Introduction
Hydroquinone is crystalline substance that consists of a benzene ring and two hydroxyl groups. It is incorporated at various concentrations in over 200 types of hair and skin cosmetic preparations and medications. It is renowned for its skin lightening capabilities but it reportedly elicits inflammatory and allergic skin responses (Stoppler, 2006). Two types of non-keratinocytes (melanocytes and Langerhans cells) seem to be the affected cells (Fernandes et al., 2004; Kooyers and Westerhof, 2006). Information on the effect of hydroquinone on the volume densities and ultrastructure of non-keratinocytes is meager or altogether lacking.
The objective of this study is to identify the effects of topical application of 4% hydroquinone on the volume densities and electron microscopic features of melanocytes and Langerhan’s cells.
Methods
The guinea pig was used for this study because of structural similarity between its epidermis and that of humans; thirty adult male animals were divided into two groups. The first group comprised 10 animals and was used to study the normal volume densities and electron microscopic features of non-keratinocytes of the thin and thick skin. The second group of 20 animals was used to study the effect of hydroquinone on non-keratinocytes. Hydrquinone cream (4%) that was daily applied onto the skin of the ear auricle of 10 animals of this group at a dose of 1 g/cm2 for 35 days. The auricle of the remaining 10 animals was treated similarly by lipobase (the vehicle). The volume density (Vv) of melanocytes was determined by point counting on skin sections stained immunohistologically for HMB45 (Fig 1) whereas NSE (Fig 2) method was used for Merkel cells. Point counting on epidermal sheets stained by the ATPase histochemical method (Fig 3) was used to determine the volume density of Langerhan’s cells. A light microscope fitted with an eyepiece graticule of 121 points was used (Fig 3) for point counting. For electron microscopy ultrathin sections of the epidermis, stained with uranyl acetate and lead citrate, were examined with a Joel transmission electron microscope. Point counting was performed on electron micrographs superimposed with a transparent sheet of 100 equidistant points. The volume densities of non-keratinocytes and their components, the relative standard error, and the minimum sample size were obtained using histomorphometric methods.
Figure 1.
melanocytes immunohistologically staining +ve (brown) to HMB45;
Figure 2.
Merkel cells staining +ve (brown) to NSE (neuron specific enolase).
Figure 3:
ATPase +ve Langerhans cells (arrows) in an epidermal sheet. Point counting graticule is superimposed.
Results
The volume density Vv of Melanocytes as revealed by this study was highest (0.06) in the epidermis of the perinasal region and lowest in the thick skin of the foot (0.03), whereas Langerhans cell Vv was highest (0.15) in the skin of the back and lowest (0.04) in the thick skin of the footpad. Merkel cells were lacking in epidermis of the ear auricle and very scarce in other locations. Application of hydroquinone induced a notable increase in the skin thickness (403.5um thick compared to a thickness of 91um in controls). There was a notable increase in Vv of the dermis (0.7 compared to 0.55 in controls). This was apparently the result of the vascular congestion and edema noted in this in the papillary dermis. Hydroquinone as seen in this study (Fig 4) resulted in an apparent decrease in melanocyte Vv (0.063 down to 0.013) and an increase in Langerhans cell Vv (0.12 up to 0.17). Moreover, it caused an apparent decrease in volume density of melanosomes within melanocytes (From 0.1 down to 0.063). These results apparently explain the skin lightening effect and the skin reactions that hydroquinone elicits. Electron microscopic examination of hydroquinone treated epidermis revealed normal keratinocytes with reduced melanin granules and normal Langerhans cells. Melanocytes, on the other hand, showed abnormal RER and melanosomes (Fig 5). These melanosomal changes augment the hypothesis that hydroquinone affect oxidation of tyrosine and hamper formation of dopaquinone and melanin (Nordlund, 2007).
Figure 4:
Comparing Vv of Langerhans cells and melanocytes in hydroquinone treated and control skin
Figure 5:
Electron micrograph of a melanocye of hydrquinone treated skin showing deformed melanosones (arrows)
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