Objective To explore the potential circular RNA(circRNA)biomarker of Uygur type 2 diabetes mellitus(T2DM).Methods A total of 120 T2DM patients and 120 subjects with normal glucose tolerance were recruited from the Department of Endocrinology and Metabolism of the First Affiliated Hospital of Shihezi University and Shihezi community from October 2020 to August 2021,and divided into four groups:60 Uygur T2DM patients(Uygur T2DM group),60 Uygur subjects with normal glucose tolerance(Uygur NC group),60 Han T2DM patients(Han T2DM group)and 60 Han subjects with normal glucose tolerance(Han NC group).Hsa_circRNA_0042817,hsa_circRNA_0006532 and hsa_circRNA_0004131 were selected as candidate circRNA,and the expression in peripheral blood were detected by RT-qPCR.Logistic regression was used to analyze the influencing factors for Uygur T2DM,and the receiver operating characteristic(ROC)curve was used to evaluate the biomarker value of circRNA in Uygur T2DM.Results The expressions of hsa_circRNA_0042817,hsa_circRNA_0006532 and hsa_circRNA_0004131 were higher in Uygur T2DM group than in Uygur NC group(P<0.05).The expression of hsa_circRNA_0042817 was higher in Uygur T2DM group than in Han T2DM group(P<0.05).Logistic regression analysis showed that hsa_circRNA_0042817 was an influencing factor for T2DM in Uygur population[OR(95%CI)3.420(1.567～7.465)].ROC curve analysis showed that the area under the curve was the largest(0.798)in hsa_circRNA_0042817.Conclusion There were up-regulated circRNA in peripheral blood in Uygur T2DM patients,and hsa_circRNA_0042817 may be a biomarker for T2DM in Uygur patients.

Objective:To analyze the viral genome sequence of novel coronavirus infected persons in Baotou City, understand the mutation characteristics of novel coronavirus genome in the process of transmission among cases, and explore the transmission rule of novel coronavirus in the clustered populations.Methods:Nine throat swabs samples (No. 1 - 7, No. 9, and No. 10), two sputum samples (No. 8, No. 11, and No. 11 sample was from No. 10 case), and one surface smear sample (No.12, and No. 12 sample was from No. 10 case) were collected from 10 confirmed cases of novel coronavirus infection in Baotou City from January 25 to February 21, 2020. Samples 1 and 3 were from single cases, and the rest were from clustered cases. The virus genome was sequenced by metagenomic next-generation sequencing (mNGS), and single nucleotide polymorphism (SNP) mutation sites were screened by comparing with NC_045512, a reference strain of novel coronavirus. Combined with relevant epidemiological information, gene mutation, virus typing, and evolutionary traceability analysis were carried out.Results:The results of viral genome mNGS showed that 76 SNP mutation sites were detected in 12 samples compared with the reference strain NC_045512, including 3 (3.95%) transitions and 73 (96.05%) reversals. There were 19 (25.00%) synonymous mutations and 57 (75.00%) non-synonymous mutations. The analysis of nucleotide and amino acid variation sites showed that mutations were found at five sites (T2821C, C6548T, T16464C, G16858A and T251C) in all the clustered cases (cases 2, 4 - 10). In the single cases, sample 1 had mutations at C9245T and A15340T, and sample 3 had mutation at C13T. The virus typing analysis showed that the samples 1 and 3 belonged to the L type of novel coronavirus, while the rest belonged to the S type of novel coronavirus. The results of genomic evolutionary relationship analysis showed that all the samples could be divided into two branches. The branches of sample 1 and 3 belonged to single cases, and the rest belonged to family clustered cases.Conclusion:The genomic characteristics of the clustered cases of novel coronavirus infection in Baotou City are basically consistent with the epidemiological investigation results, and the transmission of the virus is mainly related to close contact and family gathering.

Objective:To observe any effect of electroacupuncture (EA) on the expression of phosphorylated extracellular signal-regulated protein kinase (p-ERK1/2) and phosphorylated cyclic adenosine monophosphate response element binding protein (p-CREB) in the spinal dorsal horns of diabetics experiencing neuropathic pain.Methods:Eight rats were randomly selected from 30 healthy male Sprague-Dawley rats as the normal group (N), and the remaining twenty-two rats were treated with a single high-dose intraperitoneal injection of streptozotocin (STZ) to establish a neuropathic pain model. The rats modeled successfully were randomly divided into a model group (M, n=8) and an EA group ( n=8). In the EA group, electroacupuncture was applied at the bilateral Hou san li and Kunlun acupoints starting on the 15th day after the STZ injection. The daily sessions lasted 30 minutes for 1 week. Body weight (BW), fasting blood glucose (FBG) and paw withdrawal latency (PWL) were observed before the STZ injection and on the 7th, 14th, and 21st days afterward. The expression of p-ERK1/2 and p-CREB in the dorsal horns of the rats′ spinal cords was detected using western blotting. The count of p-CREB-positive cells in the dorsal horns and their co-localization with neurons was detected using immunofluorescence. Results:In comparison with the N group, the average BW of the M group on the 7th, 14th and 21st days after the STZ injection was significantly lower, while the average FBG was significantly higher. There was no significant difference between the M and N groups in the average PWL on the 7th day after the STZ injection, but it had decreased significantly in the M group on the 14th and 21st days. Compared with the M group, the average PWL of the EA group was significantly longer on the 21st day after the injection. The expression of p-ERK1/2 and p-CREB protein in the spines of the M group was significantly higher than in the N group. p-CREB positive cells were more numerous in the M group compared with the N group, while in the EA group they were fewer. P-CREB was co-located with neurons in the spinal dorsal horn.Conclusion:EA can alleviate neuropathic pain effectively, perhaps by inhibiting the expression of p-ERK1/2 and p-CREB in the dorsal horns of the spinal cord.

OBJECTIVE:To establish a method for simultaneous determination of tanshinol,caffeic acid,rosmarinic,salviano-lic acid B,salvianolic acid A,tanshinoneⅠ,cryptotanshinone,tanshinone ⅡA and ursolic acid in Compound xueshuantong cap-sules. METHODS:UPLC-MS/MS method was adopted. The determination was performed on ACQUITY UPLC? BEH C18 column with mobile phase consisted of acetonitrile-0.1%formic acid(gradient elution)at the flow rate of 0.2 mL/min. The column tempera-ture was 40 ℃,and the temperature of injector was 10 ℃. Analysis time was 7 min,and sample size was 5 μL. The electrospray ionization source(ESI)was used;ion source temperature was 150℃;capillary voltage was 3.5 kV;cone flow was 50 L/h;desol-vation temperature was 350 ℃;desolvation gas flow was 650 L/h;nebuliser pressure was 7 × 105 Pa;ion monitoring and multiple reaction monitoring (MRM) was performed. RESULTS:The linear ranges of tanshinol,caffeic acid,rosmarinic,salvianolic acid B,salvianolic acid A,tanshinoneⅠ,cryptotanshinone,tanshinone ⅡA and ursolic acid were 10.0-100.0 μg/mL (r=0.9998), 0.1-1.0 μg/mL(r=0.9998),4.0-40.0 μg/mL(r=0.9999),10.0-100.0 μg/mL(r=0.9999),15.0-150.0 μg/mL(r=0.9997), 8.0-80.0 μg/mL(r=0.9998),10.0-100.0 μg/mL(r=0.9997),50.0-500.0 μg/mL(r=0.9997)and 6.0-60.0 μg/mL(r=0.9998), respectively. The limits of quantitation were 40.0,9.6,38.0,88.0,130.0,39.0,4.4,3.2 and 10.0 ng/mL,separately. The limits of detection were 12.0,3.0,11.0,26.0,39.0,12.0,1.3,1.0 and 3.0 ng/mL,respectively. RSDs of precision,stability and repro-ducibility tests were all lower than 3%. The recoveries were 97.34%-103.20%(RSD=2.19%,n=6),97.22%-102.39%(RSD=2.03%,n=6),98.51%-101.70%(RSD=1.32%,n=6),97.86%-102.49%(RSD=2.09%,n=6),96.75%-103.12%(RSD=2.36%,n=6),98.43%-101.65%(RSD=1.25%,n=6), 97.59%-101.50%(RSD=1.50%,n=6), 96.45%-102.88%(RSD=2.58%,n=6),97.02%-103.11%(RSD=2.38%,n=6),separately. CONCLUSIONS:The method is simple and accurate,and can be used for simultaneous determination of 9 components in Compound xueshuantong capsules.

AIM: To investigate the expression and effects of sialyl Lewis X (SLeX) on the invasion and migration of human hepatocellular carcinoma HepG2 cells.METHODS: The expression of α1,3-fucosyltransferase VII (FUT7) in HepG2 cells and L-02 cells was detected by RT-qPCR and Western blot.The SLeX expression in HepG2 cells and L-02 cells was determined by Western blot and immunocytochemical staining.The invasion and migration abilities of the treated cells were evaluated by Transwell assay.RESULTS: The expression of FUT7 and SLeX in the HepG2 cells, but not in the L-02 cells, was observed.The invasion rates of the HepG2 cells treated with SLeX monoclonal antibody at 0.05, 0.5 and 5 mg/L were significantly decreased as compared with control group (P<0.05).The migration ability of the HepG2 cells treated with SLeX monoclonal antibody at 0.05, 0.5 and 5 mg/L was also significantly reduced as compared with control group (P<0.05).The invasion rate and migratory cell number were significantly different between any 2 groups in the HepG2 cells treated with SLeX monoclonal antibody at 0.05, 0.5 and 5 mg/L (P<0.05).CONCLUSION: HepG2 cells express SLeX.SLeX is closely related to the migration and invasion abilities of the HepG2 cells.

OBJECTIVETo investigate the role of glycogen synthase kinase 3β (GSK-3β) in the maturation and function of murine bone marrow-derived dendritic cells (BMDCs).

METHODSMature DCs (mDCs) induced by LPS were examined for GSK-3β phosphorylation level with Western blotting before and after LPS exposure. To explore the role of GSK-3β in maturation and function of DCs, we added SB216763, a selective inhibitor of GSK-3β, in the cell culture of immature DCs (iDCs), and examined CD40 and CD86 expressions in the cells by flow cytometry and the expression of IL-6, IL-12 and IL-10 mRNA by real-time PCR; the changes of the immunogenicity of the cells was evaluated by mixed lymphocyte reaction. The expression of GSK-3β and RelB was examined by Western blotting in DC2.4 cells transfected with a lentiviral vector over-expressing murine GSK-3β gene.

RESULTSLPS exposure significantly lowered GSK-3β activity in iDCs as demonstrated by increased Ser9 phosphorylation and reduced Tyr216 phosphorylation. GSK-3β inhibition induced DC maturation by increasing the expression of surface costimulatory molecules CD40 and CD86, lowered the expressions of IL-6 and IL-12 while enhanced the expression of IL-10 in iDCs, and impaired mixed lymphocyte reaction of the cells. In DC2.4 cells, lentivirus-mediated over-expression of GSK-3β obviously down-regulated the expression of RelB.

CONCLUSIONSGSK-3β is a crucial enzyme involved in the differentiation and maintenance of an immature phenotype of DCs. GSK-3β is constitutively active in iDCs to inhibit their spontaneous maturation. DCs become phenotypically mature after inhibition of GSK-3β, which also executes a proinflammatory task in DC activation. The reduction of RelB protein levels as a result of GSK-3β overexpression supports GSK-3β as a new target for inducing tolerogenic DCs.

Animals ; B7-2 Antigen ; metabolism ; CD40 Antigens ; metabolism ; Cell Differentiation ; Cells, Cultured ; Culture Media ; chemistry ; Dendritic Cells ; enzymology ; Glycogen Synthase Kinase 3 ; metabolism ; Glycogen Synthase Kinase 3 beta ; Indoles ; chemistry ; Interleukin-10 ; metabolism ; Interleukin-12 ; metabolism ; Interleukin-6 ; metabolism ; Lentivirus ; Lymphocyte Culture Test, Mixed ; Maleimides ; chemistry ; Mice ; Myeloid Cells ; enzymology ; Phosphorylation ; RNA, Messenger ; Real-Time Polymerase Chain Reaction ; Signal Transduction

Objective To investigate the role of glycogen synthase kinase 3β(GSK-3β) in the maturation and function of murine bone marrow-derived dendritic cells (BMDCs). Methods Mature DCs (mDCs) induced by LPS were examined for GSK-3βphosphorylation level with Western blotting before and after LPS exposure. To explore the role of GSK-3βin maturation and function of DCs, we added SB216763, a selective inhibitor of GSK-3β, in the cell culture of immature DCs (iDCs), and examined CD40 and CD86 expressions in the cells by flow cytometry and the expression of IL-6, IL-12 and IL-10 mRNA by real-time PCR;the changes of the immunogenicity of the cells was evaluated by mixed lymphocyte reaction. The expression of GSK-3βand RelB was examined by Western blotting in DC2.4 cells transfected with a lentiviral vector over-expressing murine GSK-3βgene. Results LPS exposure significantly lowered GSK-3βactivity in iDCs as demonstrated by increased Ser9 phosphorylation and reduced Tyr216 phosphorylation. GSK-3β inhibition induced DC maturation by increasing the expression of surface costimulatory molecules CD40 and CD86, lowered the expressions of IL-6 and IL-12 while enhanced the expression of IL-10 in iDCs, and impaired mixed lymphocyte reaction of the cells. In DC2.4 cells, lentivirus-mediated over-expression of GSK-3βobviously down-regulated the expression of RelB. Conclusion GSK-3βis a crucial enzyme involved in the differentiation and maintenance of an immature phenotype of DCs. GSK-3β is constitutively active in iDCs to inhibit their spontaneous maturation. DCs become phenotypically mature after inhibition of GSK-3β, which also executes a proinflammatory task in DC activation. The reduction of RelB protein levels as a result of GSK-3β overexpression supports GSK-3β as a new target for inducing tolerogenic DCs.

Objective To investigate the role of glycogen synthase kinase 3β(GSK-3β) in the maturation and function of murine bone marrow-derived dendritic cells (BMDCs). Methods Mature DCs (mDCs) induced by LPS were examined for GSK-3βphosphorylation level with Western blotting before and after LPS exposure. To explore the role of GSK-3βin maturation and function of DCs, we added SB216763, a selective inhibitor of GSK-3β, in the cell culture of immature DCs (iDCs), and examined CD40 and CD86 expressions in the cells by flow cytometry and the expression of IL-6, IL-12 and IL-10 mRNA by real-time PCR;the changes of the immunogenicity of the cells was evaluated by mixed lymphocyte reaction. The expression of GSK-3βand RelB was examined by Western blotting in DC2.4 cells transfected with a lentiviral vector over-expressing murine GSK-3βgene. Results LPS exposure significantly lowered GSK-3βactivity in iDCs as demonstrated by increased Ser9 phosphorylation and reduced Tyr216 phosphorylation. GSK-3β inhibition induced DC maturation by increasing the expression of surface costimulatory molecules CD40 and CD86, lowered the expressions of IL-6 and IL-12 while enhanced the expression of IL-10 in iDCs, and impaired mixed lymphocyte reaction of the cells. In DC2.4 cells, lentivirus-mediated over-expression of GSK-3βobviously down-regulated the expression of RelB. Conclusion GSK-3βis a crucial enzyme involved in the differentiation and maintenance of an immature phenotype of DCs. GSK-3β is constitutively active in iDCs to inhibit their spontaneous maturation. DCs become phenotypically mature after inhibition of GSK-3β, which also executes a proinflammatory task in DC activation. The reduction of RelB protein levels as a result of GSK-3β overexpression supports GSK-3β as a new target for inducing tolerogenic DCs.

OBJECTIVETo investigate the correlation between infection with L-form of Helicobacter pylori (Hp-L) and the expressions of macrophage migration inhibition factor (MIF), matrix metalloproteinase 9 (MMP9), and vascular endothelial growth factor (VEGF) in gastric cancer.

METHODSHp-L was examined in 80 gastric carcinoma and 50 adjacent normal tissues by Gram staining and immunohistochemical staining, and the expressions of MIF, MMP9 and VEGF were detected by immunohistochemical staining; the expression of MIF mRNA was detected by RT-PCR and the expression of MIF, MMP9 and VEGF proteins were detected by Western blotting in 30 fresh gastric cancer tissues and the corresponding adjacent tissues.

RESULTSOf the 80 gastric carcinoma tissues, 57 (71.25%) showed Hp-L positivity detected by both Gram staining and immunohistochemical staining, as compared with a rate of only 14% in the adjacent normal tissues (P<0.05). The gastric carcinoma tissues showed higher expression levels of MIF, MMP9 and VEGF proteins than the corresponding adjacent normal mucosa; the positivity MIF, MMP-9 and VEGF proteins were significantly higher in Hp-L-positive gastric carcinoma than in Hp-L-negative cases (P<0.05). Positive correlations were found between Hp-L positivity and the expressions of MIF, MMP-9 and VEGF (r=0.598, 0.292, 0.341, respectively, P<0.05). The 30 fresh gastric cancer tissues showed also significantly higher MIF mRNA expression and MIF, MMP-9 and VEGF protein expressions than the adjacent tissues (t=3.729, P<0.01). The expressions of MIF and MMP-9 were also related to the clinicopathological factors including lymph node metastasis and depth of invasion (P<0.05).

CONCLUSIONInfection with L-form of Hp-L can be an important factor that contributes to the invasion and metastasis of gastric carcinoma, the mechanism of which involves up-regulated expressions of MIF, MMP-9 and VEGF.

Adult ; Aged ; Aged, 80 and over ; Female ; Helicobacter Infections ; metabolism ; pathology ; Helicobacter pylori ; Humans ; L Forms ; Lymphatic Metastasis ; Macrophage Migration-Inhibitory Factors ; metabolism ; Male ; Matrix Metalloproteinase 9 ; metabolism ; Middle Aged ; Stomach Neoplasms ; metabolism ; microbiology ; pathology ; Vascular Endothelial Growth Factor A ; metabolism

Objective To prepare human vascular endothelial growth factor receptor 2 (VEGFR2)-targeted liquid perfluorocarbons-filled nanocapsules ultrasound contrast agent,and to investigate its ability of targeting human umbilical endothelial vein cells(HUEVC) and the peculiarity of the enhancing ultrasound imaging in vitro.Methods Rotary evaporation/emulsion technique was used to prepare biotinylated liquid perfluorocarbons-filled nanocapsules (LNCs).VEGFR2-tageted LNCs (V2-LNCs) was further made through conjugating the ligand (biotinylated anti-VEGFR2 antibody) to the surface of LNCs by biotin-avidin system and its appearance,distribution,size and zeta-potential properties were assessed.Immunofluorescent staining assay was used to identify the combination of ligand with the nanocapsules.The ability of V2-LNCs targeting with HUEVC in vitro was evaluated under the fluorescence microscope.Quantitative and statistical analysis were performed to evaluate the ultrasound contrast enhancement of the nanocapsules in vitro.Results The V2-LNCs are uniform and stabilized with size about (98.45 ± 0.07)nm.The ligation of antiVEGFR2 antibody and nanocapsules was positive in immunofluorescent straining assay.In vitro,targeting ability research showed the V2-LNCs could actively adhere to HUEVC,while the control was negative.At the same time,using anti-VEGFR2 antibody to pre-incubation with HUVEC could effectively block the interaction between V2-LNCs and HUEVC.The ultrasound images proved both V2-LNCs and LNCs can significantly enhanced ultrasound imaging contrast agent effect at a concentration of 5 mg/ml in vitro.Conclusions A stable VEGFR2-targeted liquid perfluorocarbons-filled nanocapsules was prepared successfully with biotin-avidin method and effectively bound to HUVEC specially in vitro.Also the liquid perfluorocarbons-filled nanocapsules can strongly enhance the ultrasound contrast in vitro,which might be taken as a kind of vascular ultrasound contrast agent and to get the basic experimental data for the later research.