ObjectiveTo explore the expressions of prostaglandin F2α receptor (PTGFR) and cyclooxygenase-2 (COX-2) in tissues of benign bile duct scar and their significances, and investigate the regulating effect of transforming growth factor-β1 (TGF-β1) on the expression of PTGFR in human bile duct fibroblasts cultured in vitro. MethodsThe samples of common bile duct (CBD) scars were collected from 18 patients with benign bile duct scar stricture and 6 cases of normal CBD tissues from liver transplantation donor were collected as control. The expressions of PTGFR and COX-2 were detected by immunohistochemical strept-avidin-biotin complex (SABC) method. Semiquantitative RT-PCR and ELISA methods were used to detect the mRNA and protein levels of PTGFR in bile duct fibroblasts which were effected by TGF-β1 with different concentrations (0, 10, 20, and 30 ng/ml) for 24 h. ResultsThe positive rates of PTGFR and COX-2 were 88.9% (16/18) and 83.3% (15/18) in tissues of benigh CBD scar and 33.3% (2/6) and 0 (0/6) in normal CBD tissues (Plt;0.05). The expressions of the PTGFR mRNA and protein levels became upregulated when the concentrations of the TGF-β1 became higher in human bile duct fibroblasts (Plt;0.05). And the effect was concentration dependant to some extent. ConclusionsThe high expressions of PTGFR and COX-2 play important roles in the process of benign bile duct stricture formation. TGF-β1 is able to induce higher expressions of PTGFR mRNA level and the PTGFR protein level in a concentration dependent manner, and regulate the formation of benign bile duct stricture.
【Abstract】Objective To investigate the expression of tumor growth tactor β1 (TGFβ1) and p27 in gallbladder carcinoma and their relation to the development of the carcinoma. Methods The expression of TGF-β1 and p27 in 36 cases of gallbladder carcinoma was detected by SP immunohistochemical staining. Twenty cases of chronic cholecystitis were collected as control. Results The positive rate of TGF-β1 （63.9%） was higher than that of the control （10.0%），P＜0.05， and the positive rate of p27 (47.2%) was lower than that of the control 100%(P＜0.05). The positive rate of TGF-β1 was significantly higher in metastasis or Nevin Ⅳ～Ⅴ stage cases than that of non-metastasis or Nevin Ⅰ～Ⅲ stage cases 33.3% (P＜0.05). The positive rate of p27 was statistically higher in moderate and highly differentiation (60.9%), nonmetastasis (75.0%) or Nevin’s Ⅰ～Ⅲ stage (75.0%) cases than those of poor differentiation (23.0%), metastasis (33.3%) and Nevin Ⅳ～Ⅴ stage (33.3%) cases (P＜0.05). The expression of p27 was negatively correlated with that of TGF-β1(r=-0.4473，P＜0.05). There was significant difference in survival time between patients with TGF-β1 positive and TGF-β1 negative（P＜0.05）. The difference was also found between patients with p27 positive and p27 negative. Conclusion The upregulation of TGF-β1 and downregulation of p27 in gallbladder carcinoma indicates the imbalance of TGF-β1/p27 system, which may play a role in the carcinogenesis and predict the malignant behaviors of the carcinoma.
Objective To investigate the role of IFN-γ in suppressing bleomycin-induced pulmonary fibrosis in rats.Methods Seventy-five SD rats were randomly divided into five groups (15 rats in each group)，ie.a normal group，a bleomycin-induced pulmonary fibrosis model group，a dexamethasone-treated group，a high-dose IFN-γ-treated group (150 000 U/kg) and a low-dose IFN-γ-treated group (50 000 U/kg).Five rats in each group were randomly killed in 7th day，14th day and 28th day after relative treatment respectively，and lung tissue samples were harvested for histopathology study.HE and Masson staining were used to determine the extent of alveolus inflammation and pulmonary fibrosis respectively.Histoimmunochemical method were adapted to determine protein levels of TGF-β1，CTGF，type Ⅰcollagen and type Ⅲ collagen in pulmonary tissues.Results Histopathological study showed that treatment with either dexamethasone or IFN-γ (both high dose and low dose) remarkably meliorated the extent of alveolus inflammation and suppressed pulmonary fibrosis (compared with model group，all Plt;0.05).Histoimmunochemical study suggested that both dexamethasone and IFN-γ could inhibit the expression of TGF-β1，CTGF，type Ⅰand type Ⅲ collagen (compared with model group，all Plt;0.05)，and the suppression of TGF-β1，type Ⅰand type Ⅲ collagen expression was more obvious in high-dose IFN-γ-treated group than those in low-dose group (Plt;0.05).Conclusions INF-γ possesses apparent anti-fibrosis effect that is similar to dexamethasone but with less side effect.Such effect may resulted from reduced production of type Ⅰand type Ⅲ collagen through expression inhibition of cytokines such as TGF-β1 and CTGF.
Objective To study the influence of transforming growth factor-β1(TGF-β1), dentin non-collagen proteins(dNCPs) and their complexon tissue engineering pulp system. Methods Collagen I and dentin powder were used to construct the system of pulp cells in 3dimensional culture, dentin powder was added in the gel. The tissue engineering pulp were divided TGF-β1 group, dNCPs group, TGF-β1/dNCPsgroup and control group.After3, 6 and 14 days, the appearance and the differentiation of pulp cells were observed by HE staining and immunohistochemical staining -respectively. Results Collagen I could form netted collagen gel construction. Growing condition of pulp cells in gel was similar to that of pulp cells in vivo. After the TGF-β1 and dNCPswere added, the pulp cells had some characteristics of odontoblasts and had unilateral cell process after culture 6 days. Pulp cells arranged with parallel columnar and form dentin-pulp-like complex after 14 days. Immunohistochemical staining showed dentin salivary protein(DSP) began to express in some cells.The number of positive cell was most in the TGF-β1 group. No positive cells were detected in the control group. Conclusion The transforming growth factor-β1 and noncollagen proteins can stimulate the pulp cells to transform into odontoblasts to some extent, which promote the formation of tissue engineering pulp.
Objective To investigate the effect of transforming growth factor-β1 (TGF-β1) gene transfer on the biological characteristics of osteoblasts. Methods The expression of TGF-β1 in the transfected osteoblasts was detected by in situ hybridization and assay of TGF-β1 activity in the supernatant (minklung epithelium cell growth -inhibition test). The effects of gene transfer andsupernatant of the transfected osteoblasts on the proliferation and alkaline phosphatase(ALP) activity of osteoblasts were detected by 3 H-TdR and MTT. Results The results of in situ hybridization analysis suggested that the osteoblasts transfected by TGF-β1 gene could express TGF-β1 obviously. The complex medium, which was the mixture of serum-free DMEM and the activated supernatant according to 1∶1, 1∶2, 1∶4, could inhibit growth of Mv-1-Lu evidently and the ratios ofinhibition were 16.3%, 22.7%, 28.2% respectively. TGF-β1 gene transfer hadno effect on the biological characteristics of osteoblasts, but the activated supernatant of transfected osteoblasts stimulated proliferation and inhibited ALPactivity of osteoblasts. Conclusion TGF-β1 gene transfer promotes the expression of TGF-β1 and the biological characteristics of trasfected osteoblasts are stable, which is helpful for gene therapy of bone defects in vivo.
OBJECTIVE: To explore the autocrine regulation for the transforming growth factor-beta 1(TGF-β1) by the fibroblasts in vitro. METHODS: Fibroblasts were cultured in vitro which isolated from the normal prepuce by circumcision. TGF-β1 concentration were determined with ELISA for the different passages and the different time at the same passage. RESULTS: The TGF-β1 concentration was different for the distinct passage of fibroblasts and achieved the peak (450 ng/L) at the sixth passage. Among the same fibroblasts, the TGF-β1 concentration was different in the various days and passage of the summit (680 ng/L) on the fifth day, which was 2.5 times as high as that of the first day. CONCLUSION: The fibroblasts possess the ability of autocrine of the TGF-β1 and have some regularity. It will provide the theory basis for the research about TGF-β1 regulation mechanism and the medical application about salvia miltiorrhiza.
OBJECTIVE: To investigate the mechanism of overhealing alleviation by salvia miltiorrhiza (SM) in wound healing. METHODS: Fibroblasts were cultured in vitro, and SM was applied with different concentrations (40, 80, 160 and 320 micrograms/ml) and time(the 1st, 2nd, 3rd, 4th and 5th days) to influence their autocrine. The levels of transforming growth factor-β1 (TGF-β1) and epidermal growth factor (EGF) were determined by ELIAS and radioimmunoassay respectively. RESULTS: The SM could inhibit autocrine of TGF-β1 by fibroblasts (P lt; 0.05). However, it did not affect autocrine of EGF (P gt; 0.05). CONCLUSION: The above results indicate that SM reduces overhealing by inhibiting the autocrine of TGF-β1 selectively.
OBJECTIVE: To investigate the effects of basic fibroblast growth factor (bFGF) on the promoter activities of human alpha 1(I) procollagen gene and the interaction between bFGF and transforming growth factor-beta 1 (TGF-beta 1). METHODS: Fibroblasts of the hypertrophic scar and normal skin from a 3-year-old patient were primarily cultured and subcultured in vitro. Both of the fibroblasts were transient transfected with phCOL 2.5, containing -2.5 kb of 5’f lank sequence of human alpha 1(I) procollagen gene and CAT reporter gene by FuGENE transfection reagent; and treated thereafter by 16 ng/ml bFGF, 2 ng/ml TGF-beta 1 and 16 ng/ml bFGF + 2 ng/ml TGF beta 1 for 24 hours. The relative CAT expression values were determined by CAT-ELISA. RESULTS: TGF-beta 1 bly induced the CAT expression level, however, bFGF not only inhibited the basal CAT expression but also reduced the CAT expression up-regulated by TGF-beta 1 in normal skin and hypertrophic scar fibroblasts (P lt; 0.05). CONCLUSION: bFGF can reduce the promoter activities of human alpha 1(I) procollagen gene and antagonize the role of TGF-beta 1 in up-regulating the promoter activities of human alpha 1(I) procollagen gene in normal skin and hyertrophic scar fibroblasts.
OBJECTIVE: To study the effect of overexpression of truncated type II TGF-beta receptor on transforming growth factor-beta 1(TGF-beta 1) autoproduction in normal dermal fibroblasts. METHODS: In vitro cultured dermal fibroblasts were treated with recombinant human TGF-beta 1(rhTGF-beta 1) (5 ng/ml) or recombinant adenovirus containing truncated type II TGF-beta receptor gene (50 pfu/cell). Their effects on regulating gene expression of TGF-beta 1 were observed with Northern blotting. RESULTS: rhTGF-beta 1 up-regulated the gene expression of TGF-beta 1 and type I procollagen. Overexpression of truncated receptor II down-regulated the gene expression of TGF-beta 1. CONCLUSION: Overexpression of the truncated TGF-beta receptor II decreases TGF-beta 1 autoproduction via blocking TGF-beta receptor signal. The results may provided a new strategy for scar gene therapy.
OBJECTIVE: To explore the molecular mechanisms involved in the increased collagen synthesis by platelet-derived wound healing factors (PDWHF) during wound healing in alloxan-induced diabetic rats. METHODS: Thirty-three male SD rats were divided into two groups, the normal (n = 9) (group A) and the diabetic group (n = 24). Two pieces of full-thickness skin with diameter of 1.8 cm were removed from the dorsal site of diabetic rats. PDWHF (100 micrograms/wound) was topically applied to one side of the diabetic wounds (group B) on the operation day and then once a day in the next successive 6 days. Meanwhile, bovine serum albumin (100 micrograms/wound) was applied to the other side of diabetic wound as control group (group C) in the same way. Levels of transforming growth factor-beta 1 (TGF-beta 1) and procollagen I mRNA in wound tissue were inspected by dot blotting. RESULTS: TGF-beta 1 mRNA levels in group B were 4 folds and 5.6 folds compared with those in group C after 5 and 7 days (P lt; 0.01), however, still significantly lower than those of group A (P lt; 0.05). There was no significance difference among three groups on the 10th day after wounding. The levels for procollagen I mRNA in group B amounted to 2.1, 1.8 and 2.3 folds of those in group C after 5, 7, and 10 days (P lt; 0.01), respectively. Compared with those in the group A, procollagen I mRNA levels in the group B were significantly lower after 5 and 7 days (P lt; 0.05), and no significant difference was observed between group B and A after 10 days. CONCLUSION: One important way for PDWHF to enhance the collagen synthesis in diabetic wound healing is to increase the gene expression of endogenous TGF-beta 1.