Objective To discuss application value for detecting sentinel lymphatic node(SLN) in patients with breast mass by subcutaneous injection SonoVue. Methods Forty-two cases with breast masses who were detected by conventional techniques were injected subcutaneously with 1.25 ml Sono Vue to the borderline of breast tumor at each 3,6,9, 12 points. The changes of contrast-enhanced ultrasound (CEUS) of the ipsilateral axillary SLN were observed and the start time of enhancement of lymphatic channel and lymph nodes, the position and numbers of enhancement of the lymph nodes, the time of regression were recorded. Compared with Methylene blue and surgicalbiopsy, the accuracy of detection of SLN by CEUS were evaluated. Results SLN were successfully detected in 30 of 42 patients, the detection rate was 71.4%. The total number of detected SLN was 40.Five nodes were detected in 5 of 10 (50%)patients with benign masses. Thirty-five nodes were detected in 25 of 32 (78.1%) patients with breast cancer. Fifty-seven SLN were successfully detected in 39 of 42 patients with Methylene blue, the accurate rate was 92.9%. Seven of 10 patients with benign masses and all 32 patients with breast cancer were Methylene blue positive. There were total 64 SLN found, including 11 with benign masses and 53 with lymph nodes metastasis of breast cancer cells. Conclusions The subcutaneous injection SonoVue is effective to detect SLN in patients with breast mass and apply clinical value for forecasting whether the lymph node metastases in patients with breast cancer.

Objective To investigate the methylation status of Foxp3 gene and its roles in immunological pathogenesis of Kawasaki disease(KD). Methods Thirty children with KD and eighteen agematched healthy children consented to participate in this study. Quantitative methylation specific polymerase chain reaction(MSQP) was used to assess the methylation status of Foxp3 promoter and regulatory T cells specific demethylated region(TSDR) in CD4+ T cells. The proportion of CD4+ CD25 + Foxp3 + regulatory T (Tr) cells was analyzed by flow cytometry. Transcriptional levels of CD4+ CD25 + Foxp3 + Tr associating genes (Foxp3, CTLA4, GITR, LAG3 and CCR8 ) and Foxp3-dependent molecules (UBD and LGAIS3)were measured by real-time PCR. Results ( 1 ) Demethylation level of Foxp3 promoter in CD4 + T cells from patients with KD was lower significantly than that of health subjects( P ＜ 0.01 ), and increased significantly after treated with intravenous gamma globulin therapy(IVIG) (P ＜ 0.01 ). No difference of demethylation level of TSDR region was observed among all groups ( P ＞ 0. 05 ). ( 2 ) The proportion of CD4 + CD25 + Foxp3 + Tr in peripheral blood from patients with KD , as well as mRNA levels of Foxp3 gene in CD4+ T cells, was significantly lower than those of health subjects ( P ＜0. 01 ), and increases significantly after IVIG therapy (P ＜0.01 ). Significant positive correlations between demethylation level of Foxp3 promoter and the proportion of CD4 + CD25 + Foxp3 + Tr , or expression levels of Foxp3 in CD4 + T cells, were observed during acute phase of KD (CD4+CD25+ Foxp3+ Tr: r=0.76, P＜0. 01; Foxp3: r=0.89, P＜0. 01). (3)Transcription level of CD4+ CD25+ Foxp3+ Tr associating factors, such as CTLA4, GITR, LAG3 and CCR8, was significantly down-regulated in acute phase of KD(P＜0. 01 ), and up-regulated to some extent after treated with IVIG(P ＜0. 01 ). Expression levels of Foxp3-dependent molecules UBD and LGALS3 in CD4 + T cells decreased significantly during acute phase of KD (P ＜ 0.01 ), and basically recovered to the levels of health subjects( P ＜ 0.01 ). Conclusion Decrease of demethylation level of Fopx3 promoter is correlated with immune dysfunction in Kawasaki disease.

Objective To investigate the role of IL-6/STAT3 signaling in Th17/Tr imbalance of Kawasaki disease(KD). Methods Forty-eight children with KD and eighteen age-matched healthy children were consented to participate in this study. Protein concentration of IL-6 in plasma was measured by ELISA. Transcriptional levels of IL-17A, IL-17F, RORγt, Foxp3, SOCS1 and SOCS3 were assessed by real-time PCR. The proportion of CD4+CD25+Foxp3+ regulatory T(Tr) cells and mean fluorescence intensity(MFI) for phosphorylated-STAT3(pSTAT3) protein in CD4+ T cells was analyzed by flow cytometry. A quantitative methylation specific PCR based on SYBR Green was used to evaluate methylation status of CpG islands in SOCS1 exon2, three potential bind sites for STAT3 in 5'-untraslated region(5'-UTR) of SOCS3 in CD4+ T cells. Results (1)Compared with healthy volunteers, plasma IL-6 concentration and MFI for pSTAT3 in CD4+ T cells were elevated significantly during acute phase of KD[IL-6:(54.02±20.58) pg/ml vs (8.72±2.06) pg/ml, P<0.05;pSTAT3 MFI:(55.41±15.08) vs (9.35±3.76), P<0.05], and the two items in KD patients with coronary artery lesion (KD-CAL+) were found to be higher than those in KD patients without coronary artery lesion (KD-CAL-)[IL-6:(84.76±29.35) pg/ml vs (38.65±13.76) pg/ml, P<0.05;pSTAT3 MFI:(72.36±16.81) vs (46.93±13.57), P<0.05]. (2)Transcription levels of IL-17A, IL-17F and RORγt in patients with KD were significantly elevated (P<0.05) while the proportion of CD4+CD25+Foxp3+ Treg and expression levels of Foxp3 were detected to be lower than those in normal controls (P<0.05). The mRNA levels of IL-17A, IL-17F and RORγt in KD-CAL+ group were higher than those in KD-CAL- group(P<0.05), as well as expression level of Foxp3 were found to be lower in KD-CAL+ group(P<0.05). (3)The mRNA levels of SOCS1 and SOCS3 in CD4+ T cells increased significantly during acute phase of KD(P<0.05), while the two items in KD-CAL+ group were lower than those in KD-CAL- group(P<0.05). Furthermore, CpG islands in SOCS1 exon2 and the third potential bind site for STAT3 in SOCS3 5'-UTR were hypomethylated in acute KD, while those in healthy controls were fully demethylated(P<0.05). Demethylation levels of SOCS1 exon2 and the third potential bind site for STAT3 in SOCS3 5'-UTR in KD-CAL+ group were lower than those in KD-CAL- group(P<0.05). CpG islands in the other two bind sites for STAT3 in SOCS3 5'-UTR were fully demethylated among all the groups(P>0.05). ConclusionAberrant activation of IL-6/STAT3 signaling caused by hypomethylation of SOCS1 and SOCS3 might be one contributing factor to unbalance of Th17/Tr in KD.

Objective To investigate the effect of SOCS1 and SOCS3 hypomethylation on homeostasis of Th1/Th2 in Kawasaki disease(KD). Methods Thirty-six children with KD and sixteen age-matched healthy children consented to participate in this study. Protein concentration of IL-6 in plasma was measured by ELISA. Transcriptional levels of SOCS1, SOCS3, T-bet, IFN-γ, GATA3 and IL-4 were assessed by realtime PCR. The proportion of Th1 and Th2 cells, and mean fluorescence intensity(MFI)for phosphorylated STAT3(pSTAT3)protein in CD4+ T cells was analyzed by flow cytometry. A quantitative methylation specific PCR based on SYBR Green was used to evaluate methylation status of CpG islands in SOCSl exon2, and three potential binding sites for STAT3 in 5'-untraslated region(5'-UTR)of SOCS3 in CD4+T cells. Comparisons between groups were performed with t-test. Results ①Compared with healthy volunteers, plasma IL-6 concentration[(51.8±16.3)pg/ml vs(8.6±2.0)pg/ml, respectively]and MFI for pSTAT3[(52±14)vs(10±4), respectively]in CD4+ T cells were elevated significantly during acute phase of KD(P＜0.05), and the two items in KD patients with coronary artery lesion(KD-CAL+)were found to be higher than those in KD patients without coronary artery lesion(KD-CAL-)[IL-6:(87.2±27.4)pg/ml vs(36.2±12.8)pg/ml, P＜0.05; pSTAT3 MFI:(75±15)vs(42±11), P＜0.05]. ② The proportions of Th1 and Th2 cells and transcription levels of Th-associating factors(T-bet, IFN-γ, GATA3 and IL-4)in CD4+ T cells increased significantly in acute KD(P＜0.05), while the rate of Thl div Th2 in KD patients was found to be lower than that in normal controls(P＜0.05). In addition, the proportions of Th1 and Th2 cells and expressions levels of Th-associating factors in KD-CAL+ group were higher than those in KD-CAL-group, as well as the rate of Thl div Th2 cells in KD -CAL+ group were lower than that in KD-CAL- group(P＜0.05). ③ The mRNA levels of SOCSl and SOCS3 in CD4+ T cells increased significantly during acute phase of KD(P＜0.05), while the two items in KDCAL+ group were lower than those in KD-CAL- group(P＜0.05). Furthermore, CpG islands in SOCSl exon2 and the third potential binding site for STAT3 in SOCS3 5'-UTR were hypomethylated in acute KD, while those in healthy volunteers were fully demethylated(P＜0.05). Demethylation levels of the two items mentioned above in the KD-CAL+ group were lower than those in the KD-CAL-group(P＜0.05). CpG islands in the other two binding sites for STAT3 in SOCS3 5'-UTR were fully demethylated among all the groups(P＞0.05).Conclusion Relative insufficiency of SOCS1 and SOCS3 expression caused by hypomethylation may be one contributing factor for the imbalance of Th1/Th2 in KD.

ObjectiveTo investigate the role of signal transduction of TLRs in the Henoch-Schonlein purpura (HSP). Methods Reverse-transcription PCR (RT-PCR) and real-time PCR were used to evaluate the levels of TLRs(1～10), MyD88, TRAF6, TRIF, IFN-α, IFN-β, IL-6, IL-1β, TNF-α, IP-10, RANTES,iNOS, Blys/April mRNA expression in peripheral blood mononuclear cells and their expression levels were compared using t test., while the concentration of plasma cytokines such as Blys、IFN-α、IFN-β、IL-6、IL-1、TNF-α was measured by enzyme-linked immunosorbent assay(ELISA).Expression levels of those genes were compared using t test. Results①Compared with the control group, the expression levels of TLR1, TLR2,TLR6, TLR5, TLR3, TLR7, TLR9 mRNA were up-regulated significantly(P＜0.01), while no difference of TLR4 was detected (P＞0.05).②Transcription levels of MyDg8(2.47±1.06) vs(0.73±0.22), TRAF6 (2.54±0.72)×10-3vs(0.70±0.20)×10-3, TRIF(3.18±0.86)×10-3vs(0.93±0.35)×10-3 were significantly up-regulated in acute phase of HSP (P＜0.01).③The levels of IFN-α and IFN-β protein and mRNA were remarkable increased (P＜0.01).④ The expression of cytokine/chemotactic factor such as IL-6, IL-1β, IP-10, RANTES,iNOS was higher than that of the control group(p＜0.01), while TNF-αdid not change in children with HSP (P＞0.05). ⑤ It was detected that the expression of Blys/April was higher than that of the control group(P＜0.01). ConclusionExpressions of TLR1, TLR2, TLR6, TIR5, TLR3, TLR7, TLR9, MyD88, TRAF6, and TRIF are up-regulated during acute phase of HSP, suggesting that aberrant activation of TLRs triggered by microbes may be one of the initiating factors of immune aberrance in HSP. Over expression of cytokine/chemotactic factor or Blys/April owning to the aberrant activation of TLRs, may be correlated with immunological pathogenesis of HSP.

Objective To investigate the role of T follicular helper ( Tfh) cells in the immuno-pathogenesis of persistent immune thrombocytopenic purpura ( pITP) .Methods Twenty children with pITP and twenty healthy controls were enrolled in this study .The proportion of CD4+CXCR5+ICOShigh PD-1high T ( cTfh) cells and the expression of ICOSL on CD 19+B cells in peripheral blood of the patients and healthy subjects were analyzed by flow cytometry .The expressions of Bcl-6, c-Maf, IL-21 and ICOSL at mRNA level were detected by real-time PCR.The plasma concentrations of IL-2, IL-6 and IL-21 were determined by ELISA.Results (1) Compared with the healthy controls , the proportions of cTfh cells increased signifi-cantly in patients with pITP [(17.45±9.04) %vs.(7.57±2.57) %, P<0.05], but decreased with the treatment of dexamethasone (DEX) for 7 days [(5.93±1.64) %vs.(17.45±9.04) %, P<0.05].(2) The expression of Bcl-6, c-Maf and IL-21 at mRNA level in patients with pITP were higher than those in healthy controls (P<0.05).(3) Compared with healthy controls, the expression of ICOSL at mRNA and protein levels in CD19+B cells were significantly up-regulated in patients with pITP (P<0.05), which showed no significant changes after treatment with DEX (P>0.05).(4) The plasma concentration of IL-21 was remarkably elevated in patients with pITP , regardless of DEX treatment (P<0.05).But the plasma concentrations of IL-2 and IL-6 showed no significant changes compared with control group (P>0.05).Con-clusion The immunopathogenesis of persistent immune thrombocytopenic purpura might be associated with the hyper-activation of Tfh cells caused by excessive expression of ICOSL and IL-21.The persistent high expression of ICOSL and IL-21 might be one of the important factors resulting in the recurrence of pITP in children .

Objective To investigate the effects of Yiqi Congming pill combined with western medicine on hearing and sleep quality in tinnitus patients. Methods 60 tinnitus patients caused by liver fire treated in our hospital from May 2015 to February 2017 were selected for the study and randomly divided into the observation group and the control group, with 30 cases in each group. The two groups were given conventional western medicine and Yiqi Congming pills combined with western medicine respectively. The effective rates of treatment, the speech pure-tone average (PTA), speech recognition threshold (SRT) and Pittsburgh Sleep Quality Index (PSQI) were compared between the two groups, and the adverse reactions were observed. Results The effective rate of the observation group was 93.33％, which was higher than that of the control group with 73.33％ (P<0.05). After treatment, the levels of PTA, SRT the scores of PSQI were significantly lower in the observation group than those in the control group (P<0.05). There were no differences in the incidence rates of adverse reactions between the two groups. Conclusions Yiqi Congming pill combined with western medicine can effectively treat tinnitus caused by liver fire, and improve hearing and sleep quality without obvious adverse reactions. Therefore it is worthy of clinical promotion and application.

Objective To develop a portable endoscopic thermal device which possesses the merits of compact structure,convenient operation,security and good compatibility.Methods Rapid heating elements and several materials are used,which are excellent insulation,heat conduction and can withstand high temperatures.The production process of miniaturization,anti-conglutination and constant temperatureare adopted.Results: The size of Heat probe is about 2.4mm in outer diameter and 8 mm in length.The maximum temperature will reach 250℃,heating time within 10s,output 4.5～6.5W.Conclusion Heat Probe can be compatible with various types gastrointestinal endoscopic channel.Besides hemostasis of upper gastrointestinal,it can also be applied to small polyps,erosive gastritis,etc.

Objective:To investigate the protective effect of adenosine on cochlea in acute acoustic trauma. Methods: Guinea pigs were exposed to 118 dB SPL white noise for 30 min. During the process of exposure, artificial perilymph or artificial perilymph containing adenosine (0.1 mmol/L,1 mmol/L or 5 mmol/L) was infused into the cochlea through perilymphatic perfusion. Concentration of glutamate in perilymph and thresholds of compound action potentials (CAP) were measured before and after noise exposure. Results: After exposure, the concentration of glutamate and the mean threshold shifts evoked by click were significantly higher than those of before exposure. Concentration of glutamate and thresholds of CAP in adenosine (1 mmol/L and 5 mmol/L) treated groups were significantly lower than those of the control group. Conclusion: Adenosine can inhibit the elevation of glutamate and decrease the threshold shift of CAP after exposure to noise,which can protect cochlea in acute acoustic trauma in guinea pigs.

Objective To investigate the effects of sIL-2R in plasma on regulatory T cells (Treg) in children with Kawasaki disease ( KD ) .Methods Thirty-three children with KD and fourteen age-matched healthy children were enrolled in this study .The proportions of CD4+CD25+Foxp3+Treg cells in pe-ripheral blood and mean fluorescence intensity (MFI) of phosphorylated-STAT5 (pSTAT5) protein in CD4+CD25+T cells were analyzed by flow cytometry .The concentrations of sIL-2R, IL-2, IL-7 and IL-15 in plas-ma were measured by cytometric bead array ( CBA ) .Real-time PCR was performed to detect the gene ex-pressions of Foxp3, GITR, CTLA4, IL-2Rα, IL-2Rβand IL-2Rγat mRNA level as well as the expressions of IL-17A and ROR-γt at mRNA level in CD4+CD25-T cells.Results (1) Compared with the control group, the proportions of CD4+CD25+Foxp3+Treg cells in peripheral blood from patients with KD and the expressions of associated factors including Foxp 3, GITR and CTLA-4 at mRNA level were significantly down-regulated (P<0.05).However,the expressions of IL-17A and ROR-γt at mRNA level in Th17 cells were markedly up-regulated (P<0.05), which could be recovered to some extent after treatment with IVIG (P<0.05).(2)The expressions of pSTAT5 protein in CD4+CD25+T cells from patients with acute KD were sig-nificantly decreased (P<00.5 ), but increased with IVIG intervention (P<0.05).(3)The concentrations of sIL-2R in plasma were elevated during acute KD (P<0.05), but decreased after treatment with IVIG (P<0.05).Moreover, KD patients with coronary artery lesion ( KD-CAL+) presented a high level of sIL-2R than those without coronary artery lesion (KD-CAL-) (P<0.05), but there was no significant difference in the concentrations of IL-2, IL-7 and IL-15 in plasma between two groups (P>0.05).(4)The expressions of IL-2Rαand IL-2Rβat mRNA level in CD4+CD25+T cells from patients with acute KD were lower than those of the healthy subjects (P<0.05), but up-regulated to some extent with IVIG treatment (P<0.05).There was no significant change in the expression of IL-2Rγat mRNA level (P>0.05).The concentrations of sIL-2R in plasma were negatively correlated with the expressions of IL-2Rβand Foxp3 at mRNA level and pSTAT5 at protein level (P<0.05).The expression of pSTAT5 protein had positive correlation with the ex-pression of Foxp3 at mRNA level (P<0.05).Conclusion Aberrant IL-2/STAT5 signaling pathway media-ted by significantly increased concentration of sIL-2R in plasma might be one of the factors leading to down-regulation of Treg cells in patients with acute KD .