Objective To apply fluorescence-activated cell sorting (FACS) and magnetic activated cell sorting (MACS) in sorting natural killer (NK) cells from human peripheral blood. The efficiency of these two methods were compared so that the basic conditions for the further study on the lower proportion of peripheral blood cells could be created. Methods Twenty cases of normal peripheral blood and 3 cases of post-hematopoietic stem cell transplantation (post-HSCT) were collected and NK cell were sorted by FACS and MACS. Results Before sorting the normal peripheral blood, the proportion of the NK cell in the leukocyte was (12.86±3.62) %, the purity of NK cell was up to (96.15±2.03) % and the recovery was (95.08±2.16) %after sorting by FACS, the purity was up to (93.35±3.61) % and the recovery was (94.11±3.01) % by MACS.And before sorting the post-HSCT peripheral blood, the proportion of the NK cell in the leukocyte was (11.01±2.08) %, the purity of NK cell was up to (96.22±2.16) % and the recovery was (95.27±1.18) % after sorting by FACS. The purity was up to (90.98±1.94) % and the recovery was (94.54±3.52) % by MACS. Thereis difference in purity of NK cell (t = 5.925, P ＜0.05), while no difference in rate of recovery between 2methods (t = 0.789, P ＞0.05). Conclusion FACS is more efficient, rapidly and easily than MACS in sorting NK cell, especially for post-HSCT cases.

Objective To observe the changes of CD+8CD+28,CD+8CD-28 and CD+4CDhigh25CDlow127 regulate T (Treg)cellsin peripheral blood of lung cancer patients,and to analyze the correlation between CD+8CD-28 and CD+4CDhigh25CDlow127 Treg cells to reveal the role and clinical significance of them in lung cancer patients.Methods Flow cytometry was applied to evaluate the level of CD+8CD+28,CD+8CD-28 and CD+4CDhigh25CDlow127 Treg cells in peripheral blood of 60 untreated lung cancer patients and 60 healthy controls group.The association of each term with clinical features was analyzed.Results The percentage of CD+8CD-28 and CD+4CDhigh25CDlow127 Treg cells in lung cancer group[(58.430:15.749) ％,(7.365±2.025) ％]was significantly higher than those in healthy group [(41.057±15.436)％,(6.648±1.669)％,(t=6.102,P＜0.05;t=2.115,P＜0.05)],while the percentage of CD+8CD+28cells is lower(41.570±15.739)％ than that in healthy group[(58.700±15.298)％,(t=-6.043,P＜0.05)].No close associations were found between three index and gender,age,and biological characteristics.With the increase of TNM stage,The percentage of CD+4CDhigh25 CDlow127 Treg cells increased gradually,which was remarkably higher in patients rith stage Ⅳ than that with stage ⅢA(t=-3.898,P＜0.05).The percentage of CD+4CDhigh25 CDlow127 Tregcells was uncorrelated with CD+8CD-28 cells(r=-0.169,P＞0.05).Conclusion The higher percentage of CD+8CD-28 and CD+4CDhigh25 CDlow127 Treg cells,the lower percentage of CD+8CD+28 cells may be the important reasons of immune suppression in lung cancer patients.Though there is no correlation between CD+8CD-28 and CD+4CDhigh25 CDlow127 Treg cells,it is may be helpful to understand immunologic function and it may look for more specific therapy and provide a new reference in the prognosis of lung cancer.

Objective To detect the levels of CD4+CD25HiCD127Low regulatory T cells (Treg) in the peripheral blood and its clinical significance in patients with esophageal cancer. Methods The levels of Treg in the peripheral blood were detected by three-color flow cytometry (FCM) in 80 patients with esophageal cancer and 20 healthy controls. Among the 80 patients, 30 patients were also further studied for preoperative and postoperative comparison after operation. The clinical and pathological data of each patient were collected and analyzed for the correlation with the Treg levels. Results The level of Treg in the peripheral blood of the control group was lower than that of the esophageal cancer patients [(3.36±1.14) % and (5.70±1.96) %, respectively], with significant difference (P <0.01). The levels of Treg in the peripheral blood was higher in the patients with metastasis of lymph node (n=40) than that in the patients without metastasis of lymph node (5.96±1.36) % and (4.23±1.18) %, respectively] (n=30), with significant difference (P <0.01). The levels of Treg in the peripheral blood of the patients were negatively correlated with their TNM classification. As the TNM classification advanced, the level of the Treg in the peripheral blood increased. Conclusion The levels of Treg in the peripheral blood of the patients with esophageal cancer are significantly higher than that of the healthy subjects, which is correlated with the clinical and pathological conditions. The development of esophageal cancer may relate with suppression of immune function.

Effect and safety of repeated embryo aspiration for reduction of muhifetal cervical pregnancy Was retrospectively evaluated.Three women with muhifetal cervical pregnancy successfully received conservative treatment of repeated embryo aspiration under the guidance of transvaginal ultrasound,and the gestational sac contents were aspirated completely.Local(2.5～5.0 mg injectable methotrexate,MTX) or systemic medication was used.Our findings showed that repeated embryo aspiration for reduction of cervical multifetal pregnancy could improve the coupe of disease,decrease systemic medication of MTX,and discontinue a cervical pregnancy within 8 weeks effectively and safely.However,the mminimum therapeutic dose of MTX remained to be explored.

Objective To explore the clinical significance of T lymphocyte subsets and NK cells' detection in peripheral blood of patients with non-Hodgkin' s lymphoma (NHL).Methods 62 newly diagnosed NHL and 30 healthy adults were enrolled for analyzing.T lymphocyte subsets and NK cells were deteced by flow cytometry and compared between different groups according to age,sex and pathology.Multiple linear regression was used to analyze the correlation among the levels of T lymphocyte subsets and NK cells and several clinical factors,such as clinical stage,IPI and B symptoms.Results The proportions of CD3,CD4 and NK cells in peripheral blood of NHL group [(64.13±19.83) ％,(27.10±8.20) ％,(13.51± 10.59) ％] were significantly lower than those of normal control [(65.78±10.69) ％,(31.95±12.74) ％,(18.76± 7.36) ％] (P ＜ 0.05),while the proportions of CD8 and Treg cells in peripheral blood of NHL [(32.15±13.83) ％,(10.44±3.00) ％] were significantly higher than those of normal control [(29.25±12.35) ％,(7.51±4.36) ％] (P ＜ 0.05).In NHL,the proportions of CD3 and NKT cells [(67.06±19.24) ％,(4.91±3.69) ％] in ≤60 years old group were significantly higher than those in ＞ 60 years old group [(59.18±20.33) ％,(4.89±3.05) ％] (P ＜ 0.05).The proportions of Treg cells in T-NHL was significantly higher than that in B-NHL [(8.17±6.41) ％ vs (7.11±2.53) ％] (P ＜ 0.05).The proportions of CD3,CD4,NK and NKT cells in peripheral blood of NHL were significantly correlated with clinical stage (P ＜ 0.05),that of CD8 cells were significantly correlated with IPI (P =0.000),that of Treg cells were significantly correlated with IPI and B symptoms (P ＜ 0.05).Conclusion The cellular immune is suppressed in NHL patients.The proportions of T lymphocyte subsets and NK cells in peripheral blood of patients with NHL are different between different groups according to age,sex and pathology.The correlations among that and clinical factors need to be further explored.

This study established an HPLC fingerprint of Tibetan medicine Shaji Gao from different habitats and lay a foundation for Shaji Gao varieties identification and preparation process. The chromatographic condition was as follow: Agilent zorbax SB-C18 (4.6 mm x 250 mm, 5 μm) eluted with the mobile phases of acetonitrile and 0.4% phosphoric acid water in gradient mode. The flow rate was 1.0 mL x min(-1), and the detection wavelength was set at 360 nm. The fingerprints of 15 batches Shaji Gao were carried out by similarity comparation, 7 chromatographic peaks were extracted as the common peaks of fingerprint, 3 peaks were identified, which were quercetin, kaempferol and isorhamnetin. The similarity degrees of 14 batches of samples were above 0.9 and 1 batch of samples was below 0.9. This is the first established fingerprint of Shaji Gao by using HPLC. This method has good precision, stability and repeatability that it could provide basis for quality control and evaluation of Shaji Gao.
Chromatography, High Pressure Liquid ; methods ; Medicine, Tibetan Traditional ; Quality Control

Objective To investigate the effect of idarubiein to imensify the MAC (IMAC) as preparative regimen for autologous peripheral blood stem cell transplantation(APBSCT)in acute myeioid leukemia.Methods Fourteen patients with acute myeloid leukemia who underwent APBSCT were analysed. IMAC was used as preparative regimens.Results All patients were engrafted successfully.The disease-free survival could be Ben in 8 cases(57.1%),the median disease-free survival duration were 26(8-72)months and no treatment-related mortality was present.Conclusion This study suggested that addition of Idarubicin to the MAC preparative regimens Was safe,effective and feasible for patients with acute myeloid leukemia,and may improve disease-free survival and overall survival.

Objective To investigate the effect of ventricle-peritoneal (V-P) shunt in treatment of patients who developed post-traumatic hydrocephalus (PTH) with no symptoms or with atypical symptoms due to an excessively severe traumatic brain injury (TBI).Methods A retrospective study was performed in patients who developed PTH with atypical symptoms undergone V-P shunt from January 2004 to June 2007.Patients' general information,TBI data,PTH severity and postoperative follow-up data were collected and applied to assay the improvement rate in prognosis and its associated factors.Results Thirty-one patients were involved in this study.After 12 months of follow-up,20 (65％) patients revealed significant improvements in clinical symptoms.Among 10 patients who developed PTH after decompressive craniectomy,cranioplasty was performed following V-P shunt and significant improvements were observed in nine patients.Patients' age and severity of PTH based on CT evaluation before shunt placement were strongly correlated with the surgical outcome.Conclusions Most PTH patients with atypical symptoms can benefit from V-P shunt.Additionally,younger patients and those with less severe PTH before V-P shunt are expected a better outcome.

Objective To investigate the effects of large dose of atorvastatin on the expression of Sprouty-1 in CD4 + T lymphocytes from unstable angina patients during perioperative period of PCI.Methods A total of 52 unstable angina patients enrolled were divided randomly (random number) into large-dose atorvastatin (80 mg/d,n =26) pretreated group and moderate-dose atorvastatin (20 mg/d,n =26) pretreated group.Circulating CD4 + T cells were obtained by magnetic cell sorting system (MACS) before PCI and 18-24h after PCI.For detecting the gene expression,the reverse transcription fluorescent quantitative polymerase chain reaction (RT-PCR) was used to measure the expression of Sprouty-1 mRNA in CD4 + T lymphocyte.The level of Sprouty-1 protein was detected with Western blot analysis and IL-2 was quantified by enzyme-linked immunosorbent assay (ELISA).Results ①Compared with large-dose group before PCI,the expression of Sprouty-1 mRNA and Sprouty-1 protein levels were dramatically increased in large-dose group after PCI (P ＜ 0.05).②Compared with moderate-dose group before PCI,the expression of Sprouty-1 mRNA and protein levels were slightly increased in moderate-dose group after PCI,but there was no statistical significance (P ＞ 0.05).③Compared with large-dose group before PCI,the serum level of IL-2 was decreased in large-dose group after PCI (P ＜ 0.05).Whereas the serum level of IL-2 was slightly increased in moderate-dose group after PCI compared to moderate-dose group before PCI,there was still no statistical significance (P ＞ 0.05).Conclusions Large-dose atorvastatin pretreatment reduced post-PCI myocardial inflammation through up-regulating the expression of Sprouty-1 mRNA and level of Sprouty-1 protein in CD4 + T lymphocytes.

Objective To analyse the dysfunction of immunity and clinical significance in patients with cardiac cancer.Methods The level of CD4+ CD25hi CD127low Treg cells were detected by flow cytometry (FCM),and serum IL-10 and TGF-β1 levels were determined by enzyme linked immunosorbent assay (ELISA) kit in 56 patients with cardiac cancer.15 healthy volunteers were tested as normal controls.The clinical data of each patient were collected and analyzed. Results There was a significantly higher percentage of CD4+ CD25hi CD127low Treg cells in patients with cardiac cancer (5.73±1.56)% than that (4.45±1.06)% of healthy volunteers (P<0.01).The IL-10 and TGF-β1 levels in the serum of patients with cardiac cancer were also significantly higher than that of healthy volunteers (P<0.05).There was a positive correlation between levels of IL-10.TGF-β1 and the levels of CD4+ CD25hi CD127low Treg cells.The number of CD4+ CD25hi CD127low regulatory T cells in the peripheral blood of cardiac cancer patients were significantly correlated with clinical stages and metastasis lymph node.Conclusion The CD4+ CD25hi CD127low Treg cells in the peripheral blood of cardiac cancer patients is significantly increased in comparison with that in healthy volunteers,and was also correlated with different stages.The abnormal levels of CD4+ CD25hi CD127low Treg cells may be related to tumor progression in patients with cardiac cancer.