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Objective:To investigate the feasibility of coronary CT angiography(CCTA)-feature tracking(FT) for assessing global and regional myocardial strain in patients with heart failure(HF).Methods:From July 2019 to December 2020, twenty-five patients diagnosed with HF from Peking Union Medical College Hospital were prospectively enrolled into the study. All patients underwent retrospective electrocardiogram-gated CCTA and cardiac MR (CMR) imaging within 7 days. CCTA-FT and CMR-FT were undertaken using cvi 42 dedicated commercial software to measure global and regional strain parameters, including global peak radial strain (GPRS), global peak circumferential strain (GPCS) and global peak longitudinal strain (GPLS), as well as peak radial strain (PRS), peak circumferential strain (PCS) and peak longitudinal strain (PLS) of left ventricular basal segment, middle segment and apical segment. Conventional left ventricular functional parameters were also calculated, including left ventricular ejection fraction (LVEF), left ventricular stroke volume (LVSV) and left ventricular mass index (LVMI). Paired t test or Wilcoxon signed-rank test was used to compare the differences of measurements between CCTA group and CMR group. Pearson or Spearman correlation analysis was used to analyze the correlation between the two groups. Inter-and intra-observer consistence in CCTA group was evaluated by intraclass correlation coefficient (ICC) analysis. Results:The effective radiation dose of CCTA examination was 6.00 (4.86,7.63) mSv. Inter-and intra-observer consistence in CCTA group was excellent, and the ICC value was 0.85-0.98. In the overall strain parameters, GPCS in CCTA group[-8.10%(-10.32%, -5.20%)] was significantly lower than that of CMR group[-8.49%(-13.79%, -5.95%)] ( Z=-2.15, P=0.031). There was no significant difference in GPRS and GPRS between the two measurement methods ( P>0.05). Strong correlations were observed between GPRS, GPCS and GPLS ( r=0.65, 0.63, 0.71,all P<0.001). For local strain parameters, PCS in the middle segment and apical segment of CCTA group were lower than those of CMR group ( Z=-2.17, -2.62, all P<0.05). There were no significant differences in PCS of basal segment, PRS and PLS of all segments between groups (all P>0.05). The PCS and PLS of basal segment, PRS of middle segment and PRS of apical segment were moderately correlated ( r=0.46, 0.52, 0.58, 0.53, P<0.05); The other local strain parameters were strongly correlated, the range of r value was from 0.64 to 0.70 (all P<0.001). For left ventricular functional parameters, LVEF, LVSV and LVMI showed no significant differences between groups ( P>0.05), and the correlation was extremely strong ( r=0.90, 0.89, 0.96, all P<0.001). Conclusions:The repeatability of CCTA-FT technique in measuring myocardial strain was good, and the correlation of parameters measured by CCTA-FT technique and CMR-FT technique was excellent. Therefore, CCTA-FT technique can be used as a new noninvasive and simple method to evaluate myocardial motor function.
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Objective:To document the clinical efficacy of using electrical stimulation to guide acupuncture on the peroneal longus and the peroneal brevis of patients with functional ankle instability (FAI).Methods:Seventy FAI patients were randomly divided into an observation group and a control group, each of 35. Both groups received routine balance function training, proprioception training and manual activation of the peroneal muscles. The observation group also received acupuncture at the motor points of the peroneal longus and the peroneal brevis muscles under the guidance of electrical stimulation, once a day, for 3 weeks. Before and after the treatment, any curative effect was quantified using the CAIT ankle instability assessment questionnaire, electromyography (iEMG) of the peroneal longus and the peroneal brevis, as well as those muscles′ excitation times during simulated varus walking.Results:All of the measurements after the treatment were significantly better than before the treatment, with the improvements in the observation group significantly greater than in the control group.Conclusions:Acupuncture at the motor points of the peroneal longus and the peroneal brevis under the guidance of electrical stimulation can effectively activate the related muscles and relieve the symptoms of functional ankle instability.
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【Objective】 To construct the eukaryotic expression vector carrying the human wild-type p27 and lacking nuclear localization signal p27△NLS coding sequences， and the express them in HEK293T cells， which may contribute to investigating the different locations and roles of p27 in the cytoplasm and nucleus. 【Methods】 Total RNA was prepared from human breast cancer MCF7 cells， and cDNA was obtained by reverse transcription-polymerase chain reaction （RT-PCR）. After amplification of the p27 CDs and non-NLS fragments by PCR， full length p27WT （CDKN1B， NM_004064.5） and p27△NLS coding regions were obtained. PCR products were then subcloned into the eukaryotic expression vector pCMV-Blank. After identification with bacterial PCR， double restriction enzyme digestion and sequencing， they were defined officially as pCMV-p27WT and pCMV-p27△NLS， respectively. The recombinant plasmids were transfected into HEK293T cells by electroporation. After 48 h， the levels of p27 protein in the cytoplasm and nucleus were detected by Western blotting. 【Results】 The sequencing results showed that the sequences of p27WT and p27△NLS inserted into the plasmids were both correctly consistent with that of NM_004064.5. After transfection with pCMV-p27WT， total p27 protein expression was increased and distributed in both the cytoplasm and nucleus of HEK293T cells. After transfection with pCMV-p27△NLS， p27 protein was significantly increased and almost entirely localized in the cytoplasm of HEK293T cells. 【Conclusion】 The eukaryotic expression plasmids of human p27WT and p27△NLS coding sequences were successfully constructed and overexpressed in HEK293T cells. This research may lay a foundation for investigating the biological function of p27 in the cell cycle progression of tumor cells.
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Objective:To explore the feasibility in evaluating segmental extracellular volume (ECV) using dual-layer spectral detector CT in patients with heart failure with various percentages of delayed hyper-enhanced volume, using MRI as a reference.Methods:Twenty-seven patients with heart failure in Peking Union Medical College Hospital were prospectively enrolled in this study from July 2019 to January 2021. All the patients underwent both CT late iodine enhancement and MR late gadolinium enhancement imaging within a week. According to percentages of hyper-enhanced volume on cardiac MR image, myocardial segments were classified into 3 groups: 0-4% (group A), 5%-49% (group B) and 50%-100% (group C). ECV images were independently observed by 2 experienced radiologists. The correlation and agreement between CT-ECV and MRI-ECV for the whole myocardial assessment and for the segmental assessment were evaluated using Spearman correlation and Bland-Altman analysis. Inter-observer agreement was assessed using the ICC.Results:Of the 27 patients, 432 segments were analyzed, with 199 segments in group A, 151 segments in group B and 82 segments in group C. In the whole myocardial analysis, the CT-ECV and MRI-ECV were 32.25% (28.04%, 34.28%) and 32.27% (28.35%, 34.77%), respectively. A good correlation was found between CT-ECV and MRI-ECV ( r=0.969, P<0.001). Bland-Altman analysis between CT-ECV and MRI-ECV showed a small bias (0.4%), with 95% limits of agreement of -6.9% to 7.8%. as for the segmental myocardial assessment, the CT-ECV was 28.23% (26.18%, 29.92%) for group A, 33.29% (30.16%, 35.96%) for group B and 40.22% (34.06%, 46.70%) for group C. There was statistically significant difference in ECV among three groups (all P<0.001). Good correlations between CT-ECV and MRI-ECV were found in group A ( r=0.614, P<0.001), group B ( r=0.852, P<0.001) and group C ( r=0.953, P<0.001). Bland-Altman analysis between CT-ECV and MRI-ECV showed a small bias 2.0% (95% limits of agreement:-17.6%-21.6%) in group A, a small bias -0.4% (95% limits of agreement: -13.8%-12.9%) in group B and a small bias -1.8% (95% limits of agreement: -17.4%-13.9%) in group C. The ICC values between 2 observers were 0.877 and 0.945 on CT-ECV and MRI-ECV, respectively. Conclusions:The larger percentage of hyper-enhanced volume on cardiac MR image, the stronger correlation are between the CT-ECV and MRI-ECV. CT-ECV may be taken as an alternative tool for quantitative evaluation of myocardial tissue in patients with heart failure.
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【Objective】 To construct the Rb luciferase reporter gene assay system and detect the activation ability of Rb gene for screening the targeted drugs. 【Methods】 The synthetic Rb gene sequence was annealed to form a double-stranded DNA structure and then inserted into the polyclonal site of pGL6-TA. The junction product was transformed into E.coli DH5α competent cells for expanded culture, and the constructed pGL6-Rb-Luc plasmid and pGL6-TA plasmid were transfected into HEK293 cells. The monoclonal cell line HEK293-Rb-Luc with stable expression was screened by G418, and the activation and inhibition of Rb in HEK293-Rb-Luc were tested by serum and CDK4/6 inhibitor Palbociclib. 【Results】 The sequence of Rb reaction elements in pGL6-Rb-Luc was completely correct. The recovery of serum culture significantly increased the luciferase activity in HEK293-Rb-Luc (P<0.001). Compared with 0 nmol/L, 25, 50, 75 and 100 nmol/L, CDK4/6 inhibitor Palbociclib made the inhibition rate of Rb activity rise to 6.90%, 40.23%, 50.57% and 52.07%, respectively (P<0.05). 【Conclusion】 The Rb luciferase reporter gene detection system HEK293-Rb-Luc was successfully constructed, which can effectively detect the activation level of Rb.
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【Objective】 To clone the full-length of human kidney and brain protein (KIBRA) coding sequence in eukaryotic expression vector and provide a model for studying the biological function of KIBRA in breast cancer cells. 【Methods】 Total RNA of human breast cancer cell line MCF7 was extracted. After reverse transcription, the full length of KIBRA (NM_001161661.2) coding region was amplified by PCR, and cloned into eukaryotic expression vector pCMV-Blank. After identification, it was defined officially as pCMV-KIBRA. Then it was transfected into MCF7 cells, and the expression of KIBRA was detected by real-time PCR and Western blotting after 48 hours. The primary, secondary and tertiary structures and post-transcriptional modification sites of KIBRA were analyzed with bioinformatics software. 【Results】 Bacterial PCR, double enzyme digestion and DNA sequencing results showed that the correct sequence of KIBRA was inserted into the vector pCMV-KIBRA. The mRNA and protein expressions of KIBRA were significantly increased in MCF7 cells transfected with pCMV-KIBRA. Bioinformatics analysis showed that KIBRA was composed of 1119 amino acids. There were 52 phosphorylation sites, 1 acetylation site and 5 ubiquitination sites, and the protein structure was mainly α-helix and random coil. 【Conclusion】 The eukaryotic expression vector of full-length of human KIBRA coding sequence was successfully constructed and overexpressed in breast cancer cell line MCF7, which can lay a foundation for studying the biological function of KIBRA in breast cancer.
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Objective:To evaluate the feasibility of high concentrated contrast media combined with monochromatic images to improve image quality with double low scanning in dual-layer spectral coronary CT angiography.Methods:Fifty-six patients with suspected coronary artery disease were enrolled and randomly separated into two groups. All patients were scanned at 120 kVp in step-and-shoot mode using a dual-layer detector CT (IQon spectral CT). Patients were either injected with 18 ml high concentration contrast medium(400 mgl/ml) at 2 ml/s (group A) or 45 ml contrast medium (370 mgl/ml) at 4 ml/s (group B). Forty to 80 keV monoenergetic images with 10 keV increment and conventional image (group A2) were reconstructed for group A, conventional poly-energetic image was reconstructed for group B. Region of interest was placed on aorta root (AO), middle segment of left anterior descending (LAD), left circumflex (LCX) and right coronary artery (RCA) to measure the attenuation and noise, signal-to-noise ratio (SNR) and contrast-to-noise ratio (CNR). Image quality was assessed by two reviewers independently with a 4-point scale on image quality (1-undiagnostic,4-excellent). The objective and subjective image evaluation were compared using the Kruskal-Wallis test. The Steel Dwass was used for multiple comparisons between monoenergetic images in Group A and conventional images in Group B, after the Kruskal-Wallis test.Results:There were significant differences among the attenuation, noise, SNR and CNR of group B, group A2, and monoenergetic images(all P<0.001).The CT value in the aortic root was significantly higher at 40-50 keV monoenergetic images than that in group B(all P<0.05), and similarly, the CT value in LAD,LCX and RCA at 40 keV were superior than that in group B(all P<0.001). There was no significant difference in the noise of the aortic root among 40 keV image, group A2 and group B(all P>0.05),while the noise in the 50-80 keV monoenergetic images were significantly lower than that in Group B(all P<0.001). The SNR and CNR in the aortic root were significantly higher at 40-60 keV monoenergetic images than that in group B(all P<0.001).The SNR in LAD and RCA at 40，50 keV and the CNR in LAD and RCA at 40-60 keV were higher than that in group B(all P<0.01).The SNR at 40-60 keV and CNR at 40-70 keV in LCX were superior than that in group B(all P<0.05). There were significant differences among the subjective image quality score of group B, group A2, and monoenergetic images(all P<0.001).The subjective image quality score of 40，50 keV images were not significantly different from that in group B (all P>0.05),while the score in 60-80 keV image and group A2 were lower than that of Group B(all P<0.001). Conclusion:40，50 keV low monochromatic images derived from dual-layer spectral detect CT combined with high concentrated contrast media can provide comparable or superior image quality with double low scanning in CCTA study.
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Objective To discuss the expression and clinical significance of hypoxia-inducible factor-1α (HIF-1α) and osteopontin (OPN) in esophageal cancer tissues.Methods A total of 100 patients with esophageal cancer in Enshi Tujia and Miao Autonomous Prefecture Central Hospital of Hubei Province from January 2015 to January 2017 were selected.The expressions of HIF-1α and OPN in esophageal cancer and adjacent tissues were detected by immunohistochemistry.The relationships between HIF-1α and OPN expressions and clinical pathological characteristics of esophageal cancer patients,as well as the correlation between them were analyzed.Results The positive rates of HIF-1α and OPN expression in esophageal cancer tissues were 70.00％ (70/100) and 64.00％ (64/100),which were significantly higher than the 14.00％ (14/100) and 8.00％ (8/100) in adjacent tissues,and the differences were statistically significant (x2=64.368,P ＜ 0.001;x2 =68.056,P ＜0.001).The expression of HIF-1α was correlated with TNM stage,depth of invasion and lymph node metastasis (x2 =30.159,P ＜ 0.001;x2 =17.493,P ＜ 0.001;x2 =16.357,P ＜ 0.001),but it was not correlated with sex,age,tumor diameter and differentiation (all P ＞ 0.05).The expression of OPN was correlated with TNM stage,depth of invasion and lymph node metastasis (x2 =12.558,P =0.002;/x2 =15.395,P ＜ 0.001;x2 =17.056,P ＜ 0.001),but it was not correlated with sex,age,tumor diameter and differentiation (all P ＞ 0.05).There was the positive correlation between HIF-1α and OPN expression in esophageal cancer tissues (x2 =61.144,P ＜ 0.001,C =0.616).The 2-year survival rates of HIF-1α and OPN positive patients in esophageal cancer tissues were 51.43％ and 50.00％ respectively,which were significantly lower than those of HIF-1α and OPN negative patients (80.00％ and 77.78％),and the differences were statistically significant (x2 =7.143,P =0.008;x2 =7.407,P =0.006).Conclusion HIF-1 α and OPN are highly expressed in esophageal cancer tissues,and their high expression may be related to poor survival prognosis of esophageal cancer.
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Objective To investigate the characteristics and the enhanced patterns of carotid plaque using contrast-enhanced ultrasound (CEUS) and patients with cerebral infarction.Methods The patients with carotid plaque were divided into two groups according to whether they had cerebral infarction:54 patients (62 plaques with CEUS) with cerebral infarction were included in group A,and 48 patients (54 plaques with CEUS) without cerebral infarction were included in group B.The plaques were divided into four grades according to the degree of plaque enhancement.According to the source of intraplaque contrast agents,plaque enhancement patterns were divided into adventitia enhancement,lumen enhancement and mixed enhancement.To analyze the degree and pattern of carotid plaque enhancement in the two groups.Results Carotid plaque enhancement in cerebral infarction group was mainly grade 3 (26/62) and grade 4 (22/62),while that in non-cerebral infarction group was mainly grade 2 (20/54) and grade 3 (20/54).There was significant difference between the two groups in the proportion of carotid plaque enhancement of grade 2 (P =0.019) and grade 4 (P =0.041).The proportion of plaque adventitia enhancement model in group A(27/59) was lower than that in group B (37/50),with statistically significant difference (P =0.003).While the proportion of mixed enhancement mode in group A was significantly higher than that in group B (P =0.003).Conclusions The enhancement of carotid plaque was obvious in cerebral infarction patients,and the mixed enhancement pattern was more common.It suggested that the communication between vascular cavity and plaque might be an important factor leading to cerebral infarction.
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Objective To preliminarily investigate and optimize the saline flush effects to the image quality (IQ)and effective scan rate of high-pitch CT coronary angiography (CCTA)using ultra-low contrast media (CM)protocols on third-generation dual-source CT (DCST). Methods Sixty-eight consecutive patients with suspected or known coronary artery disease (CAD)were recruited and underwent prospective high-pitch CCTA on third-generation DSCT with automatic tube voltage selection (70-80 kV)based ultra-low CM protocols:18 ml(3.0 ml/s) for 70 kV and 21 ml (3.0 ml/s) for 80 kV. All the patients were randomly divided in to two groups:Group A(n=31)with the saline flush of 3.0 ml/s(40 ml) and Group B (n=37)with the saline flush of 4.0 ml/s(40 ml). The baseline data,risk factors,quantitative image quality (IQ),qualitative IQ results,and effective dose (ED)were evaluated and compared. Results There was no significant difference in ED between two groups [(0.36±0.12)mSv vs. (0.37±0.13)mSv;t=-0.40,P=0.70]. The average ED in all patients was (0.43±0.28)mSv. However,the contrast to noise ratio (CNR) of right coronary artery distal segment in Group A was significantly higher than that in Group B(76.04±62.46 vs. 43.96±26.94;t=2.67,P=0.01) and the CNR of left circumflex artery proximal segment in Group A was significantly higher than that in Group B(69.60±43.13 vs. 44.08±24.08;t=2.96,P=0.01). No other significant difference was found in terms of baseline characteristics,and objective image quality (all P>0.05). Subjective IQ showed no significant difference between these two groups (χ=2.53,P=0.47). Conclusion Under high-pitch CCTA using ultra-low CM protocols [18-21 ml (3.0 ml/s)]on third-generation DCST,the saline flush at a flow rate of 3.0 ml/s can achieve satisfactory IQ and effective scan rate.
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Humanos , Angiografía por Tomografía Computarizada , Métodos , Vasos Coronarios , Estudios Prospectivos , Dosis de Radiación , Interpretación de Imagen Radiográfica Asistida por Computador , Cloruro de SodioRESUMEN
Objective To investigate the optimized range of body mass index (BMI) selection for patients undergoing 70 kV high-pitch dual-source coronary CT angiography (CCTA) on the third-generation dual-source CT (DSCT). Methods Patients undergoing prospective high-pitch ultra-low contrast media (CM) CCTA on the third-generation DSCT using the automatic tube voltage selection at 70 kV were included and divided into three groups:group A,with BMI≤24 kg/m;group B,with 24 kg/m<BMI<28 kg/m;and group C,with BMI≥28 kg/m. The baseline data,image quality results,and effective dose (ED) were evaluated and compared. Results No significant difference was found in age (H=2.36,P=0.78),heart rate (H=3.51,P=0.80),and ED(H=1.77,P=0.12) among three groups. The noise of the aortic root in group A was (21.66±4.00)HU,which was significantly lower than that in group C [(23.61±4.20)HU](F=2.05,P=0.03). The signal to noise ratio of aortic root in group A (18.61±5.28) was significantly better than that in group C (14.75±2.58) (F=5.31,P=0.02). No other significant difference in quantitative image quality were found among the three groups (all P >0.05). Subjective image quality in group A(Z=2.91,P=0.004) and B(Z=2.27,P=0.021) were both significantly better than that in group C. Conclusion The ultra-low tube voltage (70 kV) combined with ultra-low CM CCTA protocol on third-generation high-pitch DSCT may be better for patients with BMI<28 kg/m2 than those with BMI ≥28 kg/min China.
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Humanos , Índice de Masa Corporal , China , Angiografía por Tomografía Computarizada , Métodos , Medios de Contraste , Frecuencia Cardíaca , Estudios Prospectivos , Dosis de Radiación , Interpretación de Imagen Radiográfica Asistida por Computador , Relación Señal-RuidoRESUMEN
Objective To investigate the effects of exercise on learning and memory after cerebral ischemia. Methods Ninety Sprague-Dawley rats were randomly divided into a sham-operated group, a model group and a treadmill training group ( n=30 for each) , which were further subdivided into groups to receive 7 days, 14 days or 28 days of training with 10 rats in each. The training was treadmill running at 10 m/min for 30 min a day. Cerebral ische-mia was induced in the model and training groups using permanent, bilateral common carotid artery occlusion. The training began three days after the operation. Morris water maze tests were used to measure the rats′ learning and memory ability, and Nissl staining was employed to detect the survival rates of pyramidal neurons in the CA1 area of the hippocampus. Results The average escape latency in the treadmill trained group had shortened significantly by day 7, then further by days 14 and 28. It was significantly shorter than the model group′s average at each time point. The average platform crossing time increased significantly compared with the model group′s average. Few dead neu-rons were observed in the sham-operated group. On days 14 and 28 the average survival rate of pyramidal neurons in the model group was significantly lower than in the treadmill training group or the sham-operated group, though there was no significant difference on day 7. Conclusion Treadmill training can improve learning and memory after cere-bral ischemia, at least in rats. Better effects can be observed after longer training.
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This paper aimed to review the recent literature on the TCM treatment for stroke sequelae. The content summarized and discussed the definition of stroke sequelae, herbalmedicine, TCM patents, acupuncture and so on. Sequelae of stroke’s pathogenesis was mainly caused by deficiency generating stasis, so the treatment principles were to tonifying Qi and blood and promote circulation and remove stasis with the common prescriptions, such asBuyang-Huanwudecoction,Xuefu-Zhuyu decoction andDihuang-Yinzi. Meanwhile, Chinese medicine patents and acupuncture treatment canimprove the brain function and ADL.
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Previous studies have suggested that various kinds of inflammatory factors can influence the formation and development of tumor cells.Researche has shown that gallbladder cancer is closely linked with local inflammation,which is a risk factor for the development of gallbladder cancer.It is widely known that cholecystitis is closely correlated with gallstones,and that bile obtained from patients with gallbladder cancer contains a large variety of bacteria,such as Salmonella typhi,Helicobacter,and Escherichia coli.It is proposed that the gallbladder may be the result of the joint action of inflammation with the bacterial flora.Similarly,the inflammatory “tumor infiltrating lymphocyte” (TIL)can be observed in the tumor and its surrounding tissues,and may also play a role in tumor growth and metastasis.However,detailed mechanisms about the relationship between inflammation and gallbladder cancer is still not clear.No specific anti-inflammatory drugs for gallbladder cancer have been developed. In the near future,anti inflammatory drugs may play a more important role in gallbladder cancer prevention and treatment.
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ObjectiveInvestigate the value and feasibility of image registration with reverse rotation in lateral cerebral DSA.Methods ( 1 ) Experimental study:the target images were subtracted directly,and subtracted again after reverse rotation.Software of registration and subtraction with reverse rotation edited by the author utilizing Visual Basic.The function of the automatic angle detection by the software were evaluated to see whether it detected the angle of line.The subtraction function of DSA by the software was evaluated.(2) Clinical retrospective study:the untreated mask and target inages of 15 patients with motion along vertical axis during lateral cerebral DSA were uploaded to the software.The target images were processed with and without the software to get two sets of images.( 3 ) Evaluation:four experienced radiologists read and compared the two sets of the images,and graded their findings.Results ( 1 ) The automatic detection by the software suggested that the target images should be rotated counterclockwise 1.3°. The subtraction result of the software was satisfactory.(2)In the 15 sets of images,there were only three sets of images deemed optimal after traditional subtraction.After reverse rotation,artifacts were significantly reduced and the image sharper. There were ten cases with significant artifacts after traditional subtraction,and those images were sharper and showed more peripheral vessels after reverse rotation. The traditional subtraction images of two sets could not be interpreted,the reverse rotation registration images reached the diagnostic quality.(3)Subjective evaluation: there were more information and less noise and distortion in the registration images with reverse rotation than in the traditional subtraction. But the image resolution decreased slightly after reverse rotation registration.ConclusionThe registration of digital angiography with reverse rotation can improve the image quality in lateral cerebral DSA.
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Objective To prepare the monoclonal antibody against human perforin(HP). Methods Recombinant eukaryotic expression plasmid pCDM8-HP was extracted and purified, and the BALB/C mice were immunized with the plasmid. The hybridomas producing anti-HP McAbs were established by using hybridoma technique, then the specificity of the McAbs was identified by using immunocytochemical technique and Western blot. Results Three hybridoma cell lines secreting McAbs against human perforin were established, and the three McAbs showed positive only with LAK cells containing human perforin protein, and showed negative with inactive human peripheral blood lymphocytes (PBLs). The subclasses of the three McAbs were determined as lgG2bK. Western blot results showed that the three McAbs recognized a specific band of LAK cell lysateds with molecular weight of 70.0Kd. Conclusion The three hybridoma cell lines secreting McAbs against human perforin were established and the secreted McAbs were specific.
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Objective To express human B7.2 extracellular domain with prokaryote expression system and to evaluate its biological activity in vitro. Methods PCR was used to amplify the extracellular region of human B7. 2which contained both the IgV and IgC domains. The recombinant PGEX-4T-3/hB7. 2 (IgV+C) was obtained by cloning the PCR product into a prokaryote expression plasmid PGEX-4T-3 and was transformed into the host strain of DH5-α. The fusion protein consisted of GST and hB7.2(IgV+C) was identified by SDS-PAGE and Western blotting.T cell activation was observed by exposing purified T lymphocytes to the fusion protein and [3H]-TdR incorporation with the presence of the first signal imitated by anti-CD3 antibody. Results The fusion protein GST-hB7.2 (IgV+C) was produced and detected in inclusive body form from engineered bacterial cells. With the first signal existed,T lymphocytes proliferated when it was co-stimulated by the fusion protein. Conclusion These results indicated that the functional human B7.2(IgV+C) fusion protein can be produced in bacterial cells and the fusion protein displays the co-stimulatory activity in T lymphocytes activation.