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Insulin-Like Growth Factor Binding Protein-3: A Novel Biomarker for the Assessment of the Synthetic Capacity of Hepatocytes in Liver Cirrhosis

Department of Obstetrics & Gynecology (M.S.), Faculty of Medicine, Cairo University; Department of Medicine (M.A.F., B.A.M., S.L.), Ain Shams University, Cairo, Egypt

Address correspondence and requests for reprints to: Professor Mohamed Shaarawy, 21 El-Khalifa El-Maamoun, Roxy Building, Apt 701, Heliopolis, Cairo, Egypt.

	Abstract

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The liver is the major source of circulating insulin-like growth factor binding protein-3 (IGFBP-3). Because the hepatic tissue is deranged in cirrhotic patients, we measured serum IGFBP-3 concentrations by two-site immunoradiometric assay in sera from 37 cirrhotic patients with different stages of hepatic dysfunction. These were compared with IGFBP-3 levels from 11 healthy controls. Serum IGFBP-3 levels in patients with chronic liver disease were significantly lower than those of the control group (P < 0.0005). The mean percent decrease in cases of early liver cirrhosis, cirrhosis without, and cirrhosis with ascites were 44%, 59%, and 82% respectively, indicating that serum IGFBP-3 levels decrease as the severity of hepatic dysfunction increases.

Moreover, the decrease was more pronounced in cases with hyperbilirubinemia, elevated serum transaminases, hypoalbuminemia, and prolonged prothrombin time. There was a significant positive correlation between serum IGFBP-3 and serum albumin, as well as a significant negative correlation between serum IGFBP-3 and prothrombin time. These results indicate the close correlation of IGFBP-3 levels to worsening of hepatic functions. The determination of serum IGFBP-3 level is a clinically useful marker for the assessment of the synthetic capacity of hepatocytes in cirrhotic patients and an early predictor of impending hepatic dysfunction as well. .

	Introduction

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THE LIVER is the major source of insulin-like growth factor-1 (IGF-1). It has also been suggested to be a major source of at least two of the main binding proteins (BP) that modify the bioavailability of IGF-1 (1). IGFBP-3 binds most circulating IGFs (2) and, in association with IGF and an acid-labile protein subunit forms a stable tertiary complex (3) that prolongs IGF’s half-life. It also functions as a reservoir for IGFs and restricts their extravascular transit (4). Growth hormone (GH) has been recognized as an important regulator of serum IGFBP-3. A large study in healthy children and adolescents showed that IGFBP-3 levels reflected spontaneous GH secretion (5). In Laron-type GH insensitivity, a condition thought to be due to the absence of functional GH receptors, very low levels of IGFBP-3 were observed (6).

Six IGFBPs have been isolated and chemically characterized (7). In contrast to IGFBP-3, IGFBP-1 levels fluctuate markedly throughout the 24-h period. More than 98% of IGFs in the serum circulate bound to IGFBP-3 (8), which may also be confirmed by the fact that IGFBP-3 has the highest serum concentration among all IGFBPs. In chronic hepatic disease there are significant changes in the levels of IGFBP-1 and IGFBP-3 (9). IGF-1, IGF-2, and IGFBPs are reduced in plasma from growth-retarded children with chronic hepatic disease (10). The close correlation of IGFBP-3 levels with hepatic functions indicates a dominant regulatory role of the hepatocytes (9). In this study circulating levels of IGFBP-3 were determined in patients with chronic hepatic disease of varying degrees of severity to assess the direct effect of hepatic dysfunction on the biosynthesis of IGFBP-3 by hepatocytes. We have also examined the interrelationship between IGFBP-3 and the conventional hepatic function tests.

	Subjects and Methods

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Thirty-seven patients with hepatic cirrhosis were selected to participate in this study. All patients had histologically proven cirrhosis. They were adult males, and their ages ranged from 25–50 yr. They were segregated into 3 groups. The first group consisted of 13 patients with early hepatic cirrhosis. The second group consisted of 13 patients with hepatic cirrhosis and portal hypertension associated with disturbed hepatic function tests (low serum albumin, elevated serum transaminases, and prolonged prothrombin time). The third group consisted of 11 patients with decompensated hepatic cirrhosis (ascites, splenomegaly, portal hypertension). We excluded any patient with cardiac, respiratory, or renal dysfunction. Diabetic patients were also excluded. All patients were biochemically euthyroid. Patients with recent alcohol intake, corticosteroid therapy, vitamin K, or diuretics were also excluded. Patients with evidence of recent systemic infection or active vareceal bleeding were excluded. Patients were studied on the hospital ward and had their laboratory workup done after one overnight fast. Before the study, patients consumed a standard diet that was not modified for any research purposes. A control group consisted of 11 healthy volunteers. None was alcoholic or addicted to drugs or suffered from schistosomiasis, hepatitis, or any other medical condition. For all patients and controls height, body weight, and span were assessed. Vital data (blood pressure, radial pulse, etc.) were recorded. Clinical evaluation for signs suggestive of hepatic disease (jaundice, spider nervi, palmar erythema, lower limb edema, pallor) and of the abdomen (liver span, splenic size, ascites), chest, cardiac, and neurological conditions were recorded.

Sampling and laboratory

Fasting blood samples were obtained from the cubital vein at 0900 h. Immediately after venepuncture sera were separated by centrifugation and stored in a deep freeze at -60C until analysis. Serum IGFBP-3 was determined by a two-site immunoradiometric assay (IRMA) using DSL-6600 IGFBP-3 coated-tube IRMA kit (Diagnostic System Laboratories, Webster, TX). The intra- and interassay coefficient of variation were 2.6% and 3.1%, respectively. The minimal detection limit of the assay was 0.5 ng/mL. Conventional hepatic function tests were performed on an Hitachi clinical chemistry autoanalyzer (Boehringer Mannheim, Germany). Prothrombin time was assessed using standard thromboplastin.

Statistical analysis

IGFBP-3 levels in the patient group were not normally distributed. Statistical analyses were performed using nonparametric methods (unpaired, two-tailed Student’s t test). The criterion of significance was a value of P < 0.05. Descriptive data are expressed as mean ± SD. Correlation studies were determined by Spearman rank order correlations. Sensitivity, specificity, and predictive values of positive and negative tests were calculated according to Galen and Gambino (11).

	Results

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Serum IGFBP-3 level in cases of early liver cirrhosis was significantly lower than that of healthy controls (P < 0.0005), the mean percent decrease of serum IGFBP-3 in the pathologic group amounted to 44% (Table 1). The 95% confidence limits of the healthy reference group was 2120–5840 ng/mL. The incidence of abnormal low values of serum IGFBP-3 in cases of early liver cirrhosis (values below 2120 ng/mL) was 38.5%. Serum IGFBP-3 level in cases of liver cirrhosis without ascites was significantly lower than that of healthy controls (P < 0.0005), the mean percent decrease of serum IGFBP-3 in the pathologic group without ascites amounted to 59%. The incidence of abnormal low values of serum IGFBP-3 in cases of liver cirrhosis without ascites was 84%. Serum IGFBP-3 level in cases of liver cirrhosis with ascites was significantly lower than that of healthy controls (P < 0.0005). The mean percent decrease of serum IGFBP-3 in the pathologic group with ascites amounted to 82%. The incidence of abnormal low values of serum IGFBP-3 in cases of liver cirrhosis with ascites was 100% (all cases). Serum IGFBP-3 level in cases of liver cirrhosis with elevated serum bilirubin was significantly lower than that of cases of liver cirrhosis without elevated serum bilirubin (P < 0.0005). The mean percent decrease of serum IGFBP-3 in cases of liver cirrhosis with elevated serum bilirubin amounted to 52% (Table 2). Serum IGFBP-3 level in cases of liver cirrhosis with elevated transaminases was significantly lower than that of cases of liver cirrhosis without elevated transaminases at (P < 0.0005). The mean percent decrease of serum IGFBP-3 in liver cirrhosis with elevated SGPT amounted to 45%. Serum IGFBP-3 level in cases of liver cirrhosis with low serum albumin (<3.4 g/dL) was significantly lower than that of cases of liver cirrhosis without low serum albumin (P < 0.0005). The mean percent decrease of serum IGFBP-3 in liver cirrhosis with low serum albumin amounted to 42%. Serum IGFBP-3 level in cases of liver cirrhosis with prolonged prothrombin time was significantly lower than that of cases of liver cirrhosis without prolonged prothrombin time (P < 0.0005). The mean percent decrease of serum IGFBP-3 in liver cirrhosis with prolonged prothrombin time amounted to 55%.

View larger version (38K): [in this window] [in a new window]   Figure 1. Correlation coefficient "r" and regression line equation between the individual values of serum IGFBP-3 and liver function tests.

	Discussion

Top Abstract Introduction Subjects and Methods Results Discussion References

 
The liver is the main source of most serum proteins (12). In patients with hepatic cirrhosis, a reduction of serum albumin and other plasma proteins is a common finding. On the other hand, endocrine functions are often disturbed in patients with chronic hepatic disease (13). Growth hormone (GH) reserves in these patients were defective. Retarded linear growth during the growing period was caused by the interplay of many factors, e.g. diminished somatomedin C synthesis due to derangement of hepatic cellular functions, defective growth hormone reserves, as well as disturbed feedback mechanisms of somatomedin on the hypothalamic hypophyseal axis. The liver is the main site where synthesis of insulin-like growth factors (IGF) takes place (14). IGF-1 (somatomedin C) is more GH-dependent and more potent as a growth promotor than IGF-2. It has been established that more than 98% of IGFs in serum circulate bound to IGFBP-3 (8). The concentration of IGFBP-3 is highest in serum, the liver being its major source. Although many cell types have been shown to produce IGFBP-3 (15), the liver is by far the most significant source of IGFBP-3 production (9). In vitro studies have demonstrated IGFBP-3 production from human fetal explants (16), but not from isolated hepatocytes in primary culture, suggesting production from nonparenchymal cells (16).

Zapf et al. (7) and Donaghy et al. (9) reported that serum IGFBP-3 levels were abnormally low in patients with hepatic cirrhosis. Our study has also revealed a significant decrease of serum IGFBP-3 level even in patients with early hepatic cirrhosis when compared with that of healthy controls. Patients with more advanced cirrhosis without or with ascites showed more pronounced lower levels of serum IGFBP-3 than those of patients with early hepatic cirrhosis. This finding would suggest that the estimation of serum immunoreactive IGFBP-3 level could be a valuable and early marker of hepatic dysfunction that would parallel and compliment the Child’s clinical classification (13) of hepatic cirrhosis, which is also a prognostic indicator. To confirm this assumption we also compared changes in serum IGFBP-3 levels in the different stages of cirrhosis with other commonly used hepatic function tests. Serum IGFBP-3 levels were significantly lower in cirrhotic patients with elevated serum bilirubin than in those without significant changes in serum bilirubin. There was a significant negative correlation between serum IGFBP-3 and serum bilirubin levels. Bilirubin level may not become elevated except later in the course of many cases of cirrhosis (12). Even if bilirubin levels were elevated, the list of possible causes of its elevation would complicate the differential diagnosis, thus rendering bilirubin not a useful marker for assessing the degree of dysfunction in chronic hepatic disease, especially when compared with IGFBP-3 as a diagnostic or staging parameter.

Serum IGFBP-3 level was lower in cirrhotic patients with elevated transaminases than in cirrhotic patients without elevated transaminases. There was a significant negative correlation between serum IGFBP-3 and serum transaminases levels. Moreover, serum IGFBP-3 levels were found to be significantly lower in cirrhotic patients with hypoalbuminemia than in those without hypoalbuminemia. There was also a significant positive correlation between serum IGFBP-3 and serum albumin levels. Thus serum IGFBP-3 level seem to be as good as a marker for the hepatic synthetic capacity as serum albumin estimates. In addition, serum IGFBP-3 level demonstrated a significant decrease in cirrhotic patients with prolonged prothrombin time (PT) when compared with that of cirrhotic patients with normal prothrombin time. There was a significant negative correlation between serum IGFBP-3 levels and PT estimates. As with serum albumin level, IGFBP-3 measurement was as good a marker for hepatic function as PT measurements.

Serum IGFBP-3 seems to be a more sensitive marker than serum albumin for the assessment of the synthetic capacity of hepatocytes in liver cirrhosis. Fifty percent reduction in serum IGFBP-3 levels concords with no percent reduction in serum albumin levels in early liver cirrhosis. The sensitivity and specificity of the serum IGFBP-3 test in cases of liver cirrhosis was found to be 76% and 100% respectively. The predictive value for a positive IGFBP-3 test was 100%, whereas for a negative test it was 55%.

In summary, the present study demonstrates that IGFBP-3 levels are decreased in patients with chronic hepatic disease. This decrease is more pronounced in more advanced stages of hepatic cirrhosis. In addition serum IGFBP-3 level is significantly reduced in the early stages of the disease before the appearance of any abnormality in the conventional hepatic function tests. This may highlight the usefulness of IGFBP-3 as a novel biomarker in the early stages of hepatic cirrhosis. Moreover, the close correlation of IGFBP-3 levels to the worsening conventional hepatic function tests points to the central role of the hepatic cell mass in IGFBP-3 production. This further strengthens the reliability of serum IGFBP-3 as an early marker and predictor of liver cirrhosis. In conclusion, the estimation of IGFBP-3 in the serum is a clinically useful biomarker for the assessment of the biosynthetic capacity of hepatocytes in cirrhotic patients and is a more sensitive parameter by far than serum albumin measurement.

Received February 19, 1998.

Accepted May 19, 1998.

	References

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