The Non–neuronal and Nonmuscular Effects of Botulinum Toxin

An Opportunity for a Deadly Molecule to Treat Disease in the Skin and Beyond

S.A. Grando; C.B. Zachary

Disclosures

The British Journal of Dermatology. 2018;178(5):1011-1019. 

In This Article

Experimental Studies of Cutaneous Effects of Botulinum Neurotoxins

The results of in vivo and in vitro experiments have identified a number of direct effects of BoNT/A on non–neuronal cells in the skin, which help to explain the mechanisms and therapeutic benefits of BoNTs for nontraditional dermatological applications. Current literature on the experimental use of BoNT/A discussed below demonstrates its ability to protect skin flaps, facilitate wound healing, decrease the thickness of hypertrophic scars and produce an anti–ageing effect. Moreover, BoNT/A has been shown to decrease the infiltration of cutaneous lymphocytes and improve acanthosis in the KC–Tie2 mouse model of psoriasiform dermatitis,[59] protect from atopic dermatitis–like skin lesions of NC/Nga mice,[60] and produce an anti–itch effect in acute and mouse models of chronic dry skin itch.[61] The latter two functions may be related, in part, to the ability of BoNT/A to decrease mast–cell activity.[62]

Cutaneous Effects of Botulinum Neurotoxin A Action on Cutaneous Vasculature

In studies on skin–flap survival in rats, BoNT/A was injected into the entire flap and compared with a control group that received a saline injection.[63] The BoNT/A group showed nearly complete survival of the flaps owing to increased perfusion compared with the control group. The protective effects of BoNT/A on flap survival in rats have been independently reported by several research groups.[64–66] In one study, BoNT/A was used to prevent unfavourable effects of cigarette smoke in Wistar albino rats.[67] In another study, it increased skin–flap viability in diabetic rats, which was associated with an increase of lumen diameter, external arterial diameter and lumen/wall thickness ratio.[68] At the molecular level, BoNT/A affected the expression of vascular endothelial growth factor, platelet endothelial cell adhesion molecule 1, CD31, CD34, interleukin (IL)–1 and tumour necrosis factor–α in cutaneous flaps.[69–71] BoNT/A reduced the hypoxic area and the number of oxidative stress–associated DNA–damaged cells and apoptotic cells in the cutaneous ischaemia–reperfusion injury sites.[72] In an in vitro assay, BoNT/A significantly prevented the oxidant–induced intracellular accumulation of reactive oxygen species in vascular endothelial cells.[72]

Cutaneous Effects of Botulinum Neurotoxin A Action on Remodelling of Dermal Connective Tissue

The direct effects of BoNT/A on dermal fibroblasts are well characterized and may mediate its pharmacological effects on dermal tissue remodelling, which is characteristic of skin ageing, wound closure and scar formation. In cultured 3T3 fibroblasts, BoNT/A induced morphological changes, such as a loss of normal fibroblast morphology and the cytoplasmic retraction and spread phenomena.[73] The antiphotoageing potential of BoNT/A through a reduction in senescence–related proteins has been demonstrated in the studies of ultraviolet–B–induced premature senescence of human dermal fibroblasts in vitro. The irradiated fibroblasts that were additionally treated with BoNT/A demonstrated a decrease in the expression of senescence–associated β–galactosidase, levels of tumour suppressor and senescence–associated proteins, G1 phase cell proportion and secretion of matrix metalloproteinase (MMP)–1 and MMP–3, and also had an increase in the production of collagen types I and III.[74]

BoNT/A has been shown to alter fibroblast activities associated with wound healing. In cultures of human fibroblasts, BoNT/A inhibited expression of collagen types I and III, but enhanced the expression of MMP–2 and MMP–9.[75] Gelatin zymography experiments confirmed enhanced MMP–2 activity in collagen degradation. BoNT/A also inhibited phosphorylation of Smad2 in vitro, and inhibited silicon–induced capsule formation through the transforming growth factor (TGF)–β/Smad signalling pathway in vivo.[75] According to one study, BoNT/A may interrupt the differentiation of fibroblasts to myofibroblasts by blocking TGF–β1 signalling.[76] However, in another study, BoNT/A upregulated the expression of type I collagen and decreased the production of some MMPs.[77] In Wistar rats, BoNT/A injections reduced wound and graft contraction,[78] and in another rat model of burn wound healing, BoNT/A improved both the healing process and the cosmetic appearance of the burn scar, and was associated with faster regeneration, less inflammation and an increase in the number of fibroblasts.[79]

BoNT/A decreases the thickness of hypertrophic scars, suggesting that its use may prevent hypertrophic scars after trauma, burns or surgery. Indeed, in the rabbit ear hypertrophic scar model, BoNT/A injection decreased scar thickness.[80] The mechanisms apparently involve the ability of BoNT/A to alter major biochemical events associated with fibroblast growth and differentiation. It has been documented that BoNT/A upregulates Rac1, Cdc42 and RhoA gene expression in a dose–dependent manner in human dermal fibroblasts in vitro,[81] inhibits fibroblast proliferation and fibroblast–to–myofibroblast differentiation,[82] induces apoptosis and decreases expression of α–smooth muscle myosis and myosin–II, regulates S100A4 and collagen type I, and downregulates the TGF–β1, VEGF, MMP–1 and PDGFA genes, and some other genes relevant to invasive growth in keloid fibroblasts.[83–85] Furthermore, treatment with BoNT/A affected cell cycle distribution, slowed proliferation and inhibited connective tissue growth factor expression in fibroblasts isolated from tissue specimens of hypertrophic scar.[86–88]

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