Objective:To establish an identical UPLC-MS/MS method for determination of related substances of 15 drugs(bezafibrate,acipimox,gliquidone,glimepiride,glipizide,glibenclamide,enalapril,captopril,irbesartan,amlodipine,felodipine,nimodipine,nitrendipine,nisoldipine,nifedipine).Methods:The chromatography was performed on Waters ACQUITY UPLCHSS T3(2.1 mm × 100 mm,1.8 μm)column,mobile phase was 0.01 mmol·L-1 Ammonium formate solution containing 0.1％formic acid and methanol with gradient elution.The flow rate was 0.3 mL·min-1,and the injection volume was 2 μL.The detection was carried out by electrospray ioniza-tion(ESI).Multiple-reaction monitoring(MRM)was employed with switching electrospray ion source polarity in positive and negative mode.Results:The 21 related substances showed good linearity(r ≥0.998 35)in the tested ranges.The precision(RS D≤3.45％)and stability(RSD≤3.48％)of the method were good for the related sub-stances.The average recoveries of the related substances were in the range of 91.81％-101.84％with relative standard deviations RSD≤4.66％.Conclusion:The method is rapid,simple,accurate and highly sensitive,and can be used as a auxiliary test method and confirmatory method for current drug standards.

Objective:To establish an identical UPLC-MS/MS method for determination of related substances of 15 drugs(bezafibrate,acipimox,gliquidone,glimepiride,glipizide,glibenclamide,enalapril,captopril,irbesartan,amlodipine,felodipine,nimodipine,nitrendipine,nisoldipine,nifedipine).Methods:The chromatography was performed on Waters ACQUITY UPLCHSS T3(2.1 mm × 100 mm,1.8 μm)column,mobile phase was 0.01 mmol·L-1 Ammonium formate solution containing 0.1％formic acid and methanol with gradient elution.The flow rate was 0.3 mL·min-1,and the injection volume was 2 μL.The detection was carried out by electrospray ioniza-tion(ESI).Multiple-reaction monitoring(MRM)was employed with switching electrospray ion source polarity in positive and negative mode.Results:The 21 related substances showed good linearity(r ≥0.998 35)in the tested ranges.The precision(RS D≤3.45％)and stability(RSD≤3.48％)of the method were good for the related sub-stances.The average recoveries of the related substances were in the range of 91.81％-101.84％with relative standard deviations RSD≤4.66％.Conclusion:The method is rapid,simple,accurate and highly sensitive,and can be used as a auxiliary test method and confirmatory method for current drug standards.

Objective:To measure the morphological parameters of distal tibiofibular syndesmosis in healthy adults using multi-slice CT (MSCT) so as to provide a reference for the diagnosis of distal tibiofibular syndesmosis injury.Methods:The ankle MSCT imaging data in 110 normal adults were retrieved from the image report database of Cangzhou People′s Hospital from May 2019 to May 2021, including 56 males and 54 females; aged 18-60 years [(38.2±11.0)years]. There were 51 patients with imaging on the right ankle and 59 on the left ankle. Picture archiving and communication system (PACS) was used to measure parameters at 10 mm above the articular surface of the distal tibia on MSCT, including the anterior tibiofibular space (L1), posterior tibiofibular space (L2), middle tibiofibular space (L3), depth of fibula in notch (L4), distance of anterior tibiofibular edge (L5), distance of posterior tibiofibular edge (L6), anterior tibiofibular syndesmosis angle (A1), and fibular rotation angle (A2), and the measurements were compared by sex, age and side. The positive rate of "tibiofibular line" was observed. The morphological classification of distal tibiofibular syndesmosis was performed.Results:There was no significant difference in L1-L6, A1 and A2 among different age and side (all P>0.05). No significant difference was found in L4, L5, A1 and A2 between males and females ( P>0.05), but L1, L2, L3 and L6 were larger in males than in females ( P<0.05 or 0.01). The positive rate of "tibiofibular line" was 80.4% (45/56) in males compared to 74.1% (40/54) in females ( P>0.05), 77.2% (44/57) in the youth compared to 77.4% (41/53) in the middle-aged, and 78.0% (46/59) in the left ankle compared to 76.5% (39/51) in the right ankle (all P>0.05). Morphological classification of distal tibiofibular syndesmosis was crescent in 61 patients (55.5%), trapezoid in 14 (12.7%), I-shaped in 3 (2.7%), M-shaped in 17 (15.5%), V-shaped in 10 (9.1%), Г-shaped in 5 (4.5%). Conclusions:When L1, L2, L3 and L6 are used as references in the diagnosis of adult distal tibiofibular syndesmosis injury, gender factors rather than age or side factors should be considered. Males have wider distal tibiofibular space than females, with the fibula more forward. The "tibiofibular line" has a high positive rate and is not affected by gender, age or sides, providing a new idea for the diagnosis of distal tibiofibular syndesmosis injury and anatomical reduction. There are many variations in the morphology of distal tibiofibular syndesmosis, so it is easy to be misdiagnosed as the separation of distal tibiofibular syndesmosis on X-ray, which should be noted.

Objective:To compare the practical value of three statistical methods in establishing reference intervals by the indirect method.Methods:This is a methodological evaluation study. The data of coagulation parameters were obtained from laboratory information system, which were from pregnant women who had done prenatal examination in Peking University First Hospital from January 2017 to December 2019. The test results from 32 401 pregnant women were collected. Those healthy pregnant women were divided into three groups: early pregnancy group(n=11 151), middle pregnancy group(n=4 872) and late pregnancy group(n=16 378). Statistical analysis was performed for the result of PT, APTT, FIB, TT, D-D and FDP, and the necessity of stratification based on age in different periods of pregnancy was analyzed. Stratification based on age was necessary in three pregnancy groups for PT and APTT, in late pregnancy group for FIB and TT, and in early pregnancy group for D-D. The reference intervals of coagulation parameters were calculated by three statistical methods: non-parametric method, Hoffmann method and Q-Q plot method. Forty-two healthy pregnant women from October 2019 to January 2020 were enrolled as reference individuals for the validation of the reference intervals.The proportions of test results outside the reference intervals in the reference population calculated and compared.Results:The levels of the six coagulation assays vary significantly during the three periods of pregnancy, stratification based on age was necessary in three pregnancy groups for PT and APTT, in late pregnancy group for FIB and TT, and in early pregnancy group for D-D. If the number of test results was large, non-parametric and Hoffmann method provided more similar results, while the reference intervals calculated with Q-Q plot method was slightly wider than Hoffmann method. If the number of test results was small, reference intervals calculated with Hoffmann and Q-Q plot method were more reliable. For pregnant women during early pregnancy under the age of 35, the reference intervals of PT, APTT, FIB and TT calculated by this method were (10.44-13.11)s, (25.29-35.88)s, (2.61-4.64)g/L and (11.53-15.58)s.Conclusion:When establishing the reference interval, stratification according to pregnancy period and age was needed. Hoffmann method can be used as an alternative to the direct method.

Objective To observe the effect of febuxostat on epithelial-to-mesenchymal transition (EMT) of kidney tubules and the levels of serum IL-6 nad transforming growth factor (TGF) β1 in hyperuricemic rats.Methods Forty male SD rats were divided into 4 groups:normal control group (NC group),oteracil potassium group (OP group),oteracil potassium with febuxostat group (OF group) and oteracil potassium with benzbromarone group (OB group).Each group had 10 rats and balanced in body weights.To induce hyperuricemia,rats were given oteracil potassium by gastric garage once a day for eight weeks.Rats in OF group and OB group were given either febuxostat or benbromarone starting with oteracil potassium,and rats in NC group was given saline only.Blood samples were taken before,and at the end of 4 and 8 weeks of the treatments and serum uric acid,creatinine,blood usea nitrogen (BUN),IL-6 and TGFβ1 contents were measured at each time point.Renal pathological changes were observed via HE and Masson staining,and the expression of α-SMA and E-cadherin were detected by immunohistochemistry.Results Compared with those in NC group,the levels of serum uric acid,creatinine,BUN,IL-6 and TGFβ1 in the another three groups were increased significantly (all P ＜ 0.01).However,the IL-6 and TGFβ1 contents in OF group were much lower than those in OP group (P ＜0.01).HE and Masson staining showed that OF group had less damage and tubulointerstitial fibrosis than OP group and OB group (P ＜0.01).Moreover,the expression of α-SMA was significantly down-regulated (P ＜ 0.01) and that of E-cadherin was significantly up-regulated in OF group compared with those in OP group.Conclusion Febuxostat treatment significantly inhibited EMT and reduced the levels of IL-6 and TGFβ1 in hyperuricemia rats.

Objective To study the mechanism of some vegetables inhibiting human platelet aggregation function.Methods Some vegetable juice was mixed(tomato juice as positive control)with platelet-rich plasma(PRP),and maximum ratio of platelet aggregation was measured after induction by agonists on the aggregometer.The expression levels of platelet activation marker were detected,including fibrinogen receptor(Fib-R),P-selectin(CD62P),and the combination amount of fibrinogen on the surface of platelet by flow cytometry to validate the inhibitory effect of platelet aggregation by animal tests.Ordinary white rabbits were randomly divided into five groups,each group was fed with saline,cooked tomato juice,blanched garlic leaves juice,Chinese cabbage juice or spinach juice,respectively.And the maximum ratio of rabbit platelet aggregation in different time were observed.Results Cooked vegetable juice couldn't inhibit human platelet aggregation induced by AA,but blanched garlic leaves juice,Chinese cabbage juice and spinach juice could inhibit human platelet aggregation obviously induced by ADP.collagen or epinephrine.The inhibition intensity of platelet aggregation increased markedly along with the increase of the concentration of the cooked vegetables juice.Cooked juice of blanched garlic leaves,Chinese cabbage,spinach could not inhibit the expression of Fib-R and CD62P,Whereas they were able to significantly decrease the amount of Fib-R and fibrinogen binding. ADP-induced rabbit platelet aggregation ratios in the group fed with cooked juice of blanched garlic leaves,Chinese cabbage or spinach were significantly lower than control group after 3,5,8 weeks,respectively.The inhibition ratios of the platelet aggregation in cooked Chinese cabbage juice group were 45.7%,53.6%,48.5% after 3,5,8 weeks,respectively.Cooked juice of blanched garlic leaves group were 10.7%,66.7%,46.3%,respectively. Cooked spinach juice group were 8.7%,21.0%,42.6%,respectively.The collagen-induced rabbit platelet aggregation ratios in the groups fed with cooked juice of blanched garlic leaves or Chinese cabbage were significantly lower than control group after 5 and 8 weeks respectively. The inhibition ratios of the platelet aggregation in cooked Chinese cabbage juice group were 54.9%,56.3%after 5 and 8 weeks,respectively. Cooked juice of blanched garlic leaves group were 28.4%and 86.7%,respectively. Cooked tomato juice could not inhibit rabbit platelet aggregation in 8 weeks. Conclusions After induced by ADP or collagen,cooked juice of blanched garlic leaves,Chinese cabbage and spinach could significantly decrease the amount of Fib-R and fibrinogen binding,and inhibit platelet aggregation function significantly.It may have potential application of therapy or prlevention of thrombotic diseases.

Objective To establish a new flow cytometric immuno-bead array assay (FCIBA)to detect several platelet-specific autoantibodies simultaneously in a single serum sample.Methods A series of beads of different red fluorescent intensity were used for coating with different anti-platelet membrane protein monoclonal antibodies(anti-GP Ⅱb/a,anti-GP Ⅰ a/Ⅱ a,anti-GP Ⅳ,anti-GP Ⅰ b/Ⅸ and anti-HLAABC)to detect five platelet-specific autoantibodies in serum. The beads captured platelet antigenautoantiboay complex.Subsequently,different platelet-specific autoantibodies in a patient serum can be detected simultaneously by the flow cytometry.In addition,we evaluated the new FCIBA,and compared its results with modified antigen capture ELISA method(MACE).The new FCIBA was used to detect five platelet-specific autoantibodies in serums of autoimmune thrombocytopenic purpura(AITP)and nonautoimmune thromboeytopenic purpura patients.Results The new FCIBA can be used to detect five plateletspecific autoantibodies simultaneously(Anti-GP Ⅱ b/Ⅲ a,Anti-GP Ⅰ a/Ⅱ a,Anti-GP Ⅳ,Anti-GP Ⅰ b/Ⅸand Anti-HLA-ABC).The coefficient of variation(CV)of intra-repetition are 4.82%,6.09%,5.04%,5.73%and 5.30%,respectively.The dilution test results are in good logarithm linearity which are 0.997 2,0.996 6,0.998 8,0.996 5 and 0.998 2,respectively. The resuhs of the new FCIBA are highly correlated with those with MACE method,and the coefficient correlation were 0.928 9,0.922 4,0.889 4,0.910 0 and 0.913 4,respectively(P＜0.01).51.69%samples of AITP patients show positive for platelet-specific autoantibodies as detected by the new FCIBA.Among the AITP patients,the positivity of specific autoantibodies for anti-GPⅡb/ Ⅲ a,anti-GP Ⅰ a/Ⅱ a,anti-GP Ⅳ,anti-GP Ⅰb/Ⅸ and anti-HLA-ABC were 40.82%,24.45%,19.39%,32.65%and 17.35%.Among the 40 non-autoimmune thrombocytopenic purpura patients,none of platelet-specific autoantibodies in serum samples can be detected.Conclusion A new FCIBA is established successfully to detect five platelet-specific autoantibodies coefficient correlation.

Objective To investigate the correlation of prognosis with the immunophenotype in acute myeloid leukemia(AML)patients.Methods Immunophenotyping was performed in 131 patients with AML by mtdtieolor flow cytometry.Correlation of immunophenotype with other laboratory parameters such as initial white blood cell count(WBC),platelet count(PLT),hemoglobin(Hb),and the complete remission(CR)ratio was analyzed.Results In these AML patients,myeloid antigens CDl3,CD33 and myeloperoxidase(MPO)were more highly expressed than other antigens.Expression of CD34 and HLA-DR were lower in acute promyelocytic leukemia(M3)subtype.The expression of lymphocyte antigen CD19 and CD7 were the highest.CD7 expression was associated with age(t=-2.27,P＜0.05).CD14 was associated with initial WBC(Z=-2.284,P＜0.05).The overall CR ratio was 56.5%among all patients.CD34positive patients had a significantly lower CR rato(45.1%),compared with the CD34 negative patients whose CR ratio was 75.6%(x2=11.524,P=0.001).The CR ratio was significantly lower in cases expressing both CD34 and HLA-DR(74 patients with CR rate 41.9%)than in cases expressing only CD34 or HLA-DR(38 patients with CR rate 78.9%)and both negative(19 patients with CR rate 68.4%)(Z=-3.492,P＜0.01).However,other antigens,including CD7,CD19,CD13,CD33,CD38,CD15,CD64,CD14 and MPO had no significant association with CR ratio.Logistic regression analysis showed that age(60 years or older),initial WBC(more than 50 x 109/L),PLT(more than 30 x 109/L),Hb(less than 60g/L)and CD34 were independent adverse factors for achieving CR.Conclusions In AML patients,age,initial WBC,PLT,Hb and CD34 are associated with CR ratio.Detection of immunophenotype may help to estimate prognosis of patients with AML and guide the treatment of AML.