Mitochondrial Alpha-Keto Acid Dehydrogenase Complexes: Recent Developments on Structure and Function in Health and Disease.

The present work delves into the enigmatic world of mitochondrial alpha-keto acid dehydrogenase complexes discussing their metabolic significance, enzymatic operation, moonlighting activities, and pathological relevance with links to underlying structural features. This ubiquitous family of related but diverse multienzyme complexes is involved in carbohydrate metabolism (pyruvate dehydrogenase complex), the citric acid cycle (α-ketoglutarate dehydrogenase complex), and amino acid catabolism (branched-chain α-keto acid dehydrogenase complex, α-ketoadipate dehydrogenase complex); the complexes all function at strategic points and also participate in regulation in these metabolic pathways. These systems are among the largest multienzyme complexes with at times more than 100 protein chains and weights ranging up to ~10 million Daltons. Our chapter offers a wealth of up-to-date information on these multienzyme complexes for a comprehensive understanding of their significance in health and disease.

Structural and Biochemical Investigation of Selected Pathogenic Mutants of the Human Dihydrolipoamide Dehydrogenase.

Clinically relevant disease-causing variants of the human dihydrolipoamide dehydrogenase (hLADH, hE3), a common component of the mitochondrial α-keto acid dehydrogenase complexes, were characterized using a multipronged approach to unravel the molecular pathomechanisms that underlie hLADH deficiency. The G101del and M326V substitutions both reduced the protein stability and triggered the disassembly of the functional/obligate hLADH homodimer and significant FAD losses, which altogether eventually manifested in a virtually undetectable catalytic activity in both cases. The I12T-hLADH variant proved also to be quite unstable, but managed to retain the dimeric enzyme form; the LADH activity, both in the forward and reverse catalytic directions and the affinity for the prosthetic group FAD were both significantly compromised. None of the above three variants lent themselves to an in-depth structural analysis via X-ray crystallography due to inherent protein instability. Crystal structures at 2.89 and 2.44 Å resolutions were determined for the I318T- and I358T-hLADH variants, respectively; structure analysis revealed minor conformational perturbations, which correlated well with the residual LADH activities, in both cases. For the dimer interface variants G426E-, I445M-, and R447G-hLADH, enzyme activities and FAD loss were determined and compared against the previously published structural data.

Lipoamide dehydrogenase (LADH) deficiency: medical perspectives of the structural and functional characterization of LADH and its pathogenic variants.

(Dihydro)lipoamide dehydrogenase (LADH) deficiency is an autosomal recessive genetic metabolic disorder. It generally presents with an onset in the neonatal age and premature death. The clinical picture usually involves metabolic decompensation and lactic acidosis that lead to neurological, cardiological, and/or hepatological outcomes. Severity of the disease is due to the fact that LADH is a common E3 subunit to the pyruvate, alpha-ketoglutarate, alpha-ketoadipate, and branched-chain alpha-keto acid dehydrogenase complexes and is also part of the glycine cleavage system; hence, a loss in LADH activity adversely affects several central metabolic pathways simultaneously. The severe clinical manifestations, however, often do not parallel the LADH activity loss, which implies the existence of auxiliary pathological pathways; stimulated reactive oxygen species (ROS) production as well as dissociation from the relevant multienzyme complexes proved to be auxiliary exacerbating pathomechanisms for selected disease-causing LADH mutations. This review provides an overview on the therapeutic challenges of inherited metabolic diseases, structural and functional characteristics of the mitochondrial alpha-keto acid dehydrogenase complexes, molecular pathogenesis and structural basis of LADH deficiency, and relevant potential future medical perspectives.

Effects of Chronic Caffeine Consumption on Synaptic Function, Metabolism and Adenosine Modulation in Different Brain Areas.

Adenosine receptors mainly control synaptic function, and excessive activation of adenosine receptors may worsen the onset of many neurological disorders. Accordingly, the regular intake of moderate doses of caffeine antagonizes adenosine receptors and affords robust neuroprotection. Although caffeine intake alters brain functional connectivity and multi-omics analyses indicate that caffeine intake modifies synaptic and metabolic processes, it is unclear how caffeine intake affects behavior, synaptic plasticity and its modulation by adenosine. We now report that male mice drinking caffeinated water (0.3 g/L) for 2 weeks were behaviorally indistinguishable (locomotion, mood, memory) from control mice (drinking water) and displayed superimposable synaptic plasticity (long-term potentiation) in different brain areas (hippocampus, prefrontal cortex, amygdala). Moreover, there was a general preservation of the efficiency of adenosine A1 and A2A receptors to control synaptic transmission and plasticity, although there was a tendency for lower levels of endogenous adenosine ensuring A1 receptor-mediated inhibition. In spite of similar behavioral and neurophysiological function, caffeine intake increased the energy charge and redox state of cortical synaptosomes. This increased metabolic competence likely involved a putative increase in the glycolytic rate in synapses and a prospective greater astrocyte-synapse lactate shuttling. It was concluded that caffeine intake does not trigger evident alterations of behavior or of synaptic plasticity but increases the metabolic competence of synapses, which might be related with the previously described better ability of animals consuming caffeine to cope with deleterious stimuli triggering brain dysfunction.

Investigation of Circulating MicroRNA Levels in Antibody-Mediated Rejection After Kidney Transplantation.

BACKGROUND: One of the most important possible complications determining long-term graft survival after kidney transplant is antibody-mediated rejection (ABMR). The criterion standard approach to recognize ABMR is currently the kidney biopsy with histopathologic analysis. However, this test has limitations because of difficulties in timing of sampling, the evaluability of histology because of the questionable representativeness of specimens, and the limited number of this intervention. Hence, new reliable, noninvasive biomarkers are required to detect the development of ABMR in time. METHODS: In this study, we analyzed the clinical data of 45 kidney transplant patients (mean age of 44.51 years, 20 male and 25 female subjects). These participants were recruited into 5 subcohorts based on their clinical status, histologic findings, and level of donor-specific anti-HLA antibodies. Circulating microRNAs (miR-21, miR-181b, miR-146a, miR-223, miR-155, miR-150) in plasma samples were quantified by quantitative polymerase chain reaction and their levels were correlated with the clinical characteristics in different subgroups. RESULTS: The relative expression of plasma miR-155 (P = .0003), miR-223 (P = .0316), and miR-21 (P = .0147) were significantly higher in patients who had subsequent histology-approved ABMR with donor-specific anti-HLA antibody positivity (n = 10) than in the "triple negative" group (n = 21), and miR-155 showed the highest sensitivity (90%) and specificity (81%) to indicate ABMR development based on receiver operating characteristic analysis. CONCLUSIONS: According to our preliminary data, plasma miR-155, miR-21, and miR-223 can indicate the development of ABMR after kidney transplant in correlation with classic clinical parameters. However, future studies with larger number of participants are necessary to further evaluate the diagnostic properties of blood miRNAs in prediction of this life-threatening condition.

Population genetic features of calving interval of the Limousin beef cattle breed in Hungary.

Variance, covariance components, heritability, breeding values (BV) and genetic trends in calving interval (CI) of the Limousin population in Hungary were evaluated. A total of 3,008 CI data of 779 cows from three herds in 1996-2016 were processed. For influencing effects GLM method, for population genetic parameters and BV estimation BLUP animal model, for trend analyses linear regression was applied. The average CI obtained was 378.8 ± 3.1 days. The variance distribution components of the phenotype were as follow: age of cow at calving 34.30%, season of calving 26.09%, year of calving 23.00%, sire 7.45%, herd 3.23%, sex of calf 0.33% and type of calving 0.30%. The heritability of CI proved to be low (h2 d = 0.04 ± 0.02 and 0.03 ± 0.02; h2 m = 0.01 ± 0.02). The repeatability was low (R = 0.03 ± 0.02). Based on the phenotypic trend calculation, the CI of cows decreased by an average of 0.60 days per year (R 2 = 0.19; P < 0.05). In case of genetic trend calculation, the average BV of sires in CI increased 0.07 and 0.17 days per year (R 2 = 0.23 and 0.27; P < 0.05).

Young domestic chicks spontaneously represent the absence of objects.

Absence is a notion that is usually captured by language-related concepts like zero or negation. Whether nonlinguistic creatures encode similar thoughts is an open question, as everyday behavior marked by absence (of food, of social partners) can be explained solely by expecting presence somewhere else. We investigated 8-day-old chicks' looking behavior in response to events violating expectations about the presence or absence of an object. We found different behavioral responses to violations of presence and absence, suggesting distinct underlying mechanisms. Importantly, chicks displayed an avian signature of novelty detection to violations of absence, namely a sex-dependent left-eye-bias. Follow-up experiments excluded accounts that would explain this bias by perceptual mismatch or by representing the object at different locations. These results suggest that the ability to spontaneously form representations about the absence of objects likely belongs to the initial cognitive repertoire of vertebrate species.

Current Approaches in Molecular Enzymology.

Enzymes are the main executioners of living organisms [...].

Generation of Human Neural Progenitors from Blood Samples by Interrupted Reprogramming.

Human neuronal cell cultures are essential tools for biological and preclinical studies of our nervous system. Since we have very limited access to primary human neural samples, derivation of proliferative neural progenitor cells (NPCs) from cells harvested by minimally invasive sampling is a key issue. Here we describe a "shortcut" method to establish proliferative NPC cultures directly from peripheral blood mononuclear cells (PBMCs) via interrupted reprogramming. In addition, we provide procedures to characterize the NPC stage.

Convergence of adenosine and GABA signaling for synapse stabilization during development.

During development, neural circuit formation requires the stabilization of active γ-aminobutyric acid­mediated (GABAergic) synapses and the elimination of inactive ones. Here, we demonstrate that, although the activation of postsynaptic GABA type A receptors (GABAARs) and adenosine A2A receptors (A2ARs) stabilizes GABAergic synapses, only A2AR activation is sufficient. Both GABAAR- and A2AR-dependent signaling pathways act synergistically to produce adenosine 3',5'-monophosphate through the recruitment of the calcium­calmodulin­adenylyl cyclase pathway. Protein kinase A, thus activated, phosphorylates gephyrin on serine residue 303, which is required for GABAAR stabilization. Finally, the stabilization of pre- and postsynaptic GABAergic elements involves the interaction between gephyrin and the synaptogenic membrane protein Slitrk3. We propose that A2ARs act as detectors of active GABAergic synapses releasing GABA, adenosine triphosphate, and adenosine to regulate their fate toward stabilization or elimination.

Pharmacological Modulation of Neurite Outgrowth in Human Neural Progenitor Cells by Inhibiting Non-muscle Myosin II.

Studies on neural development and neuronal regeneration after injury are mainly based on animal models. The establishment of pluripotent stem cell (PSC) technology, however, opened new perspectives for better understanding these processes in human models by providing unlimited cell source for hard-to-obtain human tissues. Here, we aimed at identifying the molecular factors that confine and modulate an early step of neural regeneration, the formation of neurites in human neural progenitor cells (NPCs). Enhanced green fluorescent protein (eGFP) was stably expressed in NPCs differentiated from human embryonic and induced PSC lines, and the neurite outgrowth was investigated under normal and injury-related conditions using a high-content screening system. We found that inhibitors of the non-muscle myosin II (NMII), blebbistatin and its novel, non-toxic derivatives, initiated extensive neurite outgrowth in human NPCs. The extracellular matrix components strongly influenced the rate of neurite formation but NMII inhibitors were able to override the inhibitory effect of a restrictive environment. Non-additive stimulatory effect on neurite generation was also detected by the inhibition of Rho-associated, coiled-coil-containing protein kinase 1 (ROCK1), the upstream regulator of NMII. In contrast, inhibition of c-Jun N-terminal kinases (JNKs) had only a negligible effect, suggesting that the ROCK1 signal is dominantly manifested by actomyosin activity. In addition to providing a reliable cell-based in vitro model for identifying intrinsic mechanisms and environmental factors responsible for impeded axonal regeneration in humans, our results demonstrate that NMII and ROCK1 are important pharmacological targets for the augmentation of neural regeneration at the progenitor level. These studies may open novel perspectives for development of more effective pharmacological treatments and cell therapies for various neurodegenerative disorders.

Early postoperative effects of kidney transplantation on the cardiovascular system in our clinical practice / A vesetranszplantáció korai posztoperatív hatásai a szív- és érrendszeri betegségekre klinikai gyakorlatunkban.

Összefoglaló. Bevezetés: Mind a dializált, mind a veseátültetett betegek körében vezeto haláloknak számít a cardiovascularis megbetegedés. E mögött foképp bal kamrai hypertrophia, volumenterheltség, következményes szívritmuszavar, szívbillentyu-elégtelenség, fokozott atherosclerosis állhat. Célkituzés: Célunk a vesetranszplantáció hatásának vizsgálata a bal kamra pumpafunkciójára, a szívritmuszavarokat kiváltó és meghatározó tényezokre és a vitiumokra nézve. Módszerek: A 2014. december 20. és 2018. június 21. közti idointervallumban, a Debreceni Egyetem Szervtranszplantációs Tanszékén felnott betegeken végzett veseátültetéseket vizsgáltuk retrospektív analízissel (n = 184). Vesetranszplantációt megelozoen, illetve azt követoen 6 és 12 hónappal az echokardiográfiás, a laboratóriumi és a gyógyszeres terápiás értékeket tanulmányoztuk. A statisztikai elemzéseket khi-négyzet-próbával, Fisher-féle egzakt teszttel és Kruskal-Wallis-féle varianciaanalízissel (ANOVA) végeztük (szignifikancia: p<0,05). Eredmények: A bal kamra végsystolés tágassága az átültetés elott 34,67 mm volt, míg a 6 hónapos eredmény 31,82 mm, a 12 hónapos 32,68 mm volt (p = 0,01). Átültetés elott a stroke prevalenciája 7,87% volt, míg a beavatkozás után nem fordult elo szélütés (p<0,001). Transzplantáció hatására a bal pitvari átméro (43,68 mm; 41,59 mm; 41,00 mm; p = 0,0417) és a káliumszint (4,98 mmol/l; 4,49 mmol/l; 4,49 mmol/l; p = 0,01) szignifikáns változást igazolt. Mutét elott II. fokú mitralis regurgitatiót észleltünk 10,7%-nál, mely 4,3%-ra, majd 2,1%-ra csökkent (p = 0,03). Transzplantációt megelozoen a billentyumeszesedés elofordulása diabetesesek között 45% (p = 0,20), 6 hónap múlva 46,7% (p = 0,018), 12 hónap múlva 60,0% (p = 0,024) volt. Következtetés: Transzplantáció után a bal pitvari átméro, a végsystolés bal kamrai átméro regrediál, csökken a pitvari ritmuszavarok kialakulásának gyakorisága. A mitralis regurgitatio közepesen súlyos fokainál szignifikáns javulást, a diabeteses populáción belül szignifikáns emelkedést tapasztaltunk a meszes billentyuk számát tekintve. Orv Hetil. 2021; 162(26): 1052-1062. INTRODUCTION: Among the population suffering from end-stage renal failure and the population after kidney transplantation, the leading reason of death is cardiovascular triggered by left ventricular hypertrophy, volume overload, consecutive arrhythmias, valvular insufficiency and increased artherosclerosis. OBJECTIVE: This study was aimed at examining the effect of kidney transplantation on pump function of the left ventricle, arrhythmic substrates and valvular heart diseases. METHODS: At the Division of Organ Transplantation, University of Debrecen, we carried out a retrospective data analysis of adult patients (n = 184) who had kidney transplantation in the period between December 2014 and June 2018. Preoperatively and, then, postoperatively (at 6 and 12 months) we studied the echocardiographic parameters, the laboratory results. Statistical analyses were performed using the chi-square/Fisher's exact test and Kruskal-Wallis analysis of variance (ANOVA) test. The results were regarded significant if p<0.05 was found. RESULTS: Preoperatively the end-systolic diameter of the left ventricle was 34.67 mm, whereas 6 and 12 months later these values were 31.82 mm and 32.68 mm (p = 0.01). The prevalence of stroke was 7.87% preoperatively; there was no stroke detected postoperatively (p<0.001). The impact of transplantation on the left atrial diameter (43.68 mm; 41.59 mm; 41.00 mm; p = 0.04) and seral potassium level (4.98 mmol/l; 4.49 mmol/l; 4.49 mmol/l; p<0.01) showed significant improvement. Before transplantation, grade 2 mitral regurgitation was observed in 10.7% of the patients, whereas it reduced to 4.3%, then to 2.1% 6 and 12 months postoperatively (p = 0.03). Preoperative valvular calcification was detected in 45% of the diabetic study population (p = 0.20), 6 and 12 months later, in 46.7% (p = 0.018) and 60.0% (p = 0.024). CONCLUSION: After transplantation, the left atrial and the end-systolic diameter of the left ventricle regrediated, decreasing the frequency of arrhythmic episodes. The number of the middle grade mitral valve regurgitation decreased and the calcification among diabetic population increased significantly. Orv Hetil. 2021; 162(26): 1052-1062.

Effects of Kidney Transplantation on Valvular Heart Diseases.

INTRODUCTION: In patients with end-stage renal failure, hypervolemia frequently causes increased cardiac output, especially in patients who are under-dialyzed and those with cardiac decompensation. OBJECTIVE: This study aimed to examined the effect of kidney transplantation on valvular heart diseases. PATIENTS AND METHODS: This retrospective data analysis included adult patients (n = 180) who underwent kidney transplantation between February 2015 and June 2018 at the Division of Organ Transplantation, University of Debrecen, Hungary. This study examined the echocardiographic parameters and laboratory results preoperatively and postoperatively (at 6 and 12 months). Statistical analyses were performed using the χ2/Fisher exact tests and Kruskal-Wallis analysis of variance test. P < .05 was considered significant. RESULTS: No mitral regurgitation (MR) was observed preoperatively in 27% of the patients, while 62% had grade 1 MR, and 11% had grade 2 MR. Grade 2 MR was reduced from 11% to 2% twelve months after kidney transplantation (P = .03). Valvular calcification was detected preoperatively in 21.5% of the study population but was detected in 25.8% 6 months postoperation and in 35.5% 12 months postoperation (P = .09). At 12-month follow-up, 30.8% of patients without diabetes and 60% (P = .03) of patients with diabetes had valvular calcification. CONCLUSION: Significant improvement was noted in patients with moderate-stage MR because renal transplantations decrease the volume overload on the heart. After surgical intervention, elevation in the incidence of calcified valves among patients with diabetes was significant compared to patients without diabetes.

Structure of the dihydrolipoamide succinyltransferase (E2) component of the human alpha-ketoglutarate dehydrogenase complex (hKGDHc) revealed by cryo-EM and cross-linking mass spectrometry: Implications for the overall hKGDHc structure.

BACKGROUND: The human mitochondrial alpha-ketoglutarate dehydrogenase complex (hKGDHc) converts KG to succinyl-CoA and NADH. Malfunction of and reactive oxygen species generation by the hKGDHc as well as its E1-E2 subcomplex are implicated in neurodegenerative disorders, ischemia-reperfusion injury, E3-deficiency and cancers. METHODS: We performed cryo-EM, cross-linking mass spectrometry (CL-MS) and molecular modeling analyses to determine the structure of the E2 component of the hKGDHc (hE2k); hE2k transfers a succinyl group to CoA and forms the structural core of hKGDHc. We also assessed the overall structure of the hKGDHc by negative-stain EM and modeling. RESULTS: We report the 2.9 Šresolution cryo-EM structure of the hE2k component. The cryo-EM map comprises density for hE2k residues 151-386 - the entire (inner) core catalytic domain plus a few additional residues -, while residues 1-150 are not observed due to the inherent flexibility of the N-terminal region. The structure of the latter segment was also determined by CL-MS and homology modeling. Negative-stain EM on in vitro assembled hKGDHc and previous data were used to build a putative overall structural model of the hKGDHc. CONCLUSIONS: The E2 core of the hKGDHc is composed of 24 hE2k chains organized in octahedral (8 × 3 type) assembly. Each lipoyl domain is oriented towards the core domain of an adjacent chain in the hE2k homotrimer. hE1k and hE3 are most likely tethered at the edges and faces, respectively, of the cubic hE2k assembly. GENERAL SIGNIFICANCE: The revealed structural information will support the future pharmacologically targeting of the hKGDHc.

Generation of multiple iPSC clones from a male schizophrenia patient carrying de novo mutations in genes KHSRP, LRRC7, and KIR2DL1, and his parents.

Here we describe the generation of induced pluripotent stem cell lines from each member - male proband, mother, father - of a schizophrenia case-parent trio that participated in an exome sequencing study, and 3 de novo mutations were identified in the proband. Peripheral blood mononuclear cells were obtained from all three individuals and reprogrammed using Sendai virus particles carrying the Yamanaka transgenes. These 3 iPSC lines (iPSC-SZ-HU-MO 1, iPSC-SZ-HU-FA 1, and iPSC-SZ-HU-PROB 1) represent a resource for examining the functional significance of the identified de novo mutations in the molecular pathophysiology of schizophrenia.

Functional Comparison of Blood-Derived Human Neural Progenitor Cells.

Induced pluripotent stem cell (iPSC)-derived neural progenitor cells (NPCs) are promising tools to model complex neurological or psychiatric diseases, including schizophrenia. Multiple studies have compared patient-derived and healthy control NPCs derived from iPSCs in order to investigate cellular phenotypes of this disease, although the establishment, stabilization, and directed differentiation of iPSC lines are rather expensive and time-demanding. However, interrupted reprogramming by omitting the stabilization of iPSCs may allow for the generation of a plastic stage of the cells and thus provide a shortcut to derive NPSCs directly from tissue samples. Here, we demonstrate a method to generate shortcut NPCs (sNPCs) from blood mononuclear cells and present a detailed comparison of these sNPCs with NPCs obtained from the same blood samples through stable iPSC clones and a subsequent neural differentiation (classical NPCs-cNPCs). Peripheral blood cells were obtained from a schizophrenia patient and his two healthy parents (a case-parent trio), while a further umbilical cord blood sample was obtained from the cord of a healthy new-born. The expression of stage-specific markers in sNPCs and cNPCs were compared both at the protein and RNA levels. We also performed functional tests to investigate Wnt and glutamate signaling and the oxidative stress, as these pathways have been suggested to play important roles in the pathophysiology of schizophrenia. We found similar responses in the two types of NPCs, suggesting that the shortcut procedure provides sNPCs, allowing an efficient screening of disease-related phenotypes.

Investigation of de novo mutations in a schizophrenia case-parent trio by induced pluripotent stem cell-based in vitro disease modeling: convergence of schizophrenia- and autism-related cellular phenotypes.

BACKGROUND: De novo mutations (DNMs) have been implicated in the etiology of schizophrenia (SZ), a chronic debilitating psychiatric disorder characterized by hallucinations, delusions, cognitive dysfunction, and decreased community functioning. Several DNMs have been identified by examining SZ cases and their unaffected parents; however, in most cases, the biological significance of these mutations remains elusive. To overcome this limitation, we have developed an approach of using induced pluripotent stem cell (iPSC) lines from each member of a SZ case-parent trio, in order to investigate the effects of DNMs in cellular progenies of interest, particularly in dentate gyrus neuronal progenitors. METHODS: We identified a male SZ patient characterized by early disease onset and negative symptoms, who is a carrier of 3 non-synonymous DNMs in genes LRRC7, KHSRP, and KIR2DL1. iPSC lines were generated from his and his parents' peripheral blood mononuclear cells using Sendai virus-based reprogramming and differentiated into neuronal progenitor cells (NPCs) and hippocampal dentate gyrus granule cells. We used RNASeq to explore transcriptomic differences and calcium (Ca2+) imaging, cell proliferation, migration, oxidative stress, and mitochondrial assays to characterize the investigated NPC lines. RESULTS: NPCs derived from the SZ patient exhibited transcriptomic differences related to Wnt signaling, neuronal differentiation, axonal guidance and synaptic function, and decreased Ca2+ reactivity to glutamate. Moreover, we could observe increased cellular proliferation and alterations in mitochondrial quantity and morphology. CONCLUSIONS: The approach of reprograming case-parent trios represents an opportunity for investigating the molecular effects of disease-causing mutations and comparing these in cell lines with reduced variation in genetic background. Our results are indicative of a partial overlap between schizophrenia and autism-related phenotypes in the investigated family. LIMITATIONS: Our study investigated only one family; therefore, the generalizability of findings is limited. We could not derive iPSCs from two other siblings to test for possible genetic effects in the family that are not driven by DNMs. The transcriptomic and functional assays were limited to the NPC stage, although these variables should also be investigated at the mature neuronal stage.

Generation of iPSC lines from peripheral blood mononuclear cells of identical twins both suffering from type 2 diabetes mellitus and one of them additionally diagnosed with atherosclerosis.

Here we describe the generation of induced pluripotent stem cell (iPSC) lines from peripheral blood samples of identical twin sisters with type 2 diabetes mellitus (DM2). Two clonal lines from each patient (HU-DM2-A-1, HU-DM2-A-2 and HU-DM2-B-1, HU-DM2-B-2) were established via Sendai viral reprograming of peripheral blood mononuclear cells, and characterized to confirm pluripotency and genetic integrity. The established iPSC lines can help to investigate DM2 related cellular phenotypes and provide a model system for drug testing.

[Changes of lactate levels in diabetic ketoacidosis and in newly diagnosed type 1 diabetes mellitus]. / Laktátszintváltozások diabeteses ketoacidosisban és frissen diagnosztizált 1-es típusú diabetes mellitusban.

Introduction: It is known that lactate concentration is increased in diabetic ketoacidosis (DKA), however, the pathophysiology and kinetics of lactate changes are still unclear. Normally, L-lactate is the major form in the human body. According to previous data, also D- and L-lactate might be increased in hyperglycaemic disorders. Aim: We aimed to describe the kinetics and mechanisms of lactate concentration changes in ketoacidosis and newly diagnosed diabetes. Method: We performed a prospective study, including 5-18-year-old children with ketoacidosis (DKA, n = 13) and with newly diagnosed type 1 diabetes without ketoacidosis (T1DM, n = 6). We performed routine blood gas analysis 0-12-24-48 hours after admission, which also measured L-lactate levels. We also determined total venous serum lactate level by gas chromatography-mass spectrometry. Results: Initial plasma lactate concentration was increased in ketoacidosis as compared to the newly diagnosed diabetes group (p<0.05). After 12 h of rehydration, lactate levels were greatly reduced in ketoacidotic patients but after 24-48 h it was repeatedly increased (all p<0.01). In the 0-12 h phase, total serum lactate level was higher than L-lactate level, referring to D-lactate production. Conclusion: We described two L-lactate peaks in ketoacidosis. In the first 12 hours anaerobic glycolysis seems to have major role in hyperlactataemia. We assume that stimulated aerobic glycolysis leads to the second lactate peak. However, D-lactate is not routinely measured, it may contribute to the initial hyperlactataemia in both groups and is comparable to L-lactate production in ketoacidosis. Orv Hetil. 2019; 160(45): 1784-1790.

Discovery and development of extreme selective inhibitors of the ITD and D835Y mutant FLT3 kinases.

Aberrant activation of FMS-like tyrosine receptor kinase 3 (FLT3) is implicated in the pathogenesis of acute myeloid leukemia (AML) in 20-30% of patients. In this study we identified a highly selective (phenylethenyl)quinazoline compound family as novel potent inhibitors of the FLT3-ITD and FLT3-D835Y kinases. Their prominent effects were confirmed by biochemical and cellular proliferation assays followed by mice xenograft studies. Our modelling experiments and the chemical structures of the compounds predict the possibility of covalent inhibition. The most effective compounds triggered apoptosis in FLT3-ITD AML cells but had either weak or no effect in FLT3-independent leukemic and non-leukemic cell lines. Our results strongly suggest that our compounds may become therapeutics in relapsing and refractory AML disease harboring various ITD and tyrosine kinase domain mutations, by their ability to overcome drug resistance.