Objective To investigate the clinical application value of fluorescence PCR in vitro diagnosis(IVD)reagent kits in the HLA-B27 detection by comparing 2 kinds of HLA-B27 IVD reagent kit approved by CFDA.Methods A total of 573 clinical blood samples were collected and detected for HLA-B27 by the approved reagent kits based on the fluorescence PCR technique and the flow cytometry.The samples with inconsistent testing results by the two kits were further confirmed by the PCR sequencing.At the same time,about 5% samples of the positive results detected by the fluorescence PCR method were extracted for conducting the re-testing.Results Among 573 samples,191 samples were HLA-B27 positive and 382 cases were HLA-B27 negative by flow cytome-try;the same samples had 194 cases of HLA-B27 positive and 379 cases of HLA-B27 negative by real-time PCR.With flow cytome-try as reference of the final results,the positive coincidence rate of the two kinds of kit was 96.33%(184/191),the negative coinci-dence rate was 94.76%(362/382),27 samples had inconsistent results from the two kinds of assay(accounting for 4.71% of the to-tal number of samples),the total coincidence rate was 95.29% [(184+362)/573],the Kappa value was 0.896(P =0.02);the chi-square test P =0.021,the two kinds of testing method had the high consistency,but the differences existed in the testing results. The re-testing results by PCR sequencing(including 27 samples with inconsistent results by two kinds of kit)were entirely consist-ent with the fluorescence PCR testing results.Conclusion Compared with the authority method flow cytometry for HLA-B27 tes-ting in clinic,the fluorescence PCR kit may present more accurate judging ability for the HLA-B27 testing on the basis of ensuring the higher consistency of the testing results,is easier compared with the sample preparation and operating procedures,and has the stronger clinical application value and prospects s.

Objective To investigate the relationship of TCM constitution and syndrome elements based on patients with hyperhomocysteinemia. Methods A survey was conducted among 1316 patients with diabetes, coronary disease, hypertension and cerebral infarction. Totally 326 patients with hyperhomocysteinemia were incorporated in diagnostic data and their constitutions were determined. At the same time, syndrome elements identification method for syndrome elements was applied for judgment, and consistency of the constitution and syndrome elements was analyzed. Results The survey showed that prevalence rate of population with hyperhomocysteinemia was 24.77%;there were 6 cases with mild nature, and 320 cases with biased constitution;phlegm dampness, blood stasis, qi deficiency of constitutions 110 cases, 73 cases and 59 cases, respectively;blood stasis, phlegm, qi deficiency, and dampness were the main syndrome elements of hyperhomocysteinemia, of 100 cases, 98 cases, 73 cases and 59 cases, respectively;the consistency of phlegm dampness constitution and syndromes of phlegm was the most obvious (Kappa=0.89, P<0.05). Conclusion Phlegm dampness, blood stasis, and qi deficiency were the most common constitutions of hyperhomocysteinemia;blood stasis, phlegm, qi deficiency and dampness were the main syndrome elements of hyperhomocysteinemia patients. The two results were consistent, and the consistency of phlegm syndrome and phlegm dampness constitution was the most obvious.

Objective To investigate the changes in cathepsin B (CatB) expression in acutely photodamaged human dermal fibroblasts (HDFs) and their significance.Methods HDFs were isolated from the foreskin of children,and subjected to primary culture and subculture.The fourth-to eighth-passage HDFs were used in the following experiment.HDFs were divided into two groups to receive irradiation with different doses of ultraviolet A (UVA) for different durations (acutely photodamaged group) or remain unirradiated (control group).Cell counting kit-8 (CCK8) assay was conducted to evaluate the proliferative activity of HDFs after irradiation with UVA at 5,10,15,20 and 25 J/cm2 respectively.Western blot and quantitative real-time reverse transcription PCR were performed to measure the protein and mRNA expressions of CatB respectively in HDFs at 24,48 and 72 hours after exposure to UVA at 10 J/cm2,and at 48 hours after exposure to UVA at 10,15,20 and 25 J/cm2.Statistical analysis was carried out by analysis of variance and least significant difference (LSD) test using the SPSS 13.0 software.Results UVA radiation induced a decrease in the proliferative activity of HDFs.When the dose of UVA was ≤ 10 J/cm2,the survival rate of HDFs maintained higher than 85％,and significant differences were observed in cell survival rate between unirradiated and irradiated HDFs at 24,48 and 72 hours (all P ＜ 0.05).Western blot showed that the gray value of CatB protein in the acutely photodamaged group irradiated with 10 J/cm2 UVA was significantly higher than that in the control group at 24 hours (0.76 ± 0.14 vs.0.35 ± 0.01,P ＜ 0.05),48 hours (1.34 ± 0.38 vs.0.45 ± 0.12,P＜ 0.05) and 72 hours (0.82 ± 0.09 vs.0.61 ± 0.06,P＜ 0.05).Increased mRNA expressions of CatB were also observed in the acutely photodamaged group compared with the control group at 24 hours (0.149 ± 0.009 vs.0.089 ± 0.015,P ＜ 0.05),48 hous (0.173 ± 0.009 vs.0.091 ± 0.010,P ＜ 0.05) and 72 hours (0.185 ± 0.158 vs.0.111 ± 0.017,P ＜ 0.05) after UVA radiation at 10 J/cm2.The gray value of CatB protein was 0.99 ± 0.07,1.49 ± 0.14,1.89 ± 0.08,2.07 ± 0.06 in HDFs at 48 hours after exposure to UVA of 10,15,20 and 25 J/cm2,respectively,significantly higher than that in the control group (0.60 ± 0.05,all P ＜ 0.05).Similarly,the mRNA expression of CatB was up-regulated in HDFs at 48 hours after UVA radiation at 10,15,20 and 25 J/cm2 compared with the unirradiated HDFs.Conclusion The protein and mRNA expressions of CatB are up-regulated in acutely photodamaged HDFs induced by UVA radiation.

Objective To investigate the role of Toll like receptor 4/nuclear factor-κB (TLR4/NF-κB) signal pathway on myocardial dysfunction after cardiac arrest-cardiopulmonary resuscitation (CA-CPR) in animal model. Methods Twenty-six pigs were randomly divided into sham group (n = 6), CA-CPR 12 hours group (n = 10) and CA-CPR 24 hours group (n = 10). The model of CA-CPR was reproduced by endocardial electrical stimulation for 8 minutes followed by CPR, and the pigs in sham group were only given anesthesia and tracheal intubation. The changes in hemodynamics including mean arterial pressure (MAP) and cardiac output (CO), as well as morphology and ultrastructure of myocardial cells were observed before and after CPR. The levels of serum tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were determined by enzyme linked immunosorbent assay (ELISA), and protein and mRNA expressions of TLR4/NF-κB in the myocardium were determined by Western Blot and real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-PCR), respectively. Results Hemodynamic disturbance and myocardial serious injury were observed in CA-CPR groups. Compared with sham group, the levels of serum TNF-α were markedly increased 0.5 hour after return of spontaneous circulation (ROSC) in CA-CPR 12 hours and 24 hours groups (pg/L: 62.49±6.66, 48.39±2.37 vs. 10.75±0.74, both P < 0.05), and peaked at 2 hours (pg/L: 70.93±5.51, 66.03±2.60 vs. 10.87±0.91, both P < 0.05) followed by a gradual decline. The levels of serum IL-6 at 0.5 hours after ROSC in CA-CPR 12 hours and 24 hours groups were markedly higher than those of sham group (pg/L: 14.42±1.99, 11.23±1.12 vs. 8.75±0.74, both P < 0.05), and peaked at 12 hours (pg/L: 36.50±2.91, 38.15±1.26 vs. 8.88±0.62, both P < 0.05) followed by a gradual decline. The protein expressions of TLR4 and NF-κB in the myocardium were significantly increased in CA-CPR 12 hours and 24 hours groups as compared with sham group [TLR4 protein (gray value): 0.11±0.03, 0.24±0.05 vs. 0.05±0.02; NF-κB protein (gray value): 0.27±0.04, 0.24±0.03 vs. 0.09±0.02, all P < 0.05]. The mRNA levels of TLR4 in CA-CPR 12 hours and 24 hours groups were increased by approximately (9.93±1.07) folds and (9.21±1.27) folds of sham group respectively, and NF-κB mRNA expressions were increased by (4.44±0.96) folds and (6.09±0.81) folds of sham group respectively (all P < 0.01). Conclusion Activation of TLR4/NF-κB signal pathway may be one of the main pathological mechanisms of post resuscitation myocardial injury in a porcine model of CA-CPR.

BACKGROUND:Animal experiments and clinical studies have shown that, functional deprivation at the early development stage may possibly cause nerve and cognition dysfunction at adulthood. However, no research focuses on the effect of olfactory function on the neurogenesis in piriform cortex of adult guinea pigs.
OBJECTIVE:To establish olfactory deprivation models and explore the effect of olfactory experience on the distribution of doublecortin, parvalbumin, glutamic acid decarboxylase 67 and the co-localization of doublecortin with neuron-specific nuclear-binding protein in piriform cortex of adult guinea pigs.
METHODS:Twenty guinea pigs were randomly divided into two groups. Olfactory deprivation models were established through unilateral naris-occlusion and the occluded animals were al owed to survive 3 and 6 weeks, then the specimens were harvested. The distribution of doublecortin, parvalbumin, glutamic acid decarboxylase 67 was detected with immunohistochemistry. The co-localization of doublecortin with neuron-specific nuclear-binding protein in piriform cortex of adult guinea pigs was determined with immunofluorescence method.
RESULTS AND CONCLUSION:The number of doublecortin, parvalbumin, glutamic acid decarboxylase 67 positive cel s in piriform cortex at the undeprived side was significantly higher than that at the deprived side at 3 and 6 weeks (P<0.05). There was no significant difference in the co-localization of doublecortin with neuron-specific nuclear-binding protein in piriform cortex at 3 weeks group and 6 weeks group (P>0.05). The findings suggest that olfactory experience promotes the maturation and differentiation of neurons in piriform cortex of adult guinea pigs.

Objective To examine whether IFN-λ has the ability to inhibit HIV-1 infection of blood monocyte-derived macrophages and its mechanism(s). Methods Macrophages were pretreated with IFN-λ/ IFN-λ2 for 24 h before infected by HIV-1 R5 strains (Bal, Jago, and JRFL). And then the culture supernatants were detected HIV-1 reverse transcription (RT) activity and p24 protein expression by HIV-1 BT assay and ELISA. The expressions of IFN-λ receptor, CD4, CCRS, CXCR4 were evaluated by real-time PCR. Results Both IFN-λ1 and IFN-λ2, when added to macrophage cultures, inhibited HIV-1 infection and replication. This IFN-λ-mediated anti-HIV-I activity is broad, as IFN-λ could inhibit infection by both laboratory-adapted and clinical strains of HIV-1. Investigations of mechanism(s) responsible for the IFN-λ action showed that although IFN-λ had little effect on HIV-1 entry receptor CD4 and co-receptor CCR5 and CXCR4 expression, IFN-λ inhibited HIV-I infection of macrophages through connecting with IFN-λ recep-tor. Conclusion IFN-λ could inhibit HIV-I replication in macrophages. These findings indicate that IFN-λ may have a therapeutic value in the treatment of HIV-1 infection.

Objective To study the effect of inosine monophosphate dehydrogenase inhibitor (IMPDHI) on chemotaxis, migration and endocytosis of human peripheral myeloid dendritic cells (MDCs). Methods Freshly isolated peripheral blood mononuclear cells(PBMC)collected from healthy volunteers (N=15) and the study group were treated with IMPDHI. CC chemokine receptors on MDCs were analyzed by flow cytometry. The study group, control group and different chemokines were added via trans-well approach for different chemokines, stained by Lin-1/CD11c/HLA-DR and counted by flow cytometry. The migration index was calculated as a percentage of MDC migrated in response to the tested chemokine. After isolation of blood dendritic cell antigen-1+ (BDCA-1+ ), mannose receptor-mediated endocytosis was measured as the cellular uptake of FITC-dextran by the flow cytometry. Results (1) Compared to the control group, the expression of CCR1 in the study group was up-regulated significantly(17.02±3.23～30.63±9.13, P＜0.05) and the expressions of CCR3(10.26±2.25～5.81±0.97 P＜0.05) and CCR7 (9.56± 1.84～5.18±0.60 P＜0.05)were downregulated significantly. MDCs in the study group showed enhanced migratory response to inflammatory chemokine CCL2, CCL3, CCL4, CCL7 and CXCL12 (P＜0.05). (2)The endocytosis capacity in the study group was significantly higher than that in control group (P ＜0.05). Conclusion IMPDHI enhances the endocytotic capacity of MDCs and impairs the migratory response of peripheral MDCs to lymphocytic tissue by up-regulating the expression of chemokine receptor in MDCs and enhancing migratory response to inflammatory chemokines.

Adult ; Cryptococcosis ; Cryptococcus neoformans ; Humans ; Lung Diseases, Fungal ; Male

Objective To explore the abnormally functional brain region in resting state in first-episode major depression disorder patients with function magnetic resonance imaging.Methods 51 patients diagnosed with first-episode major depression disorder according to DSM-Ⅳ and 50 gender-,age-,and education-matched healthy controls completed resting state fMRI scan.The severity of depression,and unpredicted homodynamic responses across the whole brain were analyzed using Hamilton depression scale,regional homogeneity and amplitude of low-frequency fluctuation,respectively.Results Compared with control group,the right medial frontal gyms (BA6,MNI:3,-3,63,K=34) and left medial frontal gyrus(BA9,MNI:-9,36,30,K=10) (P＜0.001,uncorrected) in the case group showed higher regional homogeneity,with statistical significance.Compared with control group,right medial frontal gyrus (BA6,MNI:3,-3,63,K =35) and right posterior cingulated gyrus (BA31,MNI:3,-36,36,K =11) (P＜ 0.001,uncorrected)in the case group showed higher amplitude of low-frequency fluctuation,with statistical significance.Conclusion First-episode major depression disorder patients in resting state had several abnormally functional brain regions,which might be related to the pathological mechanism of depression disorder.