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Region-dependent effects of diabetes and insulin-replacement on neuronal nitric oxide synthase-and heme oxygenase-immunoreactive submucous neurons

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World Journal of Gastroenterology 2017年第41期23卷 7359-7368页

作者： Nikolett Bódi Zita Szalai Lalitha Chandrakumar m.ria bagyánszki Department of Physiology Anatomy and Neuro-science Faculty of Science and Informatics University of Szeged

AIm To investigate the intestinal segment-specific effects of diabetes and insulin replacement on the density of different subpopulations of submucous neurons. mETHODS Ten weeks after the onset of type 1 diabetes samples were taken from the duodenum, ileum and colon of streptozotocin-induce diabetic, insulin-treated diabetic and sex-and age-matched control rats. Whole-mount preparations of submucous plexus were prepared from the different gut segments for quantitative fluorescent immunohistochemistry. The following double-immunostainings were performed: neuronal nitric oxide synthase(n NOS) and Hu C/D, heme oxygenase(HO) 1 and peripherin, as well as HO2 and peripherin. The density of n NOS-, HO1-and HO2-immunoreactive(IR) neurons was determined as a percentage of the total number of submucous neurons. RESULTS The total number of submucous neurons and the proportion of n NOS-, HO1-and HO2-IR subpopulations were not affected in the duodenal ganglia of control, diabetic and insulin-treated rats. While the total neuronal number did not change in either the ileum or the colon, the density of nitrergic neurons exhibited a 2-and 3-fold increase in the diabetic ileum and colon, respectively, which was further enhanced after insulin replacement. The presence of HO1-and HO2-IR submucous neurons was robust in the colon of controls(38.4%-50.8%), whereas it was significantly lower in the small intestinal segments(0.0%-4.2%, P m and a decrease in the colon of the proportion of HO-IR neurons in insulin-treated diabetic animals. CONCLUSION Diabetes and immediate insulin replacement induce the most pronounced region-specific alterations of n NOS-, HO1-and HO2-IR submucous neuronal density in the distal parts of the gut.

关键词： Nitrergic neurons Heme oxygenase 1 Heme oxygenase 2 Submucous neurons Gut regionspecificity Diabetes Insulin

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World Journal of Diabetes 2012年第5期3卷 80-93页

作者： m.ria bagyánszki Nikolett Bódi Department of Physiology Anatomy and Neuroscience Faculty of Science University of Szeged H-6726 Szeged Hungary

Gastric intestinal symptoms common among diabetic patients are often caused by intestinal motility abnormalities related to enteric neuropathy. It has recently been demonstrated that the nitrergic subpopulation of myenteric neurons are especially susceptible to the development of diabetic neuropathy. Additionally, different susceptibility of nitrergic neurons located in different intestinal segments to diabetic damage and their different levels of responsiveness to insulin treatment have been revealed. These findings indicate the importance of the neuronal microenvironment in the pathogenesis of diabetic nitrergic neuropathy. The main focus of this review therefore was to summarize recent advances related to the diabetes-related selective nitrergic neuropathy and associated motility disturbances. Special attention was given to the findings on capillary endothelium and enteric glial cells. Growing evidence indicates that capillary endothelium adjacent to the myenteric ganglia and enteric glial cells surrounding them are determinative in establishing the ganglionic microenvironment. Additionally, recent advances in the development of new strategies to improve glycemic control in type 1 and type 2 diabetes mellitus are also considered in this review. Finally, looking to the future, the recent and promising results of metagenomics for the characterization of the gut microbiome in health and disease such as diabetes are highlighted.

关键词： Diabetes Enteric neurons Insulin Enteric neuropathy Nitrergic neurons

Gut region-dependent alterations of nitrergic myenteric neurons after chronic alcohol consumption

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World Journal of Gastrointestinal Pathophysiology 2015年第3期6卷 51-57页

作者： m.ria bagyánszki Nikolett Bódi Department of Physiology Anatomy and NeuroscienceUniversity of Szeged

Chronic alcohol abuse damages nearly every organ in the body. The harmful effects of ethanol on thebrain, the liver and the pancreas are well documented. Although chronic alcohol consumption causes serious impairments also in the gastrointestinal tract like altered motility, mucosal damage, impaired absorption of nu-trients and inflammation, the effects of chronically consumed ethanol on the enteric nervous system are less detailed. While the nitrergic myenteric neurons play an essential role in the regulation of gastrointestinal peristalsis, it was hypothesised, that these neurons are the first targets of consumed ethanol or its metabolites generated in the different gastrointestinal segments. To reinforce this hypothesis the effects of ethanol on the gastrointestinal tract was investigated in different rodent models with quantitative immunohistochemistry, in vivo and in vitro motility measurements, western blot analysis, evaluation of nitric oxide synthase enzyme activity and bio-imaging of nitric oxide synthesis. These results suggest that chronic alcohol consumption did not result significant neural loss, but primarily impaired the nitrergic pathways in gut region-dependent way leading to disturbed gastrointestinal motility. The gut segment-specific differences in the effects of chronic alcohol consumption highlight the significance the ethanol-induced neuronal microenvironment involving oxidative stress and intestinal microbiota.

关键词： Chronic ethanol consumption Nitrergic myenteric neurons Enteric nervous system Nitric oxide synthase Gut motility disorders Intestinal microbiota

Gut region-specific TNFR expression:TNFR2 is more affected than TNFR1 in duodenal myenteric ganglia of diabetic rats

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World Journal of Diabetes 2023年第1期14卷 48-61页

作者： Bence Pál Barta Benita Onhausz Afnan AL Doghmi Zita Szalai János Balázs m.ria bagyánszki Nikolett Bódi Department of Physiology Anatomy and NeuroscienceUniversity of SzegedSzeged 6726Hungary

BACKGROUND Cytokines are essential in autoimmune inflammatory processes that accompany type 1 *** necrosis factor alpha plays a key role among others in modulating enteric neuroinflammation,however,it has a dual role in cell degeneration or survival depending on different *** general,TNFR1 is believed to trigger apoptosis,while TNFR2 promotes cell *** importance of the neuronal microenvironment has been recently highlighted in gut region-specific diabetic enteric neuropathy,however,the expression and alterations of different TNFRs in the gastrointestinal tract has not been *** To investigate the TNFR1 and TNFR2 expression in myenteric ganglia and their environment in different intestinal segments of diabetic *** Ten weeks after the onset of hyperglycemia,gut segments were taken from the duodenum,ileum and colon of streptozotocin-induced(60 mg/body weight kg i.p.)diabetic(n=17),insulin-treated diabetic(n=15)and sex-and age-matched control(n=15)*** plexus whole-mount preparations were prepared from different gut regions for TNFR1/HuCD or TNFR2/HuCD double-labeling fluorescent ***1 and TNFR2 expression was evaluated by post-embedding immunogold electron microscopy on ultrathin sections of myenteric *** levels were measured by enzyme-linked immunosorbent assay in muscle/myenteric plexus-containing(mUSCLE-mP)tissue homogenates from different gut segments and experimental *** A distinct region-dependent TNFRs expression was detected in *** density of TNFR1-labeling gold particles was lowest,while TNFR2 density was highest in duodenal ganglia and a decreased TNFRs expression from proximal to distal segments was observed in mUSCLE-mP *** diabetics,the TNFR2 density was only significantly altered in the duodenum with decrease in the ganglia(0.32±0.02 vs 0.45±0.04,PmUSCLE-mP h

关键词： Tumor necrosis factor alpha receptors myenteric ganglia Enteric neurons Neuronal environment Diabetes Insulin

Structural and molecular features of intestinal strictures in rats with Crohn's-like disease

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World Journal of Gastroenterology 2016年第22期22卷 5154-5164页

作者： Petra Talapka Anikó Berkó Lajos István Nagy Lalitha Chandrakumar m.ria bagyánszki László Géza Puskás éva Fekete Nikolett Bódi Department of Physiology Anatomy and Neuroscience Faculty of Sciences and Informatics University of Szeged H-6726 Szeged Hungary Avidin Ltd. H-6726 Szeged Hungary

AIm: To develop a new rat model we wanted to gain a better understanding of stricture formation in Crohn’s disease (CD).mETHODS: Chronic colitis was induced locally by the administration of 2,4,6-trinitrobenzenesulfonic acid (TNBS). The relapsing inflammation characteristic to CD was mimicked by repeated TNBS treatments. Animals were randomly divided into control, once, twice and three times TNBS-treated groups. Control animals received an enema of saline. Tissue samples were taken from the strictured colonic segments and also adjacent proximally and distally to its 60, 90 or 120 d after the last TNBS or saline administrations. The frequency and macroscopic extent of the strictures were measured on digital photographs. The structural features of strictured gut wall were studied by light- and electron microscopy. Inflammation related alterations in TGF-beta 2 and 3, matrix metalloproteinases 9 (mmP9) and TImP1 mRNA and protein expression were determined by quantitative real-time PCR and western blot analysis. The quantitative distribution of caspase 9 was determined by post-embedding ***: Intestinal strictures first appeared 60 d after TNBS treatments and the frequency of them increased up to day 120. From day 90 an intact lamina epithelialis, reversible thickening of lamina muscularis mucosae and irreversible thickening of the muscularis externa were demonstrated in the strictured colonic segments. Nevertheless the morphological signs of apoptosis were frequently seen and excess extracellular matrix deposition was recorded between smooth muscle cells (SmCs). Enhanced caspase 9 expression on day 90 in the SmCs and on day 120 also in myenteric neurons indicated the induction of apoptosis. The mRNA expression profile of TGF-betas after repeated TNBS doses was characteristic to CD, TGF-beta 2, but not TGF-beta 3 was up-regulated. Overexpression of mmP9 and down-regulation of TImP1 were demonstrated. The progressive increase in the amount

关键词： Crohn’ s disease Rat model TGF-beta Intestinal strictures mmP9 TImP1

Subcellular distribution of nitric oxide synthase isoforms in the rat duodenum

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World Journal of Gastroenterology 2011年第8期17卷 1026-1029页

作者： Petra Talapka Nikolett Bódi Izabella Battonyai éva Fekete m.ria bagyánszki Department of Physiology Anatomy and Neuroscience University of Szeged

AIm:To study the cell-type specific subcellular distribution of the three isoforms of nitric oxide synthase(NOS) in the rat ***:Postembedding immunoelectronmicroscopy was performed,in which primary antibodies for neuronal NOS(nNOS),endothelial NOS(eNOS),and inducible NOS(iNOS),were visualized with protein A-gold-conjugated secondary *** ultrathin sections were examined and photographed with a Philips Cm10 electron microscope equipped with a mEGAVIEW II *** specificity of the immunoreaction in all cases was assessed by omitting the primary antibodies in the labeling protocol and incubating the sections only in the protein A-gold conjugated secondary ***:Postembedding immunoelectronmicroscopy revealed the presence of nNOS,eNOS,and iNOS immunoreactivity in the myenteric neurons,the enteric smooth muscle cells,and the endothelium of capillariesrunning in the vicinity of the myenteric plexus of the rat *** cell type-specific distributions of the immunogold particles labeling the three different NOS isozymes were *** the control experiments,in which the primary antiserum was omitted,virtually no postembedding gold particles were ***:This postembedding immunoelectronmicroscopic study provided the first evidence of celltype-specific differences in the subcellular distributions of NOS isoforms.

关键词： Postembedding immunoelectronmicroscopy Subcellular distribution Neuronal nitric oxide synthase Endothelial nitric oxide synthase Inducible nitric oxide synthase

Alleviated mucosal and neuronal damage in a rat model of Crohn's disease

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World Journal of Gastroenterology 2014年第44期20卷 16690-16697页

作者： Petra Talapka Lajos István Nagy Alexandra Pál marietta Zita Poles Anikó Berkó m.ria bagyánszki László Géza Puskás éva Fekete Nikolett Bódi Department of Physiology Anatomy and Neuroscience Faculty of Sciences and Informatics University of Szeged Avidin Ltd.

AIm:To establish a rat model suitable to investigate the repetitive relapsing inflammations(RRI)characteristic to Crohn’s ***:Colitis was induced by 2,4,6-trinitrobenzenesulfonic acid(TNBS).RRI were mimicked by repeating administrations of *** samples were taken from control,once,twice and three times treated rats from the inflamed and adjacent non-inflamed colonic segments at different timepoints during the acute intestinal *** means of the ulcerated area were measured to evaluate the macroscopic mu-cosal *** density of myenteric neurons was determined on whole mounts by Hu C/Hu D *** oxygenase-1(HO-1)expression was evaluated by molecular biological ***:TNBS-treated rats displayed severe colitis,but the mortality was negligible,and an increase of body weight was characteristic throughout the experimental *** widespread loss of myenteric neurons,and marked but transient HO-1 up-regulation were demonstrated after the first TNBS *** repeated doses the length of the recovery time and extent of the ulcerous colonic segments were markedly decreased,and the neuronal loss was on a smaller scale and was limited to the inflamed ***-1 m RNA level was notably greater than after a single dose and overexpression was sustained throughout the timepoints ***,the HO-1protein up-regulation after the second TNBS treatment proved to be *** the third treatment HO-1 protein expression could not be ***:Experimentally provoked RRI may exert a protective preconditioning effect against the mucosal and neuronal *** persistent up-regulation of HO-1 m RNA expression may correlate with this.

关键词： Crohn's disease Experimental rat model Heme oxygenase-1 myenteric neurons Repetitive re-lapsing inflammation

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World Journal of Diabetes 2021年第5期12卷 658-672页

作者： Nikolett Bódi Diána mezei Payal Chakraborty Zita Szalai Bence Pál Barta János Balázs Zsolt Rázga Edit Hermesz m.ria bagyánszki Department of Physiology Anatomy and NeuroscienceFaculty of Science and InformaticsUniversity of SzegedSzeged 6726Hungary Department of Biochemistry and molecular Biology Faculty of Science and InformaticsUniversity of SzegedSzeged 6726Hungary Department of Pathology Faculty of MedicineUniversity of SzegedSzeged 6720Hungary

BACKGROUND The importance of the neuronal microenvironment has been recently highlighted in gut region-specific diabetic enteric neuropathy. Regionally distinct thickening of endothelial basement membrane(Bm) of intestinal capillaries supplying the myenteric ganglia coincide with neuronal damage in different intestinal segments. Accelerated synthesis of matrix molecules and reduced degradation of matrix components may also contribute to the imbalance of extracellular matrix dynamics resulting in Bm thickening. Among the matrix degrading proteinases, matrix metalloproteinase 9(mmP9) and its tissue inhibitor(TImP1) are essential in regulating extracellular matrix *** To evaluate the intestinal segment-specific effects of diabetes and insulin replacement on ganglionic Bm thickness, mmP9 and TImP1 *** Ten weeks after the onset of hyperglycaemia gut segments were taken from the duodenum and ileum of streptozotocin-induced diabetic, insulin-treated diabetic and sex-and age-matched control rats. The thickness of Bm surrounding myenteric ganglia was measured by electron microscopic morphometry. Wholemount preparations of myenteric plexus were prepared from the different gut regions for mmP9/TImP1 double-labelling fluorescent immunohistochemistry. Post-embedding immunogold electron microscopy was applied on ultrathin sections to evaluate the mmP9 and TImP1 expression in myenteric ganglia and their microenvironment from different gut segments and conditions. The mmP9 and TImP1 messenger ribonucleic acid(m RNA) level was measured by quantitative polymerase chain *** Ten weeks after the onset of hyperglycaemia, the ganglionic Bm was significantly thickened in the diabetic ileum, while it remained intact in the duodenum. The immediate insulin treatment prevented the diabetes-related thickening of the Bm surrounding the ileal myenteric ganglia. Quantification of particle density showed an increasing tendency for mmP9 and a decreasing tenden

关键词： Type 1 diabetes Diabetic enteric neuropathy Neuronal microenvironment Basement membrane matrix metalloproteinase 9 Tissue inhibitor of metalloproteinase 1

A step-by-step protocol for formaldehyde-assisted isolation of regulatory elements from Arabidopsis thaliana

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Journal of Integrative Plant Biology 2014年第6期56卷 527-538页

作者： mohammad Amin Omidbakhshfard Flavia Vischi Winck Samuel Arvidsson Diego m.ria?o-Pachón Bernd mueller-Roeber University of Potsdam Institute of Biochemistry and Biology Potsdam-Golm 14476 Germany max-Planck Institute of molecular Plant Physiology Potsdam-Golm 14476 Germany Brazilian Bioethanol Science and Technology Laboratory (CTBE) Brazilian Center of Research in Energy and Materials (CNPEM) Campinas Brazil

The control of gene expression by transcriptional regulators and other types of functional y relevant DNA transactions such as chromatin remodeling and replication underlie a vast spectrum of biological processes in al organisms. DNA transactions require the control ed interaction of proteins with DNA sequence motifs which are often located in nucleosome-depleted regions （NDRs） of the chromatin. Formaldehyde-assisted isolation of regulatory elements （FAIRE） has been established as an easy-to-implement method for the isolation of NDRs from a number of eukaryotic organisms, and it has been successful y employed for the discovery of new regulatory segments in genomic DNA from, for example, yeast, Drosophila, and humans. Until today, however, FAIRE has only rarely been employed in plant research and currently no detailed FAIRE protocol for plants has been published. Here, we provide a step-by-step FAIRE protocol for NDR discovery in Arabidopsis thaliana. We demonstrate that NDRs isolated from plant chromatin are readily amenable to quantitative polymerase chain reaction and next-generation sequencing. Only minor modification of the FAIRE protocol wil be needed to adapt it to other plants, thus facilitating the global inventory of regulatory regions across species.

关键词： Arabidopsis thaliana chromatin cis-regulatory elements epigenomics FAIRE-qPCR FAIRE-seq gene expression gene regulatory network transcription factor