Objective To investigate the influence of sulfated propylene glycol alginate(PSS) and its fractions on the immunoregulation.Methods The immunoregulation activity of PSS and its fractions were investigated by using immunocyte cultivation technique in vitro.The structure activity relationship was analysed on the basis of the structure studies of PSS' fractions.Results The experimental results showed that PSS could improve spleen cell proliferation,enhance macrophage phagocytic function and inhibit T-cell and B-cell proliferation.Conclusion PSS possessed significant immunoregulation effect,whilst the immunocompetence comparison of PSS' fractions proved that the different immunocytes had different requirements for saccharides length.

Aim To study effect of the extract of Fagopynum cymosum(Fr4) on the apoptosis and telomeraseactivity in human leukemia HL-60 cells.Methods HL-60 cells were treated with Fr4 and inhibition of proliferation was measured with MTT assay.Cell morphology before and after the treatment was examined with light and electron microscopy.FILC-Annexin-V and PI double staining was used to detect cell membrane-mediated apoptosis by FCM.DNA fragmentations were analyzed with DNA gel electrophoresis.Furthermore,telomerase activity was determined with PCR-ELISA-based telomeric repeat amplification(TRAP).Results The proliferation of HL-60 cells was inhibited by Fr4 with an IC_(50) of 115 mg?L~(-1).Typical morphology and biochemical feature of apoptosis was observed with Fr4 60～240 mg?L~(-1).The percentages of early apoptosis cells were increased in a dose-dependent manner.Telomerase activity was decreased in a dose-dependent manner during the process of apoptosis.Conclusion Fr4 induced apoptosis in HL-60 cells.Down-regulation of the telomerase activity in HL-60 cells might be contributed to Fr4-induced apoptosis.

AIM:To investigate the effect of fenbendazole (FBZ) on the proliferation of human chronic myelogenous leukemia (CML) cell line K562.METHODS:The CCK-8 assay was used to detect the effect of FBZ on viability of the K562 cells and normal peripheral blood mononuclear cells (PBMC).The cell growth was measured by the method of Trypan blue exclusion.The cell cycle was analyzed by flow cytometry.The cell cycle-related proteins were detected by Western blot.RESULTS:The growth of K562 was significantly inhibited by FBZ.However, it elicited little cytotoxic effect on PBMC.Furthermore, FBZ induced G2/M phase arrest and mitotic catastrophe in the K562 cells based on the changes of nuclear morphology, DNA content, mitotic marker analysis and the number of polykaryocytes.CONCLUSION:Fenbendazole significantly inhibits the proliferation of K562 cells and induces cell cycle arrest at G2/M phase by the regulation of cell cycle-related proteins.

Objective:To investigate the effects of triptolide on radiosensitization of lung cancer A549 cells and the underlying mechanism.Methods:During June-September 2019, lung cancer A549 cells were treated with different concentrations of triptolide for 24 and 48 hours in Animal Experiment Center, Zhejiang Chinese Medical University, China. The inhibitory effects of triptolide on the proliferation of lung cancer A549 cells were determined using MTT method. Appropriate concentrations of triptolide and double distilled water were added to the experimental and control groups, respectively. The effects of triptolide on radiosensitization of lung cancer A549 cells was determined by colony formation assay. Radiosensitization ratio was calculated. Lung cancer A549 cells were divided into blank control, triptolide, radiotherapy, and radiotherapy + triptolide groups. The effects of triptolide on apoptosis and cell cycle of lung cancer A549 cells were determined by flow cytometry.Results:The 10% inhibitory concentration (IC 10) and half maximal inhibitory concentration (IC 50) of triptolide for treating lung cancer A549 cells were 36.61 nmol/L and 259.38 nmol/L, respectively at 24 hours, and they were 9.05 nmol/L and 61.49 nmol/L, respectively at 48 hours. Triptolide had an radiosensitization effect on lung cancer A549 cells, with the radiosensitization ratio of 1.135. The apoptosis rate in the radiotherapy + triptolide group was significantly higher than that in the radiotherapy [(45.47 ± 8.29)% vs. (5.25 ± 0.59)%, t = 6.847, P = 0.002]. The proportion of lung cancer A549 cells at the G2/M phase in the radiotherapy group was significantly higher than that in the radiotherapy + triptolide group [(27.82 ± 0.96)% vs. (11.98 ± 0.55)%, t = 20.176, P < 0.05]. The proportion of lung cancer A549 cells at the G2/M phase in the black group was significantly higher than that in the triptolide group [(17.31 ± 3.42)% vs. (8.05 ± 0.71)%, t = 3.749, P = 0.02]. Conclusion:Triptolide has a radiosensitization effect on lung cancer A549 cells, and the underlying mechanism may be related to its participation in cell apoptosis and cycle regulation.

Objective: To analyze the clinical value of sirolimus plus prednisone for the treatment of the refractory kaposiform hemangioendothelioma(RKHE) and Kasabach-Merritt syndrome(KMS). Method: Clinical retrospective analysis was carried out for 10 patients recruited in Children′s Hospital Affiliated to Capital Institute of Paediatrics from January 2014 to January 2017 who were non responders to or relapsers after the treatment of propranolol, prednisone, pingyangmycin and lauromacrogol(5 cases RKHE, 5 cases RKHE plus KMS, age ranged from 6 days to 9 years); patients were treated with sirolimus at the dosage of 0.035 ml/(kg·d), once a day, for 6-410 days; the diagnosis of 10 patients were confirmed by pathological biopsy and immunohistochemical examination(IHC); the difference of the coagulation parameters and the platelet counts, the size of tumor and ecchymosis at different stages of treatment were recorded and measured by scale and ultrasonography; the side effects of sirolimus were recorded as well. Result: Clinical characteristics of 10 cases (6 male and 4 female) RKHE with KMS were refractory dark red hard hemangioma or ecchymosis, the platelet counts were lower than 30.0×10⁹/L, (15±7)×10⁹/L, coagulation tests were obviously abnormal, fibrinogens were significantly decreased(0.8±0.5)g/L, the fibrin lysates and D-dimer were significantly increased(100±23)mg/L, (10 000±2 200)ng/L, the prothrombin time and activated partial thromboplastin time were prolonged(25.0±2.1)s, (58.0±3.4)s. The pathologic characteristics of the tumors were similar: spindle tumor cells, mass distribution and deeply stained nuclei tumor cells. IHC revealed positive staining for D2-40, CD31 and CD34. Stainings for factor Ⅷ and GLUT-1 were negative. In five cases RKHE plus KMS were treated with sirolimus and prednisone, after (6.5±0.7) days treatment, the platelet counts were obviously increased(72.0±0.6)×10⁹/L, coagulation parameters were obviously improved, fibrinogen significantly increased(1.5±0.2)g/L, the fibrinlysates and D-dimer significantly decreased(7±3)mg/L, (2 300±200)ng/L, the prothrombin time and activated partial thromboplastin time were prolonged(15±2.3)s, (42±3.4) s, and the sizes of tumor and ecchymosis were slightly shrunken 18%±3%, 38%±5%; after (30±5.7) days treatment, the platelet counts and coagulation parameters returned to normal(146±36)×10⁹/L, and the size of tumor and ecchymosis were obviously shrunken 73%±3%, 97%±3%; after (3±0.4) months treatment, the tumor was obviously shrunken by 93%±2% and no longer palpable. In five cases with RKHE without KMS manifested stubborn dark red hard hemangiomatous plaques, coagulation tests and platelet were obviously normal, these patients were treated with sirolimus, after (2.0±0.6) months treatment, the tumor became shrunken 8%±3%, with continuous treatment the tumor shrunk gradually, after (4.0±3.2)months(2-18 mouths) the tumor was not eliminated 51%±7%. Conclusion RKHE and KMS have typical clinical, laboratory and pathological characteristics, sirolimus plus prednisone have remarkable efficacy and minor side effects, it should be recommended for the treatment of KHE with KMS.

Rotundic acid(RA),an ursane-type pentacyclic triterpene acid isolated from the dried barks of Ilex rotunda Thunb.(Aquifoliaceae),possesses diverse bioactivities.To further study its pharmacokinetics,a simple and sensitive liquid chromatography with triple quadrupole mass spectrometry(LC-QqQ-MS/MS)method was developed and validated to quantify RA concentration in rat plasma and tissue using etofesalamide as an internal standard(IS).Plasma and tissue samples were subjected to one-step protein precipitation.Chromatographic separation was achieved on a ZORBAX Eclipse XDB-C18 col-umn(4.6 mm×50 mm,5 μm)under gradient conditions with eluents of methanol:acetonitrile(1∶1,V/V)and 5mM ammonium formate:methanol(9∶1,V/V)at 0.5mL/min.Multiple reaction monitoring transitions were performed at m/z 487.30 → 437.30 for RA and m/z 256.10 → 227.10 for IS in the negative mode.The developed LC-QqQ-MS/MS method exhibited good linearity(2-500 ng/mL)and was fully validated in accordance with U.S.Food and Drug Administration bioanalytical guidelines.Dose proportionality and bioavailability in rats were determined by comparing pharmacokinetic data after single oral(10,20,and 40 mg/kg)and intravenous(10 mg/kg)administration of RA.Tissue distribution was studied following oral administration at 20 mg/kg.The results showed that the absolute bioavailability of RA after administration at different doses ranged from 16.1％to 19.4％.RA showed good dose proportionality over a dose range of 10-40 mg/kg.RA was rapidly absorbed in a dose-dependent manner and highly distributed in the liver.In conclusion,this study is the first to systematically elucidate the absorption and distribution characteristics of RA in rats,which can provide additional information for further development and evaluation of RA in drug metabolism and pharmacokinetic studies.

Clustered regular interspaced short palindromic repeats (CRISPR) system has been widely used in recent years. Compared with traditional genome editing technology, CRISPR/Cas system has notable advantages, including high editing efficiency, high specificity, low cost and the convenience for manipulation. Type Ⅱ and Ⅴ CRISPR/Cas system only requires a single Cas9 protein or a single Cpf1 protein as effector nucleases for cutting double-stranded DNA, developed as genome editing tools. At present, CRISPR/Cas9 technology has been successfully applied to the genome editing of eukaryotes such as zebrafish, mice and human cells, whereas limited progress has been made in the genome editing of bacteria. In our review, we describe CRISPR/Cas system, its mechanism and summarize the optimization and progress of genome editing in bacteria.
Animals ; Bacteria ; CRISPR-Cas Systems ; Endonucleases ; Gene Editing ; Humans ; Mice