Chlorotyrosine protein adducts are reliable biomarkers of neutrophil-induced cytotoxicity in vivo

INTRODUCTION:A limitation for investigating the pathophysiological role of neutrophils in vivo is the lack of a reliable biomarker for neutrophil cytotoxicity in the liver. Therefore, we investigated if immunohistochemical detection of chlorotyrosine protein adducts can be used as a specific footprint for generation of neutrophil-derived hypochlorous acid in vivo.METHODS:C3Heb/FeJ mice were treated with 100 micrograms/kg endotoxin (ET) alone or in combination with 700 mg/kg galactosamine (Gal/ET). Some animals received additionally two doses of 10 mg/kg of the pancaspase inhibitor Z-VAD-fmk. An antibody against chlorotyrosine was used for the immunohistochemical analysis.RESULTS:At 6 h after Gal/ET, hepatocellular apoptosis was evident without increase in plasma ALT activities. Neutrophils accumulated in sinusoids but there was no evidence for chlorotyrosine staining. At 7 h after Gal/ET, about 54% of the sequestered neutrophils had extravasated, there was extensive necrosis and increased plasma ALT activities. Extensive immunostaining for chlorotyrosine, mainly colocalized with neutrophils, could be observed. Treatment with Z-VAD-fmk eliminated apoptosis, necrosis and the increase in plasma ALT values. Neutrophil extravasation was prevented but the overall number of neutrophils in the liver was unchanged. Chlorotyrosine staining was absent in these samples. After ET alone (7 h), sinusoidal neutrophil accumulation was similar to Gal/ET treatment but there was no apoptosis, neutrophil extravasation, ALT release or chlorotyrosine staining.CONCLUSIONS:Chlorotyrosine staining in liver samples correlated well with evidence of neutrophil-induced liver injury in the endotoxemia model. These results indicate that assessment of chlorotyrosine protein adduct formation by immunohistochemistry could be a useful marker of neutrophil-induced liver cell injury in vivo.This item is part of the UA Faculty Publications collection. For more information this item or other items in the UA Campus Repository, contact the University of Arizona Libraries at [email protected]

Validity and reliability of total body volume and relative body fat mass from a 3-dimensional photonic body surface scanner

OBJECTIVE: Three-dimensional photonic body surface scanners (3DPS) feature a tool to estimate total body volume (BV) from 3D images of the human body, from which the relative body fat mass (%BF) can be calculated. However, information on validity and reliability of these measurements for application in epidemiological studies is limited. METHODS: Validity was assessed among 32 participants (men, 50%) aged 20-58 years. BV and %BF were assessed using a 3DPS (VitusSmart XXL) and air displacement plethysmography (ADP) with a BOD POD(R) device using equations by Siri and Brozek. Three scans were obtained per participant (standard, relaxed, exhaled scan). Validity was evaluated based on the agreement of 3DPS with ADP using Bland Altman plots, correlation analysis and Wilcoxon signed ranks test for paired samples. Reliability was investigated in a separate sample of 18 participants (men, 67%) aged 25-66 years using intraclass correlation coefficients (ICC) based on two repeated 3DPS measurements four weeks apart. RESULTS: Mean BV and %BF were higher using 3DPS compared to ADP, (3DPS-ADP BV difference 1.1 +/- 0.9 L, p<0.01; %BF difference 7.0 +/- 5.6, p<0.01), yet the disagreement was not associated with gender, age or body mass index (BMI). Reliability was excellent for 3DPS BV (ICC, 0.998) and good for 3DPS %BF (ICC, 0.982). Results were similar for the standard scan and the relaxed scan but somewhat weaker for the exhaled scan. CONCLUSIONS: Although BV and %BF are higher than ADP measurements, our data indicate good validity and reliability for an application of 3DPS in epidemiological studies