Objective Co-stimulation cells is a kind of natual killer (NK)-like T cells, which can kill tumor cells.Previous studies show that lupeol , an natural plant extracts , can change the growth of NK cells ,γδT cells and their effects on tumor cells .This study aimed to investigate the effects of lupeol on human co-stimulation cells and colonic cancer cell lines SW 480 . Methods The peripheral blood mononuclear cells ( PBMC) from healthy volunteers were induced in vitro by different cytokines and transferred into Co-simulation cells.After SW480 and Co-stimulation cells were incubated with different concentrations of lupeol for different durations , methyl thiazolyl tetrazolium (MTT) was used to detect the effects of lupeol on co-stimulation cells and colonic cancer cell lines SW480. Lactate dehydrogenase was used to determine the cell-killing activity of Co-simulation cells to colonic cancer lines SW 480 . Results The concentration of lupeol in 0.1-200.0 mg/L promoted the growth of Co-stimulation cells and inhibited the colonic cancer cell lines SW480.When the concentration of lupeol is at 12.5 mg/L, the cell-killing activity of Co-simulation cells to colonic cancer lines SW 480 was enhanced significantly compared with the controls (76%vs 40%, P<0.05). Conclusion Lupeol could promote the prolifera-tion of Co-stimulation cells, inhibit the growth of cancer lines SW480, and strengthen the cytotoxicity of Co-stimulation cells against co-lonic cell lines SW480.

Objective:To study the effect of GABA and A receptor agonist THIP on proliferation and apoptosis of cytokine-induced killer (CIK) cells.Methods:CIK cells were cultured by routine method,and then treated with different concentrations of GABA and THIP.The proliferation of CIK cells were investigated by MTT assay.The apoptosis,cell cycle and immunophenotype were investigated by flow cytometry.Results:GABA could inhibit the proliferation of CIK cells in a dose-dependent manner and affect the distribution of cell cycle of CIK cells(P

Objective:To observe the effects of auto-dendritic cell vaccine on T cells of patients with condyloma acuminatum(CA) related human papilloma virus(HPV) and its clinical outcome.Methods:73 patients with condyloma acuminata(CA) in the present study were positive of HPV-DNA and have been unsuccessfully treated with freezing,laser,or electric therapy combined with interferon anti-virus treatment for half a year.The peripheral blood samples were harvested from patients and the monouclear cells were extracted to cultivate DCs in presence of cytokines.Auto DCs were sensitized with corresponding HPV virus antigen of a patient to prepare the auto-dendritic cell vaccine.Then the vaccine were cultivated with auto T-cells and the proliferation of T cells were examined.Meanwhile,the auto-DC vaccine(1-4)?107 was also injected into the patients' inguinal lymph glands and its therapeutic effectiveness was observed.Results:The prepared DC-vaccine significantly promoted the proliferation of auto T cells compared with control group(P

Purpose To compare the therapeutic effects of CO2 laser resection and traditional split laryngeal surgery on the early laryn-geal cancer, and to examine relationship between the expression of p27 and PTEN and clinical prognosis in early laryngeal squamous cell carcinoma. Methods 85 patients who underwent CO2 laser resection surgery and 46 patients who underwent split laryngeal surger-y were selected and the occurrence rates after surgery were observed. SP immunohistochemical method was uesd to detect the expression of p27 and PTEN in tumor resection marginal and their surgical margine tissues. Clinical data were collected and all patients were fol-lowed up. Results In the more than two-year follow-up study, 14 of 85 patients in CO2 laser treatment group ( recurrent group) pres-ented with local recurrence and the recurrence rate was 16. 5%, while 6 of 46 patients in split laryngeal surgery group presented with local recurrence and the recurrent rate was 13. 0%. There was no statistical significance in recurrence rate between the two groups ( P> 0. 05). 10 of all the none recurrent patients did not follow the doctor’s advice to quit smoking after the operation, while 12 in the recurrent patients did not, the difference between the two groups was statistically significant (P<0. 01). The positive rate of p27 and PTEN in laryngeal carcinoma tissues and the cancer adjacent tissues ( negative surgical margin tissues ) was 43. 5% ( 57/131 ) , 80. 2% (105/131) and 48. 9% (64/131), 83. 2% (109/131), respectively, with a significant difference (P<0. 01). The positive rate of p27 and PTEN in laryngeal carcinoma tissues of the recurrent group and non recurrent group was 20. 0% (4/20), 47. 7% (53/111) and 10. 0% (2/20), 55. 9% (62/111), respectively, with a significant difference (P<0. 05). While the positive rate of p27 and PTEN in tumor resection marginal tissues of the recurrence group and non recurrence group was 50. 0% ( 10/20 ) , 85. 6% ( 95/111) and 40. 0% (8/20), 91. 0% (101/111), the difference was also statistically significant (P<0. 01). Conclusions There was no statistically significant difference in tumor recurrence rate between CO2 laser surgery and traditional split laryngeal surgery. Postoper-ative recurrence is closely related to resume smoking. The recurrence rate of p27 and/or PTEN negative patients was higher than that of the opposite ones which should be followed up closely after treatment.

Purpose To study the expression of p53, p21 and Cdk1/p34cdc2 in the laryngeal cancer and its margin tissues and to ex-plore their relationship with local recurrence of laryngeal cancer. Methods A total of 85 patients with early laryngeal cancer were se-lected randomly during 2004 to 2010 in Tangshan Union Hospital, Hebei, China. SP immunohistochemical method was used to detect the expression of p53, p21 and Cdk1/p34cdc2 in the tumor and margin tissues. Pathological data were collected for follow-up. Results In more than 2 years of follow-up study, 14 of 85 patients with laryngeal cancer presented with recurrence (recurrent group), while 71 patients without recurrence (none recurrent group). The positive rate of p53 protein in laryngeal cancer and its margin tissues was 60. 0% and 36. 5%, respectively, the positive rate of p21 protein in laryngeal cancer and its margin tissues was 38. 8% and 21. 2%, respectively. The positive rate of Cdk1/ p34cdc2 in laryngeal cancer and its margin tissues was 70. 6% and 29. 4%, respectively. p53 protein in the surgical margin of the recurrent group and non recurrent group was 71. 4% and 29. 6% (P = 0. 003), that of p21 was 50. 0% and 15. 5%, (P =0. 004) and Cdk1/ p34cdc2 was 57. 1% and 23. 9% (P =0. 013), respectively. There was no correlation between expression of p53 with p21 protein and Cdk1/ p34cdc2 protein(P > 0. 05). Conclusion p53, p21 and Cdk1/ p34cdc2 may be involved in the occurrence, development and recurrence of laryngeal squamous cell carcinoma. Overexpression of p53, p21 and Cdk1/ p34Cdc2 in the surgical margin is closely related to local recurrence of laryngeal cancer.

Objective To investigate the method and effectiveness of the digital artery series and parallel of island flap for repairing degloved injuries of the fingertip of thumb or adjacent finger Methods Between September 2008 and July 2012,the finger artery series retrograde island flaps tiled in the repair of finger degloving injury in 13 cases,repair of adjacent finger tip degloving injury in 11 cases; 8 cases of degloved injuries of the fingertip were tiled with the digital artery parallel island flap from ulnaris middle finger and radialis ring finger of arteria digitalis communis pedicled which were from the same palm side.The size of skin and soft tissue defect ranged from 2.0 cm × 1.8 cm-7.2 cm ×5.5 cm,gutted flap minimum ranged from 1.1 cm × 1.0cm-1.5 cm × 1.3cm,and the maximum ranged from 3.0 cm × 2.2 cm-5.5 cm × 4.5 cm.The donor sites were repaired with the intermediate split thickness free skin grafts and performed with pressure dressing.Results The group of 32 cases,in 1 case the flap vasospasm occurred in operation of free process,the symptoms disappeared after local application of papaverine and hot compress ; 1 case of flap occurred disturbance of blood circulation after operation because of tight suturing for pedicle,the symptoms relieved after removing the stitches at intervals; 2 cases blisters appeared after operation,disappeared after a week.All skin flaps were survived,incision and skin graft donor sites healed by first intention.Twenty patients were followed-up from 6 to 12 months after operation.All flaps presented the satisfactory appearance and texture,recovered protective feeling.At last follow-up,the two-point discrimination was 7 to 10mm,and the flexion and extension function of wounded fingers recovered to normal.According to the functional assessment criteria of upper limb formulated by the Hand Surgery Branch of Chinese Medical Association,there were excellent in 19 cases,good in 10 cases,and moderate in 3 cases.No significant loss to the donor shape and function.Conclusion The surgery by adopted the digital artery series or parallel of island flap for repairing degloved injuries of the fingertip of thumb or adjacent finger,not only has the advantages of simpleness,safety and reliability,but also can satisfy the patients who aren't willing to accept or because of physical reasons can't accept the treatment of abdominal skin tube or nail flap from hallux toe,which performs in both high-end and low level hospitals,and deserves of general application.

Objective To evaluate the surgical methods and clinical effects of the reconstruction of soft tissue defect in distal thumbwith dorsal thumb neurocutaneous vascular flap.Methods From January,2014 to December,2016,32 patients with soft tissue defects of distal thumb were repaired with dorsal thumb neurocutaneous vascular flaps,including left thumbs in 11 cases and right thumbs in 21 cases.They were including 7 cases of nail bed defects,9 cases of pulp defects under nails,16 cases ofthe defects of tips or stump of fingers;skin and soft tissue defects range of 1.5 cm×1.0 cm-3.5 cm×3.2 cm;Flap cutting areas range of 2.0 cm×l.5 cm-4.0 cm×3.5 cm.If the donor areas could be sutured directly,be sutured;if they could not,be covered with intermediate split thickness skin grafts.All patients were followed up when they were reviewed in outpatient department,and to be evaluated the shape,texture,sensation,flexion and extension function,and donor site of the flaps.Results One case of flaps blood circulation disordereddue to tight suture,and eased after the removal of part of the sutures;One case of flaps atrophied,and the phalangette was exposed after 3 months,so we removed the end part of bone and the wound was closed;other flaps were survived,with primary wound healing.In the total 32 patients,they were followed up range of 3 to 36 months,with an average of 12 months.Eighteen cases were followed up at least 6 months,which were 4 of the 7 cases of nail bed defects,5 of the 9 cases of pulp defects under nail and 9 of the 16 cases of the defects of tip or stump of fingers.The appearances and textures of flaps were good,protective sensations were restored,and skin flap two-point discriminationswere 8-10 mm.According to the Trial Standard for Evaluation of Upper Limb Function of Chinese Society of Hand Surgery,it was excellent in 11 cases,good in 17 cases and moderate in 4 cases,with the excellent and good rate of 87.5％.Conclusion It has advantages of simple operation,low risk,high success rate and satisfactory curative effects of the use of dorsal thumb neurocutaneous vascular flap for repair of distal thumb defect of skin and soft tissue.It is not only suitable for the hospital with good technical and equipment,but also suitable for the primary hospital.

Objective To investigate the effects of dendritic cells (DCs)loading alpha-Galactosylce-ramide (α-GalCer)combined with tumor specific cytotoxic T lymphocytes (CTLs)on the growth of transplanted Heps hepatoma in mice.Methods We induced the augmentation of the DC cells and T lymphocyte derived from the mice bone marrow,and enabled them to be specific CTLs.DC cells loaded α-GalCer in vitro.First we established a Heps liver cancer xenograft model,then divided the model mice into 4 groups by random number table method (n =9):control group (intravenous injection with physiological saline),CTL group,DC loadingα-Galcer group and DC loading α-Galcer combined with CTLs group.After 2 weeks of intervention,we extrac-ted the tumor tissue,weighed the tumor and calculated the inhibition rate of tumor.The expressions of Bax/Bcl-2 cells in groups of transplanted tumor tissues were detected using immunohistochemistry and Western blotting. Results The average tumor weight of CTL group,DC loading α-GalCer group and combined treatment group were (1 .07 ±0.1 5)g,(1 .1 1 ±0.1 7)g,(0.79 ±0.1 4)g,respectively.All of them were lower than that of control group (1 .69 ±0.23)g,with significant differences (t =1 4.1 76,P =0.023;t =1 2.351 ,P =0.034;t =1 8.672,P =0.000).The average tumor weight of combined treatment group was lower than those of the CTL group and DC loading α-GalCer group,with significant differences (t =1 5.236,P =0.01 2;t =1 1 .1 76, P =0.037).Compared to the CTL group (36.69%)and DC loading α-GalCer group (34.32%),the com-bined treatment group had a higher tumor inhibition rate (53.25%;P =0.034,P =0.021 ).Immunohisto-chemical assay showed that the numbers of Bax-positive cells in CTL group,DC loading α-GalCer group and combined treatment group were 35.83 ±0.75,33.67 ±0.82,41 .1 7 ±1 .1 7 respectively,and compared with the control group (21 .67 ±2.1 6),the differences were statistically significant (t =-1 3.789,P =0.002;t =-1 5.1 1 6,P =0.001 ;t =-1 7.452,P =0.000).The numbers of Bax-positive cells in combined treatment group were different with CTL group and DC loading α-GalCer group (t =-7.730,P =0.009;t =-5.872, P =0.01 1 ).The numbers of Bcl-2-positive cells in CTL group,DC loading α-GalCer group and combined treatment group were 30.83 ±0.75,31 .67 ±1 .03,25.00 ±0.89,and compared with the control group (38.67 ±1 .21 ),the differences were statistically significant (t =9.234,P =0.007;t =1 1 .738,P =0.003;t =20.608,P =0.000).The numbers of Bcl-2-positive cells in combined treatment group were different with CTL group and DC loading α-GalCer group (t =1 1 .952,P =0.003;t =1 2.223,P =0.002).Western blot-ting test results showed that the expression levels of Bax in CTL group,DC loading α-GalCer group and com-bined treatment group were 0.46 ±0.01 ,0.42 ±0.03,0.55 ±0.01 ,and compared with the control group (0.31 ±0.02),the differences were statistically significant (t =1 .035,P =0.032;t =1 .1 24,P =0.027;t =1 .425,P =0.01 0).The expression level of Bax in combined treatment group was different with CTL group and DC loading α-GalCer group (t =1 .305,P =0.01 3;t =1 .421 ,P =0.01 0).The positive expressions of Bcl-2 in CTL group,DC loading α-GalCer group and combined treatment group were 0.34 ±0.03,0.33 ± 0.02,0.24 ±0.01 ,and compared with the control group (0.46 ±0.01 ),the differences were statistically sig-nificant (t =-1 .1 23,P =0.025;t =-1 .061 ,P =0.031 ;t =1 .278,P =0.01 4);the positive expression level of Bcl-2 in combined treatment group was different with CTL group and DC loading α-GalCer group (t =1 .1 60,P =0.021 ;t =1 .21 9,P =0.01 5).Conclusion It has synergistic killing effect on transplanted Heps hepatoma in mice using DC loading α-GalCer combined with the tumor specific CTL.

ObjectiveTo observe the effect of betulinic acid(BetA) on the growth of human cytokine induced killer(CIK) cells and the killing activity of CIK cells on the gastric cancer cells in vitro before and after induced by betulinic acid,explore its mechanism.MethodsPeripheral blood mononuclear cell (PBMC) were separated form the healthy and were induced with various of cytokine to become CIK cells in vitro.CIK cells were collected on the tenth day and were induced with betulinic acid in different concentrations,followed by 48 h,the colorimetric methyl thiazolyl tetrazolium(MTT) method assay the proliferation rate of human CIK cells.Flow cytometry (FCM) was used to detect the expression changes of perforin,granzyme B and CD107a of human CIK cells before and after betulinic acid-induced.Lactate dehydrogenase (LDH) release assay was used to measure the influence on cytotoxic activity of CIK cells induced by betulinic acid against gastric cancer cell line SGC-7901 in vitro.Western blot assay was used to measure the extracellular signal-regulated kinase1/2 (ERK1/2),and adapter proteins SH2-domain containing leukocyte protein of 76KD(SLP-76) and linker for activative of T cells(LAT) expression changes of human CIK cells before and after drug-induced.ResultsBetulinic acid can promote CIK cells growth when the concentration were in 0.08-10 μg/ml,the expression of perforin,granzyme B and CD107a of CIK cells were significantly higher than control group(P＜0.05) when the concentration of betulinic acid were in 0.3 μg/ml.In the meanwhile,the cytotoxic activity of CIK cells in vitro against gastric cancer cell line SGC-7901 were also remarkably higher than the control group (P＜0.05).The expression of SLP-76,LAT and ERK1/2 were significantly increased to a certain extent than the control group( P＜0.05 ),when CIK cells were treated with betulinic acid.ConclusionThese results suggest that betulinic acid can promote CIK cells growth in some concentrations and increase the cytotoxic activity of CIK cells against gastric cancer cell line SGC-7901,its mechanism may related with two factors,on the one hand,enhancing the activity of SLP-76,LAT and ERK1/2,on the other hand,increasing the expression of perforin,granzyme B and CD107a on the surface of CIK cells.