Objective To establish an efficient, simple, low cytotoxicity and cheap transfaction system. Methods We have used the cationic polymer polyethylenimine(PEI) to study transient transfection in MCF-7 cells by testing different conditions, including cell concentrations, DNA concentrations, the ratio of PEI nitrogen to DNA phosphate(PEI-N∶DNA-P) and the time of cells grown in serum-free culture together with PEI-DNA complex. Results The optimized cell concentrations were 2?10 5 cells seeded per well in 24-well dishes 18-24?h before transfection. The DNA concentrations and ratio of PEI-N∶DNA-P are very important for optimal transfection and they affect each other. For 1??g DNA per well, the optimal PEI-N∶DNA-P is about 33∶1, however, as for 4??g DNA, it is 9∶1. The best time of cells grown in serum-free culture together with PEI-DNA complex is about 5-7?h.Conclusion With optimized conditions, we can establish an efficient, simple, low cytotoxicity and cheap transfection system by using PEI.

BACKGROUND: Immunoloregulation of mesenchymal stem cells (MSCs) is commonly approved. Previous studies have confirmed the ability of Flk-1~+ bone marrow MSCs (BMSCs) to inhibit T/B lymphocyte proliferation in vitro. OBJECTIVE: To investigate the therapeutic effect of Flk-1~+ BMSCs in collagen-induced arthritis mice.METHODS: A total of 18 healthy male DBA-1(H-2K~q) mice aged 10 weeks were randomly divided into 3 groups. All the mice were injected at the base of the tail with bovine type II collagen (CII), and received a booster injection of CII on day 21 to establish the CIA mice model. DBA-1(H-2K~q)mouse Flk-1~+ BMSCs were isolated in vitro by the density gradient centrifugation and adherence screening. Following initial immunity, mice in the cell transplantation group were infused with Flk-1~+ BMSCs (1-2)×106 cells/mouse via the caudal vein. Mice in the cell transplantation group were injected with the same volume of Flk-1~+ BMSCs during booster. Mice in the model control group were injected with an equal volume of saline 0 or 21 days following initial immunity. Following initial immunity and booster immunization, claw pad thickening and clinical score were observed, changes of joint pathology and dynamic changes in serum factor mass concentration were determined in mice. RESULTS AND CONCLUSION: Compared with the model control group, no significant difference in claw pad thickening and mean clinical score was detected in the cell transplantation group following initial immunity (P > 0.05), with the presence of obvious damage to synovial membrane and inflammatory cell infiltration. Mass concentration of each serum cell factor was similar. The claw pad was significantly thickened (P < 0.01), mean clinical score reached 3.35 points, with severe damage to synovial membrane, proliferation of blood capillary in the cell transplantation group following booster immunization. Interleukin-6 levels were greatly increased at day 28 following initial immunity (P < 0.1), but decreased at day 35 following initial immunity (P < 0.1). Results indicated that in the collagen-induced arthritis mouse models, Flk-1~+ BMSC transplantation did not obtain prospective therapeutic efficacy, but aggravation of arthritis was observed in the cell transplantation group following booster immunization. Upregulation of interleukin-6 concentration could aggravate the behavior symptom of rheumatoid arthritis mice.

Objective To explore the feasibility and the value of ultrasound transient elastography in the estimation of intraocular pressure.Methods Thirty-two cases of glancomatous patients' and twenty-eight healthy subjects'eyeball were examined by using ultrasound transient elastography,meanwhile using Photoshop image analysis software to quantitative analysis of blue gradation ratio of the front wall tissue of eyeball,then intraocular pressure were examined by NCT.The difference of quantitative data of blue gradation ratio of the front wall tissue of eyeball between glancomatous patients and healthy subjects were compared.The relationship of quantitative value of blue gradation ratio and intraocular pressure were established.Results The blue gradation ratio of the front wall tissue of eyeball in glancomatous patients were 0.37± 0.03,while in healthy subjects were 0.33 ± 0.02,there was a significant difference between glancomatous patients and healthy subjects (P < 0.01),and there was a linear relation between blue gradation ratio and intraocular pressure(r = 0.424,P < 0.05).Conclusions The ultrasound transient elastography can be used in estimation of intra-ocular pressure in glancomatous patients,and there is a linear relation in quantitative data between blue gradation ratio and intraocular pressure.

Objective To observe and compare the changes of weight,blood pressure,glycemia, blood lipid,serum uric acid,insulin sensitivity and insulin function of laparoscopic sleeve gastrectomy (LSG)for the treatment of type 2 diabetes mellitus(T2DM)with obesity,and analysed mechanism of meta-bolic surgery.Methods Changes of weight,blood pressure,glycemia,blood lipid ,serum uric acid,in-sulin sensitivity and insulin function of 8 type 2 diabetes mellitus(T2DM)with obesity patients undergoing LSG were retrospectively analyzed.Weight,BMI,EWL%,average systolic and diastolic pressure were measured 1,2,3,6 months and fasting plasma glucose (FPG),glycosylated hemoglobin (HbAl),insulin sensitivity parameter:insulin resistance index(HOMA-IR),insulin sensitivity index(ISI),insulin area un-der the curve (AUCI),insulin function parameter:glucose disposition index(DI)were measured 1,6 months after operation and compared with preoperative levels.Meanwhile medication change was observed af-ter operation and compared with preoperation.Results ⑴Weight ,BMI at 1,2,3,and 6 months after surgery showed a decreasing trend and EWL% showed a decreasing trend over time .⑵ FBG、HbA1c showed a decreasing trend after operation.Blood lipid,uric acid of 3 patients with dyslipidemia and hyperu-ricemia decreased to normal 1 month after operation.⑶ HOMA-IR,AUCI except for 1 patient showed a de-creasing trend and ISI was contrary after operation.DI ,trends were different.⑷7 patients stopped all an-tidiabetic drugs 6 months after operation,1 patient injected insulin with good glycemic control.Blood pres-sure of 5 patients with hypertension was normal without antihypertensive drugs 6 months after operation. Conclusions LSG procedure can significantly increase insulin sensitivity of T2DM with obesity,thereby improve glycemia,cholesterol,uric acid and further promot weight loss.

Objective To explore the value of transthoracic echocardiography (TTE) and transesophageal echocardiography (TEE) in diagnosing adult multiple atrial septal defect (MASD). Methods Thirty adult patients with MASD were examined with TTE, 25 patients were examined also with TEE, 26 patients were examined with cardiac catheterization as well. Transcatheter closure of MASD was performed in 20 patients and succeeded in 18, while open-chest operation was performed in 4 patients. Results Foramen secundum atrial septal defect was diagnosed with both TTE and TEE with an accuracy rate of 60.00% (18/30) and 96.00% (24/25), respectively. The main color Doppler flow imaging (CDFI) feature of adult MASD was multiple colorful left-to-right shunt signals through the atrial septal designated, i.e. colander sign of CDFI. Conclusion TTE has some difficulties and TEE has specific value in diagnosing adult MASD. TTE can be used before open-chest operation. TEE is necessary before transcatheter occlusion to make sure of the amount and location of atrial septal defect.

Objective To investigate the feasibility and safety of transcatheter closure of coronary artery fistula (CAF)with Amplatzer vascular PlugⅡ(AVPⅡ)in pediatric patients.Methods Between June 2012 and October 2015,5 children aged 0.9 to 7.0 years old and weighted 10 to 21 kg with CAF were admitted to the Department of Pediatric Cardiology in Shandong Provincial Hospital Affiliated to Shandong University.Aortic root angiography was used first to confirm the origin,shape,branches,drainage and the diameter of the orifice of CAF by deploying the pigtail catheter.The AVPⅡwas retrogradely deployed into targeted artery through guiding catheter and aortic angiography was performed before releasing the plug.Results All the 5 children underwent transcatheter closure by AVPⅡsuccessful-ly.Two cases were involved with right coronary -right ventricular fistula,1 case of left anterior descending coronary -right ventricular fistula (residual fistula after surgical repair),and 1 case of left circumflex coronary -left atrial fistula. Four children had a single fistula,and 1 case had double fistulas.The diameter of the orifice ranged from 2.00 to 5.96 mm,and the selected occluders from 8 to 14 mm.The ratio of diameter of occluder to fistula orifice ranged from 2.3 to 3.4.All the patients were followed up for 4 to 44 months.Two patients developed instant minor and modera-te aortic re-gurgitation.No other complications such as thrombosis,embolization,residual shunt,arrhythmia,coronary dissection or perforation occurred.Conclusions Transcatheter closure of CAF by AVPⅡin pediatric patients is feasible and safe. Aortic regurgitation should be noted,especially during the procedure.

Objective To explore the clinical application and value of percutaneous transhepatic gallbladder drainage (PTGD)under CT guide in high-risk emergency.Methods In all 57 old patients with high-risk acute cholecystitis in emergency,cuff-PTGD in 39 was performed and fractional step PTGD in 18 was also used.Results PTGD was successfully in all patients.After PTGD,except for 1 patient died of severe cardiac insufficiency,the abdominal pain and fever were alleviated during 72 hours,and the complications was not demonstrated.Conclusion As a safe,noninvasive and accurate method,CT-guided PTGD may relieve symptoms quickly,reduce the mortality and improve the treatment for some old patients with high risk acute cholecystitis.

Objective To develop a double-antibody sandwich ELISA for determining the concentration of nucleoprotein (NP) of rabies virus in various products of rabies vaccine.Methods The purified rabies antibodies from a rabbit were used to coat microwell plates. Horseradish peroxidase-conjugated anti-NP monoclonal antibody was used to probe the NP bound to coated antibodies.The assay was used to quantitatively determine the concentration of NP of rabies virus.Results The results showed that the coefficient of linear correlation was higher than 0.97.The optimal linear range was 0.000625~0.01 IU/ml and the detection limit was 0.000625 IU/ml. The recovery rate was 102~109% and the coefficient of variation was only 7.2%~9.4%.No cross reactions were observed with bovine serum,bovine serum albumin,ovalbumin,refined solution of influenza vaccine,encephalitis B vaccine,and hepatitis A vaccine.Conclusions The results indicated that the assay is specific,sensitive,accurate,reproducible,and stable,and could be suitable for quantitative determination of different rabies vaccine's processes products.

Objective To study the application effect of fibrin sealant(FS) during operation for late-stage portal hypertension caused by schistosomiasis.Methods From Jun,2003 to Jun,2005,92 cases of(late-stage) portal hypertension caused by schistosomiasis treated by portal-systemic disconn-ection(PSD)(operation) were divided into two groups,namely PSD group and PSD+FS group.The early complications such as fever and exudate in splenic fossa,as well as long term complications such as recurrent bleeding,(hypertensive) gastropathy were compared in the 2 groups,and encephalopathy were compared between 64 cases of patients with FS and 28 cases of patients without FS.Results The patients undergoing PSD+FS showed decreased the exudate in splenic fossa and fever(P0 05).Conclusions FS application during operation is effective in attenuating early and long term operative complications of portal hypertension caused by(schistosomiasis),and improving outcome of surgical treatment.

Objective:To evaluate the effect of transcutaneous electrical acupoint stimulation (TEAS) on emergence agitation in the patients undergoing thoracoscopic surgery.Methods:Eighty patients of both sexes, aged 25-64 yr, with body mass index of 18-30 kg/m 2, of American Society of Anesthesiologists physical status Ⅱ or Ⅲ, undergoing elective thoracoscopic surgery under general anesthesia, were divided into 2 groups ( n=40 each) by a random number table method: TEAS group (group T) and control group (group C). In group T, the Hegu (L14), Zusanli (ST36) and Sanyinjiao (SP6) were stimulated starting from 30 min before induction of anesthesia until the end of operation, with a frequency of 2/100 Hz and disperse-dense waves.The intensity of stimulation was the maximum current that patients could tolerate.In group C, stimulating electrodes were placed at the same acupoints before induction, but no electrical stimulation was applied.Ramsay sedation score and VAS score were recorded.and emergence agitation was assessed using RSS agitation score.Blood samples from the median cubital vein or internal jugular vein were collected before anesthesia induction (T 1), at the end of surgery (T 2), immediately after removal of tracheal tube (T 3) and at 15 min after removal of tracheal tube (T 4) for determination of serum interleukin-6 (IL-6) and IL-10 concentrations by enzyme-linked immunosorbent assay.The intraoperative consumption of propofol, remifentanil and sufentanil was recorded.Adverse reactions such as tachycardia, bradycardia, hypertension, hypotension, respiratory depression, delay of awakening and nausea and vomiting were recorded after operation. Results:Compared with group C, the amount of intraoperative remifentanil consumed was significantly decreased, serum IL-6 concentrations were decreased, and the serum IL-10 concentration was increased T 2-4, Ramsay sedate score was increased, and the incidence of agitation was decreased in group T ( P<0.05). There was no significant difference in VAS score and incidence of postoperative adverse reactions between the two groups ( P>0.05). Conclusions:TEAS can reduce the development of emergence agitation in the patients undergoing thoracoscopic surgery, which is related to inhibition of inflammatory responses.