Blood Ketone Research Articles

Lactobacillus rhamnosus GG protects against atherosclerosis by improving ketone body synthesis.

Atherosclerosis (AS) is a major cause of death and morbidity worldwide. There is an increasing amount of evidence that the gut microbiota plays an important role in disorders associated with lipid metabolism, such as AS, and alterations in the composition of the gut microbiota and its metabolic potential have been identified as contributing factors in the development of AS. Recently, probiotics have attracted great interest for their excellent cholesterol-lowering ability, their capacity to improve vascular endothelial function, and their participation in the remodeling of the intestinal flora to prevent AS. The incidental findings of our other study suggest that probiotic Lactobacillus rhamnosus GG may be associated with slowing the progression of AS. Thus, we delivered strain GG into mice by oral feeding and found that strain GG could effectively inhibit AS plaque generation. We analyzed the differences in gut microbiota composition and the peripheral blood metabolome in mice after oral feeding of strain GG by 16S DNA sequencing and untargeted metabolomics, respectively. The results showed that strain GG changed the composition of the gut microbiota in mice fed a high-fat diet; elevated the abundance of beneficial bacteria, such as Bilophila and Alistipes, and decreased the abundance of harmful bacteria, such as Deltaproteobacteria. The results of enrichment analysis of the gut microbiota and the peripheral blood metabolome both indicated that the antiatherosclerotic effect of strain GG might be associated with the biosynthesis pathway of ketone bodies. In addition, strain GG attenuated endothelial injury and elevated peripheral blood ketone body content in mice but did not significantly affect low-density lipoprotein cholesterol (LDL-C) and total cholesterol (TC) content. In conclusion, our study provides new evidence that strain GG slows the progression of AS, which may be associated with its improvement of the gut microbiome and peripheral blood metabolome, its ability to increase the abundance of beneficial bacteria, and its participation in unsaturated fatty acid and ketone body synthesis and degradation. KEY POINTS: • L. rhamnosus GG attenuated endothelial injury and atherosclerotic plaque formation • L. rhamnosus GG elevated the abundance of beneficial bacteria • L. rhamnosus GG elevated peripheral blood ketone body content in mice.

Intermittent fasting and mental and physical fatigue in obese and non-obese rats.

Intermittent fasting (IF) is an alternating pattern of restricting eating. This study evaluated mental and physical fatigue secondary to IF (daily 18-hour fast, 7-days-a-week) in the high-fat diet (HFD)-induced male obese Sprague Dawley rats. Fifty-four rats were randomly assigned to a HFD (n = 28) or a standard diet (SD; n = 26). After six weeks, the HFD rats were divided into one of four groups: obese HFD ad libitum (OB-HFD-AL), obese HFD-IF (OB-HFD-IF), obese SD-AL (OB-SD-AL), and obese SD-IF (OB-SD-IF). Similarly, non-obese controls were grouped into HFD-AL (C-HFD-AL), non-obese HFD-IF (C-HFD-IF), non-obese SD-AL (C-SD-AL), and non-obese SD-IF (C-SD-IF). After 2 weeks of IF, mental and physical fatigue were measured using open field (OF) and novel object recognition (NOR) tests. Rats on IF gained weight at a slower pace (p<0.05) and had lower glucose levels (p<0.01) compared to the AL group. In non-obese rats, ketone levels were higher in the IF-HFD group than IF-SD (p<0.05) and AL-SD (p<0.01) animals. Obese rats exhibited elevated blood ketone levels in IF-SD conditions versus AL-SD rats (p<0.01). AL-HFD rats had higher ketone levels than AL-SD animals in both obese and non-obese groups (p<0.05). In conclusion, rats with higher blood ketone levels, whether they were on IF or AL, traveled a greater distance during OF suggesting a lack of physical fatigue. There was no significant difference between IF and AL during NOR indicating a lack of mental fatigue. Thus, IF results in reduced body weight and blood glucose levels but does not induce physical or mental fatigue.

Dapagliflozin and euglycaemic diabetic ketoacidosis – ‘But doctor my sugars were normal!’

Dear Editor, A ‘Dear Doctor Letter’ was issued by the manufacturer of Forxiga (dapagliflozin) in November 2021; this followed reviews by the European Medicines Agency and the UK Medicines and Healthcare products Regulatory Agency. The letter stated that ‘The removal of the T1DM indication is not due to any safety concern for dapagliflozin in any indication, including T1DM’ but must be discontinued ‘in patients with T1DM by or in consultation with a physician specialised in diabetes care and be conducted as soon as clinically practical.’1 Dapagliflozin was previously approved in the treatment of diabetes for combination with insulin therapy in type 1 diabetes and with oral medications and insulin in type 2 patients. The pathogenesis of EuDKA occurs as a consequence of absolute insulin deficiency leading to reduced glucose utilisation and enhanced lipolysis; increased delivery of free fatty acids (FFAs) to the liver coupled with raised glucagon levels promotes FFA oxidation, and subsequent production of ketone bodies.2 Rosenstock and Ferrannini also reported that following a US FDA safety communication, the majority of reports of EuDKA between 2014 and 2015 in the US were actually in patients with type 2 diabetes.2 The UK Yellow Card report did not specify the type of diabetes. A review of the UK MHRA Yellow Card reporting system on dapagliflozin in October 20223 showed the following results: total number of adverse drug reactions (ADRs) reports, 2155; serious ADR reports, 1371; fatal ADR reports, 26 (cut-off 31 August 2022). Details of ADRs which could be related to EuDKA are shown in Table 1. The number of reports submitted had risen following the letter from the manufacturer. It is expected that a large number of these reports could be related to the concomitant use of dapagliflozin and insulin therapy in patients with type 2 diabetes. Appropriate monitoring and awareness on the part of health care professionals and patients are necessary to prevent potentially harmful and fatal consequences. The advice from both the manufacturer and the medicines regulatory agencies surrounding the actions required in patients with type 1 diabetes is clear and we may prevent problems by close monitoring of patients in diabetes specialist units. Such task is facilitated by the use of electronic medical records and access to the list of all medications used by these patients. Following the research findings on the occurrence of EuDKA in patients with type 2 diabetes, we may argue that these patients should also be included in a review for EuDKA. From personal and documented reports, patients with type 2 diabetes often react negatively to the introduction of insulin therapy to their medications.4 Part of the rationale to include insulin therapy usually includes improved glycaemic control. Following a period of improvement in their self-monitored blood glucose, some patients might react by stopping or reducing their use of insulin. The resulting ketonaemia in patients with insulin deficiency taking SGLT2 inhibitors is initially asymptomatic or clinically undetectable and could gradually lead to severe ketoacidosis. There are data which document the increased incidence of chronic heart failure, especially in people with type 2 diabetes;5 as this diagnosis is also included in the newer indications for dapagliflozin it will be important to communicate the association of EuDKA and dapagliflozin use to specialists who might not necessarily be expert in managing this complication of diabetes. Most patients with type 2 diabetes are managed in primary care and initiating a community-wide alert would take time to complete and require significant human resource, increased budget, and likely lead to increased patient anxiety. Blood tests to detect low endogenous insulin secretion would also take time and increased resources. A more rapid and simple approach is required. One option could be to request that all patients with long-standing type 2 diabetes who are treated with insulin in combination with dapagliflozin or any other SGLT2 inhibitors should be advised to use blood or urine ketone testing to detect EuDKA, particularly when insulin doses are being reduced or withdrawn. Awareness of these such parameters would help health professionals to quickly identify and treat patients at risk. The Association of British Clinical Diabetologists and Diabetes UK have produced joint guidance on communication and education requirements to be provided to patients who were prescribed an SGLT2 inhibitor for the first time.6 This provides useful and practical suggestions for minimising adverse reaction to the use of these medicines. Michael Stacey, MD, FRCP, Senior Lecturer, Royal Centre for Defence Medicine, UK There are no conflicts of interest declared.

LBSUN213 An Investigation Into The Effects Of Dapagliflozin On Ketogenesis In Type 1 Diabetes

Abstract Background Treatment with sodium-glucose transporter (SGLT)-2 inhibitors in patients with type 1 diabetes (T1DM) is associated with increased ketogenesis and diabetic ketoacidosis (DKA). The exact relationship between SGLT-2 inhibitors use in T1DM and ketogenesis is not fully understood. Patients and Methods In this single center, randomized, double blind and placebo-controlled trial 70 patients with established T1DM were randomized into placebo, dapagliflozin (SGLT-2 inhibitor) alone, exenatide/Bydureon (short and long-term acting GLP-1 RA, respectively) alone or with dapagliflozin and exenatide/Bydureon (triple therapy) groups in addition to routine insulin treatment (titrated in all groups to reduce risk of hypoglycemia). Patients were tested acutely (single dose before and after 12 weeks of treatment) under extended fasting and insulinopenic conditions (stress conditions) and under basal fasting conditions during 12 weeks treatment period. Results At end of 12 weeks treatment, glycemic indices improved significantly compared to baseline in the dapagliflozin and Bydureon alone groups and improved significantly compared to placebo in the triple therapy group (HbA1c: -0.9±0.1%, p&amp;lt;0. 001). Weight loss was significant (p&amp;lt;0. 001) in all active treatment groups. Ketones (beta-hydroxybutyrate, BHB) increased significantly in blood (0.21±0. 04 to 1.57±0.21 mM) and urine under acute stress conditions in control group at baseline and at 12 weeks visits. The magnitude of this increase was reduced following a single dose of dapagliflozin or exenatide at baseline visit (average of 1.16±0.17 mM) but was further increased following a single dose in the dapagliflozin receiving groups at 12 weeks visit (average of 1.83±0.28 mM). Under acute testing conditions, single dose of dapagliflozin treatment was associated with reduced urinary BHB clearance at baseline and 12 weeks visits. After 12 weeks of treatment and under basal conditions, ketone levels in blood and urine were not significantly altered following dapagliflozin or Bydureon alone but were significantly higher in the triple therapy group (0.22±0. 04 to 0.42±0. 07 mM in blood). At 12 weeks of treatment, dapagliflozin was associated with increased glucagon and lipolysis (increased FFAs) while Bydureon was associated with decreased glucagon and lipolysis. During the 12 weeks there were no episode of DKA or hypoglycemia (&amp;lt;55mg/dl). Conclusions Adjunct therapies treatment in T1DM results in significant improvements in glycemia and HbA1c. Dapagliflozin treatment increases lipolysis and glucagon while reduces urinary excretion of BHB. This may contribute to the increased blood ketone levels following extended treatment with dapagliflozin. However, the magnitude of this increase in ketones is significantly below DKA ketones levels which indicates that other DKA triggering factors play a more significant role. Presentation: Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m.

Point-of-care Capillary Blood Ketone Measurements and the Prediction of Future Ketoacidosis Risk in Type 1 Diabetes

Diabetic ketoacidosis (DKA) remains common in T1D and the identification of those at highest risk is challenging. We aimed to determine if levels of routine capillary blood ketone measurements could predict future DKA risk. In exploratory analysis, we examined 484 participants randomly assigned to placebo in an adjunct-to-insulin medication trial program (NCT02414958, NCT02580591). Participants provided morning fasted capillary blood ketone levels twice/week through an electronic logbook during a 2-month baseline period.

LBODP037 Acute Foot Drop In An Adolescent With Type 1 Diabetes Mellitus At Onset: A Case Report

Abstract Background The incidence of type 1 diabetes (T1D) among youth is on the rise worldwide. Diabetic neuropathy is often a late manifestation of diabetes and usually results from the exposure to prolonged hyperglycemia, oxidative and inflammatory stress, and dyslipidemia. Here, we report a case of painless diabetic mononeuropathy as the presenting symptom of T1D that improved shortly after the improvement in glycemic control. Clinical case A previously healthy 15-year-old boy, presented to the emergency department (ED) with left foot weakness and dragging for 1 week. This was preceded by numbness over the left lateral leg and dorsum of the left foot. There was no history of trauma. The patient reported a history of polyuria, polydipsia, and significant weight loss for 3 months, for which he did not seek medical care. There was no family history of diabetes, hereditary neuropathy, or autoimmune diseases. Upon presentation to the ED, a general physical examination was unremarkable with a weight of 60 Kg (BMI= 20 Kg/m2), neurological examination revealed weakness in the dorsiflexion of the left foot (3/5), eversion (4/5), and normal sensation and reflexes. The remainder of the neurological examination including the right foot, upper extremities, cerebellar functions, cranial nerves and monofilament and vibration sensory testing was normal. Fundoscopic exam showed no evidence of retinopathy. The initial laboratory investigations revealed mild DKA (PH 7.28, HCO3 16mmol/L, Na 137 mEq/L, random blood glucose of 306 mg/dl, positive blood ketones) and normal kidney function. A nerve conduction study showed moderate left peroneal motor axonal neuropathy, with normal sensory nerve action potentials. Brain CT scan was unremarkable. Further tests to rule out secondary causes of neuropathy (ANA, B12 level, and thyroid function) were within normal. The diagnosis of diabetes-related mononeuropathy was confirmed by the neurology team and the patient underwent physiotherapy. After the resolution of DKA, the patient continued to use basal/bolus insulin therapy and was discharged home with close follow-up of blood glucose readings. His glycemic control has improved, and the patient reported a significant improvement in the foot drop 3 weeks later. A repeated examination by the neurology team 3 months later confirmed a complete recovery of the left foot drop. Conclusion Acute diabetic mononeuropathy can be an unusual presenting feature of T1D and may pose a diagnostic challenge. Although diabetic mononeuropathy is often a late complication of diabetes, our patient presented with acute mononeuropathy at the onset of T1D. This case highlights the importance of early diagnosis and management of diabetes to prevent and sometimes reverse diabetic neuropathy. It also shows that a complete recovery of acute diabetic mononeuropathy can be achieved by improving glycemic control in patients with newly diagnosed T1D. Presentation: No date and time listed

Effects of High-Fat and Low-Carbohydrate Diet Combined with Radiation Therapy on Tumor Microenvironment in LLC Tumor-Bearing Mice

<h3>Purpose/Objective(s)</h3> Metabolic reprogramming of tumor immune cells can regulate immune function. To add a sentence linking tumor metabolism with radioimmunity. In this study, the high heterogeneity and reprogramming of tumor cell metabolism were utilized to explore the effects of high fat and low carbohydrate diet combined with radiotherapy on tumor immune microenvironment of tumor mice bearing LLC from the perspective of tumor microenvironment. <h3>Materials/Methods</h3> Lewis lung cancer (LLC) model was established in female C57BL/6J mice aged 6 to 8 weeks and around 18g, and they were given a high fat and low carbohydrate diet (45F, fat 45.00% of energy) combined with radiotherapy. The body weight, tumor growth, blood glucose and blood ketone (βHB) values, survival time, median survival time and treatment safety of tumor-bearing mice were observed. Immunohistochemistry, multiple immunofluorescence and flow cytometry were used to detect the expression of tumor-associated microangiogenesis molecules (CD34), lymphatic microvessels (LYVE-1), collagen fibers and immune cell phenotypes (CD3, CD4, CD8, Treg and PD1). <h3>Results</h3> High fat and low carbohydrate diet combined with radiotherapy (45F+RT) group had the best tumor control, and the tumor volume was significantly smaller than the control group (p < 0.001), and tumor weight in the combined treatment group (45F+RT) was significantly lower than that in the normal diet (PT, fat 12.11% of energy) and 45F group (p < 0.001); The blood glucose in 45F group was lower than that in PT group, but there was no statistical difference (p < 0.05); It was found that the βHB value in the high-fat and low-carbohydrate diet group was higher than that in the normal diet group. There were statistically significant differences in the second and third weeks (1.00±0.20mmol/L vs 0.63±0.06mmol/L, 0.90±0.17mmol/L vs 0.70±0.10mmol/L, p < 0.05). The median survival time of mice in each group was 38 days in the PT group, 55 days in the PT+RT group and 41 days in the 45F group, but not in the 45F+RT group. Immunohistochemistry and Multiple immunofluorescences showed that CD8 expression was up-regulated and PD1, CD34, LYVE-1 and collagen fiber expression were down-regulated in 45F+RT group. Flow cytometry showed that the proportion of CD8+T cells in 45F+RT group was higher than that in PT group and PT+RT group (p < 0.05) and higher than 45F group, but the difference was not statistically significant (p < 0.05); The proportion of CD4+T cells in PT+RT and 45F+RT groups was higher than that in PT and 45F groups (p < 0.05); The proportion of Treg in T cells in 45F+RT group was lower than that in PT+RT group, but the difference was not statistically significant (p < 0.05). <h3>Conclusion</h3> High fat and low carbohydrate diet combined with radiotherapy can enhance the recruitment and function of immune effector cells in tumor microenvironment and inhibit tumor microangiogenesis, thereby inhibiting tumor growth.

Caloric Restriction Ketogenic Diets (KR) Enhance Radiotherapy Responses in Lung Cancer Xenografts

<h3>Purpose/Objective(s)</h3> Dietary interventions can change metabolite levels in the tumor microenvironment, which might then affect cancer cell metabolism to alter tumor growth. Caloric restriction (CR) and ketogenic diet (KD) are often thought to limit tumor progression. <h3>Materials/Methods</h3> Mice bearing Lewis Lung Carcinoma xenografts were randomly divided into ad libitum diet group (AL), radiation group (RT), KR diet group (KR), or KR + RT group (KT). Mice in KR and KT group were fed a KR (KetoCal 4:1 fats: proteins+carbohydrates; 70% of total energy). Mice in RT and KT group were treated with radiation (4Gy*3f). Mice weights, tumor size, blood ketone levels and survival were monitored. <h3>Results</h3> Blood ketone levels were significantly higher for the mice consuming a KR diet compared to standard diet (3.2 mmol/L vs 0.8 mmol/L at week 3, P < .001). Some mice with KR diet lost a significant amount of weight (10%ཞ15% of starting weight) by the end of the treatment. The KR diets combined with radiation resulted in slower tumor growth in Lewis Lung Carcinoma xenografts and longer survival time (P < 0.05), relative to radiation alone. The KR diet also slowed tumor growth and prolonged survival (P < 0.05), relative to control. Hematological and pathological examinations indicated that the KR diet was safe and well tolerated, and no significant cardiac, hepatic and renal toxicity was observed. <h3>Conclusion</h3> Blood ketone levels were significantly higher for the mice consuming a KR diet compared to standard diet (3.2 mmol/L vs 0.8 mmol/L at week 3, P < .001). Some mice with KR diet lost a significant amount of weight (10%ཞ15% of starting weight) by the end of the treatment. The KR diets combined with radiation resulted in slower tumor growth in Lewis Lung Carcinoma xenografts and longer survival time (P < 0.05), relative to radiation alone. The KR diet also slowed tumor growth and prolonged survival (P < 0.05), relative to control. Hematological and pathological examinations indicated that the KR diet was safe and well tolerated, and no significant cardiac, hepatic and renal toxicity was observed.

Early time-restricted eating may favorably impact cognitive acuity in university students: a randomized pilot study

Cognitive acuity is important for academic success. The preliminary efficacy of early time-restricted eating (eTRE) on cognitive acuity was examined in a sample of university students attending a large southwestern university. It was hypothesized that adherence to eTRE would result in improved cognitive acuity as assessed by the Trail Making Tests (i.e., TMT-A and TMT-B) and the Stroop Color and Word Test. Participants were randomized to an intervention group (eTRE) with a daily 6-hour eating window or a control group (CON) with a daily 16-hour eating window; 24 participants initiated the study (13 eTRE and 11 CON). Blood ketones (i.e., β-hydroxybutyrate [BHB]) and brain-derived neurotrophic factor were also assessed. The 8-week intervention was forestalled by COVID-19 laboratory closures beginning in March 2020, and only data obtained at week 0 (baseline) and week 4 were viable for statistical analyses. Study attrition was high (50% for eTRE and 23% for CON). At week 4, there was a 27% and 6% decrease in TMT-B response time for the eTRE and CON groups, respectively (P=.02). TMT-B response time was inversely correlated to plasma BHB among participants (r=-0.633; P=.008). Considering the inverse relationship between TMT-B response time and plasma BHB, these preliminary data suggest that eTRE may improve some aspects of cognitive acuity in this population. Future investigations are needed to confirm these findings and should accommodate individual preferences regarding the initiation time of the eating window while also considering the impact on social influences and exercise engagement.

Dialysis as a Novel Adjuvant Treatment for Malignant Cancers.

Simple SummaryThere is a clear need for new cancer therapies as many cancers have a very short long-term survival rate. For most advanced cancers, therapy resistance limits the benefit of any single-agent chemotherapy, radiotherapy, or immunotherapy. Cancer cells show a greater dependence on glucose and glutamine as fuel than healthy cells do. In this article, we propose using 4- to 8-h dialysis treatments to change the blood composition, i.e., lowering glucose and glutamine levels, and elevating ketone levels—thereby disrupting major metabolic pathways important for cancer cell survival. The dialysis’ impact on cancer cells include not only metabolic effects, but also redox balance, immunological, and epigenetic effects. These pleiotropic effects could potentially enhance the effectiveness of traditional cancer treatments, such as radiotherapies, chemotherapies, and immunotherapies—resulting in improved outcomes and longer survival rates for cancer patients. Cancer metabolism is characterized by an increased utilization of fermentable fuels, such as glucose and glutamine, which support cancer cell survival by increasing resistance to both oxidative stress and the inherent immune system in humans. Dialysis has the power to shift the patient from a state dependent on glucose and glutamine to a ketogenic condition (KC) combined with low glutamine levels—thereby forcing ATP production through the Krebs cycle. By the force of dialysis, the cancer cells will be deprived of their preferred fermentable fuels, disrupting major metabolic pathways important for the ability of the cancer cells to survive. Dialysis has the potential to reduce glucose levels below physiological levels, concurrently increase blood ketone body levels and reduce glutamine levels, which may further reinforce the impact of the KC. Importantly, ketones also induce epigenetic changes imposed by histone deacetylates (HDAC) activity (Class I and Class IIa) known to play an important role in cancer metabolism. Thus, dialysis could be an impactful and safe adjuvant treatment, sensitizing cancer cells to traditional cancer treatments (TCTs), potentially making these significantly more efficient.

Feasibility and Safety of a Combined Metabolic Strategy in Glioblastoma Multiforme: A Prospective Case Series.

Background Glioblastoma multiforme (GBM) may be susceptible to metabolic strategies such as fasting and ketogenic diets, which lower blood glucose and elevate ketones. Combining these two strategies may be an ideal approach for sustaining a potentially therapeutic glucose ketone index (GKI). In this prospective case series, we observed whether a combined metabolic strategy was feasible, safe, and capable of sustaining a GKI <6 in patients with GBM. Methods We provided recommendations and guidelines to 10 GBM patients at various stages of tumour progression and treatment that enabled them to complete a 5–7-day fast every 1–2 months combined with a modified ketogenic diet during the intervening weeks. Patients monitored their blood glucose and ketone levels and body weight. Adverse effects were assessed. Results Patients completed a mean of 161 ± 74 days of the combined metabolic strategy, with 34 ± 18 (21%) days of prolonged fasting (mean fast duration: 6.0 ± 1.4 days) and 127 ± 59 (79%) days on the ketogenic diet. The mean GKI for all 10 patients was 3.22 (1.28 during the fasts, 5.10 during the ketogenic diet). Body weight decreased by 8.4 ± 6.9 kg (11.2% decrease in baseline weight). The most common adverse effects attributed to the fasts and ketogenic diet were fatigue, irritability, and feeling lightheaded. The metabolic strategy did not interfere with standard oncological treatments. Conclusion This is the first study to observe the feasibility and safety of repeated, prolonged fasting combined with a modified ketogenic diet in patients with GBM. Using minimal support, patients maintained the combined metabolic strategy for 5–6 months while sustaining a potentially therapeutic mean GKI of 3.22. Weight loss was considerable. Adverse effects attributed to the metabolic strategy were mild, and it did not interfere with standard oncological treatments. Study Registration: This study is registered on the Australia New Zealand Clinical Trials Registry, number ACTRN12620001310954. The study was registered on 4 December 2020.

The Effects of A Short Period of A Ketogenic Diet on Blood Biochemistry and Immunological Status in Overweight Thai Subjects

Being overweight and obese is a fundamental health problem worldwide and is associated with many diseases. The ketogenic diet (KD) includes high fat and moderate protein, with the daily consumption of carbohydrates amounting to less than 10 % daily intake. KD is a popular choice for weight loss but has benefited many diseases, such as metabolic diseases and neurodegenerative disorders. Nonetheless, the effect of KD on overweight but healthy individuals is limited. Therefore, the objectives of the present study were to evaluate the effects of the KD program on the physical, blood biochemistry, hematological and immunological status of overweight, healthy individuals. The 9 overweight Thai women with body mass index (BMI) values over 25 kg/m2 participated in a self-conduct KD program for 28 days. The mean age of the group was 28.44 years. With 28-days of KD, a significant (p &lt; 0.01) increase in blood ketone confirmed the ketosis status of participants. The mean blood ketone concentration increased from 0.41 ± 0.061 to 1.22 ± 0.267 mmol/L. Most of the participants’ physical parameters, such as body mass index (BMI), body weight, chest circumferences, hip circumferences, and arm circumferences, significantly decreased after 28-days of KD. Furthermore, the biochemistry indices (total cholesterol, triglyceride, LDL-C, and HDL-C) did not significantly differ from the baseline. However, fasting blood sugar decreased (p &lt; 0.01) from 88.00 ± 1.014 to 83.78 ± 1.289 mg/dL. KD did not influence all hematological and immunological indices such as nitric oxide (NO) production (unstimulated and in 2.5 µg/mL PHA stimulation) and T cell subpopulations. Nonetheless, it increased NO production in 5 µg/mL PHA stimulation and mononuclear cell proliferation capacity (proliferation index). The data revealed that short-term KD in this study decreased participants’ physical body compositions and blood glucose levels. Moreover, the immune status after KD proved the immunomodulation effect of KD. HIGHLIGHTS Thai overweight subjects whose self-conducted 28-days ketogenic diet demonstrated Significantly decreased the bodyweight Significantly decreased the body compositions Significantly decreased blood sugar level. Significantly affected immune systems by increasing immune cell proliferation capacity and responding to the non-specific stimulator GRAPHICAL ABSTRACT

A retrospective analysis of the incidence, outcome and factors associated with the occurrence of euglycemic ketoacidosis in diabetic patients on sodium glucose co-transporter - 2 inhibitors undergoing cardiac surgery.

SGLT2i is a new class of drugs used for type 2 diabetes. SGLT2i are known to cause EuKA in the perioperative period. Euglycemic ketoacidosis (EuKA) can cause life-threatening metabolic acidosis in the perioperative setting. Though the event rate of SGLT2i associated diabetic ketoacidosis in nonoperative setting is low, incidence among peri-operative patients can be very high and remains unknown. The aim of this study was to find the incidence, analyze outcome, and establish correlation between risk factors and EuKA in cardiac surgical patients on SGLT2i. This is a retrospective study analyzing 24 cardiac surgical patients who were on SGLT2i for type 2 diabetes mellitus. Data collection included age, sex, BMI, preoperative HbA1C, albumin, creatinine, type of SGLT2i and timing of stopping before surgery, insulin administration in the immediate pre-operative period; use of CPB, GI infusion and inotropes in the intraoperative period; blood ketone, duration of ventilation, hydration status and length of postoperative stay in postoperative period. Patients were diagnosed to have EuKA if any one of the serially measured postoperative ketone values was more than 0.6 mmol/L (ketone positive). The collected data were used to find an association between the risk factors and the occurrence of EuKA. Of the 24 patients, 17 patients developed EuKA. (70.8.%). 10 of the 17 EuKA in our study required preoperative Insulin for diabetic control whereas none in the ketone negative patients required insulin. This was statistically significant (P = 0.019). Association of other factors to EuKA were not statistically significant. Though the event rate of SGLT2i associated Diabetic ketoacidosis in nonoperative setting is low, (17), the occurrence of EUKA in cardiac surgical patients on SGLT2i in our study was 70.8% (17 out of 24 patients). Patients who require insulin in addition to other oral hypoglycemic drugs for immediate preoperative glycemic control are at risk for the development of SGLT2 inhibitor-induced EuKA postoperatively. Missing the diagnosis of EuKA is fatal in these patients. We couldn't make a diagnosis in our first patient whom we lost. Since it was diagnosed in all our study patients by measuring serial ketone values, there was no mortality and insignificant morbidity. Cessation of SGLT2i before surgery, expectant watch for blood ketones, and treatment with GI infusion reduce morbidity and mortality in cardiac surgical patients.

Circadian clock controls rhythms in ketogenesis by interfering with PPARα transcriptional network

Ketone bodies are energy-rich metabolites and signaling molecules whose production is mainly regulated by diet. Caloric restriction (CR) is a dietary intervention that improves metabolism and extends longevity across the taxa. We found that CR induced high-amplitude daily rhythms in blood ketone bodies (beta-hydroxybutyrate [βOHB]) that correlated with liver βOHB level. Time-restricted feeding, another periodic fasting-based diet, also led to rhythmic βOHB but with reduced amplitude. CR induced strong circadian rhythms in the expression of fatty acid oxidation and ketogenesis genes in the liver. The transcriptional factor peroxisome-proliferator-activated-receptor α (PPARα) and its transcriptional target hepatokine fibroblast growth factor 21 (FGF21) are primary regulators of ketogenesis. Fgf21 expression and the PPARα transcriptional network became highly rhythmic in the CR liver, which implicated the involvement of the circadian clock. Mechanistically, the circadian clock proteins CLOCK, BMAL1, and cryptochromes (CRYs) interfered with PPARα transcriptional activity. Daily rhythms in the blood βOHB level and in the expression of PPARα target genes were significantly impaired in circadian clock-deficient Cry1,2-/- mice. These data suggest that blood βOHB level is tightly controlled and that the circadian clock is a regulator of diet-induced ketogenesis.

Post-translational dysregulation of glucose uptake during exhaustive cycling exercise in vastus lateralis muscle of healthy homozygous carriers of the ACE deletion allele.

Homozygous carriers of the deletion allele in the gene for angiotensin-converting enzyme (ACE-DD) demonstrate an elevated risk to develop inactivity-related type II diabetes and show an overshoot of blood glucose concentration with enduring exercise compared to insertion allele carriers. We hypothesized that ACE-DD genotypes exhibit a perturbed activity of signaling processes governing capillary-dependent glucose uptake in vastus lateralis muscle during exhaustive cycling exercise, which is associated with the aerobic fitness state. 27 healthy, male white Caucasian subjects (26.8 ± 1.1 years; BMI 23.6 +/− 0.6 kg m−2) were characterized for their aerobic fitness based on a threshold of 50 ml O2 min−1 kg−1 and the ACE-I/D genotype. Subjects completed a session of exhaustive one-legged exercise in the fasted state under concomitant measurement of cardiorespiratory function. Capillary blood and biopsies were collected before, and ½ and 8 h after exercise to quantify glucose and lipid metabolism-related compounds (lipoproteins, total cholesterol, ketones) in blood, the phosphorylation of 45 signaling proteins, muscle glycogen and capillaries. Effects of aerobic fitness, ACE-I/D genotype, and exercise were assessed with analysis of variance (ANOVA) under the hypothesis of a dominant effect of the insertion allele. Exertion with one-legged exercise manifested in a reduction of glycogen concentration ½ h after exercise (−0.046 mg glycogen mg−1 protein). Blood glucose concentration rose immediately after exercise in association with the ACE-I/D genotype (ACE-DD: +26%, ACE-ID/II: +6%) and independent of the fitness state (p = 0.452). Variability in total cholesterol was associated with exercise and fitness. In fit subjects, the phosphorylation levels of glucose uptake-regulating kinases [AKT-pT308 (+156%), SRC-pY419, p38α-pT180/T182, HCK-pY411], as well as cytokine/angiotensin 1-7 signaling factors [(STAT5A-pY694, STAT5B-pY699, FYN-pY420, EGFR-pY1086] were higher in angiotensin converting enzyme I-allele carriers than ACE-DD genotypes after exercise. Conversely, the AKT-S473 phosphorylation level (+117%) and angiotensin 2’s blood concentration (+191%) were higher in ACE-DD genotypes. AKT-S473 phosphorylation levels post-exercise correlated to anatomical parameters of muscle performance and metabolic parameters (p < 0.05 and │r│>0.70). The observations identify reciprocal alterations of S473 and T308 phosphorylation of AKT as gatekeeper of a post-translational dysregulation of transcapillary glucose uptake in ACE-DD genotypes which may be targeted in personalized approaches to mitigate type II diabetes.

Utility of a Handheld Blood Ketone Meter as a Postmortem Indicator of Diabetic Ketoacidosis.

When investigating a death potentially due to diabetic ketoacidosis (DKA), it is challenging to assess the glucose level in the blood, but β-hydroxybutyrate, the most prevalent ketone body in the blood, is relatively stable after death. The aim of this project is to prove that a commercially available ketone meter can be used on postmortem blood samples to aid the diagnosis of DKA in a novel setting (during coroner/medical examiner examination). Samples with acetone detected via gas chromatography were chosen retrospectively to determine whether the meter could detect ketones in postmortem blood (proof-of-concept). In all of the thawed samples, the meter detected an elevated ketone level. Samples were then obtained in a prospective manner to include those with a possible cause of death from DKA along with controls. We correctly identified 16 cases in which death was due to DKA with use of the ketone manner. The ketone levels ranged from 2.6 to 5.4 mmol/L in those cases. The diagnosis was confirmed with a greatly elevated vitreous glucose concentration or glycated hemoglobin concentration. Detecting the presence of ketones while in the autopsy suite allowed for more accurate preliminary diagnoses and utilization of resources.

A Mouse Model of Glycogen Storage Disease Type IX-Beta: A Role for Phkb in Glycogenolysis.

Glycogen storage disease type IX (GSD-IX) constitutes nearly a quarter of all GSDs. This ketotic form of GSD is caused by mutations in phosphorylase kinase (PhK), which is composed of four subunits (α, β, γ, δ). PhK is required for the activation of the liver isoform of glycogen phosphorylase (PYGL), which generates free glucose-1-phosphate monomers to be used as energy via cleavage of the α -(1,4) glycosidic linkages in glycogen chains. Mutations in any of the PhK subunits can negatively affect the regulatory and catalytic activity of PhK during glycogenolysis. To understand the pathogenesis of GSD-IX-beta, we characterized a newly created PHKB knockout (Phkb−/−) mouse model. In this study, we assessed fasting blood glucose and ketone levels, serum metabolite concentrations, glycogen phosphorylase activity, and gene expression of gluconeogenic genes and fibrotic genes. Phkb−/− mice displayed hepatomegaly with lower fasting blood glucose concentrations. Phkb−/− mice showed partial liver glycogen phosphorylase activity and increased sensitivity to pyruvate, indicative of partial glycogenolytic activity and upregulation of gluconeogenesis. Additionally, gene expression analysis demonstrated increased lipid metabolism in Phkb−/− mice. Gene expression analysis and liver histology in the livers of old Phkb−/− mice (>40 weeks) showed minimal profibrogenic features when analyzed with age-matched wild-type (WT) mice. Collectively, the Phkb−/− mouse recapitulates mild clinical features in patients with GSD-IX-beta. Metabolic and molecular analysis confirmed that Phkb−/− mice were capable of sustaining energy homeostasis during prolonged fasting by using partial glycogenolysis, increased gluconeogenesis, and potentially fatty acid oxidation in the liver.

Calorie Restriction Enhanced Glycogen Metabolism to Compensate for Lipid Insufficiency.

This study aims to investigate the metabolic phenotype and mechanism of 40% calorie restriction (CR) in mice. CR mice exhibit super-stable blood glucose, as evidenced by increased fasting blood glucose (FBG), decreased postprandial blood glucose, and reduced glucose fluctuations. Additionally, both fasting plasma insulin and the homeostasis model assessment of insulin resistance increase significantly in CR mice. Compared with control, the phosphorylation of insulin receptor substrates-1 and serine/threonine kinase decreases in liver and fat but increases in muscle of CR mice after insulin administration, indicating hepatic and adipose insulin resistance, and muscle insulin sensitization. CR reduces visceral fat much more than subcutaneous fat. The elevated FBG is negatively correlated with low-level fasting β-hydroxybutyrate, which may result from insufficient free fatty acids and diminishes ketogenic ability in CR mice. Furthermore, liver glycogen increases dramatically in CR mice. Analysis of glycogen metabolism related proteins indicates active glycogen synthesis and decomposition. Additionally, CR elevates plasma corticosterone and hypothalamic orexigenic gene expression. CR induces lipid insufficiency and stress, resulting in global physiological insulin resistance except muscle and enhances glycogen metabolism, culminating in the stability of blood glucose manifests in increased FBG, which compensates for insufficient blood ketones.

Correlation between blood ketones and exhaled acetone measured with a semiconducting gas sensor

Breath acetone (BrAce) has been reported to be useful for monitoring the pathophysiology of patients with diabetes. However, devices that measure BrAce are expensive, complex and uncommon. The FM-001, originally designed to monitor a marker of weight loss in healthy people, is a device for measuring BrAce. The FM-001 is a loading semiconducting gas sensor that is a simple and reusable device. The aim of this study was to evaluate the correlation between blood total ketone bodies (TKB) and BrAce measured with the FM-001 in patients with diabetes. Furthermore, through evaluation of that correlation, we sought to detect patients at high risk of developing diabetic ketoacidosis (DKA). Thirty-five participants (age 52 [40–57], T2DM 32, T1DM 3) were enrolled. Scatter plots and linear regression lines relating BrAce to TKB and the correlation coefficients were calculated. Receiver-operating characteristic analysis was performed to determine the cut-off for predicting patients prone to DKA. The results showed that BrAce strongly correlates with TKB (R = 0.828), and the correlation was stronger in patients whose serum C-peptide was not low. The optimal BrAce cut-off for predicting risk of developing DKA was 3400 ppb (AUC 0.924, sensitivity 73.3%, specificity 100%), which corresponds to a TKB ⩾ 1000 µmol l−1. BrAce also weakly correlated with free fatty acid. Thus, BrAce levels measured with the FM-001 strongly correlate with TKB, even in patients with diabetes. This suggests the FM-001 is a simple and potentially useful method for detecting diabetic ketosis. (UMIN-ID: UMIN000038086)

The impact of ketogenic diet on the gut microbiome and tumor growth in an in vivo epithelial ovarian cancer model (105)

<b>Objectives:</b> The ketogenic diet (KD) is hypothesized to protect against cancer progression due to metabolic reprogramming. Sustained dietary modifications alter the composition of the gut microbiome, which may influence tumor growth. Here, we present the impact of KD on the gut microbiome in relation to epithelial ovarian cancer (EOC) progression. <b>Methods:</b> C57Bl/6J ID8 (luciferase-tagged) tumor-bearing mice were fed a KD (10% protein, 0% carbohydrates, 90% fat), high fat diet (HFD) (10% protein, 15% carbohydrates, 75% fat), and control diet (CD) (10% protein, 77% carbohydrates, and 13% fat) (<i>n</i>=10 mice per group). Blood ketone and glucose levels were serially measured. EOC tumor growth was monitored with <i>in Vivo</i> Imaging System (IVIS) via intraperitoneal D-luciferin. Total tumor burden was measured in total flux (photons/second). Stool was collected at baseline, midpoint (four weeks after tumor cell injection), and endpoint (tumor mass of >150 mm³, ascites development, or weight loss >20%); 16S rDNA was isolated, and sequence reads were assigned to genus-level amplicon sequence variants (ASV) in DADA2 as well as analyzed for alpha and beta microbial diversity using Phyloseq. Statistical tests included analysis of variance (ANOVA), permutational multivariate analysis (PERMANOVA), linear regression, and Wilcoxon test. A p-value of <0.05 was considered statistically significant. <b>Results:</b> KD-fed mice exhibited an increase in ketone blood levels (2.22 mmol/L) compared to both HFD- (0.7 mmol/L, p<0.0001) and CD-fed mice (0.61 mmol/L, p<0.0001) at the endpoint. There was no significant difference in overall glucose levels or body mass composition between the groups. By day 42, there was a 9.1-fold increase in tumor growth from baseline in the KD-fed group compared to HFD-and CD-fed groups (9.1 vs 2.0 vs 3.1-fold, respectively, p<0.001 and p<0.001). By study endpoint, KD-fed mice exhibited a statistically significant increase in gut microbial alpha diversity in comparison to HFD and CD (p<0.01 at the midpoint and p<0.05 at the endpoint). Brays-Curtis dissimilarity based on microbial beta diversity demonstrated that the relative abundance of gut microbial species between the three groups was similar at baseline. However, KD-fed mice had significantly altered gut microbial composition than HFD-and CD-fed mice at both midpoint and endpoint (p = 0.001); 16S data analysis demonstrated a relative decrease in <i>Lactobacillus</i> and <i>Coriobacteriaceae</i> species and a relative increase in <i>Romboutsia</i> and <i>Akkermansia</i> species in the KD-fed mice. <b>Conclusions:</b> In our EOC murine model, KD was associated with increased EOC growth and distinct gut microbial alterations in comparison to HFD and CD. KD-fed mice exhibited increased gut microbial diversity over time, a relative depletion of bacterial species <i>Lactobacillus</i> and <i>Coriobacteriaceae</i>, and a relative enrichment of bacterial species <i>Romboutsia</i> and <i>Akkermansia</i>. Further studies are needed to understand the mechanisms underlying the relationship between KD, the gut microbiome, and EOC progression.Fig. 1