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Background Many CpGs become hyper or hypo-methylated with age. Multiple methods have been developed by Horvath et al. to estimate DNA methylation (DNAm) age including Pan-tissue, Skin & Blood, PhenoAge, and GrimAge. Pan-tissue and Skin & Blood try to estimate chronological age in the normal population whereas PhenoAge and GrimAge use surrogate markers associated with mortality to estimate biological age and its departure from chronological age. Here, we applied Horvath’s four methods to calculate and compare DNAm age in 499 subjects with type 1 diabetes (T1D) from the Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications (DCCT/EDIC) study using DNAm data measured by Illumina EPIC array in the whole blood. Association of the four DNAm ages with development of diabetic complications including cardiovascular diseases (CVD), nephropathy, retinopathy, and neuropathy, and their risk factors were investigated. Results Pan-tissue and GrimAge were higher whereas Skin & Blood and PhenoAge were lower than chronological age (p < 0.0001). DNAm age was not associated with the risk of CVD or retinopathy over 18–20 years after DNAm measurement. However, higher PhenoAge (β = 0.023, p = 0.007) and GrimAge (β = 0.029, p = 0.002) were associated with higher albumin excretion rate (AER), an indicator of diabetic renal disease, measured over time. GrimAge was also associated with development of both diabetic peripheral neuropathy (OR = 1.07, p = 9.24E−3) and cardiovascular autonomic neuropathy (OR = 1.06, p = 0.011). Both HbA1c (β = 0.38, p = 0.026) and T1D duration (β = 0.01, p = 0.043) were associated with higher PhenoAge. Employment (β = − 1.99, p = 0.045) and leisure time (β = − 0.81, p = 0.022) physical activity were associated with lower Pan-tissue and Skin & Blood, respectively. BMI (β = 0.09, p = 0.048) and current smoking (β = 7.13, p = 9.03E−50) were positively associated with Skin & Blood and GrimAge, respectively. Blood pressure, lipid levels, pulse rate, and alcohol consumption were not associated with DNAm age regardless of the method used. Conclusions Various methods of measuring DNAm age are sub-optimal in detecting people at higher risk of developing diabetic complications although some work better than the others.

OBJECTIVE To examine the persistence of the original treatment effects 10 years after the Diabetes Control and Complications Trial (DCCT) in the follow-up Epidemiology of Diabetes Interventions and Complications (EDIC) study. In the DCCT, intensive therapy aimed at near-normal glycemia reduced the risk of microvascular complications of type 1 diabetes mellitus compared with conventional therapy. METHODS Retinopathy was evaluated by fundus photography in 1211 subjects at EDIC year 10. Further 3-step progression on the Early Treatment Diabetic Retinopathy Study scale from DCCT closeout was the primary outcome. RESULTS After 10 years of EDIC follow-up, there was no significant difference in mean glycated hemoglobin levels (8.07% vs 7.98%) between the original treatment groups. Nevertheless, compared with the former conventional treatment group, the former intensive group had significantly lower incidences from DCCT close of further retinopathy progression and proliferative retinopathy or worse (hazard reductions, 53%-56%; P < .001). The risk (hazard) reductions at 10 years of EDIC were attenuated compared with the 70% to 71% over the first 4 years of EDIC (P < .001). The persistent beneficial effects of former intensive therapy were largely explained by the difference in glycated hemoglobin levels during DCCT. CONCLUSION The persistent difference in diabetic retinopathy between former intensive and conventional therapy ("metabolic memory") continues for at least 10 years but may be waning. TRIAL REGISTRATION (clinicaltrials.gov) Identifiers: NCT00360815 and NCT00360893.

The Diabetes Control and Complications Trial (1983–1993) of 1,441 subjects followed for an average of 6.5 years assessed the effects of intensive therapy aimed at maintaining near normal levels of blood glucose versus conventional therapy on the risks of diabetes complications of the eyes, kidneys, and nerves. The study was designed to test the hypothesis that the higher than normal blood glucose levels associated with conventional insulin therapy caused these complications. The study was terminated one year ahead of schedule by the monitoring board. This paper describes the medical, ethical, and statistical challenges faced by the study group and the monitoring board.

▪ Abstract The Diabetes Control and Complications Trial (DCCT) demonstrated that intensive treatment of patients with insulin-dependent diabetes mellitus (IDDM) can substantially reduce the onset and progression of diabetic retinopathy, nephropathy, and neuropathy. The major risk associated with intensive treatment is recurrent hypoglycemia. Implementation of intensive treatment recommendations is difficult but should be considered and probably recommended to most patients with IDDM. If intensive treatment is impractical, any improvement in glycemic control is probably beneficial. Improved glycemic control should be recommended to most patients with non-insulin-dependent diabetes mellitus (NIDDM). The use of insulin in patients with NIDDM is controversial, especially in patients who are overweight, overeating, and minimally symptomatic.

Dichloroacetate is known to reduce plasma cholesterol and triglyceride in patients with Fredrickson Types IIb or IV hyperlipoproteinemia. We now report the effects of chronic, oral dichloroacetate administration (as the sodium salt) in two patients with severe homozygous familial hypercholesterolemia. Dichloroacetate markedly reduced serum total and low density lipoprotein cholesterol levels and lowered the low density lipoprotein to high density lipoprotein cholesterol ratio. One patient developed a polyneuropathy while receiving dichloroacetate which resolved following discontinuation of the drug. Because of its apparent toxicity, dichloroacetate cannot be recommended for chronic oral use. Investigation of the mechanism of its lipid-lowering effect, however, may provide insight into the pathogenesis and treatment of hypercholesterolemic disorders.

The metabolic response to exercise in insulin-dependent diabetic (IDD) man was assessed during continuous insulin infusion using the subcutaneous (CSII), intravenous (CIVII), and intraperitoneal (CIPII) routes. During the basal period, plasma glucose levels were higher with CIPII (153 +/- 17 mg/dl) than with CSII (117 +/- 13 mg/dl) or CIVII (118 +/- 17 mg/dl). Basal free insulin concentrations were similar for CSII (12.3 +/- 10 microU/ml) and CIVII (12.4 +/- 1.4 MicroU/ml) but lower in CIPII (8.5 +/- 1.0 microU/ml, P less than 0.05). Exercise on a stationary bicycle at 75 W for 60 min produced a decline of plasma glucose in each protocol that was significantly only during CIVII (55 +/- 11 mg/dl, P less than 0.01). Insulin levels remained unchanged throughout the study period in all protocols. In normals, insulin values decreased during exercise and remained below basal levels through the recovery period (P less than 0.05), while plasma glucose remained unchanged. Plasma glucagon and epinephrine levels were similar in all protocols and remained unchanged with exercise, while plasma norepinephrine tended to be higher than normal in all diabetic subjects. Significant differences between normal and diabetic subjects (P less than 0.05) were observed for blood ketone bodies, while blood lactate, glycerol, and plasma FFA were similar. Normalization of intermediary metabolites occurred only with CIVII. Continuous insulin infusion provides near-normal glycemic and metabolic control before, during and following exercise in IDD man. However, to produce normal blood concentrations of intermediary metabolites during exercise, the insulin infusion rate may be excessive in terms of its hypoglycemic effect. CSII appears to be a safe, accessible, and adequate method for treating diabetic man during exercise.

The roles of basal insulin, glucagon, and free fatty acids in the regulation of ketogenesis were studied in three-day-fasted anesthetized dogs. Four protocols were employed: (1) infusion of somatostatin, resulting in combined insulin and glucagon deficiency, (2) somatostatin and intraportal glucagon replacement (1 ng./kg./min.) to produce insulin deficiency, (3) somatostatin combined with intraportal insulin replacement (350 μU./kg./min.) to produce glucagon deficiency, and (4) saline controls. Simultaneous blood sampling from the femoral artery and portal and hepatic veins allowed determination of hepatic uptake or production of free fatty acids, glycerol, ketone bodies, and glucose. When combined deficiency of insulin and glucagon was produced, no significant effect on hepatic ketone production was noted (108 ± 6 per cent of mean basal), whereas the induction of selective Insulin deficiency (basal glucagon level maintained) resulted in a rise of net hepatic ketone production (185 ± 24 per cent of mean basal, P < 0.01). Isolated glucagon lack (basal insulin maintained) did not alter net hepatic ketone production. To assess the effect of increased substrate supply on ketogenesis, all experiments included a period of acute elevation of free fatty adds (FFA), produced by infusion of Intralipld and heparin. FFA elevation in saline controls caused only a moderate stimulation of ketone production (149 ± 16 per cent of mean basal, P < 0.01) despite a threefold increase in FFA uptake. However, the combination of elevation of FFA and selective insulin deficiency (glucagon maintained) resulted in a greatly increased hepatic ketone production (357 ± 58 per cent of mean basal, P < 0.01 vs. controls), which was also significantly higher than during combined insulin and glucagon deficiency (P < 0.05). Lipolysis, as indicated by arterial glycerol levels, as well as hepatic FFA uptake was not affected by acute suppression of pancreatic hormone levels. The study demonstrated that basal insulin plays an important role in repressing ketogenesis and that basal amounts of glucagon, while ineffective in the presence of insulin, exert a stimulatory effect on ketogenesis when insulin is deficient. Glucagon's stimulatory effect on ketogenesis was due to an effect in the liver rather than to an increase in lipolysis or in hepatic FFA uptake. Increasing FFA supply per se was associated with only limited stimulation of ketogenesis, whereas the combination of insulin deficiency, basal concentrations of glucagon, and increased FFA load produced a synergistic augmentation of hepatic ketone production.

Somatostatin has recently been used as a tool to investigate the roles of insulin and glucagon in the regulation of glucose metabolism in vivo. The alterations in glucose uptake and production that somatostatin causes have been attributed to the modifications it induces in pancreatic hormone levels rather than to effects of the peptide per se, although the direct effects of the latter on glucose turnover have not been clearly assessed. The aim of the present study therefore was to determine whether or not somatostatin directly affects either the rate of glucose uptake by fat and muscle or the rate of glucose production by the liver. Glucose uptake by the perfused rat hindquarter (skeletal muscle) was unaltered by somatostatin (10-1,000 ng./ml.) whether or not the process was stimulated by insulin (250 μU./ml.). Basal and insulin-(5-40 / μU./ml.)-stimulated glucose oxidation (uptake) by isolated rat adipocytes was also unaffected by somatostatin (5-500 ng./ml.). In addition, the peptide had no effect on basal or glucagon-(10−l0–10−9 M)-stimulated glucose output by isolated rat liver cells. Somatostatin also failed to modify the actions of these sub-maximally effective levels of glucagon on either the intracellular level of cAMP or the activity of cAMP-dependent protein kinase. The present studies support the hypothesis that the acute changes in the level and turnover of glucose induced by somatostatin are attributable to its effect on the endocrine system rather than to its direct effects on glucose metabolism.

Publisher Summary Isolated fat cells have been used extensively in the study of hormone actions. This preparation offers two major advantages: a uniform cell population from which, non-adipose elements can be readily removed and sensitivity to a number of hormones at physiological concentrations. Two different types of assays have been used to measure glucose transport and the effects of various hormones (primarily insulin) upon it; of these, only one (Method 2) measures transport directly. In this method, conveniently measured end products of [ 14 C] glucose metabolism are determined in the presence and absence of the hormone being tested. The validity of the method as a measure of glucose transport rests on the demonstration in intact fat pads that, under usual circumstances, transport is a rate–limiting step in glucose metabolism and that insulin exerts its major effect on glucose metabolism at the level of membrane translocation.

Publisher Summary Receptors for polypeptide hormones are cellular components that interact with circulating hormones with great sensitivity and specificity. In recent years, investigation of polypeptide hormone receptors has focused on purified plasma membrane preparations which contain hormone specific binding sites or adenylate cyclase activity. Inactivation of hormones by receptor preparations may be significant for several reasons. The relatively slow kinetics of binding of labeled hormones to membranes has been interpreted to indicate that some of the observed binding sites may not be involved in hormone action on cells and that, therefore, they are not receptors. An alternative explanation for the presence in plasma membranes of these highly specific sites is that some of them may be involved in hormone inactivation. As the simultaneous occurrence of hormone binding and inactivation by membranes greatly complicates analysis of binding data, inactivation of hormones by receptor preparations must always be evaluated in the course of binding studies. The methods for observing hormone inactivation by receptor preparations in vitro is discussed. Definitive methods for relating this inactivation to receptor function and for establishing its physiological importance are not yet available. However, possible approaches to these two questions are discussed, and methods have been selected for presentation based on physiological considerations.