Fibrogenic polymorphisms (TGF-beta, PAI-1, AT) in Mexican patients with established liver fibrosis. Potential correlation with pirfenidone treatment.

BACKGROUND/AIM: The aim of this work was to establish a potential correlation between specific polymorphisms and presence of hepatic fibrosis in Mexican patients with established liver fibrosis (ELF). Second, necroinflammatory index improvement was correlated with Pirfenidone (PFD) treatment response and the same polymorphisms. METHODS: We analyzed TGF-beta polymorphisms in codon 25, a single basepair guanine insertion-deletion polymorphism (4G/5G) for PAI-1 and angiotensin AT-6 single nucleotide polymorphism located in -6 promoter region. Twenty patients infected with either hepatitis C virus (HCV) (n = 13) or affected by alcohol consumption (n= 7) were included. Thirty subjects with no hepatic damage were included in control group. Blood samples for genomic DNA were obtained and plasminogen activator inhibitor-1 polymorphisms were done by polymerase chain reaction-artificial introduction of a restriction site, TGF-beta by polymerase chain reaction-amplification refractory mutation system and AT by polymerase chain reaction-restriction fragment length polymorphisms. Liver biopsies were obtained at baseline and after 12 months of PFD treatment. RESULTS: Established liver fibrosis patients had the homozygote G/G TGF-beta genotype, which has been associated with increased development of fibrosis. None of our patients had the G/C genotype. All pure HCV and pure alcohol abuse subjects carried G/G TGF-beta genotype (100% vs 37% control) (P = 0.0006). The odds of having TGF-beta G/G genotype was 19.5 for HCV patients and 10.83 for alcohol consumption patients as compared with healthy subjects (P < 0.001). Established liver fibrosis patients had an improvement in necroinflammatory index after PFD treatment when correlated with plasminogen activator inhibitor-1 and angiotensinogen-6 genotypes. CONCLUSION: Our data suggested that a combination of inherited polymorphisms increased the risk of advanced fibrosis in ELF patients. Pure HCV and pure alcohol consumption patients which were homozygous G/G carriers had 19.5- and 10.8-fold higher risk to develop advanced fibrosis respectively.

Pirfenidone prevents capsular contracture after mammary implantation.

BACKGROUND: Pirfenidone (PFD), a new antifibrotic and antiinflammatory agent, prevents and resolves fibrous tissue. This study evaluated the effect of PFD on adverse events in mammary implants using an animal model. Mammary implantation, the most frequent aesthetic surgery, may present several complications after surgery such as swelling, capsule contracture, hardness, and pain. METHODS: Wistar rats underwent submammary implantation with either smooth or textured silicone gel implants and were administrated 200 mg/kg of PFD daily. The control group received saline. The animals were killed at 8 weeks. The capsular tissue of both implants was removed for histologic and molecular analyses. RESULTS: Typical postaugmentation periimplant capsules with opacity on adjacent tissues developed 8 weeks after silicone implantation. No significant differences were observed between the textured and smooth implants in any analyzed parameter. Clearly, PFD reduced capsule thickness around submmamary tissue, fibroblast-like cell proliferation, and recruitment of inflammatory cells. The total cell numbers per field were reduced as well. In contrast, the control group presented abundant mononuclear cell infiltration and fibroblast-like cell proliferation. The total content of collagen in the PFD group was 50% less than in the control group. Fibroblast cells displayed 45% less activated phenotype in the PFD group than in the control group, as determined by immunohistochemistry techniques. In the PFD animals, transforming growth factor-beta (TGF-beta) decreased 85% and collagen 1 gene expression 60%, compared with the control group. CONCLUSION: The findings show a positive effect of PFD on mammary contracture in 10 rats. Despite the small number of animals, the differences found in 10 control rats encourage the authors to propose a larger study later and to suggest PFD as a potential preventive strategy in human mammary implantation surgery.

TGF-beta1 modulates matrix metalloproteinase-13 expression in hepatic stellate cells by complex mechanisms involving p38MAPK, PI3-kinase, AKT, and p70S6k.

Transforming growth factor-beta1 (TGF-beta1), the main cytokine involved in liver fibrogenesis, induces expression of the type I collagen genes in hepatic stellate cells by a transcriptional mechanism, which is hydrogen peroxide and de novo protein synthesis dependent. Our recent studies have revealed that expression of type I collagen and matrix metalloproteinase-13 (MMP-13) mRNAs in hepatic stellate cells is reciprocally modulated. Because TGF-beta1 induces a transient elevation of alpha1(I) collagen mRNA, we investigated whether this cytokine was able to induce the expression of MMP-13 mRNA during the downfall of the alpha1(I) collagen mRNA. In the present study, we report that TGF-beta1 induces a rapid decline in steady-state levels of MMP-13 mRNA at the time that it induces the expression of alpha1(I) collagen mRNA. This change in MMP-13 mRNA expression occurs within the first 6 h postcytokine administration and is accompanied by a twofold increase in gene transcription and a fivefold decrease in mRNA half-life. This is followed by increased expression of MMP-13 mRNA, which reaches maximal values by 48 h. Our results also show that this TGF-beta1-mediated effect is de novo protein synthesis-dependent and requires the activity of p38MAPK, phosphatidylinositol 3-kinase, AKT, and p70(S6k). Altogether, our data suggest that regulation of MMP-13 by TGF-beta1 is a complex process involving transcriptional and posttranscriptional mechanisms.

Improved effects of viral gene delivery of human uPA plus biliodigestive anastomosis induce recovery from experimental biliary cirrhosis.

Gene therapy may represent a new avenue for the development of multimodal treatment for diverse forms of cirrhosis. This study explores the potential benefits of combining adenovirus-mediated human urokinase-plasminogen activator (AdHuPA) gene delivery and biliodigestive anastomosis to enhance the therapeutic efficacy of each treatment alone for cholestatic disorders resulting in secondary biliary cirrhosis. In an experimental model of secondary biliary cirrhosis, application of 6 x 10(11) vp/kg AdHuPA adenovirus vector resulted in 25.8% liver fibrosis reduction and some improvement in liver histology. The relief of bile cholestasis by a surgical procedure (biliodigestive anastomosis) combined with AdHuPA hepatic gene delivery rendered a synergistic effect, with a substantial 56.9 to 42.9% fibrosis decrease. AdHuPA transduction resulted in clear-cut expression of human uPA protein detected by immunohistochemistry and induction of up-regulation in the expression of metalloproteinases MMP-3, MMP-9, and MMP-2. Importantly, functional hepatic tests, specifically direct bilirubin, were improved. Also, hepatic cell regeneration, rearrangement of hepatic architecture, ascites, and gastric varices improved in cirrhotic rats treated with AdHuPA but not in counterpart AdGFP cirrhotic animals. We believe this might represent a novel therapeutic strategy for human cholestatic diseases.

Hepatic phenylalanine-hydroxylase and tyrosine-aminotransferase mRNA levels in rats adapted to diets with different concentrations of protein

Se estudió el efecto de la concentración de la proteína de la dieta sobre concentraciones de ARNm de la tirosina aminotransferasa (TAT) y la fenilalanina hidroxilasa (PAH) hepáticas en ratas adaptadas a consumir dietas con 18 ó 50 por ciento de caseína en un horario restringido de 7 horas (9 a 16 h) durante 5 días. Las concentraciones de ARNm de TAT de ratas adaptadas a una dieta de 18 por ciento de caseína y alimentadas en forma aguda con dietas que contenían 6, 18 ó 50 por ciento de caseína, fueron 0.15, 0.84 y 5.08 veces más altas a las 6 horas en comparación con las concentraciones de ARNm antes de la administración de la dieta. Las concentraciones de ARNm de TAT después de 17 horas de ayuno en las ratas alimentadas con 6, 18 ó 50 por ciento de caseína fueron respectivamente -0.45, 1.76 y 9.11 veces mayores en comparación con el valor basal. Las concentraciones ARNm de PAH mostraron un patrón similar; en las ratas adaptadas a 18 por ciento de caseína se observó un aumento de -.68, 1.63 y 2.5 veces en las concentraciones de ARNm de PAH en las ratas alimentadas en forma aguda con 6, 18 y 50 por ciento de casína respectivmanete y un aumento de -0.86, 2.32 y 9.33 veces después de 17 horas de ayuno. La concentraciones de ARNm de TAT y PAH en ratas adaptadas a consumir 50 por ciento de caseína y luego alimentadas con 6 ó 50 por ciento de caseína mostraron un pico máximo a las 6 horas de ayuno. Estos resultados sugieren que las concentraciones crecientes de proteína en la dieta son capaces de producir aumentos en la concentración de los ARNm de las dos enzimas, posiblemente para eliminar el exceso de aminoácidos consumidos, ya que la concentración de los ARNm dependió más del contenido de proteína de la dieta de adaptación