Different post-pancreatectomy glucagon responses to a meal test between surgical approaches.

Residual pancreatic endocrine function is important for maintaining metabolic status after pancreatectomy and is closely related to patient nutritional status and prognosis. In contrast to insulin secretion, the significance of glucagon secretion following pancreatectomy remains unclear. In this study, we assessed the changes in pancreatic glucagon secretion during pancreatectomy to determine their pathophysiological significance. We evaluated glucagon and insulin secretion using a liquid meal tolerance test before and after pancreatectomy in patients scheduled to undergo pancreaticoduodenectomy (PD) or distal pancreatectomy (DP). After pancreatectomy, fasting plasma glucagon levels were significantly decreased in both the PD (n = 10) and DP (n = 5) groups (PD: from 18.4 to 10.5 pg/mL, p = 0.037; DP: from 21.0 to 12.1 pg/mL, p = 0.043), whereas postprandial plasma glucagon levels were not changed. In the liquid meal tolerance test after pancreatectomy, 60-min plasma glucagon levels and the area under the curve (AUC) for 0-120 min of PD were significantly higher than those for DP (60-min plasma glucagon: PD 49.0 vs. DP 21.7 pg/mL, p = 0.040; AUC0-120min: PD 4,749 vs. DP 3,564 µg min/mL, p = 0.028). Postoperative plasma glucose, serum insulin, and serum C-peptide levels during the liquid meal tolerance test were not significantly different between the two groups. Although fasting plasma glucagon levels decreased, postprandial glucagon responses were maintained after both PD and DP. The difference in residual meal-stimulated glucagon response between PD and DP suggests that a relative excess of postprandial glucagon is involved in the postoperative nutritional status after PD through its impact on systemic metabolic status.

Newly discovered knowledge pertaining to glucagon and its clinical applications.

Glucagon has been defined as an 'insulin counteracting hormone', which raises blood glucose levels. Recent progress in basic research has shown that glucagon is closely involved in glucose and amino acid metabolism. Additionally, its secretion is intricately, but precisely, regulated by various mechanisms involving molecules in addition to glucose, thus showing its critical role in systemic nutrient metabolism. An innovative dual-antibody-linked immunosorbent assay for glucagon that improves measurement accuracy has been developed, and substantial clinical findings have been obtained using this new system. This discovery expanded the pathophysiological significance of glucagon and accelerated the development of its clinical applications in diabetes.

Characteristic changes in plasma glutamate levels and free amino acid profiles in Japanese patients with type 1 diabetes mellitus.

AIMS/INTRODUCTION: In addition to absolute insulin deficiency, dysregulated glucagon in type 1 diabetes is considered pathophysiologically important. Previously, we confirmed the presence of dysregulated glucagon in Japanese patients with type 1 diabetes, and found a significant correlation between plasma glucagon and blood urea nitrogen levels, suggesting an association between glucagon and amino acid metabolism. In this study, we evaluated plasma amino acid levels in Japanese patients with type 1 diabetes in the context of their functional relationship with glucagon. MATERIALS AND METHODS: We assessed plasma free amino acid levels using liquid chromatography-mass spectrometry in 77 Japanese patients with type 1 diabetes, and statistically analyzed their characteristics and relationships with clinical parameters, including glucagon. RESULTS: Participants with type 1 diabetes showed a large decrease in glutamate levels together with a characteristic change in plasma free amino acid profiles. The network structural prediction analyses showed correlations between each amino acid and glucagon in type 1 diabetes. CONCLUSIONS: Participants with type 1 diabetes showed characteristic changes in plasma glutamate levels and free amino acid profiles compared with controls and type 2 diabetes patients. Glucagon showed a closer correlation with amino acids than with parameters of glucose metabolism, suggesting that type 1 diabetes includes dysregulation in amino acids through dysregulated glucagon from remaining pancreatic α-cells, together with that in glucose by insulin deficiency.

Change in pharmacodynamic variables following once-weekly tirzepatide treatment versus dulaglutide in Japanese patients with type 2 diabetes (SURPASS J-mono substudy).

AIM: To evaluate the pharmacodynamic effects of tirzepatide, a novel dual glucagon-like peptide-1 receptor and glucose-dependent insulinotropic polypeptide receptor agonist, compared with dulaglutide in patients with type 2 diabetes. MATERIALS AND METHODS: SURPASS J-mono was a 52-week, multicentre, randomized, double-blind, parallel, active-controlled, Phase 3 study, conducted in Japan. This substudy of SURPASS J-mono evaluated postprandial metabolic variables and appetite after a meal tolerance test, and body composition measured by bioelectrical impedance analysis. RESULTS: Of 636 participants in SURPASS J-mono, 48 were included in this substudy and assigned to tirzepatide 5 mg (n = 9), tirzepatide 10 mg (n = 11), tirzepatide 15 mg (n = 9), or dulaglutide 0.75 mg (n = 19). Participants had a mean (standard deviation) age of 58.6 (7.5) years, duration of diabetes of 6.0 (6.3) years, and body mass index of 27.5 (3.5) kg/m2 . Mean glycated haemoglobin at baseline was 66 mmol/mol (8.22%). Following a standardized meal test, statistically significant differences in change from baseline in area under the concentration versus time curve from time zero to 6 h after dose for glucose, insulin, glucagon, C-peptide and triglycerides were observed in all tirzepatide treatment arms, except triglycerides at 10 mg, compared with dulaglutide at Week 32. For body composition, tirzepatide 10 mg and 15 mg resulted in a significant reduction in body weight, and all doses of tirzepatide resulted in a significant reduction in body fat mass at Week 52. CONCLUSIONS: Compared with dulaglutide, tirzepatide showed greater potential for normalizing metabolic factors after a standardized meal. Tirzepatide reduced body weight and body fat mass.

Soluble T-cadherin promotes pancreatic ß-cell proliferation by upregulating Notch signaling.

Endogenous humoral factors that link systemic and/or local insulin demand to pancreatic ß-cells have not been identified. Here, we demonstrated that T-cadherin, a unique glycosylphosphatidylinositol-anchored cadherin primarily expressed in vascular endothelial cells and cardiac and skeletal muscle cells, but not in pancreatic ß-cells, was secreted as soluble forms and was important for ß-cell proliferation. Cdh13 (T-cadherin) knockout mice exhibited impaired glucose handling due to attenuated ß-cell proliferation under high-fat diet conditions. The gene expression analyses indicated the impairment in cell cycle and Notch signaling in the islets of T-cadherin knockout mice under high-fat diet conditions. In streptozotocin-induced diabetes, the replacement of soluble T-cadherin improved ß-cell mass and blood glucose levels in T-cadherin knockout mice. Recombinant soluble T-cadherin upregulated Notch signaling in cultured murine islets. We concluded that soluble T-cadherin could work as an endogenous humoral factor whose signaling pathways including Notch signaling regulate ß-cell proliferation under diabetic conditions in mice.

Plasma lipopolysaccharide binding protein level statistically mediates between body mass index and chronic microinflammation in Japanese patients with type 1 diabetes.

Recently, it is widely recognized that microinflammation plays important roles in the pathophysiology of metabolic diseases, especially obesity-related disorders, diabetes and their complications. Lipopolysaccharide-binding protein (LBP) is a liver-derived acute-phase protein responsive to lipopolysaccharides (LPS) produced by gram-negative bacteria, thus reflects the systemic inflammation caused by the infection of those bacteria including gut dysbiosis. In this study, we evaluated the plasma LBP levels and investigated its clinical significance in 67 Japanese patients with type 1 diabetes. Univariable analysis showed that LBP levels were significantly associated with body mass index (BMI; r = 0.43, p < 0.01) and serum high-sensitivity C-reactive protein (hs-CRP; r = 0.64, p < 0.001) levels. However, there was no significant association between plasma LBP levels and diabetic complications. Mediation analysis revealed that LBP had significant mediation effects on the association between hs-CRP and BMI (0.27 [95% confidence interval 0.10-0.48]). These results suggest that the systemic condition where the LBP level increases, such as gut dysbiosis, at least partly, impacts on chronic microinflammation in patients with type 1 diabetes.

Beginning of a new era in glucagon research: Breakthrough by the new glucagon assay.

There is a new concept of diabetes as a "comprehensive nutrition disorder", caused due to both insulin and glucagon dysregulation. Dysregulated glucagon secretion in α-cells exacerbates multiple metabolic disorders: glycemic control and amino acid metabolism, together with insulin deficiency.

Consistency of plasma glucagon levels in patients with type 1 diabetes after a 1-year period.

Recent progress in research on glucagon and α-cells highlights their pathophysiological roles in diabetes. We previously showed that plasma glucagon levels measured by newly developed enzyme-linked immunosorbent assay were dysregulated in patients with type 1 diabetes with respect to plasma glucose levels, suggesting dysregulated secretion. In the current study, the annual change in plasma glucagon levels was assessed in these same patients. We found that the plasma glucagon levels in the 66 Japanese patients involved in the study were significantly correlated between both years. In addition, they were significantly associated with serum blood urea nitrogen levels in a multivariate linear regression analysis, as reported in our previous study. The statistical correlation in glucagon levels between annual checkups and the sustained significant correlation between glucagon and blood urea nitrogen suggest a constant dysregulation of glucagon in association with altered amino acid metabolism in patients with type 1 diabetes.

Positive correlation between fasting plasma glucagon and serum C-peptide in Japanese patients with diabetes.

AIMS: Glucagon plays pivotal roles in systemic glucose homeostasis mainly by promoting hepatic glucose output. Using a sandwich enzyme-linked immunosorbent assay (ELISA), we evaluated fasting plasma glucagon levels in hospitalized patients with type 1 or type 2 diabetes, and assessed the relationships between glucagon levels and various clinical parameters. METHODS: We enrolled adult Japanese diabetes patients admitted to Osaka University Medical Hospital for glycemic control between July 2017 and May 2018 in this study. After patients had fasted for 12 h, blood samples were obtained and plasma glucagon levels were measured using a sandwich ELISA. RESULTS: Total 107 patients participated in the study. The mean fasting plasma glucagon level of patients with acute onset type 1 diabetes was significantly lower than that of patients with type 2 diabetes (p < 0.05). Plasma glucagon levels were not significantly correlated with plasma glucose levels in patients with type 1 diabetes or in patients with type 2 diabetes. Multiple regression analysis indicated that fasting glucagon levels were independently and significantly correlated with fasting serum C-peptide levels in patients with type 2 diabetes. CONCLUSIONS: Our results suggest that insulin and glucagon secretion are balanced in the fasting state in patients with type 2 diabetes.

Dysregulated plasma glucagon levels in Japanese young adult type 1 diabetes patients.

Currently, the clinical dynamics of glucagon need to be revised based on previous data obtained from conventional glucagon radioimmunoassays. In the present study, we evaluated plasma glucagon levels in type 1 diabetes patients using a newly-developed sandwich enzyme-linked immunosorbent assay, and its association with clinical parameters and markers of diabetes complications were statistically assessed. The plasma glucagon level in 77 Japanese type 1 diabetes patients was 28.1 ± 17.7 pg/mL, and comparable with that reported previously for type 2 diabetes patients. However, the values were widely spread and did not correlate with plasma glucose values. Additionally, the average glucagon levels in patients in a hypoglycemic state (glucose level <80 mg/dL) did not increase (21.7 ± 12.2 pg/mL). The average glucagon level of patients experiencing hypoglycemia unawareness was significantly lower. Plasma glucagon levels evaluated using the new enzyme-linked immunosorbent assay were dysregulated in type 1 diabetes patients in respect to plasma glucose levels, suggesting dysregulation of secretion.

Alpha the versatile: Guardians of the islets.

Recent progress in α-cell research has newly revealed its versatility including secretion of multiple hormones, trans-differentiation, and abundant proliferation. A recent report from the Kushner laboratory further provided direct evidence of active proliferation of α-cells in both T1D and non-diabetic control donors. α-cells are suggested to maintain both quality and quantity of islets by serving as 'guardians' of the islets.

Skin autofluorescence is associated with vascular complications in patients with type 2 diabetes.

AIMS: Tissue accumulatedadvanced glycation end products (AGEs) can be evaluated non-invasively by an autofluorescence reader as skin autofluorescence (skin AF)·The present study investigated whether skin AF is associated with diabetic micro- and macroangiopathies in Japanese patients with type 2 diabetes mellitus (T2DM). METHODS: Skin AF was measured in 193 enrolled Japanese patients with T2DM and 24 enrolled healthy non-diabetic subjects by using the AGE reader®. Diabetic micro- and macroangiopathies were evaluated in the T2DM patients. RESULTS: Skin AF was significantly increased in patients with T2DM than in age- and sex-matched non-diabetic controls (2.35 ±â€¯0.51 [mean ±â€¯SD] and 1.91 ±â€¯0.29, respectively, p = 0.001). In subjects with T2DM, skin AF was associated with age, pack-years of smoking, and eGFR (estimated glomerular filtration rate) independently. Skin AF was significantly increased in patients with diabetic retinopathy, neuropathy, nephropathy, and macroangiopathy than in those without them, and significantly associated with the number of diabetic complications. Moreover, skin AF was an independent predictor for diabetic retinopathy, neuropathy, and nephropathy but not macroangiopathy, after adjusting for major traditional risk factors. CONCLUSIONS: Skin AF is an independent predictor for diabetic retinopathy, neuropathy and nephropathy in Japanese patients with T2DM.

Association between Subclinical Atherosclerosis Markers and the Level of Accumulated Advanced Glycation End-Products in the Skin of Patients with Diabetes.

AIM: The level of accumulated advanced glycation end-products (AGEs) in the skin has been shown to predict the risk of complications in patients with diabetes mellitus (DM). Recently, the level of accumulated fluorescent AGEs in the skin has become measurable as skin autofluorescence (skin AF) using a non-invasive apparatus, autofluorescence reader. The purpose of this study was to evaluate the association between skin AF and the subclinical atherosclerosis markers, especially endothelial dysfunction, in patients with DM. METHODS: We enrolled 140 Japanese subjects with DM who attended Osaka University Hospital, and measured the skin level of AGEs by skin AF and three subclinical atherosclerosis markers: endothelial function by flow-mediated vasodilation, FMD; carotid intima-media thickness, IMT; and brachial-ankle pulse wave velocity, baPWV. RESULTS: FMD was significantly associated with skin AF (r=－0.259, p=0.002). Furthermore, a stepwise multivariate regression analysis revealed that skin AF was an independent determinant of FMD (ß=－0.180, p=0.038). Although there were significant associations between skin AF and maximum carotid intima-media thickness (max-IMT)(r=0.298, p＜0.001) as well as baPWV (r=0.284, p= 0.001) in univariate analysis, skin AF was not an independent determinant of either carotid max-IMT or baPWV after adjustment for conventional cardiovascular risk factors. Receiver-operating characteristic curve analysis revealed that skin AF can identify the subjects whose FMD, max-IMT, and baPWV were completely within the normal range (C-statistics, 0.73; 95% confidence interval, 0.61-0.84; p＜0.001). CONCLUSIONS: Skin AF was independently associated with FMD as an indicator of endothelial dysfunction, and can be utilized as a screening marker of atherosclerosis in Japanese patients with DM.

GLP-1 signalling compensates for impaired insulin signalling in regulating beta cell proliferation in ßIRKO mice.

AIMS/HYPOTHESIS: We aimed to investigate potential interactions between insulin and glucagon-like peptide (GLP)-1 signalling pathways in the regulation of beta cell-cycle dynamics in vivo, in the context of the therapeutic potential of GLP-1 to modulate impaired beta cell function. METHODS: Beta cell-specific insulin receptor knockout (ßIRKO) mice, which exhibit beta cell dysfunction and an age-dependent decrease in beta cell mass, were treated with the dipeptidyl peptidase-4 inhibitor vildagliptin. Following this, glucose homeostasis and beta cell proliferation were evaluated and underlying molecular mechanisms were investigated. RESULTS: The sustained elevation in circulating GLP-1 levels, caused by treatment of the knockout mice with vildagliptin for 6 weeks, significantly improved glucose tolerance secondary to enhanced insulin secretion and proliferation of beta cells. Treating ßIRKO beta cell lines with the GLP-1 analogue, exendin-4, promoted Akt phosphorylation and protein expression of cyclins A, D1 and E two- to threefold, in addition to cyclin D2. Pancreases from the vildagliptin-treated ßIRKO mice exhibited increased cyclin D1 expression, while cyclin D2 expression was impaired. CONCLUSIONS/INTERPRETATION: Activation of GLP-1 signalling compensates for impaired growth factor (insulin) signalling and enhances expression of cyclins to promote beta cell proliferation. Together, these data indicate the potential of GLP-1-related therapies to enhance beta cell proliferation and promote beneficial outcomes in models with dysfunctional beta cells.

Glucotoxicity induces abnormal glucagon secretion through impaired insulin signaling in InR1G cells.

The significance of glucagon in the pathophysiology of diabetes mellitus is widely recognized, but the mechanisms underlying dysregulated glucagon secretion are still unclear. Here, we explored the molecular mechanisms of glucagon dysregulation, using an in vitro model. Hamster-derived glucagon-secreting InR1G cells were exposed to high glucose (25 mM) levels for 12 h before analyzing glucagon secretion and the activity of components involved in insulin signaling. High-glucose treatment induced increased glucagon secretion in InR1G cells, which represents a hallmark of diabetes mellitus. This treatment reduced the phosphorylation of Akt, indicating the deterioration of insulin signaling. Simultaneously, oxidative stress and JNK activity were shown to be increased. The inhibition of JNK signaling resulted in the amelioration of high-glucose level-induced glucagon secretion. Abnormally elevated glucagon secretion in diabetes can be reproduced by high-glucose treatment of InR1G cells, and the involvement of high glucose-oxidative stress-JNK-insulin signaling pathway axis has been demonstrated. These data elucidate, at least partly, the previously unclear mechanism of abnormal glucagon secretion, providing insights into a potential novel approach to diabetes treatment, targeting glucagon.

Mafa Enables Pdx1 to Effectively Convert Pancreatic Islet Progenitors and Committed Islet α-Cells Into ß-Cells In Vivo.

Among the therapeutic avenues being explored for replacement of the functional islet ß-cell mass lost in type 1 diabetes (T1D), reprogramming of adult cell types into new ß-cells has been actively pursued. Notably, mouse islet α-cells will transdifferentiate into ß-cells under conditions of near ß-cell loss, a condition similar to T1D. Moreover, human islet α-cells also appear to poised for reprogramming into insulin-positive cells. Here we have generated transgenic mice conditionally expressing the islet ß-cell-enriched Mafa and/or Pdx1 transcription factors to examine their potential to transdifferentiate embryonic pan-islet cell Ngn3-positive progenitors and the later glucagon-positive α-cell population into ß-cells. Mafa was found to both potentiate the ability of Pdx1 to induce ß-cell formation from Ngn3-positive endocrine precursors and enable Pdx1 to produce ß-cells from α-cells. These results provide valuable insight into the fundamental mechanisms influencing islet cell plasticity in vivo.

Skin Autofluorescence is Associated with Early-stage Atherosclerosis in Patients with Type 1 Diabetes.

AIM: Accumulation level of fluorescent advanced glycation end products (AGEs) in the skin can be measured non-invasively as skin autofluorescence (skin AF) by autofluorescence reader. The aim of this study was to assess possible associations between skin AF and diabetic complications, especially early-stage atherosclerosis, in Japanese type 1 diabetic patients. METHODS: Skin AF was measured by AGE reader® in 105 Japanese type 1 diabetic patients (34 men and 71 women, aged 37.4±12.4 years (±SD)) and 23 age-matched healthy non-diabetic subjects. Ultrasonic carotid intima-media thickness (IMT), ankle-brachial index (ABI), and brachial ankle pulse wave velocity (baPWV) were evaluated as indices of early-stage diabetic macroangiopathy. Urinary albumin-to-creatinine ratio (UACR), the coefficient of variation of R-R intervals (CVR-R), and presence of retinopathy were also evaluated. RESULTS: Skin AF values were significantly higher in type 1 diabetic patients than in healthy controls (2.07±0.50 (mean±SD) and 1.90±0.26, respectively, p=0.024). Skin AF was associated with carotid IMT (r=0.446, p＜0.001) and baPWV (r=0.450, p＜0.001), but not with ABI (r=－0.019, p=0.8488). Notably, skin AF was an independent risk factor for IMT thickening. Similarly, skin AF was associated with log (UACR) (r=0.194, p=0.049) and was an independent risk factor for UACR. Furthermore, skin AF values were significantly higher in patients with diabetic retinopathy than in those without (2.21±0.08 and 1.97±0.06, respectively, p=0.020). CONCLUSIONS: Skin AF was significantly associated with the presence and/or severity of diabetic complications and was an independent risk factor for carotid atherosclerosis.

Exploring the molecular mechanisms underlying α- and ß-cell dysfunction in diabetes.

Pancreatic islet dysfunction, including impaired insulin secretion in ß cells and dysregulated glucagon secretion in α cells, is the chief pathology of diabetes. In ß cells, oxidative stress, evoked by chronic hyperglycemia, was found to induce dysfunction of a critical transcription factor, PDX1, caused by its nucleocytoplasmic translocation via interactions with the insulin and JNK signaling pathways and another transcription factor, FOXO1. The significance of α-cell insulin signaling in the physiological and pathological regulation of α-cell biology was demonstrated in α-cell-specific insulin receptor knockout mice, which exhibited dysregulated glucagon secretion. Moreover, a high-glucose load directly induced excessive glucagon secretion in a glucagon-secreting cell line and isolated islets, together with impairment of insulin signaling. These findings indicate that disordered insulin signaling is central to the pathophysiology of islet dysfunction in both α and ß cells. On the other hand, certain beneficial effects of GLP-1 on dysfunctional α and ß cells indicate that it has therapeutic potential for diabetes patients who exhibit insulin resistance in islets. These studies, involving basic medical research approaches, have-at least in part-clarified the molecular mechanisms underlying α- and ß-cell dysfunction in diabetes, and offer important clues that should aid the development of future therapeutic approaches to the disease.

Significant elevation of serum dipeptidyl peptidase-4 activity in young-adult type 1 diabetes.

AIMS: Currently, inhibition of dipeptidyl peptidase-4 (DPP-4) is widely used in the treatment of type 2 diabetes. Application of this strategy is awaited as a new therapeutic approach for type 1 diabetes, but the scientific basis is still lacking. This report describes the evaluation of serum DPP-4 activity in type 1 diabetes compared with control subjects, and assessment of relationships between DPP-4 activity and diabetic complication markers and metabolic variables in type 1 diabetes. METHODS: We examined serum DPP-4 activity in Japanese young-adult type 1 diabetes (n=76, females 69.7%, age 30.9 ± 6.2 years, duration of diabetes 16.5 ± 11.1 years; mean ± SD) and healthy controls (n=22). Association of the enzymatic activity with diabetic micro- and macro- vascular complication markers and clinical parameters was also assessed. RESULTS: Subjects with type 1 diabetes displayed significantly higher serum DPP-4 activity than healthy controls (relative value, control: 1.00 ± 0.28, T1D, 1.29 ± 0.38; p=0.0011) independent of other clinical parameters. In type 1 diabetes, DPP-4 activity was positively correlated with duration of diabetes (r=0.248, p=0.031), while not correlated with HbA1c level. In univariate correlation analysis of diabetic complication markers and other metabolic parameters, coefficient of variation of R-R intervals (CVR-R) and gamma (γ)-glutamyltransferase (GGT) levels were correlated with DPP-4 activity. GGT was extracted as an independent variable of DPP-4 activity in multivariate analysis (ß=0.213, p=0.035). CONCLUSIONS: Serum DPP-4 activity is significantly elevated in Japanese type 1 diabetes, suggesting pathophysiological significance of the enzyme in type 1 diabetes.