Sequential transplantation of exosomes and BMSCs pretreated with hypoxia efficiently facilitates perforator skin flap survival area in rats.

Bone marrow mesenchymal stem cells (BMSC) are promising candidates for the treatment of trans-territory perforator flap necrosis. However, the low retention and survival rate of engrafted BMSCs limit their therapeutic efficacy. Strategies either modifying BMSCs or alleviating the inflammatory environment may solve this problem. Thus, we aimed to explore the therapeutic efficacy of sequential transplantation of exosomes and hypoxia pretreated BMSCs on flap necrosis. After the perforator flap model was created, the exosomes derived from BMSCs were injected immediately into choke zone II followed by transplantation of hypoxia pretreated BMSCs on Day 2. Gross view was performed to assess the flap survival, enzyme-linked immunosorbent assay was performed to evaluate the inflammatory factor level, microvessel number was assessed and quantitative polymerase chain reaction (qPCR) was performed to assess angiogenesis. We found that exosome delivery significantly reduced inflammatory cytokines levels on Day 1 and Day 3 and promoted the engrafted BMSCs' survival on Day 7. After combining with transplantation of hypoxia pretreated BMSCs, the flap survival rate and the angiogenesis-related gene expression were significantly higher than in the other three groups; the von Willebrand factor (vWF) vascular diameter and vWF vascular count were significantly higher than in the phosphate buffered saline (PBS) group. Thus, we concluded that sequential transplantation of exosomes and BMSCs combinatorially pretreated with hypoxia further facilitated flap survival. This sequential transplantation approach provides novel insights into the clinical treatment of flap necrosis.

[Effect of the botulinum toxin type A on myocutaneous flap expansion in minipigs model].

OBJECTIVE: To investigate the histologic effect of botulinum toxin type A (Botox A) injection on myocutaneous flap expansion in minipigs model. METHODS: Seven minipigs were included in this study. Two symmetric tattoo area, 10 cm x 6 cm in size, were selected on the bilateral flank of the pigs. The Botox A was injected into one tattoo area randomly, 4 U every point, 2 cm apart, with a total dose of 96 U. The same dose of sterile normal saline (0.9%) was injected in the same fashion on the opposite side as control. 3 days after injection , two 200 ml expanders were inserted beneath the cutaneous muscle at the tattoo area. After complete expansion of 200 ml, the specimens were drawn from both groups symmetrically and were stained by means of HE and Masson. The histologic changes of myocutaneous flap were compared. Thickness of each layer in myocutaneous flap was measured in histological section. RESULTS: The thickness of cutaneous muscle, capsule, dermis were (275.74 +/- 28.93) microm, (468.03 +/- 34.28) microm, (1990.79 +/- 102.10) microm in Botox group, and (409.13 +/- 44.63) microm, (626.55 +/- 44.05) microm, (2508.44 +/- 70.71) microm in saline group, respectively, show a significant difference between the two groups (P < 0.01). The Masson stained slice showed that collagen average gray of capsule in Botox group was 185.38 +/- 9.56, which was significantly higher than that in the saline group (120.77 +/- 10.31, P < 0.01). Light microscope (HE stained sections ) showed that muscle in Botox group was significantly atrophy and cross-section of muscle fiber decreased. The muscle fiber in saline group was generally normal. It was observed through transmission electron microscope that the light and dark band of muscle cell became fuzzy and the Z line bending in Botox group. The light and dark band in saline group arranged neatly, the Z line was clear. CONCLUSIONS: Application of Botox A in myocutaneous flap expansion can make the muscle atrophy and reduce the content of collagen in capsule layer, making the myocutaneous flap thinner which is suitable for reconstruction in face and neck.