[Effects of sodium tanshinone II A sulfonate on proliferation of fibroblasts in scar and the mRNA and protein expressions of transforming growth factor beta 1 and alpha smooth muscle actin].
OBJECTIVE
To study the effects of sodium tanshinone II A sulfonate (STS) on proliferation of fibroblasts (Fbs) in human hypertrophic scar (HS), the mRNA and protein expressions of transforming growth factor beta 1 (TGF-β1) and alpha smooth muscle actin (α-SMA), and to investigate the scar inhibition mechanism of STS.


METHODS
Fbs were isolated from HS tissues that were removed from eight patients after burn injury, and they were cultured in vitro. Cells from the 3rd to the 6th passages were used in the experiment. Fbs were divided into control group and experimental group according to the random number table, and cells in the experimental group was divided into 0.050, 0.075, 0.100, 0.125, 0.150, 0.200 mg/mL STS subgroups. Cells in each subgroup were cultured with the corresponding concentration of STS, and cells in control group were cultured in equal volume of serum-free medium. After being cultured for 24 and 48 h, cell morphology was observed with inverted phase contrast microscope; cell proliferation was determined with MTT method and the proliferation inhibition rate (IR) was calculated. After being cultured for 48 h, the protein levels of TGF-β1 and α-SMA were determined with Western blotting; the mRNA expressions of TGF-β1 and α-SMA were determined with RT-PCR (no 0.200 mg/mL STS subgroup was set for these two indicators). Data were processed with factorial analysis of variance; differences between groups were processed with LSD test or Games-Howell test for unequal variances.


RESULTS
(1) Fbs grew well in control group, but reduction in adherence and disorderly arranged Fbs were observed in experimental group. The cells in experimental group became smaller and round, with increasing intracellular particles and necrosis. A large amount of necrotic debris of cells was observed in 0.200 mg/mL STS subgroup. (2) The absorbance value of Fbs in each experimental subgroup was significantly lower than that in control group (with P values all below 0.01). Along with the increase in the concentration of STS and extension of culture time, the IR value increased, showing a certain degree of time-concentration dependence. After being cultured with STS for 24 and 48 h, IR values of cells in the experimental subgroups were respectively 23.58%, 32.11%, 37.56%, 57.98%, 79.53%, 96.69% and 34.72%, 38.48%, 47.62%, 64.40%, 89.70%, 98.01%. (3) Except for the 0.050 mg/mL STS subgroup, the protein levels of TGF-β1 and α-SMA in the other subgroups were significantly lower than those in control group (with F values respectively 57.674, 47.795, P values all below 0.001). The protein levels of TGF-β1 and α-SMA reached the nadir in 0.150 mg/mL STS subgroup, respectively 0.34 ± 0.06, 0.33 ± 0.07. The relative expression amounts of TGF-β1 and α-SMA mRNA in the experimental subgroups were obviously decreased compared with those in control group (with F values respectively 68.548, 47.522, P values all below 0.001), which was most significant in 0.150 mg/mL STS subgroup, with TGF-β1 mRNA and α-SMA mRNA respectively 0.39 ± 0.07 and 0.42 ± 0.08.


CONCLUSIONS
STS can inhibit the proliferation of Fbs, reduce the protein and mRNA expressions of TGF-β1 and α-SMA, which may be beneficial to ameliorate the formation and contracture of HS, and it is assumed as a potential drug for treating scars.
Department of Burns and Plastic Surgery, the First Affiliated Hospital of Guangxi Medical University, Nanning 530021, China.
Hua
H
Liu
Hua-lian
HL
Yang
Shu-fang
SF
Zhou
Cheng-yue
CY
Meng
D000199:Actins / Q000235:genetics / Q000378:metabolism*; D000328:Adult; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002921:Cicatrix / Q000378:metabolism / Q000473:pathology*; D005260:Female; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008297:Male; D008875:Middle Aged; D010616:Phenanthrenes / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism*; D055815:Young Adult
D000199:Actins; D010616:Phenanthrenes; D012333:RNA, Messenger; D053773:Transforming Growth Factor beta1; C024894:tanshinone II A sodium sulfonate