Effects of transforming growth factor β ₁ receptor inhibitor SD-208 on human hypertrophic scar

Objective To investigate the effects of transforming growth factor β₁ (TGF-β₁) receptor inhibitor SD-208 on human hypertrophic scar and its mechanisms. Methods Scar fibroblasts were isolated from deprecated human hypertrophic scar tissue and then sub-cultured. Cells of the fifth passage were used in the following experiments. (1) Cells were divided into blank control group (BC) and 0.5, 1.0, 3.0, and 5.0 μmol/L SD-208 groups according to the random number table (the same grouping method below), with 6 wells in each group. Cells in group BC were added with 1 μL phosphate buffer solution, while cells in the latter four groups were added with 0.5, 1.0, 3.0, and 5.0 μmol/L SD-208, respectively. After being cultured for 12 hours, the proliferation activity of cells was detected by cell counting kit 8 and microplate reader (denoted as absorbance value). Suitable amount of substance concentration of SD-208 according to the results of proliferation activity of cells was chosen for the following experiments. (2) Another batch of cells were divided into group BC and 1, 3 μmol/L SD-208 groups and treated as in (1), with 8 wells in each group. The number of migration cells was detected by transwell method. (3) Another batch of cells were grouped and treated as in (2), and the microfilament morphology of cells was observed by rhodamine-phalloidin staining. (4) Another batch of cells were grouped and treated as in (2), and the protein expression of TGF-β₁ was assessed with Western blotting. (5) Forty-eight BALB/c nude mice were divided into normal saline group (NS) and 1 μmol/L SD-208 group, and one longitudinal incision with length of 1 cm was made on their back. Then human hypertrophic scar tissue was embedded into the incision. On post injury day 7, multipoint injection of NS in a volume of 0.05 mL was performed in wounds of rats in group NS, while rats in 1 μmol/L SD-208 group were given 0.05 mL 1 μmol/L SD-208, once a day. On the day 0 (the same day), 2, 4, 6, 8, 10, 12, 14, 16, 18, and 20 post first time of injection, the weight of 8 nude mice was weighed by electronic scale, and scar area was measured by vernier caliper and the ratio of rest scar area was calculated. (6) In week 1, 2, and 3 post first time of injection, the protein expression of TGF-β₁ of human hypertrophic scar tissue was assessed with Western blotting. Data were processed with one-way analysis of variance and two independent-sample t test. Results (1) The proliferation activity of cells in group BC, 0.5, 1.0, 3.0, and 5.0 μmol/L SD-208 groups was respectively 1.00±0.03, 0.90±0.08, 0.68±0.11, 0.54±0.04, and 0.42±0.09, and the proliferation activity of cells in 0.5, 1.0, 3.0, and 5.0 μmol/L SD-208 groups was significantly lower than that in group BC (with t values from 2.9 to 22.1, P<0.05 or P<0.01). (2) The number of migration cells in 1, 3 μmol/L SD-208 groups was significantly less than that in group BC (with t values respectively 6.5 and 6.4, P values below 0.01). (3) Compared with that in group BC, fluorescence intensity of microfilaments of cells in 1, 3 μmol/L SD-208 groups was attenuated, and the pseudopod extended less. (4) The protein expressions of TGF-β₁ of cells in group BC and 1, 3 μmol/L SD-208 groups were respectively 1.00±0.08, 0.80±0.08, and 0.61±0.05, and the protein expressions of TGF-β₁ of cells in 1, 3 μmol/L SD-208 groups were significantly lower than those in group BC (with t values respectively 4.0 and 9.2, P values below 0.01). (5) The weights of nude mice in group NS and 1 μmol/L SD-208 group were similar on each time day (with t values from 0.2 to 1.1, P values above 0.05). The ratios of rest scar area of nude mice in two groups were decreased along with the injection time, and the ratios of rest scar area of nude mice in 1 μmol/L SD-208 group were significantly less than those in group NS from the day 6 to 20 post first time of injection (with t values from 1.8 to 15.9, P<0.05 or P<0.01). In week 1, 2, and 3 post first time of injection, the protein expressions of TGF-β₁ of human hypertrophic scar tissue in nude mice in two groups showed a tendency of decrease, and the protein expressions of TGF-β₁ of human hypertrophic scar tissue in nude mice in 1 μmol/L SD-208 group were significantly lower than those in group NS (with t values from 6.2 to 19.1, P values below 0.01). Conclusions SD-208 has significant inhibition effect on human hypertrophic scars, and the mechanism is correlated to the inhibition of protein expression of endogenous TGF-β₁.