Biomedical subjects

N Muthukumaran

Publications and source records attributed to N Muthukumaran.

8 recordsLinked to original sources

PubMed1991

Recipes for reconstituting skin.

Reconstituted Living Skin Equivalent (LSE) is made up of a dermal equivalent (DE) on which keratinocytes are plated where they give rise to a multilayered differentiated epidermis. The dermal equivalent develops through interactions between fibroblasts and collagen fibrils that begin to form after the cell-matrix precursor is cast. The gel that forms as a result of collagen polymerization and fluid trapping is contracted uniformly in all dimensions. By securing it at ends and edges in the mold in which it is cast, the final dimensions, strength and morphology of the forming tissue are altered. The same phenomena are seen in casting tubular tissues for the fabrication of small caliber blood vessel equivalents. The cells of the dermal equivalent are biosynthetically active and enrich the matrix to different degrees with secretory products, depending on how the cells are stimulated and on the presence or absence of an epidermis. Collagen biosynthesis by dermal cells in the DE is sensitive to growth factors, ascorbate concentrations and amino acid pools. Both ascorbate and TGF beta 1 increase total collagen biosynthesis at least two-fold by one week after tissue formation. With TGF beta 1 present, the capacity of cells in the DE to synthesize collagen increases with time, over a two-week period. If ascorbate (200 micrograms/ml) is added just after the tissue is cast and daily thereafter, contraction lattice is blocked, and collagen biosynthesis is enhanced relative to contracted controls that had received 200 micrograms/ml ascorbate once. The increase was nearly an order of magnitude over that of controls and was coordinate with a comparable increase in hyaluronate and sulfated glycosaminoglycan (GAG) production as shown by TCA-precipitable glucosamine in the intercellular matrix of the DE. Both the LSE and the Living Dermal Equivalent (LDE) exhibit complex responses to UV radiation and to various chemicals that are greatly different from responses given by monolayered cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗

PubMed1989

Purification and partial amino acid sequence of osteogenin, a protein initiating bone differentiation.

Osteogenin was purified from bovine bone matrix and its activity monitored by an in vivo bone induction assay. The purification method utilized extraction of the bone-inducing activity with 6 M urea, followed by chromatography on heparin-Sepharose, hydroxyapatite, and Sephacryl S-200. Active fractions were further purified by preparative sodium dodecyl sulfate gel electrophoresis without reduction. Osteogenin activity was localized in a zone between 30 and 40 kDa. The amino acid sequences of a number of tryptic peptides of the gel-eluted material were determined. Reduction and alkylation of purified osteogenin in 7 M guanidine hydrochloride resulted in the total loss of biological activity. Sodium dodecyl sulfate gel electrophoresis under reducing conditions revealed a broad band with an apparent molecular mass of 22 kDa.

Amino Acid Sequence↗

PubMed1989

Initiation of bone development by osteogenin and promotion by growth factors.

The cellular and molecular basis of bone development and its regulation by differentiation and growth factors is an exciting area of current research. This article briefly reviews the historical progress in the isolation of osteogenin, a novel bone differentiation factor, and its modulation by well known growth factors. Endochondral bone development is a multistep sequential cascade and the process must be operationally dissected. It has been accomplished with the demineralized bone matrix-induced bone formation model. The reproducible development of cartilage and bone in an extraskeletal site permits the study of the initiation of the first cycle of endochondral bone formation and mineralization. Recent progress in the isolation of osteogenin, a specific bone differentiation factor, by heparin affinity chromatography permits the further investigation of the commitment and clonal expansion of the putative osteoprogenitor stem cells. Once initiated, bone formation is promoted by growth factors such as platelet derived growth factor, fibroblast growth factor, insulin like growth factor, transforming growth factor beta and a plethora of non specific cytokines. Finally bone development is further modulated by systemic hormones and nutrition and a host of physical signals including electrical, gravitational and mechanical forces.

Animals↗

PubMed1988

Dose-dependence of and threshold for optimal bone induction by collagenous bone matrix and osteogenin-enriched fraction.

Subcutaneous implantation of demineralized collagenous bone matrix results in local induction of new bone formation. The dose dependence of bone induction was investigated using different amounts of rat demineralized bone matrix (DBM) and also osteogenin-enriched fraction with and without inactive collagenous bone matrix (ICBM). There is a threshold for bone induction; at least 10 mg of DBM is required. There is a dose dependent increase in bone induction between 10 to 25 mg. Exogenous type I collagen was found to be stimulatory to bone induction when suboptimal doses of DBM are employed. The activity of osteogenin-enriched dissociative extract was enhanced by addition of ICBM. These results imply that optimal bone induction requires the combined action of soluble osteogenin-enriched fraction and insoluble collagenous substratum.

Alkaline Phosphatase↗

PubMed1987

Isolation of osteogenin, an extracellular matrix-associated, bone-inductive protein, by heparin affinity chromatography.

Implantation of demineralized diaphyseal bone matrix in subcutaneous sites induces a sequence of events resulting in the local differentiation of endochondral bone. Demineralized bovine bone matrix was dissociatively extracted in 4.0 M guanidine hydrochloride and the bone-inductive proteins were purified greater than 12,000-fold. The purification steps include affinity chromatography on heparin-Sepharose, hydroxyapatite chromatography, gel filtration, and C18 reverse-phase HPLC. Since the purified protein in conjunction with insoluble collagenous bone matrix induced new bone differentiation in vivo we have designated this component osteogenin. The osteogenic potential is specific for osteogenin and is not exhibited by previously isolated growth factors.

Animals↗

PubMed1987

Biologic principles of bone induction.

This article provides a concise review of bone induction. Bone induction by demineralized bone matrix is a multistep cascade. The purification and elucidation of the chemistry of osteogens will improve bone grafting methods.

Alkaline Phosphatase↗

PubMed1985

Comparison of bone inductive proteins of rat and porcine bone matrix.

Subcutaneous implantation of demineralized bone matrix in allogenic rats induces a sequence of events resulting in de novo formation of cartilage, bone and bone marrow. In the present study endochondral bone formation by demineralized porcine matrix was studied and compared with the rat bone matrix. Endochondral bone formation was induced by 4M guanidine hydrochloride fraction IV (less than 50,000 daltons) of Sepharose CL-6B gel filtration but not by whole extract or by demineralized porcine bone matrix. Sephacryl S-200 gel filtration of the osteoinductive proteins of fraction IV showed the Porcine osteoinductive factor to be associated with protein fraction III (less than 20,000 daltons) whereas the rat with fraction II (between 20,000 and 30,000 daltons) of the chromatographic profile indicating an apparent difference in molecular weight of the osteoinductive factors between these two species.

Animals↗

PubMed1985

Bone matrix-induced local bone induction.

The sequential cellular changes in the implants in response to collagenous bone matrix-induced local bone formation include: binding of fibronectin to matrix, chemotaxis and attachment of progenitor cells, proliferation and differentiation of progenitor cells into chondrocytes, and finally osteogenesis and marrow differentiation. The cellular origin of osteogenic proteins is not clear. The present study compares the osteogenic potential of demineralized rat and porcine bone matrix by dissociative extraction and reconstitution. Judging from the Sephacryl S-200 gel filtration profiles of the dissociative extracts of rat and porcine matrix, the latter appears to be smaller. Under identical experimental conditions, the rat chondrosarcoma and osteosarcoma were examined for chondrogenic and osteogenic properties and found to be devoid of inductive potential. It is noteworthy that gel filtration fractions of rat chondrosarcoma on Sepharose CL-6B are inhibitory to bone inductive potential of demineralized rat bone matrix.

Animals↗