1b). These neutrophils frequently assembled into aggregate structures around hepatocytes with macrovesicular steatosis, resembling selleck chem Imatinib the crown-like macrophage structures found in adipose tissue of obese animals [21]. Thus, hepatic lipid accumulation triggered by a high-fat diet is associated with liver neutrophil infiltration, leading to increased hepatic MPO protein levels. Figure 1 Strong high-fat diet-induced induction of MPO in the liver. Reduced MPO Activity in LDLR?/?/MPO?/?tp Mice The accumulation of MPO in the liver upon high-fat feeding suggested that MPO might contribute to the pathogenesis of NASH. To further examine the role of MPO in NASH, we took advantage of the fact that MPO is exclusively expressed in the hematopoietic system [22], and performed bone marrow transplantation experiments that resulted in the generation of combined MPO-deficient, LDL-R-deficient mice (LDLR?/?/MPO?/?tp) mice.
LDLR?/?/MPO+/+tp controls and LDLR?/?/MPO?/?tp mice had a similar body weight, and diet-induced weight gain was comparable (LDLR?/?/MPO+/+tp from 19.9��0.4 to 21.8��0.3 g, LDLR?/?/MPO?/?tp: from 20.0��0.2 to 21.9��0.7 g). After high-fat feeding, MPO plasma levels in the LDLR?/?/MPO+/+tp mice were 324 ng/ml whereas plasma from LDLR?/?/MPO?/?tp animals contained only 22 ng/ml of MPO (p<0.01; Fig. 2a). Similar to plasma MPO, liver MPO was very significantly reduced in the LDLR?/?/MPO?/?tp group after high-fat feeding as shown by quantification of MPO-positive cell numbers, with levels well under those observed in chow-fed LDLR?/?/MPO+/+ mice(p<0.001; Fig. 2b).
Figure 2 Reduced hepatic MPO and MPO-derived nitrated proteins in LDLR?/?/MPO?/?tp mice after 8 weeks of high-fat feeding. To investigate if the marked reduction of hepatic MPO in LDLR?/?/MPO?/?tp mice translated into diminished generation of MPO-mediated cytotoxic products, we studied hepatic levels of nitrotyrosine, a protein modification generated at sites of inflammation as a result of the activity of several enzymes among which MPO, which accumulates in NAFLD [5], [8], [23]. As expected, mice in the LDLR?/?/MPO?/?tp group displayed significantly reduced levels of nitrotyrosine in the liver, consistent with reduced MPO activity (Fig. 2c; p<0.05). Characterization of NASH Severity Histological examination of the livers after H/E-staining revealed diffuse microvesicular and macrovesicular steatosis in both groups (Fig.
3). Semi-quantitative evaluation of the extent of steatosis showed no statistically significant differences between LDLR?/?/MPO+/+tp and LDLR?/?/MPO?/?tp mice (2.4��0.3 Brefeldin_A vs. 2.1��0.2, p=0.43). Inflammatory cell foci were observed in both groups, but less frequently found in the LDLR?/?/MPO?/?tp group, although the difference was not statistically significant (0.75��0.31 vs. 0.50��0.27, p=0.12). Hepatocyte ballooning, a feature of progressive human NASH, was not observed in either group.
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