Silver nanoparticles/chitosan oligosaccharide/poly(vinyl alcohol) nanofiber promotes wound healing by activating TGFβ1/Smad signaling pathway

Abstract

Wound healing occupies a remarkable place in everyday pathology and remains a challenging clinical problem. In our previous study, we prepared a silver nanoparticle/chitosan oligosaccharide/poly(vinyl alcohol) (PVA/COS-AgNPs) nanofiber via electrospinning and revealed that it could promote wound healing; however, the healing mechanism remained unknown. Therefore, we aimed to clarify the mechanism underlying the accelerated healing effect of the PVA/COS-AgNPs nanofiber. The TGFβ1/Smad signaling pathway is actively involved in wound healing. Considering the key role of this signaling pathway in wound healing, our preliminary study showed that the TGFβ1 level was significantly increased during the early stage of wound healing. Thus, in this study, hematoxylin–eosin, Masson’s trichrome, immunofluorescent staining, hydroxyproline content, quantitative real-time polymerase chain reaction, and Western blot analyses were used to analyze the wound healing in a rat model treated with gauze, the PVA/COS-AgNPs nanofiber, and the nanofiber plus SB431542 (an inhibitor of TGFβ1 receptor kinase). The results showed that the PVA/COS-AgNPs nanofiber promoted wound healing and upregulated the expression levels of cytokines associated with the TGFβ1/Smad signaling pathway such as TGFβ1, TGFβRI, TGFβRII, collagen I, collagen III, pSmad2, and pSmad3. Inhibiting this pathway with SB431542 resulted in prevention of the PVA/COS-AgNPs nanofiber-associated salutary effects on the early stage of wound healing and relative cytokines expression. In conclusion, the wound healing effect of the PVA/COS-AgNPs nanofiber involves activation of the TGFβ1/Smad signaling pathway.

Keywords:

• wound healing
• electrospinning
• nanofiber
• silver nanoparticles
• TGFβ1
• Smad proteins

Supplementary materials

Figure S1 The histology of normal skin and the histologic associated comparison of the three groups.

Notes: (A) The HE staining of normal skin. The arrows colored with red indicates hair follicles. (B) The number of blood vessels of the same multiples’ field (×100) in normal, gauze, PVA/COS-AgNPs nanofiber, PVA/COS-AgNPs nanofiber plus SB431542 at 3, 7, 12, 18 days post-wounding. (C) The relative number of hair follicles in normal, gauze, PVA/COS-AgNPs nanofiber, PVA/COS-AgNPs nanofiber plus SB431542 at 3, 7, 12, 18 days post-wounding. (D) The relative thickness of the epidermis in normal, gauze, PVA/COS-AgNPs nanofiber, PVA/COS-AgNPs nanofiber plus SB431542 at 3, 7, 12, 18 days post-wounding. Values are mean ± standard deviation. *P<0.05 and **P<0.01 vs normal group, ^#P<0.05 and ^##P<0.01 vs PVA/COS-AgNPs nanofiber plus SB431542 group.

Abbreviations: PVA, poly(vinyl alcohol); COS, chitosan oligosaccharide; AgNPs, silver nanoparticles; ND, no data; HE, hematoxylin–eosin.

Figure S2 Collagen fiber regeneration in rat skin.

Notes: (A) Masson-trichrome staining of normal skin. Blue-green staining intensity corresponds to relative quantity of deposited collagen fiber. (B) The relative quantity of collagen in normal, gauze, PVA/COS-AgNPs nanofiber, PVA/COS-AgNPs nanofiber plus SB431542 at 3, 7, 12, 18 days post-wounding. PVA/COS-AgNPs nanofiber group showed the greatest collagen synthesis. Values are mean ± standard deviation. *P<0.05 and **P<0.01 vs normal group, ^#P<0.05 and ^##P<0.01 vs PVA/COS-AgNPs nanofiber plus SB431542 group.

Abbreviations: PVA, poly(vinyl alcohol); COS, chitosan oligosaccharide; AgNPs, silver nanoparticles.

Figure S3 Increase in deposition of collagen I and collagen III over time in the three groups.

Notes: (A) Immunofluorescent staining of collagen I and collagen III of normal skin (×800). (B) The relative collagen I fluorescence intensity in normal, gauze, PVA/COS-AgNPs nanofiber, PVA/COS-AgNPs nanofiber plus SB431542 at 3, 7, 12, 18 days post-wounding. (C) The relative collagen III fluorescence intensity in normal, gauze, PVA/COS-AgNPs nanofiber, PVA/COS-AgNPs nanofiber plus SB431542 at 3, 7, 12, 18 days post-wounding. Increased deposition of collagen I and III were observed in the PVA/COS-AgNPs nanofiber group at day 18. Values are mean ± standard deviation. *P<0.05 and **P<0.01 vs normal group, ^#P<0.05 and ^##P<0.01 vs PVA/COS-AgNPs nanofiber plus SB431542 group.

Abbreviations: PVA, poly(vinyl alcohol); COS, chitosan oligosaccharide; AgNPs, silver nanoparticles.

Figure S4 VEGF expression to study the effect of angiogenesis on wound healing.

Notes: Highest VEGF expression was observed in PVA/COS-AgNPs nanofiber group. Values are mean ± standard deviation. *P<0.05 and **P<0.01 vs gauze group, ^#P<0.05 and ^##P<0.01 vs PVA/COS-AgNPs nanofiber plus SB431542 group.

Abbreviations: VEGF, vascular endothelial growth factor; PVA, poly(vinyl alcohol); COS, chitosan oligosaccharide; AgNPs, silver nanoparticles.

Acknowledgments

We gratefully acknowledge the financial support received from the Chongqing Programs for Application and Development (cstc2014yykfA110022).

Disclosure

The authors report no conflicts of interest in this work.