Porcine placenta extract promotes keratinocyte and fibroblast activities via ERK AKT and JNK growth signaling pathways

Abstract Background: Porcine Placenta Extract (PPE) has been disclosed as a biological protein-enriched compound, whereas it has been declared the stimulatory effects on cell proliferation, cell migration, angiogenesis, and various cellular mechanisms. The fibroblast and keratinocyte are epidermal cells involve in granulation tissue formation and re-epithelialization. The homeostasis of extracellular matrix (ECM) deposition/degradation also attributes wound completion by regulation of ECM protein constitution and matrix metalloprotease (MMP) activities. This study aims to investigate the stimulatory effect of PPE on keratinocyte/fibroblast proliferation and migration including MMPs and ECM protein expression. Methods: The human keratinocyte cell line (HaCaT) and human fibroblast cell line (Hs 895.Sk) were cultured with various PPE concentration and investigated cell proliferation, cell migration, MMP gene expression and ECM proteins’ gene expression by MTT assay, scratching wound assay, and qRT-PCR, respectively. ERK1/2, p-ERK1/2, Akt, p-Akt, JNK, p-JNK, and cyclin-D1 were also investigated by Western blot analysis. LY294002, PD98059, and SP600125, in which inhibitors were utilized to confirm the underlying signaling pathway of PPE stimulation. Jcl:SD rat were generated a wound and applied topical PPE with various concentration. The wound biopsy and histological staining with Masson’s trichrome were performed to investigate granulation tissue appearance. All In Vivo procedures were approved by the Ethics Committee for the Use of Animals of the Naresuan University (NU-AE630609). Results: PPE statistically significantly increased keratinocyte/fibroblast proliferation and migration with dose dependent manner. PPE 10 µg/mL statistically significantly enhanced MMP-1, MMP-2, MMP-9, MMP-10, and MMP-14 gene expression in keratinocyte. PPE 50 µg/mL also increased MMP-2, MMP-20, and MMP-14 including α-SMA, fibronectin, collagen I, and collagen III gene expression with statistically significant difference. PPE can activate phosphorylation of ERK1/2, Akt, and JNK including cyclin-D1 expression. Interestingly, PPE also stimulate granulation tissue appearance. Conclusion: PPE stimulated fibroblast/keratinocyte proliferation and migration through JNK, ERK1/2, and Akt pathway. PPE also activated MMP gene expression either keratinocyte or fibroblast and stimulated ECM protein gene expression in fibroblast. Consequently, PPE provides the potential as a complementary treatment to improve delayed wound healing and prevent scar/keloid formation.