Cytokine-induced killer cells (CIK) is the fourth largest cancer treatment after surgery,chemotherapy and radiotherapy,and it is the development direction of cancer treatment.It is a new type of immune cell,and it is named after natural killer cell samples T lymphocytes as it express CD3 and CD56.Currently,CIK treatment has a broader range of clinical applications,and it has achieved the better clinical efficacy in the blood system cancer and solid tumors,The CIK adoptive immunotherapy is considered to be a new hope for the anticancer treatment.

Objective To study the in vitro culture condition for melanoblasts from human foreskin tissue. Methods The skin tissue taken from foreskin of children was treated with 0.5% dispase Ⅱ to separate epidermis from dermis, then with trypsin to obtain single cell suspension, which was cultured in modified medium for melanoblasts, i.e., MCDB254 medium supplied with several cell growth factors. Finally, melanoblasts were obtained based on the difference of adhesion speed. The morphology and proliferation of cultured melanoblasts were observed under a light microscope. DOPA staining, immunostaining with anti- S-100 and -tyrosinase related protein 2 (TRP2) antibodies, and transmission electron microscopy were per- formed to identify the cultured melanoblasts. Results The cultured human melanocytes displayed a match-like shape, scattered arrangement, syrmnetric double poles, slim cell body, highly refractive nuclei; meanwhile, the melanoblasts exhibited plentiful cytoplasm, large volume, bipolar or irregular shape and clonal growth. Additionally, the melanocytes were positive for TRP2, S-100 and Dopa staining, while the melanoblasts were positive only for TRP2. Electron microscopy revealed the presence of mature melanin granules (stage Ⅲ-Ⅳ ) in melanocytes but immature melanin granules (stage Ⅰ ) in melanoblasts. Conclu- sion Stable pure culture of melanoblasts has been realized with the reformed medium, which may lay a foundation for the investigation into the mechanism of epidermal pigmentation.

The aim of the study is to evaluate the feasibility and safety of Bispectral index (BIS) monitoring in pediatric radio frequency catheter ablation. One hundred and six children aged 0. 6-12 years, scheduled for radio frequency catheter ablation, were randomly divided into two groups. In group A patients received BIS monitoring during the operation (n = 50), and the group B received modified Observer's Assessment of Alertness/Sedation (OAA/S) scaling (n = 56). The anesthesia was maintained with propofol target-controlled infusion. The intraoperative propefol target concentration was adjusted to maintain the BIS values between 55-65 in group A and OAA/S scale about 1 in group B respectively, The heart rate (HR), mean arterial pressure (MAP) and pulse oximetric saturation (SpO2) were measured before anesthetic induction, 1 min after induction, catheter puncturing and the end of operation respectively. The requirements of propofol, the times of supporting ventilation and recovery, the respiratory depression, nausea and vomiting postoperatively were also recorded. The intraoperative HR, MAP and SpO2 showed no differences between two groups, but the requirements of pmpofol, the times of supporting ventilation and recovery were less in group A than that of group B (P<0.05). All children didn't have nausea, vomiting and respiratory depression. The results suggest that in pediatric radio frequency catheter ablation, BIS monitoring has the advantages of timely adjustment of anesthetic depth, reducing anesthetic requirements, shortening the time of recovery, so as the perioperative safety can be improved.

Objective To study two new factor Ⅸ mutations Cys82Ser and Ile288Ser in vitro and research the molecular mechanism of haemophilia B. Methods PcDNA3. 1 ( - ) FⅨwt expression plasmid was prepared. The mutated FⅨcDNA expression plasmids, PcDNA3.1 ( - ) FⅨM1 (Cys82Ser) and PcDNA3. 1 ( - ) F Ⅸ M2 (Ile288Ser) were constructed by megaprimer method respectively. Transient expression experiments were performed using HEK293 cells transfected with the expression vectors containing the wild-type or the mutation recombinant cDNA. PcDNA3. 1 ( - ) was used as a blank control. The expression proteins were detected by ELISA, factor activity assay and flourescence stain. Results The results suggested that the two FⅨ gene mutations did not induce the reduction of the mutant FⅨ mRNA compared with the wild-type FⅨ mRNA. The FⅨ:Ag in culture media and cell lysate of wild type conduct were assigned as 100. 0%. The results of PcDNA3.1 ( - ) FⅨ M1 mutation protein were (27. 1 ± 5. 2)% and (99.4 ±4. 1)% respectively. For PcDNA3. 1( - )FⅨM2, the results were (5.3 ± 1.8)% and (31.7 ±2. 5)% respectively. The FⅨ: C in culture media of wild type conduct was also assigned as 100. 0%. Then the two types of mutant protein were ( 8. 5 ± 3.2 ) % and ＜ 1%, respectively. Immunofluorescence microscopy result suggested that the intensity of perinuclear spot was reduced in cells transfected with PcDNA3.1 ( - ) FⅨM2 while staining for PcDNA3. 1 ( - ) FⅨM1 was predominantely diffuse without perinnclear enhancement. Conclusions These results strongly suggest that the FⅨ Cys82Ser mutation protein is not been correctly folded, by any possibility. The mutation protein has secretion defect. The secretion dysfunction and the protein degradation intracellular are possiblely the molecular pathology of Ile288Ser mutant protein.

Objective To study the effects of bone marrow mesenchymal stem cells(BMSCs) transplantation combined with artesunate(Art) on rat model of liver fibrosis.Methods 20 rats from 120 SD rats were selected randomly as normal control group.10 rats from those were executed and extracted marrow to culture BMSCs.The other 90 rats were injected intraperitoneally with CCl4 to make liver fibrosis rat models.10 rats from the 90 rats were selected randomly to ensure the liver fibrosis model was made successfully after 10 weeks.Then,the rest 80 rats were divided randomly into 4 groups:model group,Art treated group,BMSCs transplantation group and Art + BMSCs transplantation group.Fibrous degeneration of the hepatic tissue was detected by Masson stain.The mRNA expression of α-SMA,TGF-β1 in liver were detected by RT-PCR.The hydroxyproline(Hyp) level in liver was detected by enzyme digestion method.Serum TNF-αand TGF-β1 were detected by ELISA.Serum hepatic fibrosis parameters were detected by chemiluminescence and serum liver functional parameters were detected by biochemistry method.Results Liver functional parameters of Art + BMSCs transplantation group were better than those of the other groups.ALB in Art + BMSCs transplantation group was (29.13 ± 3.48) g/L,which was significantly higher than (26.31 ± 2.42) g/L in Art treated group (24.35 ±3.12)g/L in BMSCs transplantation group and (20.14 ±2.86) g/L in model group.There were significant differences among the 4 groups(q =6.27 ～ 19.01,all P ＜ 0.05).Liver fibrosis parameters of Art + BMSCs transplantation group were better than other groups.Hyp in Art + BMSCs transplantation group was (1.03 ±0.16) μg/L,which was significantly lower than (1.56 ± 0.19) μg/L in Art treated group,(1.45 ± 0.24) μg/L in the BMSCs transplantation group and (3.57 ±0.91) μg/L in the model group.There were significant differences among the 4 groups (q =20.47 ～ 27.70,all P ＜ 0.05).The expression of TGF-β1 mRNA of Art + BMSCs transplantation group was lower than other groups.The index of TGF-β1 mRNA expression in Art + BMSCs transplantation group was (20.16 ± 5.21),which was lower than (58.15 ±9.16) in Art treated group,(46.31 ±6.82) in the BMSCs transplantation group and (97.08 ± 14.23) in the model group.There were significant differences among the 4 groups (q =20.35 ～ 44.93,all P ＜ 0.05).Conclusion There're synergistic effects between Art and BMSCs in the treatment of liver fibrosis.

Lung distension belongs to chronic obstructive pulmonary disease (COPD) in modern medicine.In recent years,its disease incidence and mortality have increased year by year.Supported by the traditional Chinese medicine (TCM) standardization of the State Administration of TCM project-TCM preventive treatment practice guidelines-prevention of lung distension recrudescence,the first draft of the TCM Preventive Treatment Practice Guideline on Prevention of Lung Distension Recrudescence was formulated on the basis of preventive treatment theory,evidences from both ancient and modern literatures,and through the Delphi method as well as expert consensus conference.And then,peer review comments were collected,which laid the foundation for the further improvement of this guideline.This guideline was aimed to achieve the industry consensus,to improve the quality of life (QoL) in patients of the stable period,to reduce the economic burden of patients,as well as to improve both the theoretical connotation and practical value of preventive treatment in TCM.

Objective To test synchronization of cardiac mechanical contraction by means of advanced echocardiography and investigate the correlation of left ventricular ejection fraction (LVEF) and the indexes of mechanical dyssynchrony, and the relationship between DTI, STI and RT-3DE.Methods Control group (20 cases), chronic heart failure with a widened QRS complex (12 cases) and chronic heart failure with a shortened QRS duration (10 cases) were selected. We evaluated mechanical dyssynchrony with the DTI, the STI and the RT-3DE, and analyzed the correlation between the improvement degree of cardiac function and indexes of mechanical dyssynchrony, and the correlation between DTI, STI and RT-3DE. Results (1) In CHF groups (including shortened QRS group and widened QRS group), the indexes of synchronization of cardiac mechanical contraction were higher than control group ( <0.05) . (2) However, the indexes of mechanical dyssynchrony before operation showed no statistically significant difference between the widened QRS group and the shortened QRS group (>0.05) . (3) In CHF groups (including shortened QRS group and widened QRS group), the indexes of mechanical dyssynchrony before operation were higher more than after operation ( <0.05) . (4) In postoperation, the indexes of mechanical dyssynchrony showed no statistically significant difference between the widened QRS group and the shortened QRS group ( >0.05) . (5) There was a significant negative correlation between the LVEF and the indexes of mechanical dyssynchrony (<0.01) . (6) .In the indexes of synchronization of cardiac mechanical contractions, there are significant positive correlations between the DTI, the STI and the RT-3DE ( <0.01) . Conclusion Echocardiography can be used to screen CHF patients,and patients with left ventricular synchronous (including shortened QRS duration) can also be benefited from CRT.

Objective:To provide microsurgical anatomy data in the course, branch, distribution, arterial network profile of the submental artery and the range of the flap excision in submental flap transplantation.Methods:From March, 2015 to March, 2020, a total of 36 head and neck cast specimens were studied. Acrylic-butadience-styrene plastic (ABS) filler were perfused into the external carotid artery to make cast specimens. The course, branching, distribution and the arterial framework of the submental artery under a surgical microscope were investigated.Results:The submental artery originated from the facial artery before reaching the lower edge of the mandible (1.50±0.50) cm, with a diameter of (1.50±0.85) (0.6-2.3) mm. The main trunk of submental artery was (5.5±0.5) cm in length, which ran forward along the lower edge of the mandible and branched out (9.0±3.0) (7-13) branches with diameters between 0.1-0.5 mm, and mainly distributed to skin and superficial fascia of the submental area. The main trunk of submental artery divided into ascending, horizontal and descending branches about 3.0 cm of the midline of the mandible. The ascending branch went upwards over the lower edge of the mandible and joined up with the lower labial arch or participated in the formation of the lower labial arch; the horizontal branch divided into several branches and joined up with the branches from the opposite side; the descending branch branched posteriorly and inferiorly, joined up with branches of lingual artery and superior thyroid artery. The branches of the submental artery and the branches of the peripheral arteries were joined up in the submental area to form the submental artery network. The diameter of the vessels in the network ranged 0.1-0.2 mm. The arterial network was built in the form of 1 to 3 layers, and the area of main network was about 7.0 cm×5.0 cm.Conclusion:The submental artery has a long trunk, many branches and abundant anastomoses between the branches, forming a dense submental artery network, which provides sufficient pedicle length, rich blood supply and cutting area for submental flap. The flap can be transplanted free or transposed. The best location of submental flap is near the midline of arterial network, and the appropriate area is 7.0 cm×5.0 cm.

ObjectiveTo explore the neural mechanism of attention impairment in children with primary nocturnal enuresis.MethodsERPs elicited by performing the continuous operation test(CPT) were assessed in 20 children with primary nocturnal enuresis and 20 normal children.The Go/Nogo measurements of enuretic group at central scalp(Cz) were compared with the normal children and analyzed.Results1.Behavior results: there was no significant difference in the reaction time,the correct number and the false number between primary nocturnal enuresis and control group(P>0.05).2.ERP:(1)Go stimulate:the latency of Go-N2 and P3 of the children with primary nocturnal enuresis were longer than the normal control group(Go-N2:(326.80±46.40)ms vs (295.90±38.27)ms,P3:(438.80±62.60)ms vs (402.60±39.74)ms),and the difference had statistic significance(P<0.05).(2)Nogo stimulate:①Amplitude: the amplitude of Nogo-N2 of the children with primary nocturnal enuresis were lower than that of the normal control group((-10.55±3.30)μV vs (-14.12±5.99)μV),and the difference had statistic significance(P<0.05).There was no significant difference in the amplitude of Nogo-P2 and Nogo-P3(P>0.05).②Latency: the latency of Nogo-P2 of the children with primary nocturnal enuresis was longer than that of the normal control group((214.10±27.85)ms vs (198.30±19.16)ms),and the difference had statistic significance(P<0.05).There was no significant difference in the latency of Nogo-N2 and Nogo-P3(P>0.05).ConclusionAttention impairment in children with primary nocturnal enuresis might be caused by the information processing speed and conflict monitoring function obstacle,but it is not because the reactive inhibition dysfunction,thus result in the lack of arousal function and bedwetting.

Objective To investigate function of Arg327Ile (R327I) and Arg327Ala(R327A) FⅨ mutants and to study the molecular pathogenesis of haemophilia B(HB) caused by R3271 mutation.Methods Hygromycin-resistant cell line was screened and the secretion of FⅨ antigen into the medium was measured by ELISA.The cell line with appropriate expression levels of F Ⅸ antigen was selected for culture.Recombinant F Ⅸ (rF Ⅸ ) was purified from concentrated medium by two step methods of Q-Sepharose Fast Flow and anion exchange chromatography.The concentration and purity of rF Ⅸ were determined by ELISA and SDS-PAGE,respectively.The activation of wild-type ( WT),R327I and R327A of rFⅨ by FⅦa/TF/Ca2+ or FⅪa/Ca2+ was identified by Western blot in different time periods.The FⅨa and FⅧa complex formed by interaction with different concentrations of FⅧa was used to activate F X,the apparent dissociation constant (Kd) for FⅧa binding was calculated by the kinetic results.The kinetic data of the activation of FX by WT,R327I and R327A FⅨa with or without FⅧa were calculated.Results The amount of WT,R327I and R327A rFⅨ were 450,210,64 μg,and the purity of rFⅨ was confirmed by SDSPAGE.Both R3271 and R327A could be normally activated by FⅧa/TF/Ca2+ or FⅪa/Ca2+.Kd for FⅧa binding showed that the binding capacities of R327I and R327A were 4 and 5 times lower than WT,respectively.The catalytic efficiencies of R327I and R327A F Ⅸ a for F X were 6 and 8 times lower with FⅧa,and 3 and 7.4 times lower without F Ⅷ a,respectively.Conclusions R327I and R327A rF Ⅸ mutants impair their binding to the FⅧa.The site on R327 contributes to FⅧa binding.It is partly related to the activation of FX.The low FⅧa binding to R327I FⅨa may cause HB.