OBJECTIVE: To investigate the effect of SARI overexpression on the proliferation and apoptosis of core binding factor leukemia (CBFL) cells and explore the potential molecular mechanisms. METHODS: C-KIT N822K mutation status in Kasumi-1 cell line was detected by exon 17 sequencing. Then the SARI lentiviral vector (pGC-FU-SARI) was constructed, meanwhile Kasumi-1 cells were transfected with the SARI lentiviral vector. Quantitative PCR and Western blot were employed to identify efficacy of SARI overexpression after the transfection of cells. Cells were divided into three groups, including the cells infected with pGC-FU-SARI (OE group), the cells infected with pGC-FU-GFP (NC group) and the untreated cells (blank control group). Cell proliferative activity was tested by MTT assay, cell apoptosis was measured by flow cytometry (FCM) and the expression of apoptosis-related proteins: BCL-2,BAX,Cyto C,Caspase 9,Caspase 3,cleaved-Caspase 3,PARP and cleaved-PARP as well as PI3K/Akt pathway proteins: PI3K（p85）,p-PI3K（p85）,Akt and p-Akt were detected by Western blot. RESULTS: The Kasumi-1 cells were detected to bear c-KIT N822K (T>A) mutation. The Kasumi-1 cells with SARI was overexpression were construeted successfully. Compared with NC group, the cell proliferation was decreased and cell apoptosis was increased; BCL-2 expression was reduced, BAX expression was enharued; cyto C expression appeared; the expression of Caspase 9 and Caspase 3 was down-regulated, the expression of cleaved Caspase 3 was up-regulated; the PARP expression was decreased, cleaved PARP expression was increased; the phosphorylation level of PI3K/Akt pathway proteins: p-PI3K/PI3K, p-Akt/Akt was down-regulated in OE group (P<0.05). CONCLUSION SARI gene may suppress the proliferation of CBFL cells, and induce their apoptosis through the mitochondrial pathway, which may be related with the inhibition of PI3K/Akt pathway.
Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Core Binding Factors ; Humans ; Leukemia ; Phosphatidylinositol 3-Kinases ; Proto-Oncogene Proteins c-akt

OBJECTIVE: To investigate the role of nucleophosmin (NPM) in the proliferation of chronic myeloid leukemia cells (K562 cells) and its mechanism by RNAi technology. METHODS: shRNA was used to inhibit the expression of NPM. The expression of NPM gene was detected by real-time quantitative PCR. The effect of inhibiting NPM gene on cell proliferation was detected by MTS assay. Change of cell cycle was detected by flow cytometry. Western blot was used to detect the expression of cell cycle-related proteins. RESULTS: The shRNA lentiviral vector targeting at NPM gene was successfully constructed and used to transfect the K562 cells. The results showed that compared with the control groups, suppression of NPM gene expression in K562 cells could inhibit the cell proliferation and decrease the cell colony formation. Moreover, interference of NPM gene could prolong G/G phase and arrest cell cycle, which may be related to the down-regulation of NPM gene expression and activation of p21 protein expression, thereby inhibited the formation of CDK2/ Cyclin E complex. CONCLUSION Down-regulation of NPM gene expression in K562 cells can induce cell cycle arrest and inhibit cell proliferation.
Apoptosis ; Cell Proliferation ; Gene Knockdown Techniques ; Humans ; K562 Cells ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; Nuclear Proteins

OBJECTIVE: To study the effect of soothing liver, relieving depression, invigorating spleen and reinforcing blood on reducing delirium in elderly patients with hip fracture. METHODS: From December 2014 to June 2018, 180 elderly patients with hip fracture admitted were divided into treatment group and placebo group according to the order of admission:90 patients in treatment group were treated with Jiawei Xiaoyao Powder(JWXYP), including 32 males and 58 females, with an average age of(72.12±4.92), involving 67 cases of femoral trochanter fractures and 23 cases of femoral neck fractures; 35 cases underwent dynamic hip screw fixation, 31 cases underwent intramedullary fixation and 24 cases underwent artificial hip replacement. In the placebo group, 90 patients were treated with placebo, including 37 males and 53 females, with an average age of(72.91±5.43) years old, involving 69 cases of femoral trochanteric fractures and 21 cases of femoral neck fractures, including 37 cases underwent dynamic hip screw fixation, 30 cases underwent intramedullary fixation and 23 cases underwent artificial hip replacement. The age, sex, injury site, intraoperative bleeding volume, postoperative drainage, operation time, anesthesia time, post-operative pain score, post-operative hemoglobin, post-operative CRP, delirium severity(DRS) score and delirium occurrence were observed and compared between the two groups. RESULTS: All patients were followed up until delirium returned to normal, postoperative delirium was found in 12 cases (13.33%) in the treatment group and in 39 cases(43.33%) in the placebo group, the treatment group was significantly better than the placebo group. The monitoring indexes of the two groups were compared:post-operative pain score(=0.002), post-operative hemoglobin(=0.012), post-operative CRP(=0.042). CONCLUSIONS JWXYP can relieve liver depression, invigorate spleen and invigorate blood circulation, reduce pain, inflammatory stimulation and supplement blood volume after operation, and significantly reduce the incidence of delirium after operation.
Aged ; Delirium ; prevention & control ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Fracture Fixation, Internal ; Hip Fractures ; surgery ; Humans ; Male ; Treatment Outcome

OBJECTIVE: To establish 293T cell lines stably expressing Calpain-cleavage related α3 cytoplasmic tail mutants, and to explore the effect of amino acid motifs in integrin β3 cytoplasmic tail on αⅡbβ3-mediated cell function. METHODS: 293T cell lines stably co-expressing human wild type integrin αⅡb and full length β3 or mutant β3, including β3-ΔNITY (β3 cytoplasmic tail NITY motif deleted), β3-Δ754 (β3 cytoplasmic tail TNITYRGT motif deleted) and β3-Δ759 (β3 cytoplasmic tail RGT motif deleted) were established. Spreading and adhesion of these stable cell lines on immobilized fibrinogen were tested. RESULTS: 293T-αⅡbβ3ΔNITY, 293T-αⅡbβ3Δ754, 293T-αⅡbβ3Δ759 and 293T-αⅡbβ3 cell lines were successfully established. Compared with the 293T cells, 293T-αⅡbβ3 cells which expressed full β3, possessed well adhesion and spread ability on immobilized fibrinogen, suggesting it can be as a surrogate for platelet. Compared with 293T-αⅡbβ3 cells, the 293T-αⅡbβ3ΔNITY cells showed a partial impairment of adhesion and spreadability on immobilized fibrinogen. while the 293T-αⅡbβ3Δ754 cells and 293T-αⅡbβ3Δ759 cells failed to adhere or spread on immobilized fibrinogen. CONCLUSION To the cell spreading function mediated by integrin β3, RGT motif is vital, while NITY can be dispensable. These established 293T cell lines stably expressing different β3 mutants provide a solid basis for a further analysis of mass spectrometry.
Amino Acid Motifs ; Animals ; CHO Cells ; Cell Adhesion ; Cricetinae ; Cricetulus ; HEK293 Cells ; Humans ; Integrin beta3 ; genetics ; metabolism ; Platelet Glycoprotein GPIIb-IIIa Complex ; genetics ; metabolism ; Signal Transduction

Five new polyhydroxylated furostanol saponins were isolated from the roots and rhizomes of Tupistra chinensis, and their structures were determined as tupistrosides J-N (1-5), together with four known furostanol saponins (6-9), on the basis of physico-chemical properties and spectral analysis. Among them, compounds 3 and 5 showed cytotoxicity against human cancer cell lines SW620 with IC values of 72.5 ± 2.4 and 77.3 ± 2.5 μmol·L, respectively. Compound 4 showed cytotoxicity against human cancer cell line HepG2 with IC value of 88.6 ± 2.1 μmol·L.

IMB5046 is a newly discovered nitrobenzoate functioning as a microtubule inhibitor. Here we report its synthesis and in vitro anti-angiogenic activity. IMB5046 was synthesized by conjugation of 2-morpholin-4-yl-5-nitrobenzoic acid with 4-(methylthio)benzyl alcohol via two-step reactions. The structure of the end product was verified using ¹H NMR and HR-MS spectroscopy. The effect of these compounds on cell proliferation was determined using MTT assay, and their impact on cytoskeleton was investigated using fluorescence assay. Flow cytometry was performed to examine the effect of IMB5046 on cell cycle. Cell wound scratch assay and Transwell assay were performed to examine cell migration. Endothelial tube formation assay was used to evaluate the anti-angiogenic activity of IMB5046. The results indicated that IMB5046 induced endothelial cell contraction and microtubule depolymerization, and inhibited the proliferation of endothelial cells and tumor cells, while two raw materials showed no obvious effects. IMB5046 arrested cell cycle at G₂/M phase, even at low-cytotoxic concentrations it significantly inhibited the motility of endothelial cells. IMB5046 inhibited the tube formation of endothelial cells according to the number of tubes and junctions. In conclusion, IMB5046 is a promising microtubule-targeting drug with anti-angiogenic activity.

In this study, the lipid membrane-wrapped nanoparticles loaded with metformin polymer (PolyMet) and doxorubicin (DOX) was prepared and then evaluated therapeutic effect on breast cancer. An anionic chain PGA-DOX based on γ-polyglutamic acid (PGA) with DOX was synthesized via amidation reaction and characterized by ¹H NMR. The PGA-DOX and PolyMet were loaded via electrostatic attraction to prepare the co-delivery nanoparticles system (PolyMet-DOX-NPs). Then, PolyMet-DOX-NPs were coated with cationic liposome membrane to form the core-membrane structural system (PolyMet-DOX-lipid-nanoparticles, PolyMet-DOX-LNPs). The structure and morphology of PolyMet-DOX-LNPs were observed by transmission electron microscope. The particle size, zeta potential, encapsulation efficiency (EE), drug loading (DL), release behavior in vitro of PolyMet-DOX-LNPs were investigated. The MTT assay was used to examine the cytotoxicity of PolyMet combined with DOX on 4T-1 cells. The 4T1^Fluc tumor-bearing mice model was used to evaluate the therapeutic efficacy of PolyMet-DOX-LNPs in vivo. All animal experiments were performed in line with ethical standards and approved by the Animal Experiments Ethical Committee of Zhejiang Chinese Medical University. ¹H NMR spectrum showed that PGA-DOX was successfully synthesized with DOX grafting rate of (72.03 ± 1.29) %. The EE and DL of PolyMet-DOX-LNPs was (72.76 ± 1.92) % and (1.16 ± 0.12) %, respectively. PolyMet-DOX-LNPs exhibited a suitable size of (159.3 ± 7.4) nm and positive charge of (+36.3 ± 1.9) mV with good spheroidal morphology and dispersibility. The release profiles in vitro showed that PolyMet-DOX-LNPs exhibited a slowly and maintained release behavior at physiological pH value (pH 7.4) within 48 h. Further studies showed that PolyMet combined with DOX could synergistically enhance the cytotoxicity on 4T-1 cells. Bioluminescence imaging (BLI) result showed that the luminescence signal intensity of 4T-1^Fluc cells was reduced after treatment with PolyMet-DOX-LNPs and the tumor volume growth was also inhibited. Additionally, the H&E staining and changes of body weight showed that PolyMet could reduce the toxicity of DOX. To sum up, PolyMet has a good synergistic effect with DOX in the treatment of breast cancer, which provide the foundation for this novel metformin polymer on the anti-tumor application.

BACKGROUND: Core decompression for early adult ischemic necrosis of femoral head gets the identity of most scholars, but the postoperative femoral head easily experiences collapse. How to prevent collapse is still a problem to be solved currently. OBJECTIVE: To perform biomechanical analysis of femoral head ischemic necrosis by using the finite element method and to provide biomechanical basis for the treatment of early adult femoral head necrosis. METHODS: One fresh femur specimen died of accidental death in youth and young adults was obtained, and no deformity or fracture was found. X-ray confirmed that it did not have tumor or osteoporosis. Spiral CT was used to scan normal femoral head and neck, pulp core decompression of femoral head and neck, brace device placement and bone-graft of femoral head and neck for acquiring image data from the proximal to distal vertical longitudinal axis. Scanning data were input in the Mimics software. Finite element method was utilized for biomechanical analysis of femoral head and neck of three models. RESULTS AND CONCLUSION: (1) The stress dispersal and downward conduction of normal femoral head was concentrated in the shaft of the femur and the tensile stress was concentrated in the rotor socket. (2) After pulp core decompression, the stress concentration, displacement and strain increased in the weight-bearing area of femoral head. (3) The stress of the internal bracing was similar to that of normal femoral head. (4) The stress of weight bearing area of femoral head is concentrated, and the strain is increased, so that weight bearing area is easy to collapse after pulp core decompression. The more stress distribution, more bearing load and less strain of implant and bone graft model, are conformed to the normal mechanical properties of the normal femoral head and neck.

BACKGROUND: Role of Wnt/β-catenin signaling pathway in the pathogenesis of osteoarthritis has been proved by numerous animal experiments and human specimen trials. Wnt/β-catenin signaling pathway and matrix metalloproteinase 13 (MMP-13) play important roles in pathological process of osteoarthritis, and they may be related with each other. OBJECTIVE: To detect the expression levels of β-catenin and MMP-13 in the synovial tissue of osteoarthritis, and to investigate the relationship between β-catenin and MMP-1 in the pathogenesis of osteoarthritis. METHODS: Synovial tissues were obtained from the 54 patients undergoing total knee arthroplasty or knee arthroscopic diagnosis and treatment surgery and 12 healthy patients suffering from amputation caused by trauma. The samples were divided into non-, mild-, medium-and severe-osteoarthritis groups based on advanced Mankin scores. The expression levels of β-catenin and MMP-13 in the synovial tissues were detected by immunohistochemistry, and correlation analysis was conducted. RESULTS AND CONCLUSION: Immunohistochemistry results showed that MMP-13 was negative in the non-osteoarthritis group, positive in 5 (50%) samples in the mild-osteoarthritis group, positive in 11 (65%) samples in the medium-osteoarthritis group and positive in 23 (85%) samples in the severe-osteoarthritis group. β-Catenin was positive in 1 (8%) sample in the non-osteoarthritis group, positive in 7 (70%) samples in the mild-osteoarthritis group, positive in 14 (82%) samples in the medium-osteoarthritis group and positive in 25 (93%) samples in the severe-osteoarthritis group. The expression levels of β-catenin and MMP-13 in the synovial tissues presented a positive correlation (r=0.806, P ＜ 0.001), and the levels increased with joint degeneration becoming severe. These results imply that there is a significant increase in the expression levels of β-catenin and MMP-13 in the synovial tissues of osteoarthritis compared with the non-osteoarthritis group, and β-catenin and MMP-13 exert effects in development of osteoarthritis. Beta-catenin and MMP-13 are closely associated with pathological changes such as cartilage degeneration and synovial inflammation, thus providing new theoretical direction and experimental basis for targeted gene therapy of osteoarthritis.

Objective To compare the spiral CT and pathological findings of sub-centimetre pulmonary ground glass nodules(GGNs), and to improve the aaccuracy in early diagnosis.Methods The CT findings and pathologic features of 30 patients with sub-centimetre pulmo-nary GGNs in our hospital from May 2013 to June 2016 were reviewed retrospectively.The patients were divided into 3 groups according to their pathological classification,namely the benign group,the preinvasive group and the invasive group.The shape,size,margin,vacuole sign, pleural indentation,the relations with bronchi and blood vessels of the pulmonary GGNs were analyzed.Results There was no significant difference in shape,maximum diameter and margin between the benign group and the other two groups(P>0.05).Adjacent vessels changes of the lesion showed significant differences between benign group and preinvasive or invasive groups(P<0.05).And there were significant differences in size,pleural indentation,vacuole sign and adjacent vessels changes between the preinvasive group and the invasive group(P<0.05).Conclusion The comprehensive analysis of lesions size,vacuole sign,pleural indentation and adjacent vessels changes can be helpful to improve the aaccuracy of differential diagnosis on sub-centimetre pulmonary ground glass nodules.