Objective To analyze the clinical characteristics and treatment of acne inversa.Methods Seventeen outpatients with acne inversa were collected in the Institute of Dermatology,Chinese Academy of Medical Sciences and Peking Union Medical College from January l,2012 to December 31,2012.The general condition,clinical feature and treatment of these patients were retrospectively analyzed.Results All the patients were male with the age at onset being about 20 years and disease duration varying from 2 to 50 years.Characteristic clinical manifestations were recurrent tender inflammatory papules,nodules,abscesses,fistulae and sinus tracts in the neck,axillary fossa,groin,perineum and buttocks.Among these patients,10 had a family history and seven were sporadic with mild symptoms.Oral tretinoin combined with antibiotics were the main treatment,and surgical treatment was usually used for severe patients.Conclusions Acne inversa is mainly manifested as abscess,sinus and scars in areas bearing apocrine sweat glands,and therapeutic regimen should be selected according to the severity of lesions.

Objective To detect the mRNA and protein expressions of desmoglein 1 (DSG1) and DSG3 in different types of keratinocytes (KCs).Methods Two keratinocyte cell lines HaCaT and A431,as well as primary keratinocytes from human abdomenal skin served as the object of this study.Direct immunofluorescence assay was performed to observe and quantify the expressions of DSG1 and DSG3,and quantitative PCR (qPCR) to determine the mRNA expressions of DSG1 and DSG3,in these cells.Results Both DSG1 and DSG3 were expressed in all the three types of keratinocytes,and the fluorescence intensity of DSG1 and DSG3 in HaCaT cells was higher than that in primary keratinocytes but lower than that in A431 cells.Similarly,all the keratinocytes expressed DSG1 and DSG3 mRNA,with the relative expression levels of DSG1 and DSG3 mRNA in primary keratinocytes being 291.7％ and 237.4％ of those in HaCaT cells respectively (both P ＜ 0.01),and those in A431 cells being 0.1％ and 18.8％ of those in HaCaT cells respectively (both P ＜ 0.05).Conclusions HaCaT cells,A431 cells and primary keratinocytes all can be used for the study of DSG1 and DSG3,of which,A431 cells show the strongest expressions of DSG1 and DSG3,and primary keratinocytes display the highest expressions of DSG1 and DSG3 mRNAs.

Objective:To establish a presenilin enhancer-2 (PSENEN) gene-silenced human immortalized keratinocyte (HaCaT) cell model, and to evaluate the effect of PSENEN gene silencing on the proliferation of and γ-secretase expression in HaCaT cells.Methods:Three shRNAs targeting the PSENEN gene were constructed, and inserted into the linearized LV3-pGLV-h1-GFP-puro vector to establish a recombinant lentiviral expression plasmid. After restriction enzyme digestion and sequencing, lentiviral packaging and purification were performed, and lentiviral titer was determined. Cultured HaCaT cells were divided into 5 groups: shRNA1, shRNA2 and shRNA3 groups treated with the lentivirus solutions containing PSENEN gene-targeted shRNA1, shRNA2 and shRNA3 respectively, NC group treated with the lentivirus solution containing a negative control shRNA (shNC) , and blank group treated without lentivirus solution. After transfection, inverted fluorescence microscopy was performed, and transfection efficiency was determined by flow cytometry. Cell counting kit-8 (CCK8) assay was performed to evaluate the effect of PSENEN gene silencing on the proliferation of HaCaT cells, and real-time fluorescence-based quantitative PCR (qPCR) and Western blot analysis were conducted to determine the mRNA and protein expression of PSENEN, nicastrin (NCT) , presenilin-1 (PS1) and anterior pharynx defective 1a (APH1a) genes respectively. Statistical analysis was carried out by using repeated measures analysis of variance, one-way analysis of variance, and least significant difference t test for multiple comparisons. Results:Inverted fluorescence microscopy showed that fluorescence was observed in the shRNA1 group, shRNA2 group, shRNA3 group and NC group, and flow cytometry showed that the transfection efficiency was over 98% in the above 4 groups. qPCR and Western blot analysis revealed that the mRNA and protein expression of PSENEN gene significantly decreased in the shRNA1 (0.187 ± 0.010, 0.219 ± 0.097, respectively) , shRNA2 (0.163 ± 0.022, 0.208 ± 0.014, respectively) and shRNA3 (0.174 ± 0.009, 0.185 ± 0.062, respectively) groups compared with the NC group (1.054 ± 0.272, 1.076 ± 0.075, respectively, all P < 0.001) . CCK8 assay showed that the cellular proliferative activity significantly increased in the shRNA1 group compared with the NC group at 0, 12, 36 and 48 hours (all P < 0.05) , and there was no significant difference between the 2 groups at 24 or 60 hours (both P > 0.05) ; the cellular proliferative activity was significantly higher in the shRNA2 and shRNA3 groups than in the NC group at 0, 12, 24, 36, 48 and 60 hours (all P < 0.05) . There was no significant difference in the mRNA expression of NCT, PS1 and APH1a genes among the shRNA1 group, shRNA2 group, shRNA3 group, NC group, and blank group ( F= 8.168, 4.644, 1.981, respectively, all P > 0.05) , while the relative protein expression level of mature NCT (mNCT) , immature NCT (imNCT) , carboxyl-terminal fragment of PS1 (PS1-CTF) and APH1a significantly differed among the above 5 groups ( F= 39.268, 5.929, 27.842, 20.663, respectively, all P ≤ 0.01) . Compared with the NC group, the shRNA1, shRNA2 and shRNA3 groups all showed significantly decreased protein expression of mNCT, PS1-CTF and APH1a (all P < 0.01) , but insignificant changes in imNCT protein expression (all P > 0.05) . Conclusion:The PSENEN gene-silenced HaCaT cell model was successfully constructed, and the PSENEN gene silencing could lead to an increase in the cellular proliferative activity of HaCaT cells and a decrease in the protein expression of γ-secretase subunits mNCT, PS1-CTF and APH1a.

Objective To investigate the efficacy of enteral nutrition containing glutamine on improving blood lipids and immune parameters in elderly patients with gastrointestinal cancer.Methods 98 cases of elderly patients with gastrointestinal cancer were randomly divided into observation group(n=5 1)and control group (n=47).The control group was additionally given conventional enteral nutrition,and the observation group was additionally given enteral nutrition containing glutamine.The levels of albumin (ALB),tumor necrosis factor (TNF),indicators of cellular immunity,indicators of humoral immune function and blood lipid,before and after operation,were observed in the two groups.Results 9 days after operation,the levels of ALB and total cholesterol,per-centage of CD8 + cells were lower than those in the control group,and levels of TFN,IgA,triacylglycerol,low density lipoprotein cholesterin,percentage of CD4 + cells and CD4/CD8 ratio were higher than those in the control group,and have significant differ-ences (P <0.05).While,there was no statistically significant difference of the percentage of CD3 cells between the groups(P >0.05).Conclusion Enteral nutrition containing glutamine can significantly improve blood lipids and immune function in elderly pa-tients with gastrointestinal cancer,which may have important clinical research value and be worthy of further application.

Objective Colorectal polyp is a precancerous lesion of colorectal cancer .Aim of the study was to explore the risk fac-tors of colorectal polyp malignant transformation .Methods The related information of 75 084 colonoscopies performed from 2003 to 2012 in Southwest Hospital in Chongqing were collected and the relationship between polyp malignant transformation and the pa-tient age ,sex ,polyp location ,size or histological types was analyzed .Results From 2003 to 2012 ,polyps were diagnosed in 14 806 cases of the total 75 084 patients with a 19 .72% detection rate .There were significant difference of the left-side and right-side pol-yp detection rate in different age groups ,and the frequency of polyps distributed in the whole colorectum increased with the increase of age .The rates of epithelial neoplasia and malignant transformation increased with age .At the same time ,malignant transforma-tion rate was significant higher in polyps located in left-side than that in right-side (P<0 .0167) ,in adenoma than that in inflamma-tory hyperplastic polyp (P<0 .01) .The larger diameter and the more villus ,the higher rate of malignant transformation .Conclusion Patient age ,polyp size ,location and histological type could be considered as the significant predictors of colorectal polyp malignant transformation .It may be useful to treat the polyp with endoscopy in patient with age more than 45 and adenoma whose diameter was not less than 1 cm ,located in left-side for prevention of colorectal cancer .

Objective To evaluate the reversal effect of a cholinergic receptor agonist on acantholysis in pemphigus,and to investigate its mechanism.Methods Human HaCaT keratinocytes were co-cultured with pemphigus vulgaris immunoglobulin G (PV-IgG) to establish a cell model of pemphigus,then classified into two groups to be incubated with the cholinergic receptor agonist carbachol for 12 hours (test group) or remain untreated (control group).Cell dissociation assay was performed to quantitatively estimate the reversal effect of carbachol on acantholysis,and immunofluorescence assay to qualitatively assess the changes of desmosomal proteins.Radio-immunoprecipitation assay (RIPA) lysis buffer and Triton X-100 were used to lyse HaCaT cells to obtain total proteins and cytoplasmic proteins,and Western blot was conducted to determine the expression levels of adhesion-related proteins desmoglein 3 (Dsg3) and plakoglobin (PG) on the surface of HaCaT cells,as well as the phosphorylation levels of p38 mitogen activated protein kinase (p38 MAPK) and epidermal growth factor receptor (EGFR) at different time points.Quantitative polymerase chain reaction (qPCR) was performed to detect the mRNA expressions of the above surface proteins,and coimmunoprecipitation assay to qualitatively evaluate the interaction between Dsg3 and PG.Results The number of cell debris was significantly lower in the test group than in the control group (18.67 ± 2.52 vs.46.67 ± 2.03,t =11.22,P＜0.01).Immunofluorescence assay showed that carbachol could reverse the internalization of desmosomal molecules induced by PV-IgG.In the pemphigus cell model,the levels of total Dsg3 and PG as well as non-desmosomal Dsg3 were decreased,while the level of non-desmosomal PG increased,and the interaction between Dsg3 and PG was attenuated.When the pemphigus cell model was co-cultured with carbachol,these above changes were reversed.Carbachol also increased the mRNA levels (expressed as 2-△△Ct) of Dsg3 and PG from 1.428 ± 0.215 and 1.563 ± 0.247 in the control group to 4.974 ± 0.948 (t =3.65,P =0.01) and 13.420 ± 1.715 (t =6.85,P ＜ 0.01) in the test group respectively.In phosphorylation assay,carbachol inhibited the phosphorylation of EGFR,but had no significant effect on that of p38 MAPK.Conclusions The cholinergic receptor agonist carbachol can reverse acantholysis in pemphigus,likely by inhibiting the internalization of Dsg3 and PG,enhancing their expressions and interaction,and suppressing the phosphorylation of the key signaling molecule for acantholysis,EGFR.

Objective To measure expressions of nicastrin and its downstream Notch-HES signaling pathwayassociated proteins in skin lesions of patients with acne inversa harbouring nicastrin gene mutations.Methods An immunohistochemical study was performed to measure the expressions of nicastrin and Notch-HES signaling pathwayassociated proteins in paraffin-embeded skin samples from lesions of 4 patients with acne inversa and confirmed nicastrin mutations and from normal skin of 6 human controls.Spearman correlation analysis was carried out to assess the relationship between the expressions of nicastrin and Notch-HES signaling pathway-associated proteins.Results In normal control skin samples,nicastrin was widely distributed in the full-thickness epidermis and skin appendages such as pilosebaceous units,apocrine glands and eccrine glands.However,the expressions of nicastrin and Notch-HES signaling pathway-associated proteins were markedly decreased in the epidermis and hair follicle infundibulum in lesions of patients harbouring nicastrin gene mutations compared with normal control skin.Furthermore,nicastrin expression was positively correlated with Notchl,Notch3 and HES-1 expressions (r =0.831,0.748 and 0.807,P ＜ 0.01,0.05 and 0.01 respectively),but not significantly correlated with Notch2 or HES-5 expressions (r =0.597,0.591 respectively,both P ＞0.05).Conclusion Nicastrin expression markedly decreases in lesions of patients with acne inversa harbouring nicastrin gene mutations,and is positively correlated with the expressions of several Notch-HES signaling pathway-associated proteins,suggesting that the decrease in nicastrin expression may take part in the pathogenesis of acne inversa by influencing the expression of the downstream Notch-HES signaling pathway.

Objective To analyze γ?secretase gene mutations in a pedigree with acne inversa. Methods Clinical data were collected from a pedigree with acne inversa, which contained 30 members spanning 4 generations. Of these members, 12 were affected by acne inversa, and 9 of the affected members were alive. Peripheral blood DNA was obtained from the proband, his seven relatives (including 4 affected and 3 unaffected members), and 100 unrelatedhealthy human controls. PCR was performed to amplify all the coding exons and their flanking sequences of the NCSTN, PSEN1, PSENEN, Aph1 genes followed by DNA sequencing. Results A heterozygous insertion mutation (c.229_230insCACC)of the PSENEN gene, which led to translational frameshifting and resulted in dysfunciton of the PSENEN protein, was detected in all the 5 patients, but not in unaffected members or healthy controls. Conclusion There is a novel heterozygous insertion mutation c.229_230insCACC in the PSENEN gene, which may be the molecular basis of acne inversa in this family.

Objective To construct a lentiviral vector delivering the Nicastrin (NCT) gene-targeted short hairpin RNA (shRNA) and determine gene-silencing efficiency of the vector in the human immortalized keratinocyte cell line HaCaT,and to construct a NCT gene-silenced HaCaT cell model to lay an experimental foundation for subsequently studying effects of NCT gene silencing on biological behavior of keratinocytes.Methods Three NCT gene-targeted shRNAs were designed and inserted into the pGLV3/ H1/GFP + Puro vector to construct three recombinant plasmids,which were then confirmed by sequencing.Recombinant plasmids combined with lentivirus packaging plasmids were co-transfected into 293T cells to obtain lentivirus particles,and the virus titer was determined.Cultured HaCaT cells were divided into 3 groups:blank group receiving no treatment,negative control group infected with the empty vector LV3-shNC,interference groups infected with lentivirus NCT-shRNA1,-shRNA2,-shRNA3,respectively.Flow cytometry was performed to determine transfection efficiency,and real-time fluorescence-based quantitative PCR (qRT-PCR) and Western blot analysis were conducted to determine efficiency of target gene silencing in HaCaT cells,so as to select the most efficient interference sequence.Results Sequencing analysis indicated that recombinant lentiviral vector NCT-shRNA was constructed successfully.After co-transfection of recombinant plasmids and lentivirus packaging plasmids into 293T cells,the titer of recombinant lentivirus particles was about 109 TU/ml.Flow cytometry showed that the transfection efficiency was greater than 95％.qRT-PCR revealed that the NCT mRNA expression was obviously down-regulated in the interference group compared with the negative control group,and NCT-shRNA1 was the most efficient sequence with interference efficiency being 75％.Western blot analysis showed that the inhibition rate of NCT protein expression was 71.7％ in the shRNA1 group compared with the negative control group.Conclusion The most efficient NCT-shRNA interference sequence is screened out,and the recombinant lentiviral vector NCT-shRNA and an NCT gene-silenced HaCaT cell model are both constructed successfully.

significantly save the time of diagnosis and facilitate the clinical application of large samples.