OBJECTIVE: To investigate the optimizing conditions in isolation of the side population in laryngeal carcinoma cell line Hep-2. METHOD: Single-cell suspension cells were detached from the culture flask with trypsin EDTA, at a concentration of 1 x 10(6) cells/ml. (1) The trail Samples were incubated with Hoechst33342 at a concentration of 5 microg/ml, 9 microg/ml, 10 microg/ml, 11 microg/ml for 90 minutes. (2) They were incubated with Hoechst for 50, 70, 90, 110, 130 min in water bath individually. (3) The single-cell suspension were incubated Hoechst in water bath and in thermostat each. (4) The two different density of cells were harvested, which were 100% and 70%, and then di gest into single-cell suspension. Once incubation finished, suspended in phosphate buffered saline (PBS), then test SP% by flow cytometry. Among all groups,Verapamil hydrochloride was added to the control samples, incubated at 37 degrees C for 30 minutes, the other condition were keep the same with their trial groups. RESULT: (1) The percentage of Hoechst-negative cells in trial group was (39.96 +/- 0.24)%, (26.23 +/- 0.39)%. (18.79 +/- 0.02)%, (19.01 +/- 0.14)% at the concentration of 5 microg/ml, 9 microg/ml, 10 microg/ml, 11 microg/ml respectively, when the PI-positive cells were (30.45 +/- 0.63)%, (49.9 +/- 0.42)%, (50.12 +/- 0.68)%, (64.16 +/- 0.39)% separately. (2) Varying the duration of staining incubation showed that there was a typical FACS pattern and SP% was constant when the incubation was at least 90 min. (3) Compare to water bath, SP% was more than in thermostat, the SP% was (18.67 +/- 0.45)%, (22.6 +/- 0.50)% respectively; (4) Cell density is also responsible for SP%. The low density the cell is, the less in SP%. SPSS13.0 was used in statistical analysis, the groups were compared using t-Test. P < 0.05 was considered statistically significant. CONCLUSION The optimum concentration and duration of incubation of Hoechst33342 in isolation of the side population cells in laryngeal carcinoma cell line Hep-2 is 10 microg/ml and 90 min. Incubated in water bath is better than in thermostat. The best staining cell density is around 80%-90%.
Cell Count ; Cell Line, Tumor ; Flow Cytometry ; Humans ; Laryngeal Neoplasms ; pathology ; Neoplastic Stem Cells ; cytology ; Side-Population Cells ; cytology

Apathy is a common neuropsychiatric complication after stroke,the identification of apathy after stroke and its related factors and providing an effective intervention on it may contribute to the rehabilitation in patients with stroke.This article reviews the incidence of apathy after stroke,pathophysiological mechanisms,diagnostic evaluation and treatment.

Objective To explore a novel surgical treatment for chronic suppurative otitis media (CSOM) and evaluate its treatment effect.Methods All 97 patients with chronic suppurative otitis media were chosen to be treated using this new surgical method.The skin of the external auditory canal was maintained intact.Open radical mastoidectomy was used to complete clean-up lesions ; the fascia of pedicled temporalis myofascia (PTM) was used to repair the tympanic membrane.The pedicled temporalis fascia,pedicled postauricular periosteal flap and intact skin of the external auditory canal were used in reconstruction of the posterior wall of external auditory canal.Pure tone audiometry (PTA) was performed before and after surgery,recording the air conduction and bone conduction thresholds at 0.5 kHz,1.0 kHz,2.0 kHz,4.0 kHz.The average of the patient's air and bone conduction hearing thresholds was recorded at the 4 frequencies.External auditory canal gauze was removed 3 weeks after surgery.All subjects were followed up for over 2 years.Comparison of hearing thresholds (PTA) was made ① Before and 4 weeks after surgery.② Before and 2 years after surgery.Hearing function comparison include air conduction (AC),bone conduction (BC) and air-bone gap (ABG) analysis.SPSS 16.0 was used in statistical analysis.Pre-and postoperated AC,BC and ABG were compared using T-test.P ＜ 0.05 was considered statistically significant.Results The healing rate of post-operated tympanic membrane was 95.88％ (93/97).Ninty-six ears had 2-year follow-up,and 1 patient was lost in follow-up.There were 2 patients presented with eardrum perforation during the follow-up,and the 2-year healing rate of tympanic membrane perforation was also 93.85％ (92/97).In 96 ears with 2-year followed-up,the average of pre-AC was (52.10 ±3.96) dB,the average of post-AC was (35.67 ±2.52) dB; the average of preABG was (36.6 ± 5.2) dB,and the average of post-ABG (± SD) was (12.14 ± 6.20) dB.Statistical analysis showed significant difference between preoperative and postoperative AC or ABG values (P ＜ 0.05).Conclusion The present surgical procedure broke through the existing conventional mastoidectomy of making a surgical incision in the posterior wall of the external auditory canal.This procedure cleared the lesion completely and preserved the physiological function of the external auditory canal.The acoustic systems and state of gasification to the mastoid tymnpanum were reconstructed,rehabilitated and maintained.The healing rate of hearing and tympanic membrane perforation was improved.

Objective:To experimental study the inhibiting effects of SBHL on the growth of cervical cancer cells and to investigate the mechanisms.Methods:The proliferation of ME180 cells was observed by MTT assasy; The changes of cells cycles and apoptosis of ME180 cells treated with SBHL were analyzed by agar gel electrophoresis and FCM assay;. The changes of the ultra structure of cells was observed by Transmission eleconmicrograph.Results:The proliferation of ME180 cells was inhibited after treating on SBHL for 24 h.The inhibiting effect increased granually following the concentration of SBHL(P

OBJECTIVETo observe the difference in clinical efficacy on focal vitiligo treated with heat-sensitive moxibustion in comparison with medication, and discuss its effect mechanism.

METHODSSixty-eight cases were randomized into a moxibustion group (38 cases) and a medication group (30 cases). Additionally, 20 healthy persons were selected randomly as a normal group. In the moxibustion group, the heat-sensitive moxibustion was applied to Hegu(LI 4), Quchi(LI 11), Yanglingquan(GB 34), Zusanli(ST 36), Xuehai(SP 10) and the others, once a day. In the medication group, triamcinolone acetonide cream was used externally and locally, twice a day. In the two groups, the treatment of 15 days made one session. The efficacy was observed after continuous treatment for 3 sessions. The hemorheology test was done in all of the subjects. The radioimmunoassay was adopted to determine the levels of Interleukin 2 (IL-2), Interleukin 6 (IL-6), Interleukin 10 (IL-10) and tumor necrosis factor (TNF-alpha) before and after treatment.

RESULTSThe levels of IL-6, IL-10 and TNF-alpha in vitiligo patients were higher significantly than those in the normal group (P<0. 01, P<0. 05), the level of IL-2 was lower significantly than that in the normal group (P<0. 01) before treatment. After 3 sessions treatment, IL-2 level was increased significantly in the moxibustion group and the levels of IL-6, IL-10 and TNF-alpha were reduced, without significant differences as compared with the normal group (all P>0. 05). But the differences were significant as compared with those in the medication group (all P<0. 05). The curative and remarkably effective rate was 76. 3% (29/38) after treatment in the moxibustion group, which was higher significantly than 13. 3% (4/30, P<0. 05) in the medication group.

CONCLUSIONHeat-sensitive moxibustion achieves very good clinical efficacy on focal vitiligo, which is probably via promoting blood circulation and regulating the levels of IL-6, IL-10 and TNF-alpha.

Adolescent ; Adult ; Child ; Female ; Hot Temperature ; Humans ; Male ; Middle Aged ; Moxibustion ; Triamcinolone Acetonide ; administration & dosage ; Tumor Necrosis Factor-alpha ; Vitiligo ; drug therapy ; therapy ; Young Adult

OBJECTIVE: To explore the possibility mechanism of non-side population cells (NSP) of Hep-2 be induced into stem-like cancer cells by chemotherapy drug--cisplatin. METHOD: Hep-2 cell lines were sorted by fluorescence-actived cell sorting. The acquired NSP cells in trail group were co-cultured with cisplatin for more than 48 hours,while the control group with normal saline(NS). Then identified the percentage of the side population (SP) cells by flow cytometer. The β-catenin, notch-1 mRNA in trial and control group were detected using quantitative realtime PCR, and the β-catenin, notch-1 protein in two groups were compared by Western blot. RESULT: The percentage of side population cells in two groups were (17.16 ± 0.18)%, (10.05 ± 1.20)%, respectively. There was significant difference between two groups (t = 5.844, P < 0.01). The expression of β-catenin, notch-1 was higher in trail group by qRT-PCR; the protein levels of β- catenin, notch-1 was found to inceased in the trail group by Western blot (t = 5.155, P = 0.031; t = 5.977, P = 0.004). Statistical analysis showed significant difference between two groups (P < 0.05). CONCLUSION NSP cells can be differentiated into stem-like cancer cells after being treating with cisplatin. The supposed mechanism is maybe through wnt/β-catenin, notch signaling transduction pathway abnormalities.
Antineoplastic Agents ; pharmacology ; Cell Differentiation ; Cell Line, Tumor ; Cell Separation ; Cisplatin ; pharmacology ; Flow Cytometry ; Humans ; Laryngeal Neoplasms ; pathology ; Neoplastic Stem Cells ; drug effects ; RNA, Messenger ; Signal Transduction ; beta Catenin

This paper investigate the methods to detect engraftment rate of the four patients with hematological disorders who accepted nonmyeloablative allogeneic peripheral blood stem cell transplantation(NAPBSCT). To find out the best method, their engraftment rates were detected serially at different time after NAPBSCT by means of either FISH, or conventional chromosome analysis combined with R-banding analysis concurrently.The results were carefully compared with one another. All these four sex-mismatched cases were engrafted partially,and two of them changed to full engrafment. The results show no statistically significant difference in 3 groups (conventional method, FISH for hypermetaphase, FISH for interphase nuclei). But the results strongly indicate that FISH is a rapid, precise, objective,and reliable menthod for detection of the engraftment rate,and it is suitable for sex-mismatched NAPBSCT.

Objective To summarize the experimence of nonmyeloablative allogeneic peripheral blood cell transplantation in the treatment of chronic leukemia. Methods Seven patients, including 6 cases of chronic myeloid leukemia (in chronic phase), one of chronic lymphoid leukemia (in third stage), with HLA-identical siblings donor received allogeneic peripheral blood stem cell transplantation after a nonmyeloablative conditioning. Results All of them were engrafted with donor cells (4 with full of donor cells grafted, 3 with mixed chimerism) and recovered hematopoiesis (WBC recovered to more than 0.5 ?10 9/L during postoperative 9 day to 21 day and platelet recovered to more than 30?10 9/L during postoperative 11 day to 28 day). One of them developed a GVHD of degree IV. One of them developed aGVHD of degree I. Conclusion This procedure is much safe, effective and of less complications than the myeloablative condioning regimens and may represent another new approach in the management of patients with chronic leukemia.

BACKGROUND: As a combination of reverse transcription (RT) and polymerase chain reaction (PCR), RT-PCR has been used to detect gene expression levels in cells and tissues, RNA virus contents in cells and specific gene cloned cDNA sequences.OBJECTIVE: To detect the inhibitory effcet of Stealth siRNAs on the expression of transforming growth factor β1 (TGF-β1).METHODS: There were blank control, empty vector transfection, stealth-48, stealth-166, and stealth-594 groups. Three stealth siRNAs aimed at different sequences in TGF-β1 mRNA were made, and were then transfected into BALB/c mouse lung fibroblasts in vitro. The expressions of TGF-β1 and connective tissue growth factor were detected by RT-PCR.RESULTS AND CONCLUSION: In different time periods, the TGF-β1 expression was differentially depressed by three stealth siRNAs, especially stealth-166. The inhibitory effects varied with time, which could be detective at 48 hours,reached the peak at 72 hours and then began to attenuate at 96 hours. Our findings show that the inhibitory effect of stealth siRNAs on the TGF-β1 expression in mouse lung fibroblasts can be detected by RT-PCR.

Objective To study the diagnosis and treatment of delayed radiation-induced brain injury. Methods 21 patients with delayed radiation-induced brain injury were diagnosed by CT, MRI and MRS, and were treated with dehydration, corticosteroid, neuroprective drugs and hyperbaric oxygen or surgery. Results The symptoms of 66.7% patients were obviously improved, 19.0% patients partly recovered and the symptoms of 14.3% patients were not obviously improved. Conclusion The early diagnosis and reasonable treatment could decrease delayed radiation-induced brain injury to minimal extent. The operative treatment should be performed in the patients who had operative indicators. And it was important to prevent the occurrence of delayed radiation-induced brain injury.