Observed index is a very important element in a research design, because it is a specific reflection of the effects of research factors on the research subjects and is indispensable in any research. Generally, there are two types of observed indexes: the indexes that reflect natural attributes, habits or states of the research subjects and the indexes that reflect the effects of different drugs or treatments on research subjects. This article mainly introduces the definition, characteristics, selection and observation of research indexes and the major and minor indexes.

The purpose of this study was to construct expression vectors of idiotype (Id) SmIg in patients with B-chronic lymphocytic leukemia and to express them in E.coli to obtain recombinant Id, and to investigate the effect of the protein on the proliferation and secretion of IL-2 and IFN-gamma of stimulated peripheral blood mononuclear cells (PBMC) in vitro. Light chain gene and Fd fragment of heavy chain gene were inserted into fd-tet-DOG2 vector to construct fd-tet-DOG2-Fab. Fab gene was further cloned into expression vector pHEN2 to construct the soluble expression vector pHEN2-Fab. After induction by IPTG, Fab protein was purified by Ni-NTA-chromatography. MTT was used to determine the effects of purified protein on the proliferation of stimulated PBMC in vitro and the concentrations of IL-2 and IFN-gamma in the culture supernatants were detected by ELISA. The results showed that recombinant pHEN2-Fab expression vector was constructed successfully. Fab protein was expressed in positive clone after induced by IPTG and two specific bands at 24-25 kD position were observed by SDS-PAGE electrophoresis. Proliferation of PBMC could be induced by purified Fab and the concentrations of IL-2 and IFN-gamma in culture supernatants were increased significantly after induction. It was suggested that the expression vector of SmIg Fab fragment was constructed successfully, and expressed and secreted from E. coli. The Fab protein could induce proliferation of PBMC and promote secretion of IL-2 and IFN-gamma.

Objective To investigate the comprehensive treatment of Crohn's disease complicated with anal fistula.Methods The clinical data of 33 patients with Crohn's disease complicated with anal fistula who were admitted to the Sixth Hospital of Sun Yat-Sen University from June 2007 to April 2011 were retrospectively analyzed.According to the range of the disease,surgical,medical and combined treatment were applied.Surgical treatment included thread-drawing drainage,fistulectomy,incision and drainage of the perianal abscess and enterostomy.Medical treatment include 5-aminosalicylic acid,immunosuppressive therapy,glucocorticoid,tumor necrosis factor monoclonal antibodies.All the patients were followed up till August 2012.The Crohn's disease activity index (CDAI) and perianal Crohn's disease activity index (PDAI) before and after treatment were analyzed using the independent sample t test.Results Of the 33 patients,22 received surgical and medical treatment,7 received surgical treatment,and 4 received medical treatment.The anal fistula was healed in 18 patients after the initial treatment (3 of them did not receive surgical treatment),and the mean closure time of the fistula was (3.2 ± 2.6) months (range,1-12 months) ; the condition of 8 patients was improved ; 4 patients suffered from fistula persistence ; anal fistula recurrence was observed in 6 patients,and the time for anal fistula recurrence was (35 ±56)months (range,5-148 months).The preoperative CDAI and PDAI of the 33 patients were 166 ± 100 (range,7-361) and 9.2 ± 2.6 (range,5-16),and the postoperative CDAI and PDAI of the 33 patients were 83 ± 53 (range,0-212) and 2.7 ± 3.1 (range,0-11).There were significant differences between the pre-and postoperative CDAI and PDAI (t =4.20,8.92,P ＜0.05).Of the 12 patients who were failed in the initial treatment,the anal fistula was healed in 3 patients after the treatment,and the condition of the other 9 patients remained the same.Of the 6 patients with anal fistula recurrence,5 received reoperation (4 were healed and 1 remained the same after the treatment) and the fistula of 1 patient who received medical treatment was not healed.At the end of the follow up,the anal fistulas of 20 patients were healed,and 10 remained the same.Conclusions There is no standard treatment for Crohn's disease complicated with anal fistula,surgical treatment combined with medical treatment is important for the treatment of Crohn's disease complicated with anal fistula.Surgical procedures should be chosen carefully according to the condition of the patients.

AIM: To evaluate the effects of 23G vs 20G pars plana vitrectomy ( PPV ) combined with internal limiting membrane peeling, phacoemulsification and intraocular lens implantation for macular epiretinal membrane with cataract. ·METHODS: Totally 45 eyes of 45 patients with macular epiretinal membrane and cataract were enrolled in this retrospective non-randomized controlled clinical study. All eyes were treated with PPV combined with internal limiting membrane peeling, phacoemulsification and intraocular lens implantation. There were 20 eyes in 23G PPV group, and 25 eyes in 20G PPV group. The best corrected visual acuity ( BCVA ) , intraocular pressure (IOP), counting of corneal endothelial cells ( CEC) and central retinal thickness ( CRT ) were examined before surgery. BCVA results were converted to the logarithm of the minimum angle of resolution ( LogMAR ) visual acuity. All operations were performed by the same doctor. Operation time for vitrectomy and membrane peeling, average ultrasound energy ( AVE) and effective phacoemulsification time ( EPT ) were recorded. BCVA and CRT were observed postoperatively at 30d and 90d, counting of CEC was observed postoperatively at 90d. IOP was observed postoperatively at 1d and 7d. ·RESULTS:The mean operation time for vitrectomy were 12. 57± 1. 35min in 23G group and 17. 30 ± 1. 19min in 20G group. The difference was statistically significant ( t =-12. 488, P<0. 01). There were no statistical significances in operation time for membrane peeling, AVE and EPT between 23G and 20G groups ( t=-0. 68,-1. 186,-0. 737, P=0. 500, 0. 242,0. 465). On 1d after surgery, IOP in 23G group was lower than that in 20G group, the difference was statistically significant (t= -2. 345, P=0. 024). The BCVA and CRT of the two groups both improved after operations. There were no statistically significant differences between two groups in terms of IOP, BCVA, and CRT ( F = 0. 465, 1. 895, 0. 689; P = 0. 499, 0. 176, 0. 411). IOP, BCVA and CRT were significant statistical different in different time-point within each group ( F=291. 245, 103. 06, 665. 402, P<0. 01 ). Different surgical methods of 23G and 20G had interactive effects on IOP with different time points ( F = 13. 245, P<0. 01 ), but different surgeries had no interactive effects on BCVA and CRT with different time points (F=1. 212, 2. 293;P=0. 283, 0. 129). The counting CEC in 23G group was more than that in 20G group postoperatively at 90d, the difference was statistically significant (t=2. 049, P=0. 048). ·CONCLUSION: The 23G PPV combined with internal limiting membrane peeling, phacoemulsification, intraocular lens implantation for macular epiretinal membrane with cataract is effective. Compared with 20G PPV, 23G PPV has advantages in operation time for vitrectomy and counting CEC. But lower IOP is likely in 23G PPV on 1d after surgery

Background The deficiency of limbus stem cells will cause various kinds of the disorder of eye surface,in serious condition,it will lead to corneal conjunctivalization,chronic inflammation,lasting deficiency of corneal epithelium and corneal opacity accompanying long-term visual loss.Objective The aim of this study was to investigate how to establish and evaluate the successful model of total limbal stem cell deficiency (LSCD).Methods Twenty Japanese white rabbits aged 3-5 months were selected.The limbus corneal epithelium was removed from 1 mm inner to 2 mm outer of the limbus 100-150 μm in deep,and the total central epithelium and shallow stroma were simultaneously removed in the left eyes using scalpel.The experimental eyes were examined by slitlamp 2,3,4,5 weeks after operation,and the inflammatory score was performed according to corneal opacity,corneal neovascularization and corneal flurescein dye using international universal rating criteria of LSCD model.Cornea tissue was obtained in various time points mentioned above to examine the structural change and goblet cells.The expression of cytokeratin 3(CK3) in the cornea was detected using indirect immuno-fluorescence staining.Fisher Exact Probability method was used to compare the successful rate of models in different time points.Results The successful rate of models rate was 12.5％,62.5％,81.3％ and 87.5％,respectively 2,3,4,5 weeks after operation,showing a significant difference between the 3-week group and 2-week group (P =0.009).However,no significant differences were found in the model successful rate between the 3-week group and the 4-week group,the 4-week group and the 5-week group (P =0.465,0.200).There was a little difference between the 3-week group and the 5-week group(P=0.049).The mean time of achieving successful model was(3.21 ±0.80) weeks.The result of hematoxylin & eosin staining showed the edema of cornea stroma,a mass of inflammatory cells infiltration and new vessels.Goblet cells could be found by Periodic acid-Schiff staining 5 weeks after operation.Expression of CK3 was absent in the corneas of successful model eyes.Conclusions The LSCD model can be successfully established by cutting off the epithelium of limbus and central cornea,and the average time of model formation is about 3.2 weeks.

Objective: To investigate the characteristics of traditional Chinese medicine (TCM) syndromes and their elements in patients with posthepatitic cirrhosis by analyzing the relationships between signs and symptoms and biochemical parameters. Methods: A total of 440 patients with hepatitis B cirrhosis treated in Shanghai Public Health Center and Shuguang Hospital, Longhua Hospital and Central Hospital of Putuo District Affiliated to Shanghai University of Traditional Chinese Medicine during January 2002 to January 2006 were enrolled in this study. Signs and symptoms and biochemical information of patients were collected by using a self-designed questionnaire regarding the four examinations of TCM. Signs and symptoms were firstly analyzed to find the frequency of occurrence. Then, the patients were divided into two groups according to non-existent or existent sign and symptom and the correlations between the signs and symptoms which occurred most frequently and their biochemical parameters were analyzed. Results: Sixteen symptoms which occurred most frequently were fatigue, colored urine, liver palms, opaque complexion, string-like pulse, weakness at waist and knees, dry month and bitter taste in the mouth, profuse dreaminess and poor sleepiness, heaviness of limbs, abdominal distention, yellow eyes, fine pulse, impetuosity and susceptibility to rage, splenomegaly, poor appetite, and distension and fullness in the chest and hypochondrium. A previous study on syndrome differentiation of 900 patients with posthepatitic cirrhosis showed 4 syndrome patterns: internal accumulation of dampness-heat, liver-kidney yin deficiency, internal accumulation of blood stasis-heat, and liver depression and spleen deficiency. Further analysis showed that internal accumulation of dampness-heat syndrome was characterized by obvious hepatic inflammation, poor synthesis function and more asctites. Liver-kidney yin deficiency syndrome was characterized by low-level hepatic inflammation, poor synthesis function and more ascites. Internal accumulation of blood stasis-heat syndrome was characterized by low-level hepatic inflammation, poorer synthesis function, ascites and splenomegalia, and liver depression and spleen deficiency syndrome was characterized by slight hepatic inflammation, synthesis function injury, decreased internal portal vein diameter and less ascites. Conclusion: Different syndrome patterns have different pathological features, showing the complexity and polymorphism of syndrome construction.

Objective To investigate the clinical significance of WDHD1 protein expression in esophageal squamous cell carcinoma.Methods The expression of WDHD1 protein in esophageal squamous cell carcinoma,adenocarcinoma of esophagus and normal esophagus tissues by immunohistochemical MaxVision was detected.Results The expression of WDHD1 protein in the tissues of esophageal squamous cell carcinoma was significantly higher than that in the normal esophagus tissues and adenocarcinoma of esophagus (15.68 ± 3.01 vs.3.89 ± 2.98 and 4.66 ± 2.46)(P ＜ 0.05),there was no significant between normal esophagus tissues and adenocarcinoma of esophagus (P ＞ 0.05).The expression of WDHD 1 protein in the tissues of esophageal squamous cell carcinoma,high-middle differentiation tissues was higher than poorly differentiation,TNM stage Ⅲ + Ⅳ stage was higher than Ⅰ + Ⅱ stage,depth of invasion T3 + T4 was higher than T1 + T2,with lymph node metastasis tissue was higher than without,differences was statistically significant (P ＜0.05).Conclusions The higher expression of WDHD1 protein is the marker of high invasive and low histological grade of esophageal squamous cell carcinoma,and WDHD1 in esophageal squamous cell carcinomas may play an important role in development.

OBJECTVE:To investigate the clinical effect and safety of pregabalin combined with gabapentin in the treatment of central pain after cerebral infarction.METHODS:One hundred and fifty patients with central pain after cerebral infarction in our hospital from Jan.2010 to Dec.2015 in our department were randomly divided into group A,B,C,with 50 cases in each group.Group A was given Pregabalin capsule 75 mg,bid combined with Gabapentin capsule 0.1 g,tid;group B was given Pregabalin capsule 75 mg,bid;group C was given Gabapentin capsule 0.1 g,tid;3 groups were treated for 4 weeks.VAS score,NRS score,PSQI and SF-36 score were observed among 3 groups before and after treatment to evaluate clinical efficacies of 3 groups;the occurrence of ADR were recorded in 3 groups.RESULTS:The clinical total response rate of group A,B,C were separately 94.00％,74.00％,70.00％.The clinical total response rate of group A was significantly better than that of group B and C,with statistical significance (P＜0.05).After treatment,VAS score of group A,B,C were separately(3.87 ± 0.74),(5.10 ± 1.26),(5.03 ± 1.23);NRS score were separately (3.91 ± 0.88),(5.29 ± 1.25),(5.37 ± 1.30);VAS score and NRS score of group A were signifi cantly lower than group B,C and before treatment,with statistical significance (P＜0.05);PSQI score of group A,B,C were separately(4.03 ± 0.85),(5.92 ± 1.16),(5.83 ± 1.11);SF-36 score were separately (372.84 ± 73.25),(348.07 ± 60.54),(345.67 ± 59.72);PSQI score and SF-36 score of group A were significantly better than group B,C and before treatment,with statistical sig nificance (P＜0.05).There was no statistical significance in the incidence of ADR among 3 groups (P＞0.05).CONCLUSIONS:Compared with pregabalin and gabapentin alone,pregabalin combined with gabapentin in the treatment of central pain after cerebral infarction can efficiently relieve the perceived pain,improve sleep quality and daily life quality and not increase the risk of ADR;therefore,drug combination plan is recommended for patient with central pain after cerebral infarction,especially with poor effect of two single drug.

This paper introduces a method of recognizing CD4 cell microscopic images by a computer when the images are magnified about 100 times. This method reduces the intensity non-uniformities of background, enhances and segments the cells microscopic images by means of image processing. It extracts geometric feature, colour and optical features from the cells, and a Fisher classifier which can be constructed to recognize CD4 cells.
CD4 Lymphocyte Count ; methods ; CD4-Positive T-Lymphocytes ; cytology ; Image Processing, Computer-Assisted ; methods ; Pattern Recognition, Automated

A method was proposed for the separation and determination of paraffin waxes in food by HPLC-evaporative light scattering detection (ELSD). A normal-phase column was used to separate nonparaffinic and paraffinic materials without resolving the latter into individual components. The t-test method was adopted for the evaluation of mean difference between response factors of n-alkanes in paraffin waxes on ELSD detector. No mean difference was obtained between response factors, which can be used for quantitative determination of paraffin waxes in food. The determination results obtained by HPLC-ELSD were compared with those by GC-MS. The linear range for the determination of paraffin waxes was in the range from 10 to 500 mg/L with a correlation coefficient of 0.9988, and the limit of detection was 1.0 mg/L. With the spiking level of 10, 50 and 100 mg/kg, the recovery ranged from 84.6% to 105.4% and the relative standard deviation ranging from 5.4% to 7.2%. The proposed method is simple, fast and sensitive.