We summarized our accumulated clinical and teaching experiences and explored the regularity of acupuncture needling and teaching. It is of great importance in pressing hand during inserting needle. Stroking and pressing are two crucial parts which deserve more attention, and seldom useage of pressing hand should be abolished. Operating hand needs practice before inserting needle, while it should fully relaxed during inserting. Blending "touching", "stretch" "gathering" "erupting" and "advancing" in single moment, applying appropriate dynamic mode of inserting needle such as "join 3 forces as one" "3 points in a line" expertly and naturally. In addition, enough attention should be paid on "altering direction" and "shifting point". Inserting deftly and powerfully, no/slight sensation, deqi when inserting needle are the highest reflection as an acupuncturist.
Acupuncture ; education ; Acupuncture Therapy ; instrumentation ; methods ; China ; Humans ; Teaching

Rutaecarpine (Rut) is a type of indole quinazoline alkaloid exracted from Ruticarpum. Studies showed that Rut has a wide range of pharmacological effects, such as anti-hypertension, anticancer, anti-inflammation, anti-thrombus formation. Currently, many scholars are committed to developing it into a new antihypertensive and anti-inflammatory drug with all new mechanisms. But studies found that Rut is a highly fat-soluble drug with low water and oil solubility. Its high insolubility is the main obstacle in its oral absorption and application, which greatly reduced its bioavailability. Therefore, hydroxypropyl-beta-cyclodextrin (HP-beta-CD) was used as the inclusion material to prepare Rut-HP-beta-CD inclusion complex in this experiment, in order to increase its water solubility and bioavailability. In this experiment, the inclusion complex was prepared by the stirring-freeze-dry method. The preparation process was optimized by the orthogonal test, with the inclusion rate as the index, and molar ratio between host and guest molecules, inclusion temperature, time and stirring speed as the impacting factors. Moreover, the inclusion complex was verified by detecting the apparent solubility, thin layer chromatography, microscopic identification, melting point detection and dissolution study. The results showed that under the conditions of the molar ratio between Rut and HP-beta-CD of 1: 1, temperature at 60 degrees C, inclusion time of 4h and stirring speed at 600 r x min(-1), the inclusion rate of Rut-HP-beta-CD reached 91.04%. Therefore, the preparation process of Rut-HP-beta-CD inclusion under the optimum conditions is simple and feasible, with a highest inclusion rate and reproducibility, and could significantly improve Rut's solubility and bioavailability, and provide a reliable experimental basis for its clinical application.
2-Hydroxypropyl-beta-cyclodextrin ; Alkaloids ; chemistry ; Chemistry, Pharmaceutical ; methods ; Drug Carriers ; chemistry ; Drugs, Chinese Herbal ; chemistry ; Rutaceae ; chemistry ; Solubility ; beta-Cyclodextrins ; chemistry

Objective To investigate the effect of recombinant chlamydiaphage phiCPG1 capsid protein Vp1 on Chlamydia trachomatis(Ct) after Vp1 was co-cultured with Ct (reference strains and clinical strains).Methods The recombinant chlamydiaphage phiCPG1 capsid protein Vp1 was expressed and purified.Equal amount of Ct standard strains (E/UW-5/Cx and D/UW-3/Cx) or clinical strains,which had been incubated with Vp1 protein at the concentration of 53 μg/ml for 3 h at room temperature,were inoculated into McCoy.After cell culture,idione stain and transmission electron microscope were used to observe the effect of Vp1 on the Ct.The effect of Vp1 protein on the cell line McCoy was determined by MTT assay,the responses of Escherichia coli BL21 and DH5α toward Vp1 protein were determined using broth microdilution assays.Results Vp1 had obviously inhibitive effect on Ct,the inhibition ratios were about 40％-70％in clinical strains,72％ in reference strain D and 78％ in E,respectively.Abnormally enlarged RBs were observed after Vp1-treatment and Vp1 could arrest chlamydial developmental cycle using electron microscope.There was no effect of Vp1 on McCoy cells or bacteria BL21 or DH5α.Conclusion The recombinant Vp1 from phiCPG1 has obviously inhibitive effect on the growth of Ct,it will be helpful for the treatment of Ct infection in clinic.

Antimicrobial peptides, a cluster of small peptides secreted by the majority of creatures, have been demonstrated with activity against a wide range of microorganisms including bacteria, protozoa, yeast, fungi, viruses and even tumor cells. These peptides have some features such as broad spectrum , high effi-cacy and stability, little drug resistance. A lack of new antibiotics combined with emerging multi-drug resis-tance issues demands that new antimicrobial strategies be explored for treating these infections. It has been proposed that the antimicrobial peptides might form the foundation for a new class of clinically useful an-timicrobials. We review the advantages of these molecules in construction features and bioactivity, with the focus on the mechanism and clinical applications.

OBJECTIVETo investigate the expression of osteopontin mRNA and its correlation with clinicopathologic features of gastric cancer and elucidate its role in tumor invasion and distant metastasis.

METHODSThe expression of OPN mRNA was detected by semi-quantitive RT-PCR. The relationship between the relative content of OPN mRNA and clinicopathologic features of gastric cancer was analyzed.

RESULTSIn 66 cancer tissue samples, a 330 bp band was detected in 50 cases, the positive rate of OPN mRNA expression was 75.8% (50/66). The expression in all 20 cases of normal gastric mucosa was negative. The expression was associated with the depth of tumor invasion, diameter, lymph node metastasis and but had no correlation with differentiation grades. The 66 patients were followed up for 10 approximately 27 months (mean 16 months). The OPN mRNA expression positive group (50 cases) had recurrence in 15 patients and the negative group (16 cases) had only 1 case with recurrence (P = 0.05); 10 patients died in OPN mRNA expression positive group but no patient died in OPN staining negative group (P = 0.05).

CONCLUSIONOPN mRNA is over-expressed in gastric cancer. It reflects the progression of disease and association with poor prognosis of gastric cancer. OPN may play an important role in the process of distant metastasis in gastric cancer.

Adenocarcinoma ; metabolism ; pathology ; Adult ; Aged ; Aged, 80 and over ; Female ; Humans ; Lymphatic Metastasis ; Male ; Middle Aged ; Osteopontin ; biosynthesis ; genetics ; Prognosis ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Stomach Neoplasms ; metabolism ; pathology

BACKGROUNDTo investigate the biological functions of a novel hepatitis B virus e antigen (HbeAg) interacting protein AK026018, and to use cDNA microarray technique to screen genes regulated by the protein.

METHODSThe AK026018 coding DNA fragment was amplified by reverse transcription polymerase chain reaction (RT-PCR) technique from HepG2 cell. The expressive vector of pcDNA3.1-AK was constructed by routine molecular biological methods. The HepG2 cells were transfected with pcDNA3.1 and pcDNA3.1-AK, respectively by using lipofectamine. The total RNA was isolated and reverse transcribed. The cDNA of each sample was subjected to microarray screening with 8,464 cDNA probes and analyzed by bioinformatics.

RESULTSThe expressive vector was constructed and confirmed by DNA sequencing analysis and restriction enzyme digestion. High quality mRNA and cDNA of transfected HepG2 cells had been prepared and successful microarray screening conducted. From the scanning results, there were 122 differential expression genes, of which 36 genes were down-regulated, and 16 genes were up-regulated.

CONCLUSIONMicroarray technique was successfully used to screen the genes trans-regulated by AK026018. The expression of AK026018 protein affects the expression spectrum of HepG2 cells.

Carrier Proteins ; genetics ; metabolism ; physiology ; Cell Line, Tumor ; Computational Biology ; Gene Expression Regulation, Neoplastic ; Hepatitis B e Antigens ; metabolism ; Humans ; Oligonucleotide Array Sequence Analysis ; methods ; Protein Binding ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection

The ZM-1 tissue microarrayer designed by our groups is manufactured in stainless steel and brass and contains many features that make TMA (tissue microarray) paraffin blocks construction faster and more convenient. By means of ZM-1 tissue microarrayer, biopsy needles are used to punch the donor tissue specimens respectively. All the needles with the punched specimen cylinders are arrayed into the array-board, with an array of small holes dug to fit the needles. All the specimen cylinders arraying and the TMA paraffin block shaping are finished in only one step so that the specimen cylinders and the paraffin of the TMA block can very easily be incorporated and the recipient paraffin blocks need not be made in advance, and the paraffin used is the same as that for conventional pathology purpose. ZM-1 tissue microarrayer is easy to be manufactured, does not need any precision location system, and so is much cheaper than the currently used instrument. Our method's relatively cheap and simple ZM-1 tissue microarrayer technique of constructing TMA paraffin block may facilitate popularization of the TMA technology.
Biopsy, Needle ; instrumentation ; Equipment Design ; Female ; Humans ; Immunohistochemistry ; Male ; Neoplasms ; enzymology ; Paraffin ; Tissue Array Analysis ; instrumentation

OBJECTIVETo assess protein and mRNA expression levels of heparanase and basic fibroblast growth factor (bFGF) genes in human non-small cell lung cancer (NSCLC) and their roles in tumor invasion, metastasis and prognosis.

METHODSA total of 115 paraffin-embedded and 45 fresh-frozen tissue specimens of NSCLC were studied by immunohistochemistry, Western Blot and in situ hybridization to evaluate the protein and mRNA expression status of heparanase and bFGF genes. The data was analyzed by SPSS statistical software.

RESULTSBoth human heparanase and bFGF were highly expressed in NSCLC cells, in contrast to none or a low expression in normal lung tissue. Expression of heparanase also showed a significantly higher than that in the normal tissue by Western blot (P = 0.041). Immunohistochemistry showed that heparanase expression was both cytoplasmic and membranous. The agreement between heparanase and bFGF was significant. A significant correlation was found between the expression of either protein and TNM stage, vascular invasion, lymphatic metastasis and microvascular density (MVD). Co-expression of the two proteins demonstrated an even higher correlation with the tumor stage and MVD. In addition, expression of bFGF correlated with tumor cell differentiation. Data of a multivariate analysis indicated that tumor cell differentiation, vascular invasion, lymphatic metastasis and expression of bFGF were identified as significant prognostic parameters.

CONCLUSIONSBoth heparanase and bFGF may play important roles in tumor angiogenesis, metastasis, and prognosis of NSCLC.

Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; enzymology ; metabolism ; pathology ; Cell Differentiation ; Female ; Fibroblast Growth Factor 2 ; metabolism ; Glucuronidase ; metabolism ; Humans ; Lung Neoplasms ; enzymology ; metabolism ; pathology ; Lymphatic Metastasis ; Male ; Microcirculation ; pathology ; Middle Aged ; Neoplasm Staging ; Neovascularization, Pathologic ; Prognosis ; Survival Rate

OBJECTIVETo screen and clone the genes in hepatocytes which encode protein that can interact with hepatitis B e antigen(HBeAg) by yeast-two hybridization.

METHODSRecombined HBeAg bait plasmid (pGBKT7-eAg) was transformed into yeast AH l09, followed by mating with yeast Yl87 containing liver cDNA library plasmid in 2 x YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-Ade-His) which contains X-a-gal for selecting positive blue clones. Then positive clones were selected and plasmids were prepared and sequenced. Finally, bioinformatics analysis was performed.

RESULTSTotally 245 positive colonies were selected and 101 colonies were sequenced. Through sequences alignment, 6 novel genes and 35 recorded genes were screened.

CONCLUSIONGenes of HBeAg interacting proteins have been cloned successfully, which brings some new clues for further studies on the biological functions of HBeAg and the related proteins.

Gene Library ; Hepatitis B e Antigens ; genetics ; metabolism ; Humans ; Liver ; metabolism ; Plasmids ; genetics ; Protein Binding ; Transformation, Genetic ; Two-Hybrid System Techniques

OBJECTIVETo study the effect of RNA interference (RNAi) targeting E6AP on the proliferation and apoptosis of HeLa cells.

METHODSHeLa cells were cultured and divided into 3 groups: blank control group, cells transfected with nonsense siRNA (small interference RNA), and cells transfected with specific E6AP siRNA. The expressions of E6AP mRNA and protein were detected by RT-PCR and Western blot before and after the transfection respectively. Cell proliferation was determined by methylthiazolyl tetrazolium (MTT). The cell apoptosis index was assessed by flow cytometry.

RESULTSUpon treatment with E6AP siRNA for 24, 48 and 72 h, the expression level of E6AP mRNA decreased 33%, 72% and 70% than siRNA treated group. The protein expression levels in 48 h and 72 h E6AP siRNA groups decreased 38%, 59% comparing with those of the nonsense siRNA treated group (P < 0.05). The proliferative capacity of cells transfectd with E6AP siRNA was significantly lower than blank control group (F = 101.38, P < 0.05) and siRNA treated group (F = 38.64, P < 0.05). The apoptosis index of HeLa cells treated with E6AP siRNA was significantly higher than that of the nonsense siRNA (F = 41.48, P < 0.05) and the blank control group (F = 86.36, P < 0.05).

CONCLUSIONSiRNA targeting can effectively suppress the expression levels of E6AP mRNA, corresponding with a proliferation inhibition and an enhanced apoptosis of HeLa cells.

Cell Line, Tumor ; Cell Proliferation ; Female ; Gene Expression Regulation, Neoplastic ; genetics ; Gene Silencing ; HeLa Cells ; Humans ; RNA Interference ; RNA, Small Interfering ; genetics ; pharmacology ; Reverse Transcriptase Polymerase Chain Reaction ; Ubiquitin-Protein Ligases ; antagonists & inhibitors ; metabolism ; Uterine Cervical Neoplasms ; genetics ; metabolism