Objective: To investigate the effects of heat injured keratinocytes (KC) supernatant on the expressions of collagen type I, collagen type III, and matrix metalloproteinase 1 (MMP-1) of dermal fibroblasts (Fb).
Methods: KC and Fb were isolated and cultured. Then the models of heat injured KC and Fb were reproduced in vitro, respectively. The heat injured and normal culture supernatant were collected respectively at 12 hours, and formulated as a 50% concentration of cell-conditioned medium. According to the culture medium, Fb at passage 3-5 was divided into 3 groups. Normal Fb was cultured with the conditioned medium containing 50% heat injured KC culture supernatant (group A), the conditioned medium containing 50% normal KC culture supernatant (group B), and DMEM (group C), respectively. The cells in 3 groups were collected at 24 hours. In addition, the cells in group A were collected at 0, 1, 2, 6, 12, 24, and 48 hours, respectively. Normal Fb was cultured with the conditioned medium containing 50% heat injured Fb culture supernatant. Then, the cells were collected at 0, 1, 2, 6, 12, 24, and 48 hours, respectively. The mRNA levels of the collagen type I, collagen type III, and MMP-1 of Fb were measured by real-time fluorescent quantitative PCR techniques.
Results: At 24 hours after cultured with supernatant of heat injured KC, mRNA relative expression levels of collagen type I, collagen type III, and MMP-1 in group A were significantly higher than those in groups B and C (P < 0.05). The mRNA relative expression levels of collagen type I, collagen type III, and MMP-1 in group A gradually increased with time going, showing significant differences between 0 hour and 2, 6, 12, 24, and 48 hours (P < 0.05); significant differences were found between different time points after 2 hours (P < 0.05). After Fb was treated with supernatant of heat injured Fb, the mRNA relative expression levels of MMP-1 gradually decreased with time going, showing significant differences between 0 hour and 1, 2, 6, 12, 24, and 24 hours (P < 0.05); after 2 hours of culture, significant differences were found among different time points (P < 0.05).
Conclusion: Heat injured KC supernatant may regulate the mRNA expressions of collagen type I, collagen type III, and MMP-1 of Fb.