Preclinical Models Of Type Research Articles

482-P: The GLP-1 Receptor Agonist Liraglutide Improves Glomerular Filtration Rate, Renal Inflammation, and Fibrosis in the Type 2 Diabetic SDT Fatty Rat

The Glucagon-Like Peptide 1 (GLP-1) receptor agonist liraglutide (LIRA) may reduce the rate of development and progression of diabetic kidney disease in type 2 diabetic patients. To evaluate its impact on kidney function, we evaluated the effects of LIRA on glomerular filtration rate (GFR), in both the hyperfiltration and GFR decline phases in the Spontaneously Diabetic Torii (SDT) fatty rat, a preclinical model of type 2 diabetes. SDT fatty male rats were treated subcutaneously, once daily with vehicle or LIRA 0.4mg/kg for 10 weeks. To measure the effects of LIRA on glomerular hyperfiltration, rats were injected with FITC-sinistrin at 4 weeks of treatment to measure GFR. After 5 weeks of treatment, rats underwent unilateral nephrectomy and were put on a 0.3% salt diet to induce a GFR decline for the last weeks of treatment, then GFR was measured at 10 weeks of treatment. Urine biochemistry and kidney histology analysis were also performed. Compared with vehicle, LIRA resulted in significant body weight loss and lower body weight. LIRA also reduced blood glucose levels significantly by up to ∼20%, as expected. During the hyperfiltration phase, LIRA attenuated hyperfiltration, with a 19% lower GFR vs. vehicle (vehicle: 21 ± 1.1 mL/min/kg; LIRA: 17.1 ± 1.0 mL/min/kg, p<0.05 vs. vehicle). In vehicle treated animals, unilateral nephrectomy and 0.3% salt diet induced a major reduction (-56%) in GFR, as expected. After 10 weeks of treatment, LIRA markedly attenuated this GFR decline (vehicle: 9.3 ± 0.5 mL/min/kg; LIRA: 13.3 ± 0.9 mL/min/kg, p<0.001 vs. vehicle). In line with the benefits observed on GFR, LIRA lowered urine albumin to creatinine ratio both in the hyperfiltration and GFR decline phases. Moreover, histology analysis showed that LIRA significantly reduced kidney inflammation and fibrosis scores, as compared with vehicle. In conclusion, LIRA shows significant benefits on GFR, renal inflammation and fibrosis in the SDT fatty rat model. Disclosure F. Briand: Employee; Self; Physiogenex. Stock/Shareholder; Self; Physiogenex. M. Shinohara: None. E. Brousseau: Employee; Self; Physiogenex. Y. Kageyama: None. T. Sulpice: Employee; Self; Physiogenex. Stock/Shareholder; Self; Physiogenex.

Systematic review and meta-analysis protocol for efficacy and safety of Momordica charantia L. on animal models of type 2 diabetes mellitus

BackgroundStudies on several preclinical models of type 2 diabetes mellitus have been conducted to establish the hypoglycemic activity of Momordica charantia L. Concerned with appropriateness of these models, we designed a systematic review to establish the efficacy and safety of M. charantia L. in preclinical models of type 2 diabetes mellitus.MethodsReview authors will search without language restriction in MEDLINE/PubMed, Web of Science, Embase, Scopus, and CINAHL databases through April 2019. Search filters will be applied to enhance search efficiency. The authors will search for gray literature in Google and Google Scholar, OpenGrey, and ProQuest Dissertations & Theses. Two authors will evaluate full texts, extract data, and asses risk of bias independently. The review will include randomized or non-randomized studies that assessed the efficacy or safety of M. charantia L. with vehicle control group. The primary endpoint will be fasting blood glucose level. We will use Egger’s test to assess publication biases. Chi-square test and I2 will be used to assess heterogeneity in effect size of the primary outcome. Using RevMan software version 5.3, the authors will perform a meta-analysis of quantitative data.DiscussionThe strength of evidence will be rated as high, moderate, low, or very low using GRADE framework for animal studies. This systematic review will potentially improve research practice by identifying risks of bias and design features that compromise translatability and contribute to evidence-based clinical trial design.Systematic review registrationPROSPERO CRD42019119181

A model for determining an effective in vivo dose of transplanted islets based on in vitro insulin secretion.

Allogeneic islet transplantation for the treatment of type 1 diabetes often requires multiple implant procedures, from as many as several human pancreas donors, to achieve lasting clinical benefit. Given the limited availability of human pancreases for islet isolation, porcine islets have long been considered a potential option for clinical use. Agarose-encapsulated porcine islets (macrobeads) permit long-term culture and thus a thorough evaluation of microbiological safety and daily insulin secretory capacity, prior to implantation. The goal of this study was the development of a method for determining an effective dose of encapsulated islets based on their measured in vitro insulin secretion in a preclinical model of type 1 diabetes. Spontaneously diabetic BioBreeding diabetes-prone rats were implanted with osmotic insulin pumps in combination with continuous glucose monitoring to establish the daily insulin dose required to achieve continuous euglycaemia in individual animals. Rats were then implanted with a 1×, 2× or 3× dose (defined as the ratio of macrobead in vitro insulin secretion per 24hours to the recipient animal's total daily insulin requirement) of porcine islet macrobeads, in the absence of immunosuppression. In vivo macrobead function was assessed by recipient non-fasted morning blood glucose values, continuous glucose monitoring and the presence of peritoneal porcine C-peptide. At the end of the study, the implanted macrobeads were removed and returned to in vitro culture for the evaluation of insulin secretion. Diabetic rats receiving a 2× macrobead implant exhibited significantly improved blood glucose regulation compared to that of rats receiving a 1× dose during a 30-day pilot study. In a 3-month follow-up study, 2× and 3× macrobead doses initially controlled blood glucose levels equally well, although several animals receiving a 3× dose maintained euglycaemia throughout the study, compared to none of the 2× animals. The presence of porcine C-peptide in rat peritoneal fluid 3months post-implant and the recurrence of hyperglycaemia following macrobead removal, along with the finding of persistent in vitro insulin secretion from retrieved macrobeads, confirmed long-term graft function. Increasing dosages of islet macrobeads transplanted into diabetic rats, based on multiples of in vitro insulin secretion matched to the recipient's exogenous insulin requirements, correlated with improved blood glucose regulation and increased duration of graft function. These results demonstrate the usefulness of a standardized model for the evaluation of the functional effectiveness of islets intended for transplantation, in this case using intraperitoneally implanted agarose macrobeads, in diabetic rats. The results suggest that some features of this islet-dosing methodology may be applicable, and indeed necessary, to clinical allogeneic and xenogeneic islet transplantation.

Exploration of phenylpropanoic acids as agonists of the free fatty acid receptor 4 (FFA4): Identification of an orally efficacious FFA4 agonist

The long chain free fatty acid receptor 4 (FFA4/GPR120) has recently been recognized as lipid sensor playing important roles in nutrient sensing and inflammation and thus holds potential as a therapeutic target for type 2 diabetes and metabolic syndrome. To explore the effects of stimulating this receptor in animal models of metabolic disease, we initiated work to identify agonists with appropriate pharmacokinetic properties to support progression into in vivo studies. Extensive SAR studies of a series of phenylpropanoic acids led to the identification of compound 29, a FFA4 agonist which lowers plasma glucose in two preclinical models of type 2 diabetes.

Safety, tolerability, pharmacokinetics, and pharmacodynamics of novel glucokinase activator HMS5552: results from a first-in-human single ascending dose study.

BackgroundHMS5552, a novel fourth-generation glucokinase (GK) activator, has demonstrated promising effects on glycemic control in preclinical models of type 2 diabetes. This single ascending dose study was conducted to investigate the safety, tolerability, pharmacokinetics (PK), and pharmacodynamics (PD) of HMS5552 during its first-in-human exposure.MethodsSixty healthy subjects were enrolled. In each of six dose-cohorts (5, 10, 15, 25, 35, and 50 mg), ten subjects were randomized with eight subjects receiving the same cohort-dose of HMS5552 and two receiving placebo. Plasma HMS5552 exposure, glucose, and insulin were measured repeatedly during fasting and after a standardized meal. Assessment included safety, PK, and PD endpoints.ResultsHMS5552 showed dose-proportional increases in area under the curve 0 to the last quantifiable concentration (AUC0–t) and maximum plasma concentration (Cmax). Slopes estimated by linear regression for AUC0–t and Cmax were ~1.0 (0.932 and 0.933, respectively). Geometric mean elimination half-life ranged from 4.48 to 7.51 hours and apparent clearance ranged from 11.5 to 13.1 L/h across all doses. No significant sex effect was observed in PK parameters. HMS5552 also demonstrated dose-related PD responses in terms of maximum glucose change from baseline (%) and mean glucose area under effect curve 0–4 hours change from baseline (%) (P<0.001). Fifteen adverse events were reported by nine subjects (ten with HMS5552 and five with the placebo). All adverse events were mild in intensity and resolved without any treatment.ConclusionThis first-in-human single ascending dose study provided predicted PK of HMS5552 with dose-proportional increases in AUC0–t and Cmax, as well as dose-related glucose-lowering effects over the range of 5–50 mg in healthy subjects. HMS5552 at doses up to 50 mg in healthy subjects was safe and well-tolerated.

Renal Structure‐Function Relationship in the Obese ZSF1 Rat Model of Diabetic Nephropathy

Although the obese ZSF1 rat is a preclinical model of type 2 diabetic nephropathy (DN) with marked renal dysfunction and structural derangement, the course of disease progression has not been evaluated in an age‐dependent manner. This study characterizes temporal changes in renal function relative to changes in proteinuria and glomerular and tubulointerstitial structure to further understand renal disease progression in this model.Male obese ZSF1 rats were aged to 20, 25, 30, 35, or 40 weeks of age. At those times, proteinuria (UPCR) and GFR (anesthetized, FITC‐inulin, kidney weight (KW) normalized) were measured. The left kidney was processed for histopathological analyses of glomerular lesions (% glomerulosclerosis) and tubulointerstitial damage (foci number, interstitial collagen, a‐SMA, KIM‐1, ED‐1+).GFR was steady from 20 to 30 weeks of age (avg 0.71±0.06 mL/min/g KW). At 35 and 40 weeks, GFR declined markedly to 0.29±0.11 and 0.23±0.05 mL/min/g KW, respectively. Glomerulosclerosis increased steadily from 12±1% at 20 weeks to 71±2% by 40 weeks as did UPCR, glomerular lesion severity score, interstitial lesions, a‐SMA, and ED‐1. Interstitial collagen increased modestly from 20 weeks (1.3±0.2%) to 30 weeks (3.7±0.3%) and markedly increased at 35 and 40 weeks, to 8.3±0.6% and 13.3±0.9%, respectively. KIM‐1 increased to 30 weeks (7.5±0.6%) then declined at 40 weeks.In aging obese ZSF1 rats, increased UPCR and glomerulosclerosis occurred prior to the decline in GFR and further increased as GFR fell. The decreased GFR also corresponded with a marked increase in tubulointerstitial fibrosis and injury. These results, consistent with observations in human DN, also support the obese ZSF1 rat as a relevant preclinical model of DN.

Blood glucose‐lowering effect of Tectona grandis flowers in type 2 diabetic rats: A study on identification of active constituents and mechanisms for antidiabetic action 柚木花对2型糖尿病大鼠的降糖作用：一项鉴定其活性成份与降糖作用机制的研究

AbstractBackgroundAyurveda documents antidiabetic activity of Tectona grandis flowers. However, until now, the effects of T. grandis flowers in preclinical models of type 2 diabetes (T2D) have been unknown. Hence, the aim of the present study was to evaluate the blood glucose‐lowering effect of methanol extract of T. grandis flowers (METGF) in T2D rats and to identify possible active constituents as well as the mechanisms for the antidiabetic action.MethodsRats were rendered diabetic by administration of streptozotocin–nicotinamide (65 mg/kg–110 mg/kg, i.p.). After the induction of diabetes, Groups 3 and 4 received METGF (100 and 200 mg/kg respectively) while Group 5 received glibenclamide (5 mg/kg) orally for 4 weeks. Blood glucose and body weight were determined at the end of Weeks 2 and 4. Hemoglobin, HbA1c, total protein, urea, creatinine, insulin, and lipid levels were determined in Week 4. Active constituents in METGF were identified using HPLC analysis. Antidiabetic mechanisms of METGF were evaluated by investigating the insulin sensitizing action and inhibition of α‐amylase and α‐glucosidase activity.ResultsBoth METGF and glibenclamide significantly improved body weight, reduced blood glucose, and normalized altered biochemical parameters (P &lt; 0.001) in T2D rats. The presence of polyphenolic active constituents, such as gallic acid, quercetin, rutin, ellagic acid, ferulic acid, and kaempferol, in METGF was confirmed by HPLC. In vitro, METGF, gallic acid, quercetin, and rutin exhibited significant (P &lt; 0.001) insulin sensitizing action and inhibition of α‐amylase and α‐glucosidase activity.ConclusionsPolyphenolic active constituents present in METGF are possibly responsible for the blood glucose‐lowering effect in T2D rats via an insulin‐sensitizing action, as well as inhibition of α‐amylase and α‐glucosidase activity.

Pharmacological application of carbon monoxide ameliorates islet-directed autoimmunity in mice via anti-inflammatory and anti-apoptotic effects

Recent studies have identified carbon monoxide (CO) as a potential therapeutic molecule for the treatment of autoimmune diseases owing to its anti-inflammatory and anti-apoptotic properties. We explored the efficacy and the mechanisms of action of the CO-releasing molecule (CORM)-A1 in preclinical models of type 1 diabetes. The impact of CORM-A1 on diabetes development was evaluated in models of spontaneous diabetes in NOD mice and in diabetes induced in C57BL/6 mice by multiple low-dose streptozotocin (MLDS). Ex vivo analysis was performed to determine the impact of CORM-A1 both on T helper (Th) cell and macrophage differentiation and on their production of soluble mediators in peripheral tissues and in infiltrates of pancreatic islets. The potential effect of CORM-A1 on cytokine-induced apoptosis in pancreatic islets or beta cells was evaluated in vitro. CORM-A1 conferred protection from diabetes in MLDS-induced mice and reduced diabetes incidence in NOD mice as confirmed by preserved insulin secretion and improved histological signs of the disease. In MLDS-challenged mice, CORM-A1 attenuated Th1, Th17, and M1 macrophage response and facilitated Th2 cell differentiation. In addition, CORM-A1 treatment in NOD mice upregulated the regulatory arm of the immune response (M2 macrophages and FoxP3(+) regulatory T cells). Importantly, CORM-A1 interfered with in vitro cytokine-induced beta cell apoptosis through the reduction of cytochrome c and caspase 3 levels. The ability of CORM-A1 to protect mice from developing type 1 diabetes provides a valuable proof of concept for the potential exploitation of controlled CO delivery in clinical settings for the treatment of autoimmune diabetes.

Paradoxical insights into whole body metabolic adaptations following SGLT2 inhibition

It is well known that glycemic control over time reduces microvascular and macrovascular complications in human subjects with type 2 diabetes. In addition, preclinical models of type 2 diabetes have demonstrated that long-term hyperglycemia exacerbates insulin resistance and reduces β cell function; therefore, therapies that reduce blood glucose levels are of great interest in not only controlling complications, but for restoring known defects in the pathogenesis of type 2 diabetes. Pharmacological inhibition of the sodium-glucose cotransporter 2 (SGLT2) reduces plasma glucose by limiting glucose absorption in the kidney and increasing glucose excretion in the urine. In this issue of the JCI, Merovci and colleagues and Ferrannini and colleagues independently report a paradoxical increase in endogenous glucose production in patients with type 2 diabetes following SGLT2 inhibition, despite an overall decrease in fasting plasma glucose. Together, these studies provide a unique insight into the effects of SGLT2 inhibition on whole body metabolism.

Epigenetic Mechanisms of Metabolic Memory in Diabetes

Diabetes affects 28.5 million people in the US.1 In addition, several million people with diabetes are estimated to be undiagnosed, thus increasing the risk for developing long-term complications, including macrovascular diseases and end-stage renal disease in a fairly significant fraction of the population. Type 1 diabetes, an autoimmune disease that mainly afflicts children, is characterized by the loss of insulin-producing beta cells. Until now, our understanding of this insidious disease relied on classic genetic approaches including genome wide association studies and single nucleotide polymorphisms. Although 40 immune response related genes have been implicated in type 1 diabetes, we are far from understanding how these genes render susceptibility to this complex disorder.2 Type 2 diabetes is common among people >45 years of age and characterized by the lack of serum autoantibodies, progressive insulin resistance, and beta cell insufficiency. Despite the differences in the etiology, both type 1 and type 2 diabetes are associated with microvascular complications such as diabetic nephropathy, neuropathy, and retinopathy, as well as macrovascular cardiovascular diseases including atherosclerosis, hypertension, and stroke. Tight glycemic control for 3 to 5 years may prevent cardiovascular disease but not all diabetes-related endpoints, referred to as the hyperglycemic memory3 or the legacy effect.4 Although it is unclear how the external cues such as hyperglycemia is perceived as a signal for gene transcription, several intracellular events including generation of oxidative stress and advanced glycation end products, and engagement of the mitogen activated protein kinase pathway have been implicated in metabolic memory (Figure). Recent data indicated a role for epigenetic mechanisms in a preclinical model of type 1 diabetes.5 Although gene-environment interactions have been implicated in the manifestation of type 2 diabetes, compelling evidence for the involvement of epigenetic mechanisms in the initiation and transmission of the metabolic syndrome to …

Dry olive leaf extract (DOLE) down-regulates the progression of experimental immune-mediated diabetes by modulation of cytokine profile in the draining lymph nodes

We have recently demonstrated the beneficial effects of dry olive leaf extract (DOLE) in two preclinical models of type 1 diabetes. Here we analyze the potential mechanisms underlying diabetes amelioration at the level of lymph node drainage. Treatment of C57BL/6 mice with DOLE during induction of diabetes with multiple low doses of streptozotocin (MLD-SZ) modulated cytokine expression and production in pancreatic lymph node cells, thereby changing the balance between potentially pathogenic and down-regulating cytokines. These results support the immunoregulatory potential of DOLE which takes place at the level of lymph node drainage and preserves the target tissue from autoimmune attack.

Inhibition of insulin secretion as a new drug target in the treatment of metabolic disorders.

The pattern of insulin release is crucial for regulation of glucose and lipid haemostasis. Deficient insulin release causes hyperglycemia and diabetes, whereas excessive insulin release can give rise to serious metabolic disorders, such as nesidioblastosis (Persistent Hyperinsulinemic Hypoglycemia of Infancy, PHHI) and might also be closely associated with development of type 2 diabetes and obesity. Type 2 diabetes is characterized by fasting hyperinsulinemia, insulin resistance and impaired insulin release, i.e. reduced first phase insulin release and decreased insulin pulse mass. The beta cell function of patients with type 2 diabetes slowly declines and will ultimately result in beta cell failure and increasing degrees of hyperglycemia. Type 2 diabetes, in combination with obesity and cardiovascular disorders, forms the metabolic syndrome. It has been possible to improve beta cell function and viability in preclinical models of type 1 and type 2 diabetes by reducing insulin secretion to induce beta cell rest. Clinical studies have furthermore indicated that inhibitors of insulin release will be of benefit in treatment or prevention of diabetes and obesity. Pancreatic beta cells secrete insulin in response to increased metabolism and by stimulation of different receptors. The energy status of the beta cell controls insulin release via regulation of open probability of the ATP sensitive potassium (K(ATP)) channels to affect membrane potential and the intracellular calcium concentration [Ca(2+)](i). Other membrane bound receptors and ion channels and intracellular targets that modulate [Ca(2+)](i)will affect insulin release. Thus, insulin release is regulated by e.g. somatostatin receptors, GLP-1 receptors, muscarinic receptors, cholecystokinin receptors and adrenergic receptors. Although the relationship between hyperinsulinemia and certain metabolic diseases has been known for decades, only a few inhibitors of insulin release have been characterized in vitro and in vivo. These include the K(ATP) channel openers diazoxide and NN414 and the somatostatin receptor agonist octreotide.

Successful reversal of streptozotocin-induced diabetes with stable allogeneic islet function in a preclinical model of type 1 diabetes.

The recent focus on islet transplantation as primary therapy for type 1 diabetes has heightened interest in the reversal of type 1 diabetes in preclinical models using minimal immunosuppression. Here, we demonstrated in a preclinical rhesus model a consistent reversal of all measured glycemic patterns of streptozotocin-induced type 1 diabetes. The model used single-donor islet transplantation with induction of operational tolerance. The term "operational tolerance" is used to indicate durable survival of single-donor major histocompatibility complex (MHC)-mismatched islet allografts without maintenance immunosuppressive therapy and without rejection or loss of functional islet mass or insulin secretory reserve. In this operational tolerance model, all immunosuppression was discontinued after day 14 posttransplant, and recipients recovered with excellent health. The operational tolerance induction protocol combined peritransplant anti-CD3 immunotoxin to deplete T-cells and 15-deoxyspergualin to arrest proinflammatory cytokine production and maturation of dendritic cells. T-cell deficiency was specific but temporary, in that T-cell-dependent responses in long-term survivors recovered to normal, and there was no evidence of increased susceptibility to infection. Anti-donor mixed lymphocyte reaction responses were positive in the long-term survivors, but all showed clear evidence of systemic T-helper 2 deviation, suggesting that an immunoregulatory rather than a deletional process underlies this operational tolerance model. This study provides the first evidence that operational tolerance can protect MHC nonhuman primate islets from rejection as well as loss of functional islet mass. Such an approach has potential to optimize individual recipient recovery from diabetes as well as permitting more widespread islet transplantation with the limited supply of donor islets.