Diabetic ketoacidosis (DKA), a common onset modality of type 1 diabetes mellitus (T1DM), can lead, in rare instances, to the development of cerebral edema, which is the leading cause of mortality in T1DM. Aside from the identification of several demographic and clinical risk factors for cerebral edema, attention has also been drawn to the possible link between systemic inflammation and neuroinflammation. This single-center retrospective study of 98 children with severe DKA aimed to investigate the possible relationship between neutrophil-to-lymphocyte ratio NLR) levels and the presence of cerebral edema. Patients were classified into three groups: alert (n = 28), subclinical cerebral edema (n = 59), and overt cerebral edema (n = 11). Lower blood pH and elevated NLR and blood urea were correlated with the presence of cerebral edema (p < 0.001). After a multivariable risk adjustment for possible confounding factors, such as age, pH, corrected sodium, and BUN, the NLR remained positively associated with cerebral edema (p = 0.045). As such, NLR may be an additional instrument to help practitioners target patients with a higher risk of severe cerebral edema. These patients would benefit from more rigorous neurologic surveillance, enabling the prompt identification of early signs of cerebral edema.

A growing body of evidence supports the beneficial effects of the ketone bodies (KBs), acetoacetate and β-hydroxybutyrate (BHB), on diverse physiological processes and diseases. Hence, KBs have been suggested as therapeutic tools for neurodegenerative diseases. KBs are an alternative fuel during fasting and starvation as they can be converted to Ac-CoA to produce ATP. A ketogenic diet (KD), enriched in fats and low in carbohydrates, induces KB production in the liver and favors their use in the brain. BHB is the most abundant KB in the circulation; in addition to its role as energy fuel, it exerts many actions that impact the set of proteins in the cell and tissue. BHB can covalently bind to proteins in lysine residues as a new post-translational modification (PTM) named β-hydroxybutyrylation (Kbhb). Kbhb has been identified in many proteins where Kbhb sites can be critical for binding to other proteins or cofactors. Kbhb is mostly found in proteins involved in chromatin structure, DNA repair, regulation of spliceosome, transcription, and oxidative phosphorylation. Histones are the most studied family of proteins with this PTM, and H3K9bhb is the best studied histone mark. Their target genes are mainly related to cell metabolism, chromatin remodeling and the control of circadian rhythms. The role of Kbhb on physiological processes is poorly known, but it might link KB metabolism to cell signaling and genome regulation. BHB also impacts the proteome by influencing proteostasis. This KB can modulate the Unfolded Protein Response (UPR) and autophagy, two processes involved in the maintenance of protein homeostasis through the clearance of accumulated unfolded and damaged proteins. BHB can support proteostasis and regulate the UPR to promote metabolism adaptation in the liver and prevent cell damage in the brain. Also, BHB stimulates autophagy aiding to the degradation of accumulated proteins. Protein aggregation is common to proteinopathies like Alzheimer's (AD) and Parkinson's (PD) diseases, where the KD and BHB treatment have shown favorable effects. In the present review, the current literature supporting the effects of KBs on proteome conformation and proteostasis is discussed, as well as its possible impact on AD and PD.

Acetoacetate (AA) is an important ketone body that is used as an oxidative fuel to supply energy for the cellular activities of various tissues, including the brain and skeletal muscle. We recently revealed a new signaling role for AA by showing that it promotes muscle cell proliferation in vitro, enhances muscle regeneration in vivo, and ameliorates the dystrophic muscle phenotype of Mdx mice. In this study, we provide new molecular insight into this function of AA. We show that AA promotes C2C12 cell proliferation by transcriptionally upregulating the expression of muscle-specific miR-133b, which in turn stimulates muscle cell proliferation by targeting serum response factor. Furthermore, we show that the AA-induced upregulation of miR-133b is transcriptionally mediated by MEF2 via the Mek-Erk1/2 signaling pathway. Mechanistically, our findings provide further convincing evidence that AA acts as signaling metabolite to actively regulate various cellular activities in mammalian cells.

Diabetic ketoacidosis (DKA) is a serious complication of complete insulin deficiency and insulin resistance in Type 1 diabetes (T1D). This results in the body producing high levels of serum ketones in an attempt to compensate for the insulin deficiency and decreased glucose utilization. DKA's metabolic and immunologic dysregulation results in gradual increase of systemic and cerebral oxidative stress, along with low grade systemic and cerebral inflammation and the development of pretreatment subclinical BE. During treatment the early progression of oxidative stress and inflammation is hypothesized to advance the possibility of occurrence of crisis of clinical brain edema (BE), which is the most important cause of morbidity and mortality in pediatric DKA. Longitudinal neurocognitive studies after DKA treatment show progressive and latent deficits of cognition and emphasize the need for more effective DKA treatment of this long-standing conundrum of clinical BE, in the presence of systemic osmotic dehydration, metabolic acidosis and immune dysregulation. Candidate biomarkers of several systemic and neuroinflammatory pathways prior to treatment also progress during treatment, such as the neurotoxic and neuroprotective molecules in the well-recognized tryptophan (TRP)/kynurenine pathway (KP) that have not been investigated in DKA. We used LC-MS/MS targeted mass spectrometry analysis to determine the presence and initiation of the TRP/KP at three time points: A) 6-12 hours after initiation of treatment; B) 2 weeks; and C) 3 months following DKA treatment to determine if they might be involved in the pathogenesis of the acute vasogenic complication of DKA/BE. The Trp/KP metabolites TRP, KYN, quinolinic acid (QA), xanthurnenic acid (XA), and picolinic acid (PA) followed a similar pattern of lower levels in early treatment, with subsequent increases. Time point A compared to Time points B and C were similar to the pattern of sRAGE, lactate and pyruvic acid. The serotonin/melatonin metabolites also followed a similar pattern of lower quantities at the early stages of treatment compared to 3 months after treatment. In addition, glutamate, n-acetylglutamate, glutamine, and taurine were all lower at early treatment compared to 3 months, while the ketones 3-hydroxybutaric acid and acetoacetate were significantly higher in the early treatment compared to 3 months. The two major fat metabolites, L-carnitine and acetyl-L-carnitine (ALC) changed inversely, with ALC significantly decreasing at 2 weeks and 3 months compared to the early stages of treatment. Both anthranilic acid (AA) and 3-OH-anthranilic acid (3OH-AA) had overall higher levels in the early stages of treatment (A) compared to Time points (B and C). Interestingly, the levels of AA and 3OH-AA early in treatment were higher in Caucasian females compared to African American females. There were also differences in the metabolite levels of QA and kynurenic acid (KA) between genders and between races that may be important for further development of custom targeted treatments. We hypothesize that the TRP/KP, along with the other inflammatory pathways, is an active participant in the metabolic and immunologic pathogenesis of DKA's acute and chronic insults.

Cerebral edema (CE) is a potentially devastating complication of diabetic ketoacidosis (DKA) that almost exclusively occurs in children. Since its first description in 1936, numerous risk factors have been identified; however, there continues to be uncertainty concerning the mechanisms that lead to its development. Currently, the most widely accepted hypothesis posits that CE occurs as a result of ischemia-reperfusion injury, with inflammation and impaired cerebrovascular autoregulation contributing to its pathogenesis. The role of specific aspects of DKA treatment in the development of CE continues to be controversial. This review critically examines the literature on the pathophysiology of CE and attempts to categorize the findings by types of brain injury that contribute to its development: cytotoxic, vasogenic, and osmotic. Utilizing this scheme, we propose a multifactorial pathway for the development of CE in patients with DKA.

Diabetic ketoacidosis (DKA) has been associated with cognitive impairment and structural alterations in the brain. There is increased evidence supporting the role of neuroinflammation in causing these alterations. In the present study, using human microglial cell line (CHME-5), we aimed to investigate the effect of immunoglobulins (IG) on survival, activation, reactive oxygen species (ROS) and cytokine production of microglia exposed to ketone bodies. We demonstrated that high and low dose of ketone bodies induced a significant increase in ROS within 1 h after exposure to CHME-5 cells with upregulation in mitochondrial superoxide level 5 min after exposure suggestive of early and selective impairment of mitochondrial function. A significant and delayed increase of apoptosis of CHME-5 cells was observed 4 days after ketone bodies exposure. Cytokine expression reached a peak within 1 h and persisted for 3 days after exposure to ketone bodies. IG significantly reduced ROS and transiently suppressed cytokine expression of CHME-5 cells after exposure to ketone bodies. However, no effect of IG on apoptosis was observed. Overall, these results supported that ketone bodies induced microglia activation with early and selective impairment of mitochondrial function, increased cytokines expression and delayed increase in apoptosis. IG suppressed microglia activation and transiently inhibited cytokines expression without affecting apoptosis. These results warrant further experimental work on the role of microglia and potential benefit of IG in brain structural changes induced by DKA.

There is increasing awareness that in addition to the metabolic crisis of diabetic ketoacidosis (DKA) caused by severe insulin deficiency, the immune inflammatory response is likely an active multicomponent participant in both the acute and chronic insults of this medical crisis, with strong evidence of activation for both the cytokine and complement system. Recent studies report that the matrix metalloproteinase enzymes and their inhibitors are systemically activated in young Type 1 diabetes mellitus (T1D) patients during DKA and speculate on their involvement in blood-brain barrier (BBB) disruption. Based on our previous studies, we address the question if matrix metalloproteinase 9 (MMP9) is expressed in the brain in the fatal brain edema (BE) of DKA. Our data show significant expression of MMP9 on the cells present in brain intravascular areas. The presence of MMP9 in intravascular cells and that of MMP+ cells seen passing the BBB indicates a possible role in tight junction protein disruption of the BBB, possibly leading to neurological complications including BE. We have also shown that MMP9 is expressed on neurons in the hippocampal areas of both BE/DKA cases investigated, while expression of tissue inhibitor of metalloproteinases 1 (TIMP1) was reduced in the same areas. We can speculate that intraneuronal MMP9 can be a sign of neurodegeneration. Further studies are necessary to determine the role of MMP9 in the pathogenesis of the neurologic catastrophe of the brain edema of DKA. Inhibition of MMP9 expression might be helpful in preserving neuronal function and BBB integrity during DKA.

Blood-brain interfaces comprise the cerebral microvessel endothelium forming the blood-brain barrier (BBB) and the epithelium of the choroid plexuses forming the blood-cerebrospinal fluid barrier (BCSFB). Their main functions are to impede free diffusion between brain fluids and blood; to provide transport processes for essential nutrients, ions, and metabolic waste products; and to regulate the homeostasis of central nervous system (CNS), all of which are attributed to absent fenestrations, high expression of tight junction proteins at cell-cell contacts, and expression of multiple transporters, receptors, and enzymes. Existence of BBB is an important reason that systemic drug administration is not suitable for the treatment of CNS diseases. Some diseases, such epilepsy, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and diabetes, alter BBB function via affecting tight junction proteins or altering expression and function of these transporters. This chapter will illustrate function of BBB, expression of transporters, as well as their alterations under disease status.

Due to the limited data on diabetic ketoacidosis and brain edema (DKA/BE) in children/adolescents and the lack of recent data on adults with type 1 diabetes (T1D), we addressed the question of whether neuroinflammation was present in the fatal DKA of adults. We performed immunohistochemistry (IHC) studies on the brains of two young adults with T1D and fatal DKA and compared them with two teenagers with poorly controlled diabetes and fatal DKA. C5b-9, the membrane attack complex (MAC) had significantly greater deposits in the grey and white matter of the teenagers than the young adults (p=0.03). CD59, a MAC assembly inhibitory protein was absent, possibly suppressed by the hyperglycemia in the teenagers but was expressed in the young adults despite comparable average levels of hyperglycemia. The receptor for advanced glycation end products (RAGE) had an average expression in the young adults significantly greater than in the teenagers (p=0.02). The autophagy marker Light Chain 3 (LC3) A/B was the predominant form of programmed cell death (PCD) in the teenage brains. The young adults had high expressions of both LC3A/B and TUNEL, an apoptotic cell marker for DNA fragmentation. BE was present in the newly diagnosed young adult with hyperglycemic hyperosmolar DKA and also in the two teenagers. Our data indicate that significant differences in neuroinflammatory components, initiated by the dysregulation of DKA and interrelated metabolic and immunologic milieu, are likely present in the brains of fatal DKA of teenagers when compared with young adults.

Diets that boost ketone production are increasingly used for treating several neurological disorders. Elevation in ketones in most cases is considered favorable, as they provide energy and are efficient in fueling the body's energy needs. Despite all the benefits from ketones, the above normal elevation in the concentration of ketones in the circulation tend to illicit various pathological complications by activating injurious pathways leading to cellular damage. Recent literature demonstrates a plausible link between elevated levels of circulating ketones and oxidative stress, linking hyperketonemia to innumerable morbid conditions. Ketone bodies are produced by the oxidation of fatty acids in the liver as a source of alternative energy that generally occurs in glucose limiting conditions. Regulation of ketogenesis and ketolysis plays an important role in dictating ketone concentrations in the blood. Hyperketonemia is a condition with elevated blood levels of acetoacetate, 3-β-hydroxybutyrate, and acetone. Several physiological and pathological triggers, such as fasting, ketogenic diet, and diabetes cause an accumulation and elevation of circulating ketones. Complications of the brain, kidney, liver, and microvasculature were found to be elevated in diabetic patients who had elevated ketones compared to those diabetics with normal ketone levels. This review summarizes the mechanisms by which hyperketonemia and ketoacidosis cause an increase in redox imbalance and thereby increase the risk of morbidity and mortality in patients.

There is a significantly higher incidence of cardiovascular disease (CVD) among type 1 diabetic (T1D) patients than among non-diabetic subjects. T1D is associated with hyperketonemia, a condition with elevated blood levels of ketones, in addition to hyperglycemia. The biochemical mechanism by which vitamin D (VD) may reduce the risk of CVD is not known. This study examines whether VD can be beneficial in reducing hyperketonemia (acetoacetate, AA) induced oxidative stress in endothelial cells.

HUVEC were pretreated with 1,25(OH)2D3, and later exposed to the ketone body acetoacetate.

The increases in ROS production, ICAM-1 expression, MCP-1 secretion, and monocyte adhesion in HUVEC treated with AA were significantly reduced following treatment with 1,25(OH)2D3. Interestingly, an increase in glutathione (GSH) levels was also observed with 1,25(OH)2D3 in ketone treated cells. The effects of 1,25(OH)2D3 on GSH, ROS, and monocyte-endothelial adhesion were prevented in GCLC knockdown HUVEC. This suggests that 1,25(OH)2D3 inhibits ROS, MCP-1, ICAM-1, and adherence of monocytes mediated by the upregulation of GCLC and GSH.

This study provides evidence for the biochemical mechanism through which VD supplementation may reduce the excess monocyte adhesion to endothelium and inflammation associated with T1D.

Diabetic cardiomyopathy (DC) is an independent phenotype of diabetic cardiovascular disease. The understanding of the pathogenesis of DC in young patients with type 1 diabetes (T1D) is limited. The cardiac insults of diabetic ketoacidosis (DKA) and progression of DC could include development of antibodies (Abs) to cardiac self-antigens (SAgs) such as: myosin (M), vimentin (V) and k-alpha 1 tubulin (Kα1T). The goal of this study is to determine if the insults of severe DKA and its inflammatory cascade are associated with immune responses to SAgs. Development of Abs to the SAgs were determined by an ELISA using sera collected at three time points in relation to severe DKA (pH < 7.2). Results demonstrate significant differences between the development of Abs to VIM and a previously reported diastolic abnormality (DA) during DKA and its treatment and a NDA group at 2-3 months post DKA (p = 0.0452). A significant association is present between T1D duration (<3 years) and Abs to Kα1T (p = 0.0134). Further, Abs to MYO and VIM are associated with inflammatory cytokines. We propose that severe DKA initiates the synthesis of Abs to cardiac SAgs that are involved in the early immunopathogenesis of DC in young patients with T1D.

Acetoacetate (AA) is a ketone body and acts as a fuel to supply energy for cellular activity of various tissues. Here, we uncovered a novel function of AA in promoting muscle cell proliferation. Notably, the functional role of AA in regulating muscle cell function is further evidenced by its capability to accelerate muscle regeneration in normal mice, and it ameliorates muscular dystrophy in mdx mice. Mechanistically, our data from multiparameter analyses consistently support the notion that AA plays a non-metabolic role in regulating muscle cell function. Finally, we show that AA exerts its function through activation of the MEK1-ERK1/2-cyclin D1 pathway, revealing a novel mechanism in which AA serves as a signaling metabolite in mediating muscle cell function. Our findings highlight the profound functions of a small metabolite as signaling molecule in mammalian cells.

The incidence of developing microvascular dysfunction is significantly higher in type 1 diabetic (T1D) patients. Hyperketonemia (acetoacetate, β-hydroxybutyrate) is frequently found along with hyperglycemia in T1D. Whether hyperketonemia per se contributes to the excess oxidative stress and cellular injury observed in T1D is not known.

HUVEC were treated with ketones in the presence or absence of high glucose for 24 h. NOX4 siRNA was used to specifically knockdown NOX4 expression in HUVEC.

Ketones alone or in combination with high glucose treatment cause a significant increase in oxidative stress, ICAM-1, and monocyte adhesivity to HUVEC. Using an antisense approach, we show that ketone induced increases in ROS, ICAM-1 expression, and monocyte adhesion in endothelial cells were prevented in NOX4 knockdown cells.

This study reports that elevated levels of ketones upregulate NOX, contributing to increased oxidative stress, ICAM-1 levels, and cellular dysfunction. This provides a novel biochemical mechanism that elucidates the role of hyperketonemia in the excess cellular injury in T1D. New drugs targeting inhibition of NOX seems promising in preventing higher risk of complications associated with T1D.

Hyperketonemia is a pathological condition observed in patients with type 1 diabetes and ketosis-prone diabetes (KPD), which results in increased blood levels of acetoacetate (AA) and β-hydroxybutyrate (BHB). Frequent episodes of hyperketonemia are associated with a higher incidence of vascular disease. We examined the hypothesis that hyperketonemia activates the nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways that regulate intercellular adhesion molecule 1 (ICAM-1) expression in endothelial cells.

Human umbilical vein endothelial cells (HUVECs) were cultured with AA (0-8 mM) or BHB (0-10 mM) for 0-24 hr. Western blotting was used to determine NF-κB activation in whole-cell lysates. ICAM-1 expression was measured using flow cytometry.

RESULTS show a 2.4-fold increase in NF-κB activation in cells treated with 8 mM AA compared to the control. BHB had little or no effect on NF-κB activation. Pretreatment with a reactive oxygen species (ROS) inhibitor [N-acetyl-l-cysteine (NAC)] reduced NF-κB to near-control levels. The expression of AA-induced ICAM-1 was significantly reduced when cells were pretreated with either NAC or p38 MAPK inhibitor.

These results suggest that NF-κB and p38 MAPK mediate upregulation of ICAM-1 expression in endothelial cells exposed to elevated levels of AA, which may contribute to the development of vascular disease in diabetes.

Convincing evidence exists for the early onset of diabetic cardiomyopathy and coronary artery disease (CAD) as distinct forms of cardiac disease in young patients with Type 1 diabetes mellitus (T1DM) and the pre-stages of T2DM, forms of dysregulated insulin signaling. Progression of both chronic cardiac conditions is mediated by oxidative stress and low grade inflammation. This study reports the expression of monocyte chemotactic protein-1 (MCP-1) chemokine and the interleukin (IL)-1β inflammatory cytokine in two young patients with suboptimal metabolic control and fatal diabetic ketoacidosis (DKA), two age-matched overweight/obesity cases and two age-matched controls. In addition, markers of oxidative stress, apoptosis, collagen deposition and cardiomyocyte hypertrophy were studied. Significant expression of MCP-1 and IL-1β was seen in the myocardia of the T1DM/DKA cases, with lesser amounts expressed in the overweight/obesity myocardia. All of the other markers except cardiomyocyte hypertrophy were expressed to a significantly greater extent in the T1DM/DKA and overweight/obesity cases in comparison to the age-matched controls. Cardiomyocyte hypertrophy was significantly greater in the overweight/obesity cases than in the T1DM/DKA or the control cases.

Metabolic syndrome is a condition characterized by a clustering of risk factors for cardiovascular disease. Emerging data suggest vascular integrity is disrupted in metabolic syndrome. Vascular integrity may be determined using several measurements, including pulse wave velocity, augmentation index, and flow-mediated dilation. Arterial stiffness has become an important clinical indicator of cardiovascular disease risk. Several circulating inflammatory peptides also impact vascular integrity. The present review examines the efficacy of nutritional interventions aimed at improving vascular integrity and reducing levels of associated inflammatory peptides in individuals with metabolic syndrome, with a specific focus on the effect of dietary macronutrient redistribution on these factors.

Frequent episodes of hyperketonemia are associated with a higher incidence of vascular disease. The objective of this study was to examine the hypothesis that hyperketonemia increases monocyte-endothelial cell (EC) adhesion and the development of vascular disease in diabetes. Human U937 and THP-1 monocyte cell lines and human umbilical vein endothelial cells (HUVECs) were cultured with acetoacetate (AA) (0-10 mM) or β-hydroxybutyrate (BHB) (0-10 mM) for 24 h prior to evaluating adhesion and adhesion molecule expression. The results demonstrate a significant (P < 0.01) increase in both U937 and THP-1 adhesion to HUVEC monolayers treated with 4 mM AA compared with control. Equal concentrations of BHB resulted in similar increases in monocyte-EC adhesion. Similarly, treatments of AA or BHB to isolated monocytes from human blood also show increases in adhesion to endothelial cells. intercellular adhesion molecule-1 (ICAM-1) was significantly increased on the surface of HUVECs and an increase in total protein expression with AA treatment compared with control. The expression level of lymphocyte function-associated antigen-1 (LFA-1) was increased in monocytes treated with AA, and LFA-1 affinity was altered from low to high affinity following treatment with both AA and BHB. Monocyte adhesion could be blocked when cells were preincubated with an antibody to ICAM-1 or LFA-1. Results also show a significant increase in IL-8 and MCP-1 secretion in monocytes and HUVECs treated with 0-10 mM AA. These results suggest that hyperketonemia can induce monocyte adhesion to endothelial cells and that it is mediated via increased ICAM-1 expression in endothelial cells and increased expression and affinity of LFA-1 in monocytes.

Diabetic ketoacidosis is a metabolic disorder caused by insulin deficiency and is the most important cause of mortality and morbidity in children with Type 1 diabetes mellitus. Acute neurological complications related to diabetic ketoacidosis include cerebral oedema, cerebral infarction, brain herniation, cortical venous thrombosis and cerebral haemorrhage. Cerebral infarction is rare in juvenile diabetic ketoacidosis.

We report a girl with a newly diagnosed Type 1 diabetes mellitus who presented with diabetic ketoacidosis and developed cerebral infarction with transient visual loss.

Our findings emphasize the importance of prompt evaluation and proper management of intracranial crises in diabetic ketoacidosis.

Oxidative stress is implicated as a pathogenic factor in a spectrum of chronic diseases, notably, neurodegenerative disease. Noteworthy in this regard is that type 1 diabetes mellitus (T1DM) results in oxidative stress, leading to systemic complications of T1DM. We hypothesized that oxidative stress associated with diabetic ketoacidosis (DKA) of T1DM might have measurable brain sequelae. Consistent with this hypothesis are neurohistology and neuroradiologic studies of T1DM that suggest oxidative insults are involved in the chronic complications of diabetic encephalopathy. To further address the role of oxidative stress in an acute setting, specifically in fatal brain edema (BE) associated with DKA, we studied neuronal localization and levels of oxidative stress markers reported to be increased in other neurodegenerative conditions. We demonstrated increased levels of 8-hydroxyguanosine (8OHG), 4-hydroxynonenal (HNE), and heme oxygenase-1 (HO-1) in the pyramidal neurons of the hippocampus of DKA BE in comparison to controls. However, in the cerebellum, only 8OHG was increased in the Purkinje cells and other cells of the molecular layer. These results indicate a role for oxidative stress in the pathogenesis of T1DM encephalopathy.

Altered cellular mitochondrial membrane potential (MMP) has been implicated in the increased insulin resistance and the risk for diabetes. Hyperketonemia is increasingly being identified in type 2 diabetic patients in addition to those with type 1 diabetes. No previous study has examined the effect of hyperketonemia and trivalent chromium on cellular mitochondrial membrane potential (MMP) in any cell type. Using a U937 monocyte cell culture model, this study examined the hypothesis that hyperketonemia decreases and trivalent chromium normalizes the cellular MMP level. Cells were cultured with control and ketone bodies [acetoacetate (AA), β-hydroxybutyrate (BHB)] in the absence or the presence (0.5-100 μM) of Cr(3+) at 37°C for 24 h. The MMP was determined using DiOC6 and flow cytometry. The results show a significant decrease in MMP in cells treated with AA, but not in the cells treated with BHB. The effect of AA on cellular MMP was prevented in chromium (III)-pretreated cells. Thus, hyperketonemia decreases the MMP, and supplementation with chromium (III) normalizes altered MMP, which may play a role in the improvement in glucose metabolism seen after chromium (III) supplementation in some studies with diabetic animals and patients.

Hyperglycaemia (HG), in stroke patients, is associated with worse neurological outcome by compromising endothelial cell function and the blood-brain barrier (BBB) integrity. We have studied the contribution of HG-mediated generation of oxidative stress to these pathologies and examined whether antioxidants as well as normalization of glucose levels following hyperglycaemic insult reverse these phenomena.

Human brain microvascular endothelial cell (HBMEC) and human astrocyte co-cultures were used to simulate the human BBB. The integrity of the BBB was measured by transendothelial electrical resistance using STX electrodes and an EVOM resistance meter, while enzyme activities were measured by specific spectrophotometric assays.

After 5 days of hyperglycaemic insult, there was a significant increase in BBB permeability that was reversed by glucose normalization. Co-treatment of cells with HG and a number of antioxidants including vitamin C, free radical scavengers and antioxidant enzymes including catalase and superoxide dismutase mimetics attenuated the detrimental effects of HG. Inhibition of p38 mitogen-activated protein kinase (p38MAPK) and protein kinase C but not phosphoinositide 3 kinase (PI3 kinase) also reversed HG-induced BBB hyperpermeability. In HBMEC, HG enhanced pro-oxidant (NAD(P)H oxidase) enzyme activity and expression that were normalized by reverting to normoglycaemia.

HG impairs brain microvascular endothelial function through involvements of oxidative stress and several signal transduction pathways.

Obesity is a cardiovascular risk factor associated with endothelial dysfunction, but the effect of different weight loss strategies on endothelial function is not known. The effect of diet on endothelial function in two hypocaloric diets, a very-low-carbohydrate diet (A) and a Mediterranean diet (M), was measured by brachial artery flow-mediated dilation (FMD). Design Using a longitudinal, randomized, open study design, subjects were engaged in a 2-month weight loss diet. FMD, inflammatory cytokines [interleukin-6 (IL-6) and tumour necrosis factor-alpha] and a marker of oxidative stress [8-iso-prostaglandin F2alpha (8-iso-PGF2alpha)] were measured in subjects on three occasions: before initiating the diet (T0), after 5-7 days of dieting (T5) and after 2 months of dieting (T60). The very short- and medium-term time points were established to discriminate respectively the effect of the diet itself (T5) from that of weight loss (T60). Twenty overweight/obese but otherwise healthy women (BMI: 27-34.9 kg m(-2); age 30-50 years) completed the study. Results Group A lost more weight (mean +/- SEM; -7.6 +/- 0.8 kg) than group M (-4.9 +/- 0.6 kg, P = 0.014) at T60. The FMD was not significantly different between the two groups at T0 (group A: 12.2 +/- 2.9% vs. group B: 10.3 +/- 2.3%, P = ns). In group A, FMD was significantly reduced at T5 and returned to baseline at T60; in group M, FMD increased at T5 and returned to baseline at T60 (P = 0.007 for diet x time interaction). Serum concentrations of IL-6 and 8-iso-PGF2alpha were not significantly different between the two groups at T0 and increased significantly at T5 only in group A (P < 0.001 and P < 0.005 respectively). Conclusion As endothelial dysfunction is known to be associated with acute cardiovascular events, this study suggests that the cardiovascular risk might be increased in the first days of a very-low-carbohydrate diet.

Cerebral edema is the most common neurologic complication of diabetic ketoacidosis in children. A minority of young patients with intracerebral crises in diabetic ketoacidosis present with cerebrovascular accidents. We report 2 adolescent patients with diabetic ketoacidosis who presented with coma and diffuse white matter hemorrhages in the absence of either cerebral edema or cerebrovascular accidents. These 2 cases illustrate a novel clinical and neuropathologic description of diffuse white matter hemorrhages, possibly related to a cytotoxic process as the underlying mechanism. These case descriptions emphasize that pediatric patients with diabetic ketoacidosis and coma can present with pathology not related to either cerebral edema or cerebrovascular accidents.

Elevated blood levels of the proinflammatory cytokines interleukin-6 (IL-6), interleukin-8 (IL-8), and MCP-1 (monocyte chemoattractant protein-1) increase insulin resistance and the risk of cardiovascular disease (CVD). There is no previous study that has examined the effect of ketosis and trivalent chromium on IL-6, IL-8, or MCP-1 secretion in any cell type or in human or animal model. The authors examined the hypothesis that ketosis increases and trivalent chromium decreases the levels of cytokines and oxidative stress in diabetes using a U937 monocyte cell culture model. Cells were cultured with control, high glucose (HG), and acetoacetate (AA) in the absence or presence (0.5-10 microM) of CrCl(3), chromium picolinate (Cr-P), or chromium niacinate (Cr-N) at 37 degrees C for 24 h. The data show a significant stimulation of IL-6, IL-8, and MCP-1 secretion and an increase in oxidative stress in cells treated with HG or AA. The effect of HG on cytokine secretion was reduced by Cr-N, and to a lesser extent by CrCl(3) and Cr-P. The effect of HG on oxidative stress was reduced by Cr-N and CrCl 3, but not by Cr-P. Similarly, Cr-N decreased the cytokine secretion in HG + AA-treated cells. Cr-N significantly decreased standard oxidant (H(2)O(2)) induced cytokine secretion, which suggests that reduction of cytokine secretion by Cr-N is in part mediated by its antioxidative effect. In a cell culture model, Cr-N appears to be the most effective form of chromium in inhibiting oxidative stress and proinflammatory cytokine secretion by monocytes. This study suggests that chromium niacinate supplementation may be useful in reducing vascular inflammation and the risk of CVD in diabetes.

A systemic inflammatory response (SIR) occurs prior to and during the treatment of severe diabetic ketoacidosis (DKA). IL-1beta, TNF-alpha and C5b-9 are components of SIR and have been speculated to be involved in the clinical brain edema (BE) of DKA. We studied IL-1beta, TNF-alpha, C5b-9, inducible nitric oxide (iNOS), ICAM-1, IL-10 and Hsp70 expression in the brains of two patients who died as the result of clinical BE during the treatment of DKA. IL-1beta was strongly expressed in the choroid plexus epithelium (CPE) and ependyma, and to a lesser extent in the hippocampus, caudate, white matter radiation of the pons, molecular layer of the cerebellum and neurons of the cortical gray matter. TNF-alpha was expressed to a lesser extent than IL-1beta, and only in the CP. C5b-9, previously shown to be deposited on neurons and oligodendrocytes, was found on CPE and ependymal cells. iNOS and ICAM-1 had increased expression in the CPE and ependyma. Hsp70 and IL-10 were also expressed in the CPE of the case with the shorter duration of treatment. Our data demonstrate the presence of a multifaceted neuroinflammatory cytotoxic insult of the CPE, which may play a role in the pathophysiology of the fatal brain edema of DKA.

The long-term complications of diabetes are the leading causes of morbidity and mortality in the type 1 diabetic population and remain a major public health issue. Hyperglycemia is one of the major risk factors in the development of vascular complications. A growing body of evidence indicates that hyperglycemia leads to increased oxidative stress and monocyte and endothelial cell dysfunction. In addition to hyperglycemia, type 1 diabetic patients frequently experience ketosis (hyperketonemia). The blood concentration of ketone bodies reaches higher than 25mM in diabetics with severe ketosis. Traditionally, clinical practice has considered hypertketonemia to be present only in type 1 diabetic patients. Newer data indicate that diabetic ketoaciosis or hyperketonemia co-exists with hyperglycemia among older type 2 diabetic patients and in African Americans and other minority groups with type 2 diabetes. This review will focus on the role of hyperketonemia in the etiology of oxidative stress in diabetic patients. The data presented here illustrate that the ketone body acetoacetate (AA) can generate superoxide radicals and cause increases in oxidative stress and cellular dysfunction. The data included in this review demonstrate that blood levels of markers of oxidative stress are elevated in hyperketonemic patients compared with those of normoketonemic diabetic patients. Thus, both in vitro and in vivo research indicate that ketosis can generate oxygen radicals and result in excess cellular oxidative stress in type 1 diabetic patients. Elevated oxidative stress levels in ketotic patients can play a significant role in the development of vascular inflammation and contribute to the increased incidence of vascular disease and complications associated with type 1 diabetes.

The prevalence of diabetes mellitus makes the occurrence of hyperglycemic emergencies a key component in clinical practice. The expert nurse is well positioned to manage both diabetic ketoacidosis and hyperosmolar hyperglycemic states. Patient care management includes a high index of suspicion for awareness for the possibility of diabetic ketoacidosis or hyperosmolar hyperglycemic states in patients based on a multifactorial etiology, evidence-based treatment of the emergent episode, and tertiary prevention to prevent recurrent episodes.

Recent studies support the presence of an inflammatory response during the treatment of diabetic ketoacidosis (DKA). The objectives of this study were to monitor the complement activation products C3a, C4a, Bb, and C5b-9 prior to, during, and after correction of DKA. All patients had increased levels of C3a at 6-8 h and 24 h (P<0.05). C4a was increased in only one patient. Bb showed an upward trend at 6-8 h, and was significantly elevated at 24 h (P<0.05); sC5b-9 was elevated in all patients prior to treatment or in the first 6-8 h of treatment. Results indicate that the alternative pathway may be the primary pathway of activation. These results extend the observation that both DKA and its treatment produce varying degrees of immunologic stress during the time when acute complications are most likely to occur.

Increased permeability of the cerebral microvasculature occurs during the treatment of diabetic ketoacidosis (DKA). Microvascular changes consistent with diabetic retinopathy have been reported prior to and after the treatment of DKA. This study evaluated the structural and functional aspects of the retina immediately following the correction of DKA.

Seven young patients had comprehensive ophthalmologic examinations, including fluorescein angiography, within 24 h after the correction of severe DKA (pH <7.2).

None of the patients had clinical, photographic, or angiographic evidence of a retinal abnormality.

The blood-retinal barrier (BRB) does not experience the same degree of perturbation as the blood-brain barrier (BBB) does and may be a protected site during the insult of DKA and its treatment. The greater stability of the retinal microvasculature may be due to the increased number of pericytes in the BRB in comparison with the BBB.

Cerebral edema is the most significant complication in children with diabetic ketoacidosis (DKA). Our goal was to study whether subclinical cerebral edema was preferentially vasogenic or cytotoxic. Magnetic resonance imaging (MRI)--diffusion-weighted imaging (DWI) and T2 relaxometry (T2R)--were obtained in pediatric patients presenting with severe diabetic ketoacidosis (DKA) 6-12 hours after initial DKA treatment and stabilization and 96 hours after correction of DKA. T2 relaxometry was significantly increased during treatment in both white and gray matter, in comparison to the absolute T2R values 96 hours after correction of DKA (p = .034). Classic intracellular cytotoxic edema could not be detected, based on the lack of a statistically significant decrease in ADC values. ADC values were instead elevated, implying a large component of cell membrane water diffusion, correlating with the elevated white and gray matter T2R We discuss the findings in relation to cerebral blood volume, cerebral vasoregulatory dysfunction, and cerebral hyperemia.

Diabetes is characterized by elevated levels of ketone bodies acetoacetate (AA) and 3-hydroxybutyrate (3HB). High levels of ketone bodies have been implicated in generation of cellular oxidative stress. Ketone body activation of cellular signaling pathways associated with oxidative stress, however, has not been established. Thus, ketone body effects on kinase activation in primary cultured rat hepatocytes have been examined. Treatment with AA increased the phosphorylation of extracellular signal-regulated kinase 1/2 (Erk1/2) and p38 mitogen-activated protein kinase (MAPK), maximally by approximately 2.5- and 4-fold, respectively. AA failed to activate c-Jun NH(2)-terminal kinase. AA-mediated Erk1/2 and p38 MAPK activation was detectable at 3 h post-treatment with maximal activation occurring at 12 h. In contrast, 3HB failed to activate any of these kinases. Elevated phosphorylation of Raf and MKK3/6 also occurred in response to AA. Bisindolylmaleimide, a generalized protein kinase C (PKC) inhibitor, and B581, a Ras farnesylation inhibitor, inhibited AA-mediated activation of Erk1/2 and p38 MAPK, suggesting a role for PKC and Ras in mediating such activation. Interestingly, the tyrosine kinase inhibitor genistein prevented the AA-mediated phosphorylation of Erk1/2, but not p38 MAPK. AA treatment resulted in the generation of reactive oxygen species (ROS) and the depletion of cellular glutathione levels, which was ameliorated by the antioxidants N-Acetyl-l-cysteine (NAC) and Trolox (6-hydroxy-2,5,7,8-tetramethyl-chroman-2-carboxylic acid). NAC and Trolox also ameliorated AA-mediated Erk1/2 and p38 MAPK activation, suggesting that this activation is associated with ROS and oxidative stress.

Cerebral edema during diabetic ketoacidosis (DKA) has been attributed to osmotic cellular swelling during treatment. We evaluated cerebral water distribution and cerebral perfusion during DKA treatment in children.

We imaged 14 children during DKA treatment and after recovery, using both diffusion and perfusion weighted magnetic resonance imaging (MRI). We assessed the apparent diffusion coefficients (ADCs) and measures reflecting cerebral perfusion.

The ADC was significantly elevated during DKA treatment (indicating increased water diffusion) in all regions except the occipital gray matter. Mean reductions in the ADC from initial to postrecovery MRI were: basal ganglia 4.7 +/- 2.5 x 10(-5) mm(2)/s (P=.002), thalamus 3.7 +/- 2.8 x 10(-5) mm(2)/s, (P=.002), periaqueductal gray matter 4.3 +/- 5.1 x 10(-5) mm(2)/s (P=.03), and frontal white matter 2.0 +/- 3.1 x 10(-5) mm(2)/s (P=.03). In contrast, the ADC in the occipital gray matter increased significantly from the initial to postrecovery MRI (mean increase 3.9 +/- 3.9 x 10(-5) mm(2)/s, P=.004). Perfusion MRI during DKA treatment revealed significantly shorter mean transit times (MTTs) and higher peak tracer concentrations, possibly indicating increased cerebral blood flow (CBF).

Elevated ADC values during DKA treatment suggests a vasogenic process as the predominant mechanism of edema formation rather than osmotic cellular swelling.

The authors examined the effects of high ketone body and glucose concentrations on endothelial cell (EC) function in perfused rabbit lungs. beta-Hydroxybutyrate (beta OHB), at 5 mM, decreased endothelial angiotensin-converting enzyme (eACE) activity, whereas 25 mM glucose (HG), 1 mM beta OHB, or 10 mM acetoacetate (AcAc) did not. Dry to wet weight ratios were also reduced in lungs perfused with 5 mM beta OHB, but not with AcAc. beta OHB, at 5 mM, caused massive hemorrhage and interstitial and alveolar neutrophil infiltration; AcAc only produced engorgement of septal capillaries. Thus, pulmonary EC dysfunction occurs in rabbit lungs acutely perfused with beta OHB, but not with AcAc or glucose.

The objectives of this study were to monitor plasma cytokines as markers of cellular activation and as potential markers for the progression of the acute complications of diabetic ketoacidosis (DKA). Blood samples were obtained prior to, during and after treatment of severe DKA (pH < 7.2) in six children and adolescents. Plasma IL-10, IL-1beta, TNF-alpha, IL-6, IL-8 and IL-2 cytokine levels were assayed by ELISA at each of the time points. Prior to treatment, elevations of multiple cytokines were found, the highest being IL-10. Treatment of DKA resulted in a significant decrease of IL-10 at 6-8 h (p = 0.0062), and further increases in the inflammatory cytokines at 6-8 h and/or 24 h vs 120 h (baseline): IL-1beta (p =.0048); TNF-alpha (p =.0188) and IL-8 (p =.0048). This study strengthens the hypothesis that the metabolic crisis of DKA, and its treatment, have differential effects on cellular activation and cytokine release. The time frame for the increase in inflammatory cytokines correlates with the reported progression of subclinical brain edema, interstitial pulmonary edema and the development of clinical brain edema.