Blood Level of Nitric Oxide Metabolites and Expression of Genes Regulating NO Synthesis in Early Forms of Non-Alcoholic Fatty Liver Disease.

The levels of NO metabolites in the plasma and mRNA of the NOS3, ATG9B, and NOS2 genes in peripheral blood leukocytes of healthy people and patients with early forms of non-alcoholic fatty liver disease (steatosis and weak activity non-alcoholic steatohepatitis) were studied. In patients with steatohepatitis, the concentration of NO metabolites in the blood and the level of mRNA of the NOS2 gene were higher than in patients with steatosis and healthy people. These differences can be of diagnostic value for distinguishing between steatosis and weak activity steatohepatitis in non-alcoholic fatty liver disease. A correlation between the levels of NO metabolites and the expression of the NOS2 gene in weak activity steatohepatitis was established, which indicates activation of NO synthesis in non-alcoholic steatohepatitis due to the expression of the inducible NO synthase gene. The level of the NOS2 gene mRNA in peripheral blood leukocytes of patients with weak activity steatohepatitis correlated with the level of TNFα and IL-6 cytokines. An increase in the level of NO in the blood in weak activity steatohepatitis correlated with the level of MDA, an indicator of oxidative stress.

The Role of the Soluble Interleukin-6 Receptor in the Progression of Nonalcoholic Fatty Liver Disease.

The blood level of soluble IL-6 receptor was measured in patients with different clinical and morphological forms of nonalcoholic fatty liver disease and healthy donors. The relationship of the soluble IL-6 receptor with the content of IL-6, the level of the IL6 gene mRNA, and a number of markers of hepatocyte and peripheral blood leukocyte apoptosis was assessed. It has been established for the first time that progression of nonalcoholic fatty liver disease is associated with changes in the level of soluble IL-6 receptor in the blood. In patients with high activity of nonalcoholic steatohepatitis and liver cirrhosis, the blood concentration of soluble IL-6 receptor sharply decreased in comparison with the earlier stages of progression of nonalcoholic fatty liver disease (liver steatosis, nonalcoholic steatohepatitis of weak and moderate activity). This allows considering the decrease in this indicator as a new diagnostic marker for distinguishing nonalcoholic steatohepatitis of high activity from weak and moderate activity. A close correlation between changes in the level of soluble IL-6 receptor and apoptosis of peripheral blood leukocytes and hepatocytes was revealed.

Biochemical and molecular-genetic indicators of inflammation and apoptosis in liver cirrhosis as an outcome of the progression of non-alcoholic steatohepatitis.

AIM: A comparative analysis of the complex of clinical and laboratory indicators (including the content of cytokines in blood plasma and the level of expression of TNF and IL6 genes in peripheral leukocytes, as well as the level of biochemical and molecular-genetic indicators of apoptosis, such as the content of tissue polypeptide-specific antigen (TPS) in the blood, the activity of caspases 3, 8 and 9 and the expression level of the encoding genes in peripheral blood leukocytes) in patients with non-alcoholic fatty liver disease (NAFLD) with non-alcoholic steatohepatitis (NASH) of different activity, liver cirrhosis (LC) classes A and B and in the donors of control group. MATERIALS AND METHODS: 158 patients with NAFLD were examined: 116 patients with NASH diagnosed for the first time (NASH of weak, moderate and high activity) and 42 patients with the NAFLD at the stage of liver cirrhosis diagnosed for the first time (classes A and B according to the Child-Pugh classification). The control group consisted of 54 healthy donors. The clinical blood biochemistry, cytokine profile, tissue polypeptide-specific antigen content, the level of the TNF, IL6 gene and caspase gene transcription as well as caspase activity in peripheral blood leukocytes (PBL) were evaluated. RESULTS: In the progression of NASH to LC, together with changes in general clinical parameters, the cytokine profile are changed due to an increase in the level of IL-6 and IL-1ß; in peripheral leukocytes, the activity of caspase 9 increases and the activity of caspase 8 decreases compared to NASH, and the level of the TNF gene expression decreases as compared to NASH of high activity. These parameters can be considered as promising minimally invasive markers of progression of NAFLD to LC. CONCLUSION: In nonalcoholic cirrhosis as an outcome of the progression of non-alcoholic steatohepatitis changes in clinical parameters (indicating the development of hepatocellular deficiency, violation of protein and lipid metabolism, progressive inflammation) are accompanied by specific changes in levels of biochemical and molecular-genetic indicators of apoptosis and inflammation. With the progression of NASH to LC, the cytokine profile changes due to an increase in the level of proinflammatory cytokines, the apoptosis processes triggered by the internal pathway increase and the activity of apoptosis activated via the external pathway decreases in PBL.

IL6R Gene Polymorphic Variant rs2228145(C >A) as a Marker of Genetic Liability to Nonalcoholic Steatohepatitis in the Russian Population of Karelia.

Association of IL6R gene polymorphic variant rs2228145(C>A) with the development of nonalcoholic steatohepatitis in Karelia residents is detected. The risk of nonalcoholic steatohepatitis is more than 2-fold higher in carriers of CC genotype by rs2228145 polymorphic marker than in carriers of other genotypes. Plasma levels of IL-6 and the content of IL6R gene transcripts in the peripheral blood leukocytes are higher in patients with nonalcoholic steatohepatitis than in normal subjects. No relationships between rs2228145 polymorphism and the level of IL-6 and content of IL6 and IL6R mRNA were detected. Gene IL6R polymorphic variant rs2228145(C>A) seems to be involved in genetic predisposition of the population of Karelia to nonalcoholic steatohepatitis. However, biochemical and molecular mechanisms underlying the relationship of rs2228145 with the development of nonalcoholic steatohepatitis are not yet studied.

The clinical significance of insulin resistance in non-diabetic patients with early forms of non-alcoholic fatty liver disease.

AIM: To assess the presence of insulin resistance (IR) in non-diabetic patients with early forms of non-alcoholic fatty liver disease (NAFLD) - liver steatosis (LS) and steatohepatitis (SH) of mild activity and the influence of IR on the clinical course of these diseases. MATERIALS AND METHODS: 134 patients with NAFLD were examined: 54 with LS and 80 with SH. The control group consisted of 37 healthy donors. Anthropometric parameters (body mass index (BMI), waist circumference (WC)), clinical and biochemical blood indices, including the blood level of cytokeratin-18 fragments (CK-18), TNF-α and IL-6 cytokines, insulin were evaluated. The HOMA index and the fibrosis index (NAFLD FS) were calculated. Patients were divided into groups: I - with the absence of IR (HOMA-index <2.7), II - with the presence of IR (HOMA-index> 2.7). RESULTS: Indicators of hepatic injury, inflammation, cholestasis, fibrosis and atherogenic dyslipidemia are higher in patients with LS of group II (with IR) than in group I patients (without IR). BMI, WC, γ-glutamil transpeptidase, CK-18 and fibrosis index are significantly higher in group II patients with SH compared with group I, there is no significant difference in the level of cytolysis, inflammation and dyslipidemia indices. A high incidence of IR in non-diabetic patients with LS (37.0%) and SH (55.0%) was found and the effect of IR on the clinical course of these diseases was revealed. CONCLUSION: Insulin resistance in non-diabetic patients with NAFLD was detected in SH (55.0%) with higher frequency than in LS (37.0%). In LS, IR is associated with impaired hepatic cell damage, intrahepatic cholestasis, atherogenic dyslipidemia and fibrosis. In SH, IR is combined with reliable growth in indicators of hepatocyte apoptosis, cytokine proinflammatory status and fibrosis. IR determines the progressing course of NAFLD, promoting the transformation of steatosis into steatohepatitis and steatohepatitis into fibrosis and liver cirrhosis.

Intrahepatic cholestasis in nonalcoholic fatty liver disease.

AIM: To determine the frequency of intrahepatic cholestasis and its impact on the clinical features of the different forms of non-alcoholic fatty liver disease (NAFLD). MATERIALS AND METHODS: The study involved 163 patients with NAFLD: 92 (56.4%) with hepatic steatosis (HS), 56 (34.4%) with steatohep- atitis (SH) and 15 (9.2%) with liver cirrhosis (LC). Diagnosis is based on clinical, laboratory, ultrasound and histological data. Insulin, tu- mor necrosis factor α (TNF-α), fragments of cytokeratin-18 (FCK-18) were determined by ELISA. The index of insulin resistance (HOMA- IR) was calculated. NAFLD fibrosis score (NAFLD-FS) was determined, taking into account the patient's age, body mass index, presence or absence of carbohydrate metabolism disturbances, levels of ASAT, ALAT, albumin and blood platelets. RESULTS: Cholestatic syndrome was detected in 49 (30.1%) NAFLD patients: in 23 (25%) with HS, in 19 (33.9%) with SH and in 7 (46.7%) with LC. Patients with HS, SH and LC with signs of cholestasis as compared to patients with the same forms of NAFLD without cholestasis had significantly higher levels of the following indicators: aminotransferases, triglycerides, HOMA-IR, TNF-α, FCK-18, NAFLD-FS, - the number of platelets is reduced, indirectly confirming the more rapid development of fibrosis in cholestasis. These findings were consistent with published data on the violation in cholestasis regulatory functions of bile acids, which are ligands of hepatocyte nuclear receptor, re- sponsible for normal homeostasis. CONCLUSION: In all forms of NAFLD with cholestasis were detected more pronounced liver cell inflammation, hepatocyte necrosis and apoptosis, fibrosis, disturbance of carbohydrate and lipid metabolism, which contributed to a progressive course of NAFLD and confirmed the need for medical correction of cholestasis, starting with the earliest form of NAFLD - hepatosteatosis.