BACKGROUND: Troponin I (Tn I ) could inhibit the growth of vascular endothelial cells, inhibit neovascularization,through which to inhibit the development and metastasis of solid tumor. Similar to Tn I, TnC also exists in non-muscular tissue, but does it has the analogous activity of anticancer like Tn I ?OBJECTIVE: To explore the effect of recombinant human TnC (rhTnC) on the growth of human umbilical vein endothelial cells (HUV-EC) and mouse xenograft tumor.DESIGN: Controlled observation in vivo and in vitro.SETTING: Research Institute of Medicine, Chongqing K.E.W Pharmaceutical Co., Ltd. and Department of Biochemistry of Chongqing Medical University.MATERIALS: The experiment was conducted in the Biochemical Laboratory of Research Institute of Medicine,Chongqing K.E.W Pharmaceutical Co., Ltd. from March 2003 to December 2004. 100 Kunming mice either male or female of 15-22 g purchased from Chongqing Academy of Chinese Materia Medica. E.coli BL21 (DE3)pLysS/pET3b-TnC provided by Chongqing K.E.W Pharmaceutical Co., Ltd. HUV-EC (Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences).METHODS: Human TnC cDNA was obtained from human thymus cDNA library using PCR. The colony was cloned in E.coli and a bacterial strain of gene engineering E.coli BL21 (DE3) pLysS/pET3b-TnC was obtained, which could express hTnC. The recombinant human TnC (rhTnC) was purified with affinity chromatography of Ni-NTA agarose. ①In vitro cell experiment: HUV-ECs were seeded in the 96-well plates at density of 2×103 cells per well and co-cultured with rhTnC of 1, 5, 10, and 50 mg/L for 3 days. The absorption (A value) was detected with microplate reader at 540 nm and the inhibition rate of cell growth was calculated. Meanwhile, the 50% inhibiting dose (IC50 value) was assayed by LOGIT method. ②In vivo animal experiment: Ascites tumor (S-180) that had been inoculated for 7-8 days was harvested. The tumor cells were diluted to 1 ×1010 L-1 and 0.2 rnL was subaxillarily and intraperitoneally injected into each mouse (50 mice in each group). The next day, the mice were randomly divided into 5 groups: rhTnC 20 mg/kg group, rhTnC 10 mg/kg group, rhTnC 5 mg/kg group, Cyclophosphamide (Cy) group and control group with 10 mice in each group. The rhTnC 20,10 and 5 mg/kg groups were given administration at the corresponding doses, once a day for 7 days; 50 mg/kg Cy was given the Cy group one after an interval of day, and the same volume normal saline was given to the control group. One day after the last time of administration, all mice were killed and the tumor was harvested and weighed. The inhibition rate of tumor growth was calculated: tumor inhibition rate=[(Average weight of tumor in control group-Average weight of tumor in drug group)/Average weight of control group]×100%.MAIN OUTCOME MEASURES: Inhibition rate of rhTnC to HUV-EC proliferation in cell experiment in vitro and mouse xenograft tumor in animal experiment in vivo.RESULTS: ①In vitro cell culture showed that rhTnC suppressed HUV-EC proliferation in a dose-dependent manner (IC50=7.5 mg/L). ②Similar to the result of in vitro cell experiment, after intraperitoneal administration, the inhibition rate of rhTnC 5, 10, and 20 mg/kg groups was higher than that of control group (P ＜ 0.01); after subaxillary administration, the inhibition rate of rhTnC 5, 10, and 20 mg/kg groups was also higher than that of control group (P ＜ 0.05-0.01). There was no significant difference in the inhibition rate between two administration approaches (P ＞ 0.05).CONCLUSION: rhTnC is capable of inhibiting the proliferation of HUV-EC dose-dependently, and displays the activities of inhibiting the proliferation of HUV-EC and anti-tumor.

Intervertebral disc (IVD) degeneration is one of the major causes of low back pain. As current clinical treatments are aimed at restoring biomechanical function and providing symptomatic relief, the methods focused on biological repair have aroused interest and several tissue engineering approaches using different cell types have been proposed. Owing to the unsuitable nature of degenerate cells for tissue engineering, attention has been given to the use of mesenchymal stem cells (MSCs). In this connection, we have made a study on the characteristics of MSCs derived from adult bone marrow and on the feasibility of constructing IVD tissue-engineering cell under a Three-Dimensional Pellet Culture System. The human bone marrow MSCs were isolated and purified with density gradient solution and attachment-independent culture system. MSCs isolated using this method are a homogeneous population as indicated by morphology and other criteria. They have the capacity for self-renewal and proliferation, and the multilineage potential to differentiate.
Adolescent ; Adult ; Bone Marrow Cells ; cytology ; Cell Culture Techniques ; methods ; Cells, Cultured ; Chondrogenesis ; physiology ; Humans ; Intervertebral Disc ; Intervertebral Disc Degeneration ; therapy ; Mesenchymal Stromal Cells ; cytology ; Tissue Engineering ; methods ; Young Adult

Objective:To evaluate the efficacy and safety of azacytidine (AZA) combined with homoharringtonine (HHT) and low-dose cytarabine (LDAC) in the treatment of acute myeloid leukemia (AML) patients with 3+ 7 conventional regimen intolerance.Methods:A retrospective analysis was conducted on the clinical characteristics, efficacy, prognosis, and adverse events of 33 AML patients (15 initially diagnosed and 18 relapsed/refractory) admitted to the Second Xiangya Hospital of Central South University.Results:Among the 33 AML patients treated with this regimen, the median age was 55 years old, 9 patients had a moderate cytogenetic risk, and 18 patients had a high cytogenetic risk. Among the 33 patients, 3 were lost to follow-up and 1 had incomplete data. Among the remaining 29 patients who received AZA+ HHT+ LDAC treatment, the total complete response (CR) rate was 69.0%(20/29), and the total response rate (ORR) was 79.3%(23/29); The median progression free survival (PFS) was 7.0 months. Among the subgroup analysis, including age, gender, Eastern Cooperative Oncology Group (ECOG) score, disease classification, bone marrow progenitor cells, peripheral blood leukocytes, risk stratification, and epigenetic abnormalities, only CR rates and PFS differences were statistically significant among different ECOG scoring groups ( P=0.048; P=0.021). A total of 29 patients underwent 69 AZA+ HHT+ LDAC chemotherapy cycles. Retrospective grading was performed on 69 cycles based on common toxicity criteria for adverse events (CTC AE version 5.0). The most common grade Ⅲ-Ⅳ hematological adverse events were thrombocytopenia (54/69, 78.3%) and granulocytopenia (48/69, 69.6%). Common non hematological adverse events included nausea (19/69, 27.5%), infection (17/69, 24.6%), and hypokalemia (18/69, 26.1%). Conclusions:AZA combined with HHT and LDAC has a good therapeutic effect in the treatment of acute myeloid leukemia, and adverse reaction events are controllable.

Objective To establish a two-dimensional high-performance liquid chromatography method for the determination of tigecycline in human cerebrospinal fluid, which can be used for the drug monitoring in patients with intracranial infection. Methods The quantification was carried out by an external standard method. The first-dimension column was a Aston SNX5 phenyl chromatographic column (50 mm×4.6 mm, 5 μm) with ammonium phosphate (pH was adjusted with ammonium hydroxide to 7.5)-methanol (45∶55, V/V) as the mobile phase and the flow rate was 1.2 ml/min. The second-dimension chromatographic column was Aston SC5 C₁₈ (275 mm×4.6 mm, 5 μm), with ammonium phosphate (pH was adjusted with ammonium hydroxide to 7.4)-ammonium phosphate (pH was adjusted with ammonium hydroxide to 3.0)- acetonitrile (30∶50∶20, V/V/V) as the mobile phase and the flow rate was 1.0 ml/min. The detection wavelength was 340 nm. The temperature was 40 ℃ and the injection volume was 200 μl. Results The calibration curve of tigecycline showed good linearity from 64.5 to 1 290.0 ng/ml in human cerebrospinal fluid (r=0.999 8). The RSD of intra and inter-day precision were less than 5.0% with the detection accuracy of 98.80%−106.51%. Conclusion This method is simple, quick, accurate, specific and sensitive. It meets the requirements of tigecycline determination in clinical human cerebrospinal fluid, which offers the individualized therapeutic assurance for patients with intracranial infection.