Objective: To investigate whether the expression of Apollon (an apoptosis inhibitory gene) silenced by using RNA interference (RNAi) technology in combination with tetramethylpyrazine (TMP) intervention can increase the sensitivity of human myeloid leukemia K562 cells to vincristine (VCR) and daunorubicin (DNR). Methods: Using the best small interference RNA (siRNA) targeting Apollon gene which was screened out in a previous experiment, the eukaryotic expression vector pGPHI-GFP-Neo-Apollon was constructed. Then, the K562 cells were transfected with pGPHI-GFP-Neo-Apollon by LipofectamineTM 2000 and screened out by G418. After Apollon siRNA was transfected stably into K562 cells, the expressions of Apollon mRNA and protein were detected by reverse transcription (RT)-PCR and immunofluorescence, respectively. The sensitivities to chemotherapeutic agents VCR and DNR after RNAi in combination with TMP intervention were detected by MTT method, and apoptosis rate was detected by flow cytometry. Results: pGPHI-GFP-Neo-Apollon vector was constructed successfully and it could express stably in K562 cells, which silenced the expressions of Apollon mRNA and protein effectively. The sensitivity to VCR or DNR increased significantly after RNAi in combination with TMP intervention. The half inhibitory concentration (IC50) values of VCR and DNR were significantly lower than those in the group of TMP intervention alone (P < 0.05). The apoptosis rate of K562 cells after transfection with Apollon siRNA in combination with TMP intervention was significantly increased comparing with that of the untreated control group and TMP intervention alone group (P < 0.05). Conclusion: RNAi targeting Apollon gene in combination with TMP intervention can obviously increase the sensitivity to VCR or DNR in K562 cells and significantly reduce the dosage of chemotherapeutic agents as well as improve apoptosis induced by chemotherapy. These results suggest that Apollon gene silenced by RNAi in combination with TMP intervention may reverse the drug resistance of leukemia cells. Copyright © 2013 by TUMOR.

Objective To analyse the clinical and pathological characteristics of patients with myopathies that were misdiag-nosed as myasthenia gravis because of external ophthalmoplegia ,widen the thoughts for differential diagnosis of extraocular muscle paralysis .Methods The clinical and pathological features of 11 myopathy cases with ptosis and diplopia admitted to the neurology department of the First Affiliated Hospital of Chongqing Medical University between October 2010 and May 2014 were retrospec-tively reviewed and analyzed .Results Among the 11 patients ,6 male and 5 female ,aged 16-66 years old .All had paralysis of ex-traocular muscle manifestations ,including oculopharyngeal muscular dystrophy(OPMD) in 3 cases ,oculopharyngeal distal myopa-thy in 2 cases ,chronic progressive external ophthalmoplegia(CPEO) in 6 cases .Muscle biopsy showed the characteristic pathologi-cal changes .Statistical analysis showed that the age of onset in OPMD patients was older(P<0 .05) ,creatine kinase level in oculo-pharyngeal distal myopathy patients was higher ,but the difference was not significant (P>0 .05) .Conclusion Certain rare myopa-thy should be considered for patients with external ophthalmoplegia seemingly myasthenia gravis .Muscle biopsy can provide clue for differential diagnosis .

Objective To observe the clinical efficacy and safety of nicergoline and aniracetam in treatment of mild and moderate cognitive dysfunction after cerebral infarction .Methods Infarction were enrolled after 70 cases of mild and moderate cognitive im-pairment patients were randomly divided into nicergoline treatment group (observation group) and aniracetam treatment group(con-trol group) ,all 35 cases .After 12 weeks of treatment were compared before and after treatment mini mental state examination (MMSE) ,activities of daily living scale(ADL) scores and transcranial Doppler(TCD) to improve the situation .Results After 12 weeks ,both groups after treatment compared with treatment MMSE score was significantly higher (P< 0 .05) ,and nicergoline group than aniracetam group(P< 0 .01);ADL score before treatment were significantly reduced (P< 0 .05) ,TCD improvement nicergoline group than aniracetam group (P<0 .01) .Conclusion Nicergoline mild and moderate cognitive dysfunction after cerebral in-farction have significant effects ,and better than aniracetam ,no significant adverse reactions occur for a wide range of clinical applications .

Objective There are two main functions of gastrointestinal tract,digestion and absorption,and barrier function.The latter has an important defensive effect,which keeps the body away from the invading and damaging of bacteria and endotoxin.It maintains the systemic homeostasis.Intestinal dysfunction would happen when body suffers from diseases or harmful stimulations.The more serious intestinal disorders would harm the intestinal protective mechanism,or intestinal barrier function,and bacterial/endotoxin translocation,of intestinal failure(IF) would ensue.This article provides a critical review of the evidence indicating that an increase in bacterial translocation is associated with sepsis,and even the multiple organ failure syndrome in critically ill patients.The in-transit microorganisms play an essential role in the homeostasis of local and systemic immunity.Methods All studies published from 2000 to June 2005 about intestinal permeability,bacterial translocation,and systemic inflammatory response syndrome were located by search of PubMed.Results Clinical and experimental studies investigating the correlation between bacterial translocation and systemic inflammatory response syndrome,associated with the damage of the gut barrier function.To keep the mucosal barrier function intact is one of the main issues in the prevention of bacterial translocation.This could be achieved by the adequate delivery of oxygen and nutrient supplementation to the gut.Enteral nutrition,probiotic can be a good choice.Conclusion With a better understanding of the bacteria-host interactions in health and the alterations induced by critical illness,new therapies that improve the environment of both may lead to better recovery rates in intensive care unit patients.

Objective Pathogen and antimicrobial susceptibility in children with bacterial meningitis were reviewed.Methods The positive cultures of cerebrospinal fluid samples or blood samples and its antimicrobial susceptibility were analyzed in 401 patients with the clinical diagnosis of bacterial meningitis.Results 401 cases cerebrospinal fluid samples and blood samples submitted to microbiology laboratory, 97 cases (24%) were microscopically and culturally proven to be bacterial meningitis. The most frequent pathogen was staphylococcus aureus (28%), followed by the streptococcus pneumoniae (19%) and escherichia coli (13%). Pediococci as conditioned pathogen, were found in purulent meningitis patients. One of the isolates of Staphylococcus aureus was simultaneously resistant to both vancomycin and teicoplanin. Three isolates showed simultaneous resistance to imipenem/cilastatin.Conclusions Staphylococcus aureus as the predominant pathogens of pediatric ranks first among pediatric patients of purulent meningitis. Serious drug resistance of isolated pathogenic bacteria and its in antimicrobial susceptibility in the bacterial meningitis should be considered in clinical therapy.

Objective To explore the mechanism of therapeutic action of needle knife “regulating sinews and treating bones” on knee osteoarthritis (KOA) by observing the effect of needle knife intervention on KOA rabbit behaviors, mechanical characteristics of patellar ligament (PL), and expressions of interleukin-4 (IL-4), matrix metalloproteinase-3 (MMP-3) and aggrecan in knee cartilages.Methods Forty New Zealand rabbits were randomly allocated to blank, model, needle knife and electroacupuncture groups, 10 rabbits each. A rabbit model of KOA was made by the modified Videman method of immobilization in extension position. After model making, the needle knife and electroacupuncture groups received needle knife and electroacupuncture treatments, respectively. A behavioral assessment was made using the modified Lequesne MG knee grade evaluation method in every group after model making and treatment. The samples were taken after treatment. PL tension, stress relaxation and creep state were tested using a Bose Electro Force 3300 protracted test machine. Cartilage cell IL-4 expression was examined by ELISA. MMP-3 mRNA and aggrecan mRNA expressions were detected by real-time PCR.Results After model making, there was a statistically significant difference in the Lequesne MG score between the model and blank groups (P<0.01); there was no statistically significant difference in the Lequesne MG score between the needle knife or electroacupuncture group and the model group (P>0.05). There was a statistically significant post-treatment difference in the Lequesne MG score between the needle knife or electroacupuncture group and the model group (P<0.01,P<0.05) and between the needle knife and electroacupuncture groups (P<0.05). There were statistically significant post-treatment differences in PL maximum stress, maximum displacement, elastic modulus, stress relaxation rate and creep rate between the model and blank groups (P<0.01,P<0.05). There were statistically significant post-treatment post-treatment differences in PL maximum stress, maximum displacement, elastic modulus, stress relaxation rate and creep rate between the needle knife and model groups (P<0.01,P<0.05). There was a statistically significant post-treatment difference in elastic modulus between the electroacupuncture and model groups (P<0.01). The IL-4 content and aggrecan mRNA expression decreased significantly and MMP-3 mRNA expression increased significantly in the model group after model making and there were statistically significant differences compared with the blank group (P<0.01,P<0.05). After treatment, the IL-4 content increased significantly in the needle knife and electroacupuncture groups compared with the model group (P<0.01,P<0.05) and aggrecan mRNA expression tended to increase in the two groups. The regulation of aggrecan mRNA and MMP-3 mRNA expressions was better in the needle knife group than in the electroacupuncture groups, but there was no statistically significant difference compared with the model group (P>0.05).Conclusion The mechanism of action of needle knife treatment on KOA may be that it improves ligament mechanical characteristics, regulates intra-articular stress environment, and modulates aggrecan mRNA and MMP-3 mRNA expressions and inhibits cartilage degeneration through IL-4 mechanical signal pathway, to produce the therapeutic effect of “regulating sinews and treating bones”.

DNA, Viral ; genetics ; Hep G2 Cells ; Hepatitis B virus ; drug effects ; Humans ; Oligonucleotides, Antisense ; pharmacology

To compare the activity of RGD-TRAIL in different expression systems, RGD-TRAIL in both Escherichia coli (E.coli) and Pichia pastoris was constructed and expressed. In vitro activity of RGD-TRAIL from Pichia pastoris expression system was also analyzed. Genetic engineering techniques were used to construct recombinant plasmid pET30-rgd-trail and pHBM-rgd-trail. The recombinant protein RGD-TRAIL was purified with Ni ion affinity chromatography after induction. MTT assay, ELISA, scratch wound healing, transwell migration assay and Hoechst 33342 staining were performed to detect the effects of RGD-TRAIL on proliferation, binding activity, migration and apoptosis. The expression of apoptosis-associated proteins was detected by Western blotting. Recombinant protein RGD-TRAIL was successfully expressed in a form of inclusion body in E.coli, while expressed secretorily in Pichia pastoris. It possessed more potent cytotoxicity than RGD-TRAIL in E.coli by MTT assay. The RGD-TRAIL expressed by Pichia pastoris showed powerful binding affinity with cancer cells expressing α(v), DR4, DR5 and highly potent cytotoxicity through inducing apoptosis of cancer cells. Nuclear fragmentation was examined by Hoechst 33342 staining. Cleaved PARP and caspase-3 were also detected after incubation with RGD-TRAIL. Additionally, RGD-TRAIL inhibited migration significantly in A549 and HT1080 cells. The results demonstrate that Pichia pastoris expression system is more suitable for the recombinant protein RGD-TRAIL. Its binding affinity and antitumor activity might make RGD-TRAIL a promising candidate for cancer therapy.
Antineoplastic Agents ; pharmacology ; Apoptosis ; Blotting, Western ; Cell Line, Tumor ; Chromatography, Affinity ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; Humans ; Oligopeptides ; biosynthesis ; pharmacology ; Pichia ; metabolism ; Plasmids ; Recombinant Fusion Proteins ; biosynthesis ; pharmacology ; TNF-Related Apoptosis-Inducing Ligand ; biosynthesis ; pharmacology

Electric Stimulation ; Humans ; Hypoglossal Nerve ; Sleep Apnea, Obstructive ; therapy

Objective To study the radiosensitization of histone deaeetylases inhibitor(HDACI) panobinistat in prostate cancer cells in vitro,as well as the possible mechanisms.Methods IC20 of two prostate cancer cell lines(LNCaP and PC-3)was determined using MTI assay.Cells received a single dose irradiation of 0,2,4,6,or 8 Gy using 6 MV X-ray for radiosensitivity experiment,but only 2 Gy for western blot and flow cytometry.Radiosensitization of panobinostat was investigated with clonogenic assay,and sensitizing enhancement ratio(SER)was calculated with single-hit multi-target model.Western blot was used to compare γH2AX expression.Flowcytomctry was used to detect the cell cycle distribution.Results IC20 of LNCaP and PC-3 was 2.5 and 10.0 μmol/L,respectively.SER of panobinostat at IC20 was 1.37(D0 ratio)and 1.11(Dq ratio)for LNCaP cells,and 1.78(D0 ratio)and 1.17(Dq ratio)for PC-3 cells.Expression of γH2AX gradually decreased in the 2 Gy irradiation-alone cells standing for the DSB repair,while γH2AX expression was persistent in the combination group.Irradiation triggered a G2/M arrest 6-12 hours after irradiation in LNCaP and PC-3 cells.G2/M arrest was observed when cells were treated with panobinostat for 24 hours,however,no significant change concerning cell cycle distribution was showed when cells received further irradiation.Conclusions Panobinostat Call radiosensitize prostate cancer cells,which may be related with increased DNA DSB,inhibition of DSB repair and attenuation of cell cycle modulation after irradiation.