Objective To explore if keratinocytes that stably maintain HPV11 genome can be obtained by transfection and selection methods. Methods Escherichia coil containing pBR322.HPV11 plasmid was cultured and amplified. Then the plasmid was extracted, purified and digested with BamH Ⅰ enzyme to release viral genome from the bacterial vector. After recovering from the low-melting point agarose gel by electrophoresis, the genome was self-circulated with T4 DNA ligase. The religated DNA was cotransfected with pTK-neo DNA into HaCaT keratinocytes using Lipofectamine reagent. After selection with G418 for 2 to 3 weeks, clonal and pooled cultures were expanded and analyzed. Fluorescent quantitative PCR (FQ-PCR) and nested reverse transcriptase PCR (nRT-PCR) were applied to detect HPV11 DNA and spliced HPV11 E1^E4 mRNA expression in the transfected cells. Results After the cotransfection of HPV11 genome into HaCaT keratinocytes and two-week selection,G418-resistant cell colonies were obtained with morphological features indistinguishable from normal HaCaT keratinocytes. As shown by FQ-PCR, HPV11 DNA was present in G418-selected HaCaT keratinocytes. The average viral DNA load capacity was 15.9±16.8 copies/cell in the primary culture of G418-selected HaCaT cells and 23.9±1.1 copies/cell in the third passage of the cells; there was no statistical difference between the two passages of cells (t=-0.822, P>0.05). nRT-PCR targeting HPV11 E1^E4 mRNA transcript produced a specific 628-bp fragment, which was shown by agarose gel electrophoresis. Conclusions Our data indicate that HPV11 genome can be successfully introduced into HaCaT keratinocytes by transfection and HPV11 DNA-positive cells can be obtained by G418 selection. Moreover, HPV11 DNA is still present in the third passage of transfected cells.

Objective To determine whether leflunomide could control the proinflammatory cytokine expression induced by anthralin via inhibiting the activation of nuclear factor ?B (NF-?B). Methods The expression of NF-?B inhibitory protein ? (I?B?), was analyzed by using Western blot method. MTT assay and RT-PCR were used to assess the proliferating activity and mRNA expression of intercellular adhesion molecule-1 (ICAM-1) of HaCaT keratinocytes, respectively. Results Leflunomide inhibited the degradation of I?B? by anthralin, i.e. the activation of NF-?B signaling pathway, in a dose-dependent manner. The inhibition of keratinocyte growth by anthralin did not correlate with the activation of NF-?B. Under the experimental conditions used, leflunomide was shown to be able to significantly inhibit the over-expression of ICAM-1 on keratinocytes induced by anthralin; this inhibition occurred in a dose dependent manner. Conclusions Growth inhibition by topical anti-psoriatic medication anhtralin is unrelated to the NF-?B-dependent signaling pathway, and leflunomide can control ICAM-1 expression induced by anthralin via inhibiting the activation of NF-?B.

Objective To investigate the effects of the compound T0 extracted from Tripterygium wilfordii, a traditional Chinese herb, in comparison with those of cyclosporin A(CyA), on down－ regulating the expression and activities of IL－ 1 and IL－ 2. Methods The cellular reactive systems of peritoneal macrophage (rat)－ thymocytes (rat), and spleen cells (rat)－ CTLL－ 2 (mouse) were set up. The technique of 3H－ TdR incorporation was applied. Results The production of IL－ 1 and IL－ 2 were significantly inhibited by T0 and CyA. The activities of IL－ 1 and IL－ 2 were down－ regulated by T0,rather than by CyA. Conclusion T0 and CyA are potent inhibitors of the production of IL－ 1 and IL－ 2. Effects of T0 on the activities of IL－ 1 and IL－ 2 are different from those of CyA.

Objective To Investigate the effects of antipsoriatic drugs on 5 lipoxygenase(5－LO) and set up a relevant pharmacodynamic method. Methods 5－ LO products, leukotriene B4 (LTB4) and 5 hydroxyeicosatetraenoic(5－HETE), we re determined by RP－HPLC to represent 5－LO activity. Results Cyclosporin A( CyA) and triptolide(T0) inhibited the production of LTB4 and 5－HETE in a dose dependent manner, while erythromycin did without dose dependence. The 50％ inhibitory concentration values(IC50) of CyA inhibiting LTB4 and 5－HETE were 3 8.0? g/mL and 0.96? g/mL, respectively. The IC50 of T0 inhibiting LTB4 and 5－ HETE were 2.3? 10－6? g/mL and 1.14? 10－6? g/mL, respectively. Conclusio ns The anti inflammatory effect of Tripterygium wilfordii Hook.f. may be partl y explained by its inhibition of 5－LO activity. The anti inflammatory effect of CyA has no clinical significance since the inhibitory concentration of CyA h as exceeded its pharmacological limitation. Erythromycin has no effect on 5－LO activity.

Objectives To establish an experimental model for Th1 typ e shifting and meet the requirements of studying on the mechanisms of some immun omodulators. Methods The levels of cytokines, IL-12, IFN-ice were detected by using ELISA. Sple en cells of the BALB/c mice were incubated under the following conditions: with different concentrations of T cell mitogen ConA (1 mg/mL, 0.5 mg/mL, 0.25 mg/mL, 0.125 mg/mL), mononuclear phagocyte system activator LPS (50 mg/mL, 5 mg/mL, 0. 5 mg/mL) or LPS (50 mg/mL, 5 mg/mL, 0.5 mg/mL) combined with 0.25 mg/mL ConA. Re sults LPS could induce the production of IL-12 from spleen cells. The lowest concentration that ConA could induce the measurable production of IFN-rom sp leen cells was 0.25 mg/mL. When different concentrations of LPS were combined wi th 0.25 mg/mL ConA, LPS could accelerate the production of IFN- and positively with that of IL-12. Conclusion LPS combined with ConA can induce the activation of spleen cells from mice towards Th1 type response.

Objective To determine the normal range of minimal erythema dose (MED) of normal skin to ultraviolet A (UVA) and B (UVB). Methods The definition of MED is the dose of UVA required to induce a just perceptible erythema on an individual′s skin 24 hours after irradiation. One hundred and eighteen subjects including healthy volunteers and patients with noninflammatory skin disorders were enrolled and studied with SUV1000 type UV simulator in March 2002. Results The average MED value for UVA was 55 J/cm2 (range: 18 - 95 J/cm2) in the males, and 40 J/cm2 (range: 15 - 100 J/cm2) in the females. The average MED value for UVB was 31 mJ/cm2 (range: 12 - 95 mJ/cm2) in the males and 29 mJ/cm2 (range: 8 - 95 mJ/cm2) in the females. The MED value for UVA in the males was significantly higher than that in the females (P 0.05). The MED values for UVA as well as UVB in skin type Ⅲ were significantly lower than those in skin type Ⅳ (UVA-MED: P

Objective To study the role of photoallergens and contact allergens in the pathogenesis of chronic actinic dermatitis (CAD).Methods Ba sed on the standard procedures of photopatch test recommended by the British Pho todermatology Group (BPG) and the routine procedures of patch test,photopatch a nd patch tests were performed on 56 patients with CAD,42 patients with polymorp hous light eruption (PLE) and 25 patients with chronic eczema on scalp and face by standard photopatch test series recommended by the International Contact Derm atitis Research Group (ICDRG) and home-made standard series of contact allerge ns.A set of ten Philips TL20W/09N tubes was used as the source of irradiation.Results In the 56 CAD patients,the positive rates were 46.43 %,57.14 % and 32.14% for photopatch test,patch test and both tests,respectively,which appea red to be significantly higher than those in the patients with PLE.Positive pa tch reactions were found in 65% of the patients with chronic eczema,which was s imilar to that of CAD.And the frequency of the positive allergens in chronic ec zema was the same as that in CAD,in which fragrance mixture (FM) ranked the fir st,followed by balsam of Peru (BOP),cobalt chloride,nickel sulphate.In CAD,FM and BOP were the most common allergens and photoallergens,which accounted fo r 44% and 32% of the positive reactions in patch tests,15.38% and 17.95% in pho topatch tests,respectively.Conclusions Both photoallergens and contact aller gens may play important roles in the pathogenesis of CAD.Allergens positive in patch tests and photopatch tests and related compounds which can cause cross-r eactivity with the above allergens should be avoided by the patients with CAD.

Objective To study the relationship of sunlight exposure and photoprotection with clinical activity in systemic lupus erythematosus. Methods A structured questionaire was administered to 107 SLE patients, to assess their attitudes and behavior regarding sunlight exposure and photoprotection. The clinical manifestations, laboratory findings and treatment were evaluated. Results All patients were classified into two groups based on the duration of exposure to sunlight per day. Fourty-eight (44.86%) patients were exposed to direct sunlight for an average of less than one hour per day in one group and 59 (55.14%) for one hour or more in the other group. Twenty-four (22.43%) patients reported use of photoprotective measures in spring and summer. The patients in the former group had significantly lower incidences of photosensitivity, arthritis, alopecia, exacerbations, presence of anti-dsDNA antibody, decrease of complement C3, C4 and CH50 than those in the latter group(P

Objective To investigate the effects of some compounds,bestatin,tripdiolide,etc,on leukotriene A4hydrolase activity.Methods LTB4,product of leukotriene A4hydrolase,was determined by reverse phase high performance liquid chromatography(PR-HPLC).Bestatin,an inhibitor of LTA4hydrolase was used as a positive control.Results Tripdiolide could inhibit LTA4hydrolase activity in a dose-depen-dent pattern.The50%inhibitory concentration values(IC 50 )was2.58?10 -5 ?g/mL.The other compounds,an-thralin,cyclosporin A,tretinoin,calcipotriene,clobetasol propionate,methotrexate,and erythromycin,had no effects.Conclusions Tripdiolide possibly has anti-inflammatory effect through down-regulation of leukotriene A4hydrolase in psoriasis.

Objective To investigate the effects of keratinocyte growth factor (KGF) and KGF receptor (KGFR) antisense oligonucleotide (ASODN) on cell cycle and apoptosis of HaCat cells. Methods HaCaT cell, an immortalized keratinocyte cell strain, was cultured in vitro. Flow cytometry was used to measure the cell cycle and apoptosis mediated by KGF and ASODN. Results The rates of S phase and apoptosis in the group treated with KGF increased significantly than those in the control group (both P