A ketone monoester drink reduces postprandial blood glucose concentrations in adults with type 2 diabetes: a randomised controlled trial.

AIMS/HYPOTHESIS: The aim of the present study was to conduct a randomised, placebo-controlled, double-blind, crossover trial to determine whether pre-meal ketone monoester ingestion reduces postprandial glucose concentrations in individuals with type 2 diabetes. METHODS: In this double-blind, placebo-controlled, crossover design study, ten participants with type 2 diabetes (age 59±1.7 years, 50% female, BMI 32±1 kg/m2, HbA1c 54±2 mmol/mol [7.1±0.2%]) were randomised using computer-generated random numbers. The study took place at the Nutritional Physiology Research Unit, University of Exeter, Exeter, UK. Using a dual-glucose tracer approach, we assessed glucose kinetics after the ingestion of a 0.5 g/kg body mass ketone monoester (KME) or a taste-matched non-caloric placebo before a mixed-meal tolerance test. The primary outcome measure was endogenous glucose production. Secondary outcome measures were total glucose appearance rate and exogenous glucose appearance rate, glucose disappearance rate, blood glucose, serum insulin, ß-OHB and NEFA levels, and energy expenditure. RESULTS: Data for all ten participants were analysed. KME ingestion increased mean ± SEM plasma beta-hydroxybutyrate from 0.3±0.03 mmol/l to a peak of 4.3±1.2 mmol/l while reducing 2 h postprandial glucose concentrations by ~18% and 4 h postprandial glucose concentrations by ~12%, predominately as a result of a 28% decrease in the 2 h rate of glucose appearance following meal ingestion (all p<0.05). The reduction in blood glucose concentrations was associated with suppressed plasma NEFA concentrations after KME ingestion, with no difference in plasma insulin concentrations between the control and KME conditions. Postprandial endogenous glucose production was unaffected by KME ingestion (mean ± SEM 0.76±0.15 and 0.88±0.10 mg kg-1 min-1 for the control and KME, respectively). No adverse effects of KME ingestion were observed. CONCLUSIONS/INTERPRETATION: KME ingestion appears to delay glucose absorption in adults with type 2 diabetes, thereby reducing postprandial glucose concentrations. Future work to explore the therapeutic potential of KME supplementation in type 2 diabetes is warranted. TRIAL REGISTRATION: ClinicalTrials.gov NCT05518448. FUNDING: This project was supported by a Canadian Institutes of Health Research (CIHR) Project Grant (PJT-169116) and a Natural Sciences and Engineering Research Council (NSERC) Discovery Grant (RGPIN-2019-05204) awarded to JPL and an Exeter-UBCO Sports Health Science Fund Project Grant awarded to FBS and JPL.

Ambulance clinician use of capillary blood ketone meters to improve emergency hyperglycaemia care: A stepped-wedged controlled, mixed-methods feasibility study.

AIM: To determine whether it was feasible, safe and acceptable for ambulance clinicians to use capillary blood ketone meters for 'high-risk' diabetic ketoacidosis (DKA) recognition and fluid initiation, to inform the need for a full-powered, multi-centre trial. METHODS: Adopting a stepped-wedge controlled design, participants with hyperglycaemia (capillary blood glucose >11.0 mmol/L) or diabetes and unwell were recruited. 'High-risk' DKA intervention participants (capillary blood ketones ≥3.0 mmol/L) received paramedic-led fluid therapy. Participant demographic and clinical data were collated from ambulance and hospital care records. Twenty ambulance and Emergency Department clinicians were interviewed to understand their hyperglycaemia and DKA care experiences. RESULTS: In this study, 388 participants were recruited (Control: n = 203; Intervention: n = 185). Most presented with hyperglycaemia, and incidence of type 1 and type 2 diabetes was 18.5% and 74.3%, respectively. Ketone meter use facilitated 'high-risk' DKA identification (control: 2.5%, n = 5; intervention: 6.5%, n = 12) and was associated with improved hospital pre-alerting. Ambulance clinicians appeared to have a high index of suspicion for hospital-diagnosed DKA participants. One third (33.3%; n = 3) of Control and almost half (45.5%; n = 5) of Intervention DKA participants received pre-hospital fluid therapy. Key interview themes included clinical assessment, ambulance DKA fluid therapy, clinical handovers; decision support tool; hospital DKA management; barriers to hospital DKA care. CONCLUSIONS: Ambulance capillary blood ketone meter use was deemed feasible, safe and acceptable. Opportunities for improved clinical decision making, support and safety-netting, as well as in-hospital DKA care, were recognised. As participant recruitment was below progression threshold, it is recommended that future-related research considers alternative trial designs. CLINICALTRIALS: gov: NCT04940897.

Ketogenic diet with medium-chain triglycerides restores skeletal muscle function and pathology in a rat model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is an intractable genetic disease associated with progressive skeletal muscle weakness and degeneration. Recently, it was reported that intraperitoneal injections of ketone bodies partially ameliorated muscular dystrophy by increasing satellite cell (SC) proliferation. Here, we evaluated whether a ketogenic diet (KD) with medium-chain triglycerides (MCT-KD) could alter genetically mutated DMD in model rats. We found that the MCT-KD significantly increased muscle strength and fiber diameter in these rats. The MCT-KD significantly suppressed the key features of DMD, namely, muscle necrosis, inflammation, and subsequent fibrosis. Immunocytochemical analysis revealed that the MCT-KD promoted the proliferation of muscle SCs, suggesting enhanced muscle regeneration. The muscle strength of DMD model rats fed with MCT-KD was significantly improved even at the age of 9 months. Our findings suggested that the MCT-KD ameliorates muscular dystrophy by inhibiting myonecrosis and promoting the proliferation of muscle SCs. As far as we can ascertain, this is the first study to apply a functional diet as therapy for DMD in experimental animals. Further studies are needed to elucidate the underlying mechanisms of the MCT-KD-induced improvement of DMD.

Pharmacokinetic and metabolic profiling studies of osmundacetone in rats by UPLC-MS/MS and UPLC-QE-Orbitrap-HRMS.

Osmundacetone is a potential medicinal substance existing in ferns and has excellent antioxidant effects. This research aims to obtain the pharmacokinetic data for and metabolite products of osmundacetone. An UPLC-MS/MS quantitative method was established for the measurement of osmundacetonein in rat plasma over a linear range of 6.72-860.00 ng/ml. The signal to noise ratio of the lower limit of quantification was 60:1, the precision was <9.74% and the method had good selectivity and stability. The established method was successfully applied to the pharmacokinetic study of osmundacetone for the first time. Osmundacetone reached a peak at 0.25 h with a maximum value of 3283.33 µg/L. The apparent volume of distribution not multiplied by the bioavailability was 127.96 L/kg, and the half-life of osmundacetone was 5.20 h. At the same time, an UPLC-QE-Orbitrap-HRMS method was established to identify metabolites in plasma, urine and feces for the first time. A total of 30 metabolites were identified and the metabolic profile of osmundacetone was defined. In general, we have established a mass spectrometry quantitative method for osmundacetone for the first time and characterized its metabolic characteristics in rats.

A ketogenic drink improves cognition in mild cognitive impairment: Results of a 6-month RCT.

INTRODUCTION: Counteracting impaired brain glucose metabolism with ketones may improve cognition in mild cognitive impairment (MCI). METHODS: Cognition, plasma ketone response, and metabolic profile were assessed before and 6 months after supplementation with a ketogenic drink containing medium chain triglyceride (ketogenic medium chain triglyceride [kMCT]; 15 g twice/day; n = 39) or placebo (n = 44). RESULTS: Free and cued recall (Trial 1; P = .047), verbal fluency (categories; P = .024), Boston Naming Test (total correct answers; P = .033), and the Trail-Making Test (total errors; P = .017) improved significantly in the kMCT group compared to placebo (analysis of covariance; pre-intervention score, sex, age, education, and apolipoprotein E4 as covariates). Some cognitive outcomes also correlated positively with plasma ketones. Plasma metabolic profile and ketone response were unchanged. CONCLUSIONS: This kMCT drink improved cognitive outcomes in MCI, at least in part by increasing blood ketone level. These data support further assessment of MCI progression to Alzheimer's disease.

The Combination of Cigarette Smoking and Alcohol Consumption Synergistically Increases Reactive Carbonyl Species in Human Male Plasma.

Cigarette smoking and alcohol consumption are major risk factors for lifestyle-related diseases. Although it has been reported that the combination of these habits worsens risks, the underlying mechanism remains elusive. Reactive carbonyl species (RCS) cause chemical modifications of biological molecules, leading to alterations in cellular signaling pathways, and total RCS levels have been used as a lipid peroxidation marker linked to lifestyle-related diseases. In this study, at least 41 types of RCS were identified in the lipophilic fraction of plasma samples from 40 subjects using liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS). Higher levels of 10 alkanals, 5 trans-2-alkenals, 1 cis-4-alkenal, and 3 alkadienals were detected in the smoking/drinking group (N = 10) as compared to those with either habit (N = 10 each) or without both habits (N = 10) in the analysis of covariances adjusted for age and BMI. The levels of 3 alkanals, 1 trans-2-alkenal, 1 alkadienal, and 1 4-hydroxy-2-alkenal in the smoking/drinking group were significantly higher than those in the no-smoking/drinking and no-smoking/no-drinking groups. These results strongly indicate that the combination of cigarette smoking and alcohol drinking synergistically increases the level and variety of RCS in the circulating blood, and may further jeopardize cellular function.

Capillary blood tests may overestimate ketosis: triangulation between three different measures of ß-hydroxybutyrate.

The ketone body ß-hydroxybutyrate (BHB), assessed by a point-of-care meter in venous whole blood (BHBv), was used as the main outcome in a study on nutritional ketosis in healthy older adults. Two other BHB measures were also used in the study for validation and exploratory purposes, and here we report findings on correlation and agreement between those three methods. Ketosis in the range of 0-1.5 mmol/L was induced in 15 healthy volunteers by intake of medium-chain fatty acids after a 12-h fast. BHBv was assessed at 12 time points for 4 h. The same point-of-care meter was also used to test capillary blood (BHBc) at three time points, and a laboratory test determined total ketones (TK) in plasma (BHBp + acetoacetate) at four time points. A total of 180 cases included simultaneous data on BHBv, BHBc, BHBp, and TK. TK correlated with BHBp (Pearson's r = 0.99), BHBv (r = 0.91), and BHBc (r = 0.91), all P < 0.0001. BHBv and BHBp had good agreement in absolute values. However, the slope between BHBc and BHBv, measured with the same device, was in the range of 0.64-0.78 in different regression models, indicating substantially higher BHB concentrations in capillary versus venous blood. We conclude that all three methods are valid to detect relative changes in ketosis, but our results highlight the importance of method considerations and the possible need to adjust cutoffs, e.g., in the management of ketoacidosis and in the evaluation and comparison of dietary interventions.

Accuracy improvement of electrochemical whole blood ketone sensor based on HCT compensation algorithm.

In this manuscript, we demonstrated an electrochemical test strip with HCT (hematocrit) compensation algorithm to improve the accuracy of blood ketone sensor. In the conventional electrochemical sensor, the electrochemical current was directly resolved into the concentration value of the determinant without HCT compensation. For lower or higher HCT blood sample, the measured result was inaccurate. In the proposed design, the blood impedance can be measured to estimate the HCT, which was utilized to compensate the electrochemical current to resolve the more accurate concentration of determinant. The practical blood sample tests demonstrated the proposed design can provide more believable and reliable measured result in clinical point-of-care setting.

SGLT2 inhibition versus sulfonylurea treatment effects on electrolyte and acid-base balance: secondary analysis of a clinical trial reaching glycemic equipoise: Tubular effects of SGLT2 inhibition in Type 2 diabetes.

Sodium-glucose transporter (SGLT)2 inhibitors increase plasma magnesium and plasma phosphate and may cause ketoacidosis, but the contribution of improved glycemic control to these observations as well as effects on other electrolytes and acid-base parameters remain unknown. Therefore, our objective was to compare the effects of SGLT2 inhibitors dapagliflozin and sulfonylurea gliclazide on plasma electrolytes, urinary electrolyte excretion, and acid-base balance in people with Type 2 diabetes (T2D). We assessed the effects of dapagliflozin and gliclazide treatment on plasma electrolytes and bicarbonate, 24-hour urinary pH and excretions of electrolytes, ammonium, citrate, and sulfate in 44 metformin-treated people with T2D and preserved kidney function. Compared with gliclazide, dapagliflozin increased plasma chloride by 1.4 mmol/l (95% CI 0.4-2.4), plasma magnesium by 0.03 mmol/l (95% CI 0.01-0.06), and plasma sulfate by 0.02 mmol/l (95% CI 0.01-0.04). Compared with baseline, dapagliflozin also significantly increased plasma phosphate, but the same trend was observed with gliclazide. From baseline to week 12, dapagliflozin increased the urinary excretion of citrate by 0.93 ± 1.72 mmol/day, acetoacetate by 48 µmol/day (IQR 17-138), and ß-hydroxybutyrate by 59 µmol/day (IQR 0-336), without disturbing acid-base balance. In conclusion, dapagliflozin increases plasma magnesium, chloride, and sulfate compared with gliclazide, while reaching similar glucose-lowering in people with T2D. Dapagliflozin also increases urinary ketone excretion without changing acid-base balance. Therefore, the increase in urinary citrate excretion by dapagliflozin may reflect an effect on cellular metabolism including the tricarboxylic acid cycle. This potentially contributes to kidney protection.

Ketones and pain: unexplored role of hydroxyl carboxylic acid receptor type 2 in the pathophysiology of neuropathic pain.

The mechanisms underlying neuropathic pain are poorly understood. Here we show the unexplored role of the hydroxyl carboxylic acid receptor type 2 (HCAR2) in 2 models of neuropathic pain. We used an oral treatment with dimethyl fumarate and the HCAR2 endogenous ligand ß-hydroxybutyrate (BHB) in wild-type (WT) and HCAR2-null mice. We found an up-regulation of the HCAR2 in the sciatic nerve and the dorsal root ganglia in neuropathic mice. Accordingly, acute and chronic treatment with dimethylfumarate (DMF) and BHB reduced the tactile allodynia. This effect was completely lost in the HCAR2-null mice after a 2-d starvation protocol, in which the BHB reached the concentration able to activate the HCAR2-reduced tactile allodynia in female WT mice, but not in the HCAR2-null mice. Finally, we showed that chronic treatment with DMF reduced the firing of the ON cells (cells responding with an excitation after noxious stimulation) of the rostral ventromedial medulla. Our results pave the way for investigating the mechanisms by which HCAR2 regulates neuropathic pain plasticity.-Boccella, S., Guida, F., De Logu, F., De Gregorio, D., Mazzitelli, M., Belardo, C., Iannotta, M., Serra, N., Nassini, R., de Novellis, V., Geppetti, P., Maione, S., Luongo, L. Ketones and pain: unexplored role of hydroxyl carboxylic acid receptor type 2 in the pathophysiology of neuropathic pain.

Accuracy of Flash Glucose Monitoring in a Patient with Dravet Syndrome on a Ketogenic Diet.

We herein report the case of a 3-year-old girl with severe myoclonic epilepsy of infancy known as Dravet syndrome (DS) on a ketogenic diet (KD) whose glucose concentrations were controlled by using a flash glucose monitoring system. Two-hundred ninety-three events of moderate hypoglycemia with a minimum of 45 mg/dL, not related to day or night, were recorded during the observational period. Hypoglycemia rate declined from 24.5% of all measurements to 11.8% over time; one hypoglycemia-associated seizure and one seizure due to ketone concentrations below the therapeutic range were observed. In summary, this case report broadens our understanding of hypoglycemia risk in patients with DS on a KD. Especially in childhood, the painless and easy detection of low glucose concentrations might lead to improved cognitive performance, and the reduction of hypoglycemia-induced seizures.

Acute coffee ingestion with and without medium-chain triglycerides decreases blood oxidative stress markers and increases ketone levels.

Ingestion of ketone supplements, caffeine, and medium-chain triglycerides (MCTs) may all be effective strategies to increase blood levels of the ketone body beta-hydroxybutyrate (D-BHB). However, acute ingestion of a bolus of lipids may increase oxidative stress (OS). The purpose of the study was to investigate the impact of adding varying amounts of MCTs to coffee on blood levels of D-BHB and markers of OS. Ten college-aged men ingested coffee with 0, 28, and 42 g of MCT in a randomized order. Blood samples were collected pre- as well as 2 and 4 h postprandial and analyzed for D-BHB, total cholesterol (TC), high-density lipoprotein cholesterol (HDL-c), glucose, triglycerides (TAG), insulin, and OS markers: advanced oxidation protein products (AOPP), glutathione (GSH), malondialdehyde (MDA), and hydrogen peroxide (H2O2). All three treatments resulted in a significant increase in D-BHB, HDL-c, and TC as well as a significant decrease in TAG, MDA, H2O2, and insulin. The 42 g treatment was associated with significantly higher levels of AOPP and MDA. Acute ingestion of coffee results in favorable changes to markers of cardiometabolic health that were not impacted by the addition of 28 g of MCT. However, 42 g of MCT caused significantly greater OS.

The effect of meloxicam on neonatal dairy calves: Immunoglobulin G uptake and preweaning performance.

Objectives of this study were to determine effects of meloxicam administered in 2 forms on IgG uptake, growth, and health of preweaned calves. Sixteen Holstein bulls and 14 heifers with a body weight (BW) of 44.3 ± 5.24 kg were blocked by birth date in a randomized complete block design. Calves were removed from the dam before suckling, weighed, and randomly assigned to 1 of 3 treatments: (1) colostrum replacer (CR) at 0 h with no meloxicam (control; CON), (2) 1 mg/kg of BW of meloxicam in pill form before CR (PL), or (3) 1 mg/kg of BW of meloxicam mixed in solution with CR (SL). Calves were fed 675 g of dry matter of CR, providing a volume of 3 L and 180 g of IgG. Blood samples were collected at 0 h to analyze initial IgG and ketone concentrations, and at 6, 12, 18, and 24 h to analyze IgG uptake. At 24 h, calves were fed 432 g of dry matter of 24% crude protein milk replacer (MR) split in 2 feedings, and free choice starter and water until 42 d. Weekly blood samples were analyzed for glucose, plasma urea nitrogen, and ketone concentrations. Time of consumption of MR, BW, length, hip and withers height, and heart girth were recorded weekly. All calves achieved adequate transfer of immunity. Meloxicam did not affect apparent efficiency of absorption, serum total protein, or IgG uptake at 6, 18, and 24 h; however, meloxicam-treated calves had lesser IgG concentrations at 12 h (24.40 and 22.59 g/L for PL and SL, respectively) compared with CON (28.47 g/L). Meloxicam treatment did not affect BW. Calves that received PL tended to gain length at a faster rate (0.24 cm/d) than those that received SL (0.19 cm/d). Meloxicam treatment did not affect MR intake, time of consumption of MR, total dry matter intake, or feed efficiency. Meloxicam-treated calves tended to consume more starter (560.4 and 515.4 g/d for PL and SL, respectively) than those that received CON (452.6 g/d). Ketone levels tended to be greater in meloxicam-treated calves (0.15 and 0.17 mmol/L for PL and SL, respectively), suggesting improved rumen development compared with those that received CON (0.12 mmol/L). Meloxicam treatment did not affect plasma urea nitrogen . Glucose concentrations of calves that received PL (73.2 mg/dL) were less than those that received SL (83.3 mg/dL). Results of this study suggest that meloxicam given at 0 h offers positive effects on starter intake, and possibly rumen development, of preweaned dairy calves. Treatment PL, as compared with SL, offered positive results for rumen development, indicated by lower blood glucose levels.

Preserved glucose response to low-dose glucagon after exercise in insulin-pump-treated individuals with type 1 diabetes: a randomised crossover study.

AIMS/HYPOTHESIS: This study aimed to compare the increase in plasma glucose after a subcutaneous injection of 200 µg glucagon given after 45 min of cycling with resting (study 1) and to investigate the effects of glucagon when injected before compared with after 45 min of cycling (study 2). We hypothesised that: (1) the glucose response to glucagon would be similar after cycling and resting; and (2) giving glucagon before the activity would prevent the exercise-induced fall in blood glucose during exercise and for 2 h afterwards. METHODS: Fourteen insulin-pump-treated individuals with type 1 diabetes completed three visits in a randomised, placebo-controlled, participant-blinded crossover study. They were allocated by sealed envelopes. Baseline values were (mean and range): HbA1c 54 mmol/mol (43-65 mmol/mol) or 7.1% (6.1-8.1%); age 45 years (23-66 years); BMI 26 kg/m2 (21-30 kg/m2); and diabetes duration 26 years (8-51 years). At each visit, participants consumed a standardised breakfast 2 h prior to 45 min of cycling or resting. A subcutaneous injection of 200 µg glucagon was given before or after cycling or after resting. The glucose response to glucagon was compared after cycling vs resting (study 1) and before vs after cycling (study 2). RESULTS: The glucose response to glucagon was higher after cycling compared with after resting (mean ± SD incremental peak: 2.6 ± 1.7 vs 1.8 ± 2.0 mmol/l, p = 0.02). As expected, plasma glucose decreased during cycling (-3.1 ± 2.8 mmol/l) but less so when glucagon was given before cycling (-0.9 ± 2.8 mmol/l, p = 0.002). The number of individuals reaching glucose values ≤3.9 mmol/l was the same on the 3 days. CONCLUSIONS/INTERPRETATION: Moderate cycling for 45 min did not impair the glucose response to glucagon compared with the glucose response after resting. The glucose fall during cycling was diminished by a pre-exercise injection of 200 µg glucagon; however, no significant difference was seen in the number of events of hypoglycaemia. TRIAL REGISTRATION: Clinicaltrials.gov NCT02882737 FUNDING: The study was funded by the Danish Diabetes Academy founded by Novo Nordisk foundation and by an unrestricted grant from Zealand Pharma.

Spontaneous ketonuria and risk of incident diabetes: a 12 year prospective study.

AIMS/HYPOTHESIS: Ketones may be regarded as a thrifty fuel for peripheral tissues, but their clinical prognostic significance remains unclear. We investigated the association between spontaneous fasting ketonuria and incident diabetes in conjunction with changes in metabolic variables in a large population-based observational study. METHODS: We analysed 8703 individuals free of diabetes at baseline enrolled in the Korean Genome and Epidemiology Study, a community-based 12 year prospective study. Individuals with (n = 195) or without fasting ketonuria were matched 1:4 by propensity score. Incident diabetes was defined as fasting plasma glucose ≥7.0 mmol/l, post-load 2 h glucose ≥11.1 mmol/l on biennial OGTTs, or current use of glucose-lowering medication. Using Cox regression models, HRs for developing diabetes associated with the presence of ketonuria at baseline were analysed. RESULTS: Over 12 years, of the 925 participants in the propensity score-matched cohort, 190 (20.5%) developed diabetes. The incidence rate of diabetes was significantly lower in participants with spontaneous ketonuria compared with those without ketonuria (HR 0.63; 95% CI 0.41, 0.97). Results were virtually identical when participants with fasting ketonuria were compared against all participants without ketonuria (after multivariate adjustment, HR 0.66; 95% CI 0.45, 0.96). During follow-up, participants with baseline ketonuria maintained lower post-load 1 h and 2 h glucose levels and a higher insulinogenic index despite comparable baseline values. CONCLUSIONS/INTERPRETATION: The presence of spontaneous fasting ketonuria was significantly associated with a reduced risk of diabetes, independently of metabolic variables. Our findings suggest that spontaneous fasting ketonuria may have a potential preventive role in the development of diabetes.

Profiling of carboxyl-containing metabolites in smokers and non-smokers by stable isotope labeling combined with LC-MS/MS.

Profiling of carboxyl-containing metabolites in smokers and non-smokers provides insight into the smoking-related biological events and causal relationships between exposure and adverse events. However, more comprehensive analysis of carboxyl-containing metabolites in bio-matrices with high sensitivity and accuracy is challenging. In this work, stable isotope labeling in combination with liquid chromatography-tandem mass spectrometry was used for untargeted profiling and relative quantification of carboxyl-containing metabolites in plasma of smokers and non-smokers. A pair of isotope labeling reagents, N, N-dimethylethylenediamine (DMED) and d4-DMED was used to label carboxyl-containing metabolites. Since the isotope labeled dimethylamino moieties of DMED and d4-DMED are easily fragmented and lost as characteristic neutral fragments of 45 and 49 Da, respectively, double neutral loss scans can be used to profiling of carboxyl-containing metabolites. Subsequently, based on the ion pair parameters obtained from double neutral loss scans, relative quantification method was developed. As a result, 269 carboxyl-containing metabolite candidates were discovered, and 88 metabolite candidates were found to have significant alterations between smokers and non-smokers. 7Z, 10Z-hexadecadienoic acid, myristic acid and 3ß-hydroxy-5-cholestenoic acid with significant differences confirmed by standard comparison are linked to smoking related inflammation, abnormal bile acid synthesis and cholesterol metabolism.

Simplified LC/MS assay for the measurement of isolevuglandin protein adducts in plasma and tissue samples.

Isolevuglandins (IsoLGs) are a family of highly reactive 4-ketoaldehydes formed by lipid peroxidation that modify the lysyl residues of cellular proteins. Modification of proteins by IsoLGs have been shown to contribute to disease processes such as the development of hypertension. Accurate quantitation of the extent of protein modification by IsoLGs is essential for understanding the mechanisms whereby these modifications contribute to disease and the efficacy of interventions designed to prevent this modification. The previously described LC/MS assay to quantitate IsoLG protein adducts was extremely labor-intensive and time consuming, and while it offered reasonably low intra-day variation for replicate samples, variation when replicate samples were processed on separate days was significant. These limitations significantly restricted utilization of this approach. We therefore performed a series of studies to optimize the assay. We now report a significantly simplified LC/MS assay for measurement of IsoLG protein adducts with increased sensitivity and lower intra-day and inter-day variability.

Pioglitazone prevents the increase in plasma ketone concentration associated with dapagliflozin in insulin-treated T2DM patients: Results from the Qatar Study.

Because of the unique mechanism of action of sodium-glucose co-transport inhibitors (SGLT2i), which is independent of insulin secretion and insulin action, members of this class of drugs effectively lower plasma glucose concentration when used in combination with all other antidiabetic agents, including insulin. Increased plasma ketone concentration has been reported in association with SGLT2i initiation, which, under certain clinical conditions, has developed into diabetic ketoacidosis. The daily insulin dose often is reduced at the time of initiating SGLT2i therapy in insulin-treated patients to avoid hypoglycaemia. However, reduction of insulin dose can increase the risk of ketoacidosis. In the present study, we examined the effect of the addition of dapagliflozin plus pioglitazone on plasma ketone concentration in insulin-treated T2DM patients and compared the results to the effect of dapagliflozin alone. A total of 18 poorly controlled, insulin-treated T2DM participants in the Qatar Study received dapagliflozin (10 mg) plus pioglitazone (30 mg), and 10 poorly controlled non-insulin-treated T2DM patients received dapagliflozin (10 mg) alone for 4 months. Dapagliflozin plus pioglitazone produced a robust decrease in HbA1c (-1.4%) and resulted in a 50% reduction in daily insulin dose, from 133 to 66 units, while dapagliflozin alone caused a 0.8% reduction in HbA1c. Dapagliflozin caused a four-fold increase in fasting plasma ketone concentration, while the combination of pioglitazone plus dapagliflozin was not associated with a significant increase (0.13 vs 0.15 mM) in plasma ketone concentration or in risk of hypoglycaemia. These results demonstrate that the addition of pioglitazone to dapagliflozin prevents the increase in plasma ketone concentration associated with SGLT2i therapy.

Ketogenic diet compromises vertebral microstructure and biomechanical characteristics in mice.

Ketogenic diet (KD) compromised the microstructure of cancellous bone and the mechanical property in the appendicular bone of mice, while the effects of KD on the axial bone have not been reported. This study aimed to compare the changes in the microstructure and mechanical properties of the forth lumbar (L4) vertebra in KD and ovariectomized (OVX) mice. Forty eight-week-old female C57BL/6J mice were assigned into four groups: SD (standard diet) + Sham, SD + OVX, KD + Sham, and KD + OVX groups. L4 vertebra was scanned by micro-CT to examine the microstructure of cancellous bone, after which simulative compression tests were performed using finite element (FE) analysis. Vertebral compressive test and histological staining of the L4 and L5 vertebrae were performed to observe the biomechanical and histomorphologic changes. The KD + Sham and SD + OVX exhibited a remarkable declination in the parameters of cancellous bone compared with the SD + Sham group, while KD + OVX demonstrated the most serious bone loss in the four groups. The stiffness was significantly higher in the SD + Sham group than the other three groups, but no difference was found between the remaining groups. The trabecular parameters were significantly correlated with the stiffness. Meanwhile, the OVX + Sham and KD + OVX groups showed a significant decrease in the failure load of compressive test, while there was no difference between the KD + Sham and SD + Sham groups. These findings suggest that KD may compromise the vertebral microstructure and compressive stiffness to a similar level as OVX did, indicating adverse effects of KD on the axial bone of the mice.

High fat diet attenuates hyperglycemia, body composition changes, and bone loss in male streptozotocin-induced type 1 diabetic mice.

There is a growing and alarming prevalence of obesity and the metabolic syndrome in type I diabetic patients (T1DM), particularly in adolescence. In general, low bone mass, higher fracture risk, and increased marrow adipose tissue (MAT) are features of diabetic osteopathy in insulin-deficient subjects. On the other hand, type 2 diabetes (T2DM) is associated with normal or high bone mass, a greater risk of peripheral fractures, and no change in MAT. Therefore, we sought to determine the effect of weight gain on bone turnover in insulin-deficient mice. We evaluated the impact of a 6-week high-fat (HFD) rich in medium chain fatty acids or low-fat diet (LFD) on bone mass and MAT in a streptozotocin (STZ)-induced model using male C57BL/6J mice at 8 weeks of age. Dietary intervention was initiated after diabetes confirmation. At the endpoint, lower non-fasting glucose levels were observed in diabetic mice fed with high fat diet compared to diabetic mice fed the low fat diet (STZ-LFD). Compared to euglycemic controls, the STZ-LFD had marked polydipsia and polyphagia, as well as reduced lean mass, fat mass, and bone parameters. Interestingly, STZ-HFD mice had higher bone mass, namely less cortical bone loss and more trabecular bone than STZ-LFD. Thus, we found that a HFD, rich in medium chain fatty acids, protects against bone loss in a T1DM mouse model. Whether this may also translate to T1DM patients who are overweight or obese in respect to maintenance of bone mass remains to be determined through longitudinal studies.