Fatal Ketoacidosis Research Articles

Renal Tubular Epithelial Subnuclear Vacuolization in Hypothermia and Diabetic Ketoacidosis-A Retrospective Autopsy Study.

Subnuclear vacuolization of the renal tubular epithelium refers to discrete lipid vacuoles displacing the nuclei toward the lumen. This phenomenon has been associated with conditions sharing fatal ketoacidosis as a common denominator. This retrospective study aimed to investigate renal tubular epithelial subnuclear vacuolization and other postmortem examination findings in fatal hypothermia and diabetic ketoacidosis (DKA) cases.Fourteen cases with hypothermia and 19 cases with DKA were included. More cases with DKA had focal or diffuse subnuclear vacuolization compared with hypothermia cases (89% vs 43%; P = 0.007). In 6 cases with DKA, formalin pigment was detected within subnuclear vacuoles, whereas no case with hypothermia had formalin pigment deposition. Comparative analyses of hypothermia and DKA cases revealed further differences: Vitreous beta-hydroxybutyrate was higher in the DKA group compared with the hypothermia group (P = 0.044), whereas blood ethanol concentrations were higher in the latter (P = 0.008). Hypothermia cases were older compared with the DKA cases (P = 0.022).When all cases were included in the statistical analysis, cases with subnuclear vacuolization had higher vitreous beta-hydroxybutyrate and blood ethanol concentrations (P = 0.029 and 0.023, respectively). The findings corroborate the results of previous studies suggesting a link between subnuclear vacuolization and increased levels of ketoacidosis.

Роль суммарных РНК лимфоидных и стволовых клеток в коррекции уровня глюкозы в крови при экспериментальном сахарном диабете

Цель - оценка влияния препаратов суммарной РНК лимфоидных клеток на уровень глюкозы в крови при экспериментальном аллоксановом диабете. Методика. Работа выполнена на 68 белых беспородных крысах обоего пола массой 180-220 г. Для моделирования аллоксанового сахарного диабета 1 типа животным однократно подкожно вводили полный адъювант Фрейнда в дозе 0,5 мл/крысу. Через 24 ч (на фоне 24-часового голодания, при свободном доступе к воде) однократно подкожно вводили препарат аллоксан тригидрат («La Chema», Чехия) в дозе 200 мг/кг (4% раствор в 0,9% NaCl). Для предотвращения фатального кетоацидоза, начиная с 3-х сут после введения аллоксана, все крысы получали базисную инсулинотерапию. В экспериментах использовано 8 разновидностей препаратов суммарной РНК: из лимфоидных клеток селезенки, из костного мозга и хвостовой части поджелудочной железы интактных крыс (РНКс-1, РНКкм-1 и РНКп/ж, соответственно); из лимфоидных клеток селезенки крыс, подвергнутых кровопусканию в объеме 2% от массы тела за 17 ч до выделения РНК (РНКс-2 с индуцированной повышенной морфогенетической активностью); из лимфоидных клеток селезенки и тимуса крыс, подвергнутых кровопусканию в объеме 2% от массы тела за 96 ч до выделения РНК (РНКс-3 и РНКт-3, соответственно, с высокой ингибирующей морфогенез активностью); из лимфоцитов периферической крови здорового человека (РНКлпкч), а также из стволовых клеток стромы пупочного канатика человека. Результаты. Показана возможность восстановления функции инсулярного аппарата и стойкой нормализации уровня глюкозы в крови крыс с экспериментальным аллоксановым диабетом 1 типа. Установлено наличие корригирующей способности аллогенных и ксеногенных препаратов суммарной РНК, выделенной из лимфоидных органов крыс и лимфоцитов периферической крови человека, а также из мезенхимных стромальных клеток пуповины человека, в отношении уровня глюкозы крови у крыс. Продемонстрирован различный вклад препаратов суммарной РНК, полученной из разных лимфоидных органов и стволовых клеток, в восстановлении нормального уровня глюкозы в крови животных. Установлено, что указанные препараты действуют на разные ткани-мишени в процессе восстановления функции инсулярного аппарата. Заключение. Полученные данные свидетельствуют о принципиальной возможности эффективного лечения сахарного диабета 1 и 2 типов. Доказана эффективность неинвазивного интраназального введения аллогенных и ксеногенных препаратов суммарной РНК лимфоидных и стволовых клеток. Полученные результаты ставят вопрос о необходимости разработки более адекватной экспериментальной модели, а также о перспективности поиска подходов к персонифицированному индивидуальному лечению этой патологии с учетом особенностей ее развития в каждом конкретном случае. Aim. To study the effect of total RNA from lymphoid cells on blood glucose in experimental alloxan diabetes mellitus (DM). Methods. The study was conducted on 68 white mongrel rats of both sexes weighing 180-220 g. Alloxan DM was modeled with full Freund’s adjuvant (0.5 ml/rat). After 24 h of fasting with free access to water, a single dose of alloxan trihydrate (La Chemas, Czeck Republic) was injected subcutaneously (200 mg/kg as a 4% solution in saline). All rats received basis insulin therapy starting from day 3 of the alloxan injection to prevent fatal ketoacidosis. Eight types of total RNA were used: from splenic lymphoid cells, bone marrow, and caudal part of the pancreas of intact rats; from splenic lymphoid cells of rats after blood withdrawal (2% of body weight 17 h prior to RNA isolation) (RNA after induction of increased morphogenetic activity); from splenic and thymic lymphoid cells of rats after blood withdrawal (2% of body weight 96 h prior to RNA isolation) (RNAs with high morphogenesis-inhibiting activity); and from peripheral blood lymphocytes and umbilical cord stromal stem cells of a healthy human. Results. The study showed a possibility for functional recovery of the insular apparatus and stable normalization of blood glucose level in rats with experimental alloxan DM, a model of clinical type 1 DM. Allogenic and xenogeneic total RNAs isolated from rat lymphoid organs, peripheral blood lymphocytes, and mesenchymal stromal cells of human umbilical cord were able to correct the blood glucose level in rats. Total RNAs isolated from different lymphoid organs and stem cells differently contributed to normalization of blood glucose in rats. These total RNAs were shown to influence different target tissues during restoration of the insular apparatus function. Conclusion. The study showed a principle possibility of effective therapy for types 1 and 2 DM and demonstrated the efficacy of non-invasive intranasal administration of allogenic and xenogeneic total RNAs from lymphoid and stem cells. These results indicated a need for developing a more relevant experimental model and offered a promising outlook for individualized treatment of this disease with due account of its peculiar features in each specific case. The study was conducted as a part of the Program for Basic Research of State Academies of Science in 2013-2020 on Development of Cell Models for Molecular Processes in Organs and Tissue (V.N. Orekhovich Research Institute of Biomedical Chemistry) and the complex Research Work #01201354257 on Regulation of Hemopoietic and Nonhemopoietic Functions of Bone Marrow Cells in Experiment and Clinic (Sounth Ural State Medical University).

Basal Vacuolization in Renal Tubular Epithelial Cells at Autopsy and Their Relation to Ketoacidosis.

Basal vacuolization of renal tubular epithelial cells is a useful postmortem marker for ketoacidosis. To investigate its incidence and relationship to the severity of ketoacidosis, 158 autopsy cases with elevated β-hydroxybutyrate (>1 mmol/L) over a 7-year-period were retrospectively reviewed. Sixty-eight cases (43%) exhibited basal vacuolizations (vitreous β-hydroxybutyrate: 1.16-29.35 mmol/L, mean 10.28 mmol/L), and 90 cases (57%) did not (vitreous β-hydroxybutyrate: 1.03-13.7 mmol/L, mean 2.84 mmol/L). Quantitative analysis revealed on average a fourfold elevation in β-hydroxybutyrate in cases with basal vacuolizations compared to those without; 10.3% of cases with β-hydroxybutyrate concentrations between 1.01 and 2.00 mmol/L had basal vacuolizations, and this incidence increased to 33.3% with concentrations between 4.01 and 6.00 mmol/L. A marked increase in incidence to >70% was observed with concentrations >6.00 mmol/L, and basal vacuoles were invariably present (100%) with concentrations >14.01 mmol/L. This study demonstrates that basal vacuolizations are a sensitive marker for significant ketoacidosis and reaffirms its use as an indicator for likely cases of fatal ketoacidosis at autopsy.

A gestational ketogenic diet alters maternal metabolic status as well as offspring physiological growth and brain structure in the neonatal mouse.

BackgroundThe use of the ketogenic diet (KD) among women of child-bearing age has been increasing, leading to increased interest in identifying the diet’s suitability during gestation. To date, no studies have thoroughly investigated the effect of a gestational KD on offspring growth. Since ketones have been reported to play a role in cerebral lipid and myelin synthesis, it is particularly important to investigate the diet’s impact on brain anatomy of the offspring.MethodsTo fill this knowledge gap we imaged CD-1 mouse neonates whose mothers were fed either a standard diet (SD) or a KD prior to and during gestation. Images were collected at postnatal (P) 11.5 and 21.5 using Magnetic Resonance Imaging (MRI). Maternal metabolic status was also tracked during lactation, by following their body weight, blood glucose, ketone, cholesterol, and triglyceride concentrations.ResultsThe KD dams exhibit a significant reduction in maternal fertility and litter size, as well as a high risk of developing fatal ketoacidosis by mid-lactation. To increase survival of the KD dams and offspring, fostering of P2.5 pups (from both KD and SD litters) by SD-foster dams was carried out. This resulted in stabilization of blood ketones of the KD dams, and aversion of the fatal ketoacidosis. We also note a slower and smaller weight loss for the KD compared with the SD dams. The average fostered KD pup exhibits retarded growth by P21.5 compared with the average fostered SD pup. An anatomical comparison of their brains further revealed significant structural differences at P11.5, and particularly at P21.5. The KD brain shows a relative bilateral decrease in the cortex, fimbria, hippocampus, corpus callosum and lateral ventricle, but a relative volumetric enlargement of the hypothalamus and medulla.ConclusionA gestational ketogenic diet deleteriously affects maternal fertility and increases susceptibility to fatal ketoacidosis during lactation. Prenatal and early postnatal exposure to a ketogenic diet also results in significant alterations to neonatal brain structure, and results in retarded physiological growth. These alterations could be accompanied by functional and behavioural changes in later postnatal life.

Olanzapine-induced hyperglycemic ketoacidosis and corresponding acetone concentrations post-mortem: A forensic interpretation

Olanzapine has been shown to cause or have a contributory role in the development of hyperglycemia and diabetes mellitus. Without careful monitoring for the development of these conditions and control of the resulting adverse effects, patients receiving olanzapine may be at risk of developing fatal ketoacidosis. A review of post-mortem toxicological reports has revealed an increase in the incidence of post-mortem findings of acetone in decedents who were taking olanzapine over the past decade. A review of the current literature and a comprehensive review of case histories and toxicological findings were conducted at the Centre of Forensic Sciences (Toronto, Ontario). Olanzapine concentrations ranging from <62.5 to 858 ng/mL and acetone concentrations as high as 95 mg/dL were detected concurrently. Due to the unstable nature of olanzapine, in several instances quantitation was not possible despite elevated responses during qualitative screening procedures. Five cases suggesting olanzapine-induced ketoacidosis were identified based on the case history and toxicological findings. These data have been compiled and examined with respect to acetone concentrations following olanzapine use and the forensic relevance of post-mortem olanzapine and acetone concentrations are discussed.

Structure of the Subunit Binding Domain and Dynamics of the Di-domain Region from the Core of Human Branched Chain α-Ketoacid Dehydrogenase Complex

The homo-24-meric dihydrolipoyl transacylase (E2) scaffold of the human branched-chain alpha-ketoacid dehydrogenase complex (BCKDC) contains the lipoyl-bearing domain (hbLBD), the subunit-binding domain (hbSBD) and the inner core domain that are linked to carry out E2 functions in substrate channeling and recognition. In this study, we employed NMR techniques to determine the structure of hbSBD and dynamics of several truncated constructs from the E2 component of the human BCKDC, including hbLBD (residues 1-84), hbSBD (residues 111-149), and a di-domain (hbDD) (residues 1-166) comprising hbLBD, hbSBD and the interdomain linker. The solution structure of hbSBD consists of two nearly parallel helices separated by a long loop, similar to the structures of the SBD isolated from other species, but it lacks the short 3(10) helix. The NMR results show that the structures of hbLBD and hbSBD in isolated forms are not altered by the presence of the interdomain linker in hbDD. The linker region is not entirely exposed to solvent, where amide resonances associated with approximately 50% of the residues are observable. However, the tethering of these two domains in hbDD significantly retards the overall rotational correlation times of hbLBD and hbSBD, changing from 5.54 ns and 5.73 ns in isolated forms to 8.37 ns and 8.85 ns in the linked hbDD, respectively. We conclude that the presence of the interdomain linker restricts the motional freedom of the hbSBD more significantly than hbLBD, and that the linker region likely exists as a soft rod rather than a flexible string in solution.

Natural Osmolyte Trimethylamine N-Oxide Corrects Assembly Defects of Mutant Branched-chain α-Ketoacid Decarboxylase in Maple Syrup Urine Disease

Maple syrup urine disease is caused by deficiency in the mitochondrial branched-chain alpha-ketoacid dehydrogenase (BCKD) complex. The clinical phenotype includes often fatal ketoacidosis, neurological derangement, and mental retardation. The type IA mutations Y393N-alpha, Y368C-alpha, and F364C-alpha, which occur in the E1alpha subunit of the decarboxylase (E1) component of the BCKD complex, impede the conversion of an alphabeta heterodimeric intermediate to a native alpha(2)beta(2) heterotetramer in the E1 assembly pathway. In the present study, we show that a natural osmolyte trimethylamine N-oxide (TMAO) at the optimal 1 m concentration restores E1 activity, up to 50% of the wild type, in the mutant E1 carrying the above missense mutations. TMAO promotes the conversion of otherwise trapped mutant heterodimers to active heterotetramers. This slow step does not involve dissociation/reassociation of the mutant heterodimers, which are preformed in the presence of chaperonins GroEL/GroES and Mg-ATP. The TMAO-stimulated mutant E1 activity is remarkably stable upon removal of the osmolyte, when cofactor thiamine pyrophosphate and the transacylase component of the BCKD complex are present. The above in vitro results offer the use of chemical chaperones such as TMAO as an approach to mitigate assembly defects caused by maple syrup urine disease mutations.

Naked plasmid-mediated gene transfer to skeletal muscle ameliorates diabetes mellitus.

The ability of tissues to take up naked plasmid DNA in vivo suggests an approach for reconstituting systemic metabolic deficiencies without the disadvantages of viral vectors and lipid-DNA complexes. Plasmid-mediated gene transfer into skeletal muscle was investigated as a means of providing a therapeutic source of insulin. Four plasmid constructs, each bearing a mouse furin cDNA transgene and rat proinsulin cDNA (modified for processing by furin) driven by four different promoters were injected into the calf muscles of male Balb/c mice. Insulin and C-peptide concentrations were measured by radio-immunoassays having minimal crossreactivity for proinsulin and partially processed proinsulin. Intramuscular insulin concentrations increased by up to 3.6-fold over controls seven days after single injections of CMV, beta-actin, hsp70 and myoglobin promoter constructs. The optimal dose for most constructs was 100 micrograms plasmid DNA. Intramuscular plasmid injection into streptozotocin-induced diabetic Balb/c mice raised plasma insulin and C-peptide concentrations, and reduced hyperglycaemia. Two injections (100 micrograms plasmid DNA each) caused higher plasma insulin concentrations and significantly reduced hyperglycemia in diabetic mice than a single injection. Best results were obtained when plasmid injections preceded induction of diabetes by 14 days. Skeletal muscle is a potentially useful platform for ectopic secretion of insulin using naked plasmid as a gene transfer vector. Injection at two sites 14 days before the onset of severe hyperglycemia is optimal. This approach could protect Type I diabetics from fatal ketoacidosis and enhance the action of agents that sensitize tissues to insulin in type II diabetes.

Hepatic insulin gene expression as treatment for type 1 diabetes mellitus in rats.

Type 1 diabetes mellitus is caused by a lack of insulin that results from the autoimmune destruction of the pancreatic beta-cells. Severe diabetes, if not controlled by periodic insulin injections, can lead to ketoacidosis and death. We have previously shown that sustained low level production of insulin in the liver of diabetic rats prevented their death from complications of diabetes. To test the hypothesis that there is a window of serum insulin concentrations that can prevent ketoacidosis without significant risk of hypoglycemia secondary to hyperinsulinemia, rats were infused with various doses of a recombinant retrovirus encoding an engineered rat preproinsulin-1 gene. The gene was engineered to allow processing into mature insulin by the protease furin. At the lower doses tested, fatal ketoacidosis was prevented, but the rats exhibited nonfasting hyperglycemia. At intermediate doses, which resulted in serum insulin concentrations of 1.6 mg/ml, the rats achieved near-normoglycemia and no serum ketones. These rats did not exhibit hypoglycemia even during a 24-h fast. At high virus doses, the animals achieved nonfasting normoglycemia but exhibited hypoglycemia during the fast. In conclusion, we have defined a therapeutic window of hepatic insulin expression that provides protection against ketoacidosis without significant risk of hypoglycemia. This window of sustained hepatic insulin expression might permit its development into a novel treatment modality for the prevention of ketoacidosis in patients with severe insulin-dependent diabetes mellitus.

Hepatic Insulin Gene Expression as Treatment for Type 1 Diabetes Mellitus in Rats

Type 1 diabetes mellitus is caused by a lack of insulin that results from the autoimmune destruction of the pancreatic β-cells. Severe diabetes, if not controlled by periodic insulin injections, can lead to ketoacidosis and death. We have previously shown that sustained low level production of insulin in the liver of diabetic rats prevented their death from complications of diabetes. To test the hypothesis that there is a window of serum insulin concentrations that can prevent ketoacidosis without significant risk of hypoglycemia secondary to hyperinsulinemia, rats were infused with various doses of a recombinant retrovirus encoding an engineered rat preproinsulin-1 gene. The gene was engineered to allow processing into mature insulin by the protease furin. At the lower doses tested, fatal ketoacidosis was prevented, but the rats exhibited nonfasting hyperglycemia. At intermediate doses, which resulted in serum insulin concentrations of 1.6 mg/ml, the rats achieved near-normoglycemia and no serum ketones. These rats did not exhibit hypoglycemia even during a 24-h fast. At high virus doses, the animals achieved nonfasting normoglycemia but exhibited hypoglycemia during the fast. In conclusion, we have defined a therapeutic window of hepatic insulin expression that provides protection against ketoacidosis without significant risk of hypoglycemia. This window of sustained hepatic insulin expression might permit its development into a novel treatment modality for the prevention of ketoacidosis in patients with severe insulin-dependent diabetes mellitus.

Diabetic syndrome in sand rats. IV. Morphologic changes in islet tissue.

Ultrastuctural and light microscopic studies of pancreatic islets in normal and diabetic sand rats are reported. Following the institution of a synthetic chow diet, beta cell degranulation and enhanced protein synthesis were observed. With the appearance of diabetes, glycogen infiltration occurred, with displacement of cellular organelles and eventual cytoplasmic degeneration and liquefaction. These alterations were correlated with blood glucose and serum immunoreactive insulin levels. — The sand rats are unique in that they are not able to cope with the increased caloric load of synthetic chow. They respond by marked insulin production; an increase that usually maintains the animal free of ketosis, occasionally returns them to a euglycemic state, and may rarely be terminated by beta cell degeneration and necrosis with fatal ketoacidosis.