Objective:To produce a fusion protein(ICOS-Ig)between extracellular portion of human ICOS and Fc portion of mouse IgG 2a and study on its biological activity.Methods:cDNA encoding the extracellular domain of human ICOS was prepared by RT-PCR from the RNA of the stimulated human peripherial blood mononuclear cells. The Fc portion of mouse IgG2a was cloned by PCR from the vector that contains the sequence-encoding Fc portion of mouse IgG2a.Two resulting amplified PCR products were ligated into a secrection mammalian expression vector, pSecTag2/Hygro A. The recombined vector was transfected into CHO cells by lipofectamine2000.Results:RT-PCR demonstrated the integration and mRNA synthesis of fusion gene. ELISA and Western blot analysis showed the expression of ICOS-Ig and its molecular weight was between 43-66 kD, its concentration was 5-25 ?g/ml. FACS analysis assured that ICOS-Ig had ligand specific binding activity. Addition of ICOS-Ig to MLR resulted in inhibition of T-cell proliferation and IL-2,IFN-? secretion.Conclusion:The fused gene ICOS-Ig was constructed and expressed successfully. It had the bioactivity of inhibition of T cell proliferation and cytokine secretion.

Objective:To study the bioactivity of inducible costimulator Ig(ICOS-Ig) as an inhibitor of ICOS-B7RP-1 costimulatory pathway in vitro. Methods: cDNA encoding the extracellular domain of human ICOS was prepared by RT-PCR from the RNA of the stimulated human peripheral blood mononuclear cells. The Fc portion of mouse IgG2a was cloned by PCR from the vector containing the sequence-encoding Fc portion of mouse IgG2a.The above 2 PCR products were ligated into a clone vector: pGL-3-Basic. The fusion gene was then cloned and ligated into a mammalian expression vector: pcDNA4/HisMAX A. The recombined vector was transfected into CHO cells by Lipofectamine2000 and the expression of the fusion protein was identified by Western blot. The mixture lymphoproliferation reaction(MLR) of the lymphocytes derived from BALB/c and C57BL mice was used to detect the fusion protein function in vitro. Results: Western blot analysis showed the expression of fusion protein, with the molecular weight being 43 000-66 000. FACS analysis assured that expression products had ligand specific binding activity. MLR was inhibited by the fusion protein. Conclusion: The constructed recombinant fusion protein has ligand specific binding activity and can inhibit the lymphoproliferation.

Adult ; Arterio-Arterial Fistula ; therapy ; Cardiac Catheterization ; Coronary Vessels ; pathology ; Heart Ventricles ; pathology ; Humans ; Male

The fused gene (PTH-HSA) of parathyroid hormone (PTH) gene and Human Serum Albumin(HSA) gene was amplified without linker by Overlapping PCR technology. The spliced gene was clone into Pichia pastoris secretory vector pPIC9K. With the help of promoter AOX1 and mat ? signal peptide, the PTH-HSA gene was designed to secretory expression.Linearized by restriction enzyme SalI, The recombinant plasmid pPIC9K/PTH-HSA was transformed into Pichia pastoris KM71 by electroporation. The recombinant strains which were identified by G418 and PCR analysis were induced by methanol to express protein PTH-HSA. The target protein was expressed in fermentation supernatant. Western blot analysis of the fusion protein showed that the expressed fusion protein PTH-HSA had the antigenicity of HSA.adenylate cyclase assay proved that the fused protein exhibited the bioactivity to stimulate cAMP synthesis The specific activity of broth was about 318IU/ml.

This article introduces an overview of management system for conflict of interest of the European Pharmacopoeia Commission (EPC) and the United States Pharmacopoeia Convention (USP). The EPC and USP have standardized the management system for conflict of interest in drug standard work in multiple management documents, such as the Guide for the Work, Code of Practice for the Work, Form for Declaration of Interests and Confidentiality Undertaking of the EPC, bylaws, Rules and Procedures of the Council of Experts, Code of Ethics, Standards of Conduct of the USP, in order to ensure the transparency and fairness of drug standard development, improve the credibility and rigor of drug standards. This article introduces the management system for conflict of interest of the EPC and USP, providing reference for the improvement of relevant management systems of the Chinese Pharmacopoeia Commission.

Objective: To observe the clinical efficacy of umbilicus application with Chinese medicine in treating children’s chronic diarrhea due to food intolerance and the value of healthy diet education. Methods:Eighty kids with chronic diarrhea due to food intolerance were recruited from multiple centers and divided by using the random number table into a treatment group of 40 cases and a control group of 40 cases. For both groups, based on the level of serum food-specific immunoglobulin G (Ig-G), the patients were given unified diet following the healthy diet guidance: safe, alternate, and forbidden, while the treatment group was additionally given umbilicus application with Chinese medicine. The therapeutic efficacy and symptom score of traditional Chinese medicine (TCM) were observed and compared respectively after 1-week, 2-week, 4-week, and 12-week treatment. Results:The diarrhea symptoms and coupled symptoms scores dropped significantly in the two groups after treatment ( all P<0.05), and the inter-group comparisons also showed statistical significances (P<0.05). The total effective rate was 97.5% in the treatment group versus 77.5% in the control group, and the inter-group difference was statistically significant (P<0.05). Conclusion:The healthy diet guidance based on the food intolerance test is effective in helping treat diarrhea, while umbilicus application with Chinese medicine works better in treating diarrhea for kids with chronic diarrhea due to food intolerance and can significantly enhance the therapeutic efficacy.

BACKGROUND:Currently, X-ray examination is mainly used for diagnosis of nonunion. However, this method that relies only on the clinician’s experience and degree of calus mineralization has less accuracy because it is vulnerable to projection, processing conditions and subjective factors. OBJECTIVE:To establish an animal model of nonunion and to detect the variation of biochemical markers and bone mineral density. METHODS: Twenty New Zealand white rabbits were randomly divided into two groups, and bone defect and fracture models were made in the midshaft of the forearm radius, respectively. X-ray examination of the forearm, quantitative CT measurement of bone mineral density and serological test were carried out before and at 2, 3, 4, 5, 6, 7, 8, 10, 12 weeks after surgery.RESULTS AND CONCLUSION:Postoperative X-ray films showed that the in the bone defect group, a little calus formed in three rabbits at 2 weeks, the calus formed stably at 5 weeks, but there was stil no healing at 8 weeks; in the fracture group, the fracture line was blurred at 2 weeks and a large number of caluses formed at 6-8 weeks. Compared with the fracture group, the value of bone mineral density in the bone defect group began to decrease significantly at 5 weeks after surgery. Results from the serological test showed that in the bone defect group, the activity of bone-specific alkaline phosphatase increased after surgery, reached peak at 4 weeks, began to decrease at 5 weeks and became stable at 6 weeks; the activity of tartrate-resistant acid phosphatase 5b increased after surgery, peaked at 4 weeks, then decreased and stabilized basicaly; the expression of N-terminal telopeptide of type I colagen decreased significantly at 5 weeks after surgery and became stable at 6 weeks. These findings indicate that the systematic monitoring of changes in bone mineral density and biochemical indicators such as bone-specific alkaline phosphatase, tartrate-resistant acid phosphatase 5b and type I colagen N-terminal telopeptides may help to reflect the early progress of rabbit nonunion.

Objective To review our patients who underwent reconstruction of the defect after combined cranio-maxillo-facial resection in recent years.Methods From January,2005 to January,2011,88 patients underwent reconstruction of the defect after combined cranio-maxillo-facial resection.Different reconstructive techniques were used according to the defect classifications in dura,skull base bone,and facial tissues.For dural defects,no repair (37 cases),primary closure (25 cases),and dural repair (26 cases) were performed,respectively.Dural repair materials included thigh fascia lata (2 cases),temporalis fascia (2 cases),pericranium (1 case) and artificial dural patch (21 cases).Bone reconstruction of the skull base were performed in 61 patients with titanium mesh (57 cases),free iliac bone graft (1 case),free cranial bone graft (2 cases) and Medpor (1 case),respectively.Limited facial soft and hard tissue defects in 44 patients were reconstructed with temporalis system of flaps (36 cases),sternocleidomastoid myocutaneous flap (6 cases),and pterygoid muscle flap (2 cases),respectively.Extensive facial soft and hard tissue defects in 44 patients were reconstructed with free latissimus dorsi myocutaneous flap (26 cases),free pectoralis major myocutaneous flap (12 cases),free anterolateral thigh perforator flap (5 cases cases) and free anteromedial thigh perforator flap (1 case),respectively.Results The overall success rate of 88 flaps was 100％.Cerebrospinal fluid leak was found in 4 patients,wound infection was found in 2 patients,intracranial infection was found in 2 patients,respectively.Six patients with cerebrospinal fluid leak or wound infection were cured by conservative treatment.Two patients with intracranial infection were dead although they underwent salvage surgery.Overall rate of complications was 9.1％,dead rate was 2.3％.Conclusion Successful reconstruction of the defect after combined cranio-maxillo-facial resection can be achieved by watertight dural repair,bone reconstruction of the skull base and well-vascularized tissue covered.Regional flap and free tissue transfer are both preferred reconstructive technique depending on the anatomic site and the extent of the defect.

Objective:To review our patients who underwent cranio-maxillofacial resection in the recent 10 years and explore the indication of the operation. Methods:From 2003 to 2013, 116 patients underwent cranio-maxillofacial resection in our department for the treatment of tumors involving the skull base. Tumors that involved the skull base were divided into 3 types according to skull base invasions shown in the coronal planes of CT and MRI scans. Type 1 tumor was adjacent to the skull base with free bone (n=45), type 2 tumor involved the skull base with intact dura (n=30), and type 3 tumor involved dura with free brain (n=41). All patients underwent cranio-maxillofacial resection by oral and maxillofacial surgeons and neurosurgeons. The defects after cranio-maxillofacial resection were reconstructed immediately with adjacent local or regional flaps (n=62) and free vascularized flap (n=54) according to different de-fects, respectively. Results:Cranio-maxillofacial resection was successfully performed in all patients. No intraoperative complication was found. The overall success rate of soft tissue flaps and free flaps was 98.3%and 96.4%, respectively. Three patients with intracrani-al infection (n=2) and bleeding in the internal carotid artery were dead postoperatively even though they underwent salvage surgery. The overall rate of complications was 14.7%, and the dead rate was 2.6%. Recurrence or distant metastasis was found in 36 patients dur-ing the follow-up period. Conclusion: For the indication of cranio-maxillofacial resection, the balance between tumor resection and postoperative function, survival rate, and quality of life should always be considered. This technique includes the balance between func-tion and form, survival and quality of life, donor and recipient sites, and primary and secondary functions.

Objective To evaluate the effect of revascularization of the renal artery on urinary microglobulin in patients with coronaryartery disease and significant renal artery stenosis (RAS). Methods Forty-four patients with coronary artery disease and severe RAS (luminal narrowing＞70%) underwent percutaneous transluminal renal artery angioplasty (PTRA) and stenting, as well as percutaneous coronary intervention. The urine -microglobulin (α1-MG) and β2-Cmicroglobulin (β2-MG) at baseline and at 3 months after the procedures were measured. Procedural success rate, procedural complications, serum creatinine concentration at baseline and at 3-months were also recorded. Results At 3-months after the renal revascularization therapy, there was no significant change of urine α1-MG ompared with that of the baseline, however, the urine β2-MG decreased significantly 3-months after the treatment (237±187μg/L vs 377±173 μg/L, P＜0.01). Multivariate analysis revealed that persistent elevation of urine was an independent predictor of severe events (including re-admission and renal failure) after renal revascularization therapy in patients with severe RAS (OR=3.01,95% CI 1.01-8.95, P=0.036). Conclusions In patients with coronary artery disease and severe RAS, revascularization with PTRA and stenting may improve renal tubular function, but a continuous high level of urinary microglobulins after intervention is associated with more frequent re-hospitalization and renal failure.