We investigated whether islet autoantibody profile,
genotype, and age influenced a 20-year progression to diabetes from first autoantibody positivity (autoAb
) in first-degree relatives of patients ...with type 1 diabetes.
Persistently islet autoAb
siblings and offspring (
= 462) under 40 years of age were followed by the Belgian Diabetes Registry. AutoAbs against insulin (IAA), GAD (GADA), IA-2 antigen (IA-2A), and zinc transporter 8 (ZnT8A) were determined by radiobinding assay.
The 20-year progression rate of multiple-autoAb
relatives (
= 194) was higher than that for single-autoAb
participants (
= 268) (88% vs. 54%;
< 0.001). Relatives positive for IAA and GADA (
= 54) progressed more slowly than double-autoAb
individuals carrying IA-2A and/or ZnT8A (
= 38;
= 0.001). In multiple-autoAb
relatives, Cox regression analysis identified the presence of IA-2A or ZnT8A as the only independent predictors of more rapid progression to diabetes (
< 0.001); in single-autoAb
relatives, it identified younger age (
< 0.001),
genotype (
< 0.001), and IAA (
= 0.028) as independent predictors of seroconversion to multiple positivity for autoAbs. In time-dependent Cox regression, younger age (
= 0.042),
genotype (
= 0.009), and the development of additional autoAbs (
= 0.012) were associated with more rapid progression to diabetes.
In single-autoAb
relatives, the time to multiple-autoAb positivity increases with age and the absence of IAA and
genotype. The majority of multiple-autoAb
individuals progress to diabetes within 20 years; this occurs more rapidly in the presence of IA-2A or ZnT8A, regardless of age,
genotype, and number of autoAbs. These data may help to refine the risk stratification of presymptomatic type 1 diabetes.
Inverse Relationship Between Cytotoxicity of Free Fatty Acids in Pancreatic Islet Cells and Cellular Triglyceride Accumulation
Miriam Cnop ,
Jean Claude Hannaert ,
Anne Hoorens ,
Décio L. Eizirik and
...Daniel G. Pipeleers
Diabetes Research Center, Vrije Universiteit Brussel, Brussels, Belgium
Abstract
Studies in Zucker diabetic fatty rats have led to the concept that chronically elevated free fatty acid (FFA) levels can cause
apoptosis of triglyceride-laden pancreatic β-cells as a result of the formation of ceramides, which induce nitric oxide (NO)-dependent
cell death. This “lipotoxicity” hypothesis could explain development of type 2 diabetes in obesity. The present study examines
whether prolonged exposure to FFA affects survival of isolated normal rat β-cells and whether the outcome is related to the
occurrence of triglyceride accumulation. A dose-dependent cytotoxicity was detected at 5–100 nmol/l of unbound oleate and
palmitate, with necrosis occurring within 48 h and an additional apoptosis during the subsequent 6 days of culture. At equimolar
concentrations, the cytotoxicity of palmitate was higher than that of oleate but lower than that of its nonmetabolized analog
bromopalmitate. FFA cytotoxicity was not suppressed by etomoxir (an inhibitor of mitochondrial carnitine palmitoyltransferase
I) or by antioxidants; it was not associated with inducible NO synthase expression or NO formation. An inverse correlation
was observed between the percentage of dead β-cells on day 8 and their cellular triglyceride content on day 2. For equimolar
concentrations of the tested FFA, oleate caused the lowest β-cell toxicity and the highest cytoplasmic triglyceride accumulation.
On the other hand, oleate exerted the highest toxicity in islet non–β-cells, where no FFA-induced triglyceride accumulation
was detected. In conditions without triglyceride accumulation, the lower FFA concentrations caused primarily apoptosis, both
in islet β-cells and non–β-cells. It is concluded that FFAs can cause death of normal rat islet cells through an NO-independent
mechanism. The ability of normal β-cells to form and accumulate cytoplasmic triglycerides might serve as a cytoprotective
mechanism against FFA-induced apoptosis by preventing a cellular rise in toxic free fatty acyl moieties. It is conceivable
that this potential is lost or insufficient in cells with a prolonged triglyceride accumulation as may occur in vivo.
Footnotes
Address correspondence and reprint requests to Daniel Pipeleers, Diabetes Research Center, Vrije Universiteit Brussel, Laarbeeklaan
103, 1090 Brussels, Belgium. E-mail: daniel.pipeleers{at}vub.ac.be .
Received for publication 5 October 2000 and accepted in revised form 1 May 2001.
FFA, free fatty acid; IL, interleukin; iNOS, inducible nitric oxide synthase; NO, nitric oxide; PCR, polymerase chain reaction;
RT, reverse transcriptase.
Human stem cells represent a potential source for implants that replace the depleted functional beta cell mass (FBM) in diabetes patients. Human embryonic stem cell-derived pancreatic endoderm ...(hES-PE) can generate implants with glucose-responsive beta cells capable of reducing hyperglycemia in mice. This study with device-encapsulated hES-PE (4 × 106 cells/mouse) determines the biologic characteristics at which implants establish metabolic control during a 50-week follow-up. A metabolically adequate FBM was achieved by (1) formation of a sufficient beta cell number (>0.3 × 106/mouse) at >50% endocrine purity and (2) their maturation to a functional state comparable with human pancreatic beta cells, as judged by their secretory responses during perifusion, their content in typical secretory vesicles, and their nuclear NKX6.1-PDX1-MAFA co-expression. Assessment of FBM in implants and its correlation with in vivo metabolic markers will guide clinical translation of stem cell-derived grafts in diabetes.
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•Human stem cell-derived pancreatic precursors generate functional beta cell mass•Cellular markers identify metabolically adequate human stem cell-generated implants•Significance of determining beta cell number and maturation in implants•Functional implants differ in endocrine composition from endocrine pancreas
In this article, Pipeleers and colleagues demonstrate that subcutaneous implants of device-encapsulated human stem cell-derived pancreatic endoderm can generate a functional beta cell mass that establishes sustained glucose control in mice. They identified their biologic characteristics and correlation with in vivo outcome. Data and methods are expected to guide clinical translation to beta cell replacement therapy in diabetes.
ABSTRACTBeta cell replacement has the potential to restore euglycemia in patients with insulin-dependent diabetes. Although great progress has been made in establishing allogeneic islet ...transplantation from deceased donors as the standard of care for those with the most labile diabetes, it is also clear that the deceased donor organ supply cannot possibly treat all those who could benefit from restoration of a normal beta cell mass, especially if immunosuppression were not required. Against this background, the International Pancreas and Islet Transplant Association in collaboration with the Harvard Stem Cell Institute, the Juvenile Diabetes Research Foundation (JDRF), and the Helmsley Foundation held a 2-day Key Opinion Leaders Meeting in Boston in 2016 to bring together experts in generating and transplanting beta cells derived from stem cells. The following summary highlights current technology, recent significant breakthroughs, unmet needs and roadblocks to stem cell–derived beta cell therapies, with the aim of spurring future preclinical collaborative investigations and progress toward the clinical application of stem cell–derived beta cells.
Subcutaneous implants of device-encapsulated stem cell-derived pancreatic endoderm (PE) can establish a functional beta cell mass (FBM) with metabolic control in immune-compromised mice. In a study ...with human-induced pluripotent stem cell-PE, this outcome was favored by a preformed pouch which allowed lesion-free insertion of devices in a pre-vascularized site. This was not reproduced in nude rats, known to exhibit a higher innate reactivity than mice and therefore relevant as preclinical model: a dense fibrotic capsule formed around subcutis (SC) implants with virtually no FBM formation. Placement in omentum (OM) of nude rats provided a less fibrous, better vascularized environment than SC. It resulted in less donor cell loss (56% recovery at post-transplant-PT week 3 versus 16% in SC) allowing FBM-formation. At PT week 30, 6/13 OM-recipients exhibited glucose-induced plasma hu-C-peptide to 0.1-0.4 ng/ml, versus 0/8 in SC-recipients. These levels are more than 10-fold lower than in a state of metabolic control. This shortcoming is not caused by inadequate glucose responsiveness of the beta cells but by their insufficient number. The size of the formed beta cell mass (0.4
0.2 µl) was lower than that reported in mice receiving the same cell product subcutaneously; the difference is attributed to a lower expansion of pancreatic progenitor cells and to their lower degree of differentiation to beta cells. This study in the nude rat model demonstrates that OM provides a better environment for formation of beta cells in device-encapsulated PE-implants than SC. It also identified targets for increasing their dose-efficacy.
In recent-onset type 1 diabetes, clamp-derived C-peptide predicts good response to anti-CD3. Elevated proinsulin and proinsulin/C-peptide ratio (PI/CP) suggest increased metabolic/inflammatory beta ...cell burden. We reanalyzed trial data to compare the ability of baseline acutely glucose-stimulated proinsulin, C-peptide and PI/CP to predict functional outcome.
Eighty recent-onset type 1 diabetes patients participated in the placebo-controlled otelixizumab (GSK; NCT00627146) trial. Hyperglycemic clamps were performed at baseline, 6, 12 and 18 months, involving 3 h of induced euglycemia, followed by acutely raising and maintaining glycemia to ≥ 10 mmol/l for 140 min. Plasma proinsulin, C-peptide and PI/CP were determined after acute (minute 0 at 10 mmol/l; PI
, CP
, PI/CP
) and sustained glucose stimulation (AUC between minutes 60-140). Outcome was assessed as change in AUC
C-peptide from baseline.
In multiple linear regression, higher baseline (≥median P50) PI
independently predicted preservation of beta cell function in response to anti-CD3 and interacted significantly with IAA. During follow-up, anti-CD3 tempered a further increase in PI/CP
, but not in PI
. CP
outperformed PI
and PI/CP
for post-treatment monitoring.
In recent-onset type 1 diabetes, elevated acutely glucose-stimulated proinsulin may complement or replace acutely or sustainedly stimulated C-peptide release for identifying good responders to anti-CD3, but not as outcome measure.
Alginate (Alg)-encapsulated porcine islet cell grafts are developed to overcome limitations of human islet transplantation. They can generate functional implants in animals when prepared from fetal, ...perinatal, and adult pancreases. Implants have not yet been examined for efficacy to establish sustained, metabolically adequate functional β-cell mass (FBM) in comparison with human islet cells. This study in immune-compromised mice demonstrates that subcutaneous implants of Alg-encapsulated porcine prenatal islet cells with 4 × 10
β-cells form, over 10 weeks, a FBM that results in glucose-induced plasma C-peptide >2 ng/mL and metabolic control over the following 10 weeks, with higher efficiency than nonencapsulated, while failing in peritoneum. This intracapsular FBM formation involves β-cell replication, increasing number fourfold, and maturation toward human adult β-cells. Subcutaneous Alg-encapsulated human islet cells with similar β-cell number establish implants with plasma C-peptide >2 ng/mL for the first 10 weeks, with nonencapsulated cells failing; their β-cells do not replicate but progressively die (>70%), explaining C-peptide decline and insufficient metabolic control. An Alg matrix thus helps establish β-cell functions in subcutis. It allows formation of sustained metabolically adequate FBM by immature porcine β-cells with proliferative activity but not by human adult islet cells. These findings define conditions for evaluating its immune-protecting properties.
Nutrient homeostasis is known to be regulated by pancreatic islet tissue. The function of islet beta-cells is controlled by a glucose sensor that operates at physiological glucose concentrations and ...acts in synergy with signals that integrate messages originating from hypothalamic neurons and endocrine cells in gut and pancreas. Evidence exists that the extrapancreatic cells producing and secreting these (neuro)endocrine signals also exhibit a glucose sensor and an ability to integrate nutrient and (neuro)hormonal messages. Similarities in these cellular and molecular pathways provide a basis for a network of coordinated functions between distant cell groups, which is necessary for an appropriate control of nutrient homeostasis. The glucose sensor seems to be a fundamental component of these control mechanisms. Its molecular characterization is most advanced in pancreatic beta-cells, with important roles for glucokinase and mitochondrial oxidative fluxes in the regulation of ATP-sensitive K+ channels. Other glucose-sensitive cells in the endocrine pancreas, hypothalamus, and gut were found to share some of these molecular characteristics. We propose that similar metabolic signaling pathways influence the function of pancreatic alpha-cells, hypothalamic neurons, and gastrointestinal endocrine and neural cells.
The aim of this study was to establish a gene expression blueprint of pancreatic beta cells conserved from rodents to humans and to evaluate its applicability to assess shifts in the beta cell ...differentiated state. Genome-wide mRNA expression profiles of isolated beta cells were compared to those of a large panel of other tissue and cell types, and transcripts with beta cell-abundant and -selective expression were identified. Iteration of this analysis in mouse, rat and human tissues generated a panel of conserved beta cell biomarkers. This panel was then used to compare isolated versus laser capture microdissected beta cells, monitor adaptations of the beta cell phenotype to fasting, and retrieve possible conserved transcriptional regulators.
A panel of 332 conserved beta cell biomarker genes was found to discriminate both isolated and laser capture microdissected beta cells from all other examined cell types. Of all conserved beta cell-markers, 15% were strongly beta cell-selective and functionally associated to hormone processing, 15% were shared with neuronal cells and associated to regulated synaptic vesicle transport and 30% with immune plus gut mucosal tissues reflecting active protein synthesis. Fasting specifically down-regulated the latter cluster, but preserved the neuronal and strongly beta cell-selective traits, indicating preserved differentiated state. Analysis of consensus binding site enrichment indicated major roles of CREB/ATF and various nutrient- or redox-regulated transcription factors in maintenance of differentiated beta cell phenotype.
Conserved beta cell marker genes contain major gene clusters defined by their beta cell selectivity or by their additional abundance in either neural cells or in immune plus gut mucosal cells. This panel can be used as a template to identify changes in the differentiated state of beta cells.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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