角质形成细胞与成纤维细胞对中波紫外线照射应激能力的比较研究

目的:观察两种皮肤细胞即原代角质形成细胞及成纤维细胞对中波紫外线照射的反应差异。方法:采用不同剂量中波紫外线(UVB)照射上述两种细胞,评估UVB对细胞作用的时间及剂量效应,以倒置光学显微镜观察细胞受损程度,MTT法检测细胞活性。结果:UVB照射后,细胞损伤程度均随照射剂量加大而加重。另经24、48、72小时孵育后,分别对两种细胞进行不同时间段的细胞活性(MTT)比值比较(72/24小时,72/48小时),发现细胞活性比依次递减:原代角质形成细胞(1.16和1.63),成纤维细胞(1.05和1.09)。MTT结果显示低剂量紫外线照射时,细胞损伤恢复可发生在照射后48小时;高剂量紫外线照射后,细胞的损伤程度均随孵育时间延长而加重。结论:原代角质形成细胞抵抗紫外线辐射损伤的能力较强,而成纤维细胞相对较易受损。

复合多糖抑制紫外线诱导皮肤细胞的光老化

背景:皮肤光老化除受年龄因素影响外,与日光照射亦有直接关系,由日光照射引起的皮肤老化称为光老化。近年来,越来越多的研究发现多糖类物质对皮肤光老化具有一定的抑制作用,主要作用机制可能与其具有抗氧化、保湿、抗辐射等作用相关。目的:明确复合多糖抑制皮肤细胞光老化的作用是否优于单一多糖。方法:利用紫外灯照射诱导人皮肤成纤维细胞(HSF)、人永生化角质形成细胞(HaCaT)(均购自中国科学院上海生科院细胞资源中心)出现老化损伤。在此基础上,采用MTT法和生化试剂盒分别检测相同浓度下4种单一多糖(铁皮石斛多糖、褐藻多糖、灵芝多糖、竹荪多糖)和4种多糖混合物(即复合多糖)对2种光老化表皮细胞存活率及超氧化物歧化酶、丙二醛和羟脯氨酸水平的影响。结果与结论:①与模型组相比,铁皮石斛多糖、褐藻多糖、灵芝多糖、竹荪多糖在50mg/L的质量浓度下,对光老化人皮肤成纤维细胞模型及人永生化角质形成细胞模型的抑制效果均不显著,仅有个别生化指标改善程度显著(P<0.01或P<0.05);4种单一多糖对光老化细胞模型的保护作用相近;②与模型组比较,同样在多糖质量浓度为50mg/L时,复合多糖能够显著提高2种光老化细胞模型的存活率(P<0.01)、超氧化物歧化酶及羟脯氨酸水平(P<0.01或P<0.05),降低丙二醛水平(P<0.01或P<0.05);③结果表明,在对抗皮肤成角质细胞和成纤维细胞光老化实验中,4种单一多糖抗皮肤细胞光老化的效果不显著,而其混合物复合多糖(铁皮石斛多糖+褐藻多糖+灵芝多糖+竹荪多糖)抗皮肤细胞光老化的作用显著,提示复合多糖抑制皮肤细胞光老化的作用优于单一多糖。

Skin care efficacy study of recombinant humanized collagen based on in vitro level

Studying the skin care efficacy of recombinant humanized collagen based on in vitro level.The stability of the recombinant humanized collagen was first analyzed by treating at different temperatures,then its skincare efficacy based on in vitro level was evaluated by detecting the inhibition rate of elastase,the inhibition rate of collagenase,the protein content of type I collagen in human fibroblasts,the inhibition of reactive oxygen species(ROS)with human keratinocytes,and the effects of the recombinant humanized collagen on the expression of hyaluronic acid(HA),filaggrin(FLG)and transglutaminase 1(TGM1)in keratinocytes.The results showed that recombinant humanized collagen was able to maintain stability at temperatures below 70℃.With regard to its skincare efficacy,recombinant humanized collagen could inhibit elastase and collagenase activities and promote the increase of type I collagen content in human fibroblasts.It also showed good inhibition of ROS in keratinocytes in vitro and could increase the expression of HA,FLG,and TGM1 in keratinocytes.In short,the recombinant humanized collagen exhibited a favourable skin care effect in vitro level.This study proved that it has potential firming,anti-wrinkle,moisturizing,and repairing efficacy,and is a valuable cosmetic raw material.

Antagomir Dependent MicroRNA-205 Reduction Enhances Adhesion Ability of Human Corneal Epithelial Keratinocytes

Objective To investigate the effect of microRNA-205 reduction by antagomirs on adhesion ability of normal human corneal epithelial keratinocytes(NHCEKs).Methods Antagomir-205,complementary and inhibitory to microRNA-205,was used to suppress endogenous microRNA-205 in NHCEKs.The adhesion ability of treated NHCEKs was then assessed by cell adhesion assay.Immunoblot and immunohistochemistry were conducted to determine the level of two focal adhesion-related proteins,focal adhesion kinase(FAK) and paxillin(Pax).Phalloidin staining was performed to measure the level of filamentous actin in antagomir-treated NHCEKs.Results Antagomir-205 markedly reduced the level of microRNA-205 in NHCEKs and significantly enhanced adhesion ability of NHCEKs(P<0.01).Further protein analysis validated that inhibition of mi-croRNA-205 increased the number of phosphorylated FAK and phosphorylated Pax,and decreased filamen-tous actin.Conclusion Our findings suggest that microRNA-205 has down-regulating effect on cell motility in NHCEKs.

CD13/APN expression in peripheral blood lymphocytes and skin lesions in patients with advanced psoriasis vulgaris

Objective： To observe the expression of CD13/APN in peripheral blood lymphocytes and skin lesions of patients with advanced psoriasis vulgaris, and discuss its effect on the pathogenesis of psoriasis. Methods： CD 13 expression in peripheral blood lymphocytes and skin lesions was detected by flow cytometry and imrnunohistochemical technique, respectively. Results were compared with those of healthy controls. Results： CD 13 expression was significantly higher in peripheral blood lymphocytes of patients with advanced psoriasis vulgaris than in that of healthy controls, and in skin lesions than in healthy skin tissues. The expression was mainly in the suprabasal layers of skin lesions, and positively correlated to PASI （R 0.78029）. Conclusion： The significantly higher expression of CD13 in peripheral blood lymphocytes and skin lesions of the patients with advanced psoriasis vulgaris probably is related to immunological abnormality, blood vessel abnormality and proliferation of keratinocyte in the pathogenic course of psoriasis. It may be a novel and effective way to treat psoriasis with specific CD13 inhibitors.

Some conifers from Aptian in Yanbian area of Jilin, Northeast China

Some well-preserved conifer leafy shoots and female cones from the Lower Cretaceous Changcai For- mation near Fudong of Helong, eastern Jilin, Northeast China are studied. Based on gross morphological and cuticular study, a new species, Elatides helongensis Sun et Zhao （ sp. nov. ） is described systematically. The new species is characterized by persistent, linear or slightly falcate leaves with obtusely acute apex, attached helically on the shoots. Female cones of the new species are terminal and oval, composed of persistent helically- arranged rhomboidal scales and erect seeds. One erect seed is growing on each scale. Leaf cuticles are hyposto- matic. Monocylic stomata are ellipse, composed of 2 sunken guard ceils and 4-8 subsidiary cells. Moreover, the cuticles of a young female cone of Pityostrobus yingchengensis Yang are described for the first time.

PI3-kinase-dependent activation of apoptotic machinery occurs on commitment of epidermal keratinocytes to terminal differentiation

We have investigated the earliest events in commitment of human epidermal keratinocytes to terminal differentiation. Phosphorylated Akt and caspase activation were detected in cells exiting the basal layer of the epidermis. Activation of Akt by retroviral transduction of primary cultures of human keratinocytes resulted in an increase in abortive clones founded by transit amplifying cells, while inhibition of the upstream kinase, PI3-kinase, inhibited suspension-induced terminal differentiation. Caspase inhibition also blocked differentiation, the primary mediator being caspase 8. Caspase activation was initiated by 2 h in suspension, preceding the onset of expression of the termi- nal differentiation marker involucrin by several hours. Incubation of suspended cells with fibronectin or inhibition of PI3-kinase prevented caspase induction. At 2 h in suspension, keratinocytes that had become committed to terminal differentiation had increased side scatter, were 7-aminoactinomycin D （7-AAD） positive and annexin V negative; they exhibited loss of mitochondrial membrane potential and increased cardiolipin oxidation, but with no increase in reac- tive oxygen species. These properties indicate that the onset of terminal differentiation, while regulated by PI3-kinase and caspases, is not a classical apoptotic process.

Curcumin down-regulates PCNA,cyclin D1 and Bcl-X_L expression in human keratinocyte cell lines

Objective：To evaluate the effects of curcumin on regulating the proliferation,cell cycle distribution,apoptosis and relevant mechanisms in keratinocyte cell lines.Methods：The human immortalized human keratinocyte lines（HaCaT cells） were treated with different doses of curcumin.The effects of curcumin on cell viability were measured by MTT assay,and the cell cycle distribution and apoptosis determined by flow cytometry.The mRNA expression changes of proliferating cell nuclear antigen（PCNA）,cyclin D1 and Bcl-xL were from real-time PCR analysis and the protein levels were detected by Western blotting.Results：Data obtained in the study showed that curcumin could cause significantly inhibitory effect on proliferation in HaCaT cells in a time- and dose-dependent manner.Cell arrest at G1/S phase and significant apoptosis were observed after being treated with curcumin for 24 h.In association with these,the expression of PCNA,cyclin D1 and Bcl-xL were decreased both at mRNA and protein levels for the same treatment.Conclusion：Curcumin can inhibit proliferation,induce cell arrest at G1/S phase and cause apoptosis in HaCaT cells.The decreased expression of PCNA,cyclin D1 and Bcl-xL induced by curcumin contributes to the above effects in vitro.

Effect of keratinocyte growth factor-2 on proliferation of human adult keratino cytes

To investigate the proliferative ef fect of keratinocyte growth factor (KGF 2) on human adult keratinocytes. Methods: The standard medium was keratinocyte growth medium wit hout bovine pituitary extract (BPE), hydrocortisone or epidermal growth factor ( EGF). Keratinocytes from a 48 year old subject were cultured and seeded on dis hes with standard medium of EGF in cell density of 2×10 4/32 mm 2. After 24 hours, the medium was replaced by the standard medium with 0, 4, 16, 125 and 50 0 ng/ml KGF 2, respectively. The standard medium with EGF was used as the posit ive control and the standard medium without EGF or KGF 2 was used as the negati ve controls. The growth of keratinocytes was monitored by 3 (4,5 dimethythiazo l 2 yl) 2,5 dipheyl tetrazolium bromide (MTT) assay and by photographs on day s 3, 5 and 7, respectively. Results: KGF 2 in concentrations of 4 500 ng/ml showed a sign ificant proliferative effect on days 5 and 7 as compared with that of the negati ve controls (P< 0.01 ). On day 3 the cells were prolifer ated to 1.5 2.5 fold, on day 5 to 3 5 fold and on day 7 to 3 12 fo ld in KGF 2 medium as that of the negative controls. The optimal response occur red when the concentration of KGF 2 was 125 ng/ml on day 7. Cell proliferation was also consistently higher in all KGF 2 concentrations as compared with that of the positive controls. Conclusions: KGF 2 has significant effects on the proliferatio n of adult keratinocytes, which are more effective than that of EGF. This study supports KGF 2 can improve the healing of chronic wounds in adults in clinic.

Cytotoxicity and enhancement activity of essential oil from Zanthoxylum bungeanum Maxim.as a natural transdermal penetration enhancer

The aim of this present study is to investigate the effect of Zanthoxylum bungeanum oil （essential oil from Z. bungeanum Maxim.） on cytotoxicity and the transdermal permeation of 5-fluorouracil and indomethacin. The cy- totoxicity of Z. bungeanum oil on dermal fibroblasts and epidermal keratinocytes was studied using an MTT （3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide） assay. The rat skin was employed to determine the percutaneous penetration enhancement effect of Z. bungeanum oil on hydrophilic and lipophilic model drugs, i.e., 5-fluorouracil and indomethacin. The secondary structure changes of the rat stratum comeum （SC） were determined using attenuated total reflectance-Fourier transform infrared spectroscopy （ATR-FTIR）, and saturated solubilities and SC/vehicle partition coefficients of two model drugs with and without Z. bungeanum oil were also measured to un- derstand its related mechanisms of action. It was found that the half maximal inhibitory concentration （ICs0） values of Z. bungeanum oil were significantly lower in HaCaT and CCC-ESF-1 cell lines compared to the well-established and standard penetration enhancer Azone. The Z. bungeanum oil at various concentrations effectively facilitated the percutaneous penetration of two model drugs across the rat skin. In addition, the mechanisms of permeation en- hancement by Z. bungeanum oil could be explained with saturated solubility, SC/vehicle partition coefficient, and secondary structure changes of SC.

TLR3 activation induces S100A7 to regulate keratinocyte differentiation after skin injury

Human S100A7 (psoriasin) is highly expressed in psoriasis and other inflammatory diseases; however, the function of S100A7 in wound repair remains largely unknown. Here we demonstrated that skin injury increased the expression of S100A7. Damaged cells from wounded skin induced the expression of S100A7 via the activation of Toll-like receptor 3 (TLR3) followed by the activation of p38 MAPK. S100A7, in turn, acted on keratinocytes to induce the expression of terminal differentiation marker gene loricrin through the activation of p38 MAPK and caspase-1. The differentiation of keratinocytes induced by S100A7 resulted in skin stratification, thus efficiently promoting wound closure. Taken together, our results demonstrate that the activation of TLR3 accelerates wound closure via the induction of S100A7 to induce keratinocyte differentiation. These findings also provide new insights into the development of different forms of treatment with skin wounds.

Essential oil from Zanthoxylum bungeanum Maxim. and its main components used as transdermal penetration enhancers: a comparative study

Our previous studies had confirmed that the essential oil from Zanthoxylum bungeanum Maxim. （Z. bungeanum oil） could effectively enhance the percutaneous permeation of drug molecules as a natural transdermal penetration enhancer. The aim of the present study is to investigate and compare the skin penetration enhancement effect of Z. bungeanum oil and its main components on traditional Chinese medicine （TCM） active components. Toxicities of Z. bungeanum oil and three selected terpene compounds （terpinen-4-ol, 1,8-cineole, and limonene） in epidermal keratinocytes （HaCaT） and dermal flbroblast （CCC-ESF-1） cell lines were measured using an MTT （3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide） assay. Five model drugs in TCM external preparations, namely osthole （OT）, tetramethylpyrazine （TMP）, ferulic acid （FA）, puerarin （PR）, and geniposide （GP）, which were selected based on their lipophilicity denoted by IogKo^w, were tested using in vitro permeation studies in which vertical Franz diffusion ceils and rat abdominal skin were employed. The secondary structure changes of skin stratum corneum （SC） and drug thermodynamic activities were investigated to understand their mechanisms of action using Fourier transform infrared （FTIR） spectroscopy and saturation solubility studies, respectively. It was found that Z. bungeanum oil showed lower toxicities in both HaCaT cells and CCC-ESF-1 cells compared with three terpene compounds used alone. The enhancement permeation capacities by all tested agents were in the following increasing order： terpinen-4-ol=1,8-cineole〈limonene〈Z, bungeanum oil. The mechanisms of permeation enhancement suggested that these enhancers promoted the skin permeation of drugs mainly by affecting SC lipids. These results indicated that Z. bungeanum oil exhibited better performance in enhancing the skin permeation of active components in TCM preparations.

Microenvironment-induced myofibroblast-like conversion of engrafted keratinocytes

Myofibroblasts,recognized classically by-smooth muscle actin(-SMA)expression,play a key role in the wound-healing process,promoting wound closure and matrix deposition.Although a body of evidence shows that keratinocytes explanted onto a wound bed promote closure of a skin injury,the underlying mechanisms are not well understood.The basal layer of epidermis is rich in undifferentiated keratinocytes(UKs).We showed that UKs injected into granulation tissue could switch into-SMA positive cells,and accelerate the rate of skin wound healing.In addition,when the epidermis sheets isolated from foreskin cover up the wound bed or are induced in vitro,keratinocytes located at the basal layers or adjacent sites were observed to convert into myofibroblast-like cells.Thus,UKs have a potential for myofibroblastic transition,which provides a novel mechanism by which keratinocyte explants accelerate skin wound healing.

Fractional-Order Model of the Disease Psoriasis:A Control Based Mathematical Approach

Autoimmune diseases are generated through irregular immune response of the human body. Psoriasis is one type of autoimmune chronic skin diseases that is differentiated by T-Cells mediated hyper-proliferation of epidermal Keratinocytes. Dendritic Cells and CD8＋ T-Cells have a significant role for the occurrence of this disease. In this paper, the authors have developed a mathematical model of Psoriasis involving CD4＋ T-Cells, Dendritic Ceils, CD8＋ T-Cells and Keratinocyte cell populations using the fractional differential equations with the effect of Cytokine release to observe the impact of memory on the cell-biological system. Using fractional calculus, the authors try to explore the suppressed memory, associated with the cell-biological system and to locate the position of Keratinocyte cell population as fractional derivative possess non-local property. Thus, the dynamics of Psoriasis can be predicted in a better way using fractional differential equations rather than its corresponding integer order model. Finally, the authors introduce drug into the system to obstruct the interaction between CD4＋ T-Cells and Keratinocytes to restrict the disease Psoriasis. The authors derive the Euler-Lagrange conditions for the optimality made through Matlab by developing iterative of the drug induced system. Numerical simulations are schemes.

Loss of cathepsin C enhances keratinocyte proliferation and inhibits apoptosis

Papillon-Lefevre Syndrome is a rare autosomal recessive disorder characterized by rapidly progressive periodontitis and confined palrnoplantar hyperkeratosis resulting from genetic mutations in cathepsin C （CTSC）. The present study investigated the effect of CTSC on keratinocyte proliferation and apoptosis. HaCaT keratinocytes were transfected with wild-type CTSC and CTSC-targeted siRNAs to investigate the effects of CTSC expression on cell keratosis. Real-time PCR and Western blot analyses showed that the levels of loricrin and keratin （KRT）-I, but not KRT9, was correlated with CTSC expression. Loricrin was increased in the CTSC-overex- pression group and downregulated in the CTSC-silenced group. A positive association between loricrin expression and cell apoptosis was detected in HaCaT keratinocytes. KRT1 was decreased in the CTSC-overexpression group and increased in the CTSC-silenced group. Prominent, punctuate KRT1 aggregates were present in CTSC-knockdown HaCaT cells. This study suggested that loss of CTSC contributes to keratinocyte hyperkeratosis via downregulation of loricrin and enhanced cell proliferation.