Hypothermia predicts hepatic failure after extensive hepatectomy in mice

AIM: To investigate the effect of hypothermia on the function of the liver remnant (LR) after extended hepatectomy. METHODS: We performed a 75% partial hepatectomy (PH) in male C57BL/6J mice. Body temperature was measured with a rectal probe. The study mice were prospectively grouped as hypothermic (HT) or normothermic (NT) if their body temperature was < 34 °C vs≥ 34 °C, respectively. Blood and liver samples were obtained at 24 and 48 h after 75% PH. Various factors during and after 75% PH were compared at each time point and the most important factor for a good outcome after 75% PH was determined. RESULTS: At 24 and 48 h after 75% PH, LR weight was decreased in HT mice compared with that in NT mice and the assay results in the HT mice were consistent with liver failure. NT mice had normal liver regeneration. Each intra- and post-operative factor which showed statistical significance in univariate analysis was evaluated by multivariate analysis. The most important factor for a good outcome after 75% PH was body temperature at both 24 and 48 h after surgery. CONCLUSION: Hypothermia after an extensive hepatectomy predicts impending liver failure and may be a useful clinical marker for early detection of liver failure after extended hepatectomy.     

Cloning of transforming growth factor-beta 1 (TGF-beta 1) and its type II receptor from zebrafish ovary and role of TGF-beta 1 in oocyte maturation

TGF-beta is a multifunctional factor involved in regulating a variety of cellular activities. In mammals, TGF-beta is known to regulate reproduction, including ovarian functions. The role of TGF-beta in lower vertebrates, such as fish, is poorly understood. To examine the role of TGF-beta in fish reproduction, cDNAs encoding TGF-beta 1 and the type II TGF-beta receptor (T beta RII) were cloned from the zebrafish ovary using PCR- based strategies. The mature peptide region of the zebrafish TGF-beta 1 shows 70-85% identity with TGF-beta 1 from other species. The zebrafish T beta RII cDNA sequence is the first to be reported from a fish species, and it shows a high level of conservation at the kinase domain. Using RT-PCR, we have detected mRNA expression of TGF-beta 1, T beta RII, as well as its downstream signaling molecules Smad2, 3, and 4 in ovarian follicles at different stages of development. In addition, we have examined the effect of TGF-beta 1 on oocyte maturation. TGF-beta 1 significantly inhibited both gonadotropin- and 17 alpha, 20 beta-dihydroxyprogesterone-induced oocyte maturation in a dose- and time-dependent manner. These findings demonstrate, for the first time, that TGF-beta 1 plays a role in regulating oocyte maturation in fish and suggest that a TGF-beta/Smad signaling pathway is present in the zebrafish ovary.     

Role of transforming growth factor-beta in hematologic malignancies

The transforming growth factor-beta (TGF-beta) signaling pathway is an essential regulator of cellular processes, including proliferation, differentiation, migration, and cell survival. During hematopoiesis, the TGF-beta signaling pathway is a potent negative regulator of proliferation while stimulating differentiation and apoptosis when appropriate. In hematologic malignancies, including leukemias, myeloproliferative disorders, lymphomas, and multiple myeloma, resistance to these homeostatic effects of TGF-beta develops. Mechanisms for this resistance include mutation or deletion of members of the TGF-beta signaling pathway and disruption of the pathway by oncoproteins. These alterations define a tumor suppressor role for the TGF-beta pathway in human hematologic malignancies. On the other hand, elevated levels of TGF-beta can promote myelofibrosis and the pathogenesis of some hematologic malignancies through their effects on the stroma and immune system. Advances in the TGF-beta signaling field should enable targeting of the TGF-beta signaling pathway for the treatment of hematologic malignancies.     

Thrombospondin-1 is a novel negative regulator of liver regeneration after partial hepatectomy through transforming growth factor-beta1 activation in mice

The matricellular protein, thrombospondin-1 (TSP-1), is prominently expressed during tissue repair. TSP-1 binds to matrix components, proteases, cytokines, and growth factors and activates intracellular signals through its multiple domains. TSP-1 converts latent transforming growth factor-beta1 (TGF-β1) complexes into their biologically active form. TGF-β plays significant roles in cell-cycle regulation, modulation of differentiation, and induction of apoptosis. Although TGF-β1 is a major inhibitor of proliferation in cultured hepatocytes, the functional requirement of TGF-β1 during liver regeneration remains to be defined in vivo. We generated a TSP-1-deficient mouse model of a partial hepatectomy (PH) and explored TSP-1 induction, progression of liver regeneration, and TGF-β-mediated signaling during the repair process after hepatectomy. We show here that TSP-1-mediated TGF-β1 activation plays an important role in suppressing hepatocyte proliferation. TSP-1 expression was induced in endothelial cells (ECs) as an immediate early gene in response to PH. TSP-1 deficiency resulted in significantly reduced TGF-β/Smad signaling and accelerated hepatocyte proliferation through down-regulation of p21 protein expression. TSP-1 induced in ECs by reactive oxygen species (ROS) modulated TGF-β/Smad signaling and proliferation in hepatocytes in vitro, suggesting that the immediately and transiently produced ROS in the regenerating liver were the responsible factor for TSP-1 induction. CONCLUSIONS: We have identified TSP-1 as an inhibitory element in regulating liver regeneration by TGF-β1 activation. Our work defines TSP-1 as a novel immediate early gene that could be a potential therapeutic target to accelerate liver regeneration.     

Serum level of transforming growth factor-beta1 (TGF-beta1) and the expression of TGF-beta receptor type II in peripheral blood mononuclear cells in patients with autoimmune hepatitis

BACKGROUND/AIMS: To evaluate the association of the immunosuppressive effects of transforming growth factor-beta1 (TGF-beta1) with abnormalities in immune regulation in autoimmune hepatitis (AIH), we investigated the serum level of TGF-beta1 and expression of TGF-beta receptor type II (TbetaRII) in peripheral blood mononuclear cells (PBMC) in patients with AIH. METHODOLOGY: Twenty-two patients with AIH were included in this study. Serum levels of total TGF-beta1 were determined using a specific enzyme-linked immunosorbent assay (ELISA). The expression levels of TbetaRII mRNA were semi-quantitatively determined by ribonuclease (RNase) protection assay specific for TbetaRII. RESULTS: The mean serum level of TGF-beta1 in patients with AIH (230+/-95 ng/mL) was higher than that of healthy controls (137+/-81 ng/mL, p=0.012). The expression level of TbetaRII mRNA in PBMC obtained from AIH patients (0.131+/-0.046) was lower than that in PBMC of patients with chronic hepatitis C (0.186+/-0.074, p=0.019) and that of healthy subjects (0.188+/-0.060, p=0.013). CONCLUSIONS: The present study of TbetaRII mRNA expression in PBMC from patients with AIH suggested that the decreased expression of TbetaRII might contribute partially to abnormalities in immune regulation observed in patients with AIH, despite concomitant up-regulation of TGF-beta1 production.     

Induction of transforming growth factor-beta 1 (TGF-beta 1), receptor expression and TGF-beta 1 protein production in retinoic acid-treated HL-60 cells: possible TGF-beta 1-mediated autocrine inhibition.

Treatment of HL-60 cells, a human promyelocytic leukemia cell line, with the vitamin A derivative retinoic acid (RA) for 7 days resulted in a dose-dependent decrease in proliferation and increase in granulocytic differentiation. The role of transforming growth factor-beta 1 (TGF-beta 1), a protein with pleiotropic effects on the proliferation and differentiation of various cell types, was examined during RA-induced differentiation of HL-60 cells. Although TGF-beta 1 alone had little effect on proliferation or differentiation of HL-60 cells, addition of TGF-beta 1 to HL-60 cells treated with a suboptimum concentration of RA (1.0 nmol/L) resulted in a marked decrease in proliferation with no effect on granulocytic differentiation. Studies of the mechanism of RA-induced TGF-beta sensitivity showed that although untreated HL-60 cells expressed low levels of TGF-beta 1 binding proteins on the cell surface, the levels were increased in a dose-dependent manner after RA treatment. Maximum induction was achieved after treatment with 10 nmol/L RA and consisted predominantly of the 65-Kd TGF-beta 1 receptor type. Moreover, RA treatment also resulted in a dose-dependent increase in both TGF-beta 1 steady-state mRNA expression and production of active TGF-beta with maximum induction at 10 nmol/LRA. RA treatment of HL-60 cells had no effect on TGF-beta 2 and TGF-beta 3 mRNA expression. These data suggest that the effects of RA may be mediated by a TGF-beta 1-mediated autocrine antiproliferative loop during differentiation of HL-60 cells.     

Role of neutrophils in sinusoidal endothelial cell injury after extensive hepatectomy in cholestatic rats

BACKGROUND AND AIMS: The authors have shown previously that sinusoidal endothelial cell injury developed concomitantly with the accumulation of neutrophils in the hepatic sinusoidal space in cholestatic rats after extensive hepatectomy. The aim of the present study was to investigate the role of neutrophils in the development of this kind of endothelial cell injury. METHODS: Rats underwent 78% partial hepatectomy after 2 weeks of cholestasis, and subsequent external biliary drainage for 5 days. To decrease the number of neutrophils, antirat neutrophil serum was administered intraperitoneally. Some serum parameters and histological specimens were examined 48 h after partial hepatectomy. RESULTS: Anti-neutrophil serum significantly reduced the number of accumulated neutrophils in the hepatic sinusoids. Although the purine nucleoside phosphorylase: alanine aminotransferase ratio, a marker of non-parenchymal cell injury, was increased in cholestatic-hepatectomized rats, this abnormality was significantly attenuated by the treatment with antineutrophil serum. Electron microscopically, focal detachment of cytoplasms of sinusoidal endothelial cells was observed occasionally in cholestatic-hepatectomized rats, but was not found in the antirat neutrophil serum-treated rats. CONCLUSION: These results indicate that accumulated neutrophils might be important effector cells in the pathogenesis of sinusoidal endothelial cell injury after extensive hepatectomy in cholestatic rats, even after appropriate external biliary drainage.     

Polymorphisms in the transforming growth factor-beta gene (TGF-beta) and the risk of advanced alcoholic liver disease.

BACKGROUND/AIMS: There are wide interindividual differences in the risk of developing alcoholic cirrhosis. Transforming growth factor beta(1) (TGF-beta(1)) is the main cytokine involved in liver fibrogenesis. The TGF-beta(1) gene is polymorphic at several sites and these polymorphisms are probably related to differences in the rate of TGF-beta(1) synthesis. Our aim has been to analyse the influence of the TGF-beta(1) gene polymorphisms in the predisposition to advanced alcoholic liver disease (ALD) in ethanol abusers. METHODS: TGF-beta(1) single nucleotide polymorphisms at positions -509 (C or T), +869 (C or T, codon 10), and +915 (C or G, codon 25) were examined in 165 alcoholics with advanced ALD and in 185 healthy controls. RESULTS: Among the 94 male patients with oesophageal varices, those carrying the GG genotype at position +915 were diagnosed at an older age than the remaining patients (age 52.1 years, standard deviation (SD) 9.9 vs. 45 SD 13.4, P=0.012). No other statistically significant differences were found in the distribution of the three TGF-beta(1) polymorphisms analysed individually or as combined haplotypes. CONCLUSIONS: The polymorphisms at the TGF-beta(1) gene analysed in this study are probably not related to the risk of advanced ALD.     

Serum aminoterminal peptide of type III procollagen (PIIINP) and transforming growth factor-beta1 (TGF-beta1) levels in patients with chronic hepatitis B and C

Fibrosis is the process accompanying majority of chronic diseases of liver, independent of etiological factor and leading to cirrhosis and hepatic failure. Monitoring fibrosis process by liver's biopsy is limited, so many attempts are undertaken to assess concentrations of definite proteins in blood, which could be easily accessible marker of intrahepatic process. It seems, that among others, determinations of blood concentration of aminoterminal propeptide of procollagen III--index of collagen's III synthesis and TGF-beta 1--cytokine of antiproliferative action and inhibiting hepatocytes' growth, yet inducing fibroblasts' growth and stimulating fibrosis process brings out such a possibility. The aim of the study was simultaneous determination of TGF-beta 1 and PIIINP concentration in blood of patients with chronic hepatitis B and C before interferone's therapy in comparison to healthy controls, assessment of the parameters in dependence on stage of liver fibrosis and determination of correlation between TGF-beta 1 and PIIINP. Studies were performed in 40 patients with chronic hepatitis B (CAH B) and 35 patients with chronic hepatitis C (CAH C). Significantly increased serum concentrations of TGF-beta 1 as PIIINP in both groups of patients (CAH B and CAH C; grading 2-3, staging 1-2) in comparison with control group was noted. Significant positive correlation of TGF-beta 1 and PIIINP serum concentrations in both groups of patients was observed. There was not significant changes in PIIINP serum levels in patients with hepatitis B and C in dependence on stage of liver fibrosis (staging 1 vs staging 2) but TGF-beta 1 serum levels was significantly increased in CAH B and C patients with higher stage of liver fibrosis process. On the base of obtained results, it seems that changes in TGF-beta 1 concentrations in blood reflect "grading" and "staging" and can be a marker of intensification of intrahepatic fibrosis process whereas PIIINP levels in blood have rather the relation with "grading".     

Local angiotensin II and transforming growth factor-beta1 in renal fibrosis of rats

Studies have demonstrated that local angiotensin II (Ang II) generation is enhanced in repairing kidney and that ACE inhibition or AT(1) receptor blockade attenuates renal fibrosis. The localization of ACE and Ang II receptors and their relationship to collagen synthesis in the injured kidney, however, remain uncertain. Using a rat model of renal injury with subsequent fibrosis created with chronic elevations in circulating aldosterone (ALDO), we examined the distribution and binding density of ACE and Ang II receptors in repairing kidneys, as well as their anatomic relationship to transforming growth factor-beta1 (TGF-beta1) mRNA, type I collagen mRNA, collagen accumulation, and myofibroblasts. Two groups of animals (n=7 in each group) were studied: (1) normal rats served as controls, and (2) uninephrectomized rats received ALDO (0.75 microg/h SC) and 1% NaCl in drinking water for 6 weeks. Compared with control rats, in ALDO-treated rats we found (1) significantly (P<0.01) increased blood pressure, reduced plasma renin activity, and increased plasma creatinine levels, (2) diffuse fibrosis in both renal cortex and medulla, (3) abundant myofibroblasts at these sites of fibrosis, (4) significantly increased (P<0.01) binding density of ACE and Ang II receptors (60% AT(1), 40% AT(2)) at the sites of fibrosis, and (5) markedly increased (P<0.01) expression of TGF-beta1 and type I collagen mRNAs at these same sites. Thus, in this rat model of renal repair, the enhanced expression of ACE, Ang II receptors, and TGF-beta1 is associated with renal fibrosis. Ang II generated at the sites of repair appears to have autocrine/paracrine functions in the regulation of renal fibrous tissue formation alone or through its stimulation of TGF-beta1 synthesis.     

A blocking peptide for transforming growth factor-beta1 activation prevents hepatic fibrosis in vivo

BACKGROUND/AIMS: Thrombospondin-1 is a major activator of transforming growth factor-beta1 (TGF-beta1), and a peptide derived from the latency-associated peptide, Leu-Ser-Lys-Leu (LSKL), inhibits the activation of TGF-beta1. In this study, the effects of LSKL on the hepatocyte damage and fibrogenesis in dimethylnitrosamine (DMN)-induced rat liver fibrosis were examined. METHODS: Animals were given an intraperitoneal (i.p.) injection of DMN or saline three times per week for 4 weeks, and treated with LSKL, a control peptide, or saline i.p. daily. RESULTS: Liver atrophy caused by DMN-injection was significantly inhibited in the DMN+LSKL group. The degrees of necrosis/degeneration and fibrosis scores were significantly lower in the DMN+LSKL group than in the control groups. The hydroxyproline content was significantly higher in the control groups than in the DMN+LSKL group. The amount of active TGF-beta1 was less in the DMN+LSKL group than in the control groups, and the active/total TGF-beta1 ratio in the DMN+LSKL group was suppressed in the control groups. Phosphorylation of Smad 2 in the liver was significantly decreased in the DMN+LSKL group. CONCLUSIONS: The LSKL peptide prevented the progression of hepatic damage and fibrosis through the inhibition of TGF-beta1 activation and its signal transduction in vivo.     

Reduced plasma concentrations of transforming growth factor beta1 (TGF-beta1) in obese women

OBJECTIVE: To evaluate Transforming Growth Factor beta1, (TGF-beta1) plasma concentrations and the possible relationship between this growth factor and various hormones in obese women. DESIGN: Case-control study. SETTING: Outpatient's Service for the Prevention and Treatment of Obesity at the University Hospital. SUBJECTS: Twenty-five women with mild to moderate obesity, and 15 non-obese healthy women were used as controls. MEASUREMENTS: Evaluation of TGF-beta1, insulin, prolactin, sex-hormone binding globulin, androstenedione, free triiodothyronine, free tetraiodothyronine, thyroid-stimulating hormone, dehydroepiandrosterone-sulfate, testosterone, insulin-like growth factor 1, cortisol and adrenocorticotropic hormone plasma concentrations in obese women. Blood samples were taken from fasting subjects for the determination of the above parameters. RESULTS: In obese women plasma TGF-beta1 concentrations were lower than in controls. The obese subjects also had lower GH, IGF-1 and SHBG plasma concentrations and increased insulinaemia. A positive correlation was found between TGF-beta1 and both IGF-1 (r = 0.52; P < 0.01) and DHEA-S (r = 0.43; P < 0.05), while a negative correlation was found between TGF-beta1 and SHBG (r = -0.49; P < 0.05). CONCLUSIONS: The reduction in TGF-beta1, an antilipogenic factor, may play a role in the pathogenesis of obesity and could be involved in the development of some obesity-related endocrine alterations.     

Effects of interferon-alpha on expression of hepatic stellate cell and transforming growth factor-β1 and α-smooth muscle actin in rats with hepatic fibrosis

AIM: To investigate the effect of interferon-α(IFN-α) on preventing or reversing hepatic fibrosis in rat experimental model induced by CCl4.METHODS: One hundred and ten Sprague-Dawley rats were divided into five groups: group A (normal controls,n=18), group B (fibrotic model controls, n=22), group C (IFN-α prevention, n=22) initially treated with intra-muscular injection of IFN-α in saline daily at the doses of 1&#215;105U for 6wk, group D (IFN-α treatment, n=24) treated with intra-muscular injection of IFN-α in saline daily at the doses of 1&#215;105U for 6wk after the first 6wk, group E (0.9% sodium chloride treatment control, n=24) treated with intra-muscular injection of 0.01mL/kg daily for 6wk after the first 6wk. At the end of the experiment, all rats of each group were killed. Samples of the liver obtained by biopsy were subjected to histological, immunohistochemical and electron microscopic studies for the expressions of transforming growth factor-β1(TGF-β1) and α-smooth muscle actin (α-SMA).RESULTS: The expressions of TGF-β1, the number of activated hepatic stellate cells and α-SMA in hepatic tissue of group C were significantly less than those of group B(P&lt;0.01). The degree of fibrosis score in group B was also significantly less than that of group C under light microscope (P&lt;0.01).CONCLUSION: IFN-α can inhibit the production of TGF-β1, decrease HSC activation and stimulate its apoptosis.     

Role of transforming growth factor-beta1-smad signal transduction pathway in patients with hepatocellular carcinoma

AIM:To explore the role of transforming growth factor-beta1(TGF-β1)-smad signal transduction pathway inpatients with hepatocellular carcinoma.METHODS:Thirty-six hepatocellular carcinoma speci-mens were obtained from Qidong Liver Cancer Instituteand Department of Pathology of the Second AffiliatedHospital of Nanjing Medical University.All primary anti-bodies(polyclonal antibodies)to TGF-β1,type Ⅱ Trans-forming growth factor-beta receptor(TβR-Ⅱ),nuclear fac-tor-kappaB(NF-KB),CD34,smad4 and smad7,secondaryantibodies and immunohistochemical kit were purchasedfrom Zhongshan Biotechnology Limited Company(Bei-jing,China).The expressions of TGF-β1,TβR-Ⅱ,NF-kB,smad4 and smad7 proteins in 36 specimens ofhepatocellular carcinoma(HCC)and its adjacent tissuewere separately detected by immunohistochemistry toobserve the relationship between TGF-β1 and TβR-Ⅱ,between NF-kB and TGF-β1,between smad4 and smad7and between TGF-β1 or TβR-Ⅱ and microvessel density(MVD).MVD was determined by labelling the vesselendothelial cells with CD34.RESULTS:The expression of TGF-β1,smad7 and MVDwas higher in HCC tissue than in adjacent HCC tissue(P<0.01,P<0.05,P<0.01 respectively).The expressionof TβR-Ⅱ and smad4 was lower in HCC tissue than in its adjacent tissue(P<0.01,P<0.05 respectively).Theexpression of TGF-β1 protein and NF-kB protein wasconsistent in HCC tissue.The expression of TGF-β1 andMVD was also consistent in HCC tissue.The expressionof TβR-Ⅱ was negatively correlated with that of MVD inHCC tissue.CONCLUSION:The expressions of TGF-β1,TβR-Ⅱ,NF-kB,smad4 and smad7 in HCC tissue,which are ma-jor up and down stream factors of TGF-β1-smad signaltransduction pathway,are abnormal.These factors areclosely related with MVD and may play an important rolein HCC angiogenesis.The inhibitory action of TGF-β1 isweakened in hepatic carcinoma cells because of abnor-mality of TGF-β1 receptors(such as TβR-Ⅱ)and postre-ceptors(such as smad4 and smad7).NF-kB may causeactivation and production of TGF-β1.     

Understanding the role of transforming growth factor-beta1 in intimal thickening after vascular injury

Intimal thickening is the most important cause of in-stent restenosis. The pathology of intimal thickening is attributable to a local inflammatory response after vascular injury which results in the production of cytokines. Transforming growth factor-beta1 (TGF-beta1) is a profibrotic cytokine that is involved in the induction of intimal thickening. Up-regulation of TGF-beta1 after arterial injury results in the activation of various downstream pathways which stimulate the proliferation and migration of vascular smooth muscle cells, as well as the production of local extracellular matrix proteins. Recent evidence suggests that antagonizing TGF-beta1 activity with direct or indirect inhibitors may attenuate or prevent intimal thickening. Additionally, TGF-beta1 synthesis, activation and downstream regulation may also serve as significant sources of treatment. This review attempts to show the role of TGF-beta1 in the pathology of intimal thickening and underlines the importance of TGF-beta1 as a target for therapy.