With the expanding scale of animal cell culture and increasing demand of biopharmaceutical,the development of serum-free medium based on cell lines and products has become a major task in the field of cell engineer.The statistical methods and tools are popular in the processes of design medium and can be used to evaluate the multiple factors and their interaction scientifically and effectively.The novel microarray and proteomic analysis can improve the performance of identifying the role of medium.The aim is to provide some idea for serum-free medium study through systematically summarizing the newly and commonly used serum-free medium study and design approaches and their characteristics.

Fed-batch culture is the predominant mode of the current animal cell culture process for many recombinant protein production. Fed-batch culture operation is mainly based on the nutrient continuous consumption and demand of cells to design continuous or semi-continuous concentration feed medium to maintain or support high-density cell growth and improve volumetric productivity of target protein in the reactor. The main methods to improve production efficiency of fed-batch cells culture include the optimization of medium design, selection and optimization of feed strategy and regulation of cell metabolism.

Objective To observe the efficacy of ealcitonin and bisphosphonates on renal osteopathy of maintenance hemodialysis (MHD)patients. Methods Forty-three MHD patients were raindomly divided into two groups: A group and B group. All the patients were routinely received oral calcium carbonate 1.0 g tid and calcitriol 0.25 μg qd. Calcitonin (20U) hypodermic injection was given three times a week additionally during hemodialysis in A group. Patients in B group received bisphosphonates 70 mg once a week based on the therapy of A group. Serum levels of intact parathyroid hormone (iPTH), calcium, phosphorus, alkaline phosphatase (AKP), bone mass density (BMD) of lumbar spine and femoral neck, and the degree of bone ache (visual analogue scale, VAS) were assessed before the therapy and 3, 6 and 12 months after treatment. The adverse reactions were recorded during treatment. Results The levels of AKP and iPTH in both two groups decreased significantly after treatment. The above values of pre-treatment and 12 months after treatment were as follows: AKP(U/L)of A group 244.05±41.99 and 148.35±27.71,of B group 245.60±40.86 and 143.40±28.03;PTH(ng/L) of A group 697.5±119.7 and 267.4±45.9,of B group 708.2±120.3 and 277.6±41.9 (all P<0.05). While the levels of calcium and phosphorus did not change obviously during treatment (P>0.05). BMD was not improved at 3, 6 mouths and became better at 12 mouths after treatment. As compared to pre-treatment, BMD of lumbar spine(g/cm2) in A group was 1.062±0.223 vs 1.202±0.251 ,in B group 1.033±0.152 vs 1.189±0.225; BMD of femoral neck (g/cm2)in A group was 0.993±0.108 vs 1.067±0.095,in B group 0.947±0.083 vs 1.018 ±0.217 (all P<0.05). The scores of VAS also decreased significantly at 3, 6, 12 months after treatment(P<0.05). No severe adverse reaction was found during the treatment. Conclusions Utilization of calcitonin and combination with bisphosphonates during bemodialysis can effectively preserve the BMD and prevent bone loss in MHD patients and is well tolerated. No significant difference of therapeutic effect is observed between using ealcitonin or combination with bisphosphonates.

Retroviral expression system which consists of retroviral vector,envelop protein vector and packaging cell line is an efficient expression system for recombinant protein.It has great potential in gene therapy and biopharmacy.Transcriptional active genome regions are the preferred targets for retrovirus integration.Furthermore,VSV-G protein enables this system a broader host range and makes virus integration more efficient.After infection of high-titer virus,high production clones can be selected through simple screening.So far,the research on retrovirus expression system has developed into application in bio-pharmacy industry.Here the composition of this system and the mechanism of virus transduction and summarize the application and prospect of retroviral expression system are introduced.

BACKGROUND: Intrapulmonary vascular abnormalities result in the right-to-left shunting and severe hypoxemia in liver transplantation candidates. Currently, a convenient, sensitive and effective method is absent to screen the intrapulmonary vascular dilatations.OBJECTIVE: To evaluate the role of contrast-enhanced echocardiography on clinical diagnosis of intrapulmonary shunting in liver transplantation candidates.DESIGN, TIME AND SETTING: The experiment, prospective controlled observation based on cases, was performed at the Hepatology Unit of the 458 Hospital of PLA (Guangzhou, Guangdong, China) from February 2004 to February 2006.PARTICIPANTS: Twenty-four consecutive liver transplantation candidates were recruited from the Hepatology Unit of the 458Hospital of PLA.METHODS: Routine examination was conducted under the condition without any regimen of vascular dilatation drugs.Contrast-enhanced echocardiography was applied to detect the prevalence of right-to-left shunting in the patients with end-stage liver disease. The microvesicle of the left ventricle in patients was qualitatively assessed by a score from 1+ to 3+. Accordingly, all patients were divided into two groups: intrapulmonary shunting and non-intrapulmonary shunting.MAIN OUTCOME MEASURES: The prevalence of right-to-left shunting and clinical characteristics of liver transplantation candidates were determined.RESULTS: Ten (41.7%) of 24 patients with positive contrast-enhanced echocardiography were proved to develop the intrapulmonary right-to-left shunting, including 6 for l+ and 4 for 2+ by left ventricle abnormality, which emerged after 6-10 cardiac cycles of right ventricle abnormality. There were no significant differences in age, gender, arterial blood gas analysis and liver function tests between the two groups (P > 0.05). Echocardiography results demonstrated that, the upper digestive tract hemorrhage,spleen thickness that indicated portal hypertension, pulmonary artery systolic pressure and Tei index were significandy higher in the patients of intrapulmonary shunting than in those of non-intrapulmonary shunting (P<0.05-0.01 ).CONCLUSION: Intrapulmonary vascular dilatation occurs frequently in liver transplantation candidates associated with intrapulmonary shunting but without hypoxemia. Contrast-enhanced echocardiography is a sensitive and non-invasive method for the early diagnosis of intrapulmonary vascular dilatation. The pathogenic cause is portal hypertension. Tel index can be used as an important parameter for evaluating right ventricular function in patients of intrapulmonary vascular dilatation.

The objective of this study was to develop research of cardiac cells to reestablish 3D tissue architecture in vitro, we performed studies using collagen membrane as three-dimensional scaffold for cardiac cells culture with the principles and methods of tissue engineering. The polymer scaffold provides a 3-D substrate for cell attachment and tissue formation. Cardiac cells isolated by enzymatic digestion from 1d old neonatal rats were seeded to three-dimensional collagen scaffolds and tissue culture plates. The morphology, beating rate and the metabolic indexes, including specific consumption rate of glucose (q(glu)) , specific production rate of lactate (q(lac)), lactate transform rate ( Y(lac/glu)), specific creatine kinase (CK) and lactate dehydrogenase (LDH) activities of cardiac cells cultured on three-dimensional collagen membrane and tissue culture plates were compared. It was found that cells shape and cells' CK and LDH activity was no differences between 3D and 2D cultures and cell beat rate on cell culture cluster was slower than those cells cultured on collagen membrane, However the cell glucose consumption and lactate yield rate of cells cultured on cluster was higher than those cells cultured on collagen membrane. After 5 days of cultivation, cardiac cells cultured on collagen membrane scaffolds organized into three-dimensional (3D) aggregates as opposed to the two-dimensional (2D) aggregates mosaic pattern seen in tissue culture plates, and spontaneous and rhythmical contractile 3D cultures in unison were visible to the naked eye and the area of synchronous contract three-dimensional (3D) aggregates reaches 80cm2. The mean value of q(glu), q(lac) and Y(lac/glu) of cultured on three-dimensional collagen scaffold was 7.37 micromol/10(6) cells/d, 2.92 micromol/10(6) cells/ d and 0.38 micromol/micromol, versus 7.59 micromol/10(6)cells/d, 3.83 micromol/10(6) cells/d and 0.51 micromol/micromol in tissue culture plates. These results demonstrate that cardiac cells immobilized on collagen membrane in 3D cultures maintain similar metabolic activity and contractile function when compared with native cardiac cells. The above results support the idea that engineered cardiac tissue can be used as a model of native tissue for studies of tissue development and function in vitro and eventually for tissue repair in vivo.
Animals ; Biocompatible Materials ; chemistry ; Cells, Cultured ; Collagen ; chemistry ; Flow Cytometry ; Immunohistochemistry ; Microscopy, Electron, Scanning ; Myocytes, Cardiac ; cytology ; ultrastructure ; Rats ; Rats, Wistar ; Tissue Engineering ; methods ; Tissue Scaffolds ; chemistry

Based on the different permeability of DNA-intercalant dyes YO-PRO-1(YP) and propidium iodide (PI) to the membrane of viable, apoptotic and necrotic cells, cell samples were stained with 4?mol/L YP and 4?g/ml PI for 10 min, and the fluorescence intensity of both YP and PI were measured by fluorometer at Ex/Em wavelength of 485/538nm and 530/590nm, respectively. The correlation between YP fluorescence intensity and the apoptotic cell number was confirmed by fluorescence microscope and linear regression(r=0.999,P

Aim To analyze the active ingredients of Chuanxiong, predict its target and signaling pathways in the treatment of tension-type headache, and clarify its therapeutic mechanism based on the principle of network pharmacology.Methods The effective active ingredients in Chuanxiong were retrieved from the Chinese herbal system pharmacology platform(TCMSP), and were performed by the ADME screen to collect the potential targets; the existing tension-type headache-related disease targets were collected through the GeneCards database.The targets corresponding to the active ingredients were intersected to obtain the common target as the key target.Cytoscape was used to construct and analyze the visual "drug-active ingredient-target-disease" network, and the String database was used to construct the PPI protein interaction network; through R language the GO function and KEGG pathway enrichment of common targets in the form of bubble graphs were analyzed.Lastly, molecular docking was used for preliminary verification.Results Finally 7 active ingredients, 105 compound targets and 2 139 tension-type headache-related target genes were obtained.There were 54 nodes in the protein interaction network.GO functional enrichment analysis yielded 215 entries, and KEGG pathway enrichment analysis yielded 68 signaling pathways.Molecular docking showed that FA, Chuanxiong quinone, sitosterol, ligustalin had strong affinity with CASP3, MAPK1, MAPK14.Conclusions It is suggested that Chuanxiong may treat tension-type headaches through anti-inflammatory, antioxidant and cytoprotective effects.

Adult ; Aged ; Bone Plates ; Bone Transplantation ; Female ; Fracture Fixation, Internal ; Humans ; Male ; Middle Aged ; Spinal Canal ; surgery ; Spinal Fusion ; Spinal Stenosis ; surgery ; Spondylolisthesis ; surgery

OBJECTIVETo investigate the effect of different kinds of mechanical stress on the mRNA expression of integrin beta1 subunit in cultured human periodontal ligament fibroblasts (hPDLF).

METHODSTo scalp and remove the periodontal ligament attached to the mid-third part of the fresh root of young premolars extracted for the cause of orthodontics. Cultured hPDLF by the method of digesting by I-type collagenase combining with tissue adhering. Then hPDLF was isolated and purified by cells passage. The sixth passage's cells were selected to be loaded. A new cyclic strain loading apparatus. Forcel four point bending device was used for mechanically loading. Cells were loaded by three levels (1000, 2000, 4000 microstrain) of tensional and compressive forces and collected at different times (0, 0.5, 1, 4, 8, 12 h) course after strain loading. The quantity of integrin beta1 mRNA in every group was analyzed by means of quantitative real-time PCR with the special primers of up- and down-regulated genes.

RESULTSDynamic mechanical forces down-regulated the expression of integrin beta1 subunit mRNA in hPDLF and the difference in groups by different magnitude, different kinds, and different time of mechanical forces loading were statistically significant. The stronger stimulated forces, the more down-regulated expression. Compression down-regulated the expression of integrin beta1 subunit mRNA more than tension did.

CONCLUSIONDynamic mechanical forces could regulate the expression of integrin beta1 subunit mRNA. The difference among all the groups by different magnitudes, different kinds, and different time of mechanical forces loading were statistically significant.