Objective To prepare a targeted antitumor drug delivery system using large-inner-diameter multi-walled carbon nanotubes (LID-MWCNTs) for sustained release and to study its performance. Methods LID-MWCNTs were puri?fied and oxidized,then use nanocarriers and USTs as homologous blockers. Folic acid and fluorescent labels were conjugat?ed onto the external surfaces of nanocarriers. CDDP (cisplatin) was encapsulated and ultrashort tubes (USTs) were employed to block the drug entry/exit paths. The microstructure of resulted drug delivery system (DDS) was observed, while drug load?ing efficiency and drug release profile in vitro were determined. The tumor-targeting property and cytotoxicity of DDS were also assessed. Results LID-MWCNT based sustained release targeted drug delivery system was established. Drug loading efficiency of CDDP@UST-FA-LID-MWCNTs was as high as 70.97%. A typical biphasic sustained release pattern was dem?onstrated, and the accumulating release time was 18 h. DDS exhibited a certain kind of tumor-targeting property, and inhibit?ed proliferation of tumor cells in a dose-dependent manner. Conclusion CDDP@UST-FA-LID-MWCNT drug delivery system exhibited an improved drug loading efficiency and a sustained drug release profile. It could specifically target the tu?mor cells and had a significant antitumor effect.

Objective To observe whether hepatitis B vaccine enhance the treating effect of cyto-kine induced kill(CIK) cells on hepatitis B virus transgenic(HBV-Tg) mice. Methods The HBV-Tg mice were treated with CIK cells by peritoneal injection and hepatitis B vaccine by hypodermic injection. The HBV DNA level were tested by real-time PCR,T lymphocyte subgroup were detected by flow cytometry and the pathological diversify of hepatic tissue were observed by HE staining. Results The HBV DNA loading in peripheral blood of HBV-Tg mice decreased after CIK cells were treated and CD3~+ , CD4~+ and CD8~+ cells increased which were enhanced after CIK cells combined with hepatitis B vaccine. Conclusion Hepa-titis B vaccine enhanced the treating effect of CIK on HBV-Tg mice which may be implemented by increased the blood level of CD3~+, CD4~+ and CD8~+ cells, especially CD8~+ cells level.

BACKGROUND:A mass of debris particles that can lead to cytotoxicity exist in commercial large-inner-diameter multi-walled carbon nanotubes (LID-MWCNTs).Because of the high hydrophobicity on the surface of the tube wall,the carbon tubes can be twisted and agglomerated,resulting in the low dispersion and poor biocompatibility.Therefore,to explore the effective methods of purifying and modifying LID-MWCNTs is the primary problem to develop its potency in the biomedical application.OBJECTIVE:To explore the effects of different mixed acid reflux time on LID-MWCNTs purification and biocompatibility.METHODS:After pretreatment with high temperature calcinations and hydrochloric acid pickling,LID-MWCNTs were purified under different time of mixed acid reflux time (1,2,4 hours).The mixed acid reflux time for best purification was chosen based on surface morphology and dispersibility,so as to optimize preparation technology and observe the characterization.The L929 cells and CAL-27 cells were treated with different concentrations of raw LID-MWCNTs (5,10,20,40,80 mg/L) and purified LID-MWCNTs by mixed acid reflux (5,10,20,40,80 mg/L).After 72 hours,cell counting kit-8 assay was employed to test the proliferation of L929 cells and CAL-27 cells.RESULTS AND CONCLUSION:(1) With the time of mixed acid reflux,the length of LID-MWCNTs was decreased,and the dispersion was improved.However,the external surface of the tubes after mixed acid reflux 2 and 4 hours were destroyed obviously.Especially after mixed acid reflux 4 hours,the tubes were destroyed seriously and the diameter of tubes was not uniform.However,after mixed acid reflux 1 hour,the fundamental structure and morphology of the tubes were not changed,the debris particles were undetected on the tube wall surface,and the tubes had the good dispersion.(2) Under the same concentration,the survival rate of L929 cells in the raw LID-MWCNTs group was lower than that in the purified LID-MWCNTs group.At the concentration of 10-80 mg/L,the survival rate of L929 cells in the group of mixed acids reflux 1 hour was up to 90％,higher than that in the other groups (P ＜ 0.05).(3) Under the same concentration,the survival rate of CAL-27 cells in the raw LID-MWCNTs group was lower than that in the purified LID-MWCNTs.At the concentration of 20-80 mg/L,the survival rate of CAL-27 cells in the group of mixed acids reflux 1 hour was up to 90％,higher than that in the other groups (P ＜ 0.05).These results revealed that the raw LID-MWCNTs were purified effectively after mixed acid reflux 1 hour,and the cytotoxicity was decreased.

Objective To investigate the biological effects of Wnt gene in kidney cancer Caki?2 cells. Methods The Wnt gene was silenced in kidney cancer Caki?2 cells by lentivirus vector. The cell proliferate ability of cells in each group were assayed by CCK?8 kit at different time points. The apoptosis of Caki?2 cells was observed after silencing Wnt gene by transmission electron microscope. The invasion ability of each group cells was tested using Transwell chambers. The genes expression changes of Wnt/β?catenin signaling pathway and apoptosis related gene were determined by realtime PCR. Results Compared with the other two groups,the cell proliferate ability of the cells after silencing Wntgene was suppressed,and the difference was statistically significant(P<0.05). Apoptosis increased significantly in shRNA+Caki?2+Wnt group cells with silencing of Wntgene, and apoptotic body appeared in these cells. In invasive experiment,the number of emigrated cells in shRNA+Caki?2+Wnt group were significantly lower than other groups(P<0.05). The expression of Wnt mRNA,β?catenin mRNA and Bcl?2 mRNA in shRNA+Caki?2+Wnt group cells was lower than other groups(P<0.05). Conclusion Silencing of Wnt gene of kidney cancer Caki?2 cells can affect the proliferation rate of the cells, promote the cell apoptosis,and inhibit the invasion ability,which provide certain theoretical basis for the research and development of new drugs and new therapeutic targets.

Aim To further study the molecular mecha-nism of the herbs with hot nature on the regulational action on TRPV1 channel based on the 7900 Real-time PCR instrument. Methods 7900 PCR instrument was applied to detect the intracellular flurescence of TRPV1 channel in the dorsal root ganglions ( DRG ) neurons and the effect on the TRPV1 ’ s thermo-sensational functions of the selected 11 ingredients from hot herbs was explored. Results TRPV1 channels could be ac-tivated by gradually elevated temperature; the activa-tion process could be blocked by the TRPV1 specific blocking agent capsaizepine. Most of the ingredients from hot-nature herbs had the potential to up-regualate TRPV1 channel function. Conclusions The estab-lished TRPV1 channel detection system based on PCR instrument is suitable for the analysis of regulational functions of drugs on the heat-activated TRPV1 chan-nel;the functions of hot herbs may be related to the up-regualtional effects of its active ingredients on the TRPV1 channel which will further up-regulate energy metabolism.

Objective To present the experience of laparoscopic extraperitoneal radical prostatectomy and evaluate its safety and efficacy. Methods A total of 91 patients diagnosed with localized prostate carcinoma were admitted from February 2003 to June 2008. The level of serum PSA ranged from 7. 5 - 47. 0 ng/ml(mean 14. 0 ng/ml). The volume of the prostate ranged from 35 - 75 ml(mean 52 ml). Biopsy was performed before the operation and the pathological results revealed prostate carcinoma with Gleason score no more than 8. CT, MR and ECT revealed there was no lymph node or seminal vesicle involvement and there was no bone metastasis. The procedures were performed with an-tegrade techniques and pelvic lymphadenectomies were performed in 32 cases and nerve-sparings were performed in 11 cases. Results The operation duration ranged from 105 - 270 min (mean 173 min). Intraoperative blood loss was 110 - 1200 ml(mean 315 ml). Incontinence occurred in 19 cases in early stage and 18 cases recovered within 3 months. Positive surgical margin occurred in 11 cases. There was no complication of urethra stricture during 3 - 30 months' follow-up. No lymph node was involved in 32 cases with pelvic lymphadectomy. Five of the 11 cases received nerve-sparing prostatectomy had normal erectile function during the follow-up. Conclusions Laparoscopic extraperitoneal radical prostatectomy is a safe, effective and efficient surgical procedure with the minimal invasion, less morbidity and rapid recovery. Laparoscopic radical prostatectomy is emerging as an alternative to open radical prostatectomy.

The effect of water extracts of Euphorbia hirta on the histological features and expressions of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) in the rat articular cartilage was investigated. Arthritis was induced in rats using Freund's Complete Adjuvant containing heat-killed M. tuberculosis, and treated with water extracts of E. hirta. Paraffin tissue sections of the arthritic joints were evaluated. The extent of cartilage degeneration was found to be greatest in rats treated with the highest dosage of E. hirta, followed by rats in the untreated group. Rats treated with the intermediary and low dosages of Euphorbia hirta showed improved histology. MMP-13 levels were found to be decreased with decreasing dosages of E. hirta. TIMP-1 levels were found to increase with decreasing dosages of E. hirta. MMP-3 levels fluctuated without any appreciable pattern. Low dosages of E. hirta seem to be beneficial in reducing cartilage degeneration in cases of arthritis.
Upper case ee ; Rattus norvegicus ; Euphorbia ; Water ; Degeneration, NOS

Day-old chick is unique animal model in brain development and behavior study. The intermediate medial mesopallium (IMM), a region of the chick forebrain, is intimately involved in the early learning processes, which offers the ideal opportunity to study the neural changes that underlie behavioral plasticity. In this paper, the intracellular recordings were conducted from IMM neurons in chick forebrain slices, in which electrophysiological properties, synaptic responses and long-term potentiation (LTP) were observed. Coronal sections of left forebrains (500 mum thick), containing IMM, were prepared from domestic chicks, aged 2-10 days. In 69 IMM neurons, the resting membrane potential was measured to be (-59.4+/-5.3) mV, slope membrane resistance (70.8+/-27.2) MΩ, and time constant (10.2+/-4.3) ms. The amplitude, threshold, overshoot, half-width, max rise slope and max decay slope of action potential evoked by intracellular current injection were (85.2+/-9.4) mV, (-38.7+/-7.6) mV, (25.6+/-8.9) mV, (2.1+/-0.5) ms, (150.5+/-41.2) mV/ms and (-64.3+/-14.0) mV/ms, respectively. Spike-firing frequency was increased with depolarizing current intensity in 32 of 69 tested cells [linear regression slope was (21.5+/-10.9) Hz/nA, P<0.05 in all cells]. The depolarizing synaptic responses (i.e. EPSPs), with stimulus intensity- and membrane potential-dependent properties, were elicited by dorsal (n=25) or ventral (n=62) focal electrical stimuli at 0.1 Hz in all tested IMM neurons and could be nullified reversibly by perfusion with 100 mumol/L AP5 (NMDA receptor antagonist) and 3 mumol/L DNQX (non-NMDA receptor antagonist), but enlarged by 6 mumol/L bicuculline (GABA(A) receptor antagonist). The EPSPs evoked by ventral stimulation were persistently increased after tetanic stimulation (5 Hz, 300 pulses/train, 2 trains, train interval 10 min) in 6 of 12 tested IMM neurons. The amplitude of EPSPs was potentiated to more than 120% of control level (when analyzed at 45 min of enhancement, P<0.05, n=5), which lasted at least 30 min and then could be referred to as LTP. Moreover, area under curve, duration and max rise slope of EPSPs were also enhanced (P<0.05), while no significant changes were observed in the electrophysiological parameters of IMM neurons following induction of LTP (P>0.05). These results suggest that the intracellular recording techniques in the chick brain slices can be used to perform multi-parameter analysis of synaptic responses and their LTP.
Animals ; Brain ; physiology ; Chickens ; In Vitro Techniques ; Long-Term Potentiation ; Membrane Potentials ; Receptors, N-Methyl-D-Aspartate ; antagonists & inhibitors

Objective: To investigate the relation between ser um hyaluronic acid (HA) concentration and cryopreservation-reperfusion injury. Methods: The animals were randomly assigned to 3 groups: (1 ) group A: the control; (2) group B: liver allografts were stored in lactated R inger's solution (0℃) for 2 h before implantation; (3) group C:liver allografts were stored in lactated Ringer's solution (0℃) for 4 h before implantation. Th e serum sample and liver specimen were taken up at 2 h and 4 h after transplanta tion to detect the concentration of HA, AST and LDH, and to get pathologic obser vation. Results: Serum HA increased earlier and decreased more s hortly than AST and LDH after transplantation in group A. Serum HA increased sig nificantly in group B and C, much higher than that in group A(P<0.01). The i njury of vascular endothelium and the disorder of hepatic sinuses and hepatic lo b ules were observed in group B and C. In the specimen of 4 h in group C, evident infiltration of inflammatory cell was present. Conclusion: Cryopreservation leads to injury of endothelial cell and reperfusion aggravat es this injury. The serum HA concentration indicates the degree of cold ischemia -reperfusion injury.

OBJECTIVETo investigate the effects of protein tyrosine kinase (PTK), protein tyrosine phosphatase (PTP) and protein kinase C (PKC) on apoptosis and observe the changes of cytosolic calcium ([Ca(2+)]i) of arsenic trioxide (As2O3) treated human leukemia cells NB4 and cortex neurons.

METHODS[Ca(2+)]i of NB4 cells and cortex neurons was probed with Fluo-3/AM, its changes were assayed with laser confocal microscopy in real-time after As2O3 treatment at different concentrations, the effects of PTK and PTP and the activation of PKC on these changes with confocal microscopy and phosphorus radioisotope assay. DNA ladders of NB4 cells and cortex neurons after exposed to As2O3 were observed.

RESULTSAs2O3 at 1 micromol/L could remarkably increase the [Ca(2+)]i of NB4 cells but had no effects on neurons. Vanadate, a kind of PTP inhibitor, could promote the increase of [Ca(2+)]i treated by 2, 5, 10 micromol/L As2O3 in a dose-dependent manner. The mean total increase rates at 240 seconds after exposed to As2O3 at different concentrations were (6.5 +/- 2.3)%, (21.7 +/- 2.1)%, (49.2 +/- 2.5)% for NB4 cells, and (6.7 +/- 2.1)%, (19.4 +/- 2.5)%, (52.3 +/- 2.7)% for cortex neurons, respectively. Genistein, a kind of PTK inhibitor, could decrease the increase of [Ca(2+)]i treated by 2, 5, 10 micromol/L As2O3 in a dose-dependent manner. The mean total inhibited rates at 240 seconds after As2O3 treatment at different concentrations were (6.7 +/- 2.9)%, (25.6 +/- 2.5)%, (52.2 +/- 3.5)% for NB4 cells, and (7.8 +/- 3.1)%, (18.1 +/- 2.8)%, (51.3 +/- 3.3)% for cortex neurons, respectively. The activation of PKC began to increase as exposed to As2O3 at 1 micromol/L for 3 h, and kept rising continuously in NB4 cells and at 24 h DNA ladders emerged. However, none of the above results was found in human cortex neurons, but when exposed to 2 micromol/L As2O3, the activation of PKC and DNA ladders did emerge in neurons.

CONCLUSIONSThe phosphorylation and dephosphorylation of PTK and PTP participated in nonspecific apoptosis signal transduction pathway related to As2O3, and accompanied with PKC activation. The [Ca(2+)]i elevation was closely related to increased PKC activation. There existed difference in dose tolerances to As2O3 between NB4 cell and cortex neurons.

Adult ; Apoptosis ; drug effects ; Arsenicals ; pharmacology ; Calcium ; metabolism ; Cell Line, Tumor ; Cells, Cultured ; Cerebral Cortex ; cytology ; Cytoplasm ; metabolism ; Dose-Response Relationship, Drug ; Humans ; Leukemia, Promyelocytic, Acute ; enzymology ; metabolism ; pathology ; Male ; Neurons ; cytology ; drug effects ; metabolism ; Oxides ; pharmacology ; Protein Kinase C ; metabolism ; Protein Tyrosine Phosphatases ; metabolism ; Protein-Tyrosine Kinases ; metabolism