Pigmentation In Guinea Pig Skin Research Articles

Experimental study on stromal vascular fraction mediated inhibition of skin pigmentation in guinea pigs

Pigment disorder dermatoses are common diseases with complex mechanisms. There are various methods for the clinical treatment of pigmentation diseases, but these have a poor curative effect and many adverse reactions. Currently, looking for safe and effective whitening agents is a popular research topic. Stromal vascular fractions (SVFs) are a compound cell component of adipose-derived stem cells (ADSCs) that can promote tissue regeneration, healing, and vascularization. The purpose of this experiment was to investigate the inhibitory effect of SVFs on pigmentation in guinea pigs. After guinea pig subcutaneous fat was digested and centrifuged, SVFs were isolated and quantified. SVF was injected into the pigmentation area of the prepared guinea pig pigmentation model. The amount of inducible nitric oxide synthase (iNOS) was determined using immunohistochemical analysis, histopathological staining, and the Fontana-Masson (F-M) method for measuring melanin formation. The skin of the guinea pigs obtained stable and homogenous coloration following three treatments with narrow-band ultraviolet B (NB-UVB). Hematoxylin-eosin (HE) staining revealed that compared to the control group, the cuticle, granular layer, and spinous layer were thicker and the number of epidermal melanocytes and melanin granules increased. While the quantity of pigment granules in the treated group dramatically decreased, it did not significantly change in the blank control group. F-M staining revealed that melanin granules greatly expanded following ultraviolet irradiation and were continuously distributed in basal cells and spinous layers. The entire epidermis was evenly covered in melanin granules. The level of melanin dramatically decreased following therapy. According to immunohistochemical labeling, epidermal cells' cytoplasm and membranes are where iNOS is primarily found. In the epidermis of the irradiated group, iNOS expression was much higher than in the control group, and following treatment, it decreased in the experimental group. SVFs have a reliable treatment effect on ultraviolet B (UVB)-induced pigmentation in guinea pig skin. SVFs can significantly inhibit pigmentation, effectively shorten the fading time of pigmentation, and play a role in skin whitening, providing a new breakthrough for the treatment of pigmentation diseases.

TRPA1 promotes UVB-induced skin pigmentation by regulating melanosome luminal pH.

Melanocytes stimulated by ultraviolet radiation (UVR) produce melanin and melanosomes, which causes skin pigmentation and acts as an important physiological defence process for photoprotection. Neutral luminal pH of melanosomes is critical for providing optimal conditions for the rate-limiting, pH-sensitive melanin synthesizing enzyme tyrosinase (TYR). As a major component of extraocular phototransduction pathway, transient receptor potential ankyrin1 (TRPA1) can be activated by ultraviolet B (UVB) and reported to be expressed in melanocytes. However, whether TRPA1 is involved in the regulation of melanogenesis remains unclear. Melanogenic activity of TRPA1 was evaluated in primary normal human epidermal melanocytes (HEMs) and murine B16-F10 cell cultures, and the effects of topical applications of TRPA1 specific agonist and antagonist on UVB-induced skin pigmentation were confirmed on in vivo guinea pig models. Calcium (Ca2+ ) imaging and pH imaging were performed to analyse the effects of TRPA1 on intracellular Ca2+ concentration ([Ca2+ ]ic ) and melanosome luminal pH. TRPA1 regulated melanin synthesis, UVB-induced Ca2+ influx and melanosome luminal pH in HEMs and B16-F10 cells. Topical treatment of TRPA1 specific agonist JT010 increased UVB-induced skin pigmentation in guinea pigs, while topical using of TRPA1 selective antagonist HC-030031 mitigated such pigmentation. Our results indicated that TRPA1 activated by UVB enhanced the skin pigmentation, most likely by regulating the [Ca2+ ]ic and the melanosomal pH, consequently influencing the enzymatic activity of TYR. Therefore, the results suggest TRPA1 as a potential therapeutic target in the treatment of skin pigmented disorders that are at high risk under UVB irradiation.

Comparative Study between Topical Latanoprost versus Topical 5- Fluorouracil after Dermabrasion for Treatment of Non-Segmental Stable Vitiligo

Background: Latanoprost [LT], a prostaglandin F2-alpha [PGF2α] analogue used in the treatment of glaucoma. It induces skin pigmentation in guinea pigs. The rationale of trying 5-fluorouracil [5‐FU] in vitiligo treatment may come from the first report of hyperpigmentation induced by 5‐FU as a side effect in the skin tumors and psoriasis treatment The Aim of the work: Comparison between the efficacy of topical latanoprost versus topical 5-fluorouracil after skin dermabrasion in induction of skin repigmentation in stable non-segmental vitiligo. Patients and Methods: This study included thirty Egyptian patients, 18-60 years old, with stable non-segmental vitiligo. They were divided into: group [A] patients subjected to 5 fluorouracil after dermabrasion and group [B] patients subjected to latanoprost after dermabrasion then followed up every two weeks for three months. Result: There was no statistically significant difference between both groups regarding total response to treatment and degree of repigmentation. However, there was higher response to treatment in group A than group B. In both groups patients were satisfied with the degree of improvement. As regard complication there were minimal complications only one case of koebnerization and two cases of post inflammatory hyperpigmentation Conclusion: 5- fluorouracil 5% was associated with better improvement than latanoprost. However, the difference did not reach statistical significance. Patients reported high satisfaction with the two treatment modalities.

Inhibitory effect of a genistein derivative on pigmentation of guinea pig skin

This study demonstrated the mechanisms of the inhibitory effects of GD against UVB-induced pigmentation in guinea pig skin.

Antimelanogenic chemicals with in vivo efficacy against skin pigmentation in guinea pigs.

Ultraviolet (UV) radiation under sunlight stimulates skin pigmentation through immediately affecting the oxidative modification of existing melanin pigments and the spatial redistribution of pigmented melanosomes followed by the up-regulation of melanogenic genes in delayed kinetics. However, abnormal accumulation and synthesis of melanin biopolymers are responsible for skin disorders with more pigmented patches. Chemical-based regulation of the hyperpigmented disorders has been a long-standing goal for cosmetic and pharmaceutical applications. A large number of the chemicals with antimelanogenic activity have met with limited or no success in the treatment of skin patients, since they may not overcome the challenge of penetrating the skin barrier. Guinea pig skin displays similar kinetic parameters to human skin in the transdermal absorption of numerous chemicals, thus can serve as the surrogate for human skin. Here, we provide a concise review of our current understanding of the chemical-based therapy against skin hyperpigmentation in UV-irradiated guinea pig models, suggest molecular mechanisms of the action and emphasize the translation from preclinical outcomes to skin patients.

Inhibitory Effects of Resveratrol on Melanin Synthesis in Ultraviolet B-Induced Pigmentation in Guinea Pig Skin

Resveratrol is a polyphenolic compound found in various natural products such as grapes and berries and possesses anti-cancer, anti-hyperlipidemia, and anti-aging properties. Recently, it has been reported that resveratrol inhibits α-melanocyte-stimulating hormone signaling, viability, and migration in melanoma cells. However, these effects have not been confirmed in vivo, specifically brownish guinea pigs. To evaluate the potential of resveratrol as a regulator of melanin for hyperpigmentation therapy, the influence of resveratrol on pigmentation was investigated by ultraviolet B-induced hyperpigmentation in brownish guinea pig skin. We found that resveratrol reduced the expression of melanogenesis-related proteins tyrosinase, tyrosinase-related proteins 1 and 2, and microphthalmia-associated transcription factor in melanoma cells. Furthermore, topical application of resveratrol was demonstrated to significantly decrease hyperpigmentation on ultraviolet B-stimulated guinea pig skin in vivo. Based on our histological data, resveratrol inhibits melanin synthesis via a reduction in tyrosinase-related protein 2 among the melanogenic enzymes. This study is the first to provide evidence supporting resveratrol as a depigmentation agent, along with further clinical investigation of resveratrol in ultraviolet B-induced skin disorders such as hyperpigmentation and skin photoaging.

Inhibitory effects of salidroside and paeonol on tyrosinase activity and melanin synthesis in mouse B16F10 melanoma cells and ultraviolet B-induced pigmentation in guinea pig skin

Salidroside, the major active component of Rhodiola rosea, a herb with antioxidant, free radical scavenging and tyrosinase inhibitory effects, has been recently reported in protecting the kerationcytes from the UV radiation, suggesting the potential of this component in depigmentation. Paeonol is isolated from Moutan Cortex Radicis with anti-inflammation/microbial activities, was reported to induce the down-regulation of microphthalmia-associated transcription factor and subsequently tyrosinase. To testify the potential of these compounds as melanin formation inhibitors for hyperpigmentation therapy, the influence of salidroside and paeonol on pigmentation was investigated. With arbutin as a positive control, salidroside and paeonol were evaluated for their inhibitory effect on the cell viability, tyrosinase activity and melanin synthesis in B16F10 melanoma cells, as well as their effects in UVB-induced hyperpigmentation in brown guinea pig skins. It was demonstrated that the significant inhibition of salidroside (33.0%) and paeonol (22.2–30.9%) on the tyrosinase activity is slightly lower than that of arbutin (18.4–44.7%). However, salidroside exhibited the dose-dependent inhibition (30.6–42.0%) in melanin synthesis at a low concentration of 100μM, paeonol and arbutin expressed inhibition rates of 27.4–37.2% and 25.8–45.6% within 500–1000μM. The in vivo topical application of these compounds was demonstrated to obviously decrease the hyperpigmentation on UVB stimulated guinea pig skin. This study provided the original evidence for the salidroside and paeonol as therapeutic agents for pigmentation disorder and skin lightening, with further clinical investigation of these compounds in the field of depigmentation was suggested.

Diacylglycerol kinase regulates tyrosinase expression and function in human melanocytes

Diacylglycerol increases the melanin content of human melanocytes in vitro and increases the pigmentation of guinea pig skin in vivo, but the mechanism(s) underlying those effects remain unknown. In this study, we characterized the role of diacylglycerol kinase (DGK), which phosphorylates diacylglycerol to generate phosphatidic acid, in the regulation of pigmentation. Melanin content, tyrosinase activity and tyrosinase protein levels were significantly reduced by a DGK inhibitor, but tyrosinase and MITF mRNA levels were not changed by that inhibition, and there were no effects on the expression of other melanogenesis-related proteins.

Diacylglycerol Kinase Regulates Tyrosinase Expression and Function in Human Melanocytes

Diacylglycerol increases the melanin content of human melanocytes in vitro and increases the pigmentation of guinea pig skin in vivo, but the mechanism(s) underlying those effects remain unknown. In this study, we characterized the role of diacylglycerol kinase (DGK), which phosphorylates diacylglycerol to generate phosphatidic acid, in the regulation of pigmentation. Ten isoforms of DGK have been identified, and we show that DGKζ is the most abundant isoform expressed by human melanocytic cells. Melanin content, tyrosinase activity and tyrosinase protein levels were significantly reduced by a DGK inhibitor, but tyrosinase and MITF mRNA levels were not changed by that inhibition, and there were no effects on the expression of other melanogenesis-related proteins. Isoform-specific siRNAs showed that knockdown of DGKζ decreased melanin content and tyrosinase expression in melanocytic cells. Over-expression of DGKζ increased tyrosinase protein levels, but did not increase tyrosinase mRNA levels. Glycosidase digestion revealed that inhibition of DGK reduced only the mature form of tyrosinase and the decrease of tyrosinase resulting from DGK inhibition could be blocked partially by protease inhibitors. These results suggest that DGK regulates melanogenesis via modulation of the post-translational processing of tyrosinase, which may be related with the protein degradation machinery.

Inhibitory effect of mulberroside A and its derivatives on melanogenesis induced by ultraviolet B irradiation

Mulberroside A was isolated from the ethanol extract of Morus alba roots. The enzymatic hydrolysis of mulberroside A with Pectinex® produced oxyresveratrol and oxyresveratrol-3-O-glucoside. We tested oxyresveratrol, oxyresveratrol-3-O-glucoside, and mulberroside A to determine whether they could inhibit ultraviolet B (UVB) irradiation-induced melanogenesis in brown guinea pig skin. Topical application of mulberroside A, oxyresveratrol, and oxyresveratrol-3-O-glucoside reduced the pigmentation in guinea pig skin. These compounds suppressed the expression of melanogenic enzymes tyrosinase, tyrosinase-related protein-1, and microphthalmia transcription factor. The anti-melanogenesis effect was highest with oxyresveratrol, intermediate with oxyresveratrol-3-O-glucoside, and lowest with mulberroside A. Mulberroside A is a glycosylated stilbene of oxyresveratrol; thus, the deglycosylation of mulberroside A resulted in enhanced inhibition of melanogenesis. Histological analysis with Fontana-Masson staining confirmed that these compounds significantly reduced the melanin content in the epidermis of UVB-irradiated guinea pig skin compared to the vehicle control. Thus, these compounds effectively reduced pigmentation and may be suitable cosmetic agents for skin whitening.

Dose-effect relationship between Q-switch ruby laser and skin pigmentation in Guinea pigs

Objective To investigate the dose-effect relationship between Q-switch ruby laser and skin pigmentation in Guinea pigs. Methods Ten Guinea pigs were included in this study, and the back of each pig was divided into 5 areas, negative control area receiving no UVB irradiation or laser treatment, positive control area receiving UVB irradiation but no laser treatment, low-, middle- and high-dose areas receiving UVB irradiation followed by the treatment with laser at an energy density of 1.5, 3.0 and 6.0 J/cm2 for 4, 2, and 1 session, respectively, with an interval of 1 week. Melanin content index (MCI) was determined, and confocal laser scanning microscopy (CLSM) and histopathology were performed to observe melanin granules and melanocytes in the skin tissue from Guinea pigs before the initial laser treatment, 2 and 4 weeks after the final laser treatment. Results The best response was observed in the low-dose areas, and there was a complete clearance of hyperpigmentation in these areas, which maintained 8 weeks without recurrence, while different degrees of skin pigmentation recurred in the middle- and high-dose areas. Moreover, the low-dose areas showed a significantly lower melanin content index (MCI) compared with the middle- and high-dose areas (F = 4.2,P < 0.05), together with a marked reduction in the extent of pigment deposition, melanin granules, melanocyte number and density. Conclusions The efficacy of Q-switched ruby laser on skin pigmentation in Guinea pigs is correlated with the energy density and treatment frequency, and, low intensity and high frequency are often associated with a better treatment outcome. Key words: Lasers; Skin pigmentation; Dose-response relationship, radiation; Guinea pigs

Lightening effect on ultraviolet-induced pigmentation of guinea pig skin by oral administration of a proanthocyanidin-rich extract from grape seeds.

Antioxidants such as vitamins C and E have been reported to inhibit the progression of ultraviolet (UV) radiation-induced pigmentation in the skin of hairless mice. However, little is known of the lightening effect of proanthocyanidin, a powerful polyphenolic antioxidant, on UV-induced pigmentation of the skin. We investigated the lightening effect of oral administration of a proanthocyanidin-rich grape seed extract (GSE) using guinea pigs with UV-induced pigmentation. These pigmented guinea pigs were fed diets containing 1% GSE or 1% vitamin C (w/w) for 8 weeks. GSE-feeding had an apparent lightening effect on the guinea pigs' pigmented skin. Histologic evaluation demonstrated a decrease in the number of 3,4-dihydroxyphenylalanine (DOPA)-positive melanocytes as well as 8-hydroxy-2'-deoxyguanosine (8-OHdG)-positive, Ki-67-positive, proliferating cell nuclear antigen (PCNA)-positive melanin-containing cells in the basal epidermal layer of the UV-irradiated skin in GSE-fed guinea pigs. In contrast, these parameters did not change in the skin of vitamin C-fed or control guinea pigs. GSE inhibited the activity of mushroom tyrosinase and also inhibited melanogenesis without inhibiting the growth of cultured B16 mouse melanoma cells. In conclusion, we demonstrated that oral administration of GSE is effective in lightening the UV-induced pigmentation of guinea pig skin. This effect may be related to the inhibition of melanin synthesis by tyrosinase in melanocytes and the reactive oxygen species (ROS)-related proliferation of melanocytes.

Histamine is Involved in Ultraviolet B-Induced Pigmentation of Guinea Pig Skin

We previously reported that histamine induced melanogenesis in cultured human melanocytes and that the stimulatory effect was mediated by protein kinase A activation via H2 receptors. It is well-known that ultraviolet B irradiation causes acute inflammation, known as erythema, and subsequent pigmentation, and there are several reports demonstrating an elevation of the histamine levels in ultraviolet B-irradiated skin. Thus, to evaluate the involvement of histamine in ultraviolet B-induced skin pigmentation, we examined the effect of an H2 antagonist in brownish guinea pig skin. Daily exposure to 200 mJ per cm2 ultraviolet B for 3 d evoked erythema and subsequent pigmentation in the skin samples tested. Moreover, a remarkable increase in dopa-positive melanocytes was observed in the pigmented area, which showed an increase in melanin synthesis. Topical application of famotidine, an H2 antagonist, significantly reduced pigmentation and moderated the increase of dopa-positive melanocytes in the ultraviolet B-irradiated skin. Even when the initiation of famotidine application was delayed to day 2 after irradiation, an inhibitory activity on ultraviolet B-induced pigmentation was observed; however, the ultraviolet B-induced erythema was not suppressed by topically applied famotidine. Thus, we concluded that histamine is involved in ultraviolet B-induced pigmentation and that famotidine suppressed the pigmentation by the prevention of histamine binding to H2 receptors in melanocytes but not by prevention of ultraviolet B permeability and inflammation.

Involvement of nitric oxide in UVB-induced pigmentation in guinea pig skin.

Ultraviolet (UV) B irradiation evokes erythema and delayed pigmentation in skin, where a variety of toxic and modulating events are known to be involved. Nitric oxide (NO) is generated from L-arginine by NO synthases (NOS). Production of NO is enhanced in response to UVB-stimulation and has an important role in the development of erythema. NO has recently been demonstrated as a melanogen which stimulates melanocytes in vitro, however, no known in vivo data has been reported to support this finding. In this study, we investigated the contribution of NO with UV-induced pigmentation in an animal model using an NOS inhibitor. UVB-induced erythema in guinea pig skin was reduced when an NOS inhibitor, L-NAME (N-nitro-L-arginine methylester hydrochloride), was topically applied to the skin daily, beginning 3 days before UVB-irradiation. Delayed pigmentation and an increased number of DOPA-positive melanocytes in the skin were markedly suppressed by sequential daily treatment with L-NAME. Furthermore, melanin content 13 days after UVB-irradiation was significantly lower in skin treated with L-NAME than in the controls. In contrast, D-NAME (N-nitro-D-arginine methylester hydrochloride), an ineffective isomer of L-NAME, demonstrated no effect on these UV-induced skin responses. These results suggest that NO production may contribute to the regulation of UVB-induced pigmentation.

Topical trans-4-aminomethylcyclohexanecarboxylic acid prevents ultraviolet radiation-induced pigmentation

We have studied the effect of a plasmin inhibitor, trans-4-aminomethylcyclohexanecarboxylic acid ( trans-AMCHA), on skin pigmentation induced by ultraviolet (UV) exposure in Weiser—Maples guinea pigs. When guinea pigs are exposed to UV radiation (840 mJ cm −2), skin pigmentation is clearly observed from seven days after exposure and continued to increase to 29 days. Post-exposure applications of 2 and 3% solutions of trans-AMCHA to the exposed regions prevent or inhibit the pigmentation process. When the skin is removed and stained by the Fontana—Masson method, melanin content in the basal layer of UV-exposed epidermis is significantly reduced in the regions to which 2 and 3% trans-AMCHA solutions have been applied, compared with the vehicle control. As plasmin is known to contribute to the release of arachidonic acid (AA) and the production of prostaglandins (PGs), we have examined the effects of trans-AMCHA on AA-induced pigmentation in guinea pig skin. Topical application of trans-AMCHA causes a dose-dependent decrease in AA-induced pigmentation. These results suggest that trans-AMCHA reduces melanocyte tyrosinase activity by suppressing the production of PGs, UV-induced melanogens, through the suppression of the UV-induced increase in epidermal plasmin activity.

Topically Applied Diacylglycerols Increase Pigmentation in Guinea Pig Skin

Exposure of human and murine melanocytes in vitro to the diacylglycerol (DAG) 1-oleoyl-2-acetyl-sn-glycerol (OAG) markedly increases melanin production within 24 h. To determine whether OAG can increase melanin production in vivo, increasing concentrations of OAG (10-60 mg/ml) in propylene glycol were applied daily for 5 d to shaved guinea pigs. Dose-dependent increased pigmentation was visible first on days 17-22 and persisted for 10-14 weeks. Peak epidermal melanin content in OAG-treated sites was more than twice that of untreated or vehicle-treated sites, as assessed by computerized image analysis of Fontana-Masson stained biopsy cross sections. In another experiment to assess the mechanism of DAG-mediated pigmentation, guinea pigs received twice daily separate applications of OAG, dipalmitoylglycerol (diC16), dioctanoylglycerol (diC8), each 50 mg/ml, 20 microliters/application, and propylene glycol vehicle alone for 5 d. Increased pigmentation was visible after 10 d in the OAG and diC8 sites but not in diC16 or vehicle sites. These results correlate with the reported ability of these compounds to activate protein kinase C in vitro. In a final experiment, guinea pigs received OAG 25 mg/ml three times daily to one test site, and once daily ultraviolet B (70 mJ/cm2, equivalent to 0.6 minimal erythemal dose) radiation to another for 10 d. The OAG and ultraviolet B test sites developed comparable pigmentation by both clinical and histologic criteria. Our data demonstrate that topically applied DAGs can produce a long-lasting increase in epidermal pigmentation, presumably through protein kinase C activation, which clinically and histologically closely resembles ultraviolet-induced tanning.