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BACKGROUND@#Atopic dermatitis (AD) affects approximately 10% of adults worldwide. CM310 is a humanized monoclonal antibody targeting interleukin-4 receptor alpha that blocks interleukin-4 and interleukin-13 signaling. This trial aimed to evaluate the efficacy and safety of CM310 in Chinese adults with moderate-to-severe AD.@*METHODS@#This multicenter, randomized, double-blind, placebo-controlled, phase 2b trial was conducted in 21 medical institutions in China from February to November 2021. Totally 120 eligible patients were enrolled and randomized (1:1:1) to receive subcutaneous injections of 300 mg CM310, 150 mg CM310, or placebo every 2 weeks for 16 weeks, followed by an 8-week follow-up period. The primary endpoint was the proportion of patients achieving ≥75% improvement in the Eczema Area and Severity Index (EASI-75) score from baseline at week 16. Safety and pharmacodynamics were also studied.@*RESULTS@#At week 16, the proportion of EASI-75 responders from baseline was significantly higher in the CM310 groups (70% [28/40] for high-dose and 65% [26/40] for low-dose) than that in the placebo group (20%[8/40]). The differences in EASI-75 response rate were 50% (high vs . placebo, 95% CI 31%-69%) and 45% (low vs . placebo, 95% CI 26%-64%), with both P values <0.0001. CM310 at both doses also significantly improved the EASI score, Investigator's Global Assessment score, daily peak pruritus Numerical Rating Scale, AD-affected body surface area, and Dermatology Life Quality Index compared with placebo. CM310 treatment reduced levels of thymus and activation-regulated chemokine, total immunoglobulin E, lactate dehydrogenase, and blood eosinophils. The incidence of treatment-emergent adverse events (TEAEs) was similar among all three groups, with the most common TEAEs reported being upper respiratory tract infection, atopic dermatitis, hyperlipidemia, and hyperuricemia. No severe adverse events were deemed to be attributed to CM310.@*CONCLUSION@#CM310 at 150 mg and 300 mg every 2 weeks demonstrated significant efficacy and was well-tolerated in adults with moderate-to-severe AD.
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Adult , Humans , Dermatitis, Atopic/drug therapy , Treatment Outcome , Severity of Illness Index , Antibodies, Monoclonal, Humanized/therapeutic use , Injections, Subcutaneous , Double-Blind MethodABSTRACT
Objective:To explore the prolonged therapeutic regimen for patients with plaque psoriasis, who showed a positive response to 4-week treatment with tazarotene/betamethasone dipropionate cream, but were not completely cured.Methods:A multicenter, randomized, open-labelled, parallel-controlled clinical study was conducted. A total of 232 patients with plaque psoriasis were collected, who showed a positive response to previous 4-week treatment with 0.05%/0.05% tazarotene/betamethasone dipropionate cream, but were not completely cured with the psoriasis area and severity index[PASI] improvement rate being 50%-90%. At week 5, they were randomly and equally divided into 2 groups: test group receiving treatment with 0.05%/0.05% tazarotene/betamethasone dipropionate cream once a day, and control group receiving a sequential regimen of 0.05% tazarotene gel on weekdays once a day followed by 0.05%/0.05% tazarotene/betamethasone dipropionate cream on weekends once a day. After 2-and 4-week prolonged treatment, the efficacy and safety of the 2 therapeutic regimens were evaluated and compared. Measurement data were compared between 2 groups by using covariance analysis or t test, and enumeration data were compared by using chi-square test. Results:From the 5th to the 8th week, 200 out of the 232 patients completed the treatment. Data collected from 110 patients in the test group and 112 in the control group were enrolled into the full analysis set, and those from both 113 patients in the test group and control group were enrolled into safety analysis set. After consecutive 6-and 8-week treatment, the decline rates of the PASI score were 73.05% ± 16.69% and 78.46% ± 15.40% respectively in the test group, which were significantly higher than those in the control group (66.73% ± 21.77%, 67.02% ± 34.19%, respectively, both P < 0.05) . After 6-week treatment, the proportion of subjects who achieved PASI90 was significantly higher in the test group (14 cases, 12.7%) than in the control group (5 cases, 4.5%, χ2=4.842, P=0.028) ; After 8-week treatment, the proportions of subjects who achieved PASI75 and PASI90 (61.8%, 23.6%, respectively) were significantly higher in the test group than in the control group (48.2%, 12.5%, respectively, both P < 0.05) . During the consecutive 8-week treatment, there was no significant difference in the incidence rate of adverse reactions between the test group (15.0%) and control group (23.9%, χ2=2.822, P=0.093) . Conclusion:For patients who showed a positive response to 4-week treatment with 0.05%/0.05% tazarotene/betamethasone dipropionate cream, but were not completely cured, the continuous use of 0.05%/0.05% tazarotene/betamethasone dipropionate cream for 4 weeks is a superior therapeutic regimen compared with the sequential regimen of 0.05% tazarotene gel followed by 0.05%/0.05% tazarotene/betamethasone dipropionate cream.
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Objective:To evaluate the reporting quality of clinical guidelines on skin diseases published in journals in China.Methods:The CNKI, VIP, Wanfang and SinoMed databases were searched from January 2009 to October 2019 for clinical guidelines on skin diseases published in journals in China. Two reviewers independently screened the literature, and extracted and cross-checked data. The reporting quality of these clinical guidelines was evaluated by using the Reporting Items for Practice Guidelines in Healthcare (RIGHT) , and statistical analysis was carried out with Excel 2017 software.Results:A total of 17 clinical guidelines on skin diseases were included, including 13 Western medicine guidelines and 4 Chinese medicine guidelines. Among the 13 Western medicine guidelines, the number of guidelines reporting the following areas in the RIGHT statement, namely basic information, background, evidence, recommendations, review and quality assurance, funding and declaration and management of interests, and other information, was 9, 6, 0, 4, 0, 1 and 1 respectively; among the 4 Chinese medicine guidelines, the number of guidelines reporting the above 7 areas in the RIGHT statement was 4, 3, 3, 3, 3, 2 and 2 respectively.Conclusion:There is still considerable room for improvement in the overall reporting quality of clinical guidelines on skin diseases published in journals in China during the past 10 years, and the RIGHT statement is recommended for improving the reporting quality in guideline development.
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Objective:To determine the expression of the E2F6 transcription factor in human malignant melanoma tissues and cell lines, and to evaluate the effect of E2F6 on proliferation, migration and invasion of a malignant melanoma cell line A375.Methods:Frozen tissues and paraffin-embedded tissue sections were collected from 50 cases of cutaneous malignant melanoma and 30 cases of pigmented nevus in Department of Dermatology, Chongqing Traditional Chinese Medicine Hospital from January 2012 to December 2017. Quantitative reverse transcription-PCR (qRT-PCR) was performed to determine the mRNA expression of E2F6 in the malignant melanoma and pigmented nevus tissues, as well as in 7 malignant melanoma cell lines (HM, A375, WM451, WM35, SK-MEL-1, Hs-695T and MDA-MB-435s) and pigmented nevus cells, and immunohistochemical study and Western blot analysis were conducted to determine the protein expression of E2F6 and β-catenin in the malignant melanoma tissues. An E2F6-inhibiting plasmid and a control plasmid were separately transfected into A375 cells by using a liposome-mediated transfection method, and the E2F6 gene-knockdown efficiency was verified by qRT-PCR and Western blot analysis. Cell counting kit-8 (CCK8) assay, soft-agar plate cloning assay, Transwell migration and invasion assays and 3D cell culture assay were conducted to evaluate the effect of E2F6 gene knockdown on the proliferation, migration and invasion of A375 cells, flow cytometry was performed to detect the cell cycle and apoptosis rate, and Western blot analysis was conducted to determine the protein expression of total β-catenin, activated β-catenin, c-Myc and cyclin D1. The comparison between two groups was carried out by t test, the comparison among several groups by one-way analysis of variance, and multiple comparisons by least significant difference t test; Pearson correlation coefficient was used to analyze the correlation between E2F6 and β-catenin expression in cutaneous malignant melanoma. Results:The E2F6 mRNA expression was significantly higher in the 7 malignant melanoma cell lines than in the pigmented nevus cells (all P < 0.001). qRT-PCR showed that the relative mRNA expression of E2F6 was significantly higher in the cutaneous malignant melanoma tissues (0.000 55 ± 0.000 17) than in the pigmented nevus tissues (0.000 18 ± 0.000 09, t = 3.22, P < 0.001). Both the immunohistochemical study and Western blot analysis showed significantly increased E2F6 protein expression, but decreased β-catenin protein expression in the cutaneous malignant melanoma tissues compared with the pigmented nevus tissues (all P < 0.001). Correlation analysis showed that E2F6 protein expression was negatively correlated with β-catenin expression in the malignant melanoma tissues (immunohistochemical study: r = -0.56, Western blot analysis: r = -0.63, both P < 0.01). After knockdown of the E2F6 gene in A375 cells, the mRNA and protein expression of E2F6 was significantly lower in the E2F6 inhibition group than in the control group ( t = 3.38, 2.76 respectively, both P < 0.001). CCK8 assay showed that the cellular proliferative ability was significantly lower in the E2F6 inhibition group than in the control group ( t = 4.58, P < 0.01) 48 hours after transfection; soft-agar plate cloning assay showed that the colony-formation ratio was significantly lower in the E2F6 inhibition group than in the control group ( t = 2.26, P < 0.001) ; Transwell migration and invasion assays showed that the number of cells crossing the chamber was significantly lower in the E2F6 inhibition group (165 ± 23, 96 ± 11 respectively) than in the control group (376 ± 22, 315 ± 31, t = 3.14, 2.12, respectively, both P < 0.01) ; 3D cell culture assay showed that the cell morphology markedly changed, and the invasive pseudopodia disappeared in the E2F6 inhibition group. Flow cytometry revealed that the proportion of cells at G0-G1 phase and apoptosis rate were significantly higher in the E2F6 inhibition group than in the control group (both P < 0.001). Western blot analysis showed significantly decreased protein expression of β-catenin, activated β-catenin and its downstream target proteins c-Myc and cyclin D1, but significantly increased protein expression of P21 in the E2F6 inhibition group compared with the control group (all P < 0.001) ; additionally, the E2F6 inhibition group showed significantly decreased protein expression of epithelial-mesenchymal transition-related molecules vimentin and N-cadherin, but significantly increased expression of E-cadherin compared with the control group (all P < 0.001) . Conclusions:The E2F6 transcription factor is highly expressed in malignant melanoma. Knockdown of the E2F6 gene in A375 cells can inhibit cell proliferation, migration and invasion by antagonizing the β-catenin signaling pathway.
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Objective:To analyze clinical characteristics of cosmetics-related adverse reactions and main allergenic components of cosmetics, to provide guidance for cosmetics-related adverse reaction monitoring, and to provide an objective basis for risk assessment.Methods:A total of 512 patients with suspected cosmetic adverse reactions were collected from the outpatient clinic of Chongqing Traditional Chinese Medicine Hospital from March 2018 to October 2019, including 14 males and 498 females. A uniform cosmetic adverse reaction report card was filled in, and medical history of patients and related information about the used cosmetics were recorded; 103 patients (3 males and 100 females) were subjected to patch test with their own cosmetics or cosmetic ingredients, and 48- and 72-hour patch test results were combined for comprehensive determination and analysis.Results:Among the 512 cases of suspected cosmetic adverse reactions, contact dermatitis (495 cases, 96.7%) was the most common manifestation. Cosmetic adverse reactions mainly manifested as erythema (501 cases, 97.9%), papules (313, 61.1%), edema (249, 48.6%), and scaling (166, 32.4%) ; main symptoms included itching (480, 93.8%), burning sensation (359, 70.1%), and tense sensation (297, 58.0%). Patch test with cosmetic ingredients showed positive reactions in 71 of 103 cases, and thimerosal was the allergen mostly liable to cause adverse reactions (31 cases, 30.1%), followed by sodium dodecyl sulfate (29 cases, 28.2%), Peru balsam (17 cases, 16.5%), bronopol (12 cases, 11.7%) and triethanoamine (10 cases, 9.7%). The cosmetic allergens were divided into 14 categories, and the top 4 categories with high positive patch test rates were emulsifiers (54 cases, 45.8%), preservatives (47 cases, 39.8%), fragrances (17 cases, 14.4%) and surfactants (10 cases, 8.5%). Positive patch test reactions were observed in 2 males and 69 females, and there was no significant difference in the positive rate between males and females (2/3 vs. 69/100, χ2 = 0.01, P > 0.05) ; there was also no significant difference in the positive rate among the groups aged 18 - 29 years (34%), 30 - 49 years (34%) and 50 - 70 years (32.4%; χ2 = 0.693, P > 0.05) . Conclusions:Contact dermatitis is the most common adverse reaction to cosmetics. Among the diverse allergenic components of cosmetics, thimerosal is the allergen that is mostly liable to cause adverse reactions, followed by sodium dodecyl sulfate, Peru balsam, bronopol and triethanoamine.
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Objective:To screen and evaluate sunscreen cosmetics for sensitive facial skin.Methods:From June to August in 2019, 40 subjects with positive lactic acid sting test were recruited from the staff of Chongqing Traditional Chinese Medicine Hospital, and subjected to human skin closed patch testing with 4 kinds of sunscreen cosmetics for sensitive skin (marked as products Ⅰ, Ⅱ, Ⅲ, Ⅳ respectively) separately. Then, the 40 subjects were equally divided into 2 groups to apply 2 sunscreen products with relatively higher safety (according to the above closed patch testing results) on the face respectively. Erythema, edema and desquamation were evaluated at baseline, 2 and 4 weeks after application of the 2 products, and non-invasive measurement methods were used to detect transepidermal water loss (TEWL) , stratum corneum hydration, skin melanin content and skin sebum content. In additon, the 2 products were applied on the back of the subjects separately, and an ultraviolet solar simulator was used to determine the sun protection factor (SPF, n = 12) and protection factor of UVA (PFA, n = 11) . Measurement data were compared using paired t test and one-way analysis of variance, and nonparametric data were compared using Wilcoxon signed rank test. Results:Patch testing showed that only 1 subject developed a grade 1 reaction to the sunscreen product Ⅲ, no subjects showed positive reactions to the product Ⅳ, and the safety of products Ⅲ and Ⅳ was higher than that of the other 2 products. Subjective safety evaluation revealed that the degree of erythema after 4-week application of products Ⅲ and Ⅳ was significantly lower than that before application (Wilcoxon signed rank test, Z = 4.73, 4.82 respectively, both P < 0.05) . Objective efficacy evaluation revealed that the TEWL, stratum corneum hydration and skin melanin content significantly differed among different time points (baseline, after 2- and 4-week application of products Ⅲ and Ⅳ, all P < 0.05); after 4-week application of products Ⅲ and Ⅳ, the TEWL (30.05 ± 1.47, 30.37 ± 1.28 respectively) and skin melanin content (112.58 ± 7.34, 103.47 ± 5.48 respectively) were significantly lower than those before application (all P < 0.05) , and the stratum corneum hydration (62.35 ± 2.67, 63.72 ± 2.54 respectively) was significantly higher than that before application (both P < 0.05) . At week 4, the skin melanin content was significantly lower in the product Ⅳ group (103.47 ± 5.48) than in the product Ⅲ group (112.58 ± 7.34, t = 8.45, P < 0.05) . The SPF and PFA values of the product Ⅳ (51.8 ± 2.9, 10.1 ± 1.2 respectively) were both significantly higher than those of the product Ⅲ (31.5 ± 2.6, 7.4 ± 0.7, t = 15.34, 24.66, respectively, both P < 0.05) . Conclusion:Comprehensive application of closed patch testing, long-term application test and sun protection index determination can be used to screen and evaluate the safety and efficacy of sunscreen cosmetics for sensitive facial skin.
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Objective:To evaluate the efficacy of acrivastine alone or in combination with loratadine in the treatment of chronic refractory urticaria.Methods:From March 2017 to December 2018, a multicenter, randomized, controlled clinical study was conducted in 4 centers. Patients with chronic refractory urticaria were randomly divided into two groups, i.e., combined treatment group receiving oral acrivastine capsules 8 mg thrice a day plus oral loratadine tablets 10 mg once a day, and acrivastine alone group receiving oral acrivastine capsules 8 mg thrice a day plus a placebo 10 mg once a day. The course of treatment was 4 weeks. Visits were scheduled at baseline and after 1, 2 and 4 weeks of treatment. At the same time, clinical data were collected, and adverse events were recorded. Symptom scores were evaluated based on degree of itching, number and size of wheals, duration of each attack and number of attacks per week, and symptom score reduce index (SSRI) was used to evaluate the efficacy. Repeated measures analysis of variance and chi-square test were used to evaluate the efficacy and safety.Results:Fifty-three patients in the combined treatment group and 59 in the acrivastine alone group were included in the efficacy analysis. Before treatment, there was no significant difference in symptom score or visual analogue score between the two groups. After 2 weeks of treatment, 19 patients were cured and 10 achieved marked improvement in the combined treatment group, with a response rate of 54.72%; 15 were cured and 6 achieved marked improvement in the acrivastine alone group, with a response rate of 35.59%. After 4 weeks of treatment, 23 patients were cured and 9 achieved marked improvement in the combined treatment group, with a response rate of 60.38%; 20 were cured and 2 achieved marked improvement in the acrivastine alone group, with a response rate of 37.29%. After 2 and 4 weeks of treatment, the response rates were significantly higher in the combined treatment group than in the acrivastine alone group ( χ2 = 4.13, 5.96 respectively, both P < 0.05) . The SSRI significantly differed among different follow-up time points, as well as between the 2 groups ( F = 8.62, 4.38 respectively, both P < 0.05) . Multivariate analysis of variance showed that SSRI was significantly higher in the combined treatment group (0.63 ± 0.05, 0.68 ± 0.05, respectively) than in the acrivastine alone group (0.47 ± 0.05, 0.51 ± 0.05, respectively) after 2 and 4 weeks of treatment (both P < 0.05) . Drug-related adverse reactions, including drowsiness, stomach upsets, headache and liver function abnormality, occurred in 7 patients in the combined treatment group, as well as in 3 in the acrivastine alone group. Conclusion:Acrivastine is safe and effective for the treatment of chronic refractory urticaria, and acrivastine combined with loratadine can markedly improve the efficacy.
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Objective To investigate the expression of DKK3 in human cutaneous malignant melanoma cells and tissues,and to evaluate the effect of transfection with DKK3 gene on migration and invasion of a malignant melanoma cell line A375.Methods Western blot analysis was performed to measure the relative expression of DKK3 in human cutaneous melanoma cell lines HM,A375,WM451,SK-MEL-1,Hs-695T,MDA-MB-435s and WM35,as well as pigmented nevus tissues.Real-time fluorescence-based quantitative PCR (RT-PCR) was conducted to determine the mRNA expression of DKK3 in 58 melanoma tissues (including primary melanoma and metastatic melanoma) and 30 pigmented nevus tissues from Chongqing Traditional Chinese Medicine Hospital between August 2014 and June 2017.The pcDNA3.1 (+)-Flag-Vector (control group) and pcDNA3.1 (+)-Flag-DKK3 (transfection group) were transfected into A375 melanoma cells separately.RT-PCR and Western blot analysis were performed to verify the overexpression of DKK3,and to evaluate the effect of DKK3 overexpression on the expression of molecules related to the migration and invasion of melanoma cells.Cell scratch assay,Transwell migration and invasion assay were conducted to assess the effect of DKK3 on the migration and invasion of A375 cells.Statistical analysis was done by a two-sample t-test for comparisons between two groups,one-way analysis of variance (ANOVA) for intergroup comparison,and least significant difference (LSD)-t test for multiple comparisons with the SPSS 13.0 software.Results DKK3 protein was absent or lowly expressed in the human melanoma cell lines,but highly expressed in the pigmented nevus tissues.There were significant differences in the mRNA expression of DKK3 among the primary melanoma tissues (2-ΔΔCt:[0.325 ± 0.150] × 10-3),metastatic melanoma tissues ([0.142 ± 0.210] × 103) and pigmented nevus tissues ([0.634 ±:0.120] × 10-3,F =46.57,P ＜ 0.05).In addition,the mRNA expression of DKK3 was significantly lower in the metastatic melanoma tissues than in the primary melanoma tissues and pigmented nevus tissues (LSD-t =2.48,3.12,both P ＜ 0.05).After transfection with DKK3,cell scratch assay showed that the migration rate was significantly lower in the transfection group (22.11％ ± 5.11％) than in the control group (54.36％ ± 23.22％,t =2.36,P ＜ 0.001).Transwell migration and invasion assay revealed that the number of A375 cells crossing the Transwell chamber was significantly lower in the transfection group (265 ± 33,76 ± 18 respectively) than in the control group (429 ± 41,135 ± 21 respectively;t =1.24,1.35 respectively,both P ＜ 0.001).After overexpression of DKK3 in the A375 cells in the transfection group,the mRNA and protein expression of E-cadherin were up-regulated,while the mRNA and protein expression of N-cadherin,vimentin,matrix metalloproteinase 2 (MMP2),MMP7 and MMP11 were down-regulated compared with the control group.Conclusions The expression of DKK3 is down-regulated in the melanoma cell lines and tissues,and the migration and invasion of A375 cells are markedly inhibited by overexpression of DKK3.DKK3 may be a target for inhibiting the metastasis of cutaneous malignant melanoma.
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A 28-year-old female patient presented with indurated erythema and nodules on the right lower limb for 2 years,with mild itching and pain.Skin examination showed a well-circumscribed irregular dark red patch measuring about 10 cm × 5 cm in size on the extensor aspect of the right thigh.On the patch,there were scattered or densely distributed mung bean-to soybean-sized quasi-circular violaceous nodules with a smooth surface,which were hard on palpation.Subcutaneous nodules with medium hardness were found on palpation,and hyperpigmentation was observed on the surface of some nodules.Local skin temperature was increased,with tenderness on palpation.Histopathologically,mononuclear cells showed nodular or sheet-like distribution in the middle and upper dermis,some of which had pale-staining cytoplasm.Moreover,plenty of plasma cells were observed.Immunohistochemistry revealed that histiocytes were stained strongly positive for S100.The number of IgG4-positive plasma cells increased obviously,and was more than 50 per high-power field (× 200).The proportion of IgG4-positive plasma cells in IgG-positive plasma cells was 45％.Finally,the patient was diagnosed with cutaneous Rosai-Dorfman disease with increased IgG4-positive plasma cells.
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Objective To evaluate the clinical effect of fire needle in treating vitiligo and the characteristics of vitiligo image by the confocal laser scanning microscope(CLSM).Methods The randomized self-controlled experiment design was adopted.Each patient selected two symmetric or adjacent white patches and randomly received the fire needle treatment or tacrolimus treatment. The duration of treatment was 3 months.The CLSM images of white patches were recorded before treatment and after 3,6 times of fire needle treatment.Results Among 41 cases of stable stage vitiligo,The effective rates of the fire needle group and tacrolimus group were 82.9% and 78.0% respectively,and the difference was not statistically significant(P>0.05).After fire needle treat-ment,dentritic melanin cells appeared,the pigment granules gradually appeared around the basal layer and corpora papillare,and formed the pigment ring.Conclusion Fire needle and tacrolimus have the similar effect in treating vitiligo,moreover CLSM can be used as the non-invasive,objective and reliable detection means of the recovery of vitiligo melanocyte.
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Objective To detect the expression levels of peripheral blood CXCL10 and its receptor CXCR3 and T cell subsets in of the patients with advanced vitiligo and the influence of compound Chinese medicine on it.Methods Flow cytometry was used to detect the cellular proportions of peripheral blood T cell subsets,ELISA was employed to quantify serum CXCL10 and CXCR3 expression levels before and after treatment.Results After 1 month of taking Chinese medicine,the proportions of CD3+ CD4+ cells and CD3+ CD8+ cells were increased compared before treatment(P＜0.05).The expression level of peripheral serum CXCL10 before treatment was significantly increased compare with the healthy control group(P＜0.01),and the CXCL10 level after treatment was decreased significantly compared with that before treatment(P＜0.05).The expression level of peripheral serum CXCR3 was significantly increased compared with the healthy control group(P＜0.05),while which after treatment was still significantly higher than that in the healthy control group(P＜0.05).Conclusion CXCL10,CXCR3 and T cell subsets proportion may be involved in the pathogenesis of vitiligo.The compound Chinese medicine used in this study plays the curative effect possibly by regulating T cell subsets and expression levels of CXCL10 and CXCR3.
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Objective To investigate the expression of Dickkopf-3 (DKK3) in human malignant melanoma cell lines and tissues,and to evaluate effects of DKK3 on the proliferation and apoptosis of malignant melanoma cell line A375.Methods Reverse transcription PCR (RT-PCR) and real-time fluorescence-based quantitative PCR (qRT-PCR) were performed to measure the mRNA expression of DKK3 in human malignant melanoma cell lines HM,A375,WM451,WM35,SK-MEL-1,Hs-695T and MDA-MB-435s,as well as in 38 primary melanoma tissues,4 metastatic melanoma tissues and 20 pigmented nevus tissues.Cultured malignant melanoma A375 cells were divided into 2 groups to be transfected with pcDNA3.1 (+)-Flag-DKK3 (experiment group) and pcDNA3.1 (+)-Flag-Vector (control group) respectively.The overexpression of DKK3 was verified by RT-PCR.Cell counting kit-8 (CCK8) assay and plate colony formation assay were performed to evaluate the proliferative activity of A375 cells,flow cytometry was conducted to detect apoptosis of A375 cells,and Western blot analysis was performed to determine the expression of cell cycle-and cell apoptosis-related proteins.Results The mRNA expression of DKK3 was downregulated in WM35 cells,absent in HM cells,A375 cells,WM451 cells,SK-MEL-1 cells and Hs-695T cells,but upregulated in MDA-MB-435s cells.Compared with pigmented nevus tissues,the mRNA expression of DKK3 was significantly decreased in malignant melanoma tissues (P ＜ 0.001).Compared with the control group (100％),cell colony formation was markedly suppressed in the experiment group (23.22％ ± 3.55％),and the proliferative activity of A375 cells was also significantly inhibited in the experiment group 24,48,72 hours after the transfection (all P ＜ 0.05).Flow cytometry showed that compared with the control group,A375 cells were significantly arrested in G1 phase (48.68％ ± 3.92％ vs.25.38％ + 2.92％,P ＜ 0.001),and the apoptosis rate of A375 cells was significantly increased in the experiment group (P ＜ 0.001).Compared with the control group,the experiment group showed significantly higher expression of p21,Bax,cleaved-parp and cleaved-casp3,but significantly lower expression of cyclin D1 and Bcl2 (all P ＜ 0.001).Conclusion DKK3 expression is downregulated in human malignant melanoma tissues,so it may serve as a potential tumor suppressor gene involved in the development of cutaneous malignant melanoma.
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Objective To research the change rule of peripheral serum Th1/Th2/Th17 balance and relevant cytokines in atopic dermatitis(AD) and to further study its immunologic mechanism .Methods The levels of peripheral serum IFN‐γ,IL‐4 ,IL‐17 ,IL‐21 and IL‐23 in the patients with AD were detected by the flow immunofluorescence technology and the detection results were performed the statistical analysis ,thus the change rule of Th1/Th2/Th17 balance in AD was analyzed .Results The IL‐4 level in peripheral serum of the patients with AD was increased compared with the healthy control group (P<0 .05);the IFN‐γ/IL‐4 and IFN‐γ/IL‐17 in the AD group were significantly decreased compared with the control group (P<0 .01);IL‐17 was positively corre‐lation with SCORAD scores of AD severity (P<0 .05) .Conclusion Th1/Th2 and Th1/Th17 are decreased in the AD course .Th1/Th2/Th17 balance drift may be the important mechanism of AD pathogenesis .
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Objective To clarify the cellular localization of triptolide and to explore its in-cell action sites.Methods 4-(Bro-momethyl)-7-methoxycoumarin was employed to label triptolide,then labelled triptolide was incubated with human hepatoma carci-noma cells.Subsequently,incubated cells were subjected to stain with fluorescent dye DiI or PI,which were specific to cytoplasmic membrane system and nucleus,respectively.Results Compared with the non-triptolide control,coumarin labelled triptolide shown a light blue fluorescence under UV excitation;Co-localization with DiI showed that triptolide exist in cytoplasm and(or)on cell mem-brane;Co-localization with PI showed that triptolide located in cell nucleus.Moreover,microscopic observation indicated that the fluorescence intensity in nucleus was denser than that in cytoplasm.Conclusion The presnt study demonstrate that triptolide main-ly act in nucleus,followed by acting in cytoplasm and(or)on cell membrane.
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<p><b>OBJECTIVE</b>To observe the effect of ultrasound stimulation at the acupoint Guanyuan (CV 4) on follicular development in menopausal rats.</p><p><b>METHODS</b>Menopausal female SD rats were selected by vaginal smear examinations. The rats were subjected to ultrasound stimulation at the acupoint Guanyuan with the output power of 0.1 W, working frequency of 9 MHz, and focal length of 4.5-5 mm. Electrochemiluminescence immunoassay was used to detect the serum estrogen levels of the menopausal rats. The changes in the ovarian tissue histology and the follicle number were observed.</p><p><b>RESULTS</b>Compared with the control group, a 10-day ultrasound stimulation for 10 and 5 min daily at Guanyuan significantly increased the serum estrogen levels and the numbers of primary and secondary follicles (P<0.05) and reduced the number of atretic follicles in the menopausal rats (P<0.05).</p><p><b>CONCLUSION</b>Ultrasound stimulation at the acupoint Guanyuan can increase the estrogen secretion function and promote the development of follicles in menopausal rats.</p>
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Animals , Female , Rats , Acupuncture Points , Estrogens , Blood , Menopause , Ovarian Follicle , Rats, Sprague-Dawley , SonicationABSTRACT
Objective To investigate the influence of CD4+CD25+ regulatory T cells (Tregs) on the disease progression in MRL/lps mice. Methods Tregs were separated by using magnetic beads from splenic cells of MRL/lps mice and BALB/c mice, and concentrated. Twenty-four MRLAps mice were equally divided into 3 groups, test group 1 injected with Tregs from MRL/lps mice, test group 2 injected with Tregs from BALB/c mice, and control group injected with physiological sodium chloride solution. Three weeks later, the levels of urine protein as well as serum anti-dsDNA antibody were determined; subsequently, the mice were sacrificed followed by histopathological and immunopathological examination of renal tissue. Results A significant decline was observed in the test group 1 compared with the test group 2 and control group in the urine protein score (10.63 ± 4.17 vs. 20.00 ± 5.35 and 18.75 ± 8.34, both P 0.05). There was no statistical difference between the test group 2 and control group in terms of any of the above parameters (all P > 0.05). Conclusions Injection of Tregs from homologous mice could significantly down-regulate proteinuria degree, serum anti-dsDNA antibody level, glomerular sclerosis index and IgG immune complex level in renal tissue, and thereby decelerate the progression of renal impairment in MRL/lps mice.
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Objective To investigate the therapeutic efficacy of HPV16 E7 peptide pulsed dendritic cells (DCs) in recurrent condyloma acuminate (CA). Methods A total of 32 cases of recurrent CA (more than 3 times) were divided into the treatment group and the control group. Patients (11 cases, HPV16 +and HLA A2 +) in the treatment group received treatment with DCs, while the other 21 cases in the control group were treated with interferon. A follow up of 6 months was conducted in all patients. The pathological lesions, the peripheral T cell subpopulations of the patients, and the therapeutic efficacy before and after treatment were observed. Results The size of the lesions became smaller or disappeared in the treatment group. The infiltrated lymphocytes increased, but the koilocytotic cells decreased in the lesions. No significant change in the peripheral T cell subpopulations was found before and after therapy. The recurrence rates in the treatment group and the control group were 18.2% and 61.9%, respectively. Conclusion The therapy by E7 peptide pulsed dendritic cells can improve the local immune status in the skin and reduce the recurrence rate significantly in patients with recurrent CA.
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Objective To study the effects of recombinant chimeric toxin Dsg3EC_(1-2)PE40 on T and B cells from PV patients. Methods The recombinant protein Dsg3EC_(1-2)PE40 was expressed on BL21TrxB (DE3) cells, then identified and purified. ELISPOT assay was used to detect and quantitate autoantibody-producing B cells in different concentrations of recombinant chimeric toxin, and MTT assay and ~3H-TdR assay to observe the metabolism and proliferation of T cells from PV patients in vitro. Results The purity of expressed protein Dsg3EC_(1-2)PE40 was up to 80%. The number of anti-Dsg3 antibody-producing B cells in PBMC from PV patients decreased by 40% with treatment of Dsg3EC_(1-2)PE40, which was significantly lower (P
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Objective To investigate the role of IL-2,IFN-? and IL-10 in pemphigus acantholysis. Methods Acantholysis was observed histopathologically in the skin organ culture model of pemphigus after interacting with different concentrations of IL-2, IFN-? and IL-10 for 24 h?48 h and 72 h. Results The acantholysis was promoted by IL-2 and IFN-?, and the severity of acantholysis was related to the concentrations of IL-2 and IFN-?. The effect of IFN-? was weaker than that of IL-2. IL-10 could inhibit the acantholytic effect of IFN-? significantly, and inhibit the acantholytic effect of IL-2 when its concentration was higher than 100 pg/mL. Conclusions Th1 cytokines can promote acantholysis induced by antibody of pemphigus (Pab) while Th2 cytokines can inhibit the acantholysis induced by Pab, and the effect of Th1 cytokines. Th2 lymphocytes may play an important role in the pathogenesis of pemphigus.
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Objective To analyze the clinicopathological features of angiolymphoid hyperplasia with eosinophilia (ALHE). Methods The pathological specimens of 7 cases of ALHE collected in our department from 1950 to 1999 were sectioned, stained and observed. Results There were 3 pathological characteristics in ALHE: ①massive hyperplasia of capillaries in the dermis; ②the endothelial cells proliferated and swelled, projecting into vascular cavity like tombstones; ③mixed infiltration of lymphocytes and eosinocytes in the vessels. Conclusion ALHE is a disease with local benign proliferated vessels, whose etiology and pathogenesis is still unknown. It is necessary to grasp the pathological changes of ALHE to distinguish it from other diseases.