Biochemical markers of type III and I collagen: association with retinopathy and neuropathy in type 1 diabetic subjects.

AIMS: Connective tissue alterations may contribute to the development of diabetic long-term complications in eyes, kidneys and peripheral nerves. Collagen deposition may be increased in micro- and macrovascular disease in diabetic subjects. We tested whether biochemical markers of type III and I collagen metabolism are associated with retinopathy and neuropathy in Type 1 diabetes. METHODS: A total of 28 patients, mean age 43.4 +/- 9.5 (sd) and duration of diabetes 25.2 +/- 9.7 years, were studied. Stereoscopic colour fundus photographs were taken for assessment of retinopathy which was classified as no, background or proliferative. Concentrations of aminoterminal propeptide of type III procollagen (PIIINP), carboxyterminal propeptide of type I procollagen (PICP) and carboxyterminal cross-linked telopeptide of type I collagen (ICTP) in serum and urinary excretion of cross-linked N-telopeptides of type I collagen (NTX) and deoxypyridinoline crosslinks (DPyr) into urine were measured. RESULTS: Average serum PIIINP was higher in subjects with proliferative (3.2 +/- 1.1 microg/l) than without proliferative retinopathy (2.5 +/- 0.6 microg/l) (P = 0.03). Average serum PICP was higher in subjects without retinopathy (181.7 +/- 19.5 microg/l) than in subjects with background retinopathy (132.1 +/- 42.7 microg/l) (P = 0.02). Concentrations of other collagen markers were not different in subjects with or without retinopathy. No association between collagen markers and neuropathy was found. CONCLUSIONS: The increased synthesis of type III collagen, reflecting deposition of matrix and basement membrane connective tissue, may be involved in the pathogenesis of proliferative retinopathy in Type 1 diabetic subjects. On the other hand, we observed decreased synthesis of Type I collagen, which can result in weakened vascular integrity in subjects with retinopathy.     

Type VI collagen gene expression in experimental liver fibrosis: quantitation and spatial distribution of mRNAs,and immunodetection of the protein

Type VI collagen is a minor but essential matrix component in the liver. In this study, we utilized an acute and a chronic injury model to clarify the process of liver fibrosis in rats by administration of carbon tetrachloride. Collagen gene expression, with particular emphasis on type VI collagen, was studied by molecular hybridization techniques. The α2(VI) collagen mRNA levels were markedly elevated on day 3 of acute injury and were approximately at the same high level at 7 and 14 weeks of chronic injury, as determined by Northern hybridizations and slot-blot analyses. Marked enhancement of type I collagen gene expression was similarly noted at these time points The activation of collagen gene expression in acute injury, as determined by in situ hybridization, was particularly prominent in the vicinity of the central veins. Indirect immunofluorescence demonstrated marked accumulation of type VI collagen protein as early as day 3 of acute injury, and the reaction appeared to be initiated in the proximity of central veins. These results indicate that type VI collagen gene expression, together with other connective tissue components, including type I collagen, is activated in the early stages of the fibrotic process. Type VI collagen accumulation may contribute to the distorted architecture and functional impairment of the liver in hepatic fibrosis.     

Localisation and semiquantitative assessment of hepatic procollagen mRNA in primary biliary cirrhosis

Background—Chronic liver disease ischaracterised by excessive deposition of collagen and otherextracellular matrix proteins, produced mainly, but not exclusively, byactivated hepatic stellate cells in the perisinusoidal space. Inprimary biliary cirrhosis (PBC) fibrosis is concentrated mainly aroundthe portal tracts.
Aims—To examine the hypothesisthat, in addition to hepatic stellate cells, portal tract fibroblastsmight play a significant role in the deposition of collagen in PBC.
Methods—Fifty liver biopsyspecimens from patients with PBC were studied. An in situ hybridisationtechnique was adapted to localise and measure semiquantitatively type Iprocollagen mRNA in formalin fixed, paraffin wax embedded sections,using an ³⁵S labelled cRNA probe specific for theα₁ chain of rat type I procollagen. Hepatic fibrogenicactivity was also assessed using serum type III procollagen peptide (PIIINP).
Results—In PBC, type I procollagengene expression was significantly increased. Signal was localisedmainly in and around inflamed portal tracts, to cells which had theappearances of portal fibroblasts. Signal activity in these cellscorrelated with the degree of portal fibrosis and inflammation and alsowith serum PIIINP concentrations.
Conclusions—Results are consistentwith the hypothesis that the excessive extracellular matrix, depositedwithin the liver in PBC, is synthesised not only by hepatic stellatecells but also by portal tract fibroblasts. The semiquantitativeassessment of procollagen mRNA in liver biopsy specimens may provide auseful method of evaluating the rate of synthesis of collagen andtherefore disease activity in patients with PBC.

Keywords:in situ hybridisation; procollagen peptide; primarybiliary cirrhosis; hepatic fibrosis

     

The Relationship Between Collagen Proportionate Area and Hepatitis B Surface Antigen Levels in E Antigen Positive Hepatitis B Cirrhosis

BackgroundQuantitative serum hepatitis B surface antigen (HbsAg) has been widely used as a biomarker for treatment response and prognosis in chronic hepatitis B infection, and has recently been found associated with liver histology in e-antigen positive patients. A histological measurement as a continuous variable—collagen proportionate area (CPA)—is appropriate to assess liver fibrosis degree and substages cirrhosis. We, therefore, aimed to explore the association between serum quantitative HBsAg and CPA in e antigen-positive hepatitis B cirrhosis.MethodsLiver fibrosis staging was evaluated by METAVIR semiquantitative scoring system, only patients with METAVIR fibrosis stage 4 were included. All liver sections were stained with picroSirius red for determination of collagen quantification by digital image analysis.ResultsMean CPA value was 23.46%. The percentage of patients with different classification of CPA (<20%, 20-30%, >30%) were 25.8%, 57.8%, and 16.4%, respectively. A modest correlation was found between CPA and serum HBsAg level (r = _–0.306, P =.001). Hepatitis B surface antigen level is independently associated with CPA in multivariable linear regression analyses.ConclusionSerum HBsAg levels can predict liver fibrosis determined by CPA in HBeAg-positive hepatitis B cirrhosis.     

Serum type IV collagen in various liver diseases in comparison with serum 7S collagen,laminin, and type III procollagen peptide

The clinical significance of the immunoreactive triple helical domain of type IV collagen in serum was evaluated in 73 healthy controls and 161 patients with various biopsy-proven liver diseases. Although serum levels of type III procollagen peptide were increased in all liver diseases, those of type IV collagen, 7S collagen, and laminin were principally increased in chronic liver diseases associated with hepatic fibrogenesis/fibrosis. In both non-alcoholic and alcoholic liver diseases, 7S collagen was increased in serum, while type IV collagen and laminin in serum were particularly increased in alcoholic liver diseases and in hepatocellular carcinoma, in which latter the sensitivity was greater for type IV collagen than for laminin. Gel filtration analysis in Sephacryl S-400 revealed type IV collagen in serum to be a single molecular form with a molecular weight that correspond to type IV collagen, whereas 7S collagen was recognized as several heterogeneous macromolecules. These findings indicate that serum type IV collagen is derived from the type IV protocollagen pool, and is a sensitive marker for the fibrogenetic process in hepatic basement membranes.     

Serum amino-terminal propeptide of type III procollagen and 7S domain of type IV collagen correlate with hepatic iron concentration in patients with chronic hepatitis C following alpha-interferon therapy

BACKGROUND: It has been reported that chronic infection with hepatitis C virus is associated with excess iron deposits in the liver of subjects who are neither alcoholics nor recipients of blood transfusions. However, little is known about the relationship between hepatic iron concentration (HIC) and the serum levels of hepatic fibrogenesis markers, which were caused by interferon therapy for chronic hepatitis C. Therefore, changes in the serum amino-terminal propeptide of type III procollagen (P-III-P) and the 7S domain of type IV collagen (7S-IV) in 16 patients treated with alpha-interferon (IFN-alpha) were studied, and their HIC and histological assessment evaluated. Hepatic iron concentrations were measured by using liver biopsy specimens obtained before and 6 months after the cessation of treatment. METHODS AND RESULTS: Eight subjects (50%) who had normal alanine transaminase levels at 6 months after therapy showed significantly lowered HIC, and attenuated hepatic iron staining with decreased serum levels of P-III-P and 7S-IV compared to the remaining subjects. The HIC was significantly correlated with the serum levels of P-III-P and 7S-IV in all subjects. CONCLUSIONS: These findings suggest that IFN-alpha treatment may decrease stimuli for fibrogenesis, at least in part, by reducing the hepatic iron deposition in patients with chronic hepatitis C.     

Colchicine reduces procollagen III and increases pseudocholinesterase in chronic liver disease

AIM: To test whether colchicine would be an effective antifibrotic agent for treatment of chronic liver diseases in patients who could not be treated with α-interferon.METHODS: Seventy-four patients (46 males, 28 females) aged 40-66 years (mean 53 ± 13 years) participated in the study. The patients were affected by chronic liver diseases with cirrhosis which was proven histologically (n = 58); by chronic active hepatitis C (n = 4), chronic active hepatitis B (n = 2), and chronic persistent hepatitis C (n = 6). In the four patients lacking histology, cirrhosis was diagnosed from anamnesis, serum laboratory tests, esophageal varices and ascites. Patients were assigned to colchicine (1 mg/d) or standard treatment as control in a randomized, double-blind trial, and followed for 4.4 years with clinical and laboratory evaluation.RESULTS: Survival at the end of the study was 94.6% in the colchicine group and 78.4% in the control group (P = 0.001). Serum N-terminal peptide of type III procollagen levels fell from 34.0 to 18.3 ng/mL (P = 0.0001), and pseudocholinesterase levels rose from 4.900 to 5.610 mU/mL (P = 0.0001) in the colchicine group, while no significant change was seen in controls. Best results were obtained in patients with chronic hepatitis C and in alcoholic cirrhotics.CONCLUSION: Colchicine is an effective and safe antifibrotic drug for long-term treatment of chronic liver disease in which fibrosis progresses towards cirrhosis.     

Non-Specific Iron in Patients with Beta-Thalassaemia Trait and Chronic Active Hepatitis

A non-specific iron fraction, not bound to transferrin, has been looked for in the sera of 42 never-transfused patients with beta-thalassaemia trait, 17 of whom had chronic active hepatitis, negative for HBV infection or alcohol abuse. Non-specific iron was found only in the sera of those patients with beta-thalassaemia trait plus chronic active hepatitis who had complete transferrin saturation, high serum ferritin levels and urinary iron excretion and a high degree of hepatic siderosis. In view of the known toxicity of non-transferrin iron, we suggest that this non-transferrin iron fraction may be responsible for the liver damage in these patients. Furthermore, the positive correlation between the presence and the amount of non-transferrin iron and the levels of serum ferritin suggests that this fraction is a sensitive indicator of iron-induced toxicity when severe iron overload slowly develops in patients with beta-thalassaemia trait even in the absence of any iron administration.     

人原发性肝癌及癌旁肝组织中Ⅰ、Ⅲ型胶原的基因表达

用cDNA-mRNA原位核酸杂交技术检测了2例正常人肝及12例原发性肝癌及癌旁肝组织中Ⅰ、Ⅲ型胶原的基因表达,结果发现8例原发性肝细胞癌组织中多种细胞,包括瘤细胞都有胶原基因的表达,证实人原发性肝癌组织中不仅间质细胞而且肿瘤细胞也参与胶原的合成。胶原基因的过度表达可能与细胞的活跃增生有关。     

Risk Factors for Bone Loss in Chronic Active Hepatitis and Primary Biliary Cirrhosis

Olsson R, Johansson C, Lindstedt G, Mellström D. Risk factors for bone loss in chronic active hepatitis and primary biliary cirrhosis. Scand J Gastroenterol 1994;29:753-756.

Background: Data on risk factors for bone loss in chronic active hepatitis (CAH) and primary biliary cirrhosis (PBC) are scanty and/or conflicting. Methods: Bone mineral density (BMD) in the distal forearm was measured using single-photon absorptiometry in 39 patients with CAH and 32 patients with PBC. We also attempted to identify risk factors for bone loss by means of a questionnaire and through a wide range of biochemical analyses. Results: In the CAH patients BMD is inversely related to the duration of steroid treatment and to age at menarche. In the PBC patients there was a strong correlation between BMD and serum gastrin concentrations. Conclusions: Bone loss in CAH is to some extent explained by steroid treatment and delayed menarche. Bone loss in PBC may be reduced by increased calcitonin secretion induced by gastrocalcin.     

Correlation Between Peritoneoscopic Findings and Response to Interferon in Patients with Chronic Hepatitis C

This study was designed to investigate the correlation between peritoneoscopic liver surface findings and the alanine aminotransferase (ALT) response to interferon, and to identify peritoneoscopic findings potentially useful as predictors of interferon responsiveness, in chronic hepatitis C. In all, 119 patients with hepatitis C viremia and histological evidence of chronic hepatitis received interferon alpha, in total doses of 258 to 660 million units (MU). A sustained response was seen in 29 cases (24%), a transient response with ALT relapse after therapy completion in 41 (35%) and no response in 49 (41%). Among 64 type 1 b cases, 8 (13%) showed sustained, 27 (42%) transient and 29 (45%) no response. The sustained response rate was higher with Shimada's code 200 than with code 300 or 400, even for type 1b. Interferon therapy was ineffective in more than 50% with code 300 or 400, especially in type 1b, suggesting that interferon efficacy decreases with disease progression. The sustained response rate was significantly higher, and that of no response significantly lower, in patients without reddish liver surface markings than in those with such markings (p<0.05), even in type lb. About 50% of those with reddish markings showed no response, indicating that absence of reddish markings may be a useful predictor of sustained response. The liver fibrosis score was significantly higher in the no response than in the transient and sustained response groups (p<0.02). In summary, patients with reddish liver surface markings and/or advanced fibrosis do not show a good response to interferon.     

Serum levels of tissue inhibitor of metalloproteinases-2 and of precursor form of matrix metalloproteinase-2 in patients with liver disease

Abstract: Serum levels of tissue inhibitor of metalloproteinases-2 (TIMP2) and of precursor form of matrix metalloproteinase-2 (proMMP2) were determined in patients with chronic hepatitis and hepatocellular carcinoma by a one-step sandwich enzyme immunoassay. Serum levels of TIMP2 and proMMP2 were significantly higher in patients with chronic liver disease, than in normal controls. Serum levels of TIMP2 showed a weak negative correlation with the serum albumin level and prothrombin time (PT). Serum levels of proMMP2 in patients with chronic hepatitis were strongly correlated with those of type IV collagen and were negatively correlated with PT and serum albumin levels. Serum proMMP2 levels were also significantly correlated with histological stages. These data indicate that serum levels of proMMP2 might be useful in the follow-up of patients with chronic hepatitis.     

Measurement of carboxy terminal peptide of type I procollagen in sera of patients with viral hepatitis and liver cirrhosis

In the present investigation, a radioimmunoassay for carboxy terminal peptide of human type I procollagen (type 1 C-peptide) was developed. Its clinical implication for serodiagnosis of hepatic fibrosis in 85 patients with viral hepatitis, 45 patients with post-hepatitic liver cirrhosis and 37 patients with alcoholic liver diseases was evaluated in comparison with that of the previously established amino terminal peptide (type III N-peptide) assay. Anti-sera against type I procollagen was obtained by immunization of rabbit with purified type I procollagen from culture medium of IMR-90. The serum level of type I C-peptide in normal subjects was found to be 42 ng/ml (s.d. = 19). Type I C-peptide levels in patients with acute hepatitis were within normal range, while in chronic hepatitis, the mean type I C-peptide level increased as the grade of fibrosis advanced from grade I to III. However, there was no statistically significant difference between the mean type I C-peptide level of grade III and that of liver cirrhosis. Increments of type I C-peptide levels were also observed in alcoholic liver fibrosis (fatty liver with fibrosis and liver cirrhosis). On the other hand, type III N-peptide assay appeared to reflect not only the degree of hepatic fibrosis, but also the degree of hepatic inflammation, giving the high levels in acute viral hepatitis. Collectively, the results indicate the usefulness of type I C-peptide assay for monitoring hepatic fibrosis in viral hepatitis as well as in alcoholic liver disease.     

Clinical evaluation of serum tissue inhibitor of metalloproteinases-1 levels in patients with liver diseases

Serum levels of the tissue inhibitor of metalloproteinases-1 (TIMP-1) were measured in 268 patients with liver diseases by means of a one-step sandwich enzyme immunoassay. In the cases of acute hepatitis, chronic active hepatitis (CAH), liver cirrhosis (LC) and hepatocellular carcinoma (HCC), the levels of TIMP-1 were higher than those of the control group. Tissue inhibitor of metalloproteinases-1 levels correlated with type III procollagen peptide and with type IV collagen, indicating TIMP-1 as a useful marker for hepatic fibrosis. Levels of TIMP-1 also correlated with aspartate aminotransserase and alanine aminotransferase levels and showed the highest levels in acute hepatitis. Thus, TIMP-1 might also reflect hepatic inflammation. Serum levels of α-fetoprotein and TIMP-1 had a significant positive correlation in patients with HCC. A cut-off level of TIMP-1 between LC and HCC was set at 440 ng/mL, having a low sensitivity and a high specificity. These results suggest the usefulness of TIMP-1 as a tumour marker in cases of HCC where α-fetoprotein levels are not elevated.     

The Superiority of Laparoscopic Examination in Predicting Hepatocellular Carcinoma after Inteferon Therapy for Chronic Type C Hepatitis

Abstract: To explain the appearance rates of hepatocellular carcinoma (HCC) in patients treated with interferon (IFN) and to assess the risk factors for hepatocellular carcinogenesis, we studied 442 patients with chronic type C hepatitis for one to 7.6 years (median 2.8 years). Of these 442 patients, 135 (30.5%) showed a complete response with persistent HCV-RNA negativity for more than six months after completing IFN therapy. During the observation period, HCC developed in 16 patients. We divided the patients into three groups based on laparoscopic findings of the liver surface : smooth, irregular, and nodular. The cumulative appearance rates of HCC in these three liver types were zero, 6.6% and 14%, respectively, at the end of the fifth year. As to the efficacy of IFIM, the HCC appearance rate in patients who responded was lower than that in patients who did not (p<0.05). However, multivariate Cox regression analysis showed that the laparoscopic findings correlated most strongly with liver carcinogenesis. Therefore, we conclude that laparoscopic findings before IFN therapy are significantly associated with liver carcinogenesis over the short term observation period after IFN therapy.     

Role of cell-cycle turnover and oxidative stress in telomere shortening and cellular senescence in patients with chronic hepatitis C

BACKGROUND: In addition to the telomere shortening that occurs with cell division, oxidative stress can damage or shorten telomeres and induce a condition termed premature senescence, possibly before telomeres become critically short. We investigated the effects of cell-cycle turnover and oxidative stress on cellular senescence in hepatitis C virus (HCV)-related chronic liver injury. METHOD: Using quantitative fluorescence in situ hybridization, the telomere lengths of hepatocytes in biopsy specimens from HCV-positive patients were estimated. We assessed clinicopathological parameters that reflect cell-cycle turnover, including Ki-67 positive index, serum alanine aminotransferase (ALT) level and degree of fibrosis, and also oxidative stress-related parameters, such as 8-hydroxy-2'-deoxyguanosine (8-OHdG) expression. Nuclear size and DNA content of hepatocytes were measured as morphological features of senescence. RESULTS: Telomere shortening correlated with the degree of cell turnover, hepatic fibrosis and morphological features of aging cells. Furthermore, the rate of telomere shortening per year was positively correlated with fibrosis progression. In cases of no or mild fibrosis, telomere lengths of positive patients were generally shorter than those of 8-OHdG-negative patients, and this trend achieved statistical significance in advanced-stage fibrosis. HCV carriers with persistently normal serum ALT level (PNAL) showed significantly longer telomeres than patients with active hepatitis and mild fibrosis. There was no significant difference in telomere lengths between HCV carriers with PNAL and normal controls. CONCLUSIONS: Cell-cycle turnover is the primary mechanism of telomere shortening, and can induce fibrosis progression and cellular senescence. However, oxidative stress can be an accelerator of senescence, especially in advanced-stage fibrosis.     

Comparison of Acute Physiology and Chronic Health Evaluation III and II for Prediction of Mortality in Multiple Organ Failure Patients Treated by Plasma Exchange

Abstract: This study examined the efficacy of a different scoring system for multiple organ failure (MOF) patients treated by plasma exchange (PE). Twenty-five patients with MOF resulting from fulminant hepatitis who had been treated by PE for a total number of 91 PEs in a consecutive period of 22 months were included in our study. Data were collected from each patient to compute acute physiology and chronic health evaluation (APACHE) II and APACHE (AP) III scores. The sensitivity, specificity, and correct prediction of outcomes were measured. The correct prediction of outcomes was 86.5%in AP III and 77.5% in AP II. For the prediction of patient mortality, the AP III system seems to be more reliable than AP II.—     

慢性乙型肝炎患者肝组织中巨噬细胞移动抑制因子的表达及其对炎症活动的影响

目的:检测巨噬细胞移动抑制因子(macrophage migrationinhibitory factor,MIF)在正常肝和慢性乙型肝炎(chronic hepatitis B,CHB)肝组织中表达的差异,探讨MIF的表达与CHB炎症活动及病毒载量的关系。方法:取CHB40例为实验组(HBeAg阳性CHB28例,HBeAg阴性CHB12例),正常肝组织20例为对照组,用免疫组化和原位杂交技术检测两组中MIF和MIFmRNA的表达,并测定患者ALT和AST的水平及HBV DNA载量。结果:MIF和MIF mRNA在实验组中的阳性表达例数明显高于对照组,两组相比差异有显著性意义(P<0.05);实验组肝组织中MIF和MIF mRNA表达程度与ALT和AST呈正相关,1相似文献   

Elevated carboxy terminal cross linked telopeptide of type I collagen in alcoholic cirrhosis: relation to liver and kidney function and bone metabolism

BACKGROUND: The carboxy terminal cross linked telopeptide of type I collagen (ICTP) has been put forward as a marker of bone resorption. Patients with alcoholic liver disease may have osteodystrophy. AIMS: To assess circulating and regional concentrations of ICTP in relation to liver dysfunction, bone metabolism, and fibrosis. METHODS: In 15 patients with alcoholic cirrhosis and 20 controls, hepatic venous, renal venous, and femoral arterial concentrations of ICTP, and bone mass and metabolism were measured. RESULTS: Circulating ICTP was higher in patients with cirrhosis than in controls. No overall significant hepatic disposal or production was found in the patient or control groups but slightly increased production was found in a subset of patients with advanced disease. Significant renal extraction was observed in the controls, whereas only a borderline significant extraction was observed in the patients. Measurements of bone mass and metabolism indicated only a mild degree of osteodystrophy in the patients with cirrhosis. ICTP correlated significantly in the cirrhotic patients with hepatic and renal dysfunction and fibrosis, but not with measurements of bone mass or metabolism. CONCLUSIONS: ICTP is highly elevated in patients with cirrhosis, with no detectable hepatic net production or disposal. No relation between ICTP and markers of bone metabolism was identified, but there was a relation to indicators of liver dysfunction and fibrosis. As the cirrhotic patients conceivably only had mild osteopenia, the elevated ICTP in cirrhosis may therefore primarily reflect liver failure and hepatic fibrosis.