Influence of traditional medicines on the activity of keratinocytes in wound healing: an in-vitro study.

Natural medicinal systems such as Ayurveda and folk medicine has remedies for wound management. However, the exact cellular and extracellular mechanisms involved in the healing process and its influence on keratinocytes is less discussed. Therefore, the present study was designed to evaluate the effect of certain natural wound healing medicines on the biology of the keratinocytes/HaCaT cells. Test materials such as honey (H), ghee (G), aqueous extracts of roots of Glycyrrhiza glabra (GG) and leaves of Nerium indicum (NI) were considered. The HaCaT cells were treated with the test materials singly and in combinations (H+G, all combined [Tot]) for a specific period (24, 48, and 72 hours). The cells were then subjected to cytotoxicity/proliferation and migration/scratch assays. All the test materials, except NI, were non-cytotoxic and showed increased cell proliferation at variable concentrations. Significant observations were made in the groups treated with honey (100 µg/ml at 48 hours, P<0.05; 1,000 µg/ml at 72 hours, P<0.05), GG (all concentrations at 48 hours, P<0.05; 750 µg/ml at 72 hours, P<0.05), H+G (250 µg/ml at 24 hours, P<0.001; 500 µg/ml at 48 and 72 hours, P<0.05), and Tot (50 µg/ml at 24, 48 and 72 hours, P<0.01). In the in-vitro wound healing assay, all the treated groups showed significant migration and narrowing of the scratch area by 24 and 48 hours (P<0.001) compared to control. The results obtained from the present study signifies the positive influence of these natural wound healing compounds on keratinocytes/HaCaT cells.

The Role of Natural Medicines on Wound Healing: A Biomechanical, Histological, Biochemical and Molecular Study.

BACKGROUND: Traditional medicinal systems like Ayurveda and Indian folk medicine have used Honey, Ghee, Glycyrrhiza glabra, and Nerium indicum effectively for treating wounds. The known result of these medications is faster healing. However, the mechanism of actions at the tissue level, the biochemical and molecular mechanisms of healing is not well explored and documented. This present study was therefore designed to study the efficacy of these traditional medicines singly and in combinations on excision wounds in Wistar rats. METHODS: At two different intervals (i.e., day 8 and day 16), biomechanical, histological and immunohistochemical (IHC) parameters were assessed at the wound site. IHC focused on the inflammatory rate by evaluating the level of cytokine, IL1ß and the tissue remodeling by studying the activity of myofibroblasts. RESULTS: Rapid epithelization, better remodeling, favorable inflammatory changes and an adequate myofibroblast activity at the wound site was observed in all the treated groups compared to control. CONCLUSION: This study is therefore useful in exploring the mechanism of action of these traditional medicines and providing valuable scientific evidence.

A Comparison Between Phorbol 12 Myristate 13 Acetate and Phorbol 12, 13 Dibutyrate in Human Melanocyte Culture.

INTRODUCTION: Melanocyte culture is an integral part of the studies of skin biology and cosmetic applications. After the introduction of selective medium for the culture of human melanocyte using Phorbol 12-myristate13-acetate (PMA) in 1982, a lot of methods of culturing were tried but till date PMA is a preferred mitogen because of its cost effectiveness compared to growth factors. We have tried to preliminarily evaluate the efficacy of another phorbol ester, Phorbol 12, 13-dibutyrate (PDBu) in melanocyte culture because of its less hydrophobic nature compared to PMA. This property minimizes the trace amount of mitogen in cell culture after washing off and hence does not interfere in other biological assays. AIM: To evaluate the differences in the melanocyte survival rate, morphology and mitotic index when grown in media supplemented with PMA and PDBu. MATERIALS AND METHODS: Foreskins were collected from children undergoing circumcision. Epidermal cells were isolated from foreskin and cultured using PMA and PDBu. Melanocytes in culture were monitored for the better establishment and documented. In proliferative assay, melanocytes were treated with PMA and PDBu for 24, 48 and 72 hours and proliferation was measured using 3-(4,5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay method. RESULTS: When cultured, melanocytes acquired proliferative status and bipolar morphology quicker in PDBu medium than in PMA medium. Keratinocytes survived as contamination in PMA medium whereas PDBu medium had minimal keratinocytes. MTT assay showed that PDBu has higher proliferative induction capacity than PMA. In even lower concentration of PDBu in medium, melanocytes survived till 72 hours without significant cell loss in compared to PMA medium. CONCLUSION: PDBu can be a valuable replacement for PMA in human melanocyte culture. Higher proliferation induction, unfavourable to keratinocyte survival and less hydrophobicity make PDBu a promising alternative for quicker establishment of pure human melanocyte cultures especially in cosmetic in vitro experimental dermatology.

Evaluation of Cytotoxicity of Silorane and Methacrylate based Dental Composites using Human Gingival Fibroblasts.

AIM: The effects of leached substances from the restorative dental materials may induce local and systemic adverse effects. Thus the biological and toxic properties of the restorative dental materials must be compatible with the oral tissues or with general health. Therefore, the need for biocompatible restorative dental material implies the necessity of toxicity testing. It was the purpose of this investigation to determine and compare the possible toxic effect of silorane based composite (Filtek P90) on human gingival fibroblast (HGF) in vitro using cytotoxicity measuring parameters (MTT assay) in comparison with its methacrylate counterpart (Z100) for their viability, proliferation rate. MATERIALS AND METHODS: Fresh healthy biopsy specimens of human gingival tissue of patients were obtained. For HGF, cells were cultured in Dulbecco's modified Eagle medium and grown to sub confluent monolayers. After attaining confluence, cells were treated with different doses of the Filtek P90 or Z 100 for different time point. HGF cells were observed for their proliferation, viability by MTT assay. RESULTS: The results of the cytotoxicity assay showed that, the percentage of viable cells was very good in the first 24h and marginally decreased in the next 48h period in all groups. However, the proliferation rate was never below 84% in all the groups, at any given concentration. Filtek P90 and Z100 treated cells exhibited insignificant decrease in the cell proliferation both in 24h and 48h exposure when compared to significant decrease in the cell survival rate in the positive control (Mitomycin C 250 µg/ml).) Comparison of the toxicity between Filtek P90 and Z100 in 24h & 48h separately showed that there was no significant difference (p<0.05) between these two composites in 24h and 48h' time period at all concentrations of the composites. CONCLUSION: To conclude, the new silorane based restorative composite showed comparable cytotoxic characteristics to clinically successful dimethacrylate composites suggesting the non-toxic nature in the oral environment and hence contributing to clinical success of these new restorative materials.