Objective:To design and construct the replication-deficient recombinant adenovirus Ad-siCTGF which can silence the expression of connective tissue growth factor (CTGF) by RNA interference and verified its function. Methods:A specific sequence, which was verified to be able to silence CTGF gene with high efficiency, was cloned into pSES-HUS vector to produce the shuttle plasmid pSES-siCTGF. The plasmid after Pme Ⅰ linearization was cotransduced with pAdEasy into BJ5183 E.coli strains to construct recombinant vector Ad-siCTGF. After linearization treatment with Pac Ⅰ enzyme digestion Ad-siCTGF was transfected into HEK293 cells via liposome mediation. The recombinant adenovirus was packaged. The titer of the Ad-siCTGF was increased after three times of cross-infection. 4T1 cells were infected with the adenovirus. The silencing efficiency was tested by real-time fluorescence quantitative (RFQ)-PCR and Western blotting.Results:Pac Ⅰ enzyme digestion electrophoresis indentified that recombinant adenovirus was successfully constructed. The titer of the recombinant adenovirus Ad-siCTGF was 2.6×10~(10) pfu/mL after amplification and purification. The CTGF mRNA and protein expression in 4T1 cells were decreased by 36.27% and 31.56%, respectively, compared with the control groups.Conclusion:The recombinant adenovirus which can silence the expression of CTGF was successfully constructed. It laid a good foundation for further investigation of the action mechanism of CTGF in tumor cells.

Abstract: The loop-mediated isothermal amplification (LAMP) technique is a technique for the specific and efficient amplification of target fragments at a constant temperature using two pairs of specially designed primers and a strand displacement activity DNA polymerase. LAMP technique is a simple, rapid, specific, sensitive and cost-effective nucleic acid amplification method, and therefore has a promising future in the field rapid detection of Mycobacterium tuberculosis and grassroots applications. In this review, the basic principles and characteristics of the LAMP technique, the main molecular markers for the diagnosis of tuberculosis, and the use of different molecular markers and various types of novel techniques in the diagnosis of pulmonary tuberculosis, extrapulmonary tuberculosis, and drug-resistant tuberculosis were described. The LAMP technique has been widely used in the diagnosis of tuberculosis with high sensitivity and specificity, but the technique still has some shortcomings. This paper reviews the progress of its application in tuberculosis in recent years and provides an outlook on its development, with a view to providing a rational research direction for rapid diagnosis of tuberculosis in a resource-limited environment.

BackgroundThe immunotherapy for retinoblastoma(RB) is gradually concerned recent year.To seek relative immune-associated antigen is a basis of immunotherapy.NY-ESO-1 and NY-SAR-35 are two kinds of genes of cancer testis antigen( CTA ).To understand their expressions in RB tissue can offer index for relative study.ObjectiveThis study was to investigate the expressions of two CTA NY-ESO-1 and NY-SAR-35 in RB and explore the possibility of them as potentially promising targets for antigen-specific immunotherapy of RB.Methods The samples were obtained from 15 RB eyes,12 non-tumor retinopathy eyes and 22 normal eyes with other benign eye diseases.Reverse transcription polymerase chain reaction(RT-PCR) was used to detect the expressions of NY-ESO-1 mRNA and NY-SAR-35 mRNA in the samples.Genes of positive PCR results were sequenced randomly.The relevance of the expression of the two cancer-testis antigen genes with the clinical characteristics such as tumor stage,tumor size and clinical stage were analyzed.This study was approved by Ethic Committee of Guangxi Medical University.Written informed consent was obtained from each patient before operation. Results NY-ESO-1 mRNA was positively expressed in 6 RB samples and NY-SAR-35 mRNA was expressed in 9 RB samples.In the non-tumor retinopathy samples and normal eye tissues,NY-ESO-1 mRNA and NY-SAR-35 mRNA were absent.No significant relevances were found between the expressions of the NY-ESO-1 mRNA and NY-SAR-35 mRNA with clinical characteristics such as age ( P =0.426,0.822 ),gender ( P =0.180,0.464 ),pathological classification ( P =0.744,0.582 ),tumor size ( P =0.760,0.790),and clinical stage ( P =0.868,0.707 ).Conclusions NY-ESO-1 and NY-SAR-35 have high expressing frequencies in RB tissue and their expressions in RB have specificity.These results offer a clue for the identification of targets antigen of RB.

Huanglian is a commonly used Chinese medicinal herb in the ancient and the present. It has a history of 2000 years in clinical application, having the efficacy of "Clear away heat and remove dampness, purge the sthenic fire and eliminate toxic materials", therefore can be used for various diseases or syndromes in types of dampness-heat and fire-toxin by internal or external use. Compound prescriptions mainly based on Huanglian or prescribed by Huanglian, such as Puji Xiaodu Yin, Huanglian Jiedu Tang, Zhusha Anshen Wan, Qingying Tang, Angong Niuhuang Wan, Niuhuang Qingxin Wan, Jiaotai Wan, Huanglian Ejiao Tang, Zuojin Wan, Danggui Longhui Wan, Huanglian Yanggan Wan, Wu Xiexin Tang, Lianpu Yin, Gegen Huangqin Huanglian Tang, Baitouweng Tang, Xianglian Wan etc. All of these are well-known formulas for clearing away toxin of heat-fire of heart and liver, as well as dampness-heat of stomach and intestines. Nowadays, Huanglian is generally considered as a kind of antibiotic and antivirus herb and is widely used for many infective diseases. In fact, it is also used to cure cardiovascular and cerebrovascular diseases, diabetes and cancer based on pharmacological studies. Having been using Huanglian in treating the above diseases and having conducted clinical and experimental research on cancer and liver diseases, the author observed that Huanglian and its compound prescriptions have obvious effects on liver diseases such as acute or chronic hepatitis, liver fibrosis, liver cirrhosis and liver cancer due to types of dampness-heat and fire-toxin. Part of the effects has been proved by experimental research and it is worth carrying out more research in this area for development and clinical application.
Adult ; China ; Digestive System Diseases ; drug therapy ; Drug Prescriptions ; history ; Drugs, Chinese Herbal ; therapeutic use ; Female ; History, 20th Century ; History, 21st Century ; History, Ancient ; Humans ; Liver Diseases ; drug therapy ; Male ; Medicine, Chinese Traditional ; history ; Middle Aged

BACKGROUNDAlthough video-assisted radical operation for lung cancer has been widely accepted for treatment of nonsmall cell lung cancer (NSCLC), the debate over video-assisted thoracic surgery (VATS) segmentectomy still remains. This study analyzed the clinical outcomes using VATS segmentectomy for stage I NSCLC patients to explore the safety and efficacy of VATS segmentectomy for Ia NSCLC.

METHODSRetrospective review was conducted of patients who underwent VATS segmentectomy for clinical stage I NSCLC at Shanghai Chest Hospital between November 2009 and May 2012. VATS segmentectomy was performed on 36 patients. Analyses of the patient group were performed on patient demographics and clinical characteristics, intraoperative parameters, complications, and postoperative survival.

RESULTSThirty-five of thirty-six patients underwent VATS segmentectomy with only one conversion to open thoracic surgery. There was one peri-operative mortality from the segmentectomy group and all other patients are alive with a median follow up of 327 days. The mean volume of chest tube drainage after operation for segmentectomy was 1021.4 ml. Among other parameters, the mean blood loss was 162.5 ml (50.0 - 1600.0 ml), the mean operation time 124.8 minutes (75.0 - 271.0 minutes), chest tube duration 4.1 days (2 - 8 days), and the mean length of hospital stay 6.2 days (4 - 11 days). There was one (2.8%) locoregional recurrence after segmentectomy. Two patients successfully underwent bilateral segmentectomies and are still disease free.

CONCLUSIONFor patients with stage I NSCLC, VATS segmentectomy offers a safe and equally effective option and can be applied to complicated operations such as bilateral segmentectomy.

Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; mortality ; pathology ; surgery ; Female ; Humans ; Lung Neoplasms ; mortality ; pathology ; surgery ; Male ; Middle Aged ; Neoplasm Staging ; Pneumonectomy ; methods ; Retrospective Studies ; Thoracic Surgery, Video-Assisted ; methods

OBJECTIVETo identify whether SH3GL1 gene serves as a disease associated gene of adolescent idiopathic scoliosis (AIS).

METHODSPositioning candidate cloning: "case-sibling or case-family control design" research scheme based on family constellation was designed. Fifty-six AIS patients (15 male and 41 female, mean age 15 years old, ranged from 8 to 22 years old, Cobb angle from 25 degrees to 110 degrees , average Cobb angle of 67.5 degrees ) from November 2007 to December 2008 were recruited. In all patients, blood preparation was collected, and genome DNA was extracted. According to nucleotide sequence of gene SH3GL1, primer pair for PCR amplification, cloning, and sequencing with 10 exons as emphasis was designed. Sequence comparative analysis for exon sequencing result between sib pairs or family pairs, and that between sib pair or family pairs and NCBI (National Center for Biotechnology Information) were conducted through Vector NTI Advance 10.3 software to judge whether basic group mutation occurred or not. Amino acid sequence comparative analysis for prediction was made.

RESULTSTen exons of the candidate gene SH3GL1 were successfully amplified and cloned in genome DNA of an AIS sib pair and family pairs, and the sequencing obtained positive results. Twelve basic group mutations were found in 10 exons of the candidate gene SH3GL1 of patients with AIS. These mutations were located in the second exon (3 mutations), the fourth exon (1 mutations), the fifth exon (4 mutations), the sixth exon (1 mutations), the eighth exon (1 mutations), and the tenth exon (2 mutations, noncoding region). If basic group in 515 of mRNA was mutated to T, termination codon(TAG) came into being and open reading frame was altered. The sequence of protein showed brachytmema protein was encoded, which could cause changes of primary structure.

CONCLUSIONSH3GL1 is possibly one of the disease associated genes of AIS.

Adolescent ; Base Sequence ; Child ; Exons ; genetics ; Female ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; Male ; Scoliosis ; genetics ; Sequence Alignment ; Young Adult

OBJECTIVETo prepare cryptotanshinone (CT)-cyclodextrin inclusion compound and improve dissolution of CT.

METHODInclusion ratio was determined by plotting the phase solubility curve of CT versus hydroxypropyl-beta-cyclodextrin (HPCD). CT-cyclodextrin inclusion compound was made by wet grinding method. Properties of the inclusion compound was investigated by in vitro dissolution test, DTA and IR spectrum.

RESULTInclusion ratio of CT versus HPCD was 1:1. Dissolution of CT-HPCD inclusion compound at 45 min was 21.6 times of material drug.

CONCLUSIONDissolution of CT was improved remarkably in CT-HPCD inclusion compound. The complexation force of the inclusion compound was hydrogen bond formed by carbonyl group of CT and hydroxyl group of HPCD.

2-Hydroxypropyl-beta-cyclodextrin ; Biological Availability ; Drug Carriers ; Drugs, Chinese Herbal ; chemistry ; Phenanthrenes ; chemistry ; isolation & purification ; Salvia miltiorrhiza ; chemistry ; Solubility ; Technology, Pharmaceutical ; methods ; Time Factors ; beta-Cyclodextrins ; chemistry

Female ; Humans ; Immunoglobulins, Intravenous ; therapeutic use ; Middle Aged ; Pyoderma Gangrenosum ; drug therapy ; pathology ; Skin ; pathology

Adolescent ; Adult ; Female ; Hepatitis ; therapy ; Humans ; Liver, Artificial ; Male ; Middle Aged ; Plasma Exchange

OBJECTIVETo analyze the embryo development potential after intracytoplasmic injection of sperm from azoospermia patients with different spermatogenic functions.

METHODSWe performed ICSI with sperm retrieved from azoospermia patients with different spermatogenic functions using percutaneous epididymal sperm aspiration (PESA) and testicular sperm aspiration (TESA). Then we recorded and analyzed the rates of normal fertilization, cleavages, excellent embryos and pregnancies.

RESULTSNo statistically significant differences were found between the PESA and TESA groups in the rates of normal fertilization ([74.9 +/- 19.6] vs [66.3 +/- 22.7]%, P > 0.05), cleavages ([96.7 +/- 8.6] vs [92.8 +/- 19.8]%, P > 0.05), excellent embryos ([43.5 +/- 26.2] vs [35.0 +/- 29.4]%, P > 0.05) and pregnancies (44.0 vs 52.0%, P > 0.05). The normal fertilization rates in the patients with normal spermatogenesis, mild spermatogenic dysfunction (SD), moderate SD and severe SD were (77.8 +/- 18.4), (68.4 +/- 18.5), (73.5 +/- 19.8) and (51.4 +/- 27.9)%, respectively, with significant difference between the normal spermatogenesis and mild SD groups (P < 0.05) as well as between the severe SD and the other groups (P < 0.05); the cleavage rates were (96.7 +/- 9.2), (96.5 +/- 15.0), (93.9 +/- 12.1) and (93.7 +/- 11.1)%, respectively, with no significant difference among the four groups; the excellent embryo rates were (47.1 +/- 25.8), (40.3 +/- 27.6), (36.2 +/- 23.1) and (15.0 +/- 24.6)%, respectively, with significant difference between the severe SD and the other groups; the pregnancy rates were 54.8, 50.0, 13.6 and 10.0%, respectively, with significant differences among the four groups (P < 0.001).

CONCLUSIONICSI by PESA or TESA is an effective approach to azoospermia. There are no significant differences between PESA and TESA in the rates of normal fertilization, cleavages, excellent embryos and pregnancies. The severity of spermatogenic dysfunction affects fertilization and initial development of embryos, which were shown in the rates of normal fertilization, excellent embryos and pregnancies but not that of cleavages.

Adult ; Azoospermia ; physiopathology ; therapy ; Embryonic Development ; Epididymis ; Female ; Humans ; Male ; Pregnancy ; Pregnancy Rate ; Sperm Injections, Intracytoplasmic ; Sperm Retrieval ; Spermatogenesis ; Young Adult