perilipin. These results were more significant in the extract of sigumjang fermented with isolated Bacillus amyloliquefaciens MFST compared to naturally fermented sigumjang group. SEs can be considered as a useful material for developing food with health benefits and anti-obesity properties.
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BACKGROUND: Proteinuria is typically accompanied by structural and compositional changes of the foot processes and of the slit diaphragms between podocytes. CD2-associated protein (CD2AP) in podocytes serves as an adaptor protein binding to nephrin and podocin, anchoring these slit diaphragm proteins to actin filaments of podocyte cytoskeleton and sending signals inward or outward. METHODS: In the present study, we prepared streptozotocin-induced diabetic renal tissues and cultured podocytes in diabetic conditions to investigate podocyte phenotypical changes, including quantitative and distributional changes of CD2AP protein and search for the signalling mechanisms in diabetic conditions. We prepared cultured rat glomerular epithelial cells and mouse podocytes to study how high glucose and advanced glycosylation end products (AGE) induce phenotypical changes of cultured podocyte, under (1) normal glucose (5 mM, = control), (2) high glucose (30 mM), (3) AGE-added or (4) high glucose plus AGE-added conditions. RESULTS: According to diabetic duration, density of CD2AP in renal tissue of experimental diabetic nephropathy became conglomerulated and diminished. In cultured podocytes, CD2AP co-localized with nephrin and zonula occludens-1 by confocal imaging. High glucose and high glucose plus AGE induced the relocalization and concentration of CD2AP at internal cytoplasmic and perinuclear areas of podocytes. High glucose plus AGE-added condition also decreased CD2AP protein amount and its mRNA expression compared with normal glucose or osmotic control conditions. In addition, LY294002, a phosphoinositide 3-kinase inhibitor, prevented the quantitative and distributional changes of CD2AP induced by high glucose and AGE. CONCLUSIONS: These findings suggest that diabetic conditions induce the phenotypical changes of podocyte CD2AP possibly via phosphoinositide 3-kinase/Akt signalling.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas do Citoesqueleto/genética , Diabetes Mellitus Experimental/genética , Proteína Oncogênica v-akt/fisiologia , Fosfatidilinositol 3-Quinases/fisiologia , Podócitos/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Células Cultivadas , Proteínas do Citoesqueleto/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Glucose/farmacologia , Camundongos , Proteína Oncogênica v-akt/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Podócitos/efeitos dos fármacos , Podócitos/patologia , Ratos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , EstreptozocinaRESUMO
BACKGROUND: Adenosine monophosphate-activated protein kinases (AMPKs), as a sensor of cellular energy status, have been known to play an important role in the pathophysiology of diabetes and its complications. Because AMPKs are known to be expressed in podocytes, it is possible that podocyte AMPKs could be an important contributing factor in the development of diabetic proteinuria. We investigated the roles of AMPKs in the pathological changes in podocytes induced by high-glucose (HG) and advanced glycosylation end products (AGEs) in diabetic proteinuria. METHODS: We prepared streptozotocin-induced diabetic renal tissues and cultured rat and mouse podocytes under diabetic conditions with AMPK-modulating agents. The changes in AMPKα were analyzed with confocal imaging and Western blotting under the following conditions: (1) normal glucose (5mM, =control); (2) HG (30mM); (3) AGE-added; or (4) HG plus AGE-added. RESULTS: The density of glomerularphospho-AMPKα in experimental diabetic nephropathy decreased as a function of the diabetic duration. Diabetic conditions including HG and AGE changed the localization of phospho-AMPKα from peripheral cytoplasm to internal cytoplasm and peri- and intranuclear areas in podocytes. HG reduced the AMPKα (Thr172) phosphorylation of rat podocytes, and similarly, AGEs reduced the AMPKα (Thr172) phosphorylation of mouse podocytes. The distributional and quantitative changes in phospho-AMPKα caused by diabetic conditions were preventable using AMPK activators, metformin, and 5-aminoimidazole-4-carboxamide-1ß-riboside. CONCLUSION: We suggest that diabetic conditions induce the relocation and suppression of podocyte AMPKα, which would be a suggestive mechanism in diabetic podocyte injury.
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BACKGROUND: Proteinuria results from increased glomerular permeability and is associated with retraction and effacement of the highly specialized interdigitating podocyte foot processes. In podocytes (glomerular epithelial cells [GECs]), p130Cas localizes diffusely to the cytoplasm and connects focal adhesion proteins and kinases to the glomerular basement membrane and adapter proteins of slit diaphragm insertion sites. METHODS: To investigate whether high-glucose (HG) and advanced glycosylation end products (AGEs) induce quantitative and distributional changes of podocyte p130Cas protein, a docking protein connecting F-actin fibers to the glomerular basement membrane and adapter proteins, we cultured rat GECs (r-GECs) and mouse GECs (m-GECs) under (i) normal glucose (5 mM = control), (ii) HG (30 mM), (iii) AGE-added or (iv) HG plus AGE-added conditions. We also prepared streptozotocin-induced diabetic renal tissues. RESULTS: We found that p130Cas stainings were located in peripheral cytoplasm of r-GECs and m-GECs as dots in linear alignment, partially colocalized with actin-binding proteins such as synaptopodin and alpha-actinin, and locally connected to the ends of actin filaments. In diabetic conditions, the intensities of p130Cas stainings were diminished in more pathological HG plus AGE-added condition of r-GECs and m-GECs and in chronic diabetic renal tissues. p130Cas protein was decreased significantly by HG, AGE and HG plus AGE in both cells compared with normal glucose or osmotic control conditions. p130Cas mRNA expression levels were also suppressed similarly in diabetic conditions. CONCLUSION: We suggest that diabetic conditions modulate the quantitative and distributional changes of podocyte p130Cas and therefore affect the filtration function of podocytes.
Assuntos
Proteína Substrato Associada a Crk/metabolismo , Diabetes Mellitus Experimental/metabolismo , Nefropatias Diabéticas/metabolismo , Podócitos/metabolismo , Actinas/metabolismo , Animais , Células Cultivadas , Proteína Substrato Associada a Crk/genética , Citoplasma/metabolismo , Células Epiteliais/metabolismo , Membrana Basal Glomerular/metabolismo , Produtos Finais de Glicação Avançada/metabolismo , Glomérulos Renais/patologia , Camundongos , Proteínas dos Microfilamentos/metabolismo , RNA Mensageiro/metabolismo , RatosRESUMO
BACKGROUND: The aim of this study was to enhance the bioavailability of conjugated linoleic acid (CLA), which has low water solubility, using nanoemulsion technology and to evaluate the effects of its improved bioavailability as an antiobesity agent. METHODS: The antiobesity effect of nanoemulsified water-soluble conjugated linoleic acid (N-CLA) was evaluated using in vitro and in vivo studies. Differentiated 3T3-L1 adipocytes were treated with CLA and N-CLA to assess their lipolytic effect. Further, to confirm the antiobesity effect of N-CLA, male Sprague-Dawley rats were randomly separated into four groups, ie, a group fed a normal diet, a group fed a high-fat diet (obesity rat model), a CLA-treated group, and an N-CLA-treated group. RESULTS: N-CLA showed a greater lipolytic effect on differentiated 3T3-L1 adipocytes compared with normal CLA. N-CLA enhanced the release of glycerol from triglycerides, which accumulated in differentiated 3T3-L1 adipocytes. Further, N-CLA enhanced leptin secretion to an extent similar to that of orlistat, an antiobesity agent. In an animal obesity model fed a high-fat diet, N-CLA attenuated accumulation of triglycerides, total cholesterol, and low-density lipoprotein cholesterol in serum, and also significantly decreased the volume of triglycerides and cholesterol in liver tissue. CONCLUSION: These results indicate that N-CLA has a greater antiobesity effect than CLA as a result of its improved bioavailability.
Assuntos
Fármacos Antiobesidade/farmacocinética , Ácidos Linoleicos Conjugados/farmacocinética , Nanopartículas/química , Animais , Fármacos Antiobesidade/administração & dosagem , Fármacos Antiobesidade/química , Disponibilidade Biológica , Peso Corporal/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Colesterol/sangue , Dieta Hiperlipídica , Emulsões/administração & dosagem , Emulsões/química , Emulsões/farmacocinética , Ácidos Linoleicos Conjugados/administração & dosagem , Ácidos Linoleicos Conjugados/química , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Células NIH 3T3 , Nanopartículas/administração & dosagem , Nanotecnologia , Tamanho do Órgão/efeitos dos fármacos , Tamanho da Partícula , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Solubilidade , Triglicerídeos/sangueRESUMO
Plant-derived polyhenols inhibit cancer cell proliferation and induce apoptosis. Recently, prenylflavonoids and alkyl-phloroacetophenones have been reported for their in vitro antitumor activity. In the present study, we examined the cytotoxic activity of prenyl (3-PAP) and geranyl (3-GAP) derivatives of phloroacetophenone, and xanthohumol (XN), a prenyl-chalcone, in human breast cancer (MCF-7) and human sarcoma (HT1080) cell lines in vitro. 3-GAP showed the strongest cytotoxicity in these cell lines with IC(50) values of less than 10 µM. In addition, we report that 3-GAP is a more potent anti-cancer agent for breast cancer than XN which is a well-known anticancer flavonoid. Moreover, 3-GAP did not induce cytotoxicity in the normal cell line, TCMK-1, whereas 3-PAP and XN significantly reduced TCMK-1 cell viability. In 3-GAP-treated MCF-7 cells, nuclear accumulation and transcriptional activity of p53 were increased. In addition, pro-apoptotic Bax but not B-cell lymphoma 2 (Bcl-2) expression was increased by 3-GAP. In accordance with the Bax increase, 3-GAP induced mitochondrial cytochrome c release and activated caspase-9, an initiator of the caspase cascade. In the MCF-7 cell line which does not express caspase-3, activation of caspase-7, a member of the caspase-3 subfamily, was increased by 3-GAP. The present results indicate that synthetic 3-GAP is a safe and effective anti-cancer agent, and the Bax-mediated mitochondrial pathway is the main apoptosis signaling pathway of 3-GAP in MCF-7 cells.
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Acetofenonas/uso terapêutico , Antineoplásicos Fitogênicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Mitocôndrias/efeitos dos fármacos , Fitoterapia , Sarcoma/tratamento farmacológico , Proteína X Associada a bcl-2/metabolismo , Acetofenonas/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias da Mama/metabolismo , Caspases/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Citocromos c/metabolismo , Feminino , Flavonoides/farmacologia , Flavonoides/uso terapêutico , Humanos , Concentração Inibidora 50 , Mitocôndrias/metabolismo , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Propiofenonas/farmacologia , Propiofenonas/uso terapêutico , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Sarcoma/metabolismo , Proteína Supressora de Tumor p53/metabolismoRESUMO
In this study, we investigated whether fermented rice bran (FRB) can ameliorate the oxidative stress induced by high glucose and hydrogen peroxide (H(2)O(2)) in 3T3-L1 adipocytes by analyzing reactive oxygen species (ROS), oil red O staining, as well as the expression of mRNAs related to glucose homeostasis and adipogenesis. It was first confirmed that rice bran fermented by Issatchenkia orientalis MFST1 extract increased free phenolic content compared to non-fermented rice bran. The FRB extract strongly inhibited ROS generation and upregulated the expression of PPAR-γ and adiponectin. Moreover, FRB upregulated GLUT4 related to glucose transportation and insulin sensitivity. Taken together, FRB extract ameliorated oxidative stress-induced insulin resistance by neutralizing free radicals and upregulating adiponectin in adipocytes. Our results provide information toward understanding the beneficial effects of FRB on oxidative stress.
Assuntos
Adipócitos/efeitos dos fármacos , Resistência à Insulina , Oryza , Estresse Oxidativo/efeitos dos fármacos , Fenóis/uso terapêutico , Fitoterapia , Extratos Vegetais/uso terapêutico , Células 3T3-L1 , Adipócitos/metabolismo , Adiponectina/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Fermentação , Glucose/efeitos adversos , Transportador de Glucose Tipo 4/metabolismo , Peróxido de Hidrogênio , Insulina/metabolismo , Camundongos , PPAR gama/metabolismo , Fenóis/farmacologia , Extratos Vegetais/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Sementes , Regulação para CimaRESUMO
The slit diaphragm connecting the adjacent foot processes of glomerular epithelial cells (podocytes) is the final barrier of the glomerular capillary wall and serves to prevent proteinuria. Podocytes are understood to be terminally differentiated cells and share some common features with neurons. Neurexin is a presynaptic adhesion molecule that plays a role in synaptic differentiation. Although neurexin has been understood to be specifically expressed in neuronal tissues, we found that neurexin was expressed in several organs. Several forms of splice variants of neurexin-1α were detected in the cerebrum, but only one form of neurexin-1α was detected in glomeruli. Immunohistochemical study showed that neurexin restrictedly expressed in the podocytes in kidneys. Dual-labeling analyses showed that neurexin was colocalized with CD2AP, an intracellular component of the slit diaphragm. Immunoprecipitation assay using glomerular lysate showed that neurexin interacted with CD2AP and CASK. These observations indicated that neurexin localized at the slit diaphragm area. The staining intensity of neurexin in podocytes was clearly lowered, and their staining pattern shifted to a more discontinuous patchy pattern in the disease models showing severe proteinuria. The expression and localization of neurexin in these models altered more clearly and rapidly than that of other slit diaphragm components. We propose that neurexin is available as an early diagnostic marker to detect podocyte injury. Neurexin coincided with nephrin, a key molecule of the slit diaphragm detected in a presumptive podocyte of the developing glomeruli and in the glomeruli for which the slit diaphragm is repairing injury. These observations suggest that neurexin is involved in the formation of the slit diaphragm and the maintenance of its function.
Assuntos
Glomérulos Renais/citologia , Podócitos/metabolismo , Receptores de Superfície Celular/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Sequência de Aminoácidos , Estruturas Animais/metabolismo , Animais , Cérebro/metabolismo , Proteínas do Citoesqueleto/metabolismo , Embrião de Mamíferos/metabolismo , Feminino , Expressão Gênica/genética , Glicoproteínas/genética , Guanilato Quinases/metabolismo , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Proteínas de Membrana/metabolismo , Dados de Sequência Molecular , Síndrome Nefrótica/induzido quimicamente , Síndrome Nefrótica/metabolismo , Síndrome Nefrótica/patologia , Proteínas do Tecido Nervoso/genética , Neuropeptídeos/genética , Podócitos/patologia , Ligação Proteica/fisiologia , Isoformas de Proteínas/genética , Proteinúria/urina , Ratos , Ratos Wistar , Receptores de Superfície Celular/genética , Organismos Livres de Patógenos EspecíficosRESUMO
High glucose levels can change podocyte gene expression and subsequently induce podocyte damage through altered glucose metabolism. l-Carnitine is known to play a beneficial role in diabetes; however, there are no studies on the effects of l-carnitine on podocyte alteration under high glucose conditions. This study investigated whether l-carnitine can attenuate diabetic podocyte injury through the prevention of loss of slit diaphragm proteins. The l-carnitine treatment group showed increased glucose uptakes compared to the control group, suggesting that glucose utilization in the podocytes was increased by l-carnitine. l-Carnitine treatment also prevented decreased mRNA expressions of nephrin and podocin in the high glucose-stimulated podocytes. However, mRNA expressions of CD2AP and α-actinin-4 were not significantly changed by the high glucose conditions. When these data are taken together, l-carnitine can increase glucose uptake in podocytes under high glucose conditions, and its mechanism may be at least partly related to the up-regulation of nephrin and podocin. Our results help clarify the beneficial effects of l-carnitine in diabetic nephropathy.
Assuntos
Carnitina/metabolismo , Diabetes Mellitus Experimental/metabolismo , Podócitos/metabolismo , Animais , Linhagem Celular , Diabetes Mellitus Experimental/induzido quimicamente , Nefropatias Diabéticas/prevenção & controle , Regulação da Expressão Gênica , Glucose/metabolismo , Hiperglicemia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Glomérulos Renais/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Concentração Osmolar , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Recent studies have demonstrated that the slit diaphragm of the glomerular epithelial cell (podocyte) is the structure likely to be the barrier in the glomerular capillary wall. Murine monoclonal antibody against nephrin, a molecule constituting the extracellular site of the slit diaphragm, caused severe proteinuria if injected into rats, in a complement- or inflammatory cell-independent manner. In this proteinuric state, not only nephrin but also other slit diaphragm-associated molecules are down-regulated. These observations suggest that the antibody alters the molecular composition of the slit diaphragm and, thereby, affects the glomerular permeability barrier. Recently, it was found that IP-10, SV2B, ephrin B1 and the receptors of angiotensin II were expressed in the podocyte, and that their expressions were clearly altered in anti-nephrin antibody-induced nephropathy. It is conceivable that these molecules are involved in the development of proteinuria in this model. IP-10 is assumed to play a role in maintaining the slit diaphragm function by regulating the cell cycle balance of the podocyte. SV2B and ephrin B1 play pivotal roles in the proper localization of the slit diaphragm component. In vivo and in vitro studies demonstrated that angiotensin II type 2 receptor-mediated action enhanced the expression of nephrin. We propose that these molecules could be novel therapeutic targets for proteinuria.
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Proteínas de Membrana/imunologia , Podócitos/imunologia , Proteinúria/etiologia , Animais , Quimiocina CXCL10/antagonistas & inibidores , Quimiocina CXCL10/fisiologia , Efrina-B1/antagonistas & inibidores , Efrina-B1/fisiologia , Humanos , Glicoproteínas de Membrana/antagonistas & inibidores , Glicoproteínas de Membrana/fisiologia , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/fisiologia , Proteinúria/terapia , Receptor Tipo 2 de Angiotensina/fisiologiaRESUMO
This study shows the effects of L-carnitine treatment on cell proliferation with hepa1c1c7 mouse cancer cells and NCTC 1469 normal cells. In an MTT assay, L-carnitine increased the number of dead hepa1c1c7 cells, while there was no difference in the number of NCTC 1469 cells. mRNA and protein levels of TNF-alpha, Fas, and caspase-8, which are closely related to cell apoptosis by a death ligand/receptor-dependent apoptosis pathway, were increased by L-carnitine treatment. In addition, L-carnitine treatment regulated mitochondria-dependent apoptosis pathways by inducing the up-regulation of caspase-9 and caspase-3 and the down-regulation of Bcl-2 in hepa1c1c 7 cells. Taken together, the findings of this study have demonstrated that L-carnitine could induce apoptosis in hepa1c1c7 cells by regulating Fas ligands and inhibiting the expression of Bcl-2. These results suggest that L: -carnitine treatment could be related to both a mitochondrion-dependent and a death ligand/receptor-dependent apoptosis pathway in hepa1c1c7 cells. Our results could give information for understanding the L-carnitine-induced apoptosis mechanism in some cancer cells.
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Apoptose , Carnitina/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Receptor fas/metabolismo , Animais , Caspase 3/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 8/efeitos dos fármacos , Caspase 8/metabolismo , Caspase 9/efeitos dos fármacos , Caspase 9/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Proteína Ligante Fas/efeitos dos fármacos , Proteína Ligante Fas/metabolismo , Camundongos , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Fator de Necrose Tumoral alfa/efeitos dos fármacos , Proteína Supressora de Tumor p53/efeitos dos fármacos , Proteína Supressora de Tumor p53/metabolismo , Receptor fas/efeitos dos fármacosRESUMO
The role of chemokines, especially CXCL10/interferon-gamma-inducible protein 10 kDa (IP-10), a chemokine to attract CXCR3(+) T-helper 1-type CD4(+) T cells, is largely unknown in the pathophysiology of inflammatory bowel disease; ulcerative colitis and Crohn's disease. The authors have earlier shown that IP-10 neutralization protected mice from acute colitis by protecting crypt epithelial cells of the colon. To investigate the therapeutic effect of neutralization of IP-10 on chronic colitis, an anti-IP-10 antibody was injected into mice with newly established murine AIDS (MAIDS) colitis. Anti-IP-10 antibody treatment reduced the number of colon infiltrating cells when compared to those mice given a control antibody. The treatment made the length of the crypt of the colon greater than control antibody. The number of Ki67(+) proliferating epithelial cells was increased by the anti-IP-10 antibody treatment. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)(+) apoptotic cells were observed in the epithelial cells of the luminal tops of crypts in control MAIDS colitis, whereas TUNEL(+) apoptotic epithelial cells were rarely observed with anti-IP-10 antibody treatment. In conclusion, blockade of IP-10 attenuated MAIDS colitis through blocking cellular trafficking and protecting intestinal epithelial cells, suggesting that IP-10 plays a key role in the development of inflammatory bowel disease as well as in chronic experimental colitis.
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Quimiocinas CXC/antagonistas & inibidores , Colite/prevenção & controle , Enterócitos/patologia , Síndrome de Imunodeficiência Adquirida Murina/prevenção & controle , Animais , Anticorpos Bloqueadores/farmacologia , Anticorpos Monoclonais/farmacologia , Apoptose/efeitos dos fármacos , Quimiocina CXCL10 , Quimiocinas CXC/imunologia , Doença Crônica , Colite/metabolismo , Colo/efeitos dos fármacos , Colo/metabolismo , Colo/patologia , Modelos Animais de Doenças , Enterócitos/efeitos dos fármacos , Enterócitos/metabolismo , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Marcação In Situ das Extremidades Cortadas , Antígeno Ki-67/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Síndrome de Imunodeficiência Adquirida Murina/metabolismo , Transporte Proteico/efeitos dos fármacosRESUMO
Although angiotensin II (Ang II) type 1 receptor antagonist ameliorates proteinuria, its pharmacological mechanism and the differential roles of Ang II type 1 receptor (AT1R) and type 2 receptor (AT2R) are not well understood. We analyzed the effect of Ang II type 1 receptor antagonist on proteinuria caused by antibody against nephrin, a functional molecule of glomerular slit diaphragm and dysfunction of which is involved in the development of proteinuria in several glomerular diseases. We show here that AT1R antagonist ameliorated proteinuria by preventing a reduction in the functional molecules of the slit diaphragm. We also analyzed the role of AT1R- or AT2R-mediated actions on the expression of the slit diaphragm molecules in an in vivo study of normal rat and in an in vitro study of cultured podocytes. AT1R-mediated action hampered the mRNA expression of the slit diaphragm molecules, whereas AT2R-mediated action enhanced it. These findings indicate that Ang II receptor subtypes play opposite roles in regulating the barrier function of glomerular capillary wall and that the enhancement of AT2R stimulation may serve as a potential therapeutic strategy for proteinuria.
Assuntos
Capilares , Glomérulos Renais , Podócitos/metabolismo , Receptor Tipo 1 de Angiotensina/metabolismo , Receptor Tipo 2 de Angiotensina/metabolismo , Angiotensina II/metabolismo , Bloqueadores do Receptor Tipo 1 de Angiotensina II/metabolismo , Bloqueadores do Receptor Tipo 2 de Angiotensina II , Animais , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais/toxicidade , Capilares/citologia , Capilares/metabolismo , Feminino , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Glomérulos Renais/citologia , Glomérulos Renais/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Podócitos/citologia , Proteinúria , Ratos , Receptor Tipo 1 de Angiotensina/agonistas , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 2 de Angiotensina/agonistas , Receptor Tipo 2 de Angiotensina/genética , Sialoglicoproteínas/genética , Sialoglicoproteínas/metabolismo , Proteína da Zônula de Oclusão-1RESUMO
Synaptic vesicle protein 2B (SV2B) was identified by the subtraction hybridization technique as a molecule of which mRNA expression was decreased in puromycin aminonucleoside (PAN) nephropathy by glomerular cDNA subtraction assay. The expression of SV2B was detected in glomerular lysate with Western blot analysis. Dual-labeling immunofluorescence studies with glomerular cell markers demonstrated that SV2B is expressed in glomerular visceral epithelial cells (podocytes). The expression of SV2B is detected also in cultured podocyte and in human kidney section as podocytic pattern. The decrease of SV2B mRNA was already detected before the onset of proteinuria in PAN nephropathy. The mRNA expression of SV2B clearly is altered not only in PAN nephropathy but also in another proteinuric state that is caused by an antibody against nephrin, a functional molecule of the slit diaphragm. The decreased intensity in SV2B staining was already detected before the peak of proteinuria in both models with immunofluorescence study. A reduced amount of SV2B was detected in both models also with Western blot analysis. CD2AP, another functional molecule of the slit diaphragm, was observed in cytoplasm, including the processes area of the cultured podocyte, and when the podocyte was treated with small interfering RNA for SV2B, CD2AP staining at the process area was not detected. These results suggest that SV2B is a functional molecule of podocyte, and SV2B may play a role in the expression and proper localization of CD2AP.
Assuntos
Glomerulonefrite/metabolismo , Glomérulos Renais/metabolismo , Glicoproteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Podócitos/metabolismo , Proteinúria/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Antibióticos Antineoplásicos/toxicidade , Western Blotting , Células COS , Células Cultivadas , Chlorocebus aethiops , Citoplasma/metabolismo , Proteínas do Citoesqueleto/metabolismo , DNA Complementar/genética , Feminino , Imunofluorescência , Glomerulonefrite/induzido quimicamente , Glomerulonefrite/patologia , Humanos , Glomérulos Renais/patologia , Glicoproteínas de Membrana/antagonistas & inibidores , Glicoproteínas de Membrana/metabolismo , Proteínas de Membrana/imunologia , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/metabolismo , Proteinúria/fisiopatologia , Puromicina Aminonucleosídeo/toxicidade , RNA Mensageiro/análise , RNA Interferente Pequeno/farmacologia , Ratos , Ratos Wistar , Técnica de Subtração , Proteína rab3A de Ligação ao GTP , Domínios de Homologia de srcRESUMO
Although the role of glomerular basement membrane has been emphasised as the barrier for retaining plasma proteins in the past three decades, some recent studies have demonstrated that the slit diaphragm of the glomerular epithelial cell (podocyte) is the structure likely to be the barrier in the glomerular capillary wall. Nephrin and podocin were identified as gene products mutated in Finnish-type congenital nephrotic syndrome and autosomal recessive steroid-resistant nephrotic syndrome, respectively. Nephrin s located at the outer leaflet of plasma membranes of the slit diaphragm. Podocin is reported to have an interaction with nephrin. The anti-nephrin antibody is capable of inducing massive proteinuria, which indicates that nephrin is a key functional molecule in the slit diaphragm. The expression of nephrin and podocin was reduced in glomeruli of minimal change nephrotic syndrome, which suggested that the altered expression of these molecules contributes to the development of proteinuria also in acquired diseases. Some recent studies demonstrated that CD2-associated protein (CD2AP) is also a functional molecule in the slit diaphragm, and its expression is altered in membranous nephropathy. These observations suggested that alteration of the molecular arrangement in the slit diaphragm is involved in the development of proteinuria in several kinds of glomerular diseases.
Assuntos
Podócitos/fisiologia , Animais , Humanos , Podócitos/química , Podócitos/citologiaRESUMO
Exocrinopathy and pancreatitis-like injury were developed in C57BL/6 (B6) mice infected with LP-BM5 murine leukemia virus, which is known to induce murine acquired immunodeficiency syndrome (MAIDS). The role of chemokines, especially CXCL10/interferon (IFN)-gamma-inducible protein 10 (IP-10), a chemokine to attract CXCR3+ T helper 1-type CD4+ T cells, has not been investigated thoroughly in the pathogenesis of pancreatitis. B6 mice were inoculated intraperitoneally with LP-BM5 and then injected every week with either an antibody against IP-10 or a control antibody. Eight weeks after infection, we analyzed the effect of IP-10 neutralization. Anti-IP-10 antibody treatment did not change the generalized lymphadenopathy and hepatosplenomegaly of mice with MAIDS. The treatment significantly reduced the number of IP-10- and CXCR3-positive cells in the mesenteric lymph nodes (mLNs) but not the phenotypes and gross numbers of cells. In contrast, IP-10 neutralization reduced the number of mononuclear cells infiltrating into the pancreas. Anti-IP-10 antibody treatment did not change the numbers of IFN-gamma+ and IL10+ cells in the mLN but significantly reduced their numbers, especially IFN-gamma+ and IL-10+ CD4+ T cells and IFN-gamma+ Mac-1+ cells, in the pancreas. IP-10 neutralization ameliorated the pancreatic lesions of mice with MAIDS probably by blocking the cellular infiltration of CD4+ T cells and IFN-gamma+ Mac-1+ cells into the pancreas at least at 8 wk after infection, suggesting that IP-10 and these cells might play a key role in the development of chronic autoimmune pancreatitis.