Rafael Enat

Effect of vascular endothelial growth factor on hepatic regenerative activity following partial hepatectomy in rats

BACKGROUND/AIMS Vascular endothelial growth factor (VEGF) is an angiogenic factor with a growth-promoting effect that is thought to be restricted to vascular endothelial cells. Its essential role during liver regeneration has yet to be determined. The aim of this study was to document the effect of exogenous VEGF administration on liver regeneration in rats undergoing submaximal hepatic resections. METHODS Adult male Sprague-Dawley rats (n = 4/group) undergoing 30% partial hepatectomy were administered 200 ng VEGF165 intravenously and were sacrificed at 24, 36, and 48 h postoperatively. Liver regeneration was monitored by measuring the restituted liver mass, proliferating cell nuclear antigen (PCNA) immunostaining, and hepatic PCNA protein by Western blot. RESULTS Changes in restituted liver mass 48 h postsurgery were more prominent, but did not differ statistically between VEGF-treated and control rats (47% vs. 29%; p<0.06). Nevertheless, PCNA immunostaining showed increased labeling index of hepatocytes, apparent at 36 and 48 h after partial hepatectomy (38% vs. 18% [p<0.041 and 42% vs. 11% [p<0.021], respectively). Hepatic PCNA proteins measured by Western blot showed a 3-fold increase in VEGF-treated rats 48 h postsurgery compared with controls (p<0.01). CONCLUSION Exogenous VEGF administration early after partial hepatectomy stimulates liver regeneration in rats. Whether or not VEGF165 is a direct mitogen for hepatocytes remains to be determined.

Growth hormone-stimulated insulin-like growth factor (IGF) I and IGF-binding protein-3 in liver cirrhosis

BACKGROUND/AIMS The aim of this study was to evaluate the livers potential to generate insulin-like growth factor (IGF) I and IGF-binding protein-3 (IGFBP-3), following stimulation by human recombinant growth hormone, as a possible marker for liver functional reserve in patients with liver cirrhosis. METHODS In a pilot study, 15 patients (mean age 56 years) with postnecrotic liver cirrhosis were divided into two groups according to disease severity (Child-Pugh score): Group 1 (n=8) with scores of 5-8 and Group 2 (n=7) with scores of 9-12. Five age-matched healthy subjects served as controls. Human recombinant growth hormone (0.06 mg/kg) was administered subcutaneously on 2 consecutive days. Serum levels of IGF-I and IGFBP-3 were measured before and up to 48 h after human recombinant growth hormone injection. Nutritional status was assessed by the creatinine-height index and was compared to lymphocyte count, body mass index, and muscle arm circumference. RESULTS Baseline IGF-I levels were significantly lower in patients with cirrhosis than in controls, while no differences were noted between the two patient groups. IGF-I levels increased significantly after rhGH administration to the healthy controls, to a lower degree in Group 1, while no change occurred in Group 2. IGF-I levels at 24 h and beyond correlated significantly with the nutritional status, the Child-Pugh score, and the basal levels of GH-binding protein and IGFBP-3. IGFBP-3 serum levels did not change after rhGH stimulation. CONCLUSIONS IGF-I generation after GH stimulation may provide a new dimension in the assessment of liver function and nutritional status in patients with liver cirrhosis.

Prognostic value of generation of growth hormone-stimulated insulin-like growth factor-I (IGF-I) and its binding protein-3 in patients with compensated and decompensated liver cirrhosis.

Our aim was to study the prognostic value ofgrowth hormone (GH)-stimulated insulin-like growthfactor-I (IGF-I) and IGF-binding protein-3 (IGFBP-3)generation in patients with compensated [group 1 (N = 8) with a Child-Pugh (CP) score of 5-8] anddecompensated postnecrotic liver cirrhosis [group 2 (N= 7) with a CP score of 9-12]. Serum levels of IGF-I,GH-binding protein (GHBP), and IGFBP-3 were measuredbefore and 24 hr after a single subcutaneous injectionof recombinant human GH (rhGH, 0.14 units/kg). Patients(mean age 56 years) were followed prospectively forthree years. Six patients (40%) died during the follow-up period, of whom half had a CP score<9. Mean serum IGF-I levels 24 hr after rhGHinjection (group 1 vs group 2, 17.4 ± 6.8 vs 7.4± 0.7 nmol/liter) predicted survival with 93%accuracy. Levels <10 nmol/liter portended a poorprognosis, with 15% survival at one year, whereas levels>10 nmol/liter had a 100% survival rate at one andtwo years, respectively. Baseline IGF-I (9.98 ± 2.0 vs 6.38 ± 0.8 nmol/liter), GHBP (9.2± 3 vs 5.7 ± 0.8%/50 μl), and IGFBP-3serum levels at baseline (1.7 ± 0.3 vs 0.86± 0.2 mg/liter) and at 24 hr (2.04 ± 0.38vs 0.99 ± 0.3 mg/liter) did not add to the predictive value ofstimulated IGF-I levels at 24 hr and were less accuratein predicting the outcome in comparison to CP score(80%). We conclude that stimulated IGF-1 < 10nmol/liter may be a true predictor of a negative prognosisin patients with liver cirrhosis.

Basic fibroblast growth factor is hepatotropic for rat liver in regeneration.

A role for fibroblast growth factor in liver regeneration has recently been suggested. In this study we followed the intravenous delivery of recombinant human [125I]basic fibroblast growth factor to the liver of rats following 68% partial hepatectomy. The concentration of [125I]basic fibroblast growth factor was higher in the liver (mean +/- SD, 6.8 +/- 0.89% of injected dose) and the kidney (6.7 +/- 0.2%) of sham-operated rats than in the spleen (2.8 +/- 0.45%). It increased threefold in the liver only, soon after 68% partial hepatectomy (20.3 +/- 5.3%, p < 0.001), and remained high for the first 24 h. We also studied the effect of basic fibroblast growth factor injection on the rate of [3H]thymidine incorporation into liver DNA in rats subjected to either 21% or 68% partial hepatectomy. A significant increase was seen after intramesenteric injection of 500 ng basic fibroblast growth factor into rats subjected to 21% partial hepatectomy (23.5 +/- 7.3 cpm/micrograms DNA) compared to saline-injected rats (14.5 +/- 6.4 cpm/micrograms DNA, p = 0.034). A dose of 5000-25,000 ng injected into a peripheral vein resulted in higher thymidine incorporation than in saline-injected control rats (36.9 +/- 12.7 and 9.7 +/- 6.1 cpm/micrograms DNA, respectively; p < 0.0001). No significant effect was seen after 68% partial hepatectomy. Autoradiography showed that the hepatocytes were the predominant labelled cells early after hepatectomy and basic fibroblast growth factor injection. We conclude that basic fibroblast growth factor uptake by the liver is increased after 68% partial hepatectomy and that basic fibroblast growth factor is mitogenic to liver parenchymal cells early after 21% partial hepatectomy.

A pilot study on the hemodynamic effect of short-term ursodeoxycholic acid therapy in patients with stable liver cirrhosis

Objective:Total serum bile acid concentrations are elevated in individuals with liver disease. Ursodeoxycholic acid (UDCA) therapy in such patients results in a further significant rise in plasma levels to the extent that it becomes the major circulating bile acid. In laboratory animals, bile acids, such as taurocholic acid, have also been shown to possess a diuretic-like action, as they can promote diuresis, natriuresis, and kaliuresis by inhibiting tubular sodium reabsorption. The aim of the present study was to assess the effect of 1 months UDCA therapy on cardiovascular function in cirrhotic patients.Methods:Two groups of patients with cirrhosis were studied, six with primary biliary cirrhosis (PBC) and six with postnecrotic liver cirrhosis (PNC). Cardiovascular function was assessed by determination of blood pressure, heart rate, and by two-dimensional and pulsed Doppler echocardiography.Results:In PBC patients, 1 months treatment with UDCA significantly reduced diastolic volume without changing systolic, diastolic, and mean blood pressures, heart rate, systolic and stroke volumes, ejection fraction, cardiac output, and systemic vascular resistance. In PNC patients, UDCA significantly reduced cardiac output, with a tendency to reduce left ventricular volumes, without any changes in systolic, diastolic, and mean blood pressures.Conclusion:UDCA caused reductions in diastolic volume in the PBC patients and cardiac output in the PNC patients. Such reductions are not unlike that seen in individuals treated with diuretics. This diuretic-like action deserves further study, particularly in cirrhotic patients who are also being treated with diuretics or show evidence of cardiac myopathy.

Correlation of HBV DNA and monoclonal reactivity to HBsAg in serum of patients with HBV infection

Hepatitis B virus (HBV) DNA hybridization assay, a monoclonal radioimmunoassay (M-RIA) for hepatitis B surface antigen (HBsAg) and conventional polyclonal immunoassays for HBV associated antigens were used to study sera from patients on dialysis and with acute hepatitis B. HBV DNA was detectable in hepatitis B e antigen (HBeAg) negative patients with acute hepatitis but not in HBsAg+ HBeAg- dialysis patients. In acute hepatitis, HBsAg immunoreactivity by M-RIA could still be detected even though a commercial immunoassay for HBsAg, the AUSRIA II, and the HBV DNA assay were no longer positive. Unlike in acute HBV infection, serum HBV DNA was detectable in dialysis patients who were AUSTRIA II negative but M-RIA positive. Serial determination of HBsAg by M-RIA and HBV DNA revealed episodes of HBV DNA positivity months after both the HBsAg was no longer positive by polyclonal immunoassay. Thus, the M-RIA for HBsAg and the molecular hybridization technique for HBV DNA are sensitive and specific assays for the identification of potentially infectious individuals who would not have been characterized as such based on the results of conventional polyclonal immunoassays.

Growth hormone-binding protein in partially hepatectomized rats

The role of the liver in regulating serum growth hormone-binding protein (GH-BP) was studied. We measured rat serum GH-BP and insulin-like growth factor 1 (IGF-1) 30 min to 96 h after 70% partial hepatectomy (PHP) or sham operation in adult male rats. Serum GH-BP declined sharply from 5.8 +/- 0.1% at baseline to 3.9 +/- 0.5% by 48 h following PHP. By 72 h serum GH-BP at baseline to 3.9 +/- 0.5% by 48 h following PHP. By 72 h serum GH-BP returned to baseline level and remained at that level 96 h postoperatively. In sham-operated female rats, serum GH-BP was about 2-fold higher than in males (10.5 +/- 1.46 versus 5.8 +/- 0.2%), whereas 24 h after hepatectomy a significant drop of about 50% was observed (p < 0.001). Serum IGF-1 decreased within 2-4 h postoperatively in both sham-operated and PHP groups, but thereafter was lower in the PHP rats, up to 48 h after operation, compared to sham-operated rats (p < 0.03). The study shows that the liver has an important role in the determination of serum GH-BP levels. The return to normal GH-BP level, even before the liver regained its full size following hepatectomy, suggests an increase in GH-BP production by the regenerating liver.