An In-Vitro Assay Estimating Changes in Melanin Content of Melanoma Cells due to Ultra-Dilute, Potentized Preparations

Abstract

Introduction The authors had previously conducted an in-vitro study to observe the effect of homeopathic medicines on melanogenesis, demonstrating anti-vitiligo potential by increasing the melanin content in murine B16F10 melanoma cells. A similar experiment was performed using further homeopathic preparations sourced from kojic acid (KA), hydrogen peroxide (H₂O₂; HP), 6-biopterin (BP), and [Nle⁴, D-Phe⁷]-α-melanocyte-stimulating hormone (NLE), some of which are known to induce vitiligo or melano-destruction at physiological dose.

Materials and Methods The homeopathic preparations of BP, KA, NLE, and HP were used in 30c potency. Alcohol and potentized alcohol were used as vehicle controls. Prior to starting the main experiment, the viability of B16F10 melanoma cells after treatment with study preparations was assayed. Melanin content (at 48 h and 96 h) and tyrosinase activity in melanocytes were determined.

Results At the end of 48 hours, NLE and HP in 30c potency had a significantly greater melanin content (p = 0.015 and p = 0.039, respectively) compared with controls; BP and KA in 30c potency had no significant effects. No significant changes were seen at the end of 96 hours. KA, NLE, HP, and vehicle controls showed an inhibition of tyrosinase activity.

Conclusion The study demonstrated melanogenic effects of two homeopathic preparations. Further research to evaluate the therapeutic efficacy of these medicines is warranted.

Keywords

melanin - melanoma cells - vitiligo - ultra-dilution - potentization - homeopathy

Highlights

• The present study tested the effect of four highly diluted homeopathic medicines on melanogenesis in murine melanoma cells in vitro.

• Two of the homeopathic preparations produced an increase in melanin content, demonstrating their potential in the treatment of vitiligo.

• References

• 1 Kumar S, Nayak CS, Padhi T. , et al. Epidemiological pattern of psoriasis, vitiligo and atopic dermatitis in India: hospital-based point prevalence. Indian Dermatol Online J 2014; 5: S6-S8
• 2 Shah R. Vitiligo. 1st ed. Mumbai, Maharashtra: Life Force Molecules Pvt. Ltd; 2013
• 3 Toyoda M, Luo Y, Makino T, Matsui C, Morohashi M. Calcitonin gene-related peptide upregulates melanogenesis and enhances melanocyte dendricity via induction of keratinocyte-derived melanotrophic factors. J Investig Dermatol Symp Proc 1999; 4: 116-125
• 4 Mahesh S, Mallappa M, Tsintzas D, Vithoulkas G. Homeopathic treatment of vitiligo: a report of fourteen cases. Am J Case Rep 2017; 18: 1276-1283
• 5 Joshi S, Munshi R, Talele G, Shah R. Evaluation of melanogenic and anti-vitiligo activities of homeopathic preparations on murine B16f10 melanoma cells. Eur J Pharm Med Res 2017; 4: 718-723
• 6 Chang TS. Natural melanogenesis inhibitors acting through the down-regulation of tyrosinase activity. Materials 2012; 5: 1661-1685
• 7 Liu CH, Lin CH, Tsai MJ. , et al. Melanin bleaching with dilute hydrogen peroxide. Appl Immunohistochem Mol Morphol 2013; 21: 275-279
• 8 Schallreuter KU, Büttner G, Pittelkow MR, Wood JM, Swanson NN, Körner C. Cytotoxicity of 6-biopterin to human melanocytes. Biochem Biophys Res Commun 1994; 204: 43-48
• 9 Barnetson RS, Ooi TK, Zhuang L. , et al. [Nle4-D-Phe7]-α-melanocyte-stimulating hormone significantly increased pigmentation and decreased UV damage in fair-skinned Caucasian volunteers. J Invest Dermatol 2006; 126: 1869-1878
• 10 Shah R. Standardization of the potentizing machine and quantification of impact of potentization. Indian J Res Homoeopathy 2016; 10: 126-132
• 11 Tada H, Shiho O, Kuroshima K, Koyama M, Tsukamoto K. An improved colorimetric assay for interleukin 2. J Immunol Methods 1986; 93: 157-165
• 12 Tsuboi T, Kondoh H, Hiratsuka J, Mishima Y. Enhanced melanogenesis induced by tyrosinase gene-transfer increases boron-uptake and killing effect of boron neutron capture therapy for amelanotic melanoma. Pigment Cell Res 1998; 11: 275-282
• 13 Hyun SK, Lee WH, Jeong DM, Kim Y, Choi JS. Inhibitory effects of kurarinol, kuraridinol, and trifolirhizin from Sophora flavescens on tyrosinase and melanin synthesis. Biol Pharm Bull 2008; 31: 154-158
• 14 Chikramane PS, Suresh AK, Kane SG, Bellare JR. Metal nanoparticle induced hormetic activation: a novel mechanism of homeopathic medicines. Homeopathy 2017; 106: 135-144
• 15 Shah R. Hydroquinone: Homoeopathic Pathogenetic Trial. Indian J Res Homoeopathy 2013; 7: 47-61
• 16 Allen TF. The Encyclopedia of Pure Materia Medica, Introduction. Presented by Médi-T. Available at: http://www.homeoint.org/books1/allenhandbook/a/ars-s-f.htm