AIM:To investigate the changes of CD27 and CD28 expression on cytokine-induced killer(CIK) cells and natural killer(NK) cells during cultivation.METHODS:The mononuclear cells were treated with interleukin-12(IL-2),IL-7,IL-15,stem cell factor and FLT3 ligand for four weeks. The CD27 and CD28 expression on CD3+CD56+ CIK cells and CD3-CD56+ NK cells were examined by flow cytometric analysis every week during four-week cultivation.RESULTS:The expression of CD27 on CIK and NK cells two weeks after cultivation reached the peak value at(44.57?4.07)% and(51.02?5.20)%,respectively. Then,their expression declined gradually to about 30% in the fourth week. The expression of CD28 on CIK and NK cells reached the peak at the third and the second week,the values were (4.40?0.66)% and (45.14?3.58)%,respectively,decreased rapidly and then were almost completely lost at the fourth week. CONCLUSION:According to the changes of expressions of CD27 and CD28 on CIK/NK cells,optimal harvest time of cultured CIK/NK cells should be at the third week of cultivation.

AIM: To evaluate the biological characteristics and differentiating potentials of bone marrow-derived mesenchymal stem cells(MSCs) from sensitized mice by allogeneic splenocyte transfusion in vitro.METHODS: Adherent culture method was applied for culturing the bone marrow-derived MSCs from sensitized mice.The cell morphology was examined and the surface marker profiles were analyzed by flow cytometry.The differentiating potentials of the MSCs into osteogenic,adipogenic and myogenic lineages were explored.The bone marrow-derived MSCs from the normal mice were collected and served as controls.RESULTS: Both the bone marrow-derived MSCs from sensitized and normal mice were exhibited a homogeneous distinctive morphology and were positive for the surface markers CD29,CD105,CD44 and Sca-1,negative in CD 34 and CD11b.The abilities of both MSCs to differentiate into osteogenic,adipogenic and myogenic pathways in the same condition were also observed.CONCLUSION: There is no difference in the biological characteristics and induced differentiating potentials between the sensitized mouse bone marrow-derived MSCs by allogeneic splenocytes transfusion and the MSCs from normal mice.

Liver transplantation has become the most effective treatment option for end-stage liver diseases.The liver donation after cardiac death (DCD) had been increasingly used in clinical practice.Nevertheless,the DCD donated livers inevitably exposed to long-term ischemia and hypoxia makes the implanted organs with decreased function and more postoperative complications.Different from the conventional static cold storage,machine perfusion (MP) can be continuously used in vitro liver perfusion,which could simulate the in vivo liver status.Through the observation of ischemia injury status,we could evaluate the quality of donated liver,and thus reduce ischemia related complications.This review provides an overview and evaluation on the markers that have been investigated for the assessment of graft quality and viability testing during different types of MP.

High levels of RNase,polysaccharides and polyphenol compounds make isolation of high quality RNA difficult. Thus it is presented an effective RNA extraction method based on the nuclease adsorbent macaloid,poly vinyl pyrrolidone,and high concentration of KAc and ethylene glycol monobutyl ether,which has successfully extracted high-quality RNA from many materials difficult to RNA isolation,such as RNase-rich rabbit liver,plant and microbial tissues rich in polysaccharides,lipids and polyphenol compounds. This method was found to be better than the ones in common use-Trizol and Guanidinium isothiocyanate,the yield of which was at least three time higher. Furtherly,small RNA was enriched from total RNA sample from rice seedling through by repeat deposit which deals with high concentration of LiCl,PEG8000 and NaCl. The small RNA gained was confirmed to be used for following molecular biological research by RT-PCR with the primers designed on osa-mir-156 sequence from rice miRNA.

Objective To explore the diagnostic value of virtual bronchoscopic navigation (VBN) for peripheral pulmonary lesions. Methods 200 cases with peripheral pulmonary lesions (0.8 cm < diameter < 4.0 cm) were divided into VBN and control group from June 2014 to June 2015. VBN group: VBN guided ultrafine bronchoscope to the target bronchus, control group: ultrafine bronchoscopy with chest CT as a reference guided to the target bronchus. Results There were no significant differences in the diagnosis rates between VBN group and control group in 200 patients (χ2 = 3.31, P = 0.069); But, the diagnosis rates with diameter more than 2.0 cm and less than or equal to 2.0 cm had statistically significant differences of VBN group and control group (χ2 = 13.45, 5.31, P = 0.000, 0.021, respectively); We also found that the mean time of biopsy tool reach to the lesions had significant differences between the two groups (P = 0.047). There were no significant differences in total checking time and complications (P = 0.230, 0.960, respectively). Conclusions Virtual bronchoscopic navigation did not improve the diagnosis rate of pulmonary peripheral lesions, but shortened the time to locate the lesions.

Objective To detect the expression of hypoxia-inducible factor-1α (HIF-1α) in breast can-cer and to investigate the feasibility of HIF-1α as a new therapy target for breast cancer. Methods 56 cancer tissue specimens of women with primary breast cancer admitted from Jan. 2013 to Dec. 2013 in the Fourth Hos-pital of Hebei Medical University Breast Center were selected. Immunohistochemistry was used to detect the ex-pression of HIF-1α protein in cancer tissues. The relation between HIF-1α protein expression and the clinico-pathological parameters was analyzed. Results The positive expression rate of HIF-1α protein in breast cancer was 73.2% (41/56) and its high expression rate was 41.1% (23/56), which were significantly higher than those in benign breast tumor group (P=0.000). HIF-1α protein expression was positively correlated with breast tumor size, stage, histological grade, lymph node metastasis and the expression of HER2, and it was negtively correlated with ER. Conclusions High expression of HIF-1α in breast cancer was significantly associated with poor prog-nosis. HIF-1α protein is also a risk factor for breast cancer as well as HER2, and they may have a synergistic role in the progression of breast cancer.

Objective To explore the diagnostic efficacy of bronchoscopy combined with T-SPOT in tuber-culosis. Methods Totally 248 highly suspected tuberculosis patients were included from February 2015 to April 2016 in our department. According to the different sources of sputum specimens ,they were divided into control group,atomization group and bronchoscope group;Depending on the mode of examination,they were divided into bronchoscopy group,T-SPOT group,bronchoscopy+T-SPOT group.To compare anti-acid smear positive rate,tuber-culosis diagnosis and diagnostic efficacy in each group and safety of bronchoscopy. Results The rate of sputum specimens smear-positive was the highest(37.173%) in bronchoscopy.There were statistically significant of the three groups(P=0.001);The highest diagnostic rate of pulmonary tuberculosis was(92.670%)in Bronchoscopy+T-SPOT group,three groups were statistically significant(P=0.000). The specificity,sensitivity,positive predic-tive value,negative predictive value,positive likelihood ratio and Jorden index were highest in diagnosing tuberculo-sis of bronchoscopy+T-SPOT group ,negative likelihood ratio was lowest. There were Complications of three cases of bleeding,one case of pneumothorax,one case of arrhythmia in bronchoscopy group. Conclusion Bronchoscopy combined with T-SPOT can improve the diagnostic efficacy of tuberculosis ,safe and reliable.

Objective To investigate the effects of compound radix angelicae sinensis injection on oocyte and segmentation sphere injured by subacute exposure to diesel exhaust particles (DEP) in female mice. Methods Two hundred and ten 21-day-old ICR female mice were randomly divided into 5 groups: the control group (group A), DEP group (group B), DEP+low-dose group (group C),DEP±middle-dose group (group D) and DEP+high-dose group (group E). There were 42 mice in each group. The mice were inoculated with 30 μ1 DEP suspension at 12.0 μg/μl (group B-E) or the same volume of vehicle (PBS, group A) on pharynx posterior wall by sample pipettor beginning at day 21 and repeated every 3 days for 4 times. The mice were sacrificed three days after the last exposure.Compound radix angelicae sinensis injection containing 75 (group C), 150 (group D) and 300 (group E)grams of crude drug, respectively, which was intraperitoneally administered for each mouse daily from the day of the first DEP inoculation till the day before sacrifice, consecutively for 12 days. The general conditions were observed, and the body weight and ovary/body weight ratio were tested. Superoxide dismutase (SOD) activity, malondialdehyde (MDA) and reduced glutathione (GSH) contents in ovarian tissues were assayed. Rates of survival, germinal vesicle breakdown, extrusion of the first polar body and in-vitro fertilization, and quantity of mitochondrial DNA for oocytes were investigated.Ultrastructural changes of oocytes were observed. Results ( 1 ) No significant difference of the body weight was found among all the groups (P＞0. 05). The ovary weight, ovary/body weight ratio, ovary SOD and GSH content were significantly decreased in groups B [( 1.5 ± 0. 6) mg, ( 7.2 ± 2. 5) × 10-5 ,(192. 10±23.67) nU/mg prot and (262.40 ± 31.60) nmol/mg prot], and C [( 1.7 ± 0. 2) mg,(8.9±0.6)× 10-5, (198.92±24.27) nU/mg prot and (271.66±14.58) nmol/mg prot] and D [(2. 1±0. 2) mg, (9. 8±1. 1)×10-5, (214. 37±27. 19) nU/mg prot and (285. 93±9. 55) nmol/mg prot] as comparing to groupA [(3. 3±1. 5) mg, (15.4±7.3)×10-5, (292. 30 ± 40. 03) nU/mg prot and (367.98±24.59) nmol/mg prot (P＜ 0. 05 or P＜0.01); and significantly increased in group E [(3. 7±1.1) mg, (18. 7±5. 4)× 10-5, (279. 10±12. 63) nU/mg prot ]and (353. 59±10. 61) nmol/mg prot]comparing to group B (P＜0. 01). MDA content was signi-ficantly increased for groups B, C and D [(3. 88±0.35) nmol/mg prot, (3. 62 ± 0. 19) nmol/mg prot and (2. 63 ± 0. 34) nmol/mg prot] comparing to group A [(2. 18±0. 44) nmol/mg prot](P＜0. 05 or ＜0. 01, respectively); and significantly decreased for group D and E (2. 35±0. 37 nmol/mg prot) comparing to group B (P＜0. 01). (2) In all observed time points, oocyte survival rate in group B and C, extrusion rate of the first polar body and in-vitro fertilization rate in group B, C and D were significantly lower than in group A (P ＜ 0. 05 or ＜0. 01), and all those in group E were significantly higher than in group B (P＜0. 05). Rates of germinal vesicle breakdown were 100% in all five groups. (3) Logarithmic values of mitochondrial DNA copy numbers in group C, D and E were significantly lower than in group A; whereas significantly higher in group C and D than in group B (P＜0.01). (4) Normal appearance for oocytes in group A was seen. In groups B and C, a number of cytoplasmic organelles were dramatically degenerated in part of the oocytes and some necrotic oocytes were seen. Large body of mitochondria in the oocytes swelled and vacuolized in group D, while such changes dincished to a lesser extent and scope in group E. Conclusions Compound radix angelicae sinensis injection exerts a favorable curative and protective effect on oocyte and segmentation sphere injured by subacute DEP exposure in female mice.

【Objective】To observe the efficacy and side effects of hematopietic stem cell transplantation with combination of umbilical cord blood(UCB) and neonatal peripheral blood(NPB) in the treatment of β-thalassemia major.【Methods】28 mL NPB was drawn from a HLA identical neonate within 5 hours after his birth to complement stem cell of the UCB he donated for transplantation to his sibling with β-thalassemia major.Various items of hematopoiesis reconstruction were detected in UCB and NPB respectively.After conditioning with chemotherapy by using busulfan 20 mg/kg,cyclophosphamide 200 mg/kg,melphalan 90 mg/m2 and antithymocyte globulin(ATG) 90 mg/kg,the patient received the 53 mL UCB and 28 mL NPB,achieving 5.7×107/kg nucleated cells(NC),93×105/kg CFU-GM and 3.1×105/kg CD34+CD38- cells from his HLA-identical sibling.【Results】Absolute nucleated cell(ANC) reached 0.5×109/L on 14th day post transplant,and platelets reached 20×109/L on 34th day after transplant.The heterozygosity of β-654 mutation point was detected by the PCR-RDB.The sexual chromosome changed from XX pretransplant to XY posttransplant.The patient was free red blood cell transfusion from 14th day post transplant.Her hemoglobin rose progressively from 86 g/L to 110 g/L.The patient survived for 197 days free from disease after transplantation.Following up for 9 months, the donor grew and developed normally.【Conclusion】The NPB contains a lot of stem cells.The transplantation with combination of suitable NPB and UCB is an effective tactics when the UCB cells are deficient.

BACKGROUND: Recently biodegradable hydrogel has been extensively used to delivery anticancer drug and bioactive macromolecule. However, to protect the activity of the bioactive macromolecule, we need to obtain series of hydrogel which have milder hydrogelation conditions and shorter hydroglation time.OBJECTIVE: To prepare enantiomeric poly(L-Lactic acid) (PLLA)-poly(ethylene glycol (PEG)-PLLA/ poly(D-Lactic acid) (PDLA)-PEG-PDLA stereocomplex hydrogel which has shorter hydroglation time, to physically encapsulate a model drug-lysozyme and sustained release it from the hydrogel. METHODS: Triblock copolymers of PLLA-PEG-PLLA and PDLA-PEG-PDLA were synthesized by ring-opening polymerization of L(D)-lactide using PEG as the initiator and Sn(Oct)2 as the catalyst. The triblock copolymers were characterized by 1H nuclear magnetic resonance, FT-IR and X-Ray diffractometry. A hydrogel was prepared from an aqueous mixture of PLLA20-PEG227-PLLA20 and PDLA21-PEG227-PDLA21 (10 wt% concentration) at room temperature for 12 hours. X-Ray diffractometry test was used to research the gelation mechanism. The release profile of the lysozyme as a model drug from the hydrogel was tested. The morphology of the freeze-dried hydrogel was investigated by scanning electron microscope. The cytotoxicity of the hydrogel was evaluated by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium bromide) assay.RESULTS AND CONCLUSION: Triblock copolymers of PLLA-PEG-PLLA and PDLA-PEG-PDLA were obtained. Both the PEG and PLA blocks in the copolymers could crystallize, but the crystallization of the PEG block was predominant. The stereocomplex formation between the PLLA and PDLA blocks within the hydrogel was confirmed by the X-Ray diffractometry analysis. The release profile of the lysozyme from the hydrogel exhibited a sustained-release pattern with a duration period of 7 days. The hydrogel exhibited a 3D interconnected porous structure with 50-100 μm pore size after being freeze-dried. The mouse fibroblast cell viability percentage was 99.3% after the cells contacted with the 100% extracted liquid for 72 hours.