Insulin secretion from human beta cells is heterogeneous and dependent on cell-to-cell contacts

Aims/hypothesis: We assessed the heterogeneity of insulin secretion from human isolated beta cells and its regulation by cell-to-cell contacts. Methods: Insulin secretion from single and paired cells was assessed by a reverse haemolytic plaque assay. The percentage of plaque-forming cells, the mean plaque area and the total plaque development were evaluated after 1h of stimulation with different secretagogues. Results: Not all beta cells were surrounded by a haemolytic plaque under all conditions tested. A small fraction of the beta cell population (20%) secreted more than 90% and 70% of total insulin at 2.2 and 22.2mmol/l glucose, respectively. Plaque-forming cells, mean plaque area and total plaque development were increased at 12.2 and 22.2 compared with 2.2mmol/l glucose. Insulin secretion of single beta cells was similar at 12.2 and 22.2mmol/l glucose. Insulin secretion of beta cell pairs was increased compared with that of single beta cells and was higher at 22.2 than at 12.2mmol/l glucose. Insulin secretion of beta cells in contact with alpha cells was also increased compared with single beta cells, but was similar at 22.2 compared with 12.2mmol/l glucose. Delta and other non-beta cells did not increase insulin secretion of contacting beta cells compared with that of single beta cells. Differences in insulin secretion between 22.2 and 12.2mmol/l glucose were observed in murine but not in human islets. Conclusions/interpretation: Human beta cells are highly heterogeneous in terms of insulin secretion so that a small fraction of beta cells contributes to the majority of insulin secreted. Homologous and heterologous intercellular contacts have a significant impact on insulin secretion and this could be related to the particular architecture of human islet

Wirkung von Ethanol auf die neuronale Expression von Zytokinen und Neurotrophinen in einem murinen Pneumoniemodell

Titel: Wirkung von Ethanol auf die neuronale Expression von Zytokinen und Neurotrophinen in einem murinen Pneumoniemodell. Einleitung: Chronischer Alkoholkonsum verursacht eine Funktionsstörung des Immunsystems und erhöht das Risiko postoperativer Sterblichkeit in Folge nosokominaler Infektionen. Periphere Entzündungen als Reaktion auf Operationen oder Infektionen können die Funktion des zentralen Nervensystems (ZNS) beeinträchtigen, einschließlich Gedächtnis und Kognition. Das Ziel dieser Arbeit war es, zu untersuchen, wie chronischer Alkoholkonsum die Biosynthese der proinflammatorischen Zytokine IL-1beta und TNF-alpha sowie der Neurotrophine BDNF und NT-3 während einer postoperativen Infektion mit Klebsiella Pneumoniae im Gehirn beeinflusst. Methodik: 94 weibliche 9 Wochen alte Balb/c-Mäuse wurden über 14 Tage mit Ethanol (EtOH, 3,8 mg/kg/KG) bzw. NaCl vorbehandelt (i.p.) und am 14. Tag laparotomiert. Am 16. Tag des Versuches wurde den Tieren entweder NaCl-Lösung oder Klebsiella pneumoniae nasal appliziert. 24 h bzw. 48 h danach erfolgte die Tötung und die Präparation des Gehirns. Die Expression der Zytokine und Neurotrophine wurde auf Genebene mittels quantitativer Real Time PCR, auf Proteinebene mittels Enzyme-linked immunosorbent Assay (ELISA) oder Western Blot Analyse bestimmt. Ergebnisse: Eine postoperative Infektion mit Klebsiella pneumoniae führte nach 24 h zu einer signifikant erhöhten Genexpression von IL-1beta und TNF-alpha im Gehirn der Mäuse, eine preoperative Ethanolgabe über 2 Wochen hemmt diese Genexpression signifikant. Bei allen Tieren ist die BDNF-Genexpression umgekehrt proportional zur IL-1beta-Genexpression (rxy=-0,4147, p<0,0001). Eine chronische Alkoholgabe hat keinen Einfluss auf die Genexpression von BDNF, die Genexpression von NT-3 sinkt am vierten postoperativen Tag signifikant sowie am dritten postoperativen Tag während der Pneumonie. Ein zeitlicher Vergleich zeigte bei allen operierten Tieren ohne Pneumonie eine signifikante Zunahme der IL-1beta Genexpression am 4. postoperativen Tag im Vergleich zum 3. Tag, für TNF-alpha nur bei den Sham Tieren. Schlussfolgerung: Eine Alkoholbehandlung über 2 Wochen senkt die Genexpression der proinflammatorischen Zytokine IL-1beta und TNF-alpha während einer postoperativen Infektion mit Klebsiella Pneumoniae im Gehirn von Balb/c-Mäusen. Welche Auswirkungen die Hemmung der proinflammatorischen Zytokine im Gehirn haben, könnten Zelltodmessungen sowie Verhaltenstests klären.Title: The influence of alcohol on a neuronal cytokine and neurotrophine expression in a murine pneumonia model. Background: A chronic alcohol administration causes an immune system dysfunction and increases the risk of postoperative mortality caused by nosocomial infection. Peripheral inflammation in response to surgery or infection can affect the function of the central nervous system (CNS), including memory and cognition. The purpose of this study was to analyze the influence of a chronic alcohol administration on the production of the proinflammatory cytokines IL-1beta and TNF-alpha as well as neurotrophines BDNF and NT-3 in the brain during a postoperative infection with Klebsiella pneumoniae. Methods: 94 female 9 weeks old balb/c mice were treated for 14 days with ethanol (EtOH, 3.8 mg/kg body weight) or NaCl (i.p.). On the 14th day mice underwent a laparotomy and on the 16th day of the study they were treated with intranasal NaCl-solution or a solution containing Klebsiella pneumoniae. 24 h or 48 h later the animals were killed and their brains extracted. The expression of cytokines and neurotrophines were analzyed on a gene level using quantitaive real-time PCR and on a protein level using enzyme-linked immunosorbent assay (ELISA) or western blot. Results: A postoperative infection with Klebsiella pneumoniae led after 24 h to a significant increase of a IL1-beta and TNF-alpha gene expression in brains of mice. A preoperative administration of ethanol for 2 weeks was significantly inhibiting expression of these genes. In all animals the expression of BDNF gene was inversely proportional to a IL1-beta gene expression (rxy=-0,4147, p<0,0001). A chronic alcohol administration had no influence on expression of BDNF gene. The expression of NT-3 gene was significantly reduced on a fourth postoperative day without and on the third postoperative day with pneumonia. The time analysis showed a significant increase of IL-1beta gene expression on 4th postoperative day in comparison to 3rd day in all operated animals without a pneumonia. In case of TNF-alpha this effect was observed only in the Sham-group. Conclusions: An administration of alcohol for 2 weeks reduced the gene expression of proinflammatory cytokines IL-1beta and TNF-alpha during the postoperative infection with Klebsiella pneumoniae in the brain of balb/c-mice. The influence of proinflammatory cytokine inhibition on the brain could be clarified using cell death assays as well as behavioral tests

Respective Contributions of Glycemic Variability and Mean Daily Glucose as Predictors of Hypoglycemia in Type 1 Diabetes: Are They Equivalent?

To evaluate the respective contributions of short-term glycemic variability and mean daily glucose (MDG) concentration to the risk of hypoglycemia in type 1 diabetes. People with type 1 diabetes (n = 100) investigated at the University Hospital of Montpellier (France) underwent continuous glucose monitoring (CGM) on two consecutive days, providing a total of 200 24-h glycemic profiles. The following parameters were computed: MDG concentration, within-day glycemic variability (coefficient of variation for glucose [%CV]), and risk of hypoglycemia (presented as the percentage of time spent below three glycemic thresholds: 3.9, 3.45, and 3.0 mmol/L). MDG was significantly higher, and %CV significantly lower (both P &lt; 0.001), when comparing the 24-h glycemic profiles according to whether no time or a certain duration of time was spent below the thresholds. Univariate regression analyses showed that MDG and %CV were the two explanatory variables that entered the model with the outcome variable (time spent below the thresholds). The classification and regression tree procedure indicated that the predominant predictor for hypoglycemia was %CV when the threshold was 3.0 mmol/L. In people with mean glucose ≤7.8 mmol/L, the time spent below 3.0 mmol/L was shortest (P &lt; 0.001) when %CV was below 34%. In type 1 diabetes, short-term glycemic variability relative to mean glucose (i.e., %CV) explains more hypoglycemia than does mean glucose alone when the glucose threshold is 3.0 mmol/L. Minimizing the risk of hypoglycemia requires a %CV below 34%

Toward Defining the Threshold Between Low and High Glucose Variability in Diabetes

International audienceOBJECTIVE:To define the threshold for excess glucose variability (GV), one of the main features of dysglycemia in diabetes.RESEARCH DESIGN AND METHODS:A total of 376 persons with diabetes investigated at the University Hospital of Montpellier (Montpellier, France) underwent continuous glucose monitoring. Participants with type 2 diabetes were divided into several groups-groups 1, 2a, 2b, and 3 (n = 82, 28, 65, and 79, respectively)-according to treatment: 1) diet and/or insulin sensitizers alone; 2) oral therapy including an insulinotropic agent, dipeptidyl peptidase 4 inhibitors (group 2a) or sulfonylureas (group 2b); or 3) insulin. Group 4 included 122 persons with type 1 diabetes. Percentage coefficient of variation for glucose (%CV = [(SD of glucose)/(mean glucose)] × 100) and frequencies of hypoglycemia (interstitial glucose 36% were compared with those with %CV ≤36%.CONCLUSIONS:A %CV of 36% appears to be a suitable threshold to distinguish between stable and unstable glycemia in diabetes because beyond this limit, the frequency of hypoglycemia is significantly increased, especially in insulin-treated subjects

Real world hypoglycaemia related to glucose variability and Flash glucose scan frequency assessed from global FreeStyle Libre data.

Flash glucose monitoring provides a range of glucose metrics. In the current study, we aim to identify those that indicate that glycaemic targets can be consistently met and contrast the total (t-CV) and within-day coefficient of variation (wd-CV) to guide the assessment of glucose variability and hypoglycaemia exposure. De-identified data from Flash readers were collected. The readers were sorted into 10 equally sized groups of scan frequency followed by quartiles of estimated A1c (eA1c). A similar grouping was performed for the total coefficient of variation (t-CV) and within-day coefficient of variation (wd-CV). In addition, analysis of the association of time below 54 mg/dl and glucose variability measured by t-CV and wd-CV was performed. The dataset included 1 002 946 readers. Readers sorted by 10 equal groups of scan rate and quartiles by eA1c, t-CV and wd-CV represented 25 074 readers per group. The association of lower eA1c with higher time in range and reduced time above range was clear. The correlation of eA1c quartiles and time below range was not consistent. An association between glucose variability and hypoglycaemia was found. Both wd-CV and t-CV were associated with time below range. For achieving the consensus target of &lt;1% time below 54 mg/dl, the associated wd-CV and t-CV values were 33.5% and 39.5%, respectively. The type of CV reported by the different continuous glucose monitoring systems should be acknowledged. CV &lt;36% might not be adequate to ensure low hypoglycaemia exposure. To our knowledge, the majority of continuous glucose monitoring reports the t-CV. Appropriate thresholds should be used to identify patients that would probably meet time below range targets (t-CV &lt;40% or wd-CV &lt;34%)

Precision medicine in the era of artificial intelligence: implications in chronic disease management.

Aberrant metabolism is the root cause of several serious health issues, creating a huge burden to health and leading to diminished life expectancy. A dysregulated metabolism induces the secretion of several molecules which in turn trigger the inflammatory pathway. Inflammation is the natural reaction of the immune system to a variety of stimuli, such as pathogens, damaged cells, and harmful substances. Metabolically triggered inflammation, also called metaflammation or low-grade chronic inflammation, is the consequence of a synergic interaction between the host and the exposome-a combination of environmental drivers, including diet, lifestyle, pollutants and other factors throughout the life span of an individual. Various levels of chronic inflammation are associated with several lifestyle-related diseases such as diabetes, obesity, metabolic associated fatty liver disease (MAFLD), cancers, cardiovascular disorders (CVDs), autoimmune diseases, and chronic lung diseases. Chronic diseases are a growing concern worldwide, placing a heavy burden on individuals, families, governments, and health-care systems. New strategies are needed to empower communities worldwide to prevent and treat these diseases. Precision medicine provides a model for the next generation of lifestyle modification. This will capitalize on the dynamic interaction between an individual's biology, lifestyle, behavior, and environment. The aim of precision medicine is to design and improve diagnosis, therapeutics and prognostication through the use of large complex datasets that incorporate individual gene, function, and environmental variations. The implementation of high-performance computing (HPC) and artificial intelligence (AI) can predict risks with greater accuracy based on available multidimensional clinical and biological datasets. AI-powered precision medicine provides clinicians with an opportunity to specifically tailor early interventions to each individual. In this article, we discuss the strengths and limitations of existing and evolving recent, data-driven technologies, such as AI, in preventing, treating and reversing lifestyle-related diseases

The diabetes-linked transcription factor Pax4 is expressed in human pancreatic islets and is activated by mitogens and GLP-1

We previously demonstrated that the transcription factor Pax4 is important for β-cell replication and survival in rat islets. Herein, we investigate Pax4 expression in islets of non-diabetic and diabetic donors, its regulation by mitogens, glucose and the incretin GLP-1 and evaluate its effect on human islet proliferation. Pax4 expression was increased in islets derived from Type 2 diabetic donors correlating with hyperglycaemia. In vitro studies on non diabetic islets demonstrated that glucose, betacellulin, activin A, GLP-1 and insulin increased Pax4 mRNA levels. Glucose-induced Pax4 expression was abolished by the inhibitors LY294002, PD98050 or H89. Surprisingly, increases in Pax4 expression did not prompt a surge in human islet cell replication. Furthermore, expression of the proliferation marker gene Id2 remained unaltered. Adenoviral-mediated expression of human Pax4 resulted in a small increase in Bcl-xL expression while Id2 transcript levels and cell replication were unchanged in human islets. In contrast, overexpression of mouse Pax4 induced human islet cell proliferation. Treatment of islets with 5-Aza-2′-deoxycytidine induced Pax4 without stimulating Bcl-xL and Id2 expression. Human Pax4 DNA binding activity was found to be lower than that of the mouse homologue. Thus, human pax4 gene expression is epigenetically regulated and induced by physiological stimuli through the concerted action of multiple signalling pathways. However, it is unable to initiate the transcriptional replication program likely due to post-translational modifications of the protein. The latter highlights fundamental differences between human and rodent islet physiology and emphasizes the importance of validating results obtained with animal models in human tissue

LDHA is enriched in human islet alpha cells and upregulated in type 2 diabetes

The lactate dehydrogenase isoform A (LDHA) is a key metabolic enzyme that preferentially catalyzes the conversion of pyruvate to lactate. Whereas LDHA is highly expressed in many tissues, its expression is turned off in the differentiated adult β-cell within the pancreatic islets. The repression of LDHA under normal physiological condition and its inappropriate upregulation under a diabetogenic environment is well-documented in rodent islets/β-cells but little is known about LDHA expression in human islet cells and whether its abundance is altered under diabetic conditions. Analysis of public single-cell RNA-seq (sc-RNA seq) data as well as cell type-specific immunolabeling of human pancreatic islets showed that LDHA was mainly localized in human α-cells while it is expressed at a very low level in β-cells. Furthermore, LDHA, both at mRNA and protein, as well as lactate production is upregulated in human pancreatic islets exposed to chronic high glucose treatment. Microscopic analysis of stressed human islets and autopsy pancreases from individuals with type 2 diabetes (T2D) showed LDHA upregulation mainly in human α-cells. Pharmacological inhibition of LDHA in isolated human islets enhanced insulin secretion under physiological conditions but did not significantly correct the deregulated secretion of insulin or glucagon under diabetic conditions

Proteasomal degradation of the histone acetyl transferase p300 contributes to beta-cell injury in a diabetes environment

In type 2 diabetes, amyloid oligomers, chronic hyperglycemia, lipotoxicity, and pro-inflammatory cytokines are detrimental to beta-cells, causing apoptosis and impaired insulin secretion. The histone acetyl transferase p300, involved in remodeling of chromatin structure by epigenetic mechanisms, is a key ubiquitous activator of the transcriptional machinery. In this study, we report that loss of p300 acetyl transferase activity and expression leads to beta-cell apoptosis, and most importantly, that stress situations known to be associated with diabetes alter p300 levels and functional integrity. We found that proteasomal degradation is the mechanism subserving p300 loss in beta-cells exposed to hyperglycemia or pro-inflammatory cytokines. We also report that melatonin, a hormone produced in the pineal gland and known to play key roles in beta-cell health, preserves p300 levels altered by these toxic conditions. Collectively, these data imply an important role for p300 in the pathophysiology of diabetes