Objective To develop a novel padlock probe and rolling circle amplification(RCA) to detect 5 common cell culture contaminant Mollicute( Mycoplasma arginini, Mycoplasma fermentans, Mycoplasma hyorhinis, Mycomplasma orale, and Acholeplasma laidlawii ). Methods "Universal" primers ( SPS1, SPA2 and SPS2, SPA1) were used to amplify the Mollicute 16S-23S rRNA intergenic spacer region.Amplicon was ligate Mollicute specific padlock probe. Probe amplified and monitored using a Corbett RotorGeneTM 6000 machine. Results Five reference Mollicute strains were correctly detected by RCA method.There was no cross-reaction. RCA method can sensitive detect 10 copies templates and show strong positive signal. Sixty-two cell culture specimens were detected. Thirty-seven samples were contained single specie Mollicute and 14 samples contained two Mollicutes. Eleven samples did not contained Mollicute. RCA detection results were concordant with previously species-specific PCR. Conclusion RCA can rapid, sensitive and specific detect contamination Mollicute.

Objective To examine the reliability,validity,and practicability of Cognitive Emotion Regulation Questionnaire (CERQ) in hypertensive patients in China.Methods Altogether 434 hypertensive patients and 462 healthy subjects were recruited. All the subjects were assessed with the CERQ-Chinese version (CERQ-C), Dysfunctional Attitude Scale (DAS), Mood and Anxiety Symptom Questionnaire-Short Form (MASQ-SF), and Center for Epidemiologic Studies Depression Scale (CES-D). We calculated the mean inter-item correlations for the total CERQ and for each of the subscales. Cronbach's alpha coefficient was used to analyze the inter-correlation and reliability, and confirmatory factor analysis was used to examine the 9-factor model. Results 1) Hypertension group reported significantly higher score than that of healthy ones on rumination (12.19±2.51 vs. 11.51±2.60, P<0.001), catastrophizing(8.82±2.19 vs.8.11±2.70,P<0.001),and blaming others(10.76±2.11 vs. 9.88±2.48,P<0.001), and had significantly lower score than that of healthy ones on positive reappraisal(13.80±3.55 vs.14.71±4.11,P<0.001).2)Reliability:In the hypertension group the Cronbach's alpha for the total CERQ was 0.80, and that for the 9 subscales ranged from 0.71 (self-blame) to 0.90 (rumination). In the healthy group the Cronbach's alpha for the total CERQ was 0.79, and that for the 9 subscales ranged from 0.71 (positive reappraisal) to 0.90 (rumination). The mean inter-item correlation coefficient for the 9 subscales was 0.21-0.42(the hypertension group)/0.19-0.32 (the healthy group). In the hypertension group,the test-retest reliability of the total scale was 0.82, the test-retest reliability of the 9 subscales ranged from 0.73 to 0.92. The confirmatory factor analysis showed that the 9 first-order factor data fitted both 2 samples well. Conclusion CERQ meets the psychometric standard and it is reliable and valid for cognitive emotion regulation strategies, which may be regarded as an appropriate assessment tool.

Objective To estimate the expression of ER and PR in the endometrium of both intrauterine adhesions (IUA) and non-IUA specimens. Methods The endometrium specimens from patients undergoing hysteroscopy for confirmed moderate IUA (n=20: 10 in proliferative phase, and 10 in secretory phase) were enrolled as the IUA group in Beijing Obstetrics and Gynecology Hospital from October 2014 to August 2015. The specimens scheduled for hysteroscopy due to infertility were recruited into the control group (n=26: 13 in proliferative phase, and 13 in secretory phase). Immunohistochemistry and quantificational real-time PCR (qRT-PCR) were used to detect the expression of ER-α, ER-β and PR in endometrium with different menstrual period in both groups. Results (1) Location: in both groups, the expression of ER-α, ER-β and PR appeared in the endometrial glandular epithelial cells and the stromal cells of the endometrium. The positive brown granules of ER-α, ER-β and PR appeared mainly in cell nucleus. (2) ER-α and ER-β in the endometrium:the protein expression of ER-α and ER-β in IUA group (proliferative phase: 0.657 ± 0.028, 0.493 ± 0.023; secretory phase: 0.537 ± 0.020, 0.365 ± 0.031) were significantly higher than those of control group (proliferative phase: 0.586 ± 0.025, 0.437 ± 0.022; secretory phase:0.459 ± 0.025, 0.323 ± 0.017;all P<0.01). And the ER-αand ER-βmRNA expressions in IUA group were 2.524 ± 0.296, 1.947 ± 0.339, higher than those of control group in the proliferative phase (all P<0.01), and in the secretory phase (1.977±0.333, 1.345±0.292) were also higher than those in the control group (all P<0.01). (3) PR in the endometrium: the protein expression of PR was not significantly different between IUA group (proliferative phase:0.248±0.025, secretory phase:0.194±0.024) and control group (proliferative phase: 0.234 ± 0.019, secretory phase: 0.186 ± 0.020; P=0.162, 0.359). Meanwhile, there were no statistical differences in the mRNA expression of PR in both groups with different menstrual period (proliferative phase: 1.144 ± 0.384 versus 0.981 ± 0.306, secretory phase: 0.763 ± 0.237 versus 0.631 ± 0.203; P=0.270, 0.166). (4) ER and PR expression in menstrual cycles: the expression of ER-α, ER-β and PR in the IUA group changed with the menstrual cycles, and their expression in the proliferative phase were higher than those in the secretory phase (all P<0.05). Conclusions The expression of ER-α and ER-β in the endometrium of IUA patients changes with menstrual cycle, and are higher compared with those in normal endometrium. No difference is found in the PR expression between the two groups.

AIM: To examine the effect of cholecystokinin-octapeptide (CCK-8) on focal cerebral ischemia/reperfusion injury and its underlying mechanisms. METHODS: By using the suture model of focal cerebral ischemia and reperfusion, the effects of intracerebroventricular (icv) injection of CCK-8 and proglumide, nonselective CCK receptors antagonist, on the infarct size, regional cerebral blood flow (rCBF), and the levels of nitric oxide (NO), malondialdehyde (MDA) were observed in different brain regions of rats subjected to 1 h focal cerebral ischemia followed by 24 h reperfusion. RESULTS: (1) pretreatment with different doses of CCK-8 (0.3 ?g,1.0 ?g,2.0 ?g or 4.0 ?g) could attenuate the infarct size, but the statistically significant effects of CCK-8 were obtained only at the doses of 1.0 ?g and 2.0 ?g(P

To investigate whether LL-37 and human beta defensin-2 (hBD-2) is related to the patients with psoriasis seldom having skin infections and explore the role of the two peptides and CCR6 (the receptor of hBD-2) in the pathogenesis of psoriasis, the expression levels of mRNA of LL-37, hBD-2, and CCR6 in skin lesions of patients with psoriasis vulgaris were detected by using RT-PCR. The results showed that the mRNA expression levels of the two peptides and CCR6 in psoriatic lesions all increased compared with the normal skin (P<0.001). It was suggested that up-regulated expression of LL-37 and hBD-2 might be the main reason that result in the the skin of patients with psoriasis being seldom infected, and the two peptides and CCR6 might play crucial roles in the pathogenesis of psoriasis.
Adult ; Antimicrobial Cationic Peptides ; biosynthesis ; genetics ; Female ; Humans ; Male ; Middle Aged ; Psoriasis ; genetics ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Receptors, CCR6 ; Receptors, Chemokine ; biosynthesis ; genetics ; Up-Regulation ; beta-Defensins ; biosynthesis ; genetics

The histological and immunohistological features of two cases of cutaneous lymphadenoma was studied. A single, erythematous nodule with smooth surface developed on the face of both patients. The lesion slowly progressed. Histology revealed irregular epithelial lobules in the dermis which showed a peripheral palisaded border of basaioid-like cells as well as a center composed of clear cells. Some epithelial lobules and surrounding stroma were infiltrated by numerous small lymphocytes. Immunohistological study showed that the lymphocytic infiltration was predominantly composed of T cells (CD3 positive) along with a small number of B cells (CD20 positive). Within epithelial lobules and surrounded stroma, there were numerous dendritic cells that were positive for S-100 and CDia but negative for cytokeratin 7, cytokeratin 20 or carcino-embryonic antigen. In the center of epithelial lobules in one case, a few cells positive for epithe-lial membrane antigen and CD30 was observed. The diagnosis of cutaneous lymphadenoma was made according to the pathological and immunohistochemical findings, and the infiltration was predominated by CD3-positive lymphocytes in this uncommon epithelial neoplasm.

Objective To compare the reactive oxygen species (ROS) expression in human neutrophils phagocytizing Sporothrix Schenckii and Candida albicans yeast cells,and to compare the fungicidal activity of human neutrophils against Sporothrix Schenckii and Candida albicans.Methods Human neutrophils were isolated from peripheral blood by using density gradient centrifugation method,and cultured with the presence of the yeast phase of a Sporothrix Schenckii clinical isolate and a standard strain of Candida albicans (ATCC 90028)at a multiplicity of infection of 10 or 1 for 60-210 minutes.Subsequently,flow cytometry with ROS probe (2',7'-dichlorofluorescein diacetate,DCFH-DA) was carried out to for the real time detection of intracellular ROS level,confocal laser scanning microscopy (CLSM) for the observation of ROS distribution.In addition,the fungicidal efficiency of neutrophils against Sporothrix Schenckii and Candida albicans was estimated by the number of colonies after additional culture of neutrophil lysates on brain-heart infusion agar (BHIA) medium.Statistical analysis was done by using univariate analysis of variance and LSD-t test.Results The intracellular ROS level peaked at 60 minutes in neutrophils incubated with Sporothrix Schenckii yeast cells,then decreased rapidly from 60 minutes to 210 minutes.Compared with the neutrophils incubated with Candida albicans yeast cells,those with Sporothrix Schenckii yeast cells showed a higher ROS level (expressed as mean fluorescence intensity) at 60minutes (159.67 ± 11.34 vs.112.22 ± 9.66,P＜ 0.01),but a lower ROS level at 120 minutes (89.01 ± 9.81 vs.110.25 ± 7.28,P＜ 0.05) and 180 minutes (57.63 ± 8.46 vs.109.98 ± 9.00,P＜ 0.01).CLSM revealed that ROS was mainly distributed in neutrophils with phagocytized fungal spores,and especially on the surface of phagocytized spores.Furthermore,the percentage of yeast cells killed by neutrophils was significantly lower for Sporothrix Schenckii than for Candida albicans at 180 minutes (19.21％ ± 3.68％ vs.26.63％ ± 4.97％,P ＜ 0.01).Conclusions Differential expression of intracellular ROS was observed in neutrophils after phagocytosis of Candida albicans and Sporothrix schenckii.Neutrophils exert a stronger fungicidal activity against Sporothrix Schenckii in comparison with Candida albicans,which may be associated with the rapid decrease of ROS level in neutrophils after phagocytosis.

Objective To estimate the value of BIOMED-2 primers for the detection of T cell receptor γ (TCR-γ) gene rearrangements in different types of specimens from patients with mycosis fungoides (MF).Methods Totally,15 paraffin-embedded tissue specimens,14 fresh tissue specimens and 18 whole blood specimens were obtained from 28 patients with MF,and subjected to DNA extraction.BIOMED-2 multiplex PCR tubes TCRγ (A+B) were used for the analysis of TCRγgene rearrangements.Data were processed by SPSS 13.0 software,and statistical analysis was done by chi-square test and Fisher's exact probability test.Results TCR-γ gene rearrangements were detected in 3 paraffin-embedded tissue specimens,11 fresh tissue specimens and 12 blood specimens,with significant differences in the detection rate between the three samples (x2 =13.047,P ＜ 0.01).The fresh tissue samples showed a significantly higher detection rate than the paraffin-embedded tissue samples (X2 =12.523,P ＜ 0.01).The detection rate of TCRγgene rearrangements was 3/6 in paraffin-embedded tissue samples collected in 2011,significantly higher than that in the other 9 paraffin-embedded tissue samples collected before 2011 (Fisher's exact probability test,P =0.044),but similar to that in 14 fresh tissue specimens (12/14,Fisher's exact probability test,P =0.044).Decreased detection rate of TCRγ gene rearrangements was observed in blood samples compared with fresh tissue specimens,but no statistical difference was observed between the two types of specimens (x2 =2.358,P ＞ 0.05).Conclusions BIOMED-2 multiplex PCR tubes TCRγ(A+B) are suitable for the detection of clonal rearrangements of TCRγgene in different types of specimens,especially in fresh tissue specimens,from patients with MF.

Objective To investigate the mechanism of soluble β-1, 3-D-glucan in G-test positive serum in inhibiting ROS-dependent killing of Candida albicans ( C.albicans ) mediated by neutrophil Dectin-1.Methods The expression and distribution of internalized Dectin-1 and triggered ROS in human neutrophils were detected by using confocal/two-photon laser scanning microscopy upon stimulation with C.albicans (MOI=10) which was pretreated with β-1, 3-D-glucanase (10 U/ml) or not.Abrogation test was used to analyze whether intracellular Dectin-1 was involved in C.albicans-triggered ROS production in human neutrophils.Furthermore, flow cytometry analysis was performed to detect the expression of intracel-lular Dectin-1 and ROS in neutrophils which were pretreated respectively with G-test positive serum at differ-ent dilutions for 60 min and then stimulated with C.albicans for another 60 min at 37℃.Results After stimulated with C.albicans (MOI=10) for 60 min, the expression of Dectin-1 in neutrophils was recruited to the spores of opsonophagocytized C.albicans, and partly co-localized with the triggered ROS production . However, the expression of intracellular Dectin-1 was not observed in neutrophils when stimulated with β-1, 3-D-glucanase pretreated C.albicans for 60 min at 37℃.Abrogation test further showed that C.albicans-trig-gered ROS production in neutrophils was partly and irreversibly inhibited by adding Dectin -1 blocking mAb of 5 μg/ml.In addition , both the triggered expression of intracellular Dectin-1 and ROS production in neu-trophils stimulated with C.albicans ( MOI=10 ) in the presence of G-test positive serum were significantly lower than those of neutrophils stimulated only with C.albicans (LSD-t test, P<0.01).Linear regression a-nalysis suggested that the triggered intracellular Dectin-1 and ROS production in neutrophils upon stimulation with C.albicans were both inhibited by soluble β-1, 3-D-glucan in a dose-dependent manner (Dectin-1,R2=0.702,P<0.01;ROS,R2=0.588,P<0.01 ).Conclusion Taken together, these results demonstrated that the soluble β-1, 3-D-glucan in G-test positive serum may play a role in inhibiting the ROS-dependent killing of C.albicans by interfering with internalized expression of neutrophil Dectin-1.

Objective: To assess the specificity of white dermographism in atopic dermatitis (AD) , and to investigate the relationship between its duration and severity of AD. Methods: From 1^st to 30^th March 2018, 78 patients with AD (AD group) , 100 patients with non-AD skin diseases (non-AD group) and 100 healthy controls without skin diseases (control group) were enrolled from Department of Dermatology, Wuhan No.1 Hospital. Dermographism test was conducted in each subject, and the subjects′ response and duration of white dermographism were observed. Meanwhile, the severity of skin lesions of the AD patients was evaluated. Statistical analysis was carried out by using Chi-square test, analysis of variance and linear correlation analysis. Results: Of the 78 patients in the AD group, 67 (85.90%) were positive for white dermographism, and the positive rate of white dermographism was significantly higher in the AD group than in the non-AD group (18.00% [18/100], χ² = 80.97, P<0.017) and control group (5.00% [5/100], χ² = 119.05, P<0.017) . Additionally, the positive rate of white dermographism was significantly higher in the non-AD group than in the control group (χ² = 8.30, P<0.017) . The average duration of white dermographism was 1.89 minutes in 30 patients with mild AD, 2.74 minutes in 25 patients with moderate AD, and 4.41 minutes in 12 patients with severe AD. There was a significant difference in the duration of white dermographism among the 3 AD subgroups (F = 64.588, P<0.05) . Moreover, the severity of AD was positively correlated with the duration of white dermographism (r = 0.977, P = 0.136) . Conclusion White dermographism is a specific clinical manifestation of AD, and its duration is positively correlated with the severity of AD.