ObjectiveTo explore a more perfect method for acupoint location by taking point Zusanli (ST36) as an example. MethodPoint Zusanli was located during extension and flexion of the knee joint. The selected points were not at the same place. Its reason was explained. The method of locating point Zusanli was exploredin flexion of the knee.ResultDuring extension and flexion of the knee joint, point Dubi(ST35)shifted superiorly and inferiorly. Therefore, the influence of this factor on the location of point Zusanli was considered when the acupoint was located usingpoint Dubi as a reference point.ConclusionBecause different body positions are often required when acupoints are clinically located, attention should be paid to the problem of how to correctly locate acupoints in different body positions. Again becausethe positions of acupuncture points are changeable and relative, the location of acupoints should be finally determined by acupuncturist using the method of acupoint examination under the direction of various location methods. Therefore, the relationshipbetween locating methods such as bone-length measurements and actual positions of acupoints is worthy to be considered again. Only not abandoning but also not depending on measurements is a more perfect method for acupoint location.

In addition to conventional palpation method, examination on vertebral range of motion (ROM) and mobility can help to determine a pathological displacement and thus provide solid basis for spinal tuina manipulation.

Objective To investigate rhein on podocyte nephrin gene expression and nephrin on glomerular in diabetic nephropa-thy (DN)rats .Methods DN rats mode were made by Intraperitoneal injection of streptozotocin (STZ) 60 mg/kg once .the DN model was divided into 2 groups:rhein treatment group (A) ,model group (B) ,and in addition ,set up a control group (C) .body weight ,kidney weight ,relative kidney weight ,24-hour urinary protein excretion ,serum creatinine ,blood urea nitrogen of rats were detected after 8 weeks of intervention .kidney pathological changes were observed by using light microscope ,electron microscope . immunohistochemical technique used to nephrin protein expression .Results Both the DN rats 24 -hour urinary protein excretion and podocyte nephrin gene expression of group A decreased significantly (P<0 .05) ,improvement in renal function .Conclusion Rhein on DN kidney has a protective effect ,which may be related to its podocyte nephrin gene expression.

Objective To evaluate the genetic factors for sporadic Alzheimer's disease(SAD) among a Chinese population in Beijing. Gene polymorphisms was studied: apolipoprotein E,ps-1 gene E318G missense mutation,alpha(2)-macroglobulin gene Val1000/Ile1000,mtDNA4336G mutation and methylene tetrahydro-fulek reduclase (MTHFR) C677T mutation. Methods The polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP)technique was used to analyze the genotype of 127 SAD patients and 138 non-dementia elderly controls. Results There was a significant difference in the frequency of apoE allele gene between SAD and control group. The freguencies with 2 of apoE4 ,1 of apoE4 and none of apoE4 in SAD were 2.4%,18.1%,79.5% respectively,and in normal control were 0.7%,10.1%,89.2% respectively. mtDNA4336 mutation and ps-1 E318G missense mutation were not found in either Alzheimer's disease or age-matched controls. The frequency of A2M Val1000 (GTC)/Ile1000 (ATC),G/G genetype was 0.02 in SAD and 0.01 in NC . The frequency of MTHFR C677T mutation was 46.3% in SAD and 43.8% in NC respectively, The mutation frequency of cases was not significantly increased than that of controls. Conclusions Our study indicates that apoE?4 allele gene is risk factor for SAD.

Purpose The one-step examination of myocardial perfusion imaging (MPI) combined with coronary artery calcium score (CACS) can obtain both coronary functional information and anatomical information simultaneously, this paper aims to evaluate the value of the one-step examination of MPI combined with CACS for detecting coronary artery disease (CAD). Materials and Methods 188 cases who underwent one-step examination of MPI combined with CACS and invasive coronary angiography (ICA) because of chest tightness, chest pain with suspected coronary artery disease were analyzed retrospectively, with the results of ICA used asgold standard, the diagnostic efficacy of MPI, CACS and one-step examination with combination of the two techniques for CAD was investigated. Results ①Pre-test probability of CAD was intermediate in 79.8%(150/188), and high in 20.2%(38/188) cases. Seventy-three cases were confirmed as CAD and 115 of 188 patients were negative according to ICA.②The sensitivity, specificity, accuracy, positive predictive value (PPV) and negative predictive value (NPV) for the diagnosis of CAD by MPI were 65.8%, 75.7%, 71.8%, 63.1%and 77.7%, respectively. ③ The CACS of CAD group was significantly higher than the non-CAD group (494.96±99.60 vs. 38.15±16.03, P<0.05). According to the features of the ROC curve, the best threshold for the diagnosis of CAD with CACS was 96.45, with CACS≥96.45 as the positive standard in diagnosis of CAD, the sensitivity, specificity, accuracy, PPV and NPV for the diagnosis of CAD by CACS were 60.3%, 93.9%, 80.8%, 86.3%and 78.8%, respectively. ④ The sensitivity of MPI combined with CACS were significantly higher than MPI (80.8% vs. 65.8%, P<0.05), while the specificity (71.3% vs.75.7%, P>0.05) and accuracy (75.0% vs. 71.8%, P>0.05) showed no statistically significant difference; the sensitivity of MPI combined with CACS were significantly higher than CACS (80.8%vs. 60.3%, P<0.05), while the specificity was lower than CACS (71.3%vs. 93.9%, P<0.05) and the accuracy showed no statistically significant difference (75.0%vs. 80.8%, P>0.05). Conclusion The one-step examination of MPI combined with CACS can reduce coronary heart disease misdiagnosis, improve the diagnostic sensitivity of CAD compared with the MPI or CACS, with high application value for the diagnosis of CAD, especially in moderate risk groups.

Objective To investigate the effect of needle sizes and aspiration techniques on sample quantity. Methods Aspiration was performed on porcine liver in vitro for 10 times with three different sizes of needles(19 G, 22 G and 25 G) and four different aspiration techniques[non?negative pressure(NP), 10 ml NP,20 ml NP and slow?pull], 20 mm in depth. A total of six aspirations were performed with each needle by the same aspiration technique. All the obtained specimens were fixed in formalin with the cell block method. The samples were evacuated according to our grading criteria. Results The mean±standard deviation(SD) score for 19 G,22 G, 25 G were 5?71±0?69,4?63±1?24, 3?79±1?84 respectively. The mean±SD score for methods non?NP,10 ml NP,20 ml NP and slow?pull were 4?72±1?53,4?56±1?46,4?72±1?50,4?83±1?76 respectively. The multi?analysis of variance results showed that there were statistical differences between different needles size( F=12?00,P<0?001) with 19 G being the best,followed by 22 G and the least specimen obtained by 25 G needle. There were no statistical differences among aspiration techniques ( F=0?128, P=0?943).The analysis showed that the thicker the needle was,the better sample quality was 19 G yielded to the highest quantity of specimens. The most specimens could be obtained with 19 G needle and non?NP, 22 G needle and 20 ml NP and 25 G needle and slow?pull. Conclusion In clinic, aspiration technique should be selected according to different aspiration needles. 19 G is superior to others, with non?NP method. For 22 G needle, 20 ml NP is preferred and for 25 G needle,slow?pull is preferred.

AIM:To construct a hAR and GFP fusion gene vector and to observe the AR-GFP gene expression in Hek293 cells. METHODS: A recombined vector pcDNA3.1/myc-HisA-AR-GFP (pH-AG) was constructed by gene engineering technique. The recombined vector was transfected into Hek293 cells using calcium phosphate. RESULTS: AR-GFP fusion protein was successfully expressed in Hek293 cells without biologic activity, which was confirmed by fluorescence microscopy and Western blotting. CONCLUSION: The Hek293 cells transfected by AR-GFP fusion gene can express its protein successfully. However, it is not a cellular model for ARI screening.

To investigate the effect of Kozak sequence (+4A or +4G) on expression of green fluorescent protein (GFP) gene in HEK293 cells. The eukaryotic expression vectors containing GFP gene with different Kozak sequence (+4A or +4G) were constructed by classic DNA recombination methods, including PCR, enzyme digestion, ligation, transformation, identification, et al. Two different Kozak sequences (+4A or +4G) were obtained through PCR with different mutagenic primers. The right recombinant plasmids pHGFP-A and pHGFP-G were transfected into HEK293 cells by liposome-mediated gene transfer method. The expression level of GFP was observed by fluorescent microscope, flow cytometry and Western blot. The flow cytometry revealed that the expression levels of GFP fluorescence in pHGFP-A and pHGFP-G transfected cells were about 15% and 45%, respectively. Western blot showed the specific bands of about 27 kD (GFP) both in pHGFP-G and pHGFP-A sample lanes; and the GFP expression density of pHGFP-G was about 3.87-fold as that of pHGFP-A by ImageJ software analysis. These results indicated that the +4G in Kozak sequence (when -3 site is purine base pair) plays an important role in GFP protein translation, which enhances the GFP expression up to 4-fold in HEK293 cells.
Actins ; biosynthesis ; genetics ; Cell Line ; Cloning, Molecular ; Gene Expression Regulation ; genetics ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; biosynthesis ; genetics ; Humans ; Protein Processing, Post-Translational ; RNA, Messenger ; metabolism ; Recombination, Genetic ; Transfection

ObjectiveTo study the molecular genetic mechanism of para-bombay phenotype in two individuals.MethodsThe proband was a female.When the proband donated blood,because the forward blood group wasn't coincident with her reverse blood group,the blood and saliva specimen from proband and her family members were sent to Guangzhou Blood Center for further identification.Routine serological techniques were used to determine proband's and her family members' blood group and ABH antigen in saliva.The coding regions of FUT1 and FUT2 gene,exon 6 and exon 7 of ABO gene were amplified by polymerase chain reaction using proband's and her family members' genomic DNA.All amplified products were analyzed after being directly sequenced.The two-base deletion regions of FUT1 gene were certified by cloning and haplotype sequencing.Results Proband's and her little brother's blood group were identified as para-bombay while other family members' blood group were normal.Two-base deletion heterozygous mutations of FUT1 gene were found in proband and her brother,AG deletion at position 547-552 and TT deletion at position 880-882,which caused a reading frame shift and a premature stop eodon.Meanwhile,880-882del TT heterozygous mutation was found in proband's grandfather and her father and 547-552del AG heterozygous mutation was found in proband's mother and her little sister.ResultsOf cloning and haplotype sequencing certified that these two-base deletion mutations occurred at 547-548 and 881-882 position respectively.Three new mutations were found in FUT2 gene,390C ＞ T,418A ＞ T and 749G ＞ A,which could cause the change of amino acid at position 140Ile ＞ Phe and 250Arg ＞ Gln.Conclusions Two-base deletion heterozygous mutations in different positions in FUT1 gene were found in 2 individuals,which maybe the molecular genetic mechanism of para-bombay phenotype.Heterozygous deletion mutation in one-strand DNA wouldn't change the ABO blood group.Three new mutations were also found in FUT2 gene.( Chin J Lab Med,2012,35:815-819)

OBJECTIVETo investigate the clinicopathological features of small gastrointestinal stromal tumors(GISTs) and to evaluate the efficacy of endoscopic therapy for GISTs.

METHODSClinicopathological and follow-up data of 418 patients with GISTs undergoing endoscopic therapy in the Zhongshan Hospital between January 2009 and July 2014 were analyzed retrospectively. All the cases were evaluated by the NIH risk classification and AIFP classification, and were grouped according to the tumor size and location. Nuclear atypia and mitotic count were used to evaluate the biological behavior of small GIST. Efficacy of endoscopic therapy was analyzed with follow-up data.

RESULTSOut of 418 patients, GISTs located in the esophagus was 14(3.3%), in the stomach 389(93.1%), in the duodenum 5(1.2%), in the rectum 10(2.4%). A total of 412(98.6%) patients were mainly spindle cells, and mitosis was not found in 320(76.5%) patients. In 389 small stomach GIST, 245(58.6%) were in fundic region. Cases were divided into four groups according to the size and the result revealed the bigger the size, the more the mitotic count. Nuclear atypia in the 1.5-1.9 cm group was significantly higher compared to other groups. Cases were divided into four groups according to the location and the result revealed the mitotic count was not associated to the location. While the nuclear atypia of stomach GIST was significantly higher than that of esophageal GIST and the nuclear atypia of rectum GIST was significantly higher than that of other positions. The median follow-up was 32(4-69) months. One case(gastric fundus GIST, >1.5 cm) presented local recurrence 23 months after operation and underwent endoscopic resection again. No recurrence or metastasis was found in other patients.

CONCLUSIONSEndoscopic resection technique is effective for small GISTs patients. The small GISTs with 0.4 cm diameter or less are often benign and should be followed up for long time. The small GISTs with 0.5 cm diameter or more possess the risk of malignancy, then surgical resection should be performed. Rectum small GISTs (except for 0.4 cm diameter or less) have worse biological behavior and should be removed.