RESUMO
Fibroblast growth factor 21 (FGF21) plays a key role in hepatic lipid metabolism and long-acting FGF21 analogs have emerged as promising drug candidates for the treatment of nonalcoholic steatohepatitis (NASH). It remains to characterize this drug class in translational animal models that recapitulate the etiology and hallmarks of human disease. To this end, we evaluated the long-acting FGF21 analog PF-05231023 in the GAN (Gubra Amylin NASH) diet-induced obese (DIO) and biopsy-confirmed mouse model of NASH. Male C57BL/6J mice were fed the GAN diet high in fat, fructose, and cholesterol for 34 wk before the start of the study. GAN DIO-NASH mice with biopsy-confirmed NAFLD Activity Score (NAS ≥5) and fibrosis (stage ≥F1) were biweekly administered with PF-05231023 (10 mg/kg sc) or vehicle (sc) for 12 wk. Vehicle-dosed chow-fed C57BL/6J mice served as healthy controls. Pre-to-post liver biopsy histopathological scoring was performed for within-subject evaluation of NAFLD Activity Score (NAS) and fibrosis stage. Terminal endpoints included quantitative liver histology and transcriptome signatures as well as blood and liver biochemistry. PF-05231023 significantly reduced body weight, hepatomegaly, plasma transaminases, and plasma/liver lipids in GAN DIO-NASH mice. Notably, PF-05231023 reduced both NAS (≥2-point improvement) and fibrosis stage (1-point improvement). Improvements in NASH and fibrosis severity were supported by reduced quantitative histological markers of steatosis, inflammation, and fibrogenesis as well as improvements in disease-associated liver transcriptome signatures. In conclusion, PF-05231023 reduces NASH and fibrosis severity in a translational biopsy-confirmed mouse model of NASH, supporting development of FGF21 analogs for the treatment of NASH.NEW & NOTEWORTHY It is unclear if long-acting FGF21 analogs have antifibrotic efficacy in NASH. We therefore profiled the clinically relevant FGF21 analog PF-05231023 in a translational diet-induced obese and biopsy-confirmed mouse model of NASH. We found PF-05231023 to exert hepatoprotective effects as indicated by notable improvements in plasma markers and histological hallmarks of NASH, including improved fibrosis stage. Collectively, the present study supports the continued exploration of long-acting FGF21 analogs for the treatment of NASH and other fibrotic diseases.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Camundongos , Animais , Masculino , Humanos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Cirrose Hepática/metabolismo , Camundongos Endogâmicos C57BL , Fígado/metabolismo , Obesidade/metabolismo , Dieta , Biópsia , Modelos Animais de Doenças , Dieta Hiperlipídica/efeitos adversosRESUMO
Hypertension is a critical comorbidity for progression of diabetic kidney disease (DKD). To facilitate the development of novel therapeutic interventions with the potential to control disease progression, there is a need to establish translational animal models that predict treatment effects in human DKD. The present study aimed to characterize renal disease and outcomes of standard of medical care in a model of advanced DKD facilitated by adeno-associated virus (AAV)-mediated renin overexpression in uninephrectomized (UNx) db/db mice. Five weeks after single AAV administration and 4 wk after UNx, female db/db UNx-ReninAAV mice received (PO, QD) vehicle, lisinopril (40 mg/kg), empagliflozin (20 mg/kg), or combination treatment for 12 wk (n = 17 mice/group). Untreated db/+ mice (n = 8) and vehicle-dosed db/db UNx-LacZAAV mice (n = 17) served as controls. End points included plasma, urine, and histomorphometric markers of kidney disease. Total glomerular numbers and individual glomerular volume were evaluated by whole kidney three-dimensional imaging analysis. db/db UNx-ReninAAV mice developed hallmarks of progressive DKD characterized by severe albuminuria, advanced glomerulosclerosis, and glomerular hypertrophy. Lisinopril significantly improved albuminuria, glomerulosclerosis, tubulointerstitial injury, and inflammation. Although empagliflozin alone had no therapeutic effect on renal endpoints, lisinopril and empagliflozin exerted synergistic effects on renal histological outcomes. In conclusion, the db/db UNx-ReninAAV mouse demonstrates good clinical translatability with respect to physiological and histological hallmarks of progressive DKD. The efficacy of standard of care to control hypertension and hyperglycemia provides a proof of concept for testing novel drug therapies in the model.NEW & NOTEWORTHY Translational animal models of diabetic kidney disease (DKD) are important tools in preclinical research and drug discovery. Here, we show that the standard of care to control hypertension (lisinopril) and hyperglycemia (empagliflozin) improves physiological and histopathological hallmarks of kidney disease in a mouse model of hypertension-accelerated progressive DKD. The findings substantiate hypertension and type 2 diabetes as essential factors in driving DKD progression and provide a proof of concept for probing novel drugs for potential nephroprotective efficacy in this model.
Assuntos
Anti-Hipertensivos/uso terapêutico , Compostos Benzidrílicos/uso terapêutico , Pressão Sanguínea/efeitos dos fármacos , Nefropatias Diabéticas/tratamento farmacológico , Glucosídeos/uso terapêutico , Hipertensão/tratamento farmacológico , Lisinopril/uso terapêutico , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Animais , Anti-Hipertensivos/farmacologia , Compostos Benzidrílicos/farmacologia , Nefropatias Diabéticas/complicações , Modelos Animais de Doenças , Feminino , Glucosídeos/farmacologia , Hipertensão/complicações , Lisinopril/farmacologia , Camundongos , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Resultado do TratamentoRESUMO
The pancreatic hormone amylin plays a central role in regulating energy homeostasis and glycaemic control by stimulating satiation and reducing food reward, making amylin receptor agonists attractive for the treatment of metabolic diseases. Amylin receptors consist of heterodimerized complexes of the calcitonin receptor and receptor-activity modifying proteins subtype 1-3 (RAMP1-3). Neuronal activation in response to amylin dosing has been well characterized, but only in selected regions expressing high levels of RAMPs. The current study identifies global brain-wide changes in response to amylin and by comparing wild type and RAMP1/3 knockout mice reveals the importance of RAMP1/3 in mediating this response. Amylin dosing resulted in neuronal activation as measured by an increase in c-Fos labelled cells in 20 brain regions, altogether making up the circuitry of neuronal appetite regulation (e.g., area postrema (AP), nucleus of the solitary tract (NTS), parabrachial nucleus (PB), and central amygdala (CEA)). c-Fos response was also detected in distinct nuclei across the brain that typically have not been linked with amylin signalling. In RAMP1/3 knockout amylin induced low-level neuronal activation in seven regions, including the AP, NTS and PB, indicating the existence of RAMP1/3-independent mechanisms of amylin response. Under basal conditions RAMP1/3 knockout mice show reduced neuronal activity in the hippocampal formation as well as reduced hippocampal volume, suggesting a role for RAMP1/3 in hippocampal physiology and maintenance. Altogether these data provide a global map of amylin response in the mouse brain and establishes the significance of RAMP1/3 receptors in relaying this response.
RESUMO
BACKGROUND: Animal models of non-alcoholic steatohepatitis (NASH) are important tools in preclinical research and drug discovery. Gubra-Amylin NASH (GAN) diet-induced obese (DIO) mice represent a model of fibrosing NASH. The present study directly assessed the clinical translatability of the model by head-to-head comparison of liver biopsy histological and transcriptome changes in GAN DIO-NASH mouse and human NASH patients. METHODS: C57Bl/6 J mice were fed chow or the GAN diet rich in saturated fat (40%), fructose (22%) and cholesterol (2%) for ≥38 weeks. Metabolic parameters as well as plasma and liver biomarkers were assessed. Liver biopsy histology and transcriptome signatures were compared to samples from human lean individuals and patients diagnosed with NASH. RESULTS: Liver lesions in GAN DIO-NASH mice showed similar morphological characteristics compared to the NASH patient validation set, including macrosteatosis, lobular inflammation, hepatocyte ballooning degeneration and periportal/perisinusoidal fibrosis. Histomorphometric analysis indicated comparable increases in markers of hepatic lipid accumulation, inflammation and collagen deposition in GAN DIO-NASH mice and NASH patient samples. Liver biopsies from GAN DIO-NASH mice and NASH patients showed comparable dynamics in several gene expression pathways involved in NASH pathogenesis. Consistent with the clinical features of NASH, GAN DIO-NASH mice demonstrated key components of the metabolic syndrome, including obesity and impaired glucose tolerance. CONCLUSIONS: The GAN DIO-NASH mouse model demonstrates good clinical translatability with respect to the histopathological, transcriptional and metabolic aspects of the human disease, highlighting the suitability of the GAN DIO-NASH mouse model for identifying therapeutic targets and characterizing novel drug therapies for NASH.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Animais , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Humanos , Fígado , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Obesidade/complicaçõesRESUMO
Nonalcoholic fatty liver disease (NAFLD) represents a spectrum of conditions ranging from simple steatosis (NAFL), over nonalcoholic steatohepatitis (NASH) with or without fibrosis, to cirrhosis with end-stage disease. The hepatic molecular events underlying the development of NAFLD and transition to NASH are poorly understood. The present study aimed to determine hepatic transcriptome dynamics in patients with NAFL or NASH compared with healthy normal-weight and obese individuals. RNA sequencing and quantitative histomorphometry of liver fat, inflammation and fibrosis were performed on liver biopsies obtained from healthy normal-weight ( n = 14) and obese ( n = 12) individuals, NAFL ( n = 15) and NASH ( n = 16) patients. Normal-weight and obese subjects showed normal liver histology and comparable gene expression profiles. Liver transcriptome signatures were largely overlapping in NAFL and NASH patients, however, clearly separated from healthy normal-weight and obese controls. Most marked pathway perturbations identified in both NAFL and NASH were associated with markers of lipid metabolism, immunomodulation, extracellular matrix remodeling, and cell cycle control. Interestingly, NASH patients with positive Sonic hedgehog hepatocyte staining showed distinct transcriptome and histomorphometric changes compared with NAFL. In conclusion, application of immunohistochemical markers of hepatocyte injury may serve as a more objective tool for distinguishing NASH from NAFL, facilitating improved resolution of hepatic molecular changes associated with progression of NAFLD. NEW & NOTEWORTHY Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease in Western countries. NAFLD is associated with the metabolic syndrome and can progress to the more serious form, nonalcoholic steatohepatitis (NASH), and ultimately lead to irreversible liver damage. Using gold standard molecular and histological techniques, this study demonstrates that the currently used diagnostic tools are problematic for differentiating mild NAFLD from NASH and emphasizes the marked need for developing improved histological markers of NAFLD progression.
Assuntos
Tecido Adiposo , Perfilação da Expressão Gênica/métodos , Inflamação , Cirrose Hepática , Fígado , Hepatopatia Gordurosa não Alcoólica , Obesidade , Tecido Adiposo/metabolismo , Tecido Adiposo/patologia , Índice de Massa Corporal , Progressão da Doença , Feminino , Humanos , Imuno-Histoquímica , Inflamação/imunologia , Inflamação/patologia , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica/diagnóstico , Hepatopatia Gordurosa não Alcoólica/metabolismo , Obesidade/diagnóstico , Obesidade/metabolismoRESUMO
BACKGROUND: Compounds in clinical development for nonalcoholic steatohepatitis (NASH) improve liver histopathology in diet-induced obese mouse models of biopsy-confirmed NASH. Since the biopsy section used for histopathological evaluation represents only < 1% of the whole mouse liver, we evaluated how well biopsy-based quantitative image analyses correlate to stereology-based whole-liver quantitative changes upon drug treatment. METHODS: Male leptin-deficient Lepob/Lepob mice were fed the Amylin liver NASH (AMLN) diet for 16 weeks before stratification into treatment groups using a biopsy-based evaluation of type I collagen αI (col1a1) levels. Mice were treated for 8 weeks with either vehicle (PO, QD), liraglutide (0.4 mg/kg, SC, QD), elafibranor (30 mg/kg, PO, QD) or INT-767 (10 mg/kg, PO, QD). Terminal quantitative histological assessment of liver lipid (hematoxylin-eosin staining), inflammation (galectin-3 immunohistochemistry (IHC); gal-3), and fibrosis (col1a1 IHC) was performed on terminal liver biopsies and compared with stereologically sampled serial sections spanning the medial, left and right lateral lobe of the liver. RESULTS: The distribution of liver lipid and fibrosis was markedly consistent across lobes, whereas inflammation showed some variability. While INT-767 and liraglutide significantly reduced total liver weight by 20 and 48%, respectively, elafibranor tended to exacerbate hepatomegaly in Lepob/Lepob-NASH mice. All three compounds markedly reduced biopsy-based relative liver lipid content. Elafibranor and INT-767 significantly reduced biopsy-based relative gal-3 levels (P < 0.001), whereas INT-767 and liraglutide tended to reduce relative col1a1 levels. When changes in liver weight was accounted for, both INT-767 and liraglutide significantly reduced biopsy-based total col1a1 content. Although minor differences in absolute and relative liver lipid, inflammation and fibrosis levels were observed across lobes, the interpretation of drug-induced effects were consistent with biopsy-based conclusions. Notably, the incorporation of changes in total liver mass revealed that liraglutide's efficacy reached statistical significances for all analyzed parameters. CONCLUSIONS: In conclusion, in-depth analyses of liver homogeneity demonstrated that drug-induced improvement in liver biopsy-assessed histopathology is representative for overall liver effects assessed using stereology. Importantly, these findings reveal how changes in whole-liver mass should be considered to provide a deeper understanding of apparent drug treatment efficacy in preclinical NASH studies.
Assuntos
Fígado/efeitos dos fármacos , Fígado/patologia , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/patologia , Obesidade/complicações , Animais , Ácidos e Sais Biliares/uso terapêutico , Biópsia , Peso Corporal/efeitos dos fármacos , Chalconas/uso terapêutico , Colágeno Tipo I/análise , Dieta Hiperlipídica , Galectina 3/análise , Polipeptídeo Amiloide das Ilhotas Pancreáticas/administração & dosagem , Leptina/deficiência , Lipídeos/análise , Liraglutida/uso terapêutico , Fígado/química , Cirrose Hepática/patologia , Masculino , Camundongos , Camundongos Obesos , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Tamanho do Órgão/efeitos dos fármacos , PPAR alfa/agonistas , PPAR delta/agonistas , Propionatos/uso terapêutico , Reprodutibilidade dos TestesRESUMO
BACKGROUND: There is a marked need for improved animal models of nonalcoholic steatohepatitis (NASH) to facilitate the development of more efficacious drug therapies for the disease. METHODS: Here, we investigated the development of fibrotic NASH in male Wistar rats fed a choline-deficient L-amino acid-defined (CDAA) diet with or without cholesterol supplementation for subsequent assessment of drug treatment efficacy in NASH biopsy-confirmed rats. The metabolic profile and liver histopathology were evaluated after 4, 8, and 12 weeks of dieting. Subsequently, rats with biopsy-confirmed NASH were selected for pharmacological intervention with vehicle, elafibranor (30 mg/kg/day) or obeticholic acid (OCA, 30 mg/kg/day) for 5 weeks. RESULTS: The CDAA diet led to marked hepatomegaly and fibrosis already after 4 weeks of feeding, with further progression of collagen deposition and fibrogenesis-associated gene expression during the 12-week feeding period. Cholesterol supplementation enhanced the stimulatory effect of CDAA on gene transcripts associated with fibrogenesis without significantly increasing collagen deposition. Pharmacological intervention with elafibranor, but not OCA, significantly reduced steatohepatitis scores, and fibrosis-associated gene expression, however, was unable to prevent progression in fibrosis scores. CONCLUSION: CDAA-fed rats develop early-onset progressive NASH, which offers the opportunity to probe anti-NASH compounds with potential disease-modifying properties.
Assuntos
Chalconas/uso terapêutico , Ácido Quenodesoxicólico/análogos & derivados , Colesterol/toxicidade , Hepatopatia Gordurosa não Alcoólica/induzido quimicamente , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Nutrientes/deficiência , Propionatos/uso terapêutico , Animais , Ácido Quenodesoxicólico/uso terapêutico , Colesterol/administração & dosagem , Progressão da Doença , Masculino , Hepatopatia Gordurosa não Alcoólica/patologia , Ratos , Ratos WistarRESUMO
AIMS/HYPOTHESIS: Enteroendocrine K and L cells are pivotal in regulating appetite and glucose homeostasis. Knowledge of their distribution in humans is sparse and it is unknown whether alterations occur in type 2 diabetes. We aimed to evaluate the distribution of enteroendocrine K and L cells and relevant prohormone-processing enzymes (using immunohistochemical staining), and to evaluate the mRNA expression of the corresponding genes along the entire intestinal tract in individuals with type 2 diabetes and healthy participants. METHODS: In this cross-sectional study, 12 individuals with type 2 diabetes and 12 age- and BMI-matched healthy individuals underwent upper and lower double-balloon enteroscopy with mucosal biopsy retrieval from approximately every 30 cm of the small intestine and from seven specific anatomical locations in the large intestine. RESULTS: Significantly different densities for cells positive for chromogranin A (CgA), glucagon-like peptide-1, glucose-dependent insulinotropic polypeptide, peptide YY, prohormone convertase (PC) 1/3 and PC2 were observed along the intestinal tract. The expression of CHGA did not vary along the intestinal tract, but the mRNA expression of GCG, GIP, PYY, PCSK1 and PCSK2 differed along the intestinal tract. Lower counts of CgA-positive and PC1/3-positive cells, respectively, were observed in the small intestine of individuals with type 2 diabetes compared with healthy participants. In individuals with type 2 diabetes compared with healthy participants, the expression of GCG and PYY was greater in the colon, while the expression of GIP and PCSK1 was greater in the small intestine and colon, and the expression of PCSK2 was greater in the small intestine. CONCLUSIONS/INTERPRETATION: Our findings provide a detailed description of the distribution of enteroendocrine K and L cells and the expression of their products in the human intestinal tract and demonstrate significant differences between individuals with type 2 diabetes and healthy participants. TRIAL REGISTRATION: NCT03044860.
Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Células Enteroendócrinas/metabolismo , Adulto , Idoso , Cromogranina A/metabolismo , Estudos Transversais , Feminino , Polipeptídeo Inibidor Gástrico/metabolismo , Trato Gastrointestinal/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Peptídeo YY/metabolismo , Pró-Proteína Convertase 1/metabolismo , Pró-Proteína Convertase 2/metabolismo , Pró-Proteína Convertases/metabolismoRESUMO
Novel principles for optimizing the properties of peptide-based drugs are needed in order to leverage their full pharmacological potential. We present the design, synthesis, and evaluation of a library of neoglycolipidated glucagon-like peptide 1 (GLP-1) analogues, which are valuable drug candidates for treatment of type 2 diabetes and obesity. Neoglycolipidation of GLP-1 balanced the lipophilicity, directed formation of soluble oligomers, and mediated albumin binding. Moreover, neoglycolipidation did not compromise bioactivity, as in vitro potency of neoglycolipidated GLP-1 analogues was maintained or even improved compared to native GLP-1. This translated into pronounced in vivo efficacy in terms of both decreased acute food intake and improved glucose homeostasis in mice. Thus, we propose neoglycolipidation as a novel, general method for modulating the properties of therapeutic peptides.
Assuntos
Albuminas/metabolismo , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacologia , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Glicolipídeos/sangue , Peptídeos/química , Animais , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Glucose/metabolismo , Teste de Tolerância a Glucose/métodos , Homeostase/efeitos dos fármacos , Hipoglicemiantes/química , Insulina/metabolismo , Masculino , Camundongos , Peptídeos/farmacologiaRESUMO
Bariatric surgery is currently the most effective treatment of obesity, which has spurred an interest in developing pharmaceutical mimetics. It is thought that the marked body weight-lowering effects of bariatric surgery involve stimulated secretion of appetite-regulating gut hormones, including glucagon-like peptide 1. We here report that intestinal expression of secretin is markedly upregulated in a rat model of Roux-en-Y gastric bypass, suggesting an additional role of secretin in the beneficial metabolic effects of Roux-en-Y gastric bypass. We therefore developed novel secretin-based peptide co-agonists and identified a lead compound, GUB06-046, that exhibited potent agonism of both the secretin receptor and glucagon-like peptide 1 receptor. Semi-acute administration of GUB06-046 to lean mice significantly decreased cumulative food intake and improved glucose tolerance. Chronic administration of GUB06-046 to diabetic db/db mice for 8 weeks improved glycemic control, as indicated by a 39% decrease in fasting blood glucose and 1.6% reduction of plasma HbA1c levels. Stereological analysis of db/db mice pancreata revealed a 78% increase in beta-cell mass after GUB06-046 treatment, with no impact on exocrine pancreas mass or pancreatic duct epithelial mass. The data demonstrate beneficial effects of GUB06-046 on appetite regulation, glucose homeostasis, and beta-cell mass in db/db mice, without proliferative effects on the exocrine pancreas and the pancreatic duct epithelium. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Índice Glicêmico/efeitos dos fármacos , Obesidade/tratamento farmacológico , Peptídeos/administração & dosagem , Secretina/química , Animais , Cirurgia Bariátrica , Proliferação de Células , Diabetes Mellitus Experimental/metabolismo , Modelos Animais de Doenças , Ingestão de Alimentos/efeitos dos fármacos , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Masculino , Camundongos , Obesidade/metabolismo , Obesidade/cirurgia , Peptídeos/farmacologia , Ratos , Receptores Acoplados a Proteínas G/agonistas , Receptores dos Hormônios Gastrointestinais/agonistas , Secretina/metabolismoRESUMO
Preterm infants often tolerate full enteral nutrition a few weeks after birth but it is not known how this is related to gut maturation. Using pigs as models, we hypothesized that intestinal structure and digestive function are similar in preterm and term individuals at 3-4 wk after birth and that early enteral nutrition promotes maturation. Preterm or term cesarean-delivered pigs were fed total parenteral nutrition, or partial enteral nutrition [Enteral (Ent), 16-64 ml·kg(-1)·day(-1) of bovine colostrum] for 5 days, followed by full enteral milk feeding until day 26 The intestine was collected for histological and biochemical analyses at days 0, 5, and 26 (n = 8-12 in each of 10 treatment groups). Intestinal weight (relative to body weight) was reduced in preterm pigs at 0-5 days but ENT feeding stimulated the mucosal volume and peptidase activities. Relative to term pigs, mucosal volume remained reduced in preterm pigs until 26 days although plasma glucagon-like peptide 2 (GLP-2) and glucose-dependent insulin-trophic peptide (GIP) levels were increased. Preterm pigs also showed reduced hexose absorptive capacity and brush-border enzyme (sucrase, maltase) activities at 26 days, relative to term pigs. Intestinal structure shows a remarkable growth adaptation in the first week after preterm birth, especially with enteral nutrition, whereas some digestive functions remain immature until at least 3-4 wk. It is important to identify feeding regimens that stimulate intestinal maturation in the postnatal period of preterm infants because some intestinal functions may show long-term developmental delay.
Assuntos
Digestão , Absorção Intestinal , Mucosa Intestinal/metabolismo , Animais , Animais Recém-Nascidos , Idade Gestacional , Peptídeo 2 Semelhante ao Glucagon/sangue , Hexoses/metabolismo , Mucosa Intestinal/crescimento & desenvolvimento , Mucosa Intestinal/patologia , Suínos , alfa-Fetoproteínas/metabolismoRESUMO
AIMS/HYPOTHESIS: We studied the impact of Roux-en-Y gastric bypass (RYGB) on the density and hormonal gene expression of small-intestinal enteroendocrine cells in obese patients with type 2 diabetes. METHODS: Twelve patients with diabetes and 11 age- and BMI-matched controls underwent RYGB followed by enteroscopy ~10 months later. Mucosal biopsies taken during surgery and enteroscopy were immunohistochemically stained for glucagon-like peptide-1 (GLP-1), peptide YY (PYY), cholecystokinin (CCK), glucose-dependent insulinotropic polypeptide (GIP) and prohormone convertase 2 (PC2) and the expression of GCG (encoding preproglucagon), PYY, CCK, GIP, GHRL (encoding ghrelin), SCT (encoding secretin), NTS (encoding neurotensin) and NR1H4 (encoding farnesoid X receptor) was evaluated. RESULTS: The density of cells immunoreactive for GLP-1, CCK and GIP increased in patients after RYGB and the density of those immunoreactive for GLP-1, PYY, CCK and PC2 increased in controls. In both groups, GHRL, SCT and GIP mRNA was reduced after RYGB while PYY, CCK, NTS and NR1H4 gene expression was unaltered. GCG mRNA was upregulated in both groups. CONCLUSIONS/INTERPRETATION: Numerous alterations in the distribution of enteroendocrine cells and their expression of hormonal genes are seen after RYGB and include increased density of GLP-1-, PYY-, CCK-, GIP- and PC2-positive cells, reduced gene expression of GHRL, SCT and GIP and increased expression of GCG.
Assuntos
Diabetes Mellitus Tipo 2/complicações , Células Enteroendócrinas/metabolismo , Derivação Gástrica , Obesidade Mórbida/cirurgia , Adulto , Colecistocinina/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/cirurgia , Feminino , Polipeptídeo Inibidor Gástrico/metabolismo , Grelina/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Obesidade Mórbida/complicações , Obesidade Mórbida/metabolismo , Peptídeo YY/metabolismo , Pró-Proteína Convertase 2/metabolismo , Resultado do TratamentoRESUMO
Neuromedin U (NMU) is a 25 amino acid peptide expressed and secreted in the brain and gastrointestinal tract. Data have shown that peripheral administration of human NMU decreases food intake and body weight and improves glucose tolerance in mice, suggesting that NMU receptors constitute a possible anti-diabetic and anti-obesity drug target. However, the clinical use of native NMU is hampered by a poor pharmacokinetic profile. In the current study, we report in vitro and in vivo data from a series of novel lipidated NMU analogs. In vitro plasma stability studies of native NMU were performed to investigate the proteolytic stability and cleavage sites using LC-MS. Native NMU was found to be rapidly cleaved at the C-terminus between Arg(24) and Asn(25) , followed by cleavage between Arg(16) and Gly(17) . Lipidated NMU analogs were generated using solid-phase peptide synthesis, and in vitro potency was investigated using a human embryonic kidney 293-based inositol phosphate accumulation assay. All lipidated analogs had preserved in vitro activity on both NMU receptors with potency improving as the lipidation site was moved away from the receptor-interacting C-terminal octapeptide segment. In vivo efficacy was assessed in lean mice as reduction in food intake after acute subcutaneous administration of 1, 0.3, 0.1, and 0.03 µmol/kg. These lipidated NMU analogs prolonged the anorectic effect of NMU in a dose-dependent manner. This was likely an effect of improved pharmacokinetic properties because of improved vitro plasma stability. Accordingly, the data demonstrate that lipidated NMU analogs may represent drug candidates for the treatment of obesity.
Assuntos
Ingestão de Alimentos/efeitos dos fármacos , Neuropeptídeos/síntese química , Neuropeptídeos/farmacologia , Animais , Células Cultivadas , Relação Dose-Resposta a Droga , Estabilidade de Medicamentos , Células HEK293 , Humanos , Masculino , Camundongos , Neuropeptídeos/sangue , Neuropeptídeos/química , Estabilidade ProteicaRESUMO
Antidiabetic treatments aiming to preserve or even to increase ß-cell mass are currently gaining increased interest. Here we investigated the effect of chronic treatment with the novel glucagon-like peptide-1 (GLP-1)-gastrin dual agonist ZP3022 (HGEGTFTSDLSKQMEEEAVRLFIEWLKN-8Ado-8Ado-YGWLDF-NH2) on glycemic control, ß-cell mass and proliferation, and islet number. Male db/db mice were treated with ZP3022, liraglutide, or vehicle for 2, 4, or 8 weeks, with terminal assessment of hemoglobin A1c, basal blood glucose, and plasma insulin concentrations. Pancreata were removed for immunohistochemical staining and stereological quantification of ß-cell mass, islet numbers, proliferation, and apoptosis. Treatment with ZP3022 or liraglutide led to a significant improvement in glycemic control. ZP3022 treatment resulted in a sustained increase in ß-cell mass after 4 and 8 weeks of treatment, whereas the effect of liraglutide was transient. The expansion in ß-cell mass observed in the ZP3022-treated mice appeared to be driven by increased ß-cell proliferation in existing islets rather than by formation of new islets, as mean islet mass increased but the number of islets remained constant. Our data demonstrate that the GLP-1-gastrin dual agonist ZP3022 causes a sustained improvement in glycemic control accompanied by an increase in ß-cell mass, increased proliferation, and increased mean islet mass. The results highlight that the GLP-1-gastrin dual agonist increases ß-cell mass more than liraglutide and that dual agonists could potentially be developed into a new class of antidiabetic treatments.
Assuntos
Gastrinas/agonistas , Peptídeo 1 Semelhante ao Glucagon/agonistas , Glucose/metabolismo , Homeostase/efeitos dos fármacos , Células Secretoras de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , Peptídeos/farmacologia , Sequência de Aminoácidos , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Hipoglicemiantes/farmacologia , Células Secretoras de Insulina/patologia , Ilhotas Pancreáticas/patologia , Masculino , Camundongos , Dados de Sequência MolecularRESUMO
Type 2 diabetes is characterized by impaired ß-cell function associated with progressive reduction of insulin secretion and ß-cell mass. Evidently, there is an unmet need for treatments with greater sustainability in ß-cell protection and antidiabetic efficacy. Through an insulin and ß cell-independent mechanism, empagliflozin, a specific sodium glucose cotransporter type 2 (SGLT-2) inhibitor, may potentially provide longer efficacy. This study compared the antidiabetic durability of empagliflozin treatment (10 mg/kg p.o.) against glibenclamide (3 mg/kg p.o.) and liraglutide (0.2 mg/kg s.c.) on deficient glucose homeostasis and ß-cell function in Zucker diabetic fatty (ZDF) rats. Empagliflozin and liraglutide led to marked improvements in fed glucose and hemoglobin A1c levels, as well as impeding a progressive decline in insulin levels. In contrast, glibenclamide was ineffective. Whereas the effects of liraglutide were less pronounced at week 8 of treatment compared with week 4, those of empagliflozin remained stable throughout the study period. Similarly, empagliflozin improved glucose tolerance and preserved insulin secretion after both 4 and 8 weeks of treatment. These effects were reflected by less reduction in ß-cell mass with empagliflozin or liraglutide at week 4, whereas only empagliflozin showed ß-cell sparing effects also at week 8. Although this study cannot be used to dissociate the absolute antidiabetic efficacy among the different mechanisms of drug action, the study demonstrates that empagliflozin exerts a more sustained improvement of glucose homeostasis and ß-cell protection in the ZDF rat. In comparison with other type 2 diabetic treatments, SGLT-2 inhibitors may through insulin-independent pathways thus enhance durability of ß-cell protection and antidiabetic efficacy.
Assuntos
Compostos Benzidrílicos/farmacologia , Glicemia/metabolismo , Diabetes Mellitus/sangue , Glucosídeos/farmacologia , Homeostase/fisiologia , Células Secretoras de Insulina/metabolismo , Transportador 2 de Glucose-Sódio/metabolismo , Animais , Compostos Benzidrílicos/uso terapêutico , Glicemia/efeitos dos fármacos , Diabetes Mellitus/tratamento farmacológico , Glucosídeos/uso terapêutico , Homeostase/efeitos dos fármacos , Células Secretoras de Insulina/efeitos dos fármacos , Masculino , Distribuição Aleatória , Ratos , Ratos Zucker , Inibidores do Transportador 2 de Sódio-GlicoseRESUMO
Short bowel syndrome (SBS) is a frequent complication after intestinal resection in infants suffering from intestinal disease. We tested whether treatment with the intestinotrophic hormone glucagon-like peptide-2 (GLP-2) increases intestinal volume and function in the period immediately following intestinal resection in preterm pigs. Preterm pigs were fed enterally for 48 h before undergoing resection of 50% of the small intestine and establishment of a jejunostomy. Following resection, pigs were maintained on total parenteral nutrition (TPN) without (SBS, n = 8) or with GLP-2 treatment (3.5 µg/kg body wt per h, SBS+GLP-2, n = 7) and compared with a group of unresected preterm pigs (control, n = 5). After 5 days of TPN, all piglets were fed enterally for 24 h, and a nutrient balance study was performed. Intestinal resection was associated with markedly reduced endogenous GLP-2 levels. GLP-2 increased the relative absorption of wet weight (46 vs. 22%), energy (79 vs. 64%), and all macronutrients (all parameters P < 0.05). These findings were supported by a 200% increase in sucrase and maltase activities, a 50% increase in small intestinal epithelial volume (P < 0.05), as well as increased DNA and protein contents and increased total protein synthesis rate in SBS+GLP-2 vs. SBS pigs (+100%, P < 0.05). Following intestinal resection in preterm pigs, GLP-2 induced structural and functional adaptation, resulting in a higher relative absorption of fluid and macronutrients. GLP-2 treatment may be a promising therapy to enhance intestinal adaptation and improve digestive function in preterm infants with jejunostomy following intestinal resection.
Assuntos
Peptídeo 2 Semelhante ao Glucagon/farmacologia , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/cirurgia , Nascimento Prematuro , Síndrome do Intestino Curto/tratamento farmacológico , Adaptação Fisiológica , Fenômenos Fisiológicos da Nutrição Animal , Animais , Animais Recém-Nascidos , Proliferação de Células/efeitos dos fármacos , Replicação do DNA/efeitos dos fármacos , Modelos Animais de Doenças , Nutrição Enteral , Idade Gestacional , Peptídeo 2 Semelhante ao Glucagon/sangue , Humanos , Absorção Intestinal/efeitos dos fármacos , Intestino Delgado/crescimento & desenvolvimento , Intestino Delgado/metabolismo , Intestino Delgado/patologia , Jejunostomia , Estado Nutricional , Nutrição Parenteral Total , Biossíntese de Proteínas/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Síndrome do Intestino Curto/sangue , Síndrome do Intestino Curto/etiologia , Síndrome do Intestino Curto/patologia , Síndrome do Intestino Curto/fisiopatologia , Sacarase/metabolismo , Suínos , Fatores de Tempo , Aumento de Peso/efeitos dos fármacos , alfa-Glucosidases/metabolismoRESUMO
A possible association between glucagon-like peptide-1 (GLP-1) analogs and incidences of pancreatitis has been suggested based on clinical studies. In male and female diabetic Zucker diabetic fatty (ZDF) rats, we investigated the effects of continuous administration of liraglutide and exenatide on biochemical [lipase, pancreatic amylase (P-amylase)] and histopathological markers of pancreatitis. Male and female ZDF rats were dosed for 13 wk with liraglutide (0.4 or 1.0 mg·kg(-1)·day(-1) sc once daily) or exenatide (0.25 mg·kg(-1)·day(-1) sc, Alzet osmotic minipumps). P-amylase and lipase plasma activity were measured, and an extended histopathological and stereological (specific cell mass and proliferation rate) evaluation of the exocrine and the endocrine pancreas was performed. Expectedly, liraglutide and exenatide lowered blood glucose and Hb A(1c) in male and female ZDF rats, whereas ß-cell mass and proliferation rate were increased with greatly improved blood glucose control. Whereas neither analog affected lipase activity, small increases in P-amylase activity were observed in animals treated with liraglutide and exenatide. However, concurrent or permanent increases in lipase and P-amylase activity were never observed. Triglycerides were lowered by both GLP-1 analogs. The qualitative histopathological findings did not reveal adverse effects of liraglutide. The findings were mainly minimal in severity and focal in distribution. Similarly, the quantitative stereological analyses revealed no effects of liraglutide or exenatide on overall pancreas weight or exocrine and duct cell mass or proliferation. The present study demonstrates that, in overtly diabetic male and female ZDF rats, prolonged exposure to GLP-1 receptor agonists does not affect biochemical or histopathological markers of pancreatitis, and whereas both exenatide and liraglutide increase ß-cell mass, they have no effect on the exocrine pancreas. However, clinical outcome studies and studies using primate tissues and/or studies in nonhuman primates are needed to further assess human risk.
Assuntos
Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Hipoglicemiantes/efeitos adversos , Pâncreas/efeitos dos fármacos , Pancreatite/induzido quimicamente , Animais , Glicemia/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Exenatida , Feminino , Peptídeo 1 Semelhante ao Glucagon/administração & dosagem , Peptídeo 1 Semelhante ao Glucagon/efeitos adversos , Hemoglobinas Glicadas/análise , Hipoglicemiantes/administração & dosagem , Lipase/sangue , Liraglutida , Masculino , Pâncreas/patologia , alfa-Amilases Pancreáticas/sangue , Pancreatite/patologia , Peptídeos/administração & dosagem , Peptídeos/efeitos adversos , Ratos , Ratos Zucker , Peçonhas/administração & dosagem , Peçonhas/efeitos adversosRESUMO
AIM: To validate the gubra DIO-rats as a useful animal model of human obesity. METHODS: The gubra diet-induced obesity (DIO) rat model was based on male Sprague-Dawley rats with ad libitum access to regular chow and a palatable diet rich in fat and sugar. To evaluate the versatility of the gubra DIO-rats as a valid model of human obesity syndrome, the efficacy of 2 weight loss compounds liraglutide and sibutramine with different mechanisms of action were examined in 7-month-old gubra DIO-rats. Liraglutide (200 µg/kg, sc) was administered bi-daily, and sibutramine (5 mg/kg, po) was administered once daily for 23 d. RESULTS: Both the compounds effectively reduced the food intake, body weight and total fat mass as measured by nuclear magnetic resonance. Whereas the 5-HT reuptake inhibitor/5-HT receptor agonist sibutramine reduced the intake of both chow and the gubra-diet, the GLP-1 analogue liraglutide predominantly reduced the intake of the highly palatable diet, indicating a shift in food preference. Sibutramine lowered the insulin sensitivity index, primarily via reductions in glucose-stimulated insulin secretion. CONCLUSION: This animal model responds well to 2 weight loss compounds with different mechanisms of action. Moreover, the gubra DIO-rat can be particularly useful for the testing of compounds with potential effects on diet preference.
Assuntos
Fármacos Antiobesidade/uso terapêutico , Peso Corporal/efeitos dos fármacos , Ciclobutanos/uso terapêutico , Ingestão de Alimentos/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Obesidade/tratamento farmacológico , Animais , Fármacos Antiobesidade/farmacologia , Ciclobutanos/farmacologia , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Peptídeo 1 Semelhante ao Glucagon/uso terapêutico , Teste de Tolerância a Glucose , Resistência à Insulina , Liraglutida , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Prolactin-releasing peptide (PrRP) is an endogenous neuropeptide involved in appetite regulation and energy homeostasis. PrRP binds with high affinity to G-protein coupled receptor 10 (GPR10) and with lesser activity towards the neuropeptide FF receptor type 2 (NPFF2R). The present study aimed to develop long-acting PrRP31 analogues with potent anti-obesity efficacy. A comprehensive series of C18 lipidated PrRP31 analogues was characterized in vitro and analogues with various GPR10 and NPFF2R activity profiles were profiled for bioavailability and metabolic effects following subcutaneous administration in diet-induced obese (DIO) mice. PrRP31 analogues acylated with a C18 lipid chain carrying a terminal acid (C18 diacid) were potent GPR10-selective agonists and weight-neutral in DIO mice. In contrast, acylation with aliphatic C18 lipid chain (C18) resulted in dual GPR10-NPFF2R co-agonists that suppressed food intake and promoted a robust weight loss in DIO mice, which was sustained for at least one week after last dosing. Rapid in vivo degradation of C18 PrRP31 analogues gave rise to circulating lipidated PrRP metabolites maintaining dual GPR10-NPFF2R agonist profile and long-acting anti-obesity efficacy in DIO mice. Combined GPR10 and NPFF2R activation may therefore be a critical mechanism for obtaining robust anti-obesity efficacy of PrRP31 analogues.
Assuntos
Fármacos Antiobesidade/administração & dosagem , Obesidade/tratamento farmacológico , Hormônio Liberador de Prolactina/análogos & derivados , Hormônio Liberador de Prolactina/administração & dosagem , Receptores Acoplados a Proteínas G/agonistas , Receptores de Neuropeptídeos/agonistas , Redução de Peso/efeitos dos fármacos , Acilação , Animais , Regulação do Apetite/efeitos dos fármacos , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Ingestão de Alimentos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/etiologia , Obesidade/metabolismo , Hormônio Liberador de Prolactina/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neuropeptídeos/metabolismo , Resultado do TratamentoRESUMO
Non-alcoholic steatohepatitis (NASH) has emerged as a major challenge for public health because of high global prevalence and lack of evidence-based therapies. Most animal models of NASH lack sufficient validation regarding disease progression and pharmacological treatment. The Gubra-Amylin NASH (GAN) diet-induced obese (DIO) mouse demonstrate clinical translatability with respect to disease etiology and hallmarks of NASH. This study aimed to evaluate disease progression and responsiveness to clinically effective interventions in GAN DIO-NASH mice. Disease phenotyping was performed in male C57BL/6J mice fed the GAN diet high in fat, fructose, and cholesterol for 28-88 weeks. GAN DIO-NASH mice with biopsy-confirmed NASH and fibrosis received low-caloric dietary intervention, semaglutide (30 nmol/kg/day, s.c.) or lanifibranor (30 mg/kg/day, p.o.) for 8 and 12 weeks, respectively. Within-subject change in nonalcoholic fatty liver disease (NAFLD) Activity Score (NAS) and fibrosis stage was evaluated using automated deep learning-based image analysis. GAN DIO-NASH mice showed clear and reproducible progression in NASH, fibrosis stage, and tumor burden with high incidence of hepatocellular carcinoma. Consistent with clinical trial outcomes, semaglutide and lanifibranor improved NAS, whereas only lanifibranor induced regression in the fibrosis stage. Dietary intervention also demonstrated substantial benefits on metabolic outcomes and liver histology. Differential therapeutic efficacy of semaglutide, lanifibranor, and dietary intervention was supported by quantitative histology, RNA sequencing, and blood/liver biochemistry. In conclusion, the GAN DIO-NASH mouse model recapitulates various histological stages of NASH and faithfully reproduces histological efficacy profiles of compounds in advanced clinical development for NASH. Collectively, these features highlight the utility of GAN DIO-NASH mice in preclinical drug development.