The effects of > 10 kD component of serum-free conditioned medium from murine bone marrow-derived endothelial cell line on the formation of CFU-GM and HPP-CFC were investigated. We found that > 10 kD component alone could stimulate CFU-GM colony formation in dose-dependent. Above- 10 kD component supplemented with GM-CSF not only resulted in significant increase of CFU-GM as compared with > 10 kD or GM-CSF alone, but also could stimulate HPP-CFC formation. The effect of > 10 kD component on the formation of HPP-CFC was also in dose-dependent. The component above- 10 kD combined with IL-3, IL-6, EPO, SCF or GM-CSF further enhanced HPP-CFC colony formation. The results suggest that > 10 kD component may be used as the substitutes of hematopoietic growth factors to stimulate the formation of CFU-GM and HPP-CFC.
Objective To study the changes of the microenvironment of bone marrow hematopiesis after exposure to gamma radiation. Methods Immunohistochemistry and electrophoretic mobility shift assay (EMSA) were used. Results The expression of nuclear factor kappa B(NFκB) protein in the bone marrow stromal cells (BMSCs) was elevated after exposure to 8 Gy gamma rays radiation determined with immunohistochemistry. The activity of NFκB in the BMSCs was significantly increased after irradia tion determined with EMSA and it reached the peak 4 hours after irradiation. Conclusion Our findings suggest that nuclear factor kappa B in the BMSCs is involved in the protection of BMSCs and in the reco very of hematopoiesis after exposure to radiation.
Objective To investigate the effects of combined radiation burn injury on bone marrow stromal cells in mice. Methods Mice were treated with 6.0 GY ray radiation and 15% (TBSA) Ⅲ° burn. The culture of CFU GM and bone marrow stromal adherent cells were performed. Cell cycle, DNA content and the expression of adhesion molecules were detected by flow cytometry. Results ①The adherent capacity of bone marrow stromal cells and the numbers of CFU F after combined radiation burn injury were significant decreased. ②Three days later, cultured stromal adherent cell inhibited the growth of CFU GM, but when the agar layer existed that inhibition were weaken or disappear. ③The cell proportion of G 0 and G 1 was decreased, and the proportion of S and G 2+M was increased after combined injury. ④The DNA content in the bone marrow stromal cells was significantly lower in the combined injury group after 3 days. ⑤The expression of cell adhesion molecules including vascular cell adhesion molecule 1 (VCAM 1), fibronection (Fn), laminin (Ln), and collagen type Ⅳ (Col Ⅳ) on bone marrow stromal cell were decreased in combined radiation burn injury. Conclusions The damage of bone marrow stromal cells in bone marrow hematopoietic inductive microenvironment might be one of the most important factors in hematopoietic disorder in combined radiation burn injury.
The rapid and highly-efficient gene replacing is prerequisite for studying E.coli gene engineering strains.The Red recombinant system was combined with genetic engineering techniques to substitute chloromycetin-resistant gene from genetic engineering strain and simultaneously maintain galactose-screening marker.We have successfully confirmed that chloromycetin-resistant gene could be replaced by employing Red recombinant system and simultaneously retained other characteristic of strain,thus providing important insights into genetic engineering strain with new genetic characteristics and gene function research.
The fused gene(F-HN) of the newcastle disease virus F gene and HN gene were 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 α signal peptide,the F-HN gene was designed to secretory expression.Linearized by restriction enzyme SacⅠ,The recombinant plasmid pPIC9K-F-HN was transformed into Pichia pastoris GS115 by electroporation.The recombinant strains which were identified by G418 and PCR analysis were induced by methanol to express protein F-HN.The target protein was expressed in fermentation supernatant. Western blot analysis of the fusion protein suggested that F-HN combination protein had good reactionogenicity.It provids further research efficient subunits genetic engineering vaccine in foundation.
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