Az intraluminális hidrogén peroxid elimináció mechanizmusai az endoplazmás retikulumban = Mechanisms of intraluminal hydrogen peroxide elimination in the endoplasmic reticulum

Az endoplazmás retikulum (ER) oxidatív protein foldingjának luminális H2O2 termelése hozzájárul az organellum oxidatív környezetének kialakulásához. Kimutattuk, hogy az Ero1α, az oxidatív hajtogatás és ER redox homeosztázis egyik legfontosabb szabályozója, feldúsul a MAM frakcióban, és szabályozza a Ca2+ áramokat. Az Ero1α szintjének mind növelésével mind csökkentésével módosítani lehetett az ER Ca2+ fluxusokat, amely feltárja a fehérje kulcsfontosságú szerepét a korai szekréciós kompartimentumban. Azt is megfigyeltük, hogy a máj ER luminális H2O2 szint emelkedése in vivo egerekben a mikroszómális GSH és fehérje tiol tartalom csökkenését, valamint luminális oxidoreduktázok redox állapotának eltolódását eredményezte. Az oxidatív hatás kiváltotta ar ER tágulását, mely redukálószerekkel kivédhető volt. ER-be célzott katalázt overexpresszáló, antitest termelő sejtekben az érett antitest polimerek csökkent szekrécióját, míg az antitest prekurzor monomerek/dimerek intracelluláris felhalmozódását észleltük. Az eredmények szerint a helyi H2O2 termelés elősegíti, míg a H2O2 eltávolítása rontja a diszulfidok kialakulását. Három review-t közöltünk az ER redox viszonyairól. Egy tanulmányban az oxidatív protein foldingra új paradigmát javasoltunk: a több oxidáns hipotézist. Két átfogó review-ban a jelenlegi ismereteket foglaltuk össze az ER legfontosabb redox rendszereiről. Másik két cikkben pedig a kompartimentáció jelentőségét alátámasztó eredményeket tárgyaltuk. | Oxidative protein folding in the endoplasmic reticulum (ER) results in luminal H2O2 production, contributing to the formation of the oxidative environment of the organelle. We showed that Ero1α, a key controller of oxidative folding and ER redox homeostasis, is enriched in mitochondrial-associated ER membranes (MAM) and regulates Ca2+ fluxes. Either increasing or decreasing the levels of Ero1α affected Ca2+ fluxes, which reveals a pivotal role for this oxidase in the early secretory compartment. We also observed that the elevation of hepatic ER luminal H2O2 levels of mice in vivo resulted in a decrease in microsomal GSH and protein-thiol contents and in a redox shift of certain luminal oxidoreductases. The oxidative wave was accompanied by reversible dilation of ER, prevented by concomitant reducing treatment. ER targeted catalase overexpressing antibody producing cells showed diminished secretion of mature antibody polymers, while incomplete antibody monomers/dimers were accumulated and/or secreted. The results indicate that local H2O2 production promotes, while quenching of H2O2 impairs disulfide formation. We published three reviews on the redox conditions in the ER. In a Hypothesis paper we proposed a new paradigm for the oxidative folding: the multiple oxidant hypothesis. In two comprehensive reviews we summarized the present knowledge on the major redox systems in the ER. We summarized the facts showing the importance of compartmentation in two other reviews

Kaposvár 61-es út 2. lelőhely Árpád-kori embertani anyagának paleopatológiai vizsgálata

Palaopathological investigation of an Árpádian Age series from Kaposvár road 61, Site 2. The skeletal remains of 202 individuals, 54 males, 60 females, 88 subadults and adults of undeterminable sex were analysed in an Árpádian Age series from Kaposvár Road 61, Site 2. The life expectancy at birth in the studied population was 22.7 years, the sex ratio was balanced. Enthesopathies (E), degenerative joint diseases (DJD) and Schmorl-hernia (SH) were the most common pathological alterations. The pattern of E, DJD and SH prevalence by sex and age was analysed. There was a significant difference in the frequencies of the E on the patellae between males and females, and between adultus and maturus age goups. Sex differences could be explained by the heavier physical load of the knee joints in males. A considerable difference was observed between the adultus and maturus age groups also in the prevalence of spondylarthrosis and arthrosis deformans.The reason of this fenomenon can be explained by aging and the deterioration of joint structures. Significant sexual differences were found in the prevalence of spondylosis, arthrosis deformans and spondylodiscitis, as well as spondylarthrosis. This observation could be explained by the sexual differences in the physical loads of individuals, with the exception of spondylodiscitis. Approximately one third of specimens affected by periostitis. These results, together with the high incidence of endocranial lesion, suggest the possibility of high prevalence of infectious diseases in the studied community. Hematologic disorders, particularly poroticus hyperostosis (PH) were of high prevalence in the sample. PH was present in higher rate among young females than young males. The fast growth of these individuals, menstruation, pregnancy, giving birth and also breastfeeding could have been the possible causes of this fact. Several rare diseases were also recorded; an adult male suffered from bone tuberculosis, an adult female from leprosy. An osteoblastic metastatic bone cancer of a mature female was observed as well

Novel Crizotinib–GnRH conjugates revealed the significance of lysosomal trapping in GnRH-based drug delivery systems

Several promising anti-cancer drug–GnRH (gonadotropin-releasing hormone) conjugates have been developed in the last two decades, although none of them have been approved for clinical use yet. Crizotinib is an effective multi-target kinase inhibitor, approved against anaplastic lymphoma kinase (ALK)- or ROS proto-oncogene 1 (ROS-1)-positive non-small cell lung carcinoma (NSCLC); however, its application is accompanied by serious side effects. In order to deliver crizotinib selectively into the tumor cells, we synthesized novel crizotinib analogues and conjugated them to a [d-Lys6]–GnRH-I targeting peptide. Our most prominent crizotinib–GnRH conjugates, the amide-bond-containing [d-Lys6(crizotinib*)]–GnRH-I and the ester-bond-containing [d-Lys6(MJ55*)]–GnRH-I, were able to bind to GnRH-receptor (GnRHR) and exert a potent c-Met kinase inhibitory effect. The efficacy of compounds was tested on the MET-amplified and GnRHR-expressing EBC-1 NSCLC cells. In vitro pharmacological profiling led to the conclusion that that crizotinib–GnRH conjugates are transported directly into lysosomes, where the membrane permeability of crizotinib is diminished. As a consequence of GnRHR-mediated endocytosis, GnRH-conjugated crizotinib bypasses its molecular targets—the ATP-binding site of RTKs— and is sequestered in the lysosomes. These results explained the lower efficacy of crizotinib–GnRH conjugates in EBC-1 cells, and led to the conclusion that drug escape from the lysosomes is a major challenge in the development of clinically relevant anti-cancer drug–GnRH conjugates

Decreased Nuclear Ascorbate Accumulation Accompanied with Altered Genomic Methylation Pattern in Fibroblasts from Arterial Tortuosity Syndrome Patients

Ascorbate requiring Fe2+/2-oxoglutarate-dependent dioxygenases located in the nucleoplasm have been shown to participate in epigenetic regulation of gene expression via histone and DNA demethylation. Transport of dehydroascorbic acid is impaired in the endomembranes of fibroblasts from arterial tortuosity syndrome (ATS) patients, due to the mutation in the gene coding for glucose transporter GLUT10. We hypothesized that altered nuclear ascorbate concentration might be present in ATS fibroblasts, affecting dioxygenase activity and DNA demethylation. Therefore, our aim was to characterize the subcellular distribution of vitamin C, the global and site-specific changes in 5-methylcytosine and 5-hydroxymethylcytosine levels, and the effect of ascorbate supplementation in control and ATS fibroblast cultures. Diminished nuclear accumulation of ascorbate was found in ATS fibroblasts upon ascorbate or dehydroascorbic acid addition. Analyzing DNA samples of cultured fibroblasts from controls and ATS patients, a lower global 5-hydroxymethylcytosine level was found in ATS fibroblasts, which could not be significantly modified by ascorbate addition. Investigation of the (hydroxy)methylation status of specific regions in six candidate genes related to ascorbate metabolism and function showed that ascorbate addition could stimulate hydroxymethylation and active DNA demethylation at the PPAR-gamma gene region in control fibroblasts only. The altered DNA hydroxymethylation patterns in patient cells both at the global level and at specific gene regions accompanied with decreased nuclear accumulation of ascorbate suggests the epigenetic role of vitamin C in the pathomechanism of ATS. The present findings represent the first example for the role of vitamin C transport in epigenetic regulation suggesting that ATS is a compartmentalization disease

Subcellular Localization of Thioredoxin/Thioredoxin Reductase System—A Missing Link in Endoplasmic Reticulum Redox Balance

The lumen of the endoplasmic reticulum (ER) is usually considered an oxidative environment; however, oxidized thiol-disulfides and reduced pyridine nucleotides occur there parallelly, indicating that the ER lumen lacks components which connect the two systems. Here, we investigated the luminal presence of the thioredoxin (Trx)/thioredoxin reductase (TrxR) proteins, capable of linking the protein thiol and pyridine nucleotide pools in different compartments. It was shown that specific activity of TrxR in the ER is undetectable, whereas higher activities were measured in the cytoplasm and mitochondria. None of the Trx/TrxR isoforms were expressed in the ER by Western blot analysis. Co-localization studies of various isoforms of Trx and TrxR with ER marker Grp94 by immunofluorescent analysis further confirmed their absence from the lumen. The probability of luminal localization of each isoform was also predicted to be very low by several in silico analysis tools. ER-targeted transient transfection of HeLa cells with Trx1 and TrxR1 significantly decreased cell viability and induced apoptotic cell death. In conclusion, the absence of this electron transfer chain may explain the uncoupling of the redox systems in the ER lumen, allowing parallel presence of a reduced pyridine nucleotide and a probably oxidized protein pool necessary for cellular viability

Glucose transporter type 10 - lacking in arterial tortuosity syndrome - facilitates dehydroascorbic acid transport

Loss-of-function mutations in the gene encoding GLUT10 are responsible for arterial tortuosity syndrome (ATS), a rare connective tissue disorder. In the present study GLUT10 mediated dehydroascorbic acid (DAA) transport was investigated, supposing its involvement in the pathomechanism. GLUT10 protein produced by in vitro translation and incorporated into liposomes efficiently transported DAA. Silencing of GLUT10 decreased DAA transport in immortalized human fibroblasts whose plasma membrane was selectively permeabilized. Similarly, the transport of DAA through endomembranes was markedly reduced in fibroblasts from ATS patients. Re-expression of GLUT10 in patients' fibroblasts restored DAA transport activity. The present results demonstrate that GLUT10 is a DAA transporter and DAA transport is diminished in the endomembranes of fibroblasts from ATS patients. This article is protected by copyright. All rights reserved