Abstract
BackgroundA novel in vitro system was employed to investigate liver tissue respiration (mitochondrial O2 consumption) in mice treated with concanavalin A (Con A). This study aimed to investigate hepatocyte bioenergetics in this well-studied hepatitis model.MethodsC57Bl/6 and C57Bl/6 IFN-γ−/− mice were injected intravenously with 12 mg ConA/kg. Liver specimens were collected at various timepoints after injection and analyzed for cellular respiration and caspase activation. Serum was analyzed for interferon-gamma (IFN-γ) and aminotransferases. Fluorescence activated cell sorting analysis was used to determine the phenotype of infiltrating cells, and light and electron microscopy were used to monitor morphological changes. Phosphorescence analyzer that measured dissolved O2 as function of time was used to evaluate respiration.ResultsIn sealed vials, O2 concentrations in solutions containing liver specimen and glucose declined linearly with time, confirming zero-order kinetics of hepatocyte respiration. O2 consumption was inhibited by cyanide, confirming the oxidation occurred in the respiratory chain. Enhanced liver respiration (by ≈68%, p<0.02) was noted 3 hr after ConA treatment, and occurred in conjunction with limited cellular infiltrations around the blood vessels. Diminished respiration (by ≈30%, p=0.005) was noted 12 hr after ConA treatment, and occurred in conjunction with deranged mitochondria, areas of necrosis, and prominent infiltrations with immune cells, most significantly, CD3+NKT+ cells. Increases in intracellular caspase activity and serum IFN-γ and aminotransferase levels were noted 3 hr after ConA treatment and progressed with time. The above-noted changes were less pronounced in C57Bl/6 IFN-γ−/− mice treated with ConA.ConclusionsBased on these results, liver tissue bioenergetics is increased 3 hr after ConA exposure. This effect is driven by the pathogenesis of the disease, in which IFN-γ and other cytokines contribute to. Subsequent declines in liver bioenergetics appear to be a result of necrosis and active caspases targeting the mitochondria within hepatocytes.
Highlights
A novel in vitro system was employed to investigate liver tissue respiration in mice treated with concanavalin A (Con A)
Concanavalin A (ConA) is a plant lectin from seeds of Canavalia ensiformis [Jack bean] that serves as a polyclonal T-cell mitogen and is used to induce hepatitis in mice
Targeting natural killer T (NK T) cells with glucocerebroside, a naturally occurring glycolipid that inhibits NK T cell proliferation, ameliorates ConA-induced hepatitis [5]. These findings suggest the pathological outcome of the liver in ConA-treated animals relies on initial activation of innate immune components, such as NK T cells, followed by subsequent activation and recruitment of effector immune cells, including macrophages and T-cells
Summary
A novel in vitro system was employed to investigate liver tissue respiration (mitochondrial O2 consumption) in mice treated with concanavalin A (Con A). Liver specimens were collected at various timepoints after injection and analyzed for cellular respiration and caspase activation. Other cells of the innate immune system are early events that, in turn, lead to secretion of inflammatory cytokines, such as interferon-γ (IFN-γ), tumor necrosis factor (TNF)-α, interleukin (IL)-12, and IL-18 [3]. This biochemical burst of inflammatory mediators targets multiple organs, including the liver. Its outcome is a hepatotoxicity that is characterized by mononuclear cellular infiltration and foci of necrosis [4]
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