Impact of phenothiazine derivatives on melanogenesis and antioxidant status of normal melanocytes
Phenothiazine derivatives are used in the treatment of schizophrenia, but also have antihistamine, antitussive and antiemetic activities. There have been appeared numerous articles for last ten years on new biological properties of phenothiazines, among them anticancer, antibacterial, antiviral, antiprionic activity and modifying multidrug resistance. The mechanism of phenothiazines action includes the ability to interact with dopamine (D1 and D2) adrenergic (α1 and α2), histamine (H1), muscarinic (M1) and serotonergic (5HT1A and 5HT2A) receptors. Unfortunately, phenothiazine derivatives treatment entails a whole range of side effects, including changes in skin and ocular pigmentation, especially in case of long-term and/or high-dose therapy.
Melanocytes are present in the skin, hair, eyes, inner ear, brain, heart, lung and adipose tissue, where as specialized, dendritic, pigmented cells play an important physiological role. They produce melanin pigment in multistage and complex process called melanogenesis. Melanin determines the colour of the skin, hair and eyes, protects against UV radiation and also has the ability to bind metal ions, drugs as well as to scavenge free radicals including reactive oxygen species. The melanogenesis process is regulated by many physical and biochemical factors like UV radiation, cyclic adenosine monophosphate and microphthalmia-associated transcription factor (MITF), which is involved in regulation of proliferation and differentiation
It has been established that free radicals can be both harmful and beneficial for cells depending on the environment. Beneficial effects of free radicals involve the important physiological role as regulatory mediators in cellular growth, proliferation, differentiation
and apoptosis. In contrast, at high concentrations, free radicals can lead to oxidative stress damaging cell structures, including lipids and membranes, proteins and nucleic acids. The cellular response to oxidative stress includes non-enzymatic agents and antioxidant enzymes, among them the most important are superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx).
The aim of this study was to evaluate the effect of phenothiazine derivatives
on biochemical processes in normal human melanocytes. Two key biochemical processes were evaluated in this study: melanogenesis as well as the process of oxidative stress induction
and free radicals scavenging. In order to achieve this purpose cellular viability, melanin content, MITF protein content, tyrosinase activity as well as antioxidant enzymes: SOD, CAT, GPx activity and hydrogen peroxide content, were determined.
On the basis of the obtained results it was found that viability of normal human epidermal melanocytes cultured in the presence of phenothiazine derivatives decreased
with increasing drug concentration. Taking into account the determined EC50 values, cytotoxicity order of the analyzed drugs can be presented as follows: fluphenazine (1,24μM) > thioridazine (2,24μM) ≥ chlorpromazine (2,53μM) ≥ perphenazine (2,76μM) >> prochlorperazine (18,49μM). It has been stated that chlorpromazine, prochlorperazine and thioridazine in low concentrations (0,0001μM, 0,001μM) stimulate melanogenesis in normal melanocytes, while fluphenazine, perphenazine, prochlorperazine, and thioridazine in higher concentrations (≥1μM) decrease tyrosinase activity, melanin and MITF protein content. Furthermore, it has been demonstrated that all analyzed phenothiazine derivatives
in concentrations ≥0,01μM are able to generate oxidative stress in normal human epidermal melanocytes. They cause an increase in SOD activity and H2O2 content, decrease in CAT activity and modulation of GPx activity.
The changes in level of cell viability as well as in melanogenesis process and antioxidant status of normal human melanocytes in response to phenothiazine derivatives treatment may explain the role of melanin and oxidative stress in the mechanisms of phenothiazines side effects directed to pigmented tissues.
Key words: phenothiazine derivatives, melanin, tyrosinase, MITF, SOD, CAT, GPx, H2O2