Effects of moxibustion at different temperature on wound healing and apoptosis in rats with pressure ulcer based on PI3K/AKT/mTOR signal pathway

Objective

To investigate the effects of moxibustion on wound healing and evaluate whether the efficacy of moxibustion at different temperatures varies. Additionally, to explore the mechanism by which moxibustion regulates the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian rapamycin target protein (mTOR) signaling pathway on wound healing and cell apoptosis.

Methods

A total of 90 SPF-grade SD rats were randomly divided into six groups (15 rats per group): control group, model group, recombinant bovine basic fibroblast growth factor gel (rb-bFGF) group, moxibustion group A [temperature (42 ± 1) °C], moxibustion group B [temperature (45 ± 1) °C], and moxibustion group C [ temperature (48 ± 1) °C]. Except the control group, the interventions were conducted in the other groups with pressure ulcer induced by ischemia/reperfusion injury. In the control group, povidone-iodine was administered at the knee joint. In the model group, after povidone-iodine applied at the pressure ulcer, petroleum jelly gauze was bandaged. In the rb-bFGF group, after povidone-iodine applied at the pressure ulcer, rb-bFGF was smeared and petroleum jelly gauze was bandaged. In the moxibustion group A, the suspending moxibustion with moxa stick was delivered 3 cm to 5 cm above the ulcer. The paperless temperature recorder probe was placed on the wound surface where moxibustion was delivered, and the real-time temperature was monitored and adjusted, kept at (42 ± 1) °C. The moxibustion temperature was kept at (45 ± 1) °C in the moxibustion group B, and (48 ± 1) °C in the moxibustion group C. The intervention lasted 14 days in each group. The wound healing rates were compared on the 3, 5, 7 and 14 days after the first intervention among the 6 groups. After 14-day intervention respectively, the histopathological changes were observed with HE staining, apoptosis was detected with TUNEL method and flow cytometry; VEGF-A protein expression determined with immunohistochemistry; the contents of Interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and transforming growth factor-β (TGF-β) in serum with ELISA, and PI3K/AKT/mTOR related protein and mRNA expression was with Western blotting and qRT-PCR in wound tissues of each group.

Results

Regarding the intervention effects, (1) compared with the control group, the model group showed significant inflammatory infiltration in wound tissues, higher levels of cell apoptosis, and elevated serum levels of IL-6, TNF-α and TGF-β (P < 0.05); (2) compared with the model group, the rb-bFGF group and 3 moxibustion groups showed higher wound healing rates (P < 0.05), reduced inflammatory infiltration in the wound tissues, lower levels of cell apoptosis, and decreased serum levels of IL-6, TNF-α and TGF-β (P < 0.05). Among the moxibustion groups, group C showed significantly better results than group A (P < 0.01). Regarding the mechanism, (1) compared with the control group, the model group exhibited significantly higher protein and mRNA expression levels of PI3K/AKT/mTOR signal pathway components (P < 0.05); (2) compared with the model group, the rb-bFGF group and 3 moxibustion groups showed lower protein and mRNA expression levels of PI3K/AKT/mTOR signal pathway components (P < 0.05). Among the moxibustion groups, group C showed significantly higher expression levels than group A (P < 0.01).

Conclusion

Moxibustion significantly promotes wound healing, with high-temperature moxibustion being more effective than low-temperature moxibustion in reducing the expression of inflammatory factors and cell apoptosis, thereby improving wound healing rates. It is presented that the higher moxibustion temperature, the higher wound healing rate, which may be associated with the regulation of PI3K/AKT/mTOR signal pathway with moxibustion.