中药增骨Ⅰ号对体外成纤维细胞的影响

为了探讨增骨Ⅰ 号的作用机制 ,进行了体外成纤维细胞的实验。采用酶消化法将人包皮组织进行成纤维细胞的分离 ,纯化及培养 ,并测定骨钙素、细胞钙含量。结果显示成纤维细胞在增骨Ⅰ 号组增殖迅速 ,其OD值明显高于对照组 (P <0 .0 5 ) ,但反映成骨表型表达的骨钙素及细胞钙含量未见明显增加。表明增骨 Ⅰ 号仅有激活成纤维细胞的作用 ,本实验结果为序贯疗法应用增骨 Ⅰ 号作为骨重建中的激活药物提供了理论依据。

The Antiviral Action of Polyhexamethylene Guanidine Derivatives

Although the PHMG （polyhexamethylene guanidine） and other oligomer guanidines are known as highly efficient biocides against a broad spectrum of microorganisms and eukaryotic cells, the cell protection by PHMG derivatives has been established firstly in this study. The antiviral protection was also exhibited after 15 min pretreatment of different cell cultures with low-concentration of PHMG salts. Monolayers of the continuous bovine tracheal cells culture （TCC） and primary culture of chicken embryo fibroblasts （FCE） were treated with aqueous solutions of PHMG chloride salts or PHMG succinate. The molecules of PHMG polycation adhered to the plasma membrane of the cells tested as they were treated with PHMG for 15-30 min. The viral material was added to the cell cultures after the wash-out carried out twice to rid of unbound PHMG. The viruses of Equine herpesvirus type 1, Rhinotracheitis infectious bovine and Equine infectious anemia virus were used. The protective effect from the cytopathic action of herpes and retroviruses was exhibited after 15 min pretreatment of cell monolayer with PHMG chloride at the TCC concentrations of 10^-3 - 10^-2% and FCE concentrations of 10^-5 - 10^-4%. The unique antiviral properties of PHMG salts represented in our research had never been shown before.

Surface Modification of Anionic Polyurethane with Silk Fibroin Peptide and Its Effects on the Culture of Fibroblasts

The surface modification of the anionic polyurethane(APU)film was carried out by immersing it in silk fibroin peptide(SFP)solution for 12 h and then treating with low temperature plasma glow discharge.The physical properties and moisture permeability of modified films were examined.The results showed that SFP-modified APU films had better moisture permeability than oleophilic polyurethane,as well as modified APU films kept good flexibility.Modified APU films could overcome rigid and brittle weaks of silk fibroin films.The morphology of SFP on the APU film was corpuscular aggregations.The water-contact angle measurement indicated that the change of hydrophilicity and the element chemical analysis suggested that the SFP-modified film surface was enriched with nitrogen atoms.The biocompatibility of APU films may be improved due to the change of surface components.Cell viability and proliferation of rat embryo dermal fibroblasts seeded on control films,APU films and SFP-modified APU films were evaluated by MTT assay and viable cell counts,respectively.The results indicated that the APU film modified by SFP protein showed the proliferation of fibroblasts on the film,and that the compound interface had good stability in the air.Results also showed that presoaking treatment for APU films was effective to accomplish the goal of surface modification.

Mechanistic Insights of Cells in Porous Scaffolds via Integrated Culture Technologies

This research aimed to combine 3 cell and tissue culture technologies to obtain mechanistic insights of cells in porous scaffolds. When cultivated on 2D （2-dimensional） surfaces, HDFs （human dermal fibroblasts） behaved individually and had no strict requirement on seeding density for proliferation; while HaCat cells relied heavily on initial densities for proliferation and colony formation, which was facilitated when co-cultured with HDFs. Experiments using a 3D CCIS （3-dimensional cell culture and imaging system） indicated that HDFs colonised openpores of varying sizes （125-420 ~tm） on modular substrates via bridge structures; while HaCat cells formed aperture structures and only colonised small pores （125 txm）. When co-cultured, HDFs not only facilitated HaCat attachment on the substrates, but also coordinated with HaCat cells to colonise open pores of varying sizes via bridge and aperture structures. Based on these observations, a 2-stage strategy for the culture of HDFs and HaCat cells on porous scaffolds was proposed and applied successfully on a cellulosic scaffold. This research demonstrated that cell colonisation in scaffolds was dependent on multiple factors; while the integrated 2D＆3D culture technologies and the 3D CCIS was an effective and efficient approach to obtain mechanistic insights of their influences on tissue regeneration.

Permanent myopathy caused by mutation of SCN4A Metl592Val:Observation on myogenesis in vitro and on effect of basic fibroblast growth factor on the muscle

Objective The present study is to observe in vitro the proliferation ability of the muscle cells from permanent myopathy （PM） patients of nomokalaemic periodic paralysis （normKPP）, which is caused by mutations of Metl592Val in the skeletal muscle voltage gated sodium channel （SCN4A） gene on chromosome 17q23.1. We also evaluate the possible effect of the foreign basic fibroblast growth factor （bFGF） in preventing and curing PM. Methods The gastrocnemius muscle cells were taken from two male patients with PM of the same Chinese family with Metl592Val mutation of SCN4A, determined by gene screening. Four male patients suffering from the skeletal injury without PM were taken as control. All preparations were protogenerationally cultured in vitro. Proliferation of the cultured preparations was measured by MTT. Activities of the lactic dehydrogenase （LDH）, creatine kinase （CK）, and protein content in these cells were also detected. The effects of bFGF with different doses （10 ng/mL, 20 ng/mL, 40 ng/mL, 80 ng/mL, 120 ng/mL and 160 ng/mL） on the above mentioned parameters were also evaluated. Results Cells from both PM and control subjects were successfully cultured in vitro. The cultivation of the muscle cells from PM patients in vitro was not yet seen. Results indicated the obvious stimulation of bFGF on cell proliferation, activities of LDH and CK, protein synthesis, in a dose dependent manner. The optimal dose of bFGF was 120 ng/mL （P〈0.05）, beyond which greater dose caused a less effect. The effect of bFGF on 160 ng/mL was stronger than that on 80 ng/mL, but there was no significant difference （P〉0.05）. Conclusion Myoblastic cells from patients with PM had a weaker ability of developing into the myotubules, thus they were unable to perform effective regeneration, which resulted in a progressive necrosis. The exogenous bFGF could promote the division and proliferation of the muscle cells in vitro. These results shield a light on bFGF＇s potential role in preventing and treating PM.

Reciprocal action between BMP-2 and BMP-3 in cultured fibroblast in vitro

Objective: To explore reciprocal action between BMP 2 (bone morphogenetic protein 2) and BMP 3 for better understanding of the mechanism of BMP during bone fracture union. Methods: rhBMP 2 was added into the cultured fibroblasts with the concentration of 1 200 ng/ml. The expression of BMP 3 in fibroblasts was detected by immunohistochemistry. Eukaryotic expression vector pcDNA3 BMP 3 was transfected into the fibroblasts. After the effective expression of BMP 3 was identified, BMP 2 was also detected by immunohistochemistry in BMP 3 expression cells. The fibroblasts transfected with empty vector pcDNA3 were used as the control. Results: Exogenous rhBMP 2 could promote the expression of BMP 3 in fibroblasts. BMP 3 also could be detected in these cells.Conclusions: BMP 2 and BMP 3 could reciprocally adjust the expression in fibroblasts.