Mária Bagyánszki

Background/Objectives: Glial cell line-derived neurotrophic factor (GDNF) and neurotrophin-3 (NT3) are essential neurotrophic factors that continue to be expressed in the mature intestine. Myenteric neurons are damaged by hyperglycemia to varying degrees in the different gut segments. Therefore, our aims were to identify the gut-segment-specific expression patterns of GDNF and NT3 in acute and chronic hyperglycemic rat models. Methods: Segments of the duodenum, ileum, and colon were dissected one week (acute experiment) or ten weeks (chronic experiment) after the streptozotocin-induced hyperglycemia. GDNF–HuC/HuD fluorescent immunohistochemistry, NT3 enzyme-linked immunosorbent assay, and NT3 immunogold electron microscopy were performed in rat myenteric ganglia. Results: The proportion of GDNF–immunoreactive myenteric neurons did not change in gut segments in acute hyperglycemia. However, during chronic diabetes, it was elevated in the colon and remained unchanged in the duodenum and ileum. Expressional alterations of NT3 were investigated in the small intestine. In duodenal myenteric ganglia, acute hyperglycemia induced a significant increase in the number of NT3-labeling gold particles, and its density was markedly increased in chronic diabetes, which was partially prevented by insulin treatment. In the ileum, NT3 density increased in acute hyperglycemic rats, while it decreased in chronic diabetic rats. Conclusions: Induction of neurotrophic factors was specific to different gut segments during acute and chronic hyperglycemia. These results suggest that increased colonic GDNF and duodenal NT3 expression may reflect a compensatory response of myenteric neurons to hyperglycemic states.

Background/Objectives: Enteric glial cells (EGCs) are key players in regulating enteric neurons and gastrointestinal functions including disturbed gut motility in diabetic patients. Enteric neuronal damage has been shown in type 1 diabetes, but EGCs’ vulnerability to hyperglycaemic insults requires more investigation. Therefore, we aimed to study the quantitative changes in the EGC network enmeshing enteric plexuses, intestinal smooth muscle and mucosa in streptozotocin-induced acute (1-week) and chronic (10-weeks) diabetic rat models. Methods: Fluorescent immunohistochemistry using Sox10 glial and HuC/HuD pan-neuronal markers, immunogold electron microscopy and ELISA were performed on different gut segments. Results: In the submucosal ganglia of the ileum and colon, the density of Sox10-immunoreactive EGCs was significantly reduced in acute and increased in chronic hyperglycaemic rats without any changes in the duodenum. In the myenteric ganglia, regionally distinct alterations of glial density were noted in acute hyperglycaemia; however, a remarkable decrease was observed in chronic animals. Alterations of neuronal density did not follow the pattern of glial changes, resulting in shifts in the glia/neuron ratio. The presence of Sox10-HuC/HuD-immunoreactive cells and their diabetes-related quantitative changes were also revealed in enteric plexuses. The density of Sox10-labelling gold particles was significantly increased in the duodenal myenteric glia of diabetic rats. Muscular EGC density increased only in the colon after acute hyperglycaemia and changed in all segments after chronic hyperglycaemia. Glial fibrillary acidic protein levels decreased in the small intestine of chronic hyperglycaemic rats. Conclusions: Our present findings reveal time-dependent and regionally distinct changes in the EGC network in response to hyperglycaemia, contributing to diabetic enteric neuropathy and gut motility disturbances.

The cerebral microcirculation undergoes dynamic changes regarding neurovascular responses through the perinatal period. CO ₂ is one of the most potent regulator of cerebral blood flow, however its effector mechanisms involve multiple possibly yet unknown pathways in the neonate. Acid-sensing ion channel-1A (ASIC1A) appears to play a decisive role in this cerebrovascular response in adult mice, however, cortical ASIC1A expression and its possible contribution to the response in the translational piglet model of term neonates is unknown. Therefore, we investigated neocortical ASIC1A expression and the effect of the specific ASIC1A-inhibitor psalmotoxin-1 on the neurovascular response to graded hypercapnia in piglets. Anesthetized, mechanically ventilated newborn pigs were equipped with a closed or open cranial window to assess the cortical blood flow (CoBF) with laser speckle contrast imaging or the neuronal activity with multichannel electrodes, respectively. Graded hypercapnia was elicited by ventilation of 5%–10% CO ₂ . ASIC1A was found to be ubiquitously expressed in neocortical neurons. Psalmotoxin-1 while unaffected the CoBF response to 5% CO ₂ , it significantly attenuated further increases in CoBF to 10% CO ₂ , accompanying the highly altered hypercapnia-induced changes in neuronal activity. We conclude that ASIC1A activation by hypercapnia-induced acidosis is partially responsible for the neurovascular response in the neonatal brain.

BGP-15, a poly(ADP-ribose) polymerase-1 (PARP-1) inhibitor exerts cardioprotective effects; however, the underlying mechanisms remain unclear. Therefore, our study aimed to investigate the effects of BGP-15 on the imatinib (Imtb)-induced cardiac inflammation at the biochemical level. Male rats were divided to control, Imtb-treated (60 mg/kg/day for 14 days), and Imtb + BGP-15-treated animals. In this group Imtb was co-administered with BGP-15 at the dose of 10 mg/kg/day. At the end of the experiment, nuclear factor-kappa B/p65 (NF-κB/p65), nuclear transcription factor erythroid-2 related factor (Nrf2), heme oxygenase-1 (HO-1), high mobility group box 1 (HMGB1), and myeloperoxidase (MPO) were measured by Western blot. Chemokine and interleukins (ILs) were determined by Legendplex. Additionally, cardiac specific changes were visualized by immunohistochemistry. We demonstrated that Imtb increased NF-κB/p65, IL-6, IL-1β, IL-18, MCP-1, HMGB1, as well as the expression and activity of MPO. Conversely, the expressions of antioxidant Nrf2 and HO-1 were decreased. Administration of BGP-15 effectively mitigated these inflammatory alterations by significantly reducing pro-inflammatory cytokines and MPO activity, while simultaneously restoring and enhancing the levels of Nrf2 and HO-1, thereby promoting antioxidant defenses. The immunohistochemical staining further supported these biochemical changes. Our study provides new and comprehensive biochemical insight for managing Imtb-induced inflammatory responses via BGP-15-induced PARP1 inhibition.

Several substances with antioxidant and anti-inflammatory properties are currently being investigated as potential adjunctive or standalone treatments for inflammatory bowel disease (IBD). One such substance is resveratrol (RES), also known as 3,5,4′-trihydroxy-trans-stilbene, a natural dietary polyphenol with diverse health-promoting effects. In this study, male Wistar–Hannover rats received oral RES supplementation at doses of 5, 10, or 20 mg/kg/day for 28 days. On day 25 colitis was induced using intracolonic administration of 2,4,6-trinitrobenzene sulphonic acid (TNBS). Based on histological and planimetric analysis, the 10 mg/kg dose significantly reduced colonic ulceration and pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-α) expression compared to the TNBS group. Immunohistochemistry also revealed that RES at this dose attenuated the intensity of TNF-α receptors, namely TNFR1 and TNFR2. Furthermore, the concentration of lipocalin-2 (Lcn-2) was significantly elevated in TNBS-induced colitis. In conclusion, our findings suggest that RES may exert its protective effects partly through the modulation of TNF receptor signaling in TNBS-induced colitis.

Enteric neurons regulating motility display regional damage to diabetes. By inhibiting neuroinflammation, insulin can contribute to neuronal survival, therefore, we aimed to investigate the presence of insulin in myenteric neurons and their nitrergic population in acute and chronic rat models of type 1 diabetes. One or ten weeks after the onset of hyperglycemia, gut segments and the pancreas of control, diabetic, and insulin-treated diabetic rats were investigated. In the controls, insulin-immunoreactive neurons comprised 8–9% of the total myenteric neuronal population in the ileum and colon and 2–4% in the duodenum. Except for the duodenum, this proportion was significantly increased in acute hyperglycemic rats and was decreased in the colon of the chronic ones. However, the proportion of insulin-immunoreactive nitrergic neurons remained unchanged in all segments in chronic hyperglycemia. Immunogold electron microscopy revealed an increased density of insulin-labelling gold particles in diabetic duodenal ganglia of the chronic experiment. Insulin mRNA was not detected in intestinal samples either in controls or diabetics. These findings support time-dependent and regional alterations in the proportion of insulin-immunoreactive myenteric neurons and their nitrergic subpopulation. Regionally different insulin content of myenteric neurons may contribute to their protection from diabetic damage.

Bacterial flagellin (flg22) induces rapid and permanent stomatal closure. However, its local and systemic as well as tissue‐ and cell‐specific effects are less understood. Our results show that flg22 induced local and systemic stomatal closure in intact tomato plants, which was regulated by reactive oxygen‐ and nitrogen species, and also affected the photosynthetic activity of guard cells but not of mesophyll cells. Interestingly, rapid and extensive local expression of Ethylene response factor 1 was observed after exposure to flg22, whereas the relative transcript levels of Defensin increased only after six hours, especially in systemic leaves. Following local and systemic ethylene emission already after one and six hours, jasmonic acid levels increased in the local leaves after six hours of flg22 treatment. Using immunohistochemical methods, significant defensin accumulation was found in the epidermis and stomata of flg22‐treated leaves and above them. Immunogold labelling revealed significant levels of defensins in the cell wall of the mesophyll parenchyma and guard cells. Furthermore, single cell qRT‐PCR confirmed that guard cells are able to synthesise defensins. It can be concluded that guard cells are not only involved in the first line of plant defense by regulating stomatal pore size, but can also defend themselves and the plant by producing and accumulating antimicrobial defensins where phytopathogens can penetrate.

Background/Objectives: Type 1 diabetes affects cytokines as potential inducers of NFκB signalling involved in inflammation and neuronal survival. Our goal was to assess the expression of NFκB p65 and its negative regulator, Nrf2, in myenteric neurons and adjacent smooth muscle of different gut segments after chronic hyperglycaemia and immediate insulin treatment. Methods: After ten weeks of hyperglycaemia, intestinal samples of control, streptozotocin-induced diabetic and insulin-treated diabetic rats were prepared for fluorescent immunohistochemistry, immunogold electron microscopy, ELISA and qPCR. Results: In the diabetic rats, the proportion of NFκB p65-immunoreactive myenteric neurons decreased significantly in the duodenum and increased in the ileum. The density of NFκB p65-labelling gold particles increased in the ileal but remained unchanged in the duodenal ganglia. Meanwhile, both total and nuclear Nrf2 density increased in the myenteric neurons of the diabetic duodenum. In smooth muscle, NFκB p65 and Nrf2 density increased in the small intestine of diabetic rats. While on the mRNA level, NFκB p65 and Nrf2 were induced, on the protein level, NFκB p65 increased and Nrf2 decreased in muscle/myenteric plexus homogenates. Insulin treatment had protective effects. Conclusions: Our findings reveal a segment-specific NFκB and Nrf expression in myenteric neurons and ganglionic muscular environments, which may contribute to regional neuronal survival and motility disturbances in diabetes.

Diabetes mellitus is often associated with vascular complications in which hyperglycemia-induced oxidative stress may be the cause of the impaired vessels and circulating blood cells. The aim of this study was to follow the hyperglycemia-related metabolic and morphological changes in blood and urine samples of Wistar rats. Animals were divided into streptozotocin-induced diabetic (acute and chronic), insulin-treated diabetic, reversed diabetic, and control groups. In chronic diabetic rats, decreases in albumin, total protein, and antioxidant glutation concentration were measured, while glutamic-pyruvic transaminase, alkaline phosphatase, red blood cell (RBC) count, hematocrit, and hemoglobin levels were increased. Moreover, an increased level of the phenotypic variants was detected in the RBC population of the diabetic animals. In conclusion, we verified the sensitivity of RBCs to long-lasting hyperglycemia, and to insulin deficiency, which were both accompanied with an increased level of RBC-derived parameters and the presence of eccentrocytes, hemolyzed RBCs, and codocytes. Moreover, our results show that the response of the RBC glutation system to oxidative stress depends on the duration of hyperglycemia, and that the short-term activation of this defense system is exhausted in a long-lasting oxidative environment. Insulin therapy was effective in the case of most parameters, which clearly emphasizes the importance of maintaining blood glucose at physiological level.