RESUMO
INTRODUCTION: Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease worldwide. It can progress from simple steatosis to nonalcoholic steatohepatitis and may even develop into liver fibrosis, hepatocirrhosis, or hepatocellular carcinoma, but there is no effective treatment. MATERIAL AND METHODS: Wild-type (wt) and diabetic (db/db) mouse NAFLD-induced models were used to investigate the hepatoprotective effects and potential mechanisms of dapagliflozin (a new oral hypoglycaemic drug) on type 2 diabetes mellitus (T2DM) complicated with NAFLD, and to establish wt and db/db mouse NAFLD-induced and dapagliflozin treatment models. RESULTS: Dapagliflozin reduces blood glucose, glycosylated haemoglobin, blood lipids, and serum transaminase levels in db/db mice and improves T2DM-related liver injury accompanied by NAFLD; the mechanism may be related to the decrease in dipeptidyl-peptidase-4 (DPP4) protein expression and improvement in liver enzymes. Further mechanism-related studies by our team revealed that dapagliflozin can also downregulate the expression of DPP4 proteins in the liver and reduce serum soluble DPP4 enzyme levels, thereby improving the hepatic steatosis and insulin resistance of NAFLD. CONCLUSION: Dapagliflozin may be an effective drug for the treatment of T2DM-induced NAFLD and NAFLD, providing a reliable laboratory basis and new treatment methods for the clinical treatment of NAFLD.
Assuntos
Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Hepatopatia Gordurosa não Alcoólica , Inibidores do Transportador 2 de Sódio-Glicose , Camundongos , Animais , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/complicações , Hepatopatia Gordurosa não Alcoólica/metabolismo , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Dipeptidil Peptidase 4/uso terapêutico , FígadoRESUMO
OBJECTIVE: To investigate the protective effects and molecular mechanisms of Lizhong decoction (, LZD) against non-alcoholic fatty liver disease (NAFLD). METHODS: Male Wistar rats were fed a high-fat diet for four weeks to induce NAFLD, and were administered LZD by gavage for four weeks. Potential therapeutic targets for NAFLD were analyzed using network pharmacology. Liver pathology was evaluated using Oil Red O and hematoxylin-eosin staining. Furthermore, mitochondrial function, lipid metabolism, oxidative stress, and inflammatory response were examined. RESULTS: Rats with NAFLD exhibited high levels of hepatic damage and cholesterol deposition. Moreover, apoptosis was increased, superoxide dismutase and glutathione content were reduced, malondialdehyde content was increased, and the protein expression of inflammatory cytokines and p-c-Jun N-terminal kinase was increased. The LZD treatment ameliorated mitochondrial dysfunction, reduced liver damage, inhibited oxidative stress and inflammatory response, upregulated peroxisome proliferator-activated receptor (PPAR)-γ expression, and suppressed dipeptidyl peptidase 4 (DPP4) expression in the liver. CONCLUSION: It was found that LZD alleviates NAFLD by activating PPAR-γ and inhibiting DPP4.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Animais , Dieta Hiperlipídica/efeitos adversos , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Dipeptidil Peptidase 4/uso terapêutico , Metabolismo dos Lipídeos , Fígado , Masculino , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/metabolismo , PPAR gama/genética , Ratos , Ratos WistarRESUMO
Background: Metastatic disease is a major cause of thyroid cancer-related death. However, the mechanisms responsible for thyroid cancer metastasis are unclear. Dipeptidyl peptidase-4 (DPP4) is a multifunctional cell surface glycoprotein that has been reported to be a negative prognostic factor in thyroid cancer. We explored the molecular mechanism of the role of DPP4 in thyroid cancer cell metastasis. Methods: The effects of DPP4 on thyroid cancer cell migration/invasion in vitro were assessed by transwell assays. A lung metastatic mouse model was also established to determine the effect of DPP4 on tumor metastasis in vivo. DPP4 inhibitor sitagliptin was used to test its effect on thyroid cancer cell metastasis. The mechanism of which DPP4 promotes thyroid cancer cell metastasis was explored by a series of molecular and biochemical experiments. Results: We observed that DPP4 was significantly upregulated in papillary thyroid cancers compared with control subjects, and its expression was positively associated with lymph node metastasis and BRAFV600E mutation. Functional studies showed that DPP4 knockdown significantly inhibited metastatic potential of thyroid cancer cells, and vice versa. However, DPP4 inhibitor sitagliptin did not affect the metastatic ability of thyroid cancer cells, indicating that the promoting effect of DPP4 on tumor metastasis was independent of its enzymatic activity. Mechanistically, DPP4 interacted with the α4 and ß1 integrin subunits, and stabilized the formation of integrin α4ß1 complex. DPP4-mediated integrin signal activation promoted the nuclear localization of c-Jun through the FAK/AKT pathway, thereby inducing the transcription of transforming growth factor-beta 1 (TGFB1 coding for protein TGF-ß1). TGF-ß1 then facilitated tumor metastasis by inducing the epithelial-mesenchymal transition. Conclusions: DPP4 promotes thyroid cancer cell metastasis through the integrins/FAK/AKT/c-Jun/TGF-ß1 signaling axis. These findings may have implications for an alternative therapeutic strategy for thyroid cancer.
Assuntos
Inibidores da Dipeptidil Peptidase IV , Neoplasias da Glândula Tireoide , Camundongos , Animais , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Dipeptidil Peptidase 4/genética , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Integrina alfa4beta1 , Inibidores da Dipeptidil Peptidase IV/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Transição Epitelial-Mesenquimal , Movimento Celular , Fosfato de Sitagliptina/farmacologia , Transdução de Sinais , Fatores de Crescimento Transformadores/farmacologia , Linhagem Celular TumoralRESUMO
INTRODUCTION: Human mesenchymal stromal cells (MSCs) have immunomodulatory, anti-inflammatory, and tolerogenic effects. Long-term in vitro expansion of MSCs to generate clinical grade products results in the accumulation of senescent-functionally impaired MSCs. Markers to assess the 'senescent load' of MSC products are needed. METHODS: Early and late passage human adipose tissue (AT) MSCs from pediatric and adult donors were characterized using established senescent markers [i.e., MSC size, granularity, and autofluorescence by flow cytometry; ß-galactosidase staining (SA-ß-gal); CDKN2A and CDKN1A by qRT-PCR]. In gene set enrichment analysis, DPP4 (also known as adenosine deaminase complexing protein 2 or CD26) was found as a prominent dysregulated transcript that was increased in late passage MSC(AT). This was confirmed in a larger number of MSC samples by PCR, flow cytometry, Western blotting, and immunofluorescence. In vitro immunopotency assays compared the function of CD26high and CD26low MSC(AT). The effect of senolytics on the CD26high subpopulation was evaluated in senescent MSC(AT). RESULTS: Late passage MSC(AT) had a senescence transcriptome signature. DPP4 was the most differentially enriched gene in senescent MSCs. Late passage senescent MSC(AT) had higher CD26 surface levels and total protein abundance. Moreover, CD26 surface levels were higher in early passage MSC(AT) from adults compared to pediatric donors. CD26 abundance correlated with established senescence markers. CD26high MSC(AT) had reduced immunopotency compared to CD26low MSC(AT). Senolytic treatment induced MSC apoptosis, which decreased the frequencies of CD26high MSC(AT). CONCLUSIONS: DPP4 gene expression and DPP4/CD26 protein abundance are markers of replicative senescence in MSC(AT). Samples enriched in CD26high MSC(AT) have reduced immunopotency and CD26high MSCs are reduced with senolytics.
Assuntos
Dipeptidil Peptidase 4 , Células-Tronco Mesenquimais , Tecido Adiposo/metabolismo , Adulto , Biomarcadores/metabolismo , Proliferação de Células/genética , Células Cultivadas , Senescência Celular , Criança , Dipeptidil Peptidase 4/genética , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Humanos , Células-Tronco Mesenquimais/metabolismoRESUMO
BACKGROUND: Adipose tissue inflammation is considered as one of the major mechanisms underlying the pathogenesis of insulin resistance and complications in diabetes. Here, we aimed to study the effects of adipose-derived stromal cells on diabetes-induced insulin resistance and M1 cytokine expression. METHODS: Stromal vascular fractions (SVFs) purified from the inguinal adipose tissue of diabetic mice were treated with plasma from either nondiabetic (Lepr+/+) or diabetic (Leprdb/db) mice and injected into the inguinal white adipose tissue of Leprdb/db mice. RESULTS: We found that diabetic plasma treatment induced, whereas nondiabetic plasma suppressed TNF-α, IL-1ß, and dipeptidyl peptidase 4 (DPP4) mRNA expression in SVFs in vitro. Importantly, the injection of nondiabetic plasma-treated SVFs significantly decreased TNF-α, IL-6, IL-1ß, CCL2, and IL-33 and induced IL-10 mRNA expression in adipose tissue of Leprdb/db mice in vivo. Furthermore, we observed that nondiabetic plasma-treated SVFs increased mRNA expression of Foxp3 in adipose tissue macrophages and Foxp3 in adipose CD4+ T cells, decreased CD11b+CD11c+ cells in adipose tissue, and suppressed mRNA expression of ICAM-1, FCM3, IL-6, IL-1ß, iNOS, TNF-α, and DPP4 as well as protein expression of DPP4 and phosphorylated JNK and NF-κB in the liver of Leprdb/db mice. Moreover, we found that nondiabetic plasma-treated SVFs increased Akt activation following insulin administration and attenuated glucose intolerance in Leprdb/db mice. CONCLUSIONS: Our results demonstrate that nondiabetic plasma inhibits M1 but increases M2 cytokine expression in adipose tissue of diabetic mice. Most importantly, our findings reveal that nondiabetic plasma-treated SVFs are capable of mitigating diabetes-induced plasma DPP4 activity, liver inflammation, and insulin resistance and that may be mediated through suppressing M1 cytokines but increasing IL-10 and Tregs in adipose tissue. Altogether, our findings suggest that adipose stromal cell-based therapy could potentially be developed as an efficient therapeutic strategy for the treatment of diabetes.
Assuntos
Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Resistência à Insulina , Tecido Adiposo/metabolismo , Animais , Citocinas/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/terapia , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Dipeptidil Peptidase 4/uso terapêutico , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/metabolismo , Inflamação/patologia , Resistência à Insulina/fisiologia , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/metabolismo , Células Estromais/metabolismo , Fator de Necrose Tumoral alfa/genéticaRESUMO
Rationale: Capillary leakage frequently occurs during sepsis and after major surgery and is associated with microvascular dysfunction and adverse outcome. Procalcitonin is a well-established biomarker in inflammation without known impact on vascular integrity. Objectives: We determined how procalcitonin induces endothelial hyperpermeability and how targeting procalcitonin protects vascular barrier integrity. Methods: In a prospective observational clinical study, procalcitonin levels were assessed in 50 patients who underwent cardiac surgery and correlated to postoperative fluid and vasopressor requirements along with sublingual microvascular functionality. Effects of the procalcitonin signaling pathway on endothelial barrier and adherens junctional integrity were characterized in vitro and verified in mice. Inhibition of procalcitonin activation by dipeptidyl-peptidase 4 (DPP4) was evaluated in murine polymicrobial sepsis and clinically verified in cardiac surgery patients chronically taking the DPP4 inhibitor sitagliptin. Measurements and Main Results: Elevated postoperative procalcitonin levels identified patients with 2-fold increased fluid requirements (P < 0.01), 1.8-fold higher vasopressor demand (P < 0.05), and compromised microcirculation (reduction to 63.5 ± 2.8% of perfused vessels, P < 0.05). Procalcitonin induced 1.4-fold endothelial and 2.3-fold pulmonary capillary permeability (both Ps < 0.001) by destabilizing VE-cadherin. Procalcitonin effects were dependent on activation by DPP4, and targeting the procalcitonin receptor or DPP4 during sepsis-induced hyperprocalcitonemia reduced capillary leakage by 54 ± 10.1% and 60.4 ± 6.9% (both Ps < 0.01), respectively. Sitagliptin before cardiac surgery was associated with augmented microcirculation (74.1 ± 1.7% vs. 68.6 ± 1.9% perfused vessels in non-sitagliptin-medicated patients, P < 0.05) and with 2.3-fold decreased fluid (P < 0.05) and 1.8-fold reduced vasopressor demand postoperatively (P < 0.05). Conclusions: Targeting procalcitonin's action on the endothelium is a feasible means to preserve vascular integrity during systemic inflammation associated with hyperprocalcitonemia.
Assuntos
Dipeptidil Peptidase 4 , Sepse , Animais , Permeabilidade Capilar , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Dipeptidil Peptidase 4/uso terapêutico , Endotélio Vascular/metabolismo , Inflamação/metabolismo , Camundongos , Pró-Calcitonina , Sepse/tratamento farmacológico , Sepse/metabolismoRESUMO
Catalpol (CA) is widely used in the protection of cardiomyocytes. Nevertheless, the mechanism of CA in alleviating ischemia-reperfusion-induced injury of cardiomyocytes remains unclear. Human cardiomyocyte AC16 cells were subjected to hypoxia/reoxygenation (H/R) injury. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blot analysis were applied to detect tumor necrosis factor-alpha (TNF-α) mRNA, interleukin-6 (IL-6) mRNA, interleukin-1beta (IL-1ß) mRNA, microRNA-22-3p (miR-22-3p), dipeptidyl peptidase 4 (DPP4) mRNA, and DPP4 protein expressions. The cell viability and apoptosis were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry, respectively. Lactate dehydrogenase (LDH) and creatine kinase (CK-MB) were examined by enzyme-linked immunosorbent assay (ELISA) kits. A dual-luciferase reporter gene assay was performed to confirm the binding sequence between miR-22-3p and DPP4 mRNA 3'-untranslated region (3'UTR). CA promoted the viability and reduced cell apoptosis of AC16 cells and repressed the release of inflammatory cytokines TNF-α, IL-6, and IL-1ß, and inhibited the leakage of myocardial injury markers LDH and CK-MB. Furthermore, CA enhanced the expression of miR-22-3p in cardiomyocytes, and DPP4 was validated to be the target gene of miR-22-3p. The inhibition of miR-22-3p and augmentation of DPP4 reversed the above effects of CA. CA protects A16 cells from H/R injury by regulating the miR-22-3p/DPP4 axis.
Assuntos
MicroRNAs , Traumatismo por Reperfusão , Regiões 3' não Traduzidas , Apoptose , Dipeptidil Peptidase 4/genética , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Humanos , Hipóxia/metabolismo , Interleucina-6/metabolismo , Glucosídeos Iridoides , MicroRNAs/metabolismo , Miócitos Cardíacos/metabolismo , Traumatismo por Reperfusão/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
ß-Boswellic acid (ß-BA) and 11-keto-ß-boswellic acid (ß-KBA) are crucial bioactive compounds, mostly isolated from frankincense. These compounds are known for their potent anticancer and anti-inflammatory activities. Herein, we have explored the complete anti-diabetic potential of ß-BA and ß-KBA with detailed parameters. This research revealed that treatment with ß-BA and ß-KBA at a dose of 1, 2, and 10 mg/kg body weight for 21 days significantly improved body weight loss, water consumption, and specifically the concentration of blood glucose level (BGL) in diabetic animals, which indicated that the ß-BA and ß-KBA possess strong anti-diabetic activities. Serum total superoxide dismutase (SOD) and malondialdehyde (MDA) assays were also performed to evaluate the antioxidant effects. The biochemical analysis revealed that these compounds improve an abnormal level of several biochemical parameters like serum lipid values including total cholesterol (TC), triacylglycerol (TG), low-density lipoprotein cholesterol (LDL-C) to a normal level and the high-density lipoprotein cholesterol level (HDL-C). To understand the mechanism of action of ß-BA and ß-KBA, their most probable biological targets were searched through the inverse docking approach. Our computational analysis reflects that among other probable targets, the Dipeptidyl peptidase 4 (DPP-4) enzyme could be one of the possible binders of ß-BA and ß-KBA to produce their anti-diabetic activities. These in-silico results were validated by an in-vitro experiment. It indicates that the anti-diabetic effects of ß-BA and ß-KBA are produced by the inhibition of DDP-4. Thus, these anti-diabetic, antioxidant, and anti-hyperlipidemic effects of ß-BA and ß-KBA suggest these compounds as potential therapeutics for diabetic conditions.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Extratos Vegetais/farmacologia , Triterpenos/farmacologia , Animais , Glicemia/efeitos dos fármacos , Boswellia , Dipeptidil Peptidase 4/farmacologia , Relação Dose-Resposta a Droga , Lipídeos/sangue , Malondialdeído/metabolismo , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Estreptozocina , Superóxido Dismutase/efeitos dos fármacos , Triterpenos/administração & dosagem , Redução de Peso/efeitos dos fármacosRESUMO
Rheumatoid arthritis (RA) is an autoimmune disorder that is characterized by increasing levels of proinflammatory cytokines. The ubiquitous enzyme dipeptidyl peptidase-4 (DPP4, also known as CD26) regulates different immune disorders, although the effects of DPP4 in RA are uncertain. Here, we found lower levels of DPP4 in RA synovial tissues compared with normal tissues. DPP4 levels were also lower in a rat collagen-induced arthritis model than in control (healthy) rats. Overexpression of DPP4 or exogenous treatment of RA synovial fibroblasts with DPP4 reduced levels of proinflammatory interleukin (IL)-1ß, IL-6, and IL-13, and increased anti-inflammatory IL-10 synthesis, while DPP4 inhibitors sitagliptin and vildagliptin increased proinflammatory cytokine production, indicating an enhanced risk of RA development. The evidence suggests that increasing DPP4 expression is a novel strategy for RA disease.
Assuntos
Artrite Reumatoide/tratamento farmacológico , Citocinas/efeitos dos fármacos , Dipeptidil Peptidase 4 , Fibroblastos/efeitos dos fármacos , Animais , Artrite Reumatoide/metabolismo , Citocinas/metabolismo , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/farmacologia , Fibroblastos/metabolismo , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/metabolismoRESUMO
The biological influence of antidiabetic drugs on cancer cells and diabetic cancer patients has not yet been completely elucidated. We reported that a dipeptidyl peptidase (DPP)-4 inhibitor accelerates mammary cancer metastasis by inducing epithelial-mesenchymal transition (EMT) through the CXCL12/CXCR4/mTOR axis. Metformin has been shown to inhibit the mTOR signaling pathway. In this study, we investigated whether metformin mitigates breast cancer metastasis induced by a DPP-4 inhibitor via suppression of mTOR signaling. In cultured mouse mammary and human breast cancer cells, metformin suppressed DPP-4 inhibitor KR62436 (KR)-induced EMT and cell migration via suppression of the mTOR pathway associated with AMPK activation. For the in vivo study, metformin intervention was performed in an allograft 4T1 breast cancer model mouse with or without KR. We also analyzed mice transplanted with shRNA-mediated DPP-4 knockdown 4T1 cells. Treatment with metformin inhibited the lung metastasis of DPP-4-deficient 4T1 mammary tumor cells generated by either KR administration or DPP-4 knockdown. Immunostaining of primary tumors indicated that DPP-4 suppression promoted the expression of EMT-inducing transcription factor Snail through activation of the CXCR4-mediated mTOR/p70S6K pathway in an allograft breast cancer model; metformin abolished this alteration. Metformin treatment did not alter DPP-4-deficiency-induced expression of CXCL12 in either plasma or primary tumors. Our findings suggest that metformin may serve as an antimetastatic agent by mitigating the undesirable effects of DPP-4 inhibitors in patients with certain cancers. IMPLICATIONS: Metformin could combat the detrimental effects of DPP-4 inhibitor on breast cancer metastasis via mTOR suppression, suggesting the potential clinical relevance. VISUAL OVERVIEW: http://mcr.aacrjournals.org/content/molcanres/19/1/61/F1.large.jpg.
Assuntos
Neoplasias da Mama/induzido quimicamente , Dipeptidil Peptidase 4/efeitos adversos , Perfilação da Expressão Gênica/métodos , Metformina/uso terapêutico , Serina-Treonina Quinases TOR/metabolismo , Animais , Neoplasias da Mama/patologia , Dipeptidil Peptidase 4/farmacologia , Feminino , Humanos , Metformina/farmacologia , Camundongos , Metástase Neoplásica , Transdução de SinaisRESUMO
As more and more protein biotherapeutics enter the drug discovery pipelines, there is an increasing interest in tools for mechanistic drug metabolism investigations of biologics in order to identify and prioritize the most promising candidates. Understanding or even predicting the in vivo clearance of biologics and to support translational pharmacokinetic modeling activities is essential, however there is a lack of effective and validated in vitro cellular tools. Although different mechanisms have to be adressed in the context of biologics disposition, the scope is not comparable to the nowadays widely established tools for early characterization of small molecule disposition. Here, we describe a biotransformation study of the fusion protein tetranectin apolipoprotein A1 by cellular systems. The in vivo biotransformation of tetranectin apolipoprotein A1 has been described previously, and the same major biotransformation product could also be detected in vitro, by a targeted and highly sensitive detection method based on chymotrypsin digest. In addition, the protease responsible for the formation of this biotransformation product could be elucidated to be DPP4. To our knowledge, this is one of the first reports of an in vitro biotransformation study by cells of a therapeutic protein.
Assuntos
Apolipoproteína A-I/genética , Biotransformação/genética , Dipeptidil Peptidase 4/química , Lectinas Tipo C/genética , Proteínas Recombinantes de Fusão/genética , Apolipoproteína A-I/química , Quimotripsina/farmacologia , Dipeptidil Peptidase 4/farmacologia , Descoberta de Drogas , Humanos , Lectinas Tipo C/química , Processamento de Proteína Pós-Traducional , Proteômica/métodos , Proteínas Recombinantes de Fusão/químicaRESUMO
Compared to neutrophil chemoattractants, relatively little is known about the mechanism neutrophils use to respond to chemorepellents. We previously found that the soluble extracellular protein dipeptidyl peptidase IV (DPPIV) is a neutrophil chemorepellent. In this report, we show that an inhibitor of the protease activated receptor 2 (PAR2) blocks DPPIV-induced human neutrophil chemorepulsion, and that PAR2 agonists such as trypsin, tryptase, 2f-LIGRL, SLIGKV, and AC55541 induce human neutrophil chemorepulsion. Several PAR2 agonists in turn block the ability of the chemoattractant fMLP to attract neutrophils. Compared to neutrophils from male and female C57BL/6 mice, neutrophils from male and female mice lacking PAR2 are insensitive to the chemorepulsive effects of DPPIV or PAR2 agonists. Acute respiratory distress syndrome (ARDS) involves an insult-mediated influx of neutrophils into the lungs. In a mouse model of ARDS, aspiration of PAR2 agonists starting 24 h after an insult reduce neutrophil numbers in the bronchoalveolar lavage (BAL) fluid, as well as the post-BAL lung tissue. Together, these results indicate that the PAR2 receptor mediates DPPIV-induced chemorepulsion, and that PAR2 agonists might be useful to induce neutrophil chemorepulsion.
Assuntos
Dipeptidil Peptidase 4/farmacologia , Pulmão/imunologia , Neutrófilos/imunologia , Receptor PAR-2/fisiologia , Síndrome do Desconforto Respiratório/imunologia , Tripsina/farmacologia , Triptases/farmacologia , Animais , Células Cultivadas , Quimiotaxia de Leucócito , Modelos Animais de Doenças , Feminino , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Síndrome do Desconforto Respiratório/tratamento farmacológico , Síndrome do Desconforto Respiratório/metabolismoRESUMO
Salmon gelatin (Salmo salar, SG) enzymatic hydrolysates were generated using Alcalase 2.4L, Alcalase 2.4L in combination with Flavourzyme 500L, Corolase PP, Promod 144MG and Brewer's Clarex. The hydrolysate generated with Corolase PP for 1h (SG-C1) had the highest angiotensin converting enzyme (ACE, IC50=0.13±0.05mgmL-1) and dipeptidyl peptidase IV (DPP-IV, IC50=0.08±0.01mgmL-1) inhibitory activities, and oxygen radical absorbance capacity (ORAC, 540.94±9.57µmolTEg-1d.w.). The in vitro bioactivities of SG-C1 were retained following simulated gastrointestinal digestion. Administration of SG and SG-C1 (50mgkg-1 body weight) to spontaneously hypertensive rats (SHR) lowered heart rate along with systolic, diastolic and mean arterial blood pressure. The SG-C1 hydrolysate was fractionated using semi-preparative RP-HPLC and the fraction with highest overall in vitro bioactivity (fraction 25) was analysed by UPLC-MS/MS. Four peptide sequences (Gly-Gly-Pro-Ala-Gly-Pro-Ala-Val, Gly-Pro-Val-Ala, Pro-Pro and Gly-Phe) and two free amino acids (Arg and Tyr) were identified in this fraction. These peptides and free amino acids had potent ACE and DPP-IV inhibitory, and ORAC activities. The results show that SG hydrolysates have potential as multifunctional food ingredients particularly for the management of hypertension.
Assuntos
Anti-Hipertensivos , Antioxidantes , Dipeptidil Peptidase 4 , Proteínas de Peixes , Peptídeos , Salmo salar , Animais , Anti-Hipertensivos/química , Anti-Hipertensivos/farmacologia , Antioxidantes/química , Antioxidantes/farmacologia , Produtos Biológicos/química , Produtos Biológicos/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Dipeptidil Peptidase 4/química , Dipeptidil Peptidase 4/farmacologia , Proteínas de Peixes/química , Proteínas de Peixes/farmacologia , Gelatina , Masculino , Peptídeos/química , Peptídeos/farmacologia , Hidrolisados de Proteína/química , Ratos , Ratos Endogâmicos SHR , Espectrometria de Massas em TandemRESUMO
An increasing prevalence of diabetes is known as a main risk for human health in the last future worldwide. There is limited evidence on the potential management of type 2 diabetes mellitus using bioactive peptides from marine organisms, besides from milk and beans. We summarized here recent advances in our understanding of the regulation of glucose metabolism using bioactive peptides from natural proteins, including regulation of insulin-regulated glucose metabolism, such as protection and reparation of pancreatic ß-cells, enhancing glucose-stimulated insulin secretion and influencing the sensitivity of insulin and the signaling pathways, and inhibition of bioactive peptides to dipeptidyl peptidase IV, α-amylase and α-glucosidase activities. The present paper tried to understand the underlying mechanism involved and the structure characteristics of bioactive peptides responsible for its antidiabetic activities to prospect the utilization of rich marine organism proteins.
Assuntos
Organismos Aquáticos/química , Diabetes Mellitus Tipo 2/tratamento farmacológico , Peptídeos/farmacologia , Glicemia/efeitos dos fármacos , Dipeptidil Peptidase 4/química , Dipeptidil Peptidase 4/farmacologia , Glucose/metabolismo , Humanos , Hipoglicemiantes/química , Hipoglicemiantes/farmacologia , Insulina/metabolismo , Peptídeos/química , alfa-Amilases/química , alfa-Amilases/farmacologia , alfa-Glucosidases/química , alfa-Glucosidases/farmacologiaRESUMO
Multifunctional tailorable composite systems, specifically designed for oral dual-delivery of a peptide (glucagon-like peptide-1) and an enzymatic inhibitor (dipeptidyl peptidase 4 (DPP4)), were assembled through the microfluidics technique. Both drugs were coloaded into these systems for a synergistic therapeutic effect. The systems were composed of chitosan and cell-penetrating peptide modified poly(lactide-co-glycolide) and porous silicon nanoparticles as nanomatrices, further encapsulated in an enteric hydroxypropylmethylcellulose acetylsuccinate polymer. The developed multifunctional systems were pH-sensitive, inherited by the enteric polymer, enabling the release of the nanoparticles only in the simulated intestinal conditions. Moreover, the encapsulation into this polymer prevented the degradation of the nanoparticles' modifications. These nanoparticles showed strong and higher interactions with the intestinal cells in comparison with the nonmodified ones. The presence of DPP4 inhibitor enhanced the peptide permeability across intestinal cell monolayers. Overall, this is a promising platform for simultaneously delivering two drugs from a single formulation. Through this approach peptides are expected to increase their bioavailability and efficiency in vivo both by their specific release at the intestinal level and also by the reduced enzymatic activity. The use of this platform, specifically in combination of the two antidiabetic drugs, has clinical potential for the therapy of type 2 diabetes mellitus.
Assuntos
Dipeptidil Peptidase 4/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Microfluídica/métodos , Nanopartículas/química , Células CACO-2 , Sobrevivência Celular/efeitos dos fármacos , Peptídeos Penetradores de Células/química , Quitosana/química , Técnicas de Cocultura , Dipeptidil Peptidase 4/química , Dipeptidil Peptidase 4/farmacologia , Composição de Medicamentos/métodos , Liberação Controlada de Fármacos , Sinergismo Farmacológico , Peptídeo 1 Semelhante ao Glucagon/química , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Células HT29 , Humanos , Concentração de Íons de Hidrogênio , Cinética , Metilcelulose/análogos & derivados , Metilcelulose/química , Nanopartículas/ultraestrutura , Permeabilidade , Poliglactina 910/química , Porosidade , Silício/químicaRESUMO
CD26 is a 110-kDa multifunctional molecule having dipeptidyl peptidase IV (DPPIV) enzyme activity and is present on the surface of human T cells. Soluble CD26 (sCD26) exists in human blood and enhances the proliferation of peripheral T lymphocytes induced by tetanus toxoid (TT). The mechanisms by which CD26 enhances the activation of T cells and monocytes remain to be fully elucidated. In this study, we compared the stimulation of THP-1 cells and isolated human monocytes with a combination of recombinant sCD26 and lipopolysaccharide (LPS) and the stimulation of these cells with LPS alone. We found that addition of sCD26 increased TNF-α and IL-6 mRNA and protein expression and enhanced ERK1/2 levels in the cytosol as well as c-Fos, NF-κB p50, NF-κB p65, and CUX1 levels in the nuclei of these cells. On the other hand, the selective DPPIV inhibitor sitagliptin inhibited the increase in TNF-α mRNA and protein expression as well as the increase in ERK, c-Fos, NF-κB p50, NF-κB p65, and CUX1 levels. However, it did not inhibit the increase in IL-6 mRNA and protein expression. We then demonstrated that sCD26 enhanced binding of transcription factors to the TNF- and IL-6 promoters and used reporter assays to demonstrate that transcription factor binding enhanced promoter activity. Once again, we observed differential activities at the TNF- and IL-6 promoters. Finally, we demonstrated that CUX-1 overexpression enhanced TNF- production on sCD26/LPS stimulation, while CUX-1 depletion had no effect. Neither CUX-1 overexpression nor CUX-1 depletion had an effect on IL-6 stimulation. These results are discussed in the context of a model that describes the mechanisms by which stimulation of monocytic cells by sCD26 and LPS leads to elevation of TNF- and IL-6 expression. CUX-1 is identified as a new transcription factor that differently regulates the activities of the TNF- and IL-6 promoters.
Assuntos
Dipeptidil Peptidase 4/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Interleucina-6/biossíntese , Monócitos/metabolismo , Regiões Promotoras Genéticas , Transcrição Gênica/efeitos dos fármacos , Fator de Necrose Tumoral alfa/biossíntese , Adulto , Linhagem Celular Tumoral , Feminino , Humanos , Lipopolissacarídeos/farmacologia , Masculino , Monócitos/citologia , Fatores de Transcrição/metabolismoRESUMO
Glucagon-like peptide-1 (GLP-1) [GLP-1 (7-36)-amide] plays a fundamental role in regulating postprandial nutrient metabolism. GLP-1 acts through a G-protein-coupled receptor present on the membranes of many tissues, including myocardium and endothelium. GLP-1 is cleaved by the dipeptidyl peptidase-4 enzyme to its metabolite GLP-1 (9-36)-amide within 1-2 min of its release into the circulation. Investigations have been done in humans and in animal models to determine whether GLP-1 has effects on the myocardium. Infusions of GLP-1 increase cardiac function in ischemic and non-ischemic cardiovascular disease. In humans and animal models, constant infusions of GLP-1 decrease the size of infarction and improve myocardial function in ischemic/reperfusion injury. In cardiomyopathy and heart failure, infusions of GLP-1 improve myocardial function. These beneficial effects of GLP-1 on cardiac function are mediated by both GLP-1 receptor activation and GLP-1 receptor independent actions. Infusions of the metabolite GLP-1 (9-36)-amide improve cardiac function in experimental animals with cardiovascular disease even though the metabolite does not bind to the GLP-1 receptor. The beneficial effects of GLP-1 on the heart occur in the presence of a GLP-1 receptor antagonist and in animals devoid of GLP-1 receptors. Preliminary data in animals with available GLP-1 receptor agonists and cardiac disease suggest that exenatide has beneficial effects in porcine models of ischemic heart disease. The animal data with liraglutide are inconclusive. Clinical trials with exenatide and liraglutide show significant improvements in weight, systolic blood pressure, lipid profiles, and other cardiovascular risk factors. Whether these will decrease cardiovascular events is currently under investigation.
Assuntos
Doenças Cardiovasculares/tratamento farmacológico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Angiopatias Diabéticas/tratamento farmacológico , Dipeptidil Peptidase 4/farmacologia , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Peptídeos/farmacologia , Peçonhas/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/fisiopatologia , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/fisiopatologia , Angiopatias Diabéticas/sangue , Angiopatias Diabéticas/fisiopatologia , Dipeptidil Peptidase 4/uso terapêutico , Cães , Exenatida , Feminino , Peptídeo 1 Semelhante ao Glucagon/uso terapêutico , Humanos , Lipídeos/sangue , Liraglutida , Masculino , Camundongos , Peptídeos/uso terapêutico , Ratos , Peçonhas/uso terapêuticoRESUMO
The multiple physiological characterizations of glucagon-like peptide-1 (GLP-1) make it a promising drug candidate for the therapy of type 2 diabetes. However, the biological half-life of GLP-1 is short in vivo due to degradation by dipeptidyl peptidase-IV (DPP-IV) and renal clearance. The stabilization of GLP-1 is critical for its utility in drug development. In this study, several GLP-1 mutants containing an N-terminal cyclic conformation were prepared in that the existence of cyclic conformation is predicted to increase the stabilization of GLP-1 in vivo. In this study, the binding capacities of the mutants were determined, the stabilities of the mutants were investigated and the physiological functions of the mutants were compared with those of wild-type GLP-1 in animals. The results indicated that the mutant (GLP1N8) remarkably raised the half-life in vivo; it also showed better glucose tolerance and higher HbA(1c) reduction than GLP-1 and exendin-4 in rodents. These results suggest that the GLP-1 analog (GLP1N8) which contains an N-terminal cyclic structure might be utilized as possible potent anti-diabetic drugs in the treatment of type 2 diabetes mellitus.
Assuntos
Glicemia/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Dipeptidil Peptidase 4/farmacologia , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Hemoglobinas Glicadas/efeitos dos fármacos , Hipoglicemiantes/farmacologia , Animais , Diabetes Mellitus Tipo 2/tratamento farmacológico , Desenho de Fármacos , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Teste de Tolerância a Glucose , Meia-Vida , Injeções Subcutâneas , Masculino , RatosRESUMO
Glucagon-like peptide-1 (GLP-1) was once considered as an ideal anti-diabetic candidate for its important role in maintaining glucose homeostasis through the regulation of islet hormone secretion, as well as hepatic and gastric function. However, the major therapeutic obstacle for using native GLP-1 as a therapeutic agent is its very short half-life primarily due to their degradation by the enzyme dipeptidyl peptidase IV (DPP-IV). In this study, GLP-1 analogues with modifications in amino acid site 8, 22 and 23 were synthesized using solid phase peptide synthesis. Resistance of these analogues to DPP-IV cleavage was investigated in vitro by incubation of the peptides with DPP-IV or human plasma. Glucoregulating efficacy of the analogues was evaluated in normal Kunming mice using intraperitoneal glucose tolerance model. Glucose lowering effect of combination therapy (analogue plus Vildagliptin) has also been studied. In vitro studies showed that the modified analogues were much more stable than native GLP-1 (nearly 100% of the peptide keep intact after 4 h incubation). In vivo biological activity evaluation revealed that His8-EEE (the most potent GLP-1 analogues in this study) exhibited significantly improved glycemic control potency (approximately 4.1-fold over saline and 2.5-fold over GLP-1) and longer time of active duration (at least 5 h). Combination therapy also showed the trend of its superiority over mono-therapy. Modified analogues showed increased potency and biological half-time compared with the native GLP-1, which may help to understand the structure-activity relationship of GLP-1 analogues.
Assuntos
Dipeptidil Peptidase 4/metabolismo , Peptídeo 1 Semelhante ao Glucagon/farmacocinética , Peptídeo 1 Semelhante ao Glucagon/uso terapêutico , Adamantano/administração & dosagem , Adamantano/análogos & derivados , Adamantano/farmacocinética , Adamantano/uso terapêutico , Animais , Dipeptidil Peptidase 4/farmacologia , Combinação de Medicamentos , Avaliação Pré-Clínica de Medicamentos , Estabilidade de Medicamentos , Peptídeo 1 Semelhante ao Glucagon/administração & dosagem , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Teste de Tolerância a Glucose , Meia-Vida , Hipoglicemiantes/administração & dosagem , Hipoglicemiantes/farmacocinética , Hipoglicemiantes/uso terapêutico , Masculino , Camundongos , Nitrilas/administração & dosagem , Nitrilas/farmacocinética , Nitrilas/uso terapêutico , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Proteólise , Pirrolidinas/administração & dosagem , Pirrolidinas/farmacocinética , Pirrolidinas/uso terapêutico , Relação Estrutura-Atividade , VildagliptinaRESUMO
The biological effects of endomorphins (EMs) are short-lasting due to their rapid degradation by endogenous enzymes. Competing enzymatic degradation is an approach to prolong EM bioavailability. In the present study, a series of tetra- and tripeptides of similar to EMs structure was synthesized and tested in vitro and in vivo for their ability to inhibit degradation of EMs. The obtained results indicated that, among the series of analogs, the tetrapeptide Tyr-Pro-d-ClPhe-Phe-NH(2) and the tripeptide Tyr-Pro-Ala-NH(2), which did not bind to the µ-opioid receptors, were potent inhibitors of EM catabolism in rat brain homogenate. In vivo, these two peptides significantly prolonged the analgesic and antidepressant-like effects, induced by exogenous EMs, by blocking EM degrading enzymes. These new potent inhibitors may therefore increase the level and the half life of endogenous EMs and could be used in a new therapeutic strategy against pain and mood disorders, based on increasing of EM bioavailability.