Objective: To investigate the expression of T ⁃cell immunoglobulin and immunoreceptor tyrosine⁃based inhibitory motif domain (TIGIT) and programmed death 1 (PD1) on follicular T helper cells (TFH) from the patients with rheumatoid arthritis (RA) and to analyze the clinical relevance to disease severity. Methods : The expression of TIGIT and PD1 on TFH cells were examined from 79 RA patients and 45 healthy controls (HC) by the technique of flow cytometry. The expression of TIGIT and PD1 including percentage of TIGIT + TFH , PD1 - TFH , TIGIT + PD1 - TFH , TIGIT + PD1 + TFH , TIGIT - PD1 + TFH cells and the mean fluorescence intensity (MFI) on theses TFH cells were compared between RA patients and HC. Moreover, its correlation with laboratory inspection was analyzed. Results: ① The percentage of PD1 + TFH and the MFI of PD1 on PD1 + TFH from RA patients significantly increased compared with HC (P < 0. 01) . ② The percentage of TIGIT + PD1 - TFH significantly decreased compared with HC (P < 0. 01) , whereas the percentage of TIGIT + PD1 - TFH , the MFI of PD1 on TIGIT + PD1 -TFH , the percentage of TIGIT + PD1 + TFH and the MFI of PD1 on TIGIT + PD1 + TFH significantly increased (P <0. 05) . ③ The MFI of PD1 on TIGIT + PD1 + TFH in RA patients was positively associated with rheumatoid factors (RF) (P < 0. 05) . The percentage of TIGIT - PD1 + TFH , the MFI of PD1 on TIGIT - PD1 + TFH , the MFI of PD1 on TIGIT + PD1 + TFH in RA patients were positively associated with increased RF (P < 0. 05) . ④ The percentage of TIGIT + PD1 - TFH in RA patients was negatively associated with erythrocyte sedimentation rate ( ESR) ( P <0. 05) . The percentage of TIGIT - PD1 + TFH in RA patients was positively associated with ESR (P < 0. 05) . The percentage of TIGIT + PD1 + TFH in RA patients was negatively associated with lymphocyte count ( Lym ) ( P <0. 05) . The MFI of PD1 on TIGIT + PD1 + TFH in RA patients was negatively associated with lymphocyte percentage (Lym% ) , and positively associated with neutrophil count ( Net ) , neutrophil percentage ( Net% ) , neutrophiltolymphocyte ratio (NLR) , systemic immune inflammation index (SII) , derived neutrophil to lymphocyte ratio (dN⁃LR) , ESR , C ⁃reactive protein (CRP) ( P < 0. 05) . The MFI of PD1 on TIGIT - PD1 + TFH in RA patients was positively associated with Net , ESR , CRP ( P < 0. 05 ) . ⑤ The percentage of TIGIT + PD1 - TFH in RA patients was negatively associated with disease activity score 28 (DAS28) ⅣESR (P < 0. 05) . The MFI of PD1 on TIGIT + PD1 + TFH and the MFI of PD1 on TIGIT - PD1 + TFH in RA patients was positively associated with DAS28⁃ESR ,DAS28 ⁃CRP , tender joint count (TJC) , swollen joint count (SJC) , patient visual analogue scale ( VAS) ( P <0. 05) . ⑥ Decreased percentage of TIGIT + PD1 - TFH and increased percentage of TIGIT + PD1 + TFH were risk of RA. Conclusion The decreased percentage of TIGIT + PD1 - TFH , increased percentage of TIGIT + PD1 + TFH and TIGIT - PD1 + TFH were observed in patients with RA. Aberrations level of TIGIT + PD1 - TFH , TIGIT + PD1 + TFH , TIGIT - PD1 + TFH were associated with the production of antibody , inflammation , and also disease activity.

Objective : To investigate the expression of programmed death 1 (PD1) on CXCR5 - CD4 + T cells from the patients with systemic lupus erythematosus ( SLE) and to analyze the clinical relevance to disease severity. Methods : The expression of PD1 on CXCR5 - CD4 + T cells was examined from 47 SLE patients and 35 healthy controls (HC) by the technique of flow cytometry. The expression of PD1 including percentage of PD1 + CXCR5 -CD4 + T cells and mean fluorescence intensity (MFI) on CXCR5 - CD4 + T cells was compared between SLE patients and HC. And its correlation with clinical indicators , laboratory inspection and the percentage of plasmablasts was analyzed. Moreover, the predictive value of the expression of PD1 on CXCR5 - CD4 + T cell was explored. Results: ① The percentage of PD1 + CXCR5 - CD4 + T cells from SLE patients significantly elevated compared with HC (P= 0. 008 3 , U = 540. 5) , and the MFI of PD1 on CXCR5 - CD4 + T cells from SLE patients significantly elevated compared with HC (P < 0. 000 1 , U = 187. 0) . ② The percentage of PD1 + CXCR5 - CD4 + T cells was associated with C3 ( rs = - 0. 335 2 , P = 0. 022 8) , anti - SSA (P = 0. 016 6 , t = 2. 5) , anti - histone (P = 0. 030 3 , t =2. 3) , treatment (P = 0. 020 2 , t = 3. 4) , plasmablasts ( rs = 0. 387 1 , P = 0. 0072) in SLE patients. The MFI of PD1 on CXCR5 - CD4 + T cells was associated with SLEDAI ( rs = 0. 403 1 , P = 0. 005 0) , ESR ( rs = 0. 356 1 , P= 0. 017 7) , CRP ( rs = 0. 337 4 , P = 0. 028 9) , RBC ( rs = - 0. 297 0 , P = 0. 042 6) , HGB ( rs = - 0. 302 9 , P= 0. 038 5) , HCT ( rs = - 0. 381 6 , P = 0. 008 1) , L ( rs = - 0. 393 7 , P = 0. 006 2) , L% ( rs = - 0. 391 2 , P= 0. 006 5) , N% ( rs = 0. 315 0 , P = 0. 031 1) , NLR ( rs = 0. 373 0 , P = 0. 009 8) , LMR ( rs = - 0. 431 5 , P =0. 002 5) , dNLR ( rs = 0. 315 0 , P = 0. 031 1) , anti⁃Ro52 (P = 0. 029 5 , t = 63. 5) , treatment (P = 0. 035 5 , W= 21) , plasmablasts ( rs = 0. 315 8 , P = 0. 030 6) . ③ Logistic regression analysis showed that the MFI of PD1 on CXCR5 - CD4 + T cells was a risk factor for SLE. ④ ROC analysis showed the AUC of the MFI of PD1 on CXCR5 -CD4 + T cells was 0. 886. A further established model based on combination of the MFI of PD1 on CXCR5 - CD4 + T cells and HGB showed predictive value in distinguishing SLE from HC with AUC of 0. 979. And predictive value was positively associated with SLEDAI ( rs = 0. 313 6 , P = 0. 030 3) . Conclusion Increased percentage of PD1 + CXCR5 - CD4 + T cells and increased MFI of PD1 on CXCR5 - CD4 + T cells in SLE are associated with disease severity and activity , and a model based on combination of the MFI of PD1 on CXCR5 - CD4 + T cells and HGB shows prominent value for predicting SLE.

Objective:To evaluate the association of maternal nutrition status in the first trimester with gestational diabetes mellitus (GDM) and macrosomia.Methods:378 pregnant women who took prenatal care in Shunyi Women′s and Children′s Hospital of Beijing Children′s Hospital were enrolled in the study. Blood samples were collected at first prenatal visit (<12 gestation weeks) to measure the level of hemoglobin and iron status indexes including serum iron, ferritin, transferrin, total iron binding capacity, iron saturation, transferrin saturation. The incidence of GDM and macrosomia were collected and Logistic regression was used to evaluate the associations of maternal nutrients status in the first trimester with GDM and macrosomia.Results:The incidence rate of GDM was16.9%,the incidence of anemia and iron deficiency in the first trimester were2.4% and 2.5%, respectively. After adjustment for variables such as maternal age, pre-pregnancy BMI, family history of diabetes, and parity, Logistic regression showed that in the first trimester, iron saturation>50% ( OR=0.238, 95% CI 0.068-0.831), transferrin saturation>50% ( OR=0.08, 95% CI 0.010-0.677) were protective factors of GDM; iron saturation 25%-50% ( OR=0.361, 95% CI 0.143-0.908); transferrin saturation 25%-50% ( OR=0.383, 95% CI 0.165-0.891); ferritin>30 ng/ml ( OR=0.418, 95% CI0.186-0.939) were protective factors of macrosomia. Conclusion:Maternal iron status in the first trimester might be associated with GDM and macrosomia. Thus, maternal iron status assessment in the first trimester is necessary.

Objective : To analyze circular RNA (circRNA) expression profiles in oral squamous cell carcinoma (OSCC) and its clinical significance. Methods : The expression of circRNA was detected with circRNA microarray assay in three samples of OSCC tumor and matched adjacent tissues. Quantitative real-time PCR (RT-qPCR) was used to verify the expression of circRNA in 45 pair OSCC tissues and normal adjacent tissues. The relationship between the expression of circRNA and the clinicopathological characteristics of OSCC was analyzed. circRNAs/miRNAs interaction were predicted using Arraystar' s home-made miRNA target prediction software. Results : 155 circRNAs were differentially expressed between the OSCC tissues and matched adjacent tissues, of which 45 circRNAs were up-regulated and 110 circRNAs were down-regulated in OSCC tissues (fold changes ≥1.5 and P < 0.05). In the selected three circRNAs that were most significantly upregulated or downregulated in OSCC, the RT-qPCR results showed that hsa_circ_0001874, hsa_circ_0001971 and has_circ_0067934 were increased, while hsa_circ_0000520, hsa_circ_0023944 and hsa_circ_0000140 were decreased in OSCC tissues versus normal adjacent tissues (P < 0.001). The results were generally consistent with the microarray data. Among the circRNA expression profiles in OSCC, the up-regulation of hsa_circ_0001874 was the highest and the expression of hsa_circ_0001874 was significantly correlated with TNM stage and tumor grade. The result of Arraystar' s home-made miRNA target prediction software indicated that miR-103a-3p, miR-107, miR-593-5p, miR-661 and miR-662 may be potential target genes of hsa_circ_0001874. Conclusion The differentially expressed circRNAs in OSCC tissues and normal adjacent tissues were identified, and these dysregulated circRNAs and their potential target gents may play important roles in the development of OSCC.

Objective: To explore the expression and clinical significance of circular RNA circHIPK3 in oral squamous cell carcinoma (OSCC), analyze the effect of circHIPK3 on the proliferation of OSCC cells. Methods: The expression of circHIPK3 in OSCC tissues, adjacent non-cancerous tissues and OSCC cell lines were detected by quantitative real-time polymerase chain reaction (qPCR). The correlations between the expression of circHIPK3 in OSCC tissues and the clinicopathological features were analyzed as well. circHIPK3-specific siRNA si-circHIPK3 and negative control siRNA si-NC were designed and synthesised and used to transfect CAL27 and SCC15 cells respectively. The proliferation capacity of CAL27 and SCC15 cells after transfection with si-circHIPK3 was detected by CCK-8 assay. The expression of miR-124 in OSCC was detected by qPCR, and the correlation between expression of circHIPK3 and the expression of miR-124 was analyzed. Using qPCR to detect the expression of miR-124 in CAL27 and SCC15 cells after transfection with si-circHIPK3 and si-NC respectively. Furthermore, using CCK-8 assay to detect the proliferation capacity of CAL27 and SCC15 cells after transfection with si-NC, si-circHIPK3, miR-124 mimic, si-circHIPK3+miR-124 inhibitor. Results: The expression of circHIPK3 in OSCC tissues [2.23 (1.86, 3.00)] was significantly higher than that of the adjacent non-cancerous tissues [1.05 (0.85, 1.26)] (U=1 094, P=0.000). The expression of circHIPK3 in CAL27 (3.02±0.51) and SCC15 cells (3.16±0.75) was higher than those of human normal oral keratinocytes (hNOK) (1.26±0.30) (P=0.000). The expression of circHIPK3 was found to be closely associated with TNM stage (P<0.05) and tumor grades (P<0.05). Knockdown of circHIPK3 can inhibit proliferation of CAL27 and SCC15 cells (P<0.05). The expression of miR-124 in OSCC tissues (0.61±0.35) was significantly lower than that in adjacent non-cancerous tissues (1.13+0.39) (t=-5.36, P<0.05). Correlation analysis showed that the expression of circHIPK3 in OSCC was negatively correlated with the expression of miR-124 (r=-0.767, P<0.001). Moreover, down-regulation of miR-124 rescued the phenotype induced by knockdown of circHIPK3. Conclusions The expression of circHIPK3 in OSCC was increased, and silencing of circHIPK3 expression can inhibit the proliferation of OSCC cells. Our results suggest that circHIPK3 may play a key role in the occurrence and development process of OSCC through the regulation of miR-124 expression.

Objective Detecting plasma level of circular RNA(circRNA)hsa_circ_0009024 in pulmonary tuberculosis(TB)patients,and evaluating its diagnostic value for TB.Methods From January 2016 to December 2016, a hosptial-based, case-control study was performed, which include 90 untreated active pulmonary tuberculosis patients(TB group),75 healthy people(healthy control)and 84 patient with other diseases(other disease group).Plasma level of circRNA hsa_circ_0009024 was detected by real-time quantitative polymerase chain reaction.Furthermore, the 90 patients with TB were divided into different subgroups according to cavity formation and the lung fields involvement: patients without lung cavity(55 cases)vs those with lung cavity(35 cases),patients with involvement of <2 lung fields(49 cases)vs≥2 lung fields(41 cases).Plasma levels of hsa _circ_0009024 of 41 TB patientswere monitored andcomparedbefore and after 3 months anti-TB therapy.The sensitivity and specificity of plasma hsa_circ_0009024 were analyzed by using the receiver operating characteristic(ROC)curve.The comparison between two groups was performed with Mann-Whitney U test and the comparison among multigroupswas conducted with Kruskal-Wallis H test.Results Plasma levels of hsa_circ_0009024 in TB patients[1.98(1.42, 2.71)]were significantly higher than healthy controls[1.03(0.78,1.33)]and other disease groups[1. 13(0.77,1.51)](H=76.58,P<0.0001).Plasma levels of hsa_circ_0009024 in cavity pulmonary TB patients were higher than pulmonary TB patients without cavity(U=392.50,P<0.0001).Plasma levels of hsa_circ_0009024 in TB patients with involvement of ≥2 lung fields were higher than <2 lung fields(U=590.50,P=0.0008).As compared to pre -treatment[2.01(1.41, 2.71)], the plasma hsa_circ_0009024 levels decreased significantly in 3 months[1.22(0.85,1.47)](U=292.50,P<0.0001)after anti-TB therapy.The area under the receiver operating characteristics curve(AUC)of plasma hsa_circ_0009024 in discriminating the patients with TB from normal controls, pneumonia patients and lung cancer patients were 0.841 and 0.811, respectively.Conclusion The hsa_circ_0009024 can be used as a potential biomarker in TB diagnosis and monitoring.

Objective To investigate the expression of T cell immunoreceptor with Ig and immunoreceptor tyrosine-based inhibitory domains (TIGIT) on peripheral CD4+ T and CD8+ T cells from patients with rheumatoid arthritis (RA) and its significance in order to clarify its role in the development of RA.Methods Peripheral blood samples were collected from 81 patients with RA and 33 healthy controls (HC).Expression of TIGIT on the surface of peripheral blood leukocytes was detected by flow cytometry.Differences in TIGIT expression between RA and HC groups were comparatively analyzed.Correlations of TIGIT expression on CD4+ T and CD8+ T cells with several laboratory indexes were analyzed.All data were statistically analyzed.Results (1) The percentage of TIGIT-expressing CD3+ T cells in patients with RA was significantly higher than that in HC (P<0.01).Moreover, the median fluorescence intensity (MFI) of TIGIT on CD3+ T cells was significantly elevated in patients with RA as compared with that in HC (P<0.01).No significant difference in the expression of TIGIT on B cells, monocytes or neutrophils was observed between RA and HC groups.(2) The percentages of TIGIT-expressing CD4+ T and CD8+ T cells were significantly elevated in patients with RA as compared with those in HC (P<0.01).Moreover, the MFI of TIGIT on CD4+ T and CD8+ T cells were significantly elevated in patients with RA as compared with those in HC (P<0.01).(3) The percentages of both TIGIT-expressing CD4+ T cells and TIGIT-expressing CD8+ T cells in patients with RA were positively correlated with erythrocyte sedimentation rate (ESR) (rs=0.355, P<0.01;rs=0.277, P=0.013).(4) The percentages of both TIGIT-expressing CD4+ T cells and TIGIT-expressing CD8+ T cells in patients with RA were positively correlated with rheumatoid factor (RF) (rs=0.265, P=0.017;rs=0.366, P<0.01).The MFI of TIGIT on CD4+ T cells was positively correlated with RF in RA group (rs=0.226, P=0.043).The percentage of TIGIT-expressing CD4+ T cells was positively correlated with anti-cyclic citrullinated peptide (anti-CCP) antibody in patients with RA who were positive for anti-CCP antibody (rs=0.324, P=0.012).(5) The percentage of TIGIT-expressing CD4+ T cells as well as the MFI of TIGIT on CD4+ T cells in patients with RA was positively correlated with Disease Activity Score-28 (DAS28) (r=0.232, P=0.038;r=0.343, P<0.010).Conclusion The expression of TIGIT on T cells is elevated in patients with RA and correlated with inflammatory markers, antibody production and disease activity.

Objective To detect the serum level of long non-coding RNA (lncRNA ) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in pulmonary tuberculosis (TB) patients ,and to evaluate its diagnostic value .Methods The expression of serum MALAT1 in 56 hospitalized TB patients , 35 latent TB infection (LTBI) individuals and 40 healthy controls were detected by real-time quantitative PCR .Serum levels of MALAT1 before and 3 ,6 months after anti-TB therapy in 16 TB patients were determined .Receiver operating characteristic (ROC) curve analysis was used to evaluate the sensitivity and specificity of serum MALAT 1 .The comparison between two groups was performed by Student t-test , and the comparison among three groups was performed with one-way analysis of variance test .Results Serum level of MALAT1 in TB patients was (2 .10 ± 1 .05) ,which was significantly higher than those in LTBI individuals (1 .16 ± 0 .51) and healthy controls (1 .02 ± 0 .44 ,F= 28 .53 ,P< 0 .01) .The MALAT1 level in TB patients with positive sputum smear was significantly higher than that in patients with negative sputum smear (2 .42 ± 1 .03 vs 1 .43 ± 0 .74 ,t= 2 .66 ,P< 0 .01) .Compared with pre-treatment (2 .28 ± 0 .79) ,the serum MALAT 1 levels decreased significantly in 3 months (1 .35 ± 0 .39) and 6 months (1 .05 ± 0 .30) after anti-TB therapy (t= 4 .33 ,6 .05 ;both P< 0 .01) .The area under the curve (AUC) of serum MALAT1 was 0 .821 , with sensitivity and specificity of 0 .732 and 0 .850 , respectively .Conclusion The expression of MALAT1 is up-regulated in TB patients , and could be used as potential novel biomarkers for the diagnosis of TB .

Objective To analyze the expression profile of long non-coding RNA (lncRNA) in the lipopolysaecharide (LPS)-induced inflammation of monocyte-derived macrophages.Methods Peripheral blood mononuclear cells were derived from healthy donor and induced into macrophages. The macrophages were divided into blank control group and LPS (1 mg/L) stimulated 12 hours group. Culture supernatants and cell pellets were harvested in each group, enzyme linked immunosorbent assay (ELISA) was used to assay the production changes of interleukins (IL-1β and IL-6), and tumor necrosis factor-α (TNF-α) in the supernatant. The technique of lncRNA microarray was used to test the lncRNA expression profile in LPS-induced inflammation of macrophages and control macrophages. The raw data of lncRNA were pretreated for normalization. Five lncRNA expressions were validated by real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Furthermore, qRT-PCR was used to detect the expression of NR_028034 in macrophages after LPS-induced inflammation.Results ① The contents of IL-1β (ng/L:562.93±61.17 vs. 59.74±15.68), IL-6 (ng/L: 702.46±92.31 vs. 71.66±18.25) and TNF-α (ng/L: 794.50±63.89 vs. 85.12±22.07) in the LPS group were significantly higher than those in the blank control group (allP < 0.01). These results indicated that the inflammatory model of human macrophages was constructed successfully. ② Compared with blank control group, and 1479 lncRNA which have more than 2 folds variation and significant difference (P < 0.05) by statistical analysis was defined as lncRNA with differential expression. Among these lncRNA, LPS group showed 953 up- regulated and 526 down- regulated genes by 2 folds and 49 up- regulated and 35 down- regulated genes by 5 folds. ③ qRT-PCR results were generally consistent with the microarray data. ④ The expression of NR_028034 was increased by (4.41±0.65), (11.56±2.04), (18.58±1.36) folds compared with blank control group at 3, 6, 12 hours after LPS stimulation (allP < 0.01).Conclusions These data show a significantly altered lncRNA expression profile in the LPS-induced inflammation of monocyte-derived macrophages, suggesting that lncRNA may be involved in regulation of macrophages inflammatory response.

Objective To analyze the expression profile of circular RNA (circRNA) in LPS-treated murine peritoneal macrophages (MPMs) and to investigate the effects of mmu_circ_0000790 (circ790) on the secretion of pro-inflammatory cytokines in MPMs following LPS stimulation.Methods MPMs were isolated from C57BL/6 male mice and then stimulated with (LPS treatment group) or without (blank control group) 1 mg/L of LPS for 12 hours.Supernatants of cell culture and cell pellets were collected from each group.Enzyme linked immunosorbent assay (ELISA) was used to measure the changes in the secretion of IL-1β,IL-6 and TNF-α in the supernatants.Expression profile of circRNA in macrophages from the two groups (n=3) was analyze by circRNA microarray.Some differentially expressed circRNAs were validated by real-time PCR (RT-PCR).Moreover,RT-PCR was also performed to detect the expression of circ790 in MPMs after LPS stimulation.Small interfering RNA (siRNA) oligo specific for circ790 was designed and synthesized.Then the synthesized siRNA oligo and normal control (NC) oligo were transfected into MPMs by LipofectamineTM2000.The transfected MPMs were treated with LPS.ELISA was used to detect the levels of IL-1β,IL-6 and TNF-α in the supernatants.Arraystar's home-made microRNA target prediction software was used to predict circ790/microRNA.Results The concentrations of IL-1β,IL-6 and TNF-α in the LPS treatment group were significantly higher than those in the blank control group (P<0.01).These results indicated that the inflammatory model of MPMs was successfully constructed.Statistical differences in 34 differentially expressed circRNAs were found between the two groups (P<0.05).Among them,12 circRNAs were up-regulated and the other 22 circRNAs were down-regulated in the LPS treatment group.RT-PCR results were generally consistent with the microarray data.The expression of circ790 was increased by (1.94±0.15),(3.18±0.13) and (4.21±0.22) folds as compared with that of the blank control group at 3,6 and 12 h after LPS stimulation (P<0.05).After transfecting the MPMs with circ790-siRNA,the secretion of IL-6 was significantly decreased (P<0.05) without notable influence on the secretion of IL-1β and TNF-α.circ790 could function as a microRNA sponge to regulate the gene expression.Conclusion These data show a significantly altered circRNA expression profile in the LPS-treated MPMs.circ790 may be involved in the regulation of IL-6 secreted by macrophages.