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Sideroblastic anaemias are a diverse group of congenital and acquired bone marrow failure disorders marked by the presence of ring sideroblasts, ineffective erythropoiesis, and systemic iron overload. Congenital Sideroblastic anaemia (CSA) is mainly caused by gene mutations associated with heme synthesis, iron-sulfur [Fe-S] cluster, and mitochondrial protein synthesis pathways. The most prevalent form of CSA is caused by mutations in the erythroid-specific -amino levulinate synthase (ALAS2) gene, which encodes the first enzyme in the heme synthesis pathway in red blood cells. The second most prevalent form of CSA is caused by a mutation in the Solute carrier family 25 member 38 (SLC25A38) gene, which codes for an erythroid-specific protein of the inner mitochondrial membrane. Additionally, 15-20 genes are altogether associated with CSA. In this study, we aim to identify the CSA patients, understand their genetics and establish genotype-phenotype correlation. We have identified fifteen cases of CSA using our targeted NGS (t-NGS) panel. The major clinical findings in our cohort were microcytic anaemia, ring sideroblasts, and dyserythropoiesis in the bone marrow. Currently, two patients are responsive to pyridoxine, while the rest are on blood transfusion support. We have identified ten variants in three different genes of CSA (ALAS2, SLC25A38 & HSPA9). Five patients harbour four hemizygous variants- p.Ala282Ser, p.Arg170Cys, p.Arg204Gln and exon 2 duplication in the ALAS2 gene. In seven patients, we have identified three homozygous mutations - p.Pro190Arg, p.Arg187Gln and p.Arg134Cys in the SLC25A38 gene. These mutations have been predominantly identified in the European population. Three patients revealed three heterozygous variants p. Thr463Ile, D326Tyr, and Arg284Trp in the HSPA9 gene. PyMoL was used to evaluate the functional effects of these variations and understand their effect on the structure of the protein. We believe that by combining a bone marrow examination with genetic sequencing, CSA patients can acquire a definitive diagnosis., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Congenital amegakaryocytic thrombocytopenia (CAMT) is a rare, genetic, autosomal recessive disorder characterized by severe thrombocytopenia, due to inefficient bone marrow megakaryopoiesis eventually leading to aplasia. Majority of the cases are due to homozygous or compound heterozygous mutations in MPL gene encoding for thrombopoietin (THPO) receptor protein. CAMT can be diagnosed at early phase of life, with major complication of transfusion dependency and hematopoietic transplantation as only curative treatment. We have investigated the sequence variations in MPL gene of 7 bone marrow failure (BMF) subjects, who presented with clinically diverse phenotypes, through next generation sequencing (NGS). Plasma THPO levels were estimated using ELISA. Insilico sequence and structure-based analyses were performed to understand the structural and functional implications of mutations, identified through NGS. We studied 7 CAMT subjects suspected of BMF, who presented with severe thrombocytopenia followed by pancytopenia, bleeding manifestation and physical anomalies. The plasma THPO levels were significantly elevated (p<0.05) in all the cases. Molecular analysis by NGS identified 9 genomic mutations in MPL gene. These included 7 non-synonymous substitution, 1 nonsense substitution and 1 in-del mutations, of which 4 are novel mutations. Insilico analysis predicted damaging effects on THPO-R and its reduced affinity for THPO for all the identified mutations. CAMT is a rare disorder with diverse clinical phenotypes and diagnosis is challenging. The elevated plasma THPO levels should be considered for the primary diagnosis and prognosis of the disease. However, molecular analysis of MPL gene is important for the diagnosis and management of the disease through genetic counselling. Though the cytokines, THPO-R agonist are used for the treatment of CAMT, HSCT is the only curative therapy., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The global epidemic of type 2 diabetes has prompted numerous studies and public health efforts to reduce its development. A variety of interventions, including lifestyle modifications and pharmacological agents directed at ameliorating the major risk factors for type 2 diabetes, are of proven efficacy in reducing the development of type 2 diabetes in people with impaired glucose tolerance. While prevention of the hyperglycaemia characteristic of diabetes is arguably an important, clinically relevant outcome, a more compelling outcome with greater clinical significance is the prevention or reduction of the relatively diabetes-specific microvascular and less-specific cardiovascular disease (CVD) complications associated with diabetes. These complications cause the majority of morbidity and excess mortality associated with diabetes. Any reduction in diabetes should, logically, also reduce the occurrence of its long-term complications; however, most diabetes prevention trials have not been of sufficient duration to allow such an evaluation. The limited long-term data, largely from the Da Qing Diabetes Prevention Study (DQDPS) and the Diabetes Prevention Program (DPP) and their respective follow-up studies (DQDPOS and DPPOS), suggest a reduction in microvascular complications and amelioration of CVD risk factors. Only the DQDPOS and Study to Prevent Non-Insulin-Dependent Diabetes Mellitus (STOP-NIDDM) studies have shown a reduction in CVD events and only DQDPOS has demonstrated a decrease in CVD and overall mortality. While these limited data are promising, whether diabetes prevention directly reduces complication-related morbidity and mortality remains unclear. Longer follow-up of prevention studies is needed to supplement the limited current clinical trial data, to help differentiate the effects of diabetes prevention itself from the means used to reduce diabetes development and to understand the balance among benefits, risks and costs of prevention.

The incretins are gut hormones secreted in response to nutrient/carbohydrate ingestion and act on the pancreatic beta cell to amplify glucose-stimulated insulin secretion. Incretin hormone-based treatments for patients with type 2 diabetes represent a major advance in diabetes therapeutics. The ability of the incretin agents (glucagon-like peptide 1 [GLP-1] agonists and dipeptidyl peptidase IV [DPP-4] inhibitors) to improve glycaemia with a low associated risk of hypoglycaemia, together with beneficial/neutral effects on body weight, offers a significant advantage for both patients and treating clinicians. In this edition of 'Then and Now,' it is useful to look back 25 years and reflect upon the developments in this field since Nauck and colleagues published two seminal papers. In 1986 they first documented a reduced incretin effect in patients with type 2 diabetes (Diabetologia 29:46-52), and then in 1993 they demonstrated that, in patients with poorly controlled type 2 diabetes, a single exogenous infusion of an incretin (GLP-1) increased insulin levels in a glucose-dependent manner and normalised fasting hyperglycaemia (Diabetologia 36:741-744). In the ensuing 26 years, progress in the field of incretin hormones has resulted in a greater understanding of the relative roles of GLP-1 and glucose-dependent insulinotropic polypeptide secretion and activity in the pathogenesis of type 2 diabetes and the important recognition that native GLP-1 is quickly degraded by the ubiquitous protease DPP-4. This has led to the development of GLP-1 agonists that are resistant to degradation by DPP-4 and of selective inhibitors of DPP-4 activity as therapeutic agents. GLP-1 agonists (exenatide and liraglutide) and DPP-4 inhibitors (sitagliptin, vildagliptin, saxagliptin and linagliptin) currently represent effective treatment options for patients with type 2 diabetes. Several additional agents are in the pipeline, including longer acting DPP-4-resistant GLP-1 agonists. More exciting, however, is the increasing recognition that the incretin agents have numerous extra-glycaemic effects that could translate into potential cardiovascular and other benefits.

Aims/hypothesis: The aim of the study was to examine the determinants of oral glucose tolerance in 602 persons with impaired glucose tolerance (IGT) who participated in the Actos Now for Prevention of Diabetes (ACT NOW) study., Methods: In addition to the 602 IGT participants, 115 persons with normal glucose tolerance (NGT) and 50 with impaired fasting glucose (IFG) were identified during screening and included in this analysis. Insulin secretion and insulin sensitivity indices were derived from plasma glucose and insulin during an OGTT. The acute insulin response (AIR) (0-10 min) and insulin sensitivity (S(I)) were measured with the frequently sampled intravenous glucose tolerance test (FSIVGTT) in a subset of participants., Results: At baseline, fasting plasma glucose, 2 h postprandial glucose (OGTT) and HbA(1c) were 5.8 +/- 0.02 mmol/l, 10.5 +/- 0.05 mmol/l and 5.5 +/- 0.04%, respectively, in participants with IGT. Participants with IGT were characterised by defects in early (DeltaI (0-30)/DeltaG (0-30) x Matsuda index, where DeltaI is change in insulin in the first 30 min and DeltaG is change in glucose in the first 30 min) and total (DeltaI(0-120)/DeltaG(0-120) x Matsuda index) insulin secretion and in insulin sensitivity (Matsuda index and S(I)). Participants with IGT in whom 2 h plasma glucose was 7.8-8.3 mmol/l had a 63% decrease in the insulin secretion/insulin resistance (disposition) index vs participants with NGT and this defect worsened progressively as 2 h plasma glucose rose to 8.9-9.94 mmol/l (by 73%) and 10.0-11.05 mmol/l (by 80%). The Matsuda insulin sensitivity index was reduced by 40% in IGT compared with NGT (p < 0.005). In multivariate analysis, beta cell function was the primary determinant of glucose AUC during OGTT, explaining 62% of the variance., Conclusion: Our results strongly suggest that progressive beta cell failure is the main determinant of progression of NGT to IGT.

Aims/hypothesis: To determine the independent and potentially synergistic effects of agonists for PPAR gamma and RXR on glucose and lipid metabolism, as well as gene expression, in human skeletal muscle cell cultures., Methods: Fully differentiated myotubes from non-diabetic subjects and subjects with Type II (non-insulin-dependent) diabetes mellitus were chronically (2 days) treated with LG100268 (4 mumol/l), an RXR agonist, or troglitazone (4.6 mumol/l), a PPAR gamma agonist or both, to determine the effects on glucose uptake, activity of glycogen synthase and palmitate oxidation., Results: The combination of both agents increased glucose uptake (60 +/- 9% compared to control subjects) but not either agent alone (16 +/- 9 and 26 +/- 6% for LG100268 and troglitazone, p < 0.01, respectively). The agent LG100268 alone had little effect on the activity of glycogen synthase but the effect of troglitazone increased with LG100268 (p < 0.05). With chronic exposure, LG100268 upregulated palmitate oxidation (53 +/- 12% increase, p < 0.005), in a way similar to troglitazone (68 +/- 23%, p < 0.005). Synergism was observed when both agonists were combined (146 +/- 38%, p < 0.005 vs either agent alone). Treatment with either agent led to about a twofold increase in the expression of fatty acid transporter (FAT/CD36). Troglitazone upregulated PPAR gamma protein expression, whereas LG100268 had no effect. Furthermore, neither LG100268 nor troglitazone had any effect on the protein expression of RXR isoforms or PPAR alpha., Conclusion/interpretation: Co-activation of PPAR gamma and RXR results in additive or synergistic effects on glucose and lipid metabolism in skeletal muscle, but unlike troglitazone, LG100268 does not alter expression of its own receptor.

Aims/hypothesis: To evaluate the tissue distribution and possible role of the peroxisome proliferator-activated receptors (PPARs) in insulin action in fat and muscle biopsy specimens from lean, obese and subjects with Type II (non-insulin-dependent) diabetes mellitus., Methods: We measured PPAR alpha, PPAR beta (delta) and PPAR gamma protein expression by western blot analysis. The PPAR gamma protein was also measured in muscle before and after 3-h hyperinsulinaemic (300 mU.m-2.min-1) euglycaemic clamps., Results: The PPAR alpha protein was expressed preferentially in muscle relative to fat (more than sevenfold). The PPAR beta protein was similar in fat and muscle. The amount of PPAR gamma protein found in muscle was, on average, two-thirds of that present in fat. There was no statistically significant difference between non-diabetic and diabetic subjects in baseline (preclamp) muscle PPAR (alpha, beta or gamma) protein expression. Subgroup analysis showed, however, significantly higher PPAR gamma protein in the most insulin resistant diabetic subjects with glucose disposal rates of 3-6 mg.kg-1.min-1 compared with their age and weight matched counterparts with glucose disposal rates of 6-9 (147 +/- 23 vs 88 +/- 10 AU/microgram protein, p < or = 0.01 in diabetic and vs 94 +/- 15, p < or = 0.04 in non-diabetic subjects). Muscle PPAR gamma protein and glucose disposal rates were inversely correlated in diabetic subjects (r = -0.47, p < or = 0.05)., Conclusion/interpretation: All PPARs (alpha, beta or gamma) are present in skeletal muscle and adipose tissue with different relative distributions. The PPAR gamma protein is abundant in skeletal muscle as well as adipose tissue. The altered expression of skeletal muscle PPAR gamma is consistent with a role for this nuclear protein in the impaired insulin action of Type II diabetes.