RESUMO
OBJECTIVE: To study the clinical characteristics of Hennekam syndrome. METHODS: We described a case of long-term iron deficiency anemia, characteristic facial anomalies, growth retardation, and intestinal lymphangiectasia. To our knowledge, this is the first case of Hennekam syndrome reported in China. Meanwhile, relevant literature was also reviewed. RESULTS: A total of 35 cases of Hennekam syndrome were identified, consisting of 18 males and 17 females (age ranging 0 - 40 years old). Lymphangiectasia, lymphedema, facial anomalies (hypertelorism, flat nasal bridge and flat face) and developmental retardation were the major clinical manifestations of the syndrome. CCBE1 mutation may have played an important role in the pathogenesis of the syndrome. Long-term moderate-to-severe iron deficiency anemia was a distinctive feature of our case. Lymphangiography revealed lymphangiectasia of small intestine and lower limb, as well as thoracic outlet obstruction. Complete elimination of anemia and significant increase of serum albumin level were observed several months after the adhesiolysis procedure of the distal end of thoracic duct. However, anemia and severe hypoalbuminemia relapsed after taking greasy food. CONCLUSIONS: Hennekam syndrome is a rare autosomal recessive syndrome characterized by defective lymphatic development. Congenital lymphangiectasia should be considered in the patients with unexplained developmental retardation and hypoalbuminemia. Moreover, intestinal lymphangiectasia can be a rare cause of gastrointestinal bleeding.
Assuntos
Anormalidades Craniofaciais , Doenças dos Genitais Masculinos , Linfangiectasia Intestinal , Linfedema , Feminino , Humanos , Adulto JovemRESUMO
OBJECTIVE: To investigate the role of focal adhesion kinase (FAK) in adhesion and migration of hepatic stellate cells (HSC). METHODS: Two recombinant plasmids expressing short hairpin RNAs (shRNAs) targeting FAK were constructed and one plasmid substantially suppressing FAK expression in HSC was selected. Real-time PCR and Western blot were used to detect the knockdown effects of FAK gene. After 48-hour treatment with FAK shRNA, toluidine blue colorimetric assay was used to detect the cell adhesion. Wound-healing assay and improved Boyden double-chamber were used to detect the cell migration induced by FN. RESULTS: The recombinant plasmid expressing FAK shRNA was successfully constructed and transfected into HSC. Compared with the controls, the expression of FAK mRNA and protein in HSC treated with FAK shRNA was markedly down-regulated by 76.82% and 72.53%, respectively. The expression of p-FAK (Tyr397) protein was also decreased by 62.71% 48 h posttransfection. The adhesion of HSC was inhibited by 58.69% at 48 h after shRNA transfection. FAK gene silencing could also dramatically inhibit FN-stimulated HSC migration, and the cell migration distance and the cell number of crossing membrane were decreased by 58.27% and 83.70%, respectively. CONCLUSIONS: FAK gene silencing suppresses adhesion and migration of HSC, and FAK may be a potential target for novel anti-fibrosis therapies.
Assuntos
Quinase 1 de Adesão Focal/metabolismo , Células Estreladas do Fígado/enzimologia , Cirrose Hepática/prevenção & controle , Interferência de RNA , Animais , Western Blotting , Adesão Celular , Linhagem Celular , Movimento Celular , Regulação para Baixo , Fibronectinas , Quinase 1 de Adesão Focal/genética , Vetores Genéticos , Células Estreladas do Fígado/citologia , Cirrose Hepática/patologia , Plasmídeos/genética , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , TransfecçãoRESUMO
OBJECTIVES: To investigate the effects of FAK-related non-kinase (FRNK) on expressions of type I collagen and matrix metalloproteinase-2 (MMP-2) mRNA and tissue inhibitor of metalloproteinase-2 (TIMP-2) mRNA in rat hepatic stellate cells (HSC). METHODS: Using in vitro cell culture technique, FRNK plasmids were transfected into HSC mediated by cationic liposome. Type I collagen synthesis capability in HSC was examined by 3H-Pro incorporation assay. The levels of FRNK in HSC were assayed by Western blot, and the expressions of MMP-2 and TIMP-2 were assayed by RT-PCR on mRNA levels. RESULTS: The exposure of HSC to FRNK caused the expression of FRNK protein to be up-regulated, and the FRNK protein contents reached the highest point at 48 h after the transfection, P less than 0.05. The expressions of MMP-2 mRNA were up-regulated by FRNK; the expressions of TIMP-2 mRNA were down-regulated by FRNK; the ratios of MMP-2 mRNA/TIMP-2 mRNA were enhanced by FRNK. CONCLUSION: After FRNK was transfected, the capability of type I collagen synthesis in HSC was inhibited, which may be related to the up-regulation of MMP-2 mRNA/TIMP-2 mRNA.
Assuntos
Células Estreladas do Fígado/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Proteínas Tirosina Quinases/genética , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Animais , Células Cultivadas , Colágeno Tipo I/metabolismo , Células Estreladas do Fígado/citologia , Plasmídeos , RNA Mensageiro/genética , Ratos , TransfecçãoRESUMO
OBJECTIVE: To explore the effect of Salvia miltiorrhiza monomer IH764-3 on apoptosis in hydrogen peroxide (H2O2)-stimulated hepatic stellate cells (HSCs). METHODS: HSCs were cultured in medium with different IH764-3 doses (10 mg/L, 20 mg/L, 30 mg/L, 40 mg/L) and without IH764-3. Direct cell count, 3H-thymidine incorporation, Annexin-V/Propidium Iodide double-labeled flow cytometry, TUNEL and transmission electron microscopy were employed to estimate the influence of IH764-3 on proliferation and apoptosis of HSCs. The expression of extracellular signal-regulated kinase 1 (ERK1) mRNA and protein in HSCs were detected using RT-PCR and Western blot respectively. RESULTS: It was showed that H2O2 could promote HSC proliferation. In contrast, IH764-3 at concentrations of 10 mg/L, 20 mg/L, 30 mg/L and 40 mg/L inhibited its proliferation. The inhibition rates were 7.13%, 28.36%, 53.80% and 73.10% (P < 0.01). And the inhibition rates of IH764-3 at concentrations of 30 mg/L at 12 h, 24 h and 48 h were 22.24%, 40.51% and 61.65%. Furthermore, IH764-3 could also induce the HSC apoptosis in dose-dependent an dtime-dependent manners (P < 0.01). In addition, after exposed of HSCs to IH764-3 for 24 h, ERK production decreased and ERK1 mRNA was down-regulated earlier about 2 h after exposure to IH764-3. CONCLUSION: IH764-3 may inhibit the proliferation and induce apoptosis of HSCs in both dose-dependent and time-dependent manners, which may be related to down-regulation of ERK expression.
Assuntos
Apoptose/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Células Estreladas do Fígado/citologia , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Salvia miltiorrhiza/química , Apoptose/fisiologia , Linhagem Celular , Regulação para Baixo/efeitos dos fármacos , Medicamentos de Ervas Chinesas/isolamento & purificação , Humanos , Peróxido de Hidrogênio/farmacologia , Proteína Quinase 3 Ativada por Mitógeno/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
AIM: To investigate the effects and mechanism of disruption of focal adhesion kinase (FAK) expression on collagen metabolism in rat hepatic stellate cells (HSC). METHODS: The plasmids expressing FAK short hairpin RNA (shRNA) were transfected into HSC-T6 cells, and the level of FAK expression was determined by both real-time quantitative polymerase chain reaction (Q-PCR) and Western blotting analysis. The production of type I collagen and type III collagen in FAK-disrupted cells was analyzed by real-time Q-PCR. The level of collagen metabolism proteins, including matrix metalloproteinases-13 (MMP-13) and tissue inhibitors of metalloproteinases-1 (TIMP-1) was also determined by both real-time Q-PCR and Western blotting analysis. RESULTS: The transfection of FAK shRNA plasmids into HSC resulted in disrupted FAK expression. Compared with the HK group, the levels of type I collagen and type III collagen mRNA transcripts in FAK shRNA plasmid group were significantly decreased (0.69 +/- 0.03 vs 1.96 +/- 0.15, P = 0.000; 0.59 +/- 0.07 vs 1.62 +/- 0.12, P = 0.020). The production of TIMP-1 in this cell type was also significantly reduced at both mRNA and protein levels (0.49 +/- 0.02 vs 1.72 +/- 0.10, P = 0.005; 0.76 +/- 0.08 vs 2.31 +/- 0.24, P = 0.000). However, the expression of MMP-13 mRNA could be significantly up-regulated by the transfection of FAK shRNA plasmids into HSC (1.74 +/- 0.20 vs 1.09 +/- 0.09, P = 0.000). CONCLUSION: These data support the hypothesis that shRNA-mediated disruption of FAK expression could attenuate extracellular matrix (ECM) synthesis and promote ECM degradation, making FAK a potential target for novel anti-fibrosis therapies.
Assuntos
Colágeno Tipo III/metabolismo , Colágeno Tipo I/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/genética , Células Estreladas do Fígado/metabolismo , RNA Interferente Pequeno/metabolismo , Animais , Linhagem Celular , Colágeno Tipo I/genética , Colágeno Tipo III/genética , Células Estreladas do Fígado/citologia , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/metabolismo , RNA Interferente Pequeno/genética , Ratos , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , TransfecçãoRESUMO
The gene phosphatase and tensin homolog deleted on chromosome 10 (PTEN) codes for a tumor-suppressor phospholipid phosphatase. Deletion, mutation or abnormal expression of PTEN is commonly found in many kinds of malignant tumors. At the time of this study, though, the role of PTEN expression in the pathology of hepatic fibrosis remains unclear. In this study, we investigate the dynamic expression of PTEN in a rat model of hepatic fibrosis, with special emphasis on the activation and proliferation of hepatic stellate cells (HSC) in vivo. The rat model of hepatic fibrosis used in this study employed common bile duct ligation. At four time points, the expression of PTEN in hepatic tissues and activated HSC in rat liver tissues was measured by immunohistochemical staining, Western blotting, real-time fluorescent quantitative PCR and immunofluorescence confocal laser scanning microscopy, respectively. Further, alpha-smooth muscle actin (alpha-SMA), an activated HSC marker in rat liver tissues, was detected by immunohistochemical staining. This study showed that aggravation of hepatic fibrosis led to gradually decreasing expression of PTEN in the hepatic tissues. Further, as hepatic fibrosis worsens, PTEN-expressing activated HSC accounts for an increasingly smaller percentage of all activated HSC. In contrast, the percentage of alpha-SMA-expressing HSC cells increases significantly. In conclusion, expression of PTEN mRNA and protein is down-regulated in fibrogenic rat liver tissue, and its expression in HSC in vivo also decreases with progression of fibrosis. Thus, these results show that the dynamic expression of PTEN in hepatic tissues negatively correlates with activation and proliferation of HSC.