The Bone Defect Repairment in Vivo with Co-cultured EPCs and BMSCs / 昆明医科大学学报

Objective To reconstruct the deformity of appearance and function of patients with bone defect, co-cultured system with two stem cells were combined with partial deproteinized biological bone to reconstruct the defect of tibia which is one of the main weight-bearing bone. Methods The bone marrow and peripheral blood were harvested form 18 New Zealand rabbits to isolated bone marrow stem cells and epithelial progenitor cells, and engineering bone was constructed with co-cultured system with these two stem cells and partial deproteinized biological bones; about 1 CM of bone defect of each rabbit was made with bone rongeur, then engineering bones were transplanted into the defect area, the osteogenesis and bone defect recovery were observed on day 14, 28 and month 2.Results The difference of absorbance values of BMSCs group, co-cultured cell group and blank group at each time point and between groups were all statistically significant (P＜0.001), and the collagen content of bone tissue increased gradually after implantation of tissue engineered bone, and the difference between each group was statistically significant (P ＜0.001). The repairment of bone defect with the PDPBB combined with BMSCs and EPCs system has the strongest ability to repair the structure andfunction of the tibial defect area. Conclusion The engineering bone constructed with two stem cells and partial deproteinized bone is a good material for bone defect reconstruction.

The Correlation Analysis between Hyperuricemia and Dyslipidemia in Rural Areas,Yunnan Province / 昆明医科大学学报

Objective To provide evidence－based prevention of chronic disease and nutritional interventions by investigating the development of dyslipidemia and hyperuricemia in rural areas of Yunnan and analyzing the relationship between hyperuricemia and dyslipidemia. Methods The cross－sectional study recruited 513 residents over 18 years old in 2016. The morning fasting venous blood of all subjects were obtained to detect uric acid, serum total cholesterol (TC), triglycerides (TG) and high－density lipoprotein cholesterol (HDL－C) . Each food factor's blood lipid to the hyperuricemia was determined through multivariate logistic regression. Results The prevalence rates of hyperuricemia and dyslipidemia were 18.7% and 44.7% respectively. Females were more likely to have hyperuricemia than males (P＜0.05) . Males were more likely to develop dyslipidemia than females (P＞0.05) . The prevalence of hyperuricemia and dyslipidemia increased with age. Logistic regression analysis showed that the level of TG and LDL－C were risk factors and the level of HDL－C was protective factor of hyperuricemia. Conclusion Regularly testing the levels of uric acid and blood lipid, enhancing the knowledge of reasonable diet and developing healthy dietary habits have significant importance to prevent chronic diseases.

Effects of arsenic trioxide combined with bortezomib on proliferation and apoptosis of K562 cells and their mechanism / 中国实验血液学杂志

This study was aimed to investigate the effect of arsenic trioxide (As(2)O(3)) alone and in combination with bortezomib (Bor) on proliferation and apoptosis of leukemia cell line K562, and to analyze the potential mechanism. K562 cells were treated with different concentrations of As(2)O(3) or Bor (alone or combination) for 24, 48 h. MTT method was used to detect the cell proliferation. After K562 cells were treated with 0.5 µmol/L As(2)O(3) alone or in combination with 10 nmoL/L Bor, the apoptosis rate and cell cycle were measured by flow cytometry, and the activity of NF-κB was analyzed by SP immunohistochemistry. The results indicated that the different concentrations of As(2)O(3) and Bor could inhibit the K562 cell proliferation in a time- and dose-dependent manners (P < 0.05). The IC(50) of Bor and As(2)O(3) in 48 h were 20 nmol/L and 0.6 µmol/L respectively. When K562 cells were treated with As(2)O(3) or Bor alone for 24 h, the apoptotic rate of K562 cells increased, and the apoptotic rate in combination group was higher than that in As(2)O(3) or Bor group. The cells were apparently arrested in G(2)/M phase in Bor group and G(0)/G(1) phase in As(2)O(3) group. The activity of NF-κB decreased significantly in As(2)O(3) or Bor group (P < 0.05), this effect was most significant in the combination group (P < 0.01). It is concluded that both As(2)O(3) and Bor can inhibit the proliferation and induce apoptosis of K562 cells, a synergistic effect can be observed when a low dose of As(2)O(3) combined with low dose of Bor. The different cell cycle block site and the decrease of activity of NF-κB may be one of the mechanisms underlying their synergic effect.

Effects of hyperbaric oxygenation combined with As(2)O(3) on proliferation of K562 cells and associated mechanism / 中国实验血液学杂志

This study was aimed to explore the effects of hyperbaric oxygenation (HBO) alone or combined with As(2)O(3) on proliferation, apoptosis and expression of HIF-1a, VEGF, caspase-3 mRNA of K562 cells, and the molecular mechanism of As(2)O(3) enhancing the anti-leukemic effect of HBO so as to provide a scientific basis for clinical treatment of chronic myeloid leukemia. The effects of drugs on proliferation of K562 cells was assayed by MTT method, the apoptosis rate of K562 cells was detected by flow cytometry with Annexin V/PI double staining, the expressions of HIF-1a, VEGF, caspase-3 mRNA of K562 cells were determined by real-time quantitative PCR. The results showed that as compared with As(2)O(3) alone, HBO combined with As(2)O(3) could increase inhibitory rate of K562 cell proliferation, and enhance apoptotic effect, obviously down-regulate expressions of HIF-1a and VEGF mRNA, up-regulate expression of caspase-3 mRNA. The effect of HBO combined with As(2)O(3) was higher then effect of As(2)O(3) alone, and their effects were synergistic (P < 0.05). It is concluded that HBO combined with As(2)O(3) can increase the expression of caspase 3 mRNA and decrease the expression of HIF-1a and VEGF mRNA, which may be one of molecular mechanisms underlying their synergistic antileukemia efficacy.

Expression of miR-21 in multiple myeloma and its clinical significance / 中国实验血液学杂志

This study was aimed to explore the expression of microRNA-21 (miR-21) in multiple myeloma (MM), and its correlation with plasma β2-microglobulin (β2-MG) and staging of MM by Durie-Salmon (D-S) classification. The expression level of miR-21 in bone marrow mono-nuclear cells (BMMNC) of 43 patients with MM and 20 healthy individuals was examined by real-time polymerase chain reaction (real-time PCR), and the correlations among the expression level of miR-21 and related clinical pathologic features, plasma β2-MG and staging of MM by D-S classification were analyzed. The results showed that the expression of miR-21 in BMMNC of MM patients was obviously higher than that in normal controls (P < 0.05). The expression of miR-21 in BMMNC of relapsed/refractory MM patients was obviously higher than that in newly diagnosed MM patients. The expression of miR-21 in MM patients decreased after chemical therapy, especially in effective group (P < 0.05), there was no significant change of miR-21 expression level in ineffective/progressive group before and after chemical therapy (P > 0.05). The expression of miR-21 was related with staging of MM and plasma β2-MG level. It is concluded that the expression levels of miR-21 in BMMNC of MM patients are significantly higher than in normal bone marrow, these data indicated that miR-21 may play an important role in the development of MM. Super expression of miR-21 is positively correlated with level of plasma β2-MG and staging of MM by D-S classification.

Expression and significance of B cell-activating factor of TNF family (BAFF) and B cell lymphoma/leukemia-2 (BCL-2) in multiple myeloma / 中国实验血液学杂志

This study was aimed to explore the expression of B cell-activating factor of TNF family (BAFF) and B cell lymphoma/leukemia-2 (BCL-2) in bone marrow mononuclear cells (BMMNC) of multiple myeloma (MM) and the significance of BAFF and BCL-2 for occurrence, development and prognosis of MM. The bone marrow of 40 cases of MM and 10 healthy persons was collected, the mononuclear cells (MNC) were isolated, the expression of BAFF and BCL-2 mRNA in BMMNC was detected by real-time PCR; the plasma was simultaneously collected and the β2-MG level was determined; the clinical staging of MM patients was performed according to Durie-Salmon (D-S) staging criterion. The results indicated that the expression level of BAFF and BCL-2 mRNA in MM patients increased, as compared with normal controls, the difference was statistical significant (p < 0.05); the expression level of BAFF and BCL-2 mRNA in plateau stage after treatment obviously decreased. The expression level of BAFF and BCL-2 mRNA in relapsed/refractory MM patients was significantly higher than that in normal controls and patients reached plateau stage (p < 0.05), there was no statistically significant difference between newly diagnosed and relapsed/refractory MM patients (p > 0.05). The expression of BAFF and BCL-2 mRNA related with D-S staging and β2-MG level. It is concluded that the expression levels of BAFF and BCL-2 mRNA increase, moreover the expression levels of BAFF and BCL-2 mRNA in newly diagnosed and relapsed/refractory MM patients are higher than those in patients reached plateau stage, which suggest the BAFF and BCL-2 may be involved in occurrence and development of MM; the relation of expression level of BAFF and BCL-2 mRNA to MM load is positive, which indicates the expression level of BAFF and BCL-2 mRNA may be a new indicator for evaluating the prognosis of MM patients.

Applications of antisense technology in discovery and development of novel antibacterial drugs / 国际药学研究杂志

Antibiotic-resistance has become a serious problem to human health. Innovative approaches are urgently required for both antimicrobial drug discovery and reversal of resistance. Great success has been achieved since the introduction of antisense technology. This review focuses on the applications of antisense technology in the aspects including identification and validation of novel antibacterial drug targets, screening of antibacterial drugs from natural products using hypersensitized strains engineered by inducible antisense RNA, and inhibition of antibiotic-resistance genes to reverse susceptibility of antibiotic-resistant strains to currently used antibiotics. In addition, the antibacterial activities, the shortcomings and prospects as antibacterial drugs of synthetic antisense oligodeoxynucleotides and their analogs are also assessed in this review.

Differentiation of smooth muscle progenitor cells in peripheral blood and its application in tissue engineered blood vessels / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

<p><b>BACKGROUND</b>A major shortcoming in tissue engineered blood vessels (TEBVs) is the lack of healthy and easily attainable smooth muscle cells (SMCs). Smooth muscle progenitor cells (SPCs), especially from peripheral blood, may offer an alternative cell source for tissue engineering involving a less invasive harvesting technique.</p><p><b>METHODS</b>SPCs were isolated from 5-ml fresh rat peripheral blood by density-gradient centrifugation and cultured for 3 weeks in endothelial growth medium-2-MV (EGM-2-MV) medium containing platelet-derived growth factor-BB (PDGF BB). Before seeded on the synthesized scaffold, SPC-derived smooth muscle outgrowth cell (SOC) phenotypes were assessed by immuno-fluorescent staining, Western blot analysis, and reverse transcription polymerase chain reaction (RT-PCR). The cells were seeded onto the silk fibroin-modified poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (SF-PHBHHx) scaffolds by 6x10(4) cells/cm2 and cultured under the static condition for 3 weeks. The growth and proliferation of the seeded cells on the scaffold were analyzed by 3-(4,5-dimethylthiazol-2-yl)-diphenyltetrazolium bromide (MTT) assay, scanning electron microscope (SEM), and 4,6-diamidino-2-phenylindole (DAPI) staining.</p><p><b>RESULTS</b>SOCs displayed specific "hill and valley" morphology, expressed the specific markers of the SMC lineage: smooth muscle (SM) alpha-actin, calponin and smooth muscle myosin heavy chain (SM MHC) at protein and messenger ribonucleic acid (mRNA) levels. RT-PCR results demonstrate that SOCs also expressed smooth muscle protein 22alpha (SM22alpha), a contractile protein, and extracellular matrix components elastin and matrix Gla protein (MGP), as well as vascular endothelial growth factor (VEGF). After seeded on the SF-PHBHHx scaffold, the cells showed excellent metabolic activity and proliferation.</p><p><b>CONCLUSION</b>SPCs isolated from peripheral blood can be differentiated into the SMCs in vitro and have an impressive growth potential in the biodegradable synthesized scaffold. Thus, SPCs may be a promising cell source for constructing TEBVs.</p>

Construction of tissue-engineered heart valves by using decellularized scaffolds and endothelial progenitor cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Tissue-engineered heart valves have the potential to overcome the limitations of present heart valve replacements. This study was designed to develop a tissue engineering heart valve by using human umbilical cord blood-derived endothelial progenitor cells (EPCs) and decellularized valve scaffolds.</p><p><b>METHODS</b>Decellularized valve scaffolds were prepared from fresh porcine heart valves. EPCs were isolated from fresh human umbilical cord blood by density gradient centrifugation, cultured for 3 weeks in EGM-2-MV medium, by which time the resultant cell population became endothelial in nature, as assessed by immunofluorescent staining. EPC-derived endothelial cells were seeded onto the decellularized scaffold at 3 x 10(6) cells/cm(2) and cultured under static conditions for 7 days. Proliferation of the seeded cells on the scaffolds was detected using the MTT assay. Tissue-engineered heart valves were analyzed by HE staining, immunofluorescent staining and scanning electron microscopy. The anti-thrombogenic function of the endothelium on the engineered heart valves was evaluated by platelet adhesion experiments and reverse transcription-polymerase chain reaction (RT-PCR) analysis for the expression of endothelial nitric oxide synthase (eNOS) and tissue-type plasminogen activator (t-PA).</p><p><b>RESULTS</b>EPC-derived endothelial cells showed a histolytic cobblestone morphology, expressed specific markers of the endothelial cell lineage including von Willebrand factor (vWF) and CD31, bound a human endothelial cell-specific lectin, Ulex Europaeus agglutinin-1 (UEA-1), and took up Dil-labeled low density lipoprotein (Dil-Ac-LDL). After seeding on the decellularized scaffold, the cells showed excellent metabolic activity and proliferation. The cells formed confluent endothelial monolayers atop the decellularized matrix, as assessed by HE staining and immunostaining for vWF and CD31. Scanning electron microscopy demonstrated the occurrence of tight junctions between cells forming the confluent monolayer. Platelets adhesion experiments suggested that the neo-endothelium was non-thrombogenic. The expression levels of eNOS and t-PA genes in the neo-endothelium were quite similar to those in human umbilical vein endothelial cells.</p><p><b>CONCLUSIONS</b>EPCs isolated from the human umbilical cord blood can differentiate into endothelial cells in vitro and form a functional endothelium atop decellularized heart valve scaffolds. Thus, EPCs may be a promising cell source for constructing tissue-engineered heart valves.</p>

Clinical study on drug-seperated moxibustion at Shenque (CV 8) for treatment of ulcerative colitis / 中国针灸

<p><b>OBJECTIVE</b>To observe the therapeutic effect of drug-separated moxibustion at Shenque (CV 8) on ulcerative colitis (UC) and the influence on autoimmunity level, and study on the mechanism.</p><p><b>METHODS</b>Sixty cases were randomly divided into a treatment group and a control group, 30 cases in each group. The treatment group were treated with drug-separated moxibustion at Shenque (CV 8) and the control group with oral administration of sulphasalazine and metronidazole tablets. The main symptoms, pathological changes of the intestinal mucosa and the total therapeutic effect in the two groups before and after treatment, and the changes of the content of blood serum immunoglobulin, peripheral blood T-cell subgroup and NK cell were observed.</p><p><b>RESULTS</b>The markedly effective rate and the total effective rate in the treatment group were 60.0% and 86.7% respectively, much higher than the control group (P<0.05, P<0.01). IgG content in the treatment group after treatment significantly decreased with a significant difference as compared with the control group (P<0.05); there were no significant differences in IgA and IgM before and after the treatment (P>0.05). After treatment, peripheral blood T-cell subgroup and NK cell in the treatment group increased to a certain extent, which was correlated positively to the therapeutic effect.</p><p><b>CONCLUSION</b>The drug-separated moxibustion at Shenque (CV 8) is a satisfactory method for treatment of ulcerative colitis, and it exerts therapeutic effect possibly through regulating immunological function of the organism.</p>

Clinical observation on medicine-separated moxibustion for treatment of primary dysmenorrhea and study on the mechanism / 中国针灸

<p><b>OBJECTIVE</b>To observe clinical therapeutic effect of medicine-separated moxibustion on primary dysmenorrhea and study on the mechanism.</p><p><b>METHODS</b>Ninety-six cases of primary dysmenorrhea were randomly divided into a treatment group and a control group, 48 cases in each group. They were treated respectively with medicine-separated moxibustion and Yueyueshu Perfusing Powder for 3 consecutive months.</p><p><b>RESULTS</b>The markedly effective and cured rate and the total effective rate were 87.5% and 100.0% in the treatment group, and 29.2% and 83.8% in the control group with significant differences between the two groups (P < 0.01 and P < 0.05). After treatment, blood prosglanding F2 content in menorrhea blood and oxytocin level in plasma during menstruation decreased significantly as compared with those before treatment (P < 0.01).</p><p><b>CONCLUSION</b>The medicine-separated moxibustion has a reliable therapeutic effect on primary dysmenorrhea and the therapeutic effect is exerted possibly by regulating abnormal prosglanding and oxytocin levels in the patient.</p>

Molecular cloning and expression of anti-tumor adhesion peptide (beta3) / 生物工程学报

To block tumor cell adhesion, inhibit tumor metastasis and recurrence, the anti-adhesion peptide-trimeric beta peptide (DLYYLMDLSYSMKGGDLYYLMDLSYSMKGGDLYYLMDLSYSMK, beta3) was designed. The DNA fragment of beta3 was cloned into expression vector pET-His and the fusion protein His-beta3 was expressed in E. coli. BL21(DE3)plysS. After 1.5 hours' induction with IPTG, His-beta3 peptide was expressed significantly amounting to 10% of the insoluble proteins and 4% of the total proteins. 20mg of beta3 peptide was obtained from one litter culture medium after purification by using metal-chelating sepharose 6B FF. The purity of beta3 is 92.2% according to Gel-Pro analysis. The anti-adhesion effects of beta3 peptide, beta1 peptide (DLYYLMDLSYSMK) and GRGDS on the hepatocellular carcinoma cell line SMMC-7721 and the high metastasis hepatocellular carcinoma cell line HCCLM6 were studied. The result showed the beta3 blocked the adhesion of HCCLM6 cells and SMMC-7721 cells to fibronectin (FN) specifically. The inhibition effect was dose-dependent and time-dependent and the inhibition rate of beta3 was higher than three times concentration of beta1 and GRGDS. This suggested that pET-His-beta3/BL21(DE3)plysS was a suitable expression system for beta3, and the expressed beta3 specially inhibited the adhesion of cancer cells.