Effects of immune responses mediated by topological structures of three-dimensional bioprinted scaffolds on hair follicle cycle in mice

Abstract

Objective: To explore the effects of the immune responses mediated by topological structures of three-dimensional bioprinted scaffolds on hair follicle cycle in mice. Methods: The study was an experimental research. The alginate-gelatin composite hydrogels were printed into scaffolds using a three-dimensional bioprinter and named T45 scaffolds, T60 scaffolds, and T90 scaffolds according to the 3 topological structures of the scaffolds (the rotation angles of the printhead during printing were 45°, 60°, and 90°, respectively), and the morphology of the three scaffolds was observed after cross-linking by naked eyes. Nine 8-week-old female C57BL/6J mice were divided into T45 group, T60 group, and T90 group, according to the random number table, with three mice in each group, and the T45, T60, and T90 scaffolds were subcutaneously implanted on the back of mice, respectively. On post implantation day (PID) 7, the hair growth in the dorsal depilated area of mice was observed, the thickness of the fiber capsule around the scaffolds was observed by hematoxylin-eosin staining, and the expression levels of CD68, bone morphogenetic protein-2 (BMP-2), and tumor necrosis factor (TNF) protein in the tissue surrounding the scaffolds were observed by immunofluorescence staining. The samples of the above experiments were all 3. Results: The topological structures of the three scaffolds were all clear with high fidelity after cross-linking. On PID 7, the hair growth was obvious in the dorsal depilated area of mice in T45 group and T90 group, while hair growth was slow in the scaffold implantation area of mice in T60 group, which was significantly different from that of the unimplanted area. On PID 7, compared with (18±4) μm in T90 group, the thickness of both the fiber capsule around the scaffolds ((39±4) and (55±8) μm) of mice in T45 group and T60 group was significantly increased (P<0.05); the thickness of the fiber capsule around the scaffolds of mice in T60 group was also significantly increased compared with that in T45 group (P<0.05). On PID 7, the expression level of CD68 protein in the tissue surrounding the scaffolds of mice in T60 group was significantly higher than the levels in T45 group and T90 group (with both P values <0.05). The expression level of BMP-2 protein in the tissue surrounding the scaffolds of mice in T60 group was significantly higher than the levels in T45 group and T90 group (with both P values <0.05), and the expression level of BMP-2 protein in the tissue surrounding the scaffolds of mice in T45 group was significantly higher than that in T90 group (P<0.05). The expression level of TNF protein in the tissue surrounding the scaffolds of mice in T60 group was significantly lower than the levels in T45 group and T90 group (with both P values <0.05). Conclusions: Three-dimensional bioprinted scaffolds with different topological structures mediate different degrees of immune responses after being implanted in mice. A moderate immune response promotes hair growth in depilated area of mice, while an excessive immune response results inhibits the hair follicle entering into the anagen phase.