Objective To study the significance of level changes of serum hydrogen sulfide(H2 S) and interleukin-6(IL-6)in children with hand,foot and mouth disease(HFMD).Methods Nity-two cases with HFMD were enrolled and divided into severe HFMD group(48 cases)and common HFMD group(44 cases).Forty-six healthy children were enrolled as healthy control group.The serum H2 S and IL-6 were detected.Results Acute phase:compared to the healthy control group[H2 S(55.76 ±7.80)μmol /L,IL-6 (61.31 ±13.43)ng /L],the level of serum H2 S significantly reduced and the level of IL-6 significantly increased in severe HFMD group[H2 S(21.72 ±7.52)μmol /L;IL-6(131.33 ±17.82)ng /L]and common HFMD group[H2 S(31.86 ±8.41 )μmol /L;IL-6(95.48 ±15.07)ng /L](P ﹤0.01 ),and there was signifi-cantly difference between the severe HFMD group and common HFMD group(P ﹤0.01 ).Recovery phase:compared to the healthy control group,the serum H2 S level[(34.54 ±13.21 )μmol /L]was lower and IL-6 [(92.73 ±15.25)ng /L]was higher in severe HFMD group(P ﹤0.01 ).The serum H2 S level was negatively correlated with IL-6 in severe HFMD group and common HFMD group(r =-0.31 ,P ﹤0.01 ;r =-0.45, P ﹤0.01 ).Conclusion The serum H2 S and IL-6 participate in the pathological process of HFMD,and the level change can be used as one of indicators of early diagnosis.

Objective: To observe the cellular damage of low-dose combined exposure to Hg, Pb and Cd on hippocampal neurons in rat. Methods: SH-SY5Y cells were randomly divided into 8 groups by 2×2×2 factorial design: control group, Pb exposure group, Hg exposure group, Pb+Hg exposure group, Pb+Cd exposure group, Hg+Cd exposure group and Pb+Cd+Hg exposure group. And the cell viabilities were measured. On this basis, an animal model was established. Twenty eight-week-old SD pregnant rats were randomly divided into four groups by random number table, and five in each group: the control group(distilled water), 1-fold metal mixture exposure group (1×MM, poisoning solution containing mercury chloride 0.15 mg/L, lead acetate trihydrate 25 mg/L, cadmium chloride 7.5 mg/L), 5-fold metal mixture exposure group (5×MM, poisoning solution containing mercury chloride 0.75 mg/L, lead acetate trihydrate 125.00 mg/L, cadmium chloride 37.50 mg/L), 10-fold metal mixture exposure group (10×MM, poisoning solution containing mercury chloride 1.50 mg/L, lead acetate trihydrate 250.00 mg/L, cadmium chloride 75.00 mg/L). Pregnant rats drank water until delivery. Twenty male pups were selected and exposed to these metals through breast milk until weaned. The heavy metals dose of poisoning water was adjusted, and then the weaned rats were exposed to heavy metals via drinking poisoning water until adulthood (postnatal day 83). The blood samples and brain hippocampus samples were collected to observe the ultrastructural changes of hippocampus, and to determine the levels of Hg, Pb and Cd in blood. In addition, apoptosis rate and fluorescence intensity of reactive oxygen species and intracellular free calcium concentration ([Ca²⁺]_i) in hippocampal neurons were measured. Results: Cellular factorial design analysis showed that Hg+Pb+Cd (at no observed adverse effect level, 1.0, 0.5 and 0.1 μmol/L, respectively)had a interaction on cell viability after 48 or 72 hours of combined exposure (P<0.05). The results of ultrastructure showed that mitochondria decreased, ridges and matrixes gradually dissolved in rat hippocampal neurons of 5×MM group; nuclear chromatin aggregated, more ridges and matrixes dissolved and the mitochondria also decreased in rat hippocampal neurons of 10×MM group. The concentration of Hg, Pb and Cd in the blood of 1×MM group, 5×MM group and 10×MM group were higher than those in the control group, and the differences were statistically significant (P<0.001). There was no significant difference in apoptosis rate between the 1×MM group and the control group. The apoptosis rate of 5×MM group and 10×MM group was higher than that in the control group, and the differences were statistically significant (P<0.001). There was no statistically significant difference in the fluorescence intensity of reactive oxygen species in hippocampal neurons of the 1×MM group and the control group. The fluorescence intensity of reactive oxygen species in the 5×MM group and the 10×MM group was higher than that in the control group, the difference was statistically significant (P<0.05). There was no significant difference in the fluorescence intensity of [Ca²⁺]_i between the 1×MM group and the control group. The fluorescence intensity values of [Ca²⁺]_i in the 5×MM group and the 10×MM group were higher than the control group, the differences were statistically significant (P<0.001). Conclusion Low-level combined exposure to Hg, Pb, and Cd caused synergistic neurotoxic damage, and the process may be related to the changes of neuronal apoptosis, reactive oxide species, and [Ca²⁺]_i levels.

Objective:To analyze the etiological distribution and phylogenetic characteristics of hand, foot and mouth disease (HFMD) in Qujing city of Yunnan Province in 2020.Methods:Stool samples were collected from HFMD cases in Qujing city in 2020 and virus RNA was extracted directly from treated stool suspensions. Virus VP4 gene sequences were firstly amplified using MD91/OL68-1 primer pairs and sequenced, then the virus serotypes were determined by BLAST search on the GenBank. Virus entire VP1 gene sequences were amplified and sequenced. Virus serotypes were identified online using Enterovirus Genotyping Tool Version 0.1. Sequences of reference virus genotypes/sub-genotypes were downloaded according to references. Phylogenetic trees were constructed by MEGA5.2 software and the genetic characteristics were analyzed.Results:A total of 47 strains of enteroviruses (EVs) were detected with a detection rate of 10.22% (47/460). The detected viruses were coxsackievirus A4 (CVA4, 0.65%, 3/460), CVA6 (7.83%, 36/460), CVA10 (0.87%, 4/460) and CVA16 (0.87%, 4/460). All were enterovirus species A (EVA), while other group viruses were not detected. The predominant virus was CVA6, accounting for 7.83% (36/460). EVA71 was not detected. CVA4 strains of C2 and C4 subgenotypes were co-circulating strains in Qujing city. CVA6 subgenotype D3a and CVA16 subgenotype B1a were also circulated in Qujing city. All CVA10 strains were in a separate lineage.Conclusions:Similar to the previous situation in China, the detection rates of EVA71 and CVA16 were very low, even zero. This study showed that CVA6 was the predominant virus, indicating a HFMD outbreak caused by CVA6 in Qujing city in 2020. The phylogenetic analysis showed CVA10 isolates belonged to a separate lineage, which might be unique to Qujing city. Laboratory and molecular epidemiological surveillance of non-EVA71 and non-CVA16 viruses, especially CVA6 and CVA10 should be strengthened in the future.

Objective To investigate the expression of T cell immunoglobulin and mucindomain-containing molecule-3 (TIM3) on PBMCs,and the plasma concentrations of soluble forms of Galetcin9 and their clinical relationship with rheumatoid arthritis (RA).Methods Peripheral blood samples were collected from 39 patients,25 osteoarthritis (OA) patients and 20 healthy subjects (HC).The expressions of TIM3 on peripheral blood mononuelear cells (PBMCs) were detected by flow cytometry.The concentrations of soluble Galetcin9 were assessed by enzyme linked immunosorbent assay (ELISA).And the relationship between their expression levels and clinical manifestations were analyzed.Levene F test was used for statistical analysis,normal distribution data were compared by t test and Pearson correlation analysis,while Mann-Whitney U test and Spearman correlation analysis were used for non-normal distribution data.Results The expression of TIM3 on CD4+ T cells was significantly higher than that of the HC [(14.7±3.2)％ vs (5.1±0.8)％,t=2.339,P=0.022 7],while there was no statistical difference between the RA group and the OA group [(14.7±3.2)％ vs (5.8±0.4)％,t=1.928,P=0.058 9].The expression of TIM3 was significantly correlated with the concentration of RF in the serum and the corresponding DAS28 score (r=0.325 8,P=0.043 0;r=0.407 5,P=0.010 0).The expression of TIM3 on CD8 + T cells in RA patients was significantly higher than that in the HC and OA [(21.1±3.4)％ vs (8.3±1.5)％,t=2.531,P=0.0142;(21.1±3.4)％ vs (10.7±1.0)％,t=2.314,P=0.024 0] which was significantly correlated with the concentration of RF in serum and the corresponding DAS28 score (r=0.451 5,P=0.003 9;r=0.524 1,P=0.000 6) as well.However,the expression of TIM3 on CD56+ NK cells was not significantly different from that of either OA or HC[(56.4±3.4)％ vs (50.6±3.8)％,t=1.047,P=0.299 8;(56.4± 3.4)％ vs (56.1±3.4)％,t=0.048,P=0.961 9],the concentration of serum RF and the corresponding DAS28 score were not significantly related.We also found that plasma Galectin 9 concentrations in RA patients were significantly higher than those of OA patients [(4.24±0.22) ng/ml vs (3.15±0.18) ng/ml,t=3.187,P=0.024] and healthy subjects [(4.24±0.22) ng/ml vs (2.55±0.14) ng/ml,t=5.567,P＜0.01],which was correlated with RF and DAS28 (r=0.479 2,P=0.002 0;r=0.353 0,P=0.027 5) while there was no correlation with CRP (r=0.176 3,P=0.283 1).Conclusion The upregulated expressions of TIM3 on peripheral lymphocytes and the high levels of plasma concentration of soluble Galectin 9 are closely correlated with the severity of the disease,suggesting that TIM3/Galectin 9 pathway may play a critical role in the pathogenesis of RA.

Objective:To investigate the effect of B7-H3 gene on the biological function of fibr-oblastlike synoviocytes (FLS) in osteoarthritis (OA).Methods:Synovial tissue of five cases of OA and synovial tissue of 4 normal knee were obtained, and the primary cell lines were isolated and cultured. The expression of B7-H3 in OA synovial tissue and primary OA-FLS were studied by immunohi-stochemistry, real time-poly merase chain reaction (PCR) and FACS. According to sites 996 and 1041 of B7-H3, corresponding siRNA was designed and the expression of B7-H3 in FLS was silenced and down-regulated. The inhibition of B7-H3 and its protein in target cells was determined by Western blot and FACS. The migration and invasion ability of B7-H3 in target cells were analyzed by scratch assay and Transwell assay. CCK8 assay was used to detect cell proliferation ability, and CBA assay was used to detect cytokines and chemokines in cell culture supernatant. GraphPad Prism 8.0 software was used to analyze the experimental data. The normal distribution data was expressed as mean±standard deviation ( Mean± SD). The comparison between data was performed by T test, and P<0.05 was considered statistically significant. Results:The abnormally high expression of B7-H3 in fibro-blast-like synoviocytes of OA was detected. Compared with siNC, si996 and si1041 inhibited the expression of B7-H3 in OA-FLS. In the Transwell migration experiment, the mean cells number of random view in the siNC group, the si996 group, and the si1041 group indicating decreased migration ability of OA-FLS [siNC vs si996 (100.3±3.7) /view vs (48.7±1.2) /view, t=13.24, P<0.001; siNC vs si1041 (100.3±3.7) /view vs (59.7±1.9) /view, t=9.80, P<0.001). In the Transwell invasion experiment, the mean cells number of random view in the siNC group, in the si996 group, and in the si1041 group indicating decreased invasion ability of OA-FLS [siNC vs si996 (127.3±5.6) /view vs (39.7±3.3) /view, t=13.49, P<0.001; siNC vs si1041 (127.3±5.6) /view vs (57.3±1.9) /view, t=11.85, P<0.001]. The secretion of IL-6 [siNC vs si996 (248±21) pg/ml vs (111±12) pg/ml, t=24.08, P=0.002; siNC vs si1041 (248±21) pg/ml vs (46±5) pg/ml, t=13.21, P=0.006], IL-8 [siNC vs si996 (118.1±15.6) pg/ml vs (47.1±5.4) pg/ml, t=6.68, P=0.022; siNC vs si1041 (118.1±15.6) pg/ml vs (10.0±1.3) pg/ml, t=13.08, P=0.006], CXCL8 [siNC vs si996 (178.8±6.4) ng/ml vs (83.2±2.7) ng/ml, t=13.77, P=0.005; siNC vs si1041 (178.8±6.4) ng/ml vs (93.5±2.8) ng/ml, t=12.23, P=0.007] and CCL2 [siNC vs si996 [(184.1±5.1) ng/ml vs (109.4±5.9) ng/ml, t=9.57, P=0.011; siNC vs si1041 (184.1±5.1) ng/ml vs (97.1±1.5) ng/ml, t=16.39, P=0.004] was decreased . Conclusion:B7-H3 may regulate the migration, invasion, cytokine secretion and other biological functions of OA-FLS, providing clues for further study of B7-H3's involvement in the pathogenesis of OA.