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ObjectiveTo investigate the effect of heat treatment combined with narrow band ultraviolet B(NB-UVB) on cultured normal human melanocytes in vitro.MethodsMelanocytes were isolated from the foreskin of normal human,cullured in vitro,and irradiated with NB-UVB of different doses(20,30,50,70,90,120 and 180 mJ/cm2).Then,MTT assay was performed to evaluate the proliferation and activity of melanocytes to determine the optimal dose of UVB for the next experiment.Melanocytes were classified into 3 groups to be treated with heat at 42 ℃ for 1 hour (heat group),irradiated with UVB at 50 mJ/cm2 (UVB group),or irradiated with UVB at 50 mJ/cm2 followed by heat treatment at 42 ℃ for 1 hour (combination group),daily for 3 successive days; those receiving no treatment served as the control.After 24-hour culture following the last treatment,tyrosinase activity was evaluated with L-dopa as the substrate,melanin content was detected by NaOH assay,and cell cycle stages were determined by flow cytometry.ResultsNB-UVB irradiation decreased the viability of melanocytes in a dose-dependent manner,and the optimum dose of UVB was 50 mJ/cm2.The tyrosinase activity of melanocytes was 0.244 ± 0.018 and 0.310 ± 0.015 respectively in the UVB group and combination group,and increased by 3.8％ (P ＜ 0.05) and 31.9％ (P ＜ 0.05) respectively compared with the control group (0.235 ± 0.018); the melanin content was 0.201 ± 0.016 and 0.286 ± 0.019,respectively in the UVB group and combination group,and increased by 17.5％ (P ＜ 0.05 ) and 67.3％ (P ＜ 0.05) compared with the control group (0.171 ± 0.016).In comparison with the control group,the percentage of melanocytes in G1 phase was decreased by 23.94％ in the UVB group(P＜ 0.05) and 33.51％ in the combination group(P ＜ 0.05),while that in S phase and G2 phase increased by 15.35％ (P ＜ 0.05 ) and 11.93％ (P ＜ 0.05),respectively in the UVB group,and 17.76％ (P ＞ 0.05) and 16.08％ (P ＞ 0.05),respectively in the heat group.ConclusionHeat treatment and NB-UVB can synergistically enhance the tyrosinase activity and accelerate melanogenesis,proliferation and differentiation,of melanocytes.
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ObjectiveTo explore the effects of heat treatment and ultraviolet B (UVB) radiation alone or in combination on the expression of heat shock protein (HSP) 72 in human epidermal melanocytes.Methods Melanocytes were obtained from human foreskin,and subjected to primary culture.After 3 to 5 passages,the melanocytes were classified into 4 groups:control group (receiving no treatment),heat treatment group (treated with heat at 42 ℃ for 1 hour every day for 3 days),UVB group(irradiated with UVB at 50 mJ/cm2 daily for 3days),combination group(treated with heat at 42 ℃ for 1 hour followed by irradiation with UVB at 50 mJ/cm2daily for 3 days).After another 2- to 6-hour culture following the last treatment,melanocytes were collected and subjected to real time PCR and Western blot for the detection of HSP72 mRNA and protein expression,respectively.ResultsThe mRNA and protein expressions of HSP72 were significantly higher in the heat treatment group and combination group than in the control group (mRNA:6.584 ± 0.871 and 7.269 ± 0.454 vs.0.975 ± 0.089,both P ＜ 0.001; protein:2.022 ± 0.058 and 2.080 ± 0.045 vs.0.532 ± 0.033,both P ＜ 0.001 ),but was similar between the UVB group and control group (mRNA:0.832 ± 0.084 vs.0.975 ± 0.089,P ＞ 0.05;protein:0.546±0.021 vs.0.532 ± 0.033,P ＞ 0.05).The ANOVA of factorial design showed that neither heat treatment nor UVB irradiation had interaction effect on the mRNA or protein expression of HSP72 (F =2.106,1.399 respectively,both P ＜ 0.05).ConclusionsHeat treatment can cause an increase in the expression of HSP72,which may enhance the function of melanocytes and protect melanocytes from UVB induced damage.
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ObjectiveTo observe the increasing effect of infrared irradiation on tyrosinase activity and melanin content in cultured normal human epidermal melanocytes in vitro and to explore the optimal dose of infrared irradiation.MethodsEpidermal melanocytes were isolated from normal human foreskin tissue,and subjected to primary culture.Methyl thiazolyl tetrazolium(MTT) assay was performed to evaluate the effect of different doses(0,20,60,80,100,140,240,320 J/cm2)of infrared light on the proliferation of melanocytes and to select the optimal irradiation dose.Then,melanocytes were irradiated with infrared light at the optimal dose for 3 consecutive days followed by the determination of tyrosinase activity,melanin content,and cell cycle via dopa oxidation assay,NaOH solubilization method and flow cytometry,respectively.ResultsThe best intervention dose of infrared light was 80 J/cm2.The tyrosinase activity(A492 nm) and melanin content(A492 nm)were 0.3601 ± 0.0301 and 0.2748 ± 0.0243 respectively in melanocytes after irradiation with infrared light of 80 J/cm2 for 3 days,significantly higher than those in unirradiated melanocytes(0.3114 ± 0.0341,0.2325 ±0.0254,respectively,both P ＜ 0.05),with an increase rate of 15.64％ and 18.19％ respectively.Cell cycle analysis revealed a decline in cell percentage in G1 phase(P ＜ 0.01 ) but a concomitant increase in cell percentage in G2 and S phase (both P ＜ 0.05) in irradiated melanocytes compared with unirradiated melanocytes.ConclusionsThe optimal dose of infrared light is 80 J/cm2 for the irradiation of melanocytes,and this dose of infrared light can increase melanin content,tyrosinase activity,differentiation and proliferation of melanocytes.
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Objective To investigate the effect of heat treatment on the proliferation of, melanin synthesis and tyrosinase activity in cultured normal human melanocytes. Methods Normal human foreskin tissue was obtained by sterile circumcision and melanocytes were harvested by using methods for epidermal cell culture. Basic fibroblast growth factor (bFGF) was utilized as the primary mitogen to establish the culture system of normal human epidermal melanocytes. Masson-Fontana staining was proformed to identify melanocytes.Third-passage melanocytes were treated with hyperthermia at various temperatures (39 ℃, 41 ℃, 42 ℃, 43 ℃ and 45℃) for 1 hour a day for consecutive 3 days followed by the measurement of cell viability with MTT assay. The hyperthermia at optimized temperature was used to treat fourth-passage melanocytes for 1 hour a day for consecutive 3 days; subsequently, the tyrosinase activity were detected with L-Dopa as the substrate, and melanin content was determined in heat-treated and untreated (control) melanocytes. Results The hyperthermia at 42 ℃ exhibited the strongest promotive effect on the proliferation of melanocytes among these 5 hyperthermia conditions. After treatment with hyperthermia at 42 ℃ for 1 hour a day for consecutive 3 days, melanocytes showed an increment in tyrosinase activity by 36.4% and melanin synthesis by 78% compared with the untreated melanocytes (both P＜0.05). Conclusions Heat treatment can enhance the proliferation of cultured human melanocytes, promote their melanin synthesis and tyrosinase activity.
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Objective To investigate the early diagnosis of vitiligo and its differential diagnosis from other depigmentated diseases using polarized light dermoscopy(PD)imaging analysis.Methods Patients with localized depigmented macules were enrolled into this study.PD was used to observe the micromorphology,feature and color of skin lesions.Histopathology was performed to confirm the diagnosis of all cases except for those of pityriasis versicolor which were confirmed by clinical and laboratory examination.Results Of the 176 patients.97 were diagnosed as vitiligo.Residual perifollicular pigmentation Was observed in 91.94%(57/62)of patients with progressing vitiligo and 62.86%(22/35)of those with stable vitiligo,with significant difference between the two groups of patients(P<0.05).However.residual perifollicular pigmentation was absent in the 79 non-vitiligo depigrnented cases.The presence of telangiectasia,early reservoirs of pigmentation and perilesional hyperpigmentation were related to the stage of vitiligo and treatment history of patients.Conclusions PD,which efficiently eliminates the interference of reflected light on skin lesions of vililigo,is an imaging technique that allows for the visualization of minor structures and features of the skin lesions that are indiscernible to naked eyes.In a nutshell,the application of PD has offered references to the early diagnosis of vitiligo and its differential diagnosis from other depigmentation diseases.
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Objective To investigate the value of image analysis technology of polari-light dermoscopy (PD) in diagnosing head and face tumors. Methods We randomly collected the head and face tumors from the patients in this hospital, and then diagnosed them with naked eyes and PD, respectively. After making sure the diagnosis by histopathology, we analyzed the results retrospectively. Results We summarized the diagnostic signs of 211 head and face tumors on dermoscopy, which shown a great specificity. Its diagnosis rate reached to 92.89 %, obviously higher than that (69.67 %)with naked eyes. Due to clinical misdiagnosis and erroneous treatment, the recurrence rate of skin lesions was 19.91 %. The top 3 of head and face tumors, in turn, was seborrheic keratosis, melanocytic tumor and basal cell carcinoma. Conclusions PD is a non-invasive image analysis technology, which is especially applicable to the early diagnosis of head and face tumors. It can make sense in decreasing blind biopsy, directing surgical excision areas effectively, and selecting reasonable remedy. It also has great significance in the recovery and beauty of skin lesions.
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Objective To investigate the role of heat shock poteins(HSPs)in the pathogenesis of psoriasis.Methods Expression of HSP27,HSP70and HSP60was detected by immunohistochemical and image analysis in epidermal keratinocytes of pre-and post-treatment psoriatic lesional,and non-lesional skin in25psoriatic patients and6healthy controls.Results There was constitutive expression of HSP27and HSP70in epidermal keratinocytes of psoriatic non-lesional skin and normal controls,and the weak expression in psoriatic lesions.There was expression of HSP60in keratinocytes of psoriatic skin,but no expression of HSP60in non-lesional and normal skin.Expression of HSP27and HSP70recovered gradually in post-treatment psoriatic lesions,and no expression of HSP60was observed in lesional skin after treatment.Conclusion HSPs may play a certain role in psoriatic stress protective mechanism.
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Objective To investigate the effects of pituitary adenylate cyclase activating polypeptide (PACAP) and its receptor (PACAP-R) on the pathogenesis of psoriasis. Methods The expression of PACAP and PACAP-R in the skin from 10 normal controls, 25 psoriatic lesions and non-lesional skins was measured by immunohistochemical technique. Results The expression of PACAP and PACAP-R was significantly lower in the psoriatic lesional skins than that of the non-lesional skins. The area density and mean absorbance of PACAP and PACAP-R in the lesional skins were significantly lower compared with those in the non-lesional skins (P