Fig 1: Effects of Hst1-MAD on acute wound healing. a Representative photographs of acute skin wounds in mice without treatment or treated with 10 µmol/L Hst or 1 µmol/L Hst1-MAD1 for 3 d (n = 9/group). b Wound healing percentages (%) in all groups on days 3, 5 and 10 post-surgeries. c Collagen fibers (blue stained) expressed in the newly formed dermal layers on day 5 post-surgery. The expression levels of collagen that were quantified using the function “count/size” of Image Pro plus and calculated using the formula: mean optical density (MOD) = integrated option density (IOD) sum/area sum. The sections were colored using Masson's trichrome staining. Scare bar = 100 µm (n = 6/group). d CD31-positive vessels and the positive expression of vascular endothelial growth factor (VEGF, green arrows) on day 10 post-surgery. The sections were immunohistochemically stained using corresponding antibodies to CD31 (GB13063; 1:300; Servicebio Inc., Boston, MA, USA) and VEGF (MA5-13182; 1:100; Thermo Fisher Scientific Co., CA, USA) and further counterstained with eosin. Scale bar = 50 µm. Angiogenesis were evaluated by the fold changes of the surface area of CD31-positive vessels and the IOD of VEGF (n = 6/group). e Expression of claudin 1 and claudin 2 (red color) in the newly formed epidermal layer (delineated in yellow curve) on day 10 post-surgery. The sections were immunofluorescently stained using respective antibodies to caludin 1 (37–4900; 1:100; Thermo Fisher Scientific Co., Shanghai, China) and claudin 2 (32–5600; 1:200; Thermo Fisher Scientific Co., Shanghai, China) (n = 6/group). f Ratio of M2 to M1 macrophages in the acute wound-surrounding tissues on day 5 post-surgery (n = 6/group). g Expression levels of endogenous antioxidant NAD(P)H quinone oxidoreductase1 (NQO1; ab28947; 1:1000; Abcam Trade Co., Shanghai, China) and a series of pro-inflammatory cytokines, such as tumor necrosis factor-a (TNF-a; ab6671; 1:1000; Abcam Trade Co., Shanghai, China), interleukin-6 (IL-6; ab9324; 1:1000; Abcam Trade Co., Shanghai, China) and macrophage inflammatory protein-1ß (MIP-1ß; C04131; 1:1000; Signalway Antibody Co., Nanjin, China). The expression of inflammatory factors of Hst1 has been detected in our previously published paper. Therefore, we did not evaluate it again in this study (n = 6/group). h Illustration of how Hst1-MAD enhanced the healing of acute skin wound. The diagram showing acute skin wound treated with Hst1-MAD (left) or without treatment (right). Data were presented as mean ± standard deviation. Statistical analyses in b, c, d, e and f were performed using one-way analysis of variance (ANOVA) with Bonferroni test as post-hoc comparison, in g were performed using t-test. *P < 0.05, **P < 0.01, ***P < 0.001 vs. control group, #P < 0.05 vs. 10 µmol/L Hst1 group