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1.
Mol Biotechnol ; 46(2): 118-26, 2010 Oct.
Article in English | MEDLINE | ID: mdl-20499289

ABSTRACT

Bone morphogenetic protein-7 (BMP-7) is a secreted multifunctional growth factor of the TGF-beta superfamily, which is predominantly known for its osteoinductive properties and emerging potential for treatment of kidney diseases. The mature 34-38 kDa disulfide-linked homodimer protein plays a key role in the differentiation of mesenchymal cells into bone and cartilage. In this study, the full-length sequence of hBMP-7 was amplified and, then, cloned, expressed, and purified from the conditioned medium of 293T cells stably transfected with a lentiviral vector. The mature protein dimer form was properly secreted and recognized by anti-BMP-7 antibodies, and the protein was shown to be glycosilated by treatment with exoglycosidase, followed by western blotting. Moreover, the activity of the purified protein was demonstrated both in vitro, by alkaline phosphatase activity in C2C12 cells, and in vivo by induction of ectopic bone formation in Balb/c Nude mice after 21 days, respectively. This recombinant protein platform may be very useful for expression of different human cytokines and other proteins for medical applications.


Subject(s)
Bone Morphogenetic Protein 7/biosynthesis , Bone Morphogenetic Protein 7/isolation & purification , Recombinant Proteins/biosynthesis , Recombinant Proteins/isolation & purification , Animals , Biological Assay/methods , Bone Morphogenetic Protein 7/chemistry , Bone Morphogenetic Protein 7/genetics , Bone Morphogenetic Protein 7/pharmacology , Chromatography, Affinity , Genetic Vectors , HEK293 Cells , Humans , Lentivirus , Mice , Mice, Inbred BALB C , Plasmids , Recombinant Proteins/chemistry , Recombinant Proteins/pharmacology
2.
Mol Biotechnol ; 39(2): 89-95, 2008 Jun.
Article in English | MEDLINE | ID: mdl-18327551

ABSTRACT

Social and economical development is closely associated with technological innovation and a well-developed biotechnological industry. In the last few years, Brazil's scientific production has been steadily increasing; however, the number of patents is lagging behind, with technological and translational research requiring governmental incentive and reinforcement. The Cell and Molecular Therapy Center (NUCEL) was created to develop activities in the translational research field, addressing concrete problems found in biomedical and veterinary areas and actively searching for solutions by employing a genetic engineering approach to generate cell lines over-expressing recombinant proteins to be transferred to local biotech companies, aiming at furthering the development of a national competence for local production of biopharmaceuticals of widespread use and of life-saving importance. To this end, mammalian cell engineering technologies were used to generate cell lines over-expressing several different recombinant proteins of biomedical and biotechnological interest, namely, recombinant human Amylin/IAPP for diabetes treatment, human FVIII and FIX clotting factors for hemophilia, human and bovine FSH for fertility and reproduction, and human bone repair proteins (BMPs). Expression of some of these proteins is also being sought with the baculovirus/insect cell system (BEVS) which, in many cases, is able to deliver high-yield production of recombinant proteins with biological activity comparable to that of mammalian systems, but in a much more cost-effective manner. Transfer of some of these recombinant products to local Biotech companies has been pursued by taking advantage of the São Paulo State Foundation (FAPESP) and Federal Government (FINEP, CNPq) incentives for joint Research Development and Innovation partnership projects.


Subject(s)
Biopharmaceutics , Interdisciplinary Communication , Recombinant Proteins/biosynthesis , Technology Transfer , Amyloid/biosynthesis , Animals , Baculoviridae/metabolism , Biotechnology , Bone Morphogenetic Proteins/biosynthesis , Brazil , Cell Line , Factor IX/biosynthesis , Factor VIII/biosynthesis , Follicle Stimulating Hormone/biosynthesis , Genetic Engineering , Genetic Vectors/biosynthesis , Humans , Islet Amyloid Polypeptide , Research/economics , Research/organization & administration , Spodoptera/virology
3.
Braz. j. med. biol. res ; 38(10): 1463-1473, Oct. 2005. ilus
Article in English | LILACS | ID: lil-409280

ABSTRACT

Bone morphogenetic proteins (BMPs) are multi-functional growth factors belonging to the transforming growth factor ß superfamily. Family members are expressed during limb development, endochondral ossification, early fracture, and cartilage repair. The activity of BMPs was first identified in the 1960s but the proteins responsible for bone induction were unknown until the purification and cloning of human BMPs in the 1980s. To date, about 15 BMP family members have been identified and characterized. The signal triggered by BMPs is transduced through serine/threonine kinase receptors, type I and II subtypes. Three type I receptors have been shown to bind BMP ligands, namely: type IA and IB BMP receptors and type IA activin receptors. BMPs seem to be involved in the regulation of cell proliferation, survival, differentiation and apoptosis, but their hallmark is their ability to induce bone, cartilage, ligament, and tendon formation at both heterotopic and orthotopic sites. This suggests that, in the future, they may play a major role in the treatment of bone diseases. Several animal studies have illustrated the potential of BMPs to enhance spinal fusion, repair critical-size defects, accelerate union, and heal articular cartilage lesions. Difficulties in producing and purifying BMPs from bone tissue have prompted the attempts made by several laboratories, including ours, to express these proteins in the recombinant form in heterologous systems. This review focuses on BMP structure, molecular mechanisms of action and significance and potential applications in medical, dental and veterinary practice for the treatment of cartilage and bone-related diseases.


Subject(s)
Animals , Humans , Bone Morphogenetic Proteins/physiology , Protein Conformation , Bone Diseases/therapy , Bone Morphogenetic Proteins/chemistry , Bone Morphogenetic Proteins/therapeutic use , Clinical Trials as Topic , Cartilage Diseases/therapy , Meta-Analysis as Topic , Signal Transduction/genetics , Signal Transduction/physiology
4.
Braz J Med Biol Res ; 38(10): 1463-73, 2005 Oct.
Article in English | MEDLINE | ID: mdl-16172739

ABSTRACT

Bone morphogenetic proteins (BMPs) are multi-functional growth factors belonging to the transforming growth factor ss superfamily. Family members are expressed during limb development, endochondral ossification, early fracture, and cartilage repair. The activity of BMPs was first identified in the 1960s but the proteins responsible for bone induction were unknown until the purification and cloning of human BMPs in the 1980s. To date, about 15 BMP family members have been identified and characterized. The signal triggered by BMPs is transduced through serine/threonine kinase receptors, type I and II subtypes. Three type I receptors have been shown to bind BMP ligands, namely: type IA and IB BMP receptors and type IA activin receptors. BMPs seem to be involved in the regulation of cell proliferation, survival, differentiation and apoptosis, but their hallmark is their ability to induce bone, cartilage, ligament, and tendon formation at both heterotopic and orthotopic sites. This suggests that, in the future, they may play a major role in the treatment of bone diseases. Several animal studies have illustrated the potential of BMPs to enhance spinal fusion, repair critical-size defects, accelerate union, and heal articular cartilage lesions. Difficulties in producing and purifying BMPs from bone tissue have prompted the attempts made by several laboratories, including ours, to express these proteins in the recombinant form in heterologous systems. This review focuses on BMP structure, molecular mechanisms of action and significance and potential applications in medical, dental and veterinary practice for the treatment of cartilage and bone-related diseases.


Subject(s)
Bone Morphogenetic Proteins/physiology , Protein Conformation , Animals , Bone Diseases/therapy , Bone Morphogenetic Proteins/chemistry , Bone Morphogenetic Proteins/therapeutic use , Cartilage Diseases/therapy , Clinical Trials as Topic , Humans , Meta-Analysis as Topic , Signal Transduction/genetics , Signal Transduction/physiology
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