Production of inflammatory cytokines by in vitro stimulated peripheral blood cells in children with chronic viral hepatitis C

Abstract

Chronic viral hepatitis C (ChVHC) is still a relevant problem for healthcare worldwide, despite significant advances in the diagnosis and treatment of this disease. Cytokines (CTs) play an important role in the pathogenesis of ChVHC, since they are universal regulators of immune response mechanisms. Studying the production of CTs by blood cells under the influence of various stimulants in vitro provides in-depth information about functioning of the producer cells. In this regard, the aim of this work was to study the production of cytokines by in vitro stimulated peripheral blood leukocytes from patients with ChVHC. The study group included 22 children with ChVHC and 29 conditionally healthy children, (mean age: 10 years, and 4 months; 10 years and 9 months, respectively). Samples of heparinized blood were diluted with RPMI-1640 medium. The following series were prepared: the control sample without stimulant; a specimen supplied with phytohemagglutinin (Sigma); a sample stimulated with agonistic antibodies to CD3 (aCD3, Biocon, Russia); a specimen treated with agonistic antibodies to CD3 (Biocon, Russia) and to CD28 (aCD28, Beckman Coulter, USA). The blood samples were incubated for 24 hours (37 °C, 5% CO2). The concentration of IL-6, IL-8, and IL-17 was determined in the cell culture supernatants using ELISA technique (Vector-Best, Russia). Statistical analysis was performed by the Mann–Whitney test and the sign test. It was found that spontaneous production of IL-6 and IL-17 by blood cells did not differ between healthy children and patients with ChVHC, whereas IL-8 levels were higher in viral infection. In vitro exposure to various stimulants led to increased concentrations of all studied CTs complexes in both patient groups. The IL-8 concentration in the ChVHC group was lower than the control values in all three stimulation variants. In cell cultures from ChVHC patients with combined aCD3 and aCD28 exposure, IL-6 and IL-17 levels were lower than in healthy controls. IL-6 contents were also lower than in controls exposed to aCD3 alone. Thus, patients with ChVHC had elevated spontaneous IL-8 concentrations and decreased stimulated levels of all studied proinflammatory CTs. This finding suggests a decreased reserve capacity of CT-producing blood cells during chronic infection.