Objective: To obtain polycystin-1 intracellular region. Methods: cDNA of polycystin-1 intracellular region was generated by PCR and then cloned into pProEX Hta, which was prokaryotic expression vector. After verified by sequencing, the recombinant was transformed into E.coli host to express and purify the fusion protein by affinity chromatography. Results: 660 bp cDNA of polycystin-1 intracellular region and 2.6×104 fusion protein were obtained. Conclusion: The fusion protein containing polycystin-1 intracellular region is obtained and is helpful for preparing anti-polycystin-1 monoclonal antibody.

Objective: To clone human augmenter of liver regeneration (hALR) cDNA, construct the recombinant expression vector, express and purify its product. Methods: The hALR cDNA was obtained by using RT PCR method with total RNA extracted from the fetal hepatic tissue. Then it was cloned into the pGEM T vector, and subcloned into expression vector pGEX 4T 3.After proved to be correct by sequencing, recombinant expression plasmid pGEX 4T 3(hALR) was transformed into E.coli BL21(DE3). The fusion protein GST hALR was produced by IPTG induction, isolated by affinity chromatography glutathione Sepharose 4B and cleaved by Thrombin. Results and Conclusion: Recombinant expression plasmid pGEX 4T 3(hALR) is constructed. The hALR is highly expressed in E.coli . The fusion protein in the plasma is resoluble. The procedure of purification and cleavage of fusion protein is easy and simple.

Objective: To clone hGLP-1 cDNA in the pBS SK(+/-)vector and construct the expression vector of pGEX-4T-3/hGLP-1cDNA to express GST-hGLP-1 fusion protein. Methods: The hGLP-1 cDNA was constructed by 6 synthetic oligonucleotides fragments, followed by the procedure of annealing and ligation with oligonucleotides fragments. The hGLP-1 cDNA was cloned into the pBS SK(+/-) vector, and was selected by α-complementation. It was confirmed by DNA sequening, then inserted into the MCS of the fusion expression vector pGEX-4T-3. The recombinant vector was transformed into E. coli TG1. Results: The recombinant plasmid DNA was digested with restrictive endonuclease BamHⅠand XhoⅠ. The result demonstrated that the hGLP-1 cDNA was successfully inserted into the pGEX-4T-3 vector and fusion protein GST-hGLP-1 had been expressed in SDS-PAGE. Conclusion： Expression of GST-hGLP-1 fusion protein can provide foundation for obtaining a larger quantity of recombinant hGLP-1 for experimental and clinic studies.

BACKGROUND: A new material of porous carbonated hydroxyapatite cement (PCHC) is discovered using foaming technique.The new material characterizes original solidification and forms porous structure.OBJECTIVE: To investigate the biomechanical effect of PCHC on repairing cancellous bone defect.METHODS: Among 30 New Land rabbits, 25 ones were considered as surgery group, whose bilateral condyles of femur was used to establish bone defect model (5.5 mm diameter and 12 mm depth). PCHC was implanted into the left side, which was considered as the experimental group, and carbonated hydroxyapatite cement (CHC) was implanted into the right side, which was considered as the control group. Another 5 rabbits were used as normal mechanical control group. Both PCHC and CHC were dip in simulated body fluid (SBF) to test mechanical intension. PCHC and CHC were then implanted into muscles of back in the surgery group. Rabbits Were-sacrificed after 2, 4, 8, 12, and 16 weeks postoperatively. Mechanical analysis was tested following intra-bone and intramuscular implantation, and compressive strength was then tested following dipping into SBF.RESULTS AND CONCLUSION: PCHC: Intra-bone mechanical strength was lower at 2 weeks, the lowest at 4 weeks, but then closed to intension of normal cancellated bone at 8 weeks, higher than normal cancellated bone at 12 weeks, and recovered to the level of normal cancellated bone at 16 weeks. CHC: Intra-bone strength was higher than that of PCHC at 2 weeks, decreased at 4 weeks, gradually increased at 8, 12, and 16 weeks, but still lower than intension of normal cancallated bone. Compressive strength of both PCHC and CHC was not changed following dipping in SBF; however, compressive strength was changed remarkably following intramuscular implantation. The results demonstrated that PCHC characterized by immobilization in situ and mechanical supporting. Thus it could be used for one kind of bone substitute material to repair the bone defect.

BACKGROUND: Carbonated hydroxyapatite cement (CHC) s a new kind of biomaterial for bone defect, which is made of powder and fluid, and can be mixed to be pasty to repair various bone defects.OBJECTIVE: To observe the improvement of vertebrae height and pain in patients with osteoporosis vertebral compression fracture (VCF) after vertebroplasty by using a new kind of bone graft biomaterial, taking CHC as the filling material to reinforce the vertebral body.DESIGN: A contrast observation trial taking patients as subjects.SETTING: Department of Orthopaedics, General Hospital of Chinese PLA.PARTICIPANTS: Totally 34 patients with thoracic or lumbar osteoporosis VCF who received the treatment in the Department of Orthopaedics, General Hospital of Chinese PLA between October 2000 and August 2003. Inclusive criteria: ①Definite diagnosis by CT; ② Informed consents were obtained from the patients. Exclusive criteria: The patients with osteoporosis vertebral compression fractures who suffered vertebral posterior wall fracture. There were 6 males and 28 females, and they were aged (72±13)years; Among the patients, 27 were diagnosed as postmenopausal osteoporosis, 1 as cortical hormone-induced osteoporosis and 6 male patients weresenile osteoporosis.METHODS: ①All the patients were randomly divided into two groups: Experimental group (n =23) and control group (n=11). All the patients were performed percutaneous operation with local anesthenia. All cases were performed percutaneous operation under local anesthesia. Under the C-arm monitored, one side pedicle puncture was performed to enter the anterior column of the involved VCF. Patients of the experimental group were filled with CHC. Patients of control group were filled with polymethyl Methacrylate (PMMA) with the same way. ② Referred to McGill-Melzack scoring. Among the scale 0-100 mm (0 was no pain, 100 was acute pain), the value indicated the painful intensity and mental assault degree. ＜ 30 scores indicated good, 30-40 basically satisfied and ≥ 50 poor .③ Referred to the method from Lee et al, the preoperative height (A1) and postoperative height (A2) of compression fracture position of VCF were measured according to the lateral X-ray film. At the same time, the upper vertebral height (A3) and the inferior vertebral height (A4) were measured at the same position. The original height (A) of the involved vertebra was calculated as (A)= (A3+A4)/2,and the preoperative vertebral compression rate =(A-A1 )/A, the postoperative vertebral compression rate =(A-A2)/A, the restoring rate = (the preoperative vertebral compression rate-the postoperative vertebral compression rate)/the preoperative vertebral compression rate. ④ The wounds of the patients were observed after operation. The levels of blood routine, serum calcium and serum phosphorus were detected before, one day and one week after operation. MAIN OUTCOME MEASURES: ① Preoperative and postoperative VAS scoring. ② The vertebral compression rate and restoring rate. ③ Wounds were observed after operation. The blood routine, the serum calcium and serum phosphorus were detected before, one day and one week after operation.RESULTS: Totally 34 patients were involved in the result analysis. ①The preoperative visual analogue scale (VAS) score of experimental group were (91.5±21.7) points, and the postoperative ones were (44.5±27.2) points. The difference of VAS score reduced gradually along with the postoperative time. There was no difference of VAS score between experimental group and the control group 4 weeks after operation. ② The biocompatibility of CHC in the vertebral body was fine. The vertebral compression rate of experimental group was recovered from (43.1±21.4)％ preoperatively to (27.3± 18.5)％ postoperatively. The rate of restored heights was (27.3±18.5)％. ③ All patients obtained Ⅰ stage wound healing, and none of them had infection, inflammatory secretion and nervous symptom. There were no differences in blood routine test, serum calcium, serum phosphorus between patients in two groups. One case filled by PMMA and two cases filled by CHC presented leakage, and none had nervous symptom.CONCLUSION: As the filling materials for vertebropalsty, CHC can restore the vertebral heights and relieve pain safely and effectively, however, its efficacy to relieve pain is not significant as PMMA in the short term.

BACKGROUND: Carbonated hydroxyapatite cement is a new type material for skeletal repair and hydroxyapatites have been applied in the clinical treatment of skeletal defect.OBJECTIVE: To observe the effective characteristics of carbonated hydroxyapatite cement on repair of skeletal defect by animal experiment.DESIGN: Paired design, self-controlled and verified experiment was applied in the research.SETTING: Orthopedic Institute and Animal Experimental Center of Chinese PLA.MATERIALS: The experiment was performed in Orthopedic Institute and Animal Experimental Center of Chinese PLA from May 2002 to January 2003, in which, 10 healthy adult male mongrel dogs were applied, body mass weighted varied from 20 to 22 kg.METHODS: Animal model of skeletal defect was prepared on proximal ends of humeri of 10 mongrel dogs thydroxyapatitet were randomized into experimental side and control side. Ceramics repair of skeletal defect was done by carbonated hydroxyapatite cement and high-temperature sintered hydroxyapatite respectively. The animals were sacrificed on the 5th day, 4th, 8th, 12th and 16th weeks successively after operation. The repair effects were performed with X-ray and histological observation.staining.Results of stereomicroscopic and X-ray observations on bilateral skeletal defect: Osseointegration with carbonated hydroxyapatite cement was tight on the experimental side and the interface became unclear gradually with time lasting. The interface between hydroxyapatite and bone was still clear on the and eosin staining and thydroxyapatitet of ground bone with Gimsa staining:On the 8th week on the experimental side, the new bone grew into carbonated hydroxyapatite cement, on the 16th week, the two parts were intermixed and integrated and the bone island was formed around newly generated vessels in carbonated hydroxyapatite cement. On the control side, hydroxyapatite still maintained integrated and the bone interface was clear between hydroxyapatite and bone. On the 16th week, the aggradation of newly generated bone presented on hydroxyapatite surface.CONCLUSION: Carbonated hydroxyapatite cement possesses solidification property in situ, biocompatibility and osseous conductive activity. It is the satisfactory new type material for repair of skeletal defect.

COVID-19 is caused by a novel coronavirus-severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), which has being spreading around the world, posing a serious threat to human health and lives. Neutralizing antibodies and small molecule inhibitors for virus replication cycle are the main antiviral treatment for novel coronavirus recommended in China. To further promote the rational use of antiviral therapy in clinical practice, the National Center for Infectious Diseases (Beijing Ditan Hospital Capital Medical University and the First Affiliated Hospital, Zhejiang University School of Medicine) invited experts in fields of infectious diseases, respiratory and intensive care to develop an Expert Consensus on Antiviral Therapy of COVID-19 based on the Diagnosis and Treatment Guideline for COVID-19 ( trial version 10) and experiences in the diagnosis and treatment of COVID-19 in China. The consensus is concise, practical and highly operable, hopefully it would improve the understanding of antiviral therapy for clinicians and provide suggestions for standardized medication in treatment of COVID-19.

Objective: To investigate the clinical effect and safety of long-acting pegylated interferon-α-2b (Peg-IFN-α-2b) (Y shape, 40 kD) injection (180 μg/week) in the treatment of HBeAg-positive chronic hepatitis B (CHB) patients, with standard-dose Peg-IFN-α-2a as positive control. Methods: This study was a multicenter, randomized, open-label, and positive-controlled phase III clinical trial. Eligible HBeAg-positive CHB patients were screened out and randomized to Peg-IFN-α-2b (Y shape, 40 kD) trial group and Peg-IFN-α-2a control group at a ratio of 2:1. The course of treatment was 48 weeks and the patients were followed up for 24 weeks after drug withdrawal. Plasma samples were collected at screening, baseline, and 12, 24, 36, 48, 60, and 72 weeks for centralized detection. COBAS® Ampliprep/COBAS® TaqMan® HBV Test was used to measure HBV DNA level by quantitative real-time PCR. Electrochemiluminescence immunoassay with Elecsys kit was used to measure HBV markers (HBsAg, anti-HBs, HBeAg, anti-HBe). Adverse events were recorded in detail. The primary outcome measure was HBeAg seroconversion rate after the 24-week follow-up, and non-inferiority was also tested. The difference in HBeAg seroconversion rate after treatment between the trial group and the control group and two-sided confidence interval (CI) were calculated, and non-inferiority was demonstrated if the lower limit of 95% CI was > -10%. The t-test, chi-square test, or rank sum test was used according to the types and features of data. Results: A total of 855 HBeAg-positive CHB patients were enrolled and 820 of them received treatment (538 in the trial group and 282 in the control group). The data of the full analysis set showed that HBeAg seroconversion rate at week 72 was 27.32% in the trial group and 22.70% in the control group with a rate difference of 4.63% (95% CI -1.54% to 10.80%, P = 0.1493). The data of the per-protocol set showed that HBeAg seroconversion rate at week 72 was 30.75% in the trial group and 27.14% in the control group with a rate difference of 3.61% (95% CI -3.87% to 11.09%, P = 0.3436). 95% CI met the non-inferiority criteria, and the trial group was non-inferior to the control group. The two groups had similar incidence rates of adverse events, serious adverse events, and common adverse events. Conclusion In Peg-IFN-α regimen for HBeAg-positive CHB patients, the new drug Peg-IFN-α-2b (Y shape, 40 kD) has comparable effect and safety to the control drug Peg-IFN-α-2a.

Objective: To investigate the efficacy and safety of the new investigational drug pegylated interferon α-2b (Peg-IFN-α-2b) (Y shape, 40 kD) injection (180 µg/week) combined with ribavirin in the treatment of patients with genotype 1/6 chronic hepatitis C (CHC), with standard-dose Peg-IFN-α-2a combined with ribavirin as a positive control. Methods: A multicenter, randomized, open-label, and positive-controlled phase III clinical trial was performed. Eligible patients with genotype 1/6 CHC were screened out and randomly divided into Peg-IFN-α-2b(Y shape, 40kD) group and Peg-IFN-α-2a group at a ratio of 2:1. The patients in both groups were given oral ribavirin for 48 weeks in addition and then followed up for 24 weeks after drug withdrawal. Abbott Real Time HCV Genotype II was used to determine HCV genotype, and Cobas TaqMan quantitative real-time PCR was used to measure HCV RNA level at 0, 4, 12, 24, 48, and 72 weeks. Adverse events were recorded in detail. The primary efficacy endpoint was sustained virological response (SVR), and a non-inferiority test was also performed. Results: A total of 561 patients with genotype 1/6 CHC were enrolled, among whom 529 received treatment; 90.9% of these patients had genotype 1 CHC. The data of the full analysis set showed that SVR rate was 69.80% (95% CI 65.00%-74.60%) in the trial group and 74.16% (95% CI 67.73%-80.59%) in the control group (P = 0.297 0). The data of the per protocol set (PPS) showed that SVR rate was 80.63% (95% CI 76.04%-85.23%) in the trial group and 81.33% (95% CI 75.10%-87.57%) in the control group (P = 0.849 8), and the 95% CI of rate difference conformed to the non-inferiority standard. The analysis of the PPS population showed that of all subjects, 47.9% achieved rapid virologic response, with a positive predictive value of 93.8%. The incidence rate of adverse events was 96.30% in the trial group and 94.94% in the control group, and the incidence rate of serious adverse events was 5.13% in the trail group and 5.06% in the control group. Conclusion In the regimen of Peg-IFN-α combined with ribavirin for the treatment of genotype 1/6 CHC, the new investigational drug Peg-IFN-α-2b(Y shape, 40 kD) has comparable clinical effect and safety to the control drug Peg-IFN-α-2a.