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Biomedical subjects

Thomas J Koob

Publications and source records attributed to Thomas J Koob.

12 recordsLinked to original sources
PubMed2006

Changes in perlecan during chondrocyte differentiation in the fetal bovine rib growth plate.

Perlecan is a heparan sulfate proteoglycan present in the growth plate and essential for endochondral ossification. We evaluated the synthesis and structure of perlecan in the different zones of the growth plate. The growth plates from fetal bovine ribs were isolated and sequentially sliced into 1-mm sections containing the hypertrophic zone, lower proliferative zone, upper proliferative zone, intermediate zone, and resting zone, respectively. The slices were then either incubated in culture medium with 35SO4 to measure total sulfated proteoglycan synthesis and perlecan synthesis, extracted for perlecan core protein analysis by Western blot, or extracted for perlecan isolation and subsequent characterization of glycosaminoglycan size and disaccharide composition. 35SO4 incorporation into perlecan was three-fourfold higher in the proliferating/hypertrophic zone than the resting zone. Western blot showed perlecan content was greatest in the lower and upper proliferating zones and that a perlecan fragment lacking portions of the N- and C-terminal domains containing heparan sulfate was also present in all zones. Purified perlecan from the hypertrophic/lower proliferative zone had larger chondroitin sulfate chains and a different composition of CS and HS disaccharides than the perlecan isolated from the resting zone. These results indicate perlecan deposition is increased and is turned over during proliferation to be replaced by a perlecan with a different sulfation pattern.

Animals↗
PubMed2006

Material properties and biochemical composition of mineralized vertebral cartilage in seven elasmobranch species (Chondrichthyes).

Elasmobranchs, particularly sharks, function at speed and size extremes, exerting large forces on their cartilaginous skeletons while swimming. This casts doubt on the generalization that cartilaginous skeletons are mechanically inferior to bony skeletons, a proposition that has never been experimentally verified. We tested mineralized vertebral centra from seven species of elasmobranch fishes: six sharks and one axially undulating electric ray. Species were chosen to represent a variety of morphologies, inferred swimming speeds and ecological niches. We found vertebral cartilage to be as stiff and strong as mammalian trabecular bone. Inferred swimming speed was a good, but not infallible, predictor of stiffness and strength. Collagen content was also a good predictor of material stiffness and strength, although proteoglycan was not. The mineral fraction in vertebral cartilage was similar to that in mammalian trabecular bone and was a significant predictor of material properties.

Animals↗
PubMed2005

Structure and permeability of the egg capsule of the bonnethead shark, Sphyrna tiburo.

In the viviparous bonnethead shark, Sphyrna tiburo, a fluid-filled, acellular egg capsule surrounds fertilized eggs and developing embryos throughout gestation. Like other placental shark species, the capsule remains intact even at the placental implantation site. Although its intervention between the uterine and embryonic tissues of the placenta has long been thought to mediate physiological exchange, little information is available concerning even its basic structure or permeability to solutes. The 1 mum thick capsule wall consists of an inner layer of gelatinous material and an outer layer consisting of at least three laminae of orthogonally arranged fibrous material. These fibers are irregular and often branched. Permeability experiments showed that solutes less than 1,355 Da diffuse across the egg capsule whereas those greater than 6,000 Da do not pass through the membrane. Solute movement across the capsule is a concentration-dependent phenomenon indicating diffusion rather than active transport. Experimental data also suggest that there is an increase in the permeability of the egg capsule to low molecular weight materials during mid- and late gestation. These observations are discussed in relation to the function of the egg capsule as a mediator of maternal-embryonic interactions in matrotrophic sharks.

Animal Structures↗
PubMed2004

Indentation properties of growing femoral head following ischemic necrosis.

Little is known about the mechanical properties of the growing femoral head as it develops deformity following ischemic injury. The purpose of this study was to determine the indentation stiffness of growing femoral head following ischemic injury and to correlate the changes in stiffness with radiographic and histopathologic changes in the femoral head as it develops deformity. Following the induction of ischemia in 24 piglets, indentation testing of whole femoral heads was performed at 2, 4, and 8 weeks, as well as on femoral heads from eight sham operated animals. At 2 weeks, a 52% reduction of indentation stiffness was observed in the infarcted femoral heads compared to the control heads (p=0.004). The bony epiphyses in infarcted femoral heads were smaller due to growth arrest but they were not deformed. Histologically, no evidence of repair was seen. At 4 and 8 weeks, the indentation stiffness in the infarcted femoral heads was reduced by 75% (p<0.000001) and 72% (p=0.001) respectively compared to the control heads. Variable degree of femoral head deformity and repair was observed at 4 weeks. Severe deformity with extensive revascularization and repair were observed at 8 weeks. Although epiphyseal cartilage was thickened on the infarcted femoral heads only a weak correlation was found between the increase in the cartilage thickness and the decrease in the indentation stiffness (R(2)=0.55). These results indicate that the indentation properties of growing femoral head were significantly affected by ischemic injury, prior to the presence of repair process and deformity. A further decrease in the indentation stiffness was concomitant with repair of the infarcted head. These findings suggest that a reduction in the mechanical properties of the infarcted femoral head include both a cartilage and a bony component, which cannot be differentiated at this point. The study validates early institution of treatments that are aimed at limiting the mechanical loading of the affected hip. The study also suggests that in order to minimize the mechanical compromise of the infarcted femoral head, early institution of treatments aimed at stimulating new bone formation and retarding osteoclastic bone resorption may be beneficial.

Animals↗
PubMed2003

A novel fibrocartilaginous tendon from an elasmobranch fish ( Rhinoptera bonasus).

Tendons of the jaw adductor muscles of a hard prey crushing stingray exhibit similar adaptations to compressive and shear loads as those seen in mammalian tendons. Ventral intermandibular tendon from the cownose ray, Rhinoptera bonasus, has a prominent fibrocartilaginous pad that lies between a fibrous region of the tendon and the mineralized tissue of the jaw. Histologically the pad is similar to the fibrocartilaginous meniscus of mammals, and these tissues also share some biochemical traits. Proteoglycan (PG) content in the fibrocartilaginous pad is nearly four times higher than in the linearly arrayed tendinous tissue. The predominant PGs appear to be an aggrecan-like molecule and a decorin-like molecule. The decorin-like molecule is quite small when compared to mammalian decorin (20-80 kDa vs. 100-200 kDa). This study is the first to document adaptations to compressive/shear loading in tendon from a cartilaginous fish, and the similarities to the mammalian condition argue for the early evolution of this reactive ability of tendinous tissue.

Animals↗
PubMed2003

Mechanical and thermal properties of novel polymerized NDGA-gelatin hydrogels.

Nordihydroguaiaretic acid (NDGA), an antioxidant with two functional ortho-catechols from the creosote bush, has been shown to increase the mechanical properties of synthetic collagen fibers, producing biologically based, biocompatible fibers with material properties in uniaxial tensile tests to failure that are comparable to those of native tendon (Koob and Hernandez, Biomaterials 23 (2002) 203; Koob et al., J Biomed Mater Res, 56 (2001) 31; 56 (2001) 40). The NDGA polymerization scheme was applied to gelatin hydrogels to determine whether it could provide a viable approach for producing gelatin based biological materials with advantageous mechanical and thermal properties. NDGA treatment eliminated gelatin solubilization from hydrogels in chaotropic agents and increased the thermal stability of gelatin hydrogels from less than 37 degrees C to over 80 degrees C. NDGA caused a dose dependent increase in the compressive stiffness and fracture load of gels ranging in concentration from 2.5% to 40% gelatin in uniaxial, unconfined compression tests to failure. Maximum fracture load averaged 0.5+/-0.1MPa and the compressive modulus averaged 4.4+/-1.4MPa for all gelatin concentrations, however, the concentration of NDGA that produced maximum strength and stiffness varied inversely with gelatin concentration. The compressive strength and stiffness of 5% gelatin hydrogels treated with NDGA were independent of temperature up to 52 degrees C. These results indicate that NDGA polymerization renders gelatin hydrogels thermally and mechanically stable and thereby potentially useful for surgical procedures that would benefit from biocompatible, stable and mechanically competent gelatin-based biomaterials.

Biocompatible Materials↗
PubMed2002

Isolation and identification of the major heparan sulfate proteoglycans in the developing bovine rib growth plate.

Heparan sulfate proteoglycans are thought to mediate the action of growth factors. The heparan sulfate-containing proteoglycans in extracts of the bovine fetal rib growth plate were detected using the monoclonal antibody 3G10, which recognizes a neoepitope generated by heparitinase digestion (David, G., Bai, X. M., Van der Schueren, B., Cassiman, J. J., and Van den Berghe, H. (1992) J. Cell Biol. 119, 961-975). The heparan sulfate proteoglycans that react with this antibody were identified using antisera to known proteoglycans; purified using CsCl density gradient centrifugation, molecular sieve, and ion exchange chromatography; and then characterized. The major heparan sulfate proteoglycans in the growth plate had core proteins of 200 kDa and larger and were identified as perlecan and aggrecan. These two heparan sulfate proteoglycans could be effectively separated from each other by CsCl density gradient centrifugation alone. Perlecan contained 25% heparan sulfate and 75% chondroitin sulfate. The heparan sulfate chains on growth plate perlecan were considerably smaller than the chondroitin sulfate chains, and the heparan sulfate disaccharide content was different than that found for heparan sulfate from either kidney, tumor tissue, or growth plate aggrecan. Aggrecan contained only 0.1% heparan sulfate, which was localized to the CS-1 domain of aggrecan. These results indicate that perlecan and aggrecan would be the principal candidate proteoglycans involved in the action of heparan sulfate-binding proteins in the developing growth plate.

Amino Acid Sequence↗
PubMed2002

Material properties of polymerized NDGA-collagen composite fibers: development of biologically based tendon constructs.

Methods for stabilizing collagen-based materials with catechol containing monomers were developed in order to produce fibers with mechanical properties in tension comparable to those of normal tendon. Fibers produced from pepsin solubilized, bovine tendon type I collagen were polymerized with the di-catechol nordihydroguaiaretic acid (NDGA). Polymerization was based on the chemical oxidation of the constituent o-catechols to reactive o-quinone functionalities. NDGA caused a dose dependent increase in the tensile strength and stiffness of the type I collagen fibers. A second treatment with NDGA improved the tensile properties significantly. Comparison of the effects of NDGA with those of biologically relevant mono-catechols indicated that the bi-catechol functionality of NDGA was responsible for generation of the superior tensile properties. Elimination of unreacted intermediates from the treated fibers with ethanol increased the effectiveness of the cross-linking process while simultaneously sterilizing the material. Catalyzing oxidation by saturating the reaction buffer with oxygen increased the effectiveness of polymerization and the resulting tensile properties of the treated fibers. The ultimate tensile strength of the optimized NDGA-treated fibers averaged 90 MPa; the elastic modulus of these fibers averaged 580 MPa. Both values are comparable to native tendon. The material properties of the NDGA cross-linked fibers exceed the properties of collagen fibers treated with other cross-linking strategies such as glutaraldehyde and carbodiimide. These results indicate that NDGA cross-linking may provide a viable approach to stabilizing collagenous materials for use in repair of ruptured, lacerated or surgically transected tendons, as well as other biomaterial constructs for surgical repair of musculoskeletal injuries and disease.

Biocompatible Materials↗
PubMed2002

The evolution of tendon--morphology and material properties.

Phylogenetically, tendinous tissue first appears in the invertebrate chordate Branchiostoma as myosepta. This two-dimensional array of collagen fibers is highly organized, with fibers running along two primary axes. In hagfish the first linear tendons appear and the myosepta have developed specialized regions with unidirectional fiber orientation-a linear tendon within the flat sheet of myoseptum. Tendons react to compressive load by first forming a fibrocartilaginous pad, and under severe stress, sesamoid bones. Evidence for this ability to react to load first arises in the cartilaginous fish, here documented in a tendon from the jaw of a hard-prey crushing stingray. Sesamoid bones are common in bony fish and also in tetrapods. Tendons will also calcify under tensile loads in some groups of birds, and this reaction to load is seen in no other vertebrates. We conclude that the evolutionary history of tendon gives us insight into the use of model systems for investigating tendon biology. Using mammal and fish models may be more appropriate than avian models because of the apparent evolution of a novel reaction to tensile loads in birds.

Animals↗
PubMed2002

Biomimetic approaches to tendon repair.

The linear organization of collagen fibers in tendons results in optimal stiffness and strength at low strains under tensile load. However, this organization makes repairing ruptured or lacerated tendons extremely difficult. Current suturing techniques to join split ends of tendons, while providing sufficient mechanical strength to prevent gapping, are inadequate to carry normal loads. Immobilization protocols necessary to restore tendon congruity result in scar formation at the repair site and peripheral adhesions that limit excursion. These problems are reviewed to emphasize the need for novel approaches to tendon repair, one of which is the development of biomimetic tendons. The objective of the empirical work described here was to produce biologically-based, biocompatible tendon replacements with appropriate mechanical properties to enable immediate mobilization following surgical repair. Nor-dihydroguaiaretic acid (NDGA), a di-catechol from creosote bush, caused a dose dependent increase in the material properties of reconstituted collagen fibers, achieving a 100-fold increase in strength and stiffness over untreated fibers. The maximum tensile strength of the optimized NDGA treated fibers averaged 90 MPa; the elastic modulus of these fibers averaged 580 MPa. These properties were independent of strain rates ranging from 0.60 to 600 mm/min. Fatigue tests established that neither strength nor stiffness were affected after 80 k cycles at 5% strain. Treated fibers were not cytotoxic to tendon fibroblasts. Fibroblasts attached and proliferated on NDGA treated collagen normally. NDGA-fibers did not elicit a foreign body response nor did they stimulate an immune reaction during six weeks in vivo. The fibers survived 6 weeks with little evidence of fragmentation or degradation. The polymerization scheme described here produces a fiber-reinforced NDGA-polymer with mechanical properties approaching an elastic solid. The strength, stiffness and fatigue properties of the NDGA-treated fibers are comparable to those of tendon. These fibers are biocompatible with tendon fibroblasts and elicit little rejection or antigenic response in vivo. These results indicate that NDGA polymerization may provide a viable approach for producing collagenous materials that can be used to bridge gaps in ruptured or lacerated tendons. The tendon-like properties of the NDGA-fiber would allow early mobilization after surgical repair. We predict that timely loading of parted tendons joined by this novel biomaterial will enhance mechanically driven production of neo-tendon by the colonizing fibroblasts and result in superior repair and rapid return to normal properties.

Animals↗
PubMed2002

The notochord of hagfish Myxine glutinosa: visco-elastic properties and mechanical functions during steady swimming.

To determine the possible locomotor functions of the hagfish notochord, we measured its flexural stiffness EI (N m(-2)) and flexural damping C (kg m(3) s(-1)), under in vitro conditions that mimicked the body curvature and bending frequency measured during steady undulatory swimming. To assess the notochord's contribution to the mechanical behavior of the whole body, we also measured EI and C of the whole body, the body with skin removed, and the notochord with the outer fibrous sheath removed. When subjected to dynamic bending at angular frequencies from pi to 6pi rad s(-1) and midline curvatures from 11 to 40 m(-1), 1 cm in situ body segments (N=4), located at an axial position of 37% of the body length, showed significant changes in EI, C, the Young's modulus or material stiffness (E, MPa), the net work to bend the body over a cycle (W, J) and resilience (R, % energy return). When skin, muscles and the outer fibrous sheath of the notochord were removed sequentially, each structural reduction yielded significant changes in mechanical properties: C decreased when the skin was removed, E increased when the muscles were removed, and EI and R decreased when the outer fibrous sheath was removed. Although occupying only a small portion of the cross-sectional area, the notochord provides the body with 75% of its total EI and 80% of total C, by virtue of its high E, ranging from 4 to 8 MPa, which is an order of magnitude greater than that of the whole body. Thus, as the body's primary source of EI and C, the notochord determines the passive (i.e. internal, non-muscular) mechanical behavior of the swimming hagfish. EI and C covary inversely and non-linearly such that as C increases, EI decreases. However, the bending moments M (Nm) produced by each property increase proportionally, and the ratio of stiffness to damping moments, also known as the amplification ratio at resonance, is nearly invariant (approximately 7) with changes in driving frequency. If the body operates in life at or near resonance, the variables EI and C interact over a range of swimming speeds to produce passive mechanical stability.

Animals↗