Individuals with Primary Sclerosing Cholangitis Have Elevated Levels of Biomarkers for Apoptosis but Not Necrosis

BACKGROUND AND AIM: Hepatocyte apoptosis or necrosis from accumulation of bile salts may play an important role in the disease progression of primary sclerosing cholangitis (PSC). The aim of the current study was to measure serum markers of hepatocyte apoptosis (cytokeratin-18 fragments--K18) and necrosis (high-mobility group protein B1--HMGB1) in adults with PSC and examine the relationship with disease severity. METHODS: We measured serum levels of K18 and HMGB1 in well-phenotyped PSC (N = 37) and 39 control subjects (N = 39). Severity of PSC was assessed biochemically, histologically, and PSC Mayo risk score. Quantification of hepatocyte apoptosis was performed using TUNEL assay. RESULTS: The mean age of the study cohort was 49.7 ± 13.3 years and comprised of 67% men and 93% Caucasian. Serum K18 levels were significantly higher in the PSC patients compared to control (217.4 ± 78.1 vs. 157.0 ± 58.2 U/L, p = 0.001). However, HMGB1 levels were not different between the two groups (5.38 ± 2.99 vs. 6.28 ± 2.85 ng/mL, p = 0.15). Within the PSC group, K18 levels significantly correlated with AST (r = 0.5, p = 0.002), alkaline phosphatase (r = 0.5, p = 0.001), total bilirubin (r = 0.61, p ≤ 0.001), and albumin (r = -0.4, p = 0.02). Serum K18 levels also correlated with the level of apoptosis present on the liver biopsy (r = 0.8, p ≤ 0.001) and Mayo risk score (r = 0.4, p = 0.015). CONCLUSION: Serum K18 but not HMGB1 levels were increased in PSC and associated with severity of underlying liver disease and the degree of hepatocyte apoptosis

Liver injury and fibrosis induced by dietary challenge in the ossabaw miniature Swine

BACKGROUND: Ossabaw miniature swine when fed a diet high in fructose, saturated fat and cholesterol (NASH diet) develop metabolic syndrome and nonalcoholic steatohepatitis (NASH) characterized by liver injury and fibrosis. This study was conducted to further characterize the development of NASH in this large animal model. METHODS: Ossabaw swine were fed standard chow (control group; n = 6) or NASH diet (n = 6) for 24 weeks. Blood and liver tissue were collected and liver histology were characterized at 0, 8, 16 and 24 weeks of dietary intervention. Hepatic apoptosis and lipid levels were assessed at week 24. RESULTS: The NASH diet group developed metabolic syndrome and progressive histologic features of NASH including: (a) hepatocyte ballooning at 8 weeks which progressed to extensive ballooning (>90% hepatocytes), (b) hepatic fibrosis at week 16, which progressed to moderate fibrosis, and (c) Kupffer cell accumulation with vacuolization at 8 weeks which progressed through week 24. The NASH diet group showed increased hepatocyte apoptosis that correlated with hepatic total and free cholesterol and free fatty acids, but not esterified cholesterol or triglycerides. CONCLUSIONS: This report further characterizes the progression of diet-induced NASH in the Ossabaw swine model. In Ossabaw swine fed the NASH diet: (a) hepatocyte injury and fibrosis can occur without macrovesicular steatosis or excess triglyceride accumulation; (b) hepatocyte ballooning generally precedes the development of fibrosis; (c) there is increased hepatocyte apoptosis, and it is correlated more significantly with hepatic free cholesterol than hepatic free fatty acids and had no correlation with hepatic triglycerides

The roles of the formal and informal sectors in the provision of effective science education

For many years, formal school science education has been criticised by students, teachers, parents and employers throughout the world. This article presents an argument that a greater collaboration between the formal and the informal sector could address some of these criticisms. The causes for concern about formal science education are summarised and the major approaches being taken to address them are outlined. The contributions that the informal sector currently makes to science education are identified. It is suggested that the provision of an effective science education entails an enhanced complementarity between the two sectors. Finally, there is a brief discussion of the collaboration and communication still needed if this is to be effective

Liver Lipid Quantification in NASH and Control Swine at Week 24.

<p>Quantification of lipid species in livers of NASH and control group swine at Week 24 of dietary intervention. Shown is the mean ± standard deviation.</p><p>Liver Lipid Quantification in NASH and Control Swine at Week 24.</p

Liver histology by electron microscopy (EM) (A-D 21,400X).

<p><b>A and C:</b> An EM section from a control swine hepatocyte at week 24 shows no accumulation of abnormal material. <b>B and D</b>: An EM section of the liver from a swine fed NASH diet at week 24. Hepatocytes were filled with electron-dense material arranged as whorled lamellar structures, located within membrane-bound vesicles (white arrows) consistent with autophagolysosomes.</p

Liver total, free, and esterified cholesterol measurement and their correlation with liver caspase 3/7 activity at week 24.

<p>Total cholesterol <b>(A)</b>, free cholesterol <b>(B)</b> and esterified cholesterol <b>(C)</b> were higher in the NASH diet fed group than control group. Positive correlations were observed between caspase 3/7 activity and total cholesterol (R² = 0.3770, p = 0.0338) <b>(D)</b> and free cholesterol (R² = 0.4961, p = 0.0105) <b>(E)</b> levels, but not esterified cholesterol (R² = 0.0098, p = 0.7593) <b>(F)</b>. * indicates statistically significance difference between the NASH diet and the control diet groups.</p

Liver Lipid Quantification in NASH and Control Swine at Week 24.

<p>Quantification of lipid species in livers of NASH and control group swine at Week 24 of dietary intervention. Shown is the mean ± standard deviation.</p><p>Liver Lipid Quantification in NASH and Control Swine at Week 24.</p

Quantification of hepatic apoptosis at week 24.

<p><b>A:</b> TUNEL assay was performed on paraffin embedded sections. The NASH diet group had significantly higher apoptotic index (number of TUNEL positive cells per 1000 cells) (p = 0.0125). <b>B:</b> Hepatic caspase 3/7 activity shown in fluorescence arbitrary units. Caspase 3/7 activity is significantly higher in NASH diet group than the control diet fed group (p = 0.0397). All data was presented as Mean ± SEM. * indicates statistically significance difference between the NASH diet and the control diet groups.</p

Correlation of hepatic free Fatty acid and triglyceride levels with apoptosis at week 24.

<p><b>A:</b> Hepatic free fatty acid levels were more than 5-fold higher in the NASH diet group than control group although it did not reach statistical significance (p = 0.0667). <b>B:</b> There was no difference in hepatic triglyceride concentration between two groups (p = 0.2134). <b>C</b>: A positive correlation was observed between caspase 3/7 activity and fatty acid levels (R² = 0.3096, p = 0.0603). <b>D:</b> There was no correlation between hepatic triglyceride concentration and hepatic caspase 3/7 activity (R² = 0.0684, p = 0.4115).</p