Hepatic insulin synthesis increases in rat models of diabetes mellitus type 1 and 2 differently.

Insulin-positive (+) cells (IPCs), detected in multiple organs, are of great interest as a probable alternative to ameliorate pancreatic beta-cells dysfunction and insulin deficiency in diabetes. Liver is a potential source of IPCs due to it common embryological origin with pancreas. We previously demonstrated the presence of IPCs in the liver of healthy and diabetic rats, but detailed description and analysis of the factors, which potentially can induced ectopic hepatic expression of insulin in type 1 (T1D) and type 2 diabetes (T2D), were not performed. In present study we evaluate mass of hepatic IPCs in the rat models of T1D and T2D and discuss factors, which may stimulate it generation: glycaemia, organ injury, involving of hepatic stem/progenitor cell compartment, expression of transcription factors and inflammation. Quantity of IPCs in the liver was up by 1.7-fold in rats with T1D and 10-fold in T2D compared to non-diabetic (ND) rats. We concluded that ectopic hepatic expression of insulin gene is activated by combined action of a number of factors, with inflammation playing a decision role.

Accelerated Generation of Extra-Islet Insulin-Producing Cells in Diabetic Rats, Treated with Sodium Phthalhydrazide.

ß-cells dysfunction plays an important role in the pathogenesis of type 2 diabetes (T2D), partially may be compensated by the generation of extra-islet insulin-producing cells (IPCs) in pancreatic acini and ducts. Pdx1 expression and inflammatory level are suggested to be involved in the generation of extra-islet IPCs, but the exact reasons and mechanisms of it are unclear. Macrophages are key inflammatory mediators in T2D. We studied changes in mass and characteristics of extra-islet IPCs in rats with a streptozotocin-nicotinamide model of T2D and after i.m. administration of 20 daily doses of 2 mg/kg b.w. sodium aminophthalhydrazide (APH). Previously, we found that APH modulates macrophage production and increases the proliferative activity of pancreatic ß-cells. Expressions of insulin and Pdx1, as well as F4/80 (macrophage marker), were detected at the protein level by immunohistochemistry analysis, the concentration of pro- and anti-inflammatory cytokines in blood and pancreas-by ELISA. Diabetic rats treated with APH showed an increasing mass of extra-islet IPCs and the content of insulin in them. The presence of Pdx1+ cells in the exocrine pancreas also increased. F4/80+ cell reduction was accompanied by increasing TGF-ß1 content. Interestingly, during the development of diabetes, the mass of ß-cells decreased faster than the mass of extra-islet IPCs, and extra-islet IPCs reacted to experimental T2D differently depending on their acinar or ductal location.

Accelerated liver recovery after acute CCl4 poisoning in rats treated with sodium phthalhydrazide.

Pharmacotherapy of hepatobiliary disorders is an important issue due to the high prevalence of liver failure, toxic and viral hepatitis and cirrhosis. The number of stimuli that can potentially induce or accelerate liver recovery is limited; in our study we selected sodium phthalhydrazide, which has been found to promote liver regeneration after partial hepatectomy. We examined the effects of phthalhydrazide on liver morphometric, histological and biochemical parameters in rats intoxicated with CCl4. Accelerated liver recovery after CCl4 intoxication in phthalhydrazide-treated animals was evidenced by increased number of liver sinusoidal cells, reduced focal necrosis of hepatocytes and reduced perifocal leukocyte infiltration. Decreased plasma levels of pro-inflammatory cytokines TNF-α and IL-18 and decreased concentrations of IL-6 and IFN-γ in liver homogenates were associated with reduced severity of cholestasis and normalized hepatic protein synthesis in CCl4-intoxicated rats exposed to phthalhydrazide. Anti-inflammatory and immunomodulating properties of phthahlhydrazide can be an important factor contributing to accelerated liver recovery at early stages of acute CCl4-toxic liver impairment.

Recruitment of macrophages and bone marrow stem cells to regenerating liver promoted by sodium phthalhydrazide in mice.

Pharmacological interventions which could be hepatoprotective, depending on bioavailability, anti-inflammatory and macrophage-targeting potential of drugs, are still at early preclinical stages. Existing evidence from many animal models of liver injury, as well as from human data, indicate that pharmacological and/or phytochemical interventions have limited impact on liver recovery. Recent studies on stem cell therapies focused on different cell subsets involved in tissue repair, including monocytes/macrophages and bone marrow cells migrating to the injured liver. Partial hepatectomy (PH) resulted in a rapid increase of monocytes/macrophages in bone marrow and liver, which could be further enhanced by prior treatment of animals with sodium phthalhydrazide. Increased number of proliferating Ki67+ hepatocytes, increased total protein and albumin content in regenerating liver, recruitment of CD172a+ macrophages and more differentiated CD45lowCD117+ bone marrow cells, could be further promoted by the treatment of animals with 2 mg/kg b.w. phthalhydrazide, considered immunomodulatory, antioxidant and macrophage-silencing. Phenotypic polarization of macrophages can possibly explain the macrophage reparative capacities, protective against liver injury. Enhanced macrophage cell recruitment from bone marrow to regenerating liver can be possibly one of important events in hepatic recovery.

Partial recovery from alloxan-induced diabetes by sodium phthalhydrazide in rats.

In the commonly used experimental model of diabetes, a cytotoxic glucose analogue alloxan can selectively destruct pancreatic ß-cells, with characteristics similar to the type-1 diabetes (T1D) in humans. Treatment of diabetic rats with sodium phthalhydrazide partially reversed diabetogenic pathology in the alloxan-induced diabetes. The alloxan-treated rats with permanent hyperglycemia, which further received i.p. twenty daily doses 2mg/kg b.w. phthalhydrazide, showed at 60days of the experiment a significant amelioration of the diabetes status. Hyperglycemia was decreased by 52%, glycated haemoglobin HbA1c returned to control value, insulin concentration significantly increased from 45,4% (alloxan group) to 59,5% (alloxan+phthalhydrazide) of the control values. Importantly, phthalhydrazide treatment of alloxan-treated diabetic rats markedly decreased the concentrationof interleukin-6 (IL-6) and corticosterone level. Morphometric analysis revealed a marked increase in the number of pancreatic islets/mm2, and a number of cells/mm2 in the pancreatic islets. These changes, including 3-fold increase in the number of insulin-producing cells and 2-fold decrease in blood glucose levels, correlated with the increased proliferative activity of pancreatic ß-cells in the diabetic phthalhydrazide-treated animals. Interestingly, the number of CD68+ cells/macrophages in the pancreatic islets, which was relatively high in the alloxan group (63,9+- 16.4/mm2), markedly decreased after the phthalhydrazide treatment (23,6+-7,2/mm2). Taking together with the previous data on the phthalhydrazide-related macrophage silencing, restriction of macrophage quantity in the alloxan-affected pancreatic islets can be possibly one of important events leading to the partial recovery from the ß-cell disruption.

A tetrahydrophthalazine derivative 'sodium nucleinate" exerts a potent suppressive effect upon LPS-stimulated mononuclear cells in vitro and in vivo.

We described the use of a new chemical substance Sodium nucleinate (SN) as an immunomodulatory substance exhibiting antiinflammatory properties. Sodium nucleinate (SN) registrated in Russian Federation as Tamerit, is 2-amino-1,2,3,4-tetrahydrophthalazine-1,4-dione sodium salt dihydrate, derivative of well known chemical substance luminol. To comprehend the mechanisms of SN immunomodulatory activity, we examined the SN modulation of the innate inflammatory cytokine response of human PBMC stimulated with LPS in vitro. Furthermore, we studied the immunomodulatory effects of SN in mice challenged with E. coli LPS in vivo to investigate a possible novel approach to therapy of excessive inflammation that interfere with the response to endotoxin and inflammatory mediators. Our results demonstrated that SN is an efficient inhibitor of sepsis development in mice model of LPS-induced sepsis. The changes induced by SN include decreased mice plasma inflammatory cytokine production. Simmilary we demonstrated a decreased TNF-alpha, IFN-gamma and IL-6 response in human LPS-stimulated PBMNCs. SN was therefore shown to be a promising inhibitor of multiple inflammatory cytokine secretion.