Objective To evaluate the application of high-performance liquid chromatography (HPLC) in diagnosis and screening of thalassemia. Methods Automated HPLC was used to measure HbF and HbA2 in 100 genetically diagnosed thalas-semic patients and 35 normal children. The results were compared with those from traditional tests including alkali denaturation test and cellulose acetate electrophoresis. The diagnose accordance rates, sensitivity and specificity were compared. Results Seventy-fourβthalassemia, 64 were heterozygous with single mutations and 10 were compound heterozygous with double muta-tions. Twenty-sixαthalassemia, 25 were compound mutations and one was heterozygous with single mutation. The HbF percent-age from HPLC was higher than that from alkali denaturation tests in either thalassemia or normal children (P<0.01). HbF level from HPLC inα-thalassemia was signiifcantly different from that in the normal children (P=0.011). The percentage of HbA2 from HPLC was higher than that from cellulose acetate electrophoresis (P=0.010). HbA2 in the single heterozygousβ-thalassemia were twice higher than that in the double heterozygous mutatedβ-thalassemia (P<0.01). The combination of HbF-HbA2 (≥4.0%) from HPLC with MCV (<80 lf) and MCH (<27 pg) had high accordance rates (99.3%), sensitivity (99.0%) and speciifcity (100.0%) in diagnosis of thalassemia. Conclusions When the results of HPLC are combined with MCV and MCH, it can be applied to the diagnosis of thalassemia with high speciifcity, high sensitivity and has high diagnostic accordance rate with genetic results. HPLC can be an ideal approach to screenβthalassemia.

Aim To investigate the effect of paeoniflorin(PF)on TLR2/4 pathway in AGEs-induced RAW264.7 macrophages.Methods RAW264.7 macrophages were incubated at different time points in AGEs stimulation,as well as different concentrations of PF,to optimize experimental conditions.RAW264.7 macrophages were randomly divided into five groups: control group(DMEM),bull serum albumin(BSA)group(200 mg·L-1 BSA),AGEs group(200 mg·L-1 AGEs),paeoniflorin group(200 mg·L-1 AGEs+10-5 mol·L-1 PF)and TLR2/4 inhibitor group(200 mg·L-1 AGEs+30 mg·L-1 OxPAPC).The expression of Toll-like receptor 2(TLR2),Toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88),p-IRAK1,TIR-domain containing adaptor protein-inducing IFN-β(TRIF),interferon regulatory factor 3(IRF3),p-IRF3,NF-κB p-p65,NF-κB p65,inducible nitric oxide synthase(iNOS),tumor necrosis factor-α(TNF-α),interleukin-l β(IL-1β)and monocyte chemotactic protein-1(MCP-1)were measured by Western blot.Real-time PCR was used to detect the expression of TLR2 and TLR4 mRNA,while TNF-α,IL-1β and MCP-1 levels in cell supernatant were measured by ELISA.Results Compared with control group,AGEs significantly increased the expression of TLR2,TLR4,MyD88,p-IRAK1,TRIF,IRF3,p-IRF3,NF-κB p-p65,NF-κB p65,iNOS,TNF-α,IL-1β and MCP-1 proteins(P<0.01),as well as TLR2 and TLR4 mRNA(P<0.01).TNF-α,IL-1β and MCP-1 contents were also elevated in cell supernatant(P<0.01).The effects induced by AGEs were decreased significantly in PF and TLR2/4 inhibitor group(P<0.01).Conclusion PF plays an anti-inflammatory effect via inhibiting TLR2/4 pathway on macrophages,which may provide a new theoretical basis for the treatment of diabetic nephropathy.

Objective:To investigate the expression of HIF-1α, HIF-2αand MT in human papillary thyroid carcinoma ( PTC) and the association of their expression with clinicopathological indicators .Methods:Immunohistochemistry was used to analyze the ex-pression of HIF-1α, HIF-2αand MT in 70 PTC samples .The correlations of HIF-1α, HIF-2αand MT expression with one another , and with several clinicopathological indicators were statistically analyzed .Results:In 70 PTC samples, the positive expression rates of HIF-1α, HIF-2αand MT were 52.86%(37/70), 50.00%(35/70) and 44.29%(31/70), respectively.HIF-1α, HIF-2αand MT expression had significant correlations with cervical lymph node metastasis (P=0.034, P=0.022, and P=0.032, respectively). Meanwhile, HIF-1αexpression had a positive correlation with HIF-2α(rs =0.258, P=0.031) and MT (rs =0.266, P=0.026). HIF-2αand MT expression were positively correlated (rs=0.259, P=0.030).Concomitant expression of any two or all of the three molecules had stronger correlation with lymph node metastasis than did each alone ( P=0.004 for HIF-1α/HIF-2α, P=0.024 for HIF-1α/MT, P=0.029 for HIF-2α/MT, P=0.017 for HIF-1α/HIF-2α/MT).Conclusion:HIF-1α, HIF-2αand MT expression in PTC samples have a closely correlation , which are related to cervical lymph node metastasis .Therefore, the expression of HIF-1α, HIF-2αand MT might be used as biomarkers for cervical lymph node metastasis of PTC .

Objective: To investigate arthroscopic treatment for acute acromioclavicular dislocation by using Twin Tail TightRope combined with distal joint capsular repair. Methods: The clinical data of 40 patients with acromioclavicular dislocation treated between February 2016 and December 2017 were retrospectively analyzed. The patients were divided into arthroscopic group (20 cases, using arthroscopic Twin Tail TightRope combined with distal joint capsular repair for anatomical repair of stable structure of acromioclavicular joint) and control group (20 cases, treated with clavicular hook plate internal fixation) according to different surgical methods. There was no significant difference in gender, age, cause of injury, Rockwood classification, time from injury to operation, preoperative visual analogue scale (VAS) score and Constant score between the two groups ( P>0.05), which were comparable. Postoperative VAS score and Constant score were used to assess shoulder function and re-dislocation was also observed. Results: The incisions of the two groups healed by first intention, and no early postoperative complications occurred. All patients were followed up 12-18 months (mean, 13.5 months). Postoperative X-ray films showed good anatomical reduction in both groups, but the clavicular hook had a presense in the subacromial space in control group. All patients in arthroscopic group achieved satisfactory shoulder function and returned to work after operation; there was no obvious pain, no complications such as exposure of implant after operation, and no need to remove the implant. In the control group, 4 patients had obvious subacromial impingement pain after operation, and 1 patient had re-dislocation after removal of internal fixator at 1 year after operation; the rest had no complications related to internal fixation, and the internal fixators were removed at 1.0-1.5 years after operation, without re-dislocation. The VAS score and Constant score at 3 months and 1 year after operation in both groups significantly improved when compared with those before operation, and further improved at 1 year after operation ( P<0.05). The VAS score and Constant score at 3 months and 1 year after operation in arthroscopic group were significantly better than those in control group ( P<0.05). Conclusion: Arthroscopic treatment for acute acromioclavicular joint dislocation by using Twin Tail TightRope combined with distal capsular repair is more effective than traditional incision surgery and can obtain more satisfactory results in patient compliance and function recovery because of minimally invasive surgery.

Objective:To establish an HPLC-MS/MS method for the determination of vinblastine in rat plasma. Methods:Aceto-nitrile was used to precipitate protein in the samples after the addition of internal standard, and then the concentration was analyzed by HPLC/MS/MS. All the separations were carried out on an Ultimate C18 column (150 mm × 2. 1 mm, 5. 0 μm). The mobile phase was composed of acetonitrile and 10 mmol·L-1 ammoniumacetate (containing 0. 1% formic acid) (49 ∶51) and was pumped at a flow rate of 0. 3 ml·min-1 under 40 °C. The detection was performed with multiple reactions monitoring (MRM) using electrospray ioniza-tion (ESI). The precursor/product ion transitions were monitored at m/z 811. 4→m/z 224. 2 (positive ion mode) for vinblastine and m/z 825. 4→m/z 807. 4(positive ion mode) for internal standard vincristine. Results:Good linearity of vinblastine was obtained with-in the range of 0. 457-950 ng·ml-1(r=0. 997 1). The lower limit of quantification was 0. 457 ng·ml-1. The extraction recoveries were within the range of 89. 15%-95. 28%. The precision of intra-and inter-day was not more than 7. 95%. T1/2 of vinblastine in rats was (5. 86 ± 2. 37) h, and AUC(0-t) and AUC(0-∞) was (68. 45 ± 14. 51) and (95. 03 ± 33. 09)μg·L-1 ·h, respectively. Conclu-sion:The method is fast, sensitive and accurate, which provides research basis for the development of vinblastine and transporters re-search in medicine. The concentration of vinblastine in rats is low, and the half-life is long.

OBJECTIVE:To establish physiological pharmacokinetic (PBPK) model of cefdinir in healthy volunteers,and to predict pharmacokinetic process of cefdinir in volunteers after oral administration. METHODS:Using“toubao dini”“cefdinir”“logP”“pKa”as keywords,related literatures about physico-chemical constants of cefdinir were retrieved from CNKI,ScienceDi-rect,PubMed and other databases;according to related guidelines and preliminary clinical trial plan of FDA,GastroPlusTM 8.6 soft-ware was used to establish PBPK model of oral administration of cefdinir;the effectiveness of the model was evaluated by multiple error. The model was used to simulate the absorption of cefdinir in the gastrointestinal tracts. The bioequivalence of test preparation and reference preparation were evaluated through single and population(n=500)simulation tests using cmax and AUC0-∞ of cefdinir reference preparation (capsule and granular formulation) as factors when release rate t85%=15 min (i.e. accumulatively released 85% within 15 min). RESULTS:The blood concentration-time curves of cefdinir predicted by PBPK model fitted well with mea-sured value(R2≥0.95);the pharmacokinetic parameters(cmax,tmax,AUC0-∞)were close to measured results,and the multiple er-rors were less than 2. After oral administration,cefdinir was mainly absorbed by the intestinal tract (45.6%),especially by seg-ment 1 of jejunum(14.8%);the absorption amount was significantly lower than the release amount of absorption site,and reached the maximal value(about 40%)within 4 h. The results of single simulation test showed that there was no statistical significance in cmax and AUC0-∞ between cefdinir test and reference preparations (P>0.05). The results of population simulation test showed that the relative bioavailability of cefdinir test particle and test capsule respectively were 99.01%-102.99% and 97.60%-105.90%;90%CI of cmax and AUC0-∞ values were within 80%-125% of reference preparation. CONCLUSIONS:The PBPK model is accurate and reliable in this study,can provide reference for pharmacokinetic study and bioequivalence evaluation of cefdinir preparations. Test preparation and reference preparation are equivalent.

Objective:To establish an LC–MS/MS method for the determination of the active metabolite(SN-38) and secondary metabolite(SN-38G) of irinotecan in rat liver microsomes incubation system, and optimize the incubation conditions. Methods:Meth-anol was selected to precipitate protein in the samples, and then the concentrations were analyzed by LC–MS/MS. All the separation was carried out on a ZORBAX Eclipse XDB-C18 column(2. 1 mm × 50 mm, 3. 5 μm) with the mobile phase of acetonitrile – water (containing 0. 1% formic acid) (23 :77) at a flow rate of 0. 3 ml·min-1. The mass spectrometer was operated with multiple reac-tions monitoring ( MRM) using electrospray ionization ( ESI) . The incubation conditions were optimized by single factor design. Re-sults:SN-38 and SN-38G showed a good linearity ( r≥0. 9972) respectively within the range of 2. 3-920 ng·ml-1 and 2. 5-1000 ng ·ml-1. The intra-and inter-day RSD was below 14. 6%(n=6). The average recovery was within the range of 74. 1%-123. 4% with RSD below 13. 5% (n=6). The optimal incubation conditions were as follows:the concentration of liver microsomal protein was 0. 3 mg·ml-1 and the incubation time was 30 min. Conclusion:The method is rapid, sensitive and accurate in the quantification of SN-38 and SN-38G in the incubation system,which provides methodological basis for the activity determination of UGT1A1 enzyme in vitro.

Objective:To explore effect of human umbilical cord mesenchymal stem cells(hUC-MSCs)on macrophage M1/M2 polarization.Methods:hUC-MSCs were co-cultured with pTHP-1 cells which were macrophage-like cells induced by PMA and tran-scriptome sequencing data were analyzed.Differentially expressed genes were screened and analyzed by GO and KEGG enrichment analysis.Effect of hUC-MSCs on pTHP-1 cells proliferation was analyzed by cell proliferation assay(CCK-8 and EdU).Flow cytometry was used to verify influence of hUC-MSCs on relative contents of inflammatory cytokine TNF-α and anti-inflammatory cytokine IL-10 in pTHP-1 cells which were interaction with LPS.Effect of hUC-MSCs on M1/M2-related molecular phenotype of pTHP-1 cells was studied by qRT-PCR and flow cytometry.Results:Transcriptome sequencing data analysis showed that M1-related genes TNF-α(P<0.05)and HLA-DRA(P<0.01)decreased to a great extent and M2-related gene ARG1(P<0.05)increased to a great extent in pTHP-1 cells after co-culture with hUC-MSCs,suggesting that hUC-MSCs inhibited macrophage M1 polarization.GO and KEGG analysis showed that these dysregulated genes regulated inflammation and immune response.hUC-MSCs inhibited proliferation of pTHP-1 cells,reduced content of TNF-α and increased content of IL-10(P<0.001).qRT-PCR and flow cytometry showed mRNA expressions of HLA-DRA(P<0.05)and CD68(P<0.01)and CD14+CD11c+M1 macrophage percentage were down-regulated,while mRNA expressions of CD163(P<0.001),CD206(P<0.001)and CD14+CD163+M2 macrophage percentage were significantly up-regulated in pTHP-1 cells after co-culture with hUC-MSCs.Conclusion:hUC-MSCs inhibit macrophage polarization to M1 and promote polariza-tion to M2 in vitro.

Objective To evaluate the inhibitory effect of asiaticoside on bleomycin-induced skin cicatrization. Methods Thirty male C57BL/6 mice were randomly divided into three groups:negative control group,model control group,and asiaticoside group,ten in each group.In model control group and asiaticoside group,1 mg·mL-1bleomycin was subcutaneously injected into the dorsal skin of mice every day;4 h later,1 mL 0.9% sodium chloride solution 1 mL asiaticoside(20 mg·mL-1) was injected into the lesion skin in the model control group and the asiaticoside group,respectively.In the negative control group, the same volume of 0.9% sodium chloride solution was subcutaneously injected into the dorsal skin of the mice at the two time points every day.After 21 days,skin specimens were harvested to observe the histomorphology and detect myofibroblast proliferation and expression of inflammatory factors. Results The skin scar was significantly attenuated in the asiaticoside group as compared with the model control group,and the dermal thickness measured exhibited a gradual decrease in asiaticoside group.The expression of α-antismooth muscle antisbidy and infiltration of inflammatory cells were significantly lower in the asiaticoside group than in the model control group. Conclusion Asiaticoside inhibits the development of skin scar of mice by regulating proliferation and differentiation of myofibroblasts and down-regulating inflammatory cells.

Objective:To investigate the efficacy and safety of intravenous thrombolysis with recombinant tissue plasminogen activator (rt-PA) in patients with maintenance hemodialysis (MHD) and acute ischemic stroke.Methods:The clinical data of 235 patients with acute ischemic stroke receiving MHD were collected in our hospital from March 2018 to October 2021. According to the treatment methods chosen by themselves, these patients were divided into control group ( n=70, only receiving standardized secondary stroke prevention), rt-PA low-dose group ( n=85, receiving rt-PA intravenous thrombolysis, 0.6 mg/kg) and rt-PA standard-dose group ( n=80, receiving rt-PA intravenous thrombolysis, 0.9 mg/kg). The effective rate 24 h after treatment, good efficacy rate 7 d after treatment, and good prognosis rate and mortality 90 d after treatment were used to evaluate the effectiveness. The incidences of intracranial hemorrhage, symptomatic intracranial hemorrhage, and severe extracranial hemorrhage 90 d after treatment were used to evaluate the safety. Results:There was no statistical difference in the good prognosis rate 90 d after treatment among the rt-PA low-dose group, the rt-PA standard-dose group and the control group (71.8%, 68.8%, and 64.3%; P>0.05), but the effective rate 24 h after treatment and good efficacy rate 7 d after treatment in the rt-PA low-dose group and rt-PA standard-dose group (44.7%, 57.7%; 46.3%, 62.5%) were both significantly higher than those in the control group (27.1%, 38.6%; P<0.05). The mortality 90 d after treatment in the rt-PA low-dose group (7.1%) was significantly lower than that in the rt-PA standard-dose group (22.5%) and control group (21.4%, P<0.05). The incidences of intracranial hemorrhage and symptomatic intracranial hemorrhage in the rt-PA low-dose group (8.2%, 3.5%) were significantly lower than those in the rt-PA standard-dose group (22.5%, 16.3%; P<0.05), and the incidences of extracranial complications and gastrointestinal bleeding (5.9%, 1.2%) were significantly lower than those in the rt-PA standard-dose group (18.8%, 10.0%; P<0.05). Conclusion:Intravenous thrombolytic therapy with 0.6 mg/kg rt-PA is recommended for acute ischemic stroke patients receiving MHD.