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Background: Managing metabolism for optimal training, performance, and recovery in medium-to-high-level endurance runners involves enhancing energy systems through strategic nutrient intake. Optimal carbohydrate intake before, during, and after endurance running can enhance glycogen stores and maintain optimal blood glucose levels, influencing various physiological responses and adaptations, including transitory post-endurance inflammation. This randomized trial investigates the impact of a high-dose 2:1 maltodextrin–fructose supplementation to medium-to-high-level endurance runners immediately before, during, and after a 15 km run at 90% VO2max intensity on post-exercise inflammatory stress. Methods: We evaluated inflammatory biomarkers and lipidomic profiles before the endurance tests and up to 24 h after. We focused on the effects of high-dose 2:1 maltodextrin–fructose supplementation on white blood cell count, neutrophil number, IL-6, cortisol, and CRP levels, as well as polyunsaturated fatty acids, ω-3 index, and AA/EPA ratio. Results: This supplementation significantly reduced inflammatory markers and metabolic stress. Additionally, it may enhance the post-activity increase in blood ω-3 fatty acid levels and reduce the increase in ω-6 levels, resulting in a lower trend of AA/EPA ratio at 24 h in the treated arm. Conclusions: Adequate carbohydrate supplementation may acutely mitigate inflammation during a one-hour endurance activity of moderate-to-high intensity. These effects could be beneficial for athletes engaging in frequent, high-intensity activities.

Los trastornos autoinmunes representan una familia de al menos 80 condiciones diferentes que surgen de una respuesta aberrante del sistema inmunológico resultando finalmente en la destrucción de tejidos y órganos específicos del cuerpo. Es importante destacar que durante las últimas tres décadas los estudios epidemiológicos han proporcionado evidencia de un aumento constante en la incidencia y prevalencia de trastornos autoinmunes. En los últimos años, varios estudios han demostrado que la vitamina D y los ácidos grasos poliinsaturados (AGPs) omega-3 ejercen propiedades inmunomoduladoras y antiinflamatorias sinérgicas que pueden aprovecharse positivamente para la prevención y el tratamiento de trastornos autoinmunes. En este sentido, el reciente ensayo clínico denominado VITAL (ensayo de vitamina D y omega 3); un estudio a gran escala, aleatorizado, doble ciego, controlado con placebo encontró que la suplementación conjunta de vitamina D y AGPs omega-3 (VIDOM) puede reducir la incidencia de enfermedades autoinmunes. En esta revisión de la literatura, resumimos los mecanismos moleculares detrás de las propiedades inmunomoduladoras y antiinflamatorias de la vitamina D y los AGPs omega-3, así como la posible interacción bidireccional entre el metabolismo de la vitamina D y el metabolismo de los AGPs omega-3 que justifica la co- suplementación VIDOM en trastornos autoinmunes.

IntroductionIslet transplantation (ITx) shows promise in treating T1D, but the role of islet autoantibodies on graft survival has not been clearly elucidated. We aimed to analyze the effect of GAD65 and IA2 autoantibody status on graft survival and attainment of insulin independence in subjects with T1D who underwent ITx.MethodWe conducted a retrospective cohort study on 47 ITx recipients from 2000 to 2018. Islet infusion was performed via intrahepatic portal (n=44) or onto the omentum via laparoscopic approach (n=3). Immunosuppression involved anti-IL2 receptor antibody, anti-TNF, and dual combinations of sirolimus, tacrolimus, or mycophenolate mofetil (Edmonton-like) in 38 subjects (80.9%). T-cell depletion induction with Edmonton-like maintenance was used in 9 subjects (19%). GAD65 and IA2 autoantibodies were assessed pre-transplant and post-transplant (monthly) until graft failure, and categorized as persistently negative, persistently positive, or seroconverters. Graft survival was analyzed using U-Mann-Whitney test, and Quade’s nonparametric ANCOVA adjusted for confounders. Kaplan-Meier and Log-Rank tests were employed to analyze attainment of insulin independence. P value

Spontaneous bacterial peritonitis (SBP) is a severe complication in patients with decompensated liver cirrhosis and is commonly treated with broad spectrum antibiotics. However, the rise of antibiotic resistance requires alternative therapeutic strategies. As recently shown, human amnion-derived mesenchymal stem cells (hA-MSCs) are able, in vitro, to promote bacterial clearance and modulate the immune and inflammatory response in SBP. Our results highlight the upregulation of FOXO1, CXCL5, CXCL6, CCL20, and MAPK13 in hA-MSCs as well as the promotion of bacterial clearance, prompting a shift in the immune response toward a Th17 lymphocyte phenotype after 72 h treatment. In this study, we used an in vitro SBP model and employed omics techniques (next-generation sequencing) to investigate the mechanisms by which hA-MSCs modify the crosstalk between immune cells in LPS-stimulated ascitic fluid. We also validated the data obtained via qRT-PCR, cytofluorimetric analysis, and Luminex assay. These findings provide further support to the hope of using hA-MSCs for the prevention and treatment of infective diseases, such as SBP, offering a viable alternative to antibiotic therapy.

Islet cell transplantation (ITx) is an effective therapeutic approach for selected patients with type 1 diabetes with hypoglycemia unawareness and severe hypoglycemia events. In organ transplantation, human leukocyte antigen (HLA) mismatching between donor and recipient negatively impacts transplant outcomes. We aimed to determine whether HLA matching has an impact on islet allograft survival. Forty-eight patients were followed up after islet transplantation at our institution from 2000 to 2020 in a retrospective cohort. Patients underwent intrahepatic ITx or laparoscopic omental approach. Immunosuppression was dependent upon the protocol. We analyzed HLA data restricted to A, B, and DR loci on allograft survival using survival and subsequent multivariable analyses. Patients were aged 42.8 ± 8.4 years, and 64.3% were female. Diabetes duration was 28.6 ± 11.6 years. Patients matching all three HLA loci presented longer graft survival ( P = 0.030). Patients with ≥1 HLA-B matching had longer graft survival compared with zero matching ( P = 0.025). The number of HLA-B matching was positively associated with time of graft survival (Spearman’s rho = 0.590; P = 0.034). Analyses adjusted for confounders showed that ≥1 matching for HLA-B decreased the risk of allograft failure ( P = 0.009). Our data suggest that HLA-B matching between recipients and donors improved islet allograft survival. Matching all three HLA loci (A, B, and DR) was also associated with prolonged islet allograft survival. Prospective studies and a larger sample size are warranted to validate our findings.

Islet transplantation for type 1 diabetes management is hindered by the life-long need for immunosuppressive medications. Here, the authors report an islet encapsulation device with local anti-rejection drug release that achieves long-term diabetes reversal in male rats and reduces drug-related toxicity.

ABSTRACT Objective: The occurrence of partial remission (honeymoon phase) in type 1 diabetes (T1D) has been associated with a reduced risk of chronic microvascular complications of diabetes. We have published case reports showing that a combination therapy with the DPP-4 inhibitor sitagliptin plus vitamin D3 (VIDPP-4i) can prolong the honeymoon phase in patients with new-onset T1D. In the present case-control study, we investigated the frequency of occurrence of clinical remission (CR) in patients with new-onset T1D after VIDPP-4i treatment. Subjects and methods: In this case-control study, we collected data spanning 10 years from medical records of 46 patients (23 females) recently diagnosed with T1D. Overall, 27 participants with CR (insulin dose-adjusted glycated hemoglobin [IDAA1c] ≤ 9) at 12 or 24 months composed the case group, and 19 participants without CR served as the control group. Chi-square with Yates correction was used to analyze the association between VIDPP-4i use and CR, and odds ratio (OR) was used to determine the chance of CR due to VIDPP-4i treatment exposure. Results: In all, 37 patients (80.4%) experienced CR at some time over 24 months. The mean CR duration was 13.15 ± 9.91 months. Treatment with VIDPP-4i was significantly associated with CR. At 24 months, the OR of CR after VIDPP-4i exposure was 9.0 (95% confidence interval [CI] 2.21-30.18, p = 0.0036). Additionally, 9 (33.6%) and 4 (14.8%) patients in the VIDPP-4i group experienced insulin-free CR at 12 and 24 months, respectively. Conclusion: Therapy with VIDPP-4i was associated with a higher frequency and duration of the honeymoon phase. Randomized controlled trials are needed to confirm these findings.

Diabetic foot ulcer (DFU) is associated with neuropathy and/or peripheral artery disease of the lower limb in diabetic patients, affecting quality of life and leading to repeated hospitalizations and infections [...]

Abstract The pathophysiology of coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and especially of its complications is still not fully understood. In fact, a very high number of patients with COVID-19 die because of thromboembolic causes. A role of plasminogen, as precursor of fibrinolysis, has been hypothesized. In this study, we aimed to investigate the association between plasminogen levels and COVID-19-related outcomes in a population of 55 infected Caucasian patients (mean age: 69.8 ± 14.3, 41.8% female). Low levels of plasminogen were significantly associated with inflammatory markers (CRP, PCT, and IL-6), markers of coagulation (D-dimer, INR, and APTT), and markers of organ dysfunctions (high fasting blood glucose and decrease in the glomerular filtration rate). A multidimensional analysis model, including the correlation of the expression of coagulation with inflammatory parameters, indicated that plasminogen tended to cluster together with IL-6, hence suggesting a common pathway of activation during disease’s complication. Moreover, low levels of plasminogen strongly correlated with mortality in COVID-19 patients even after multiple adjustments for presence of confounding. These data suggest that plasminogen may play a pivotal role in controlling the complex mechanisms beyond the COVID-19 complications, and may be useful both as biomarker for prognosis and for therapeutic target against this extremely aggressive infection.

Abstract Acute respiratory distress syndrome (ARDS) in COVID‐19 is associated with high mortality. Mesenchymal stem cells are known to exert immunomodulatory and anti‐inflammatory effects and could yield beneficial effects in COVID‐19 ARDS. The objective of this study was to determine safety and explore efficacy of umbilical cord mesenchymal stem cell (UC‐MSC) infusions in subjects with COVID‐19 ARDS. A double‐blind, phase 1/2a, randomized, controlled trial was performed. Randomization and stratification by ARDS severity was used to foster balance among groups. All subjects were analyzed under intention to treat design. Twenty‐four subjects were randomized 1:1 to either UC‐MSC treatment (n = 12) or the control group (n = 12). Subjects in the UC‐MSC treatment group received two intravenous infusions (at day 0 and 3) of 100 ± 20 × 106 UC‐MSCs; controls received two infusions of vehicle solution. Both groups received best standard of care. Primary endpoint was safety (adverse events [AEs]) within 6 hours; cardiac arrest or death within 24 hours postinfusion). Secondary endpoints included patient survival at 31 days after the first infusion and time to recovery. No difference was observed between groups in infusion‐associated AEs. No serious adverse events (SAEs) were observed related to UC‐MSC infusions. UC‐MSC infusions in COVID‐19 ARDS were found to be safe. Inflammatory cytokines were significantly decreased in UC‐MSC‐treated subjects at day 6. Treatment was associated with significantly improved patient survival (91% vs 42%, P = .015), SAE‐free survival (P = .008), and time to recovery (P = .03). UC‐MSC infusions are safe and could be beneficial in treating subjects with COVID‐19 ARDS.

Background: Obesity is a pandemic disease characterized by excessive severe body comorbidities. Reduction in fat accumulation represents a mechanism of prevention, and the replacement of white adipose tissue (WAT) with brown adipose tissue (BAT) has been proposed as one promising strategy against obesity. In the present study, we sought to investigate the ability of a natural mixture of polyphenols and micronutrients (A5+) to counteract white adipogenesis by promoting WAT browning. Methods: For this study, we employed a murine 3T3-L1 fibroblast cell line treated with A5+, or DMSO as control, during the differentiation in mature adipocytes for 10 days. Cell cycle analysis was performed using propidium iodide staining and cytofluorimetric analysis. Intracellular lipid contents were detected by Oil Red O staining. Inflammation Array, along with qRT-PCR and Western Blot analyses, served to measure the expression of the analyzed markers, such as pro-inflammatory cytokines. Results: A5+ administration significantly reduced lipids’ accumulation in adipocytes when compared to control cells (p < 0.005). Similarly, A5+ inhibited cellular proliferation during the mitotic clonal expansion (MCE), the most relevant stage in adipocytes differentiation (p < 0.0001). We also found that A5+ significantly reduced the release of pro-inflammatory cytokines, such as IL-6 and Leptin (p < 0.005), and promoted fat browning and fatty acid oxidation through increasing expression levels of genes related to BAT, such as UCP1 (p < 0.05). This thermogenic process is mediated via AMPK-ATGL pathway activation. Conclusion: Overall, these results demonstrated that the synergistic effect of compounds contained in A5+ may be able to counteract adipogenesis and then obesity by inducing fat browning.

The ability to culture live pancreatic tissue slices for long periods of time would enable longitudinal studies ex vivo. Here the authors culture human and mouse pancreatic slices in a perfluorocarbon-based culture system and show stable endocrine and exocrine function for up to ten days in culture.

Summary: The scarcity of donors and need for immunosuppression limit pancreatic islet transplantation to a few patients with labile type 1 diabetes. Transplantation of encapsulated stem cell-derived islets (SC islets) might extend the applicability of islet transplantation to a larger cohort of patients. Transplantation of conformal-coated islets into a confined well-vascularized site allows long-term diabetes reversal in fully MHC-mismatched diabetic mice without immunosuppression. Here, we demonstrated that human SC islets reaggregated from cryopreserved cells display glucose-stimulated insulin secretion in vitro. Importantly, we showed that conformally coated SC islets displayed comparable in vitro function with unencapsulated SC islets, with conformal coating permitting physiological insulin secretion. Transplantation of SC islets into the gonadal fat pad of diabetic NOD-scid mice revealed that both unencapsulated and conformal-coated SC islets could reverse diabetes and maintain human-level euglycemia for more than 80 days. Overall, these results provide support for further evaluation of safety and efficacy of conformal-coated SC islets in larger species. : Scarcity of donors and need for immunosuppression limit pancreatic islet transplantation to a few patients with labile type 1 diabetes. Islet encapsulation may eliminate the need for chronic immunosuppression. Conformal coating seeks to overcome limitations of traditional microencapsulation. Transplantation of conformal-coated stem cell-derived islets might extend the applicability of islet transplantation to a larger cohort of patients. Keywords: encapsulation, β cells, polyethylene glycol, alginate, insulin, Type 1 Diabetes, islet transplantation, stem cell-derived islets, conformal coating, thiol-ene click chemistry

Clinical islet allotransplantation has been successfully regulated as tissue/organ for transplantation in number of countries and is recognized as a safe and efficacious therapy for selected patients with type 1 diabetes mellitus. However, in the United States, the FDA considers pancreatic islets as a biologic drug, and islet transplantation has not yet shifted from the experimental to the clinical arena for last 20 years. In order to transplant islets, the FDA requires a valid Biological License Application (BLA) in place. The BLA process is costly and lengthy. However, despite the application of drug manufacturing technology and regulations, the final islet product sterility and potency cannot be confirmed, even when islets meet all the predetermined release criteria. Therefore, further regulation of islets as drugs is obsolete and will continue to hinder clinical application of islet transplantation in the US. The Organ Procurement and Transplantation Network together with the United Network for Organ Sharing have developed separately from the FDA and BLA regulatory framework for human organs under the Human Resources & Services Administration to assure safety and efficacy of transplantation. Based on similar biologic characteristics of islets and human organs, we propose inclusion of islets into the existing regulatory framework for organs for transplantation, along with continued FDA oversight for islet processing, as it is for other cell/tissue products exempt from BLA. This approach would reassure islet quality, efficacy and access for Americans with diabetes to this effective procedure.

Summary: The transplantation of human embryonic stem cell (hESC)-derived insulin-producing β cells for the treatment of diabetes is finally approaching the clinical stage. However, even with state-of-the-art differentiation protocols, a significant percentage of undefined non-endocrine cell types are still generated. Most importantly, there is the potential for carry-over of non-differentiated cell types that may produce teratomas. We sought to modify hESCs so that their differentiated progeny could be selectively devoid of tumorigenic cells and enriched for cells of the desired phenotype (in this case, β cells). Here we report the generation of a modified hESC line harboring two suicide gene cassettes, whose expression results in cell death in the presence of specific pro-drugs. We show the efficacy of this system at enriching for β cells and eliminating tumorigenic ones both in vitro and in vivo. Our approach is innovative inasmuch as it allows for the preservation of the desired cells while eliminating those with the potential to develop teratomas. : In this article, Domínguez-Bendala and colleagues present a strategy to modify pluripotent stem cells to allow for the selective destruction of tumorigenic escapees and/or fully developed teratomas, as well as for the specific selection of differentiated insulin-producing β-like cells. Cells engineered in this manner feature a double fail-safe mechanism designed to minimize the risks associated with their clinical transplantation. Keywords: human embryonic stem cells, beta cell differentiation, suicide genes, teratoma, transplantation

Type 1 diabetes (T1D), which is caused by the autoimmune destruction of insulin-secreting pancreatic beta cells, represents a high-risk category requiring COVID-19 vaccine prioritization. Although COVID-19 vaccination can lead to transient hyperglycemia (vaccination-induced hyperglycemia; ViHG), its influence on the course of the clinical remission phase of T1D (a.k.a. “honeymoon phase”) is currently unknown. Recently, there has been an increasing concern that COVID-19 vaccination may trigger autoimmune phenomena. We describe the case of a 24-year-old young Italian man with T1D who received two doses of the BNT162b2 mRNA (Pfizer-BioNTech) COVID-19 vaccine during a prolonged honeymoon phase. He experienced a transient impairment in glucose control (as evidenced by continuous glucose monitoring) that was not associated with substantial changes in stimulated C-peptide levels and islet autoantibody titers. Nonetheless, large prospective studies are needed to confirm the safety and the immunometabolic impact of the BNT162b2 vaccine in T1D patients during the honeymoon phase. Thus far, T1D patients who are going to receive COVID-19 vaccination should be warned about the possible occurrence of transient ViHG and should undergo strict postvaccination surveillance.

Systemic inflammation represents a shared pathophysiological mechanism which underlies the frequent clinical associations among chronic inflammatory rheumatic diseases (CIRDs), insulin resistance, type 2 diabetes (T2D), and chronic diabetes complications, including cardiovascular disease. Therefore, targeted anti-inflammatory therapies are attractive and highly desirable interventions to concomitantly reduce rheumatic disease activity and to improve glucose control in patients with CIRDs and comorbid T2D. Therapeutic approaches targeting inflammation may also play a role in the prevention of prediabetes and diabetes in patients with CIRDs, particularly in those with traditional risk factors and/or on high-dose corticosteroid therapy. Recently, several studies have shown that different disease-modifying antirheumatic drugs (DMARDs) used for the treatment of CIRDs exert antihyperglycemic properties by virtue of their anti-inflammatory, insulin-sensitizing, and/or insulinotropic effects. In this view, DMARDs are promising drug candidates that may potentially reduce rheumatic disease activity, ameliorate glucose control, and at the same time, prevent the development of diabetes-associated cardiovascular complications and metabolic dysfunctions. In light of their substantial antidiabetic actions, some DMARDs (such as hydroxychloroquine and anakinra) could be alternatively termed “diabetes-modifying antirheumatic drugs”, since they may be repurposed for co-treatment of rheumatic diseases and comorbid T2D. However, there is a need for future randomized controlled trials to confirm the beneficial metabolic and cardiovascular effects as well as the safety profile of distinct DMARDs in the long term. This narrative review aims to discuss the current knowledge about the mechanisms behind the antihyperglycemic properties exerted by a variety of DMARDs (including synthetic and biologic DMARDs) and the potential use of these agents as antidiabetic medications in clinical settings.

Parkinson’s disease (PD) is second-most common disabling neurological disorder worldwide, and unfortunately, there is not yet a definitive way to prevent it. Polyphenols have been widely shown protective efficacy against various PD symptoms. However, data on their effect on physio-pathological mechanisms underlying this disease are still lacking. In the present work, we evaluated the activity of a mixture of polyphenols and micronutrients, named A5+, in the murine neuroblastoma cell line N1E115 treated with 6-Hydroxydopamine (6-OHDA), an established neurotoxic stimulus used to induce an in vitro PD model. We demonstrate that a pretreatment of these cells with A5+ causes significant reduction of inflammation, resulting in a decrease in pro-inflammatory cytokines (IFN-γ, IL-6, TNF-α, and CXCL1), a reduction in ROS production and activation of extracellular signal-regulated kinases (ERK)1/2, and a decrease in apoptotic mechanisms with the related increase in cell viability. Intriguingly, A5+ treatment promoted cellular differentiation into dopaminergic neurons, as evident by the enhancement in the expression of tyrosine hydroxylase, a well-established dopaminergic neuronal marker. Overall, these results demonstrate the synergic and innovative efficacy of A5+ mixture against PD cellular pathological processes, although further studies are needed to clarify the mechanisms underlying its beneficial effect.

Here, we present a case that required a supplemental “old school” islet purification for a safe intraportal infusion. Following pancreas procurement from a brain-dead 26-year-old male donor (body mass index: 21.9), 24.6 ml of islet tissue was isolated after continuous density gradient centrifugation. The islet yield was 504,000 islet equivalent (IEQ), distributed among the following three fractions: 64,161 IEQ in 0.6 ml of pellet, 182,058 IEQ in 10 ml, and 258,010 IEQ in 14 ml with 95%, 20%, and 10% purity, respectively. After a 23-h culture, we applied supplemental islet purification, based on the separation of tissue subfractions during unit gravity sedimentation, a technique developed over 60 years ago (“old school”). This method enabled the reduction of the total pellet volume to 11.6 ml, while retaining 374,940 IEQ with a viability of over 90%. The final islet product was prepared in three infusion bags, containing 130,926 IEQ in 2.6 ml of pellet, 108,079 IEQ in 4 ml of pellet, and 135,935 IEQ in 5 ml of pellet with 65%, 40%, and 30% purity, respectively, and with the addition of unfractionated heparin (70 units/kg body weight). Upon the islet infusion from all three bags, portal pressure increased from 7 to 16 mmHg. Antithrombotic prophylaxis with heparin was continued for 48 h after the infusion, with target activated partial thromboplastin time 50–60 s, followed by fractionated heparin subcutaneous injections for 2 weeks. β-Cell graft function assessed on day 75 post-transplantation was good, according to Igls criteria, with complete elimination of severe hypoglycemic episodes and 50% reduction in insulin requirements. Time spent within the target glucose range (70–180 mg/dl) improved from 42% to 98% and HbA1c declined from 8.7% to 6.7%. Supplemental “old school” islet purification allowed for the safe and successful utilization of a robust and high-quality islet preparation, which otherwise would have been discarded.

Standardized islet characterization assays that can provide results in a timely manner are essential for successful islet cell transplantation. A critical component of islet cell quality is β-cell function, and perifusion-based assessments of dynamic glucose-stimulated insulin secretion (GSIS) are the most informative method to assess this, as they provide the most complex in vitro evaluation of GSIS. However, protocols used vary considerably among centers and investigators as they often use different low- and high-glucose concentrations, exposure-times, flow-rates, oxygen concentrations, islet numbers, analytical methods, measurement units, and instruments, which result in different readouts and make comparisons across platforms difficult. Additionally, the conditions of islet storage and shipment prior to assessment may also affect islet function. Establishing improved standardized protocols for perifusion GSIS assays should be an integral part of the ongoing effort to increase the rigor of human islet studies. Here, we performed detailed evaluation of GSIS of human islets using a fully automated multichannel perifusion instrument following various warm-up recovery times after cold storage that corresponds to current shipping conditions (8°C). We found that recovery times shorter than 18 h (overnight) resulted in impaired insulin secretion. While the effects were relatively moderate on second-phase insulin secretion, first-phase peaks were restored only following 18-h incubation. Hence, the biphasic profile of dynamic GSIS was considerably affected when islets were not allowed to recover for a sufficient time after being maintained in cold. Accordingly, while cold storage might improve islet cell survival during shipment and prolong the length of culture, functional assessments should be performed only after allowing for at least overnight recovery at physiological temperatures.

Background: A physiological hallmark of patients with type 2 diabetes mellitus (T2DM) is β cell dysfunction. Despite adequate treatment, it is an irreversible process that follows disease progression. Therefore, the development of novel therapies that restore β cell function is of utmost importance. Methods: This study aims to unveil the mechanistic action of mesenchymal stem cells (MSCs) by investigating its impact on isolated human T2DM islets ex vivo and in vivo. Findings: We propose that MSCs can attenuate β cell dysfunction by reversing β cell dedifferentiation in an IL-1Ra-mediated manner. In response to the elevated expression of proinflammatory cytokines in human T2DM islet cells, we observed that MSCs was activated to secret IL-1R antagonist (IL-1Ra) which acted on the inflammed islets and reversed β cell dedifferentiation, suggesting a crosstalk between MSCs and human T2DM islets. The co-transplantation of MSCs with human T2DM islets in diabetic SCID mice and intravenous infusion of MSCs in db/db mice revealed the reversal of β cell dedifferentiation and improved glycaemic control in the latter. Interpretation: This evidence highlights the potential of MSCs in future cell-based therapies regarding the amelioration of β cell dysfunction. Keywords: Mesenchymal stem cells, Type 2 diabetes mellitus, β cell dysfunction, Inflammation, β cell dedifferentiation

Summary: Treatment of human pancreatic non-endocrine tissue with Bone Morphogenetic Protein 7 (BMP-7) leads to the formation of glucose-responsive β-like cells. Here, we show that BMP-7 acts on extrainsular cells expressing PDX1 and the BMP receptor activin-like kinase 3 (ALK3/BMPR1A). In vitro lineage tracing indicates that ALK3+ cell populations are multipotent. PDX1+/ALK3+ cells are absent from islets but prominently represented in the major pancreatic ducts and pancreatic duct glands. We identified the purinergic receptor P2Y1 (P2RY1) as a surrogate surface marker for PDX1. Sorted P2RY1+/ALK3bright+ cells form BMP-7-expandable colonies characterized by NKX6.1 and PDX1 expression. Unlike the negative fraction controls, these colonies can be differentiated into multiple pancreatic lineages upon BMP-7 withdrawal. RNA-seq further corroborates the progenitor-like nature of P2RY1+/ALK3bright+ cells and their multilineage differentiation potential. Our studies confirm the existence of progenitor cells in the adult human pancreas and suggest a specific anatomical location within the ductal and glandular networks. : Qadir et al. describe and characterize a population of multipotent, BMP-7-responsive progenitor-like cells within the human exocrine pancreas. These cells are characterized by the expression of PDX1 and ALK3, a canonical BMP receptor. Their findings shed new light on potential regenerative pathways in the human pancreas. Keywords: human pancreatic progenitor cells, ALK3, PDX1, islet regeneration, beta cell regeneration

Polyphenols have been widely studied for their antiviral effect against respiratory virus infections. Among these, resveratrol (RV) has been demonstrated to inhibit influenza virus replication and more recently, it has been tested together with pterostilbene against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In the present work, we evaluated the antiviral activity of polydatin, an RV precursor, and a mixture of polyphenols and other micronutrients, named A5+, against influenza virus and SARS-CoV-2 infections. To this end, we infected Vero E6 cells and analyzed the replication of both respiratory viruses in terms of viral proteins synthesis and viral titration. We demonstrated that A5+ showed a higher efficacy in inhibiting both influenza virus and SARS-CoV-2 infections compared to polydatin treatment alone. Indeed, post infection treatment significantly decreased viral proteins expression and viral release, probably by interfering with any step of virus replicative cycle. Intriguingly, A5+ treatment strongly reduced IL-6 cytokine production in influenza virus-infected cells, suggesting its potential anti-inflammatory properties during the infection. Overall, these results demonstrate the synergic and innovative antiviral efficacy of A5+ mixture, although further studies are needed to clarify the mechanisms underlying its inhibitory effect.

Ischemic stroke (IS) is still among the leading causes of death and disability worldwide. The pathogenic mechanisms beyond its development are several and are complex and this is the main reason why a functional therapy is still missed. The beneficial effects of natural compounds against cardiovascular diseases and IS have been investigated for a long time. In this article, we reviewed the association between the most studied polyphenols and stroke protection in terms of prevention, effect on acute phase, and rehabilitation. We described experimental and epidemiological studies reporting the role of flavonols, phenolic acid, and stilbens on ischemic mechanisms leading to stroke. We analyzed the principal animal models used to evaluate the impact of these micronutrients to cerebral blood flow and to molecular pathways involved in oxidative stress and inflammation modulation, such as sirtuins. We reported the most significant clinical trials demonstrated as the persistent use of polyphenols is clinically relevant in terms of the reduction of vascular risk factors for IS, such as Atrial Fibrillation. Interestingly, different kinds of polyphenols provide brain protection by activating different pathways and mechanisms, like inducing antithrombotic effect, such as Honokiol. For this reason, we discussed an appropriate integrative use of them as a possible therapeutic alternative against stroke.

The Food and Drug Administration (FDA) has been regulating human islets for allotransplantation as a biologic drug in the US. Consequently, the requirement of a biological license application (BLA) approval before clinical use of islet transplantation as a standard of care procedure has stalled the development of the field for the last 20 years. Herein, we provide our commentary to the multiple FDA’s position papers and guidance for industry arguing that BLA requirement has been inappropriately applied to allogeneic islets, which was delivered to the FDA Cellular, Tissue and Gene Therapies Advisory Committee on 15 April 2021. We provided evidence that BLA requirement and drug related regulations are inadequate in reassuring islet product quality and potency as well as patient safety and clinical outcomes. As leaders in the field of transplantation and endocrinology under the “Islets for US Collaborative” designation, we examined the current regulatory status of islet transplantation in the US and identified several anticipated negative consequences of the BLA approval. In our commentary we also offer an alternative pathway for islet transplantation under the regulatory framework for organ transplantation, which would address deficiencies of in current system.

Abstract Type 1 diabetes mellitus (T1DM) results from an autoimmune attack against the insulin-producing ß cells which leads to chronic hyperglycemia. Exosomes are lipid vesicles derived from cellular multivesicular bodies that are enriched in specific miRNAs, potentially providing a disease-specific diagnostic signature. To assess the value of exosome miRNAs as biomarkers for T1DM, miRNA expression in plasma-derived exosomes was measured. Nanoparticle tracking analysis and transmission electron microscopy confirmed the presence of plasma-derived exosomes (EXOs) isolated by differential centrifugation. Total RNA extracted from plasma-derived EXOs of 12 T1DM and 12 control subjects was hybridized onto Nanostring human v2 miRNA microarray array and expression data were analyzed on nSolver analysis software. We found 7 different miRNAs (1 up-regulated and 6 down-regulated), that were differentially expressed in T1DM. The selected candidate miRNAs were validated by qRT-PCR analysis of cohorts of 24 T1DM and 24 control subjects. Most of the deregulated miRNAs are involved in progression of T1DM. These findings highlight the potential of EXOs miRNA profiling in the diagnosis as well as new insights into the molecular mechanisms involved in T1DM.

COVID-19 is without any doubt the worst pandemic we have faced since the H1N1 virus outbreak. Even if vaccination against SARS-CoV-2 infection is becoming increasingly available, a more feasible approach for COVID-19 prevention and therapy is still needed. Evidence of a pathological link between metabolic diseases and severe forms of COVID-19 has stimulated critical reflection and new considerations. In particular, an abnormal immune response observed in certain patients with SARS-CoV-2 infection suggested possible common predisposing risk factors with autoimmune diseases such as Type 1 Diabetes (T1D). Correct supplementation with dietary factors may be key to preventing and counteracting both the underlying metabolic impairment and the complications of COVID-19. A set of agents may inhibit the cytokine storm and hypercoagulability that characterize severe COVID-19 infection: vitamin D3, omega-3 polyunsaturated fatty acids, polyphenols like pterostilbene, polydatin and honokiol, which can activate anti-inflammatory and antioxidant sirtuins pathways, quercetin, vitamin C, zinc, melatonin, lactoferrin and glutathione. These agents could be highly beneficial for subjects who have altered immune responses. In this review, we discuss the antiviral and metabolic effects of these dietary factors and propose their combination for potential applications in the prevention and treatment of COVID-19. Rigorous studies will be fundamental for validating preventive and therapeutic protocols that could be of assistance to mitigate disease progression following SARS-CoV-2 infection.

Oxidative stress plays critical roles in the pathogenesis of diabetes. This study tested the hypothesis that by protecting β-cells against oxidative stress and inflammation, an Nrf2 activator, dimethyl fumarate (DMF), may prevent or delay the onset of type 1 diabetes in non-obese diabetic (NOD) mice. Firstly, islet isolation was conducted to confirm the antioxidative effects of DMF oral administration on islet cells. Secondly, in a spontaneous diabetes model, DMF (25 mg/kg) was fed to mice once daily starting at the age of 8 weeks up to the age of 22 weeks. In a cyclophosphamide-induced accelerated diabetes model, DMF (25 mg/kg) was fed to mice twice daily for 2 weeks. In the islet isolation study, DMF administration improved the isolation yield, attenuated oxidative stress and enhanced GCLC and NQO1 expression in the islets. In the spontaneous model, DMF significantly reduced the onset of diabetes compared to the control group (25% vs. 54.2%). In the accelerated model, DMF reduced the onset of diabetes from 58.3% to 16.7%. The insulitis score in the islets of the DMF treatment group (1.6 ± 0.32) was significantly lower than in the control group (3.47 ± 0.21). The serum IL-1α, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-9, IL-12p70, IFN-γ, TNF-α, MCP-1 and CXCL16 levels in the DMF-treated group were lower than in the control group. In conclusion, DMF may protect islet cells and reduce the incidence of autoimmune diabetes in NOD mice by attenuating insulitis and proinflammatory cytokine production.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The inflammasome is a key contributor to the inflammatory innate immune response after stroke. We have previously shown that inflammasome proteins are released in extracellular vesicles (EV) after brain and spinal cord injury. In addition, we have shown that inflammasome proteins offer great promise as biomarkers of central nervous system (CNS) injury following brain trauma. In the present study, we used a Simple Plex Assay (Protein Simple), a novel multi-analyte automated microfluidic immunoassay platform, to analyze serum and serum-derived EV samples from stroke patients and control subjects for inflammasome protein levels of caspase-1, apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC), Interleukins (IL)-1β, and (IL)-18. Receiver operator characteristic (ROC) curves with associated confidence intervals obtained from the analysis of serum samples revealed that the area under the curve (AUC) for ASC was 0.99 with a confidence interval between 0.9914 and 1.004, whereas the AUC for caspase-1, IL-1β, and IL-18 were 0.75, 0.61, and 0.67, respectively. Thus, these data indicate that ASC is a potential biomarker of stroke and highlight the role of the inflammasome in the inflammatory response after brain ischemia.

BACKGROUND:Type 1 and Type 2 diabetes mellitus (T1DM and T2DM) are caused by beta(β)-cell loss and functional impairment. Identification of mechanisms of β-cell death and therapeutic interventions to enhance β-cell survival are essential for prevention and treatment of diabetes. Oxidative stress is a common feature of both T1DM and T2DM; elevated biomarkers of oxidative stress are detected in blood, urine and tissues including pancreas of patients with DM. Islet transplantation is a promising treatment for diabetes. However, exposure to stress (chemical and mechanical) and ischemia-reperfusion during isolation and transplantation causes islet loss by generation of reactive oxygen species (ROS). Human intracellular antioxidant enzymes and related molecules are essential defenses against ROS. Antioxidant enzyme levels including superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPX) have been shown to be low in islet cells. However, little is known about the expression and function of antioxidant enzymes within islet cell subsets. We evaluated the expression of the key antioxidant enzymes in β- and alpha(α)-cell and accessed effects of oxidative stress, islet isolation and transplantation on β/α-cell ratio and viability in human islets. METHODS:Human pancreata from T1DM, T2DM and non-diabetic deceased donors were obtained and analyzed by confocal microscopy. Isolated islets were (I) transplanted in the renal sub-capsular space of streptozotocin-induced diabetic nude mice (in vivo bioassay), or (II) exposed to oxidative (H2O2) and nitrosative (NO donor) stress for 24 hrs in vitro. The ratio, % viability and death of β- and α-cells, and DNA damage (8OHdG) were measured. RESULTS AND CONCLUSIONS:Catalase and GPX expression was much lower in β- than α-cells. The β/α-cell ratio fells significantly following islet isolation and transplantation. Exposure to oxidative stress caused a significantly lower survival and viability, with higher DNA damage in β- than α-cells. These findings identified the weakness of β-cell antioxidant capacity as a main cause of vulnerability to oxidative stress. Potential strategies to enhance β-cell antioxidant capacity might be effective in prevention/treatment of diabetes.

A variety of tissue engineering techniques utilizing different cells and biomaterials are currently being explored to construct urinary bladder walls de novo, but so far no approach is clearly superior. The aim of this study was to determine whether mesenchymal stem cells (MSCs) isolated from different sources, (bone marrow [BM-MSCs] and adipose tissue [ADSCs]), differ in their potential to regenerate smooth muscles in tissue-engineered urinary bladders and to determine an optimal number of MSCs for urinary bladder smooth muscle regeneration. Forty-eight rats underwent hemicystectomy and bladder augmentation with approximately 0.8 cm 2 graft. In the first and second groups, urinary bladders were reconstructed with small intestinal submucosa (SIS) seeded with 10 × 10 6 or 4 × 10 6 ADSCs/cm 2 , respectively. In the third and fourth groups, urinary bladders were augmented with SIS seeded with 10 × 10 6 or 4 × 10 6 BM-MSCs/cm 2 , respectively. In the fifth group, urinary bladders were augmented with SIS without cells. The sixth group (control) was left intact. Smooth muscle regeneration was evaluated by real-time polymerase chain reaction (RT-PCR) and histological examinations. Histologically, there were no significant differences between urinary bladders augmented with ADSCs and BM-MSCs, but there was a marked increase in smooth muscle formation in bladders augmented with grafts seeded with MSCs in higher density (10 × 10 6 /cm 2 ) compared to lower density (4 × 10 6 /cm 2 ). Molecular analysis revealed that bladders reconstructed with ADSC-seeded grafts expressed higher levels of smooth muscle myosin heavy chain, caldesmon, and vinculin. Bladders augmented with unseeded SIS were fibrotic and devoid of smooth muscles. ADSCs and BM-MSCs have comparable smooth muscle regenerative potential, but the number of MSCs used for graft preparation significantly affects the smooth muscle content in tissue-engineered urinary bladders.

Vitamin D and omega 3 fatty acid (ω-3) co-supplementation potentially improves type 1 diabetes (T1D) by attenuating autoimmunity and counteracting inflammation. This cohort study, preliminary to a randomized control trial (RCT), is aimed at evaluating, in a series of T1D children assuming Mediterranean diet and an intake of cholecalciferol of 1000U/day from T1D onset, if ω-3 co-supplementation preserves the residual endogen insulin secretion (REIS). Therefore, the cohort of 22 “new onsets” of 2017 received ω-3 (eicosapentenoic acid (EPA) plus docosahexaenoic acid (DHA), 60 mg/kg/day), and were compared retrospectively vs. the 37 “previous onsets” without ω-3 supplementation. Glicosilated hemoglobin (HbA1c%), the daily insulin demand (IU/Kg/day) and IDAA1c, a composite index (calculated as IU/Kg/day × 4 + HbA1c%), as surrogates of REIS, were evaluated at recruitment (T0) and 12 months later (T12). In the ω-3 supplemented group, dietary intakes were evaluated at T0 and T12. As an outcome, a decreased insulin demand (p < 0.01), particularly as pre-meal boluses (p < 0.01), and IDAA1c (p < 0.01), were found in the ω-3 supplemented group, while HbA1c% was not significantly different. Diet analysis in the ω-3 supplemented group, at T12 vs. T0, highlighted that the intake of arachidonic acid (AA) decreased (p < 0.01). At T0, the AA intake was inversely correlated with HbA1c% (p < 0.05; r;. 0.411). In conclusion, the results suggest that vitamin D plus ω-3 co-supplementation as well as AA reduction in the Mediterranean diet display benefits for T1D children at onset and deserve further investigation.

Type 1 diabetes (T1D) is a chronic autoimmune disease leading to immune-mediated destruction of pancreatic beta cells, resulting in the need for insulin therapy. The incidence of T1D is increasing worldwide, thus prompting researchers to investigate novel immunomodulatory strategies to halt autoimmunity and modify disease progression. T1D is considered as a multifactorial disease, in which genetic predisposition and environmental factors interact to promote the triggering of autoimmune responses against beta cells. Over the last decades, it has become clear that vitamin D exerts anti-inflammatory and immunomodulatory effects, apart from its well-established role in the regulation of calcium homeostasis and bone metabolism. Importantly, the global incidence of vitamin D deficiency is also dramatically increasing and epidemiologic evidence suggests an involvement of vitamin D deficiency in T1D pathogenesis. Polymorphisms in genes critical for vitamin D metabolism have also been shown to modulate the risk of T1D. Moreover, several studies have investigated the role of vitamin D (in different doses and formulations) as a potential adjuvant immunomodulatory therapy in patients with new-onset and established T1D. This review aims to present the current knowledge on the immunomodulatory effects of vitamin D and summarize the clinical interventional studies investigating its use for prevention or treatment of T1D.

Sirtuins (SIRTs) are seven nicotinamide adenine dinucleotide (NAD+)-dependent protein deacetylases enzymes (SIRT1−7) that play an important role in maintaining cellular homeostasis. Among those, the most studied are SIRT1 and SIRT3, a nuclear SIRT and a mitochondrial SIRT, respectively, which significantly impact with an increase in mammals’ lifespan by modulating metabolic cellular processes. Particularly, when activated, both SIRT1 and 3 enhance pancreatic β-cells’ insulin release and reduce inflammation and oxidative stress pancreatic damage, maintaining then glucose homeostasis. Therefore, SIRT1 and 3 activators have been proposed to prevent and counteract metabolic age-related diseases, such as type 2 diabetes mellitus (T2DM). Physical activity (PA) has a well-established beneficial effect on phenotypes of aging like β-cell dysfunction and diabetes mellitus. Recent experimental and clinical evidence reports that PA increases the expression levels of both SIRT1 and 3, suggesting that PA may exert its healthy contribute even by activating SIRTs. Therefore, in the present article, we discuss the role of SIRT1, SIRT3, and PA on β-cell function and on diabetes. We also discuss the possible interaction between PA and activation of SIRTs as a possible therapeutic strategy to maintain glucose hemostasis and to prevent T2DM and its complications, especially in the elderly population.

For isolated pancreatic islet cell preparations, it is important to be able to reliably assess their mass and quality, and for clinical applications, it is part of the regulatory requirement. Accurate assessment, however, is difficult because islets are spheroid-like cell aggregates of different sizes (

Selection of an optimal donor pancreas is the first key task for successful islet isolation. We conducted a retrospective multicenter study in 11 centers in North America to develop an islet donor scoring system using donor variables. The data set consisting of 1,056 deceased donors was used for development of a scoring system to predict islet isolation success (defined as postpurification islet yield >400,000 islet equivalents). With the aid of univariate logistic regression analyses, we developed the North American Islet Donor Score (NAIDS) ranging from 0 to 100 points. The c index in the development cohort was 0.73 (95% confidence interval 0.70–0.76). The success rate increased proportionally as the NAIDS increased, from 6.8% success in the NAIDS < 50 points to 53.7% success in the NAIDS ≥ 80 points. We further validated the NAIDS using a separate set of data consisting of 179 islet isolations. A comparable outcome of the NAIDS was observed in the validation cohort. The NAIDS may be a useful tool for donor pancreas selection in clinical practice. Apart from its utility in clinical decision making, the NAIDS may also be used in a research setting as a standardized measurement of pancreas quality.

BACKGROUND:T-cell depleting strategies have become an integral part of immunosuppressive regimens in organ transplantation. Alemtuzumab is a humanized monoclonal antibody against CD52, a cell-surface antigen on several immune cells. It has been suggested that lymphocyte depletion increases the risk of serious infections. However, this has not been observed with short-term alemtuzumab treatment in an organ transplant setting. For induction therapy using alemtuzumab following liver transplantation, we found that T- and B-cell numbers declined rapidly after alemtuzumab therapy; however, the natural killer (NK) cell number was sustained. NK cells are important effectors of innate immunity. Since the effects of alemtuzumab on NK cell functions, especially those of liver NK cells, are unknown, this study aimed to investigate this in detail. METHODS:To assess the effect of alemtuzumab on NK cells, samples were obtained from 7 organ donors and examined by flow cytometry using Annexin V and propidium iodide. Phenotypical and functional differences within subsets of NK cells with different levels of CD52 expression were determined by flow cytometry and in vitro cytotoxicity assays. RESULTS:CD52 expression on NK cells was lower than that on other lymphocyte subsets. The liver contained a large number of CD52- NK cells compared with the peripheral blood. In vitro treatment of liver-derived NK cells with alemtuzumab did not result in cell death. In contrast, co-incubation with alemtuzumab induced cell death in peripheral blood mononuclear cells and non-NK cells in the liver. Furthermore, CD52- liver NK cells were more cytotoxic and produced more IFN-γ than CD52+ NK cells after cytokine activation. CONCLUSION:The liver contains a large number of CD52- NK cells. These cells are refractory to alemtuzumab and have robust activity. These findings indicate that CD52- NK cells persist and could protect against infection after alemtuzumab-based lymphocyte depletion.

BACKGROUNDAllogeneic human islet transplantation is an effective therapy for the treatment of patients with Type 1 Diabetes (T1D). The low number of islet transplants performed worldwide and the different transplantation protocols used limit the identification of the most effective therapeutic options to improve the efficacy of this approach.METHODSWe present a retrospective analysis on the data collected from 44 patients with T1D who underwent islet transplantation at our institute between 2000 and 2007. Several variables were included: recipient demographics and immunological characteristics, donor and transplant characteristics, induction protocols, and additional medical treatment received. Immunosuppression was induced with anti-CD25 (Daclizumab), alone or in association with anti-tumor necrosis factor alpha (TNF-α) treatments (Etanercept or Infliximab), or with anti-CD52 (Alemtuzumab) in association with anti-TNF-α treatments (Etanercept or Infliximab). Subsets of patients were treated with Filgrastim for moderate/severe neutropenia and/or Exenatide for post prandial hyperglycemia.RESULTSThe analysis performed indicates a negative association between graft survival (c-peptide level ≥ 0.3 ng/ml) and islet infusion volume, with the caveat that, the progressive reduction of infusion volumes over the years has been paralleled by improved immunosuppressive protocols. A positive association is instead suggested between graft survival and administration of Exenatide and Filgrastim, alone or in combination.CONCLUSIONThis retrospective analysis may be of assistance to further improve long-term outcomes of protocols for transplant of islets and other organs.

Pancreatic islets are known to contain low level of antioxidants that renders them vulnerable to oxidative stress. Nrf2 is the master regulator of numerous genes, encoding antioxidant, detoxifying, and cytoprotective molecules. Activation of Nrf2 pathway induces up-regulation of numerous genes encoding antioxidant and phase II detoxifying enzymes and related proteins. However, little is known regarding the role of this pathway in human islet cells. The aim was to investigate the effect of Nrf2 activator (dh404, CDDO-9,11-dihydro-trifluoroethyl amide) on human islet cells.Human islets were obtained from cadaveric donors. After dh404 treatment, Nrf2 translocation, mRNA expression, and protein abundance of its key target gene products were examined. The proportion of dh404-treated or non-treated viable islet beta cells was analyzed using flowcytemetry. The cytoprotective effects against oxidative stress and production of inflammatory mediators, and in vivo islet function after transplantation were determined.Nrf2 nuclear translocation was confirmed by con-focal microscope within 2 hours after treatment, which was associated with a dose-dependent increase in mRNA expression of anti-oxidants, including NQO1, HO-1, and GCLC. Enhanced HO-1 expression in dh404 treated islets was confirmed by Western Blot assay. Islet function after transplantation (2000 IEQ/mouse) to diabetic nude mice was not affected with or without dh404 treatment. After induction of oxidative stress with hydrogen peroxide (200 μM) the proportion of dh404-treated viable islet cells was significantly higher in the dh404-treated than untreated islets (74% vs.57%; P

The nonobese diabetic (NOD) mouse represents a well-established experimental model analogous to human type 1 diabetes mellitus (T1D) as it is characterized by progressive autoimmune destruction of pancreatic β-cells. Experiments were designed to investigate the impact of moderate-intensity training on T1D immunomodulation and inflammation. Under a chronic exercise regime, NOD mice were trained on a treadmill for 12 weeks (12 m/min for 30 min, 5 d/wk) while age-matched, control animals were left untrained. Prior to and upon completion of the training period, fed plasma glucose and immunological soluble factors were monitored. Both groups showed deteriorated glycemic profiles throughout the study although trained mice tended to be more compensated than controls after 10 weeks of training. An exercise-induced weight loss was detected in the trained mice with respect to the controls from week 6. After 12 weeks, IL-6 and MIP-1β were decreased in the trained animals compared to their baseline values and versus controls, although not significantly. Morphometric analysis of pancreata revealed the presence of larger infiltrates along with decreased α-cells areas in the control mice compared to trained mice. Exercise may exert positive immunomodulation of systemic functions with respect to both T1D and inflammation, but only in a stringent therapeutic window.

The discovery of insulin changed the course of history in the treatment of diabetes. However, despite tremendous progress in insulin formulations and treatment strategies, insulin treatment still cannot fully prevent chronic complications and intensive insulin treatment to improve metabolic control, has often paralleled an increased risk of severe hypoglycemia. A cure for Type 1 diabetes should include: ? Restoration of self tolerance, to prevent recurrence of autoimmunity ? Restoration of physiologic metabolism by replacement of the biologic function (insulin producing cells) that was partially of completely impaired as a result of the disease process. ? Prevention of destruction of the new insulin producing cells by the recipient immune system in the absence of any treatment that could introduce an additional risk to the patient, which could be comparable to, or higher than the risk imposed by disease progression under exogenous insulin treatment. Pancreatic islets could be considered an ideal and more physiologic alternative to insulin, as they can restore metabolic control after transplantation, preventing the development of chronic complications. In fact, islets are capable of perfectly timed insulin release and can keep glucose levels in the normal range, functioning for an entire lifetime, if they are not destroyed by the recipient's immune system. Significant progress has been recently reported in the translational research approach towards the development of a cure for Type 1 diabetes. There is now strong evidence for the technical feasibility of human islet isolation and purification procedures. Proof of function of isolated human islets has been clearly established both in animal models and in pilot clinical trials of human islet allotransplantation in patients with surgical and Type 1 diabetes. Additional research in now needed to improve the current clinical results in terms of long term prevention of rejection and recurrence of autoimmunity, the development of safe, non diabetogenic immunomodulation strategies and ultimately the achievement of donor specific immune tolerance. If success will be achieved in these areas, then we will face the critical challenge of identifying sufficient and suitable sources of insulin producing tissue to treat the increasing number of patients who could benefit from this form of therapy and which would not be limited to Type 1 diabetes. That is why the work on xenogeneic islets, embryonic and adult stem cells, islet regeneration and proliferation, as well as engineering of insulin producing cells must be continued, to identify the most ideal source of insulin producing tissue that could be utilized on a large scale once the impediments and limitations of immunosuppression will be resolved.

We evaluated whether 1,5-anhydroglucitol (1,5-AG) (GlycoMark ® ), a test for measuring postprandial glucose and glucose variability, could be a tool for assessing short-term glycemic control in islet cell transplant (ICT) subjects. Data of 21 subjects, with type 1 DM and allogenic islet transplantation, who had concomitant fructosamine, HbA1c, 1,5-AG ( n = 85 samples), and capillary glucose self-monitoring measurements ( n = 2,979) were analyzed retrospectively at different time points after ICT. A significant negative association was observed between 1,5-AG and HbA1c ( p = 0.02), but not with fructosamine. When HbA1c was divided in quartiles as 6.2, a decrease of an estimated 0.70 ± 0.30 μg/ml in 1,5-AG was associated with each quartile of increase in HbA1c ( p < 0.0001). There was a significant decline of 1.64 ± 0.3mg/dl in postprandial glucose values for each 1 unit increase in 1,5-AG ( p < 0.0001). For those with HbA1c ≥ 6.0% when 1,5-AG was ≥8.15 μg/ml, the mean estimated glucose level was 103.71 ± 3.66 mg/dl, whereas it was 132.12 ± 3.71 mg/ dl when 1,5-AG was

Our objective is to evaluate if there is an association between liver fat accumulation after islet transplantation (ITx) and graft survival. A cohort study was conducted in 34 subjects with type 1 diabetes postallogeneic ITx. Liver fat content was evaluated by magnetic resonance imaging (MRI) (change in liver signal intensity on in-phase and opposed-phase images). Kaplan–Meier curves and Cox regression analysis were performed with islet dysfunction duration as the dependent variable and fat liver content as an independent one. Values of p < 0.05 were significant (SSPS ® 18.0 and MedCalc ® 12.5). Patients' mean age was 40 ± 8 years (diabetes duration: 31 ± 12 years; male: 41%). Islet survival did not differ in patients without (51 months, 95% CI 40–62 months) or with steatosis (48 months, 95% CI 38–58 months; p = 0.55) during islet dysfunction period. Nevertheless, survival curves appear to separate late in the follow-up, and after 40 months steatosis was associated with shorter graft survival ( p log rank = 0.049). This association remained (RR 23.5, 95% CI 1.1–516.0; p = 0.045) after adjustments for possible confounding factors. In this sample of subjects with type 1 diabetes submitted to ITx, steatosis was not associated with islet failure in the whole cohort. However, in subjects with functional islets after 40 months, a shorter graft survival was observed in those with steatosis during the islet dysfunction period, even after adjustments to variables known to be associated with islet failure.

Adipose tissue contains multipotent elements with phenotypic and gene expression profiles similar to human mesenchymal stem cells (hMSCs) and pericytes. The chance of clinical translation of the multilineage potential of these cells is delayed by the poor/negligible cell survival within cryopreserved lipoaspirates, the difficulty of ex vivo expansion, and the complexity of current Good Manufacturing Practice (cGMP) requirements for expanded cells. Hence, availability of a minimally manipulated, autologous, hMSC/pericyte-enriched fat product would have remarkable biomedical and clinical relevance. Here, we present an innovative system, named Lipogems, providing a nonexpanded, ready-to-use fat product. The system uses mild mechanical forces in a completely closed system, avoiding enzymes, additives, and other manipulations. Differently from unprocessed lipoaspirate, the nonexpanded Lipogems product encompasses a remarkably preserved vascular stroma with slit-like capillaries wedged between adipocytes and stromal stalks containing vascular channels with evident lumina. Immunohistochemistry revealed that Lipogems stromal vascular tissue included abundant cells with pericyte/hMSC identity. Flow cytometry analysis of nonexpanded, collagenase-treated Lipogems product showed that it was comprised with a significantly higher percentage of mature pericytes and hMSCs, and lower amount of hematopoietic elements, than enzymatically digested lipoaspirates. Differently from the lipoaspirate, the distinctive traits of freshly isolated Lipogems product were not altered by cryopreservation. Noteworthy, the features of fresh product were retained in the Lipogems product obtained from human cadavers, paving the way to an off-the-shelf strategy for reconstructive procedures and regenerative medicine. When placed in tissue culture medium, the Lipogems product yielded a highly homogeneous adipose tissue-derived hMSC population, exhibiting features of hMSCs isolated from other sources, including the classical commitment to osteogenic, chondrogenic, and adipogenic lineages. Moreover, the transcription of vasculogenic genes in Lipogems-derived adipose tissue hMSCs was enhanced at a significantly greater extent by a mixture of natural provasculogenic molecules, when compared to hMSCs isolated from enzymatically digested lipoaspirates.

Conventional culture vessels are not designed for physiological oxygen (O 2 ) delivery. Both hyperoxia and hypoxia—commonly observed when culturing cells in regular plasticware—have been linked to reduced cellular function and death. Pancreatic islets, used for the clinical treatment of diabetes, are especially sensitive to sub- and supraphysiological O 2 concentrations. A result of current culture standards is that a high percentage of islet preparations are never transplanted because of cell death and loss of function in the 24–48 h postisolation. Here, we describe a new culture system designed to provide quasiphysiological oxygenation to islets in culture. The use of dishes where islets rest atop a perfluorocarbon (PFC)-based membrane, coupled with a careful adjustment of environmental O 2 concentration to target the islet physiological pO 2 range, resulted in dramatic gains in viability and function. These observations underline the importance of approximating culture conditions as closely as possible to those of the native microenvironment, and fill a widely acknowledged gap in our ability to preserve islet functionality in vitro. As stem cell-derived insulin-producing cells are likely to suffer from the same limitations as those observed in real islets, our findings are especially timely in the context of current efforts to define renewable sources for transplantation.

Clinical islet transplantation has demonstrated success in treating type 1 diabetes. A current limitation is the intrahepatic portal vein transplant site, which is prone to mechanical stress and inflammation. Transplantation of pancreatic islets into alternative sites is preferable, but challenging, as it may require a three-dimensional vehicle to confer mechanical protection and to confine islets to a well-defined, retrievable space where islet neovascularization can occur. We have fabricated biostable, macroporous scaffolds from poly(dimethylsiloxane) (PDMS) and investigated islet retention and distribution, metabolic function, and glucose-dependent insulin secretion within these scaffolds. Islets from multiple sources, including rodents, nonhuman primates, and humans, were tested in vitro. We observed high islet retention and distribution within PDMS scaffolds, with retention of small islets (

microRNAs (miRNAs) play an important role in pancreatic development and adult β-cell physiology. Our hypothesis is based on the assumption that each islet cell type has a specific pattern of miRNA expression. We sought to determine the profile of miRNA expression in α-and β-cells, the main components of pancreatic islets, because this analysis may lead to a better understanding of islet gene regulatory pathways. Highly enriched (>98%) subsets of human α-and β-cells were obtained by flow cytometric sorting after intracellular staining with c-peptide and glucagon antibody. The method of sorting based on intracellular staining is possible because miRNAs are stable after fixation. MiRNA expression levels were determined by quantitative high throughput PCR-based miRNA array platform screening. Most of the miRNAs were preferentially expressed in β-cells. From the total of 667 miRNAs screened, the Significant Analysis of Microarray identified 141 miRNAs, of which only 7 were expressed more in α-cells (α-miRNAs) and 134 were expressed more in β-cells (β-miRNAs). Bioinformatic analysis identified potential targets of β-miRNAs analyzing the Beta Cell Gene Atlas, described in the T1Dbase, the web platform, supporting the type 1 diabetes (T1D) community. cMaf, a transcription factor regulating glucagon expression expressed selectively in α-cells (TFα) is targeted by β-miRNAs; miR-200c, miR-125b and miR-182. Min6 cells treated with inhibitors of these miRNAs show an increased expression of cMaf RNA. Conversely, over expression of miR-200c, miR-125b or miR-182 in the mouse alpha cell line αTC6 decreases the level of cMAF mRNA and protein. MiR-200c also inhibits the expression of Zfpm2, a TFα that inhibits the PI3K signaling pathway, at both RNA and protein levels.In conclusion, we identified miRNAs differentially expressed in pancreatic α- and β-cells and their potential transcription factor targets that could add new insights into different aspects of islet biology and pathophysiology.

While regional heterogeneity in islet distribution has been well studied in rodents, less is known about human pancreatic histology. To fill gaps in our understanding, regional differences in the adult human pancreas were quantitatively analyzed including the pathogenesis of type 2 diabetes (T2D). Cadaveric pancreas specimens were collected from the head, body and tail regions of each donor, including subjects with no history of diabetes or pancreatic diseases (n = 23) as well as patients with T2D (n = 12). The study further included individuals from whom islets were isolated (n = 7) to study islet yield and function in a clinical setting of islet transplantation. The whole pancreatic sections were examined using an innovative large-scale image capture and unbiased detailed quantitative analyses of the characteristics of islets from each individual (architecture, size, shape and distribution). Islet distribution/density is similar between the head and body regions, but is >2-fold higher in the tail region. In contrast to rodents, islet cellular composition and architecture were similar throughout the pancreas and there was no difference in glucose-stimulated insulin secretion in islets isolated from different regions of the pancreas. Further studies revealed preferential loss of large islets in the head region in patients with T2D. The present study has demonstrated distinct characteristics of the human pancreas, which should provide a baseline for the future studies integrating existing research in the field and helping to advance bi-directional research between humans and preclinical models.

MicroRNAs regulate gene expression by inhibiting translation or inducing target mRNA degradation. MicroRNAs regulate organ differentiation and embryonic development, including pancreatic specification and islet function. We showed previously that miR-7 is highly expressed in human pancreatic fetal and adult endocrine cells. Here we determined the expression profile of miR-7 in the mouse-developing pancreas by RT-PCR and in situ hybridization. MiR-7 expression was low between embryonic days e10.5 and e11.5, then began to increase at e13.5 through e14.5, and eventually decreased by e18. In situ hybridization and immunostaining analysis showed that miR-7 colocalizes with endocrine marker Isl1, suggesting that miR-7 is expressed preferentially in endocrine cells. Whole-mount in situ hybridization shows miR-7 highly expressed in the embryonic neural tube. To investigate the role of miR-7 in development of the mouse endocrine pancreas, antisense miR-7 morpholinos (MO) were delivered to the embryo at an early developmental stage (e10.5 days) via intrauterine fetal heart injection. Inhibition of miR-7 during early embryonic life results in an overall downregulation of insulin production, decreased β-cell numbers, and glucose intolerance in the postnatal period. This phenomenon is specific for miR-7 and possibly due to a systemic effect on pancreatic development. On the other hand, the in vitro inhibition of miR-7 in explanted pancreatic buds leads to β-cell death and generation of β-cells expressing less insulin than those in MO control. Therefore, in addition to the potential indirect effects on pancreatic differentiation derived from its systemic downregulation, the knockdown of miR-7 appears to have a β-cell-specific effect as well. These findings suggest that modulation of miR-7 expression could be utilized in the development of stem cell therapies to cure diabetes.