简介：Objective:Toinvestigatetherelationshipbetweenapoptosisandproliferatingcellnuclearantigen(PCNA)expressionofkeratinocytesinCondylomataacuminata(CA).Methods:PCNAexpressionwasobservedbyimmunohistochemistrytechnique(ABCmethod)in51CAspecimensand18normalspecimensofforeskinorvaginalmucosae.55specimens(40intheCAgroupand15inthecontrolgroup)wererandomlysampledforinsitulabelingofapoptoticcellsusingtheTUNELmethod.Results:PositiveexpressionofPCNAinCAandcontrolgroupswere90.2%and77.8%,respectively,andtheproliferationindexinCAgroupwassignificantlyhigherthanthatinthecontrolgroup(P<0.001).Thepositiverateofapoptosiswas42.5%intheCAgroupand53.3%inthecontrolgroup,andtherewerenosignificantdifferencesintheapoptoticindexandapoptosis-proliferationratiobetweentwogroups(P>0.05).Theproliferationindexshowedasignificantnegativecorrelationwiththeapoptosis-proliferationratio(r=-0.62,P=0.01)intheCAgroup.Conclusion:ItissuggestedthattheproliferativeappearanceofCAcouldbeduetotheimbalancebetweencellgrowthandcelldeathwhichiscausedbymoreproliferationandlessapoptosisinkeratinocytes.

简介：AbstractKeratinocytes, located in the outer part of human skin, are the main epidermal cell type and play an essential role in skins defense against infection. Besides creating a physical barrier between the environment and the internal body, keratinocytes exert powerful immune function in anti-pathogen infection in the skin. At the recognition stage, pattern recognition receptors (PRRs) expressed by keratinocytes sense pathogen-associated molecular patterns (PAMPs) existing in pathogens. Toll like receptors (TLRs) are the most important PRRs in keratinocytes. Other PRRs such as dectin-1 and nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) are also found to participate in this process. Activated PRRs enhance the secretion of cytokines, chemokines and the production of antimicrobial peptides (AMPs). Proinflammatory cytokines tumor necrosis factor-α, interleukin (IL)-1α, IL-6, IL-1β and IL-18, chemokines (C-X-Cmotif) ligand (CXCL) 1, CXCL2, CCL20, CCL2 and IL-8, AMPs human β-defensin (HBD) 2, HBD3 and LL37 are the main molecules expressed in this procedure. Thymic stromal lymphopoietin (TSLP), IL-36γ, IL-17 family member IL- 17C and anti-inflammatory cytokine IL-10 can also be secreted. Some molecules produced by keratinocytes such as ribonuclease 5 and 7, S100 proteins own antimicrobial properties. Keratinocytes defense responses can be regulated by internal and external factors. This review summarizes recent advances on the innate immune function of keratinocytes against infection, promoting the finding of a new direction for avoiding severe skin infection as well as the potential treatment of keratinocyte-associated inflammatory dermatosis.

简介：Objective:Toinvestigatetheproliferativeeffectofkeratinocytegrowthfactor(KGF-2)onhumanadultkeratinocytes.Methods:Thestandardmediumwaskeratinocytegrowthmediumwithoutbovinepituitaryextract(BPE),hydrocortisoneorepidermalgrowthfactor(EGF).Keratinocytesfroma48-year-oldsubjectwereculturedandseededondisheswithstandardmediumofEGFincelldensityof2×104/32mm2.After24hours,themediumwasreplacedbythestandardmediumwith0,4,16,125and500ng/mlKGF-2,respectively.ThestandardmediumwithEGFwasusedasthepositivecontrolandthestandardmediumwithoutEGForKGF-2wasusedasthenegativecontrols.Thegrowthofkeratinocyteswasmonitoredby3-(4,5-dimethythiazol-2-yl)-2,5dipheyltetrazoliumbromide(MTT)assayandbyphotographsondays3,5and7,respectively.Results:KGF-2inconcentrationsof4-500ng/mlshowedasignificantproliferativeeffectondays5and7ascomparedwiththatofthenegativecontrols(P<0.01).Onday3thecellswereproliferatedto1.5-2.5-fold,onday5to3-5-foldandonday7to3-12-foldinKGF-2mediumasthatofthenegativecontrols.TheoptimalresponseoccurredwhentheconcentrationofKGF-2was125ng/mlonday7.CellproliferationwasalsoconsistentlyhigherinallKGF-2concentrationsascomparedwiththatofthepositivecontrols.Conclusions:KGF-2hassignificanteffectsontheproliferationofadultkeratinocytes,whicharemoreeffectivethanthatofEGF.ThisstudysupportsKGF-2canimprovethehealingofchronicwoundsinadultsinclinic.

简介：微数组技术是的模仿的微严肃(SMG)生物反应器和DNA识别“玩的空间基因”的强大的工具在对微严肃的细胞的反应给角色调音。我们使用了这些生物工学工具由用生物反应器在正常严肃为15，50，和60d由恢复culturing跟随了的高方面比率容器为3，4，9，和10d暴露房间到SMG在人的表皮的keratinocytes上调查SMG和post-SMG恢复效果。作为结果，我们鉴别162差别表示了基因，其32是“最一致地在路线试验的时间被影响的中心基因”。十一中心基因从综合压力反应小径并且是并列地下面调整的。七个中心基因，都是金属硫蛋白MT-I和MT-IIisoforms，是并列地起来调整的。另外，HLA-G，在细胞的有免疫力的反应抑制的关键基因，被发现在恢复阶段期间显著地起来调整。总的来说，超过80%差别从更短的暴露表示了基因(/=9d)暴露，超过50d被需要恢复到更短的暴露的影响水平。数据显示那更短的SMG暴露持续时间将从微严肃效果导致更快、更完全的恢复。

简介：AbstractObjective:Acne inversa is a multifactorial chronic debilitating disease. Genetic factors are involved in 40% of patients, especially the nicastrin (NCSTN) gene. However, the role of the mutated NCSTN gene in the pathogenesis of acne inversa remains unclear. Retinoic acid is recommends to treat moderate to severe acne inversa, therefor we conduct this in vitro research to study the association between NCSTN gene mutation and the retinoic acid signaling pathway in human immortalized skin keratinocyte (HaCaT) cells.Methods:HaCaT cells were infected with a lentivirus-mediated short hairpin RNA (shRNA) expression plasmid specifically targeting the NCSTN gene. Real-time polymerase chain reaction (PCR) and Western blotting were used to detect the interference efficiency of NCSTN. RNA sequencing was used to detect differential genes in the NSCTN-deficient HaCaT cells. Based on bioinformatics analysis and clinical treatment data, the retinoic acid signal pathway was selected for screening. Quantitative PCR was used to verify the changes in the expressions of retinoic acid signaling pathway-related receptors and molecules in the HaCaT cell line after NCSTN silencing. The Student t test and one-way analysis of variance were used to evaluate intergroup differences.Results:Sequencing showed that the NCSTN-shRNA lentiviral recombinant expression plasmid was successfully constructed. After lentivirus infection of HaCaT cells, real-time PCR results showed significantly reduced NCSTN mRNA expression in the interference group compared with the negative control group, and the interference efficiency was 75.0%. Western blotting showed that the inhibition rate of NCSTN protein expression in the shRNA group was 71.7%. RNA sequencing revealed significant differential expression of some genes, and changes in signaling pathways. Compared with the control group, the group with the silenced NCSTN showed significantly decreased expression of retinoic acid receptors (RARα: F=23.482, RARβ: F=603.241, RXRα: F=69.689, and RARRES1: F=167.482, and all P < 0.001), and peroxisome proliferator-activated receptor γ (F=8.138, P < 0.01).Conclusion:Defective function of the NCSTN gene leads to an impaired retinoic acid signaling pathway in HaCaT cells, which suggests that the retinoic acid signaling pathway may play a role on the onset of acne inversa caused by NCSTN gene mutation.

简介：AbstractObjective:The plant polyphenol resveratrol (3,4′,5-trihydroxystilbene) (RSV) has been proposed for use because of its protective effect on ultraviolet (UV)-induced skin disorders. In UVB-induced skin damage, cell autophagy and apoptosis have been approved to prevent the damage and to contribute to the cytoprotective role of RSV; however, the detailed mechanism remains unknown. So, we conducted this study to investigate the cytoprotective effects of RSV on UVB-irradiated human epidermal keratinocytes (HEKs) and its undergoing mechanisms.Methods:Secretion of thirty-six inflammatory cytokines of HEKs induced by 50 mJ/cm2 UVB at 0, 12, 24, and 48 hours were detected by a human cytokine assay and the interleukin (IL)-8 protein level in the culture media were determined by ELISA. Next, HEKs were treated with or without 100 μmol/L RSV in the presence or absence of 50 mJ/cm2 UVB, and activator protein 1 and NF-κB-related proteins were measured by Western blot. Furthermore, cells exposed to UVB radiation were treated with apoptosis activators procaspase-activating compound 1 (PAC-1), apoptosis activator 2 (AA2) or RSV to investigate the effect of RSV on the percentage of apoptotic cells by flow cytometry. Then cells were treated with autophagy inhibitors LY294002, 3-methyladenine (3-MA) or RSV in the presence of UVB and chloroquine (CQ) to investigate the effect of RSV on autophagy through detecting microtubule-associated protein 1 light chain 3 (LC3) expression by western blot. Finally, the effect of LY294002, 3-MA, ATG5 siRNA, PAC-1, and AA2 on RSV-mediated reduction of IL-8 expression was determined by ELISA assay.Results:RSV treatment decreased the secretion of IL-8 (UVB vs. UVB+ RSV: [1454.05 pg/mL ± 52.95 pg/mL] vs. [553.68 pg/mL ± 206.03 pg/mL], P < 0.001), and downregulated the protein level of c-Fos in UVB-irradiated HEKs (UVB vs. UVB+ RSV: [0.103 ± 0.009] vs. [0.048 ± 0.015], P < 0.01). In UVB-irradiated HEKs, the result of western blot showed that LY294002 and 3-MA inhibited RSV-induced LC3 II accumulation (UVB + CQ + RSV vs. UVB + CQ + 3-MA+ RSV vs. UVB + CQ + LY294002+ RSV: [1.15 ± 0.03] vs. [0.77 ± 0.13] vs. [0.67 ± 0.13], P < 0.01), and the result of flow cytometry showed that PAC-1 and AA2 prevented RSV from reducing cell apoptosis (UVB+ RSV vs. UVB+ PAC-1 + RSV vs. UVB+ AA2+ RSV: [19.56% ± 0.62%] vs. [94.33% ± 0.15%] vs. [94.97% ± 1.91%], P < 0.001). The data of ELSA assay showed that LY294002, 3-MA, and ATG5 siRNA reversed the RSV-mediated inhibition of IL-8 protein secretion by UVB-irradiated HEKs (UVB + LY294002 vs. UVB + LY294002 + RSV: [3283.00 pg/mL ± 444.05 pg/mL] vs. [1608.58 pg/mL ± 128.42 pg/mL], P < 0.05; UVB + 3-MA vs. UVB+ 3-MA+ RSV: [2941.88 pg/mL ± 103.80 pg/mL] vs. [1867.51 pg/mL ± 153.84 pg/mL], P < 0.01; UVB+ siATG5 vs. UVB+ siATG5+ RSV: [2530.11 pg/mL ± 685.34 pg/mL] vs. [3011.42 pg/mL ± 435.69 pg/mL], P > 0.05), whereas neither PAC-1 nor AA2 exerted similar effects.Conclusion:RSV may regulate autophagic flux to inhibit IL-8 expression in UVB-challenged keratinocytes.

简介：AbstractObjective:This study was performed to investigate the effects of honokiol on the activation of transient receptor potential channel V1 (TRPV1) and the secretion of thymic stromal lymphopoietin (TSLP) in a human benign epidermal keratinocyte line (HaCaT).Methods:HaCaT keratinocytes were cultivated and divided into six groups: capsaicin-induced model control group, capsazepine control group, solvent control group, and three honokiol treatment groups (7.81, 15.63, and 31.25 mg/L of honokiol). The effect of honokiol on calcium (Ca2+) influx was measured by a Ca2+ fluorescence imaging system. The fluorescence intensity (F) of cells was measured. The rate of change in F (ΔF/F0) was calculated, and the ΔF/F0-time curve was constructed. HaCaT keratinocytes were stimulated with polyinosinic:polycytidylic acid, recombinant human tumor necrosis factor α, and recombinant human interleukin 4. Different concentrations of honokiol (15.63, 7.81, and 3.91 mg/L) were added to the cells in the respective honokiol groups; 20 mg/L of dexamethasone or 0.5% dimethyl sulfoxide was added to the cells in the positive control group or solvent control group. The TSLP concentration in the HaCaT keratinocytes of each group was detected by enzyme-linked immunosorbent assay. Statistical analysis was performed by one-way analysis of variance and Dunnett t test.Results:The capsaicin-induced Ca2+ fluorescence intensity in HaCaT keratinocytes was significantly inhibited in the 31.25 mg/L honokiol group; ΔF/F0 at 45 second was 0.76 in the model control group and 0 in the 31.25 mg/L honokiol group. The TSLP level in the 15.63 and 7.81 mg/L honokiol groups was lower than that in the solvent control group (t= 7.382, P= 0.003, and t= 2.766, P= 0.023, respectively), while the TSLP level in the 3.91 mg/L honokiol group was not significantly different from that in the solvent control group (t= 1.872, P= 0.124).Conclusions:Honokiol inhibited the Ca2+ influx induced by capsaicin (TRPV1 agonist) in HaCaT keratinocytes. Honokiol has an inhibitory effect on TSLP secretion in HaCaT keratinocytes.

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简介：Objective:ToinvestigatetheexpressionofHPV16mRNAinnormalhumankeratinocytestransfectedwithpSV2-neo/16.Firsthumankeratinocyteswereculturedintheserum-freemediumM154.Second,theplasmidpSV2-neo/16wastransfectedintothehumankeratinocytesusingatransfectingreagent.Third,RT-PCRandSouthernBlottingwereusedtodetecttheexpressionofHPV16mRNAandDNAinthetransfectedkeratinocytes,respectively.Results:TheexpressionofHPV16mRNAwassuccessfullyamplifiedandan110bpwasditectedbyRT-PCR.A7.9kbfragmentwasconfirmedinthetransfectedkeratinocytesbySouthernBlotanalysis.Conclusion:HPV16mRNAandDNAweresuccessfullydetectedinthehumankeratinocytes.