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Biomedical subjects

H C Anderson

Publications and source records attributed to H C Anderson.

At least 19 recordsLinked to original sources
PubMed1996

Healing of a segmental defect in the rat femur with use of an extract from a cultured human osteosarcoma cell-line (Saos-2). A preliminary report.

Devitalized extracts from cultured human osteosarcoma cells (Saos-2) can induce ectopic bone formation. The ability of an extract from Saos-2 cells to stimulate healing of an operatively created four-millimeter defect in the femoral diaphyses of rats was compared with that of collagen and that of autogenous bone graft. Forty adult rats were randomized into four groups of ten each. In Group 1 (controls), no material was placed in the defect; in Group 2, the defect was filled with pure bovine collagen; in Group 3, it was filled with autogenous graft obtained by morseling of the resected segment of the femur; and in Group 4, it was filled with ten milligrams of extract from Saos-2 cells that was mixed with an equal amount of bovine collagen. Five rats from each group were killed at four weeks and the remaining five, at eight weeks. Each femoral defect was analyzed radiographically and histologically for osseous healing. There was no evidence of healing at either four or eight weeks in Groups 1 and 2. Although there was some new-bone formation in Group 3, none of the defects had united at eight weeks. There was early, almost complete union in all five four-week specimens in Group 4 and complete healing of the defect in four of the five rats assessed at eight weeks. The Saos-2 cell extract was found to be the most effective agent, promoting union by mature lamellar bone within eight weeks.

Animals↗
PubMed1995

Effects of zinc and divalent cation chelators on ATP hydrolysis and Ca deposition by rachitic rat matrix vesicles.

Elsewhere it has been shown that zinc is highly concentrated in the hypertrophic zone of epiphyseal cartilage. It has also been shown that zinc deficiency can result in abnormal bone development, suggesting a direct or indirect role for zinc in calcification. Because matrix vesicles have been implicated in the initiation of calcification, we tested the effect of zinc and its chelators, such as EGTA and phenanthroline, on ATP-dependent Ca uptake by rat matrix vesicles. EGTA pretreatment of matrix vesicles inhibited ATP-dependent Ca uptake by 50%. To see if zinc depletion by EGTA pretreatment is responsible for decreased levels of ATP-dependent Ca uptake, ZnCl2 concentrations, ranging from 5 to 100 mumol/L, were tested for their ability to restore Ca deposition. Zinc exerted a striking enhancing effect on ATP-dependent Ca uptake of both untreated and EGTA-pretreated matrix vesicles in a dose-dependent manner. A 50% activation occurs at about 16 mumol/L Zn2+. At 63 mumol/L Zn2+, there was a fourfold increase in Ca-depositing activity. Addition of an excess amount of phenanthroline relative to Zn2+ concentration to the reaction mixture failed to abolish activation of Ca uptake by Zn2+, indicating that the putative chelator-Zn2+ complex formation did not prevent activation. The observed partial inhibition of ATPase and the activation of ATP-dependent Ca uptake of Zn2+ suggest that, in addition to ATPase, some other Ca and/or Pi uptake activators responsive to Zn2+ treatment are present in mammalian matrix vesicles.

Adenosine Triphosphatases↗
PubMed1995

Use of monoclonal antibody to detect bone morphogenetic protein-4 (BMP-4).

A monoclonal antibody that reacts with murine and human bone morphogenetic protein-4 (BMP-4) has been developed using recombinant BMP-4 as an immunogen. The antibody that bound most tightly to recombinant murine (rm)BMP-4 was selected, subcloned, and characterized. The specificity of the antibody was confirmed using Western blot analysis and enzyme-linked immunosorbent assay (ELISA). The antibody reacts with murine and human BMP-4 in both the reduced and nonreduced condition; however, this antibody shows cross-reactivity with neither human BMP-2 nor TGF-beta 1. Thus, the produced antibody could recognize the disulfide-linked dimeric structure of bioactive BMP-4, regardless of the species. Immunocytochemical study using this antibody successfully shows the cytosolic localization of BMP-4 in osteoinductive cells; i.e., BFO and Saos-2 in which the level of mRNA for BMP-4 was proved to be constitutively high by Northern blot analysis. In addition, the antibody could demonstrate the presence of BMP-4 in developmental bone formation in the alveolar bone of rat embryo by immunohistochemistry. The antibody could be used for a more sensitive approach for quantitative analysis of BMP-4.

Animals↗
PubMed1995

Molecular biology of matrix vesicles.

Matrix vesicles are extracellular 100-nanometer-diameter membrane-invested particles selectively located within the matrix of bone, cartilage, and predentin. They serve as the initial site of calcification in all skeletal tissues. Matrix vesicle biogenesis occurs by polarized budding and pinching off of vesicles from specific regions of the outer plasma membrane of chondrocytes, osteoblasts, and odontoblasts. Seeding of selected areas of matrix with matrix vesicles explains the localized distribution of subsequent zones of mineralization. Matrix vesicle biogenesis in the growth plate is linked to the chondrocyte cell cycle and reflects a stage in programmed cell death (apoptosis). Generation of initial hydroxyapatite mineral crystals occurs within the matrix vesicle membrane during Phase 1 of biologic mineralization. Phase 1 is controlled by phosphatases (including alkaline phosphatase) and Ca-binding molecules with which the matrix vesicles are well endowed. Phase 2 of biologic mineralization begins with breakdown of matrix vesicle membranes, exposing preformed hydroxyapatite to the extracellular fluid after which mineral crystal proliferation is governed by extracellular conditions. Phase 1 and Phase 2 of mineralization are under cellular control. Phase 1 is initiated by cells generating calcifiable matrix vesicles and releasing them into sites of intended calcification. Phase 2 is controlled by cells regulating extracellular ionic conditions and matrix composition.

Alkaline Phosphatase↗
PubMed1995

The mechanism of bone induction and bone healing by human osteosarcoma cell extracts.

Saos-2 cultured human osteosarcoma cells contain an extractable bone inducing agent that can induce heterotopic bone in the muscle of Nu/Nu mice. A semipurified GuHCl extract of Saos-2 cells also can promote healing and complete bony union in otherwise non-healing surgically induced defects of rat femur. Northern blot analyses indicate expression of mRNAs for bone morphogenetic proteins (BMP)-1, 2, 3, 4, 6 and transforming growth factor beta (TGF beta) in Saos-2 cells, and BMP-2, 3, 4, 5, 7 and TGF beta in nonosteoinductive U20S human osteosarcoma cells. Saos-2 cells exceeded U20S cells in expression levels of BMP-1, 3, 4 and TGF beta, whereas U20S cells expressed higher levels of BMP-2, 6 and also expressed trace amounts of BMP-5 and 7 not seen in Saos-2 cells. The authors hypothesize that Saos-2 cells contain an optimal admixture of known bone growth factors plus possible other unknown components that, acting alone or in combination with bone morphogenetic protein and/or TGF beta, can induce bone. Although bone inducing agent-induced heterotopic bones have half lives of only a few weeks, the reparative bone induced by bone inducing agent in femoral defects gives every indication of being permanent and self-sustaining. This suggests a fundamental difference between heterotopic and orthotopic osteoprogenitor cells with those involved in orthotopic bone repair more closely resembling the committed or determined osteoprogenitor cells of marrow as described by Friedenstein.

Animals↗
PubMed1994

Changes in subcellular localization and surface expression of L-selectin, alkaline phosphatase, and Mac-1 in human neutrophils during stimulation with inflammatory mediators.

The localization of the adhesion protein L-selectin in human neutrophils was determined by subcellular fractionation and immunoelectron microscopy and compared with the localization of Mac-1 (alpha m beta 2) and alkaline phosphatase, the marker for secretory vesicles. L-selectin was found to be localized exclusively on the plasma membrane of unstimulated cells and also of stimulated cells, although markedly diminished. This was in contrast to Mac-1, which was also localized in secretory vesicles and in specific/gelatinase granules as shown previously [Sengeløv, H., et al. J. Clin. Invest. (1993) 92, 1467-1476]. Stimulation of neutrophils with inflammatory mediators such as tumor necrosis factor (TNF), platelet-activating factor (PAF), or f-Met-Leu-Phe (fMLP), induced parallel up-regulation of the surface membrane content of alkaline phosphatase and Mac-1 and down-regulation of L-selectin, as evidenced by flow cytometry. Preimbedding immunoelectron microscopy confirmed that L-selectin was present mainly on tips of microvilli in unstimulated cells and showed that alkaline phosphatase and Mac-1 were randomly distributed on the surface membrane of fMLP-stimulated cells. These studies indicate that the transition of neutrophils from L-selectin-presenting cells to Mac-1-presenting cells induced by inflammatory mediators is mediated by incorporation of secretory vesicle membrane, rich in Mac-1 and devoid of L-selectin, into the plasma membrane.

Alkaline Phosphatase↗
PubMed1992

Familial syndrome of endocrine and neuroectodermal abnormalities.

We report on a previously undescribed combination of endocrine and neuroectodermal abnormalities in four sibs from Burma. These abnormalities include low growth hormone levels in response to provocative stimuli, delayed puberty associated with prepubertal levels of gonadotropins in the males and pubertal levels of gonadotropins in the females, type II diabetes mellitus with elevated insulin levels, mild mental retardation, sensori-neural deafness, and alopecia without pili torti. They also had a characteristic facial appearance and fleshy hands and feet. This family appears to have a previously undescribed combination of endocrine and neuroectodermal abnormalities.

Adolescent↗
PubMed1992

Comparison of normal and rachitic rat matrix vesicles.

Accumulated information about MVs in PO4-deficient rachitic rats shows them to be normal in most aspects. Both normal and rachitic MVs show the same ultrastructural features and selective spatial distribution in the growth plate, and both contain a nearly identical array of major proteins. Rachitic and normal MVs show the same avidity to calcify in vivo and in vitro. A slightly greater specific activity of ALP in rachitic MVs may enhance their calcifiability. We conclude that rachitic rat MVs are essentially 'normal' and can be used as an adequate model to study the mechanism of biological calcification.

Alkaline Phosphatase↗
PubMed1992

Presence and specific concentration of carbonic anhydrase II in matrix vesicles.

Matrix vesicles were isolated from the epiphyseal growth plates of normal weanling rats, and the presence of carbonic anhydrase II was demonstrated by Western blotting and ultrastructural immunolocalization using the immunogold technique. Total carbonic anhydrase activity was assayed and showed a statistically significant increase in matrix vesicles as compared to normal rat chondrocytes derived from the same growth plates. These results are the first to establish the presence of carbonic anhydrase in matrix vesicles.

Animals↗
PubMed1992

Bone-inducing agent (BIA) from cultured human Saos-2 osteosarcoma cells.

The Saos-2 line of human osteosarcoma cells was established in culture in 1975. These cells produce a large amount of alkaline phosphatase but little or no matrix in vitro, and are unable to grow when transplanted into athymic mice. We decided to test our local strain of Saos-2 cells for bone-inducing ability in the skeletal muscle of athymic mice by implanting freeze-dried, acetone-defatted cells, with and without a collagen carrier. A bone-inducing activity (BIA) thus was demonstrated in 88% of 90 implants of devitalized Saos-2 cells. In further studies, we have used guanidinium hydrochloride (Gu-HCl) to extract, solubilize, and remove the Saos bone-inducing agent(s) in an active state which when reprecipitated by aqueous dialysis was able to induce ultrastructurally typical endochondrial bone formation in nude mouse muscle in 92% of 48 implants. This preliminary report is offered to alert investigators to the presence of an extractable BIA in Saos-2 cells.

Animals↗
PubMed1992

Immunolocalization of alkaline phosphatase in osteoblasts and matrix vesicles of human fetal bone.

A monoclonal antibody raised against alkaline phosphatase (ALP) of human osteosarcoma was used to localize this enzyme in human fetal bone tissue. For light microscopy, the presence of alkaline phosphatase in osteoblasts and osteocytes was demonstrated by use of an avidin-biotin immunoperoxidase procedure. Electron microscopic immunolocalization was accomplished with an indirect immunoperoxidase method which revealed a concentration of the enzyme on matrix vesicle and osteoblast plasma membranes. In addition, many vesicular protrusions arising from areas of plasma membrane on the lateral surfaces of adjacent osteoblasts were strongly immunolabeled. Immunostaining for ALP was absent in vesicles which contained fine crystallites. Alkaline-phosphatase-rich matrix vesicles may play a significant role in the mineralization of the extracellular matrix.

Alkaline Phosphatase↗
PubMed1991

Monoclonal antibody against human bone alkaline phosphatase.

A monoclonal antibody which reacts preferentially with human bone alkaline phosphatase (AP) has been developed using human osteosarcoma bone AP as an immunogen. The antibody with the highest selectivity shows about three- and a-half fold greater binding to bone than to liver AP. The selectivity was confirmed using mixtures of authentic samples of the bone and liver isoenzymes. Its selectivity was also shown by measurements of bone AP in sera from patients with either bone or liver diseases. Such a selective antibody could lead to the development of a monoclonal antibody highly specific for bone AP which can be used for a more sensitive approach for quantitative analysis of the isoenzyme.

Alkaline Phosphatase↗
PubMed1991

Cardiovascular implant calcification: a survey and update.

Calcification of cardiovascular prosthetic implants is a common and important problem. This review provides an update based upon the Conference on Cardiovascular Implant Calcification held as part of the 13th World Congress of the International Society for Heart Research, 1989. A variety of cardiovascular prostheses are affected clinically by calcification, including bioprosthetic heart valves, aortic homografts and trileaflet polymeric valve prostheses. In addition, experimental studies have demonstrated calcification of artificial heart devices in ventricular assist systems in long-term calf studies. The pathophysiology of this disease process is incompletely understood. A common element between the various types of cardiovascular implant calcification is the localization of calcific deposits to devitalized cells and membranous debris. Prevention of cardiovascular implant calcification by either biomaterial modifications or regional drug therapy (controlled release) is being investigated.

Animals↗
PubMed1991

Increased matrix vesicle protein in rachitic rat epiphyseal growth plates.

Extracellular, membrane-bound vesicles are widely regarded to be the initial site of calcification in a variety of tissues under normal and pathological conditions. Alkaline phosphatase is believed to play a vital role in this process by hydrolysing ester phosphates or mineral inhibitors, e.g. inorganic phosphates. In the present study, matrix vesicles from normal and rachitic rat growth plates were compared with regard to specific activity of alkaline phosphatase, total vesicle protein and ultrastructural distribution of alkaline phosphatase activity. Matrix vesicles were released from normal or rachitic growth plates by collagenase digestion and isolated by differential centrifugation. Enzyme cytochemical localization involving a cerium capture method was performed on vesicles collected by vacuum filtration on Millipore filters. SDS gels and Western blots on fractions of both normal and rachitic matrix vesicles showed major proteins to be almost identical and confirmed the presence of alkaline phosphatase in both. Total matrix vesicle protein ((mg total matrix vesicle protein/rat) x 10(2)) per rat was significantly greater for the rachitic animals (9.0 +/- 2.0 vs. 4.0 +/- 1.0), P less than 0.0001. Alkaline phosphatase specific activity (units alkaline phosphatase/mg vesicle protein) in the rachitic and normal matrix vesicles was 25.29 +/- 9.36 and 18.78 +/- 3.37, respectively (0.05 less than P less than 0.1). Electron dense cerium phosphate deposits were localized to the outer membrane surface of matrix vesicles derived from both types of rats. This data, the first to quantify the relationship between rickets, matrix vesicle protein and alkaline phosphatase specific activity, suggests that matrix vesicles from rachitic and normal rats have biochemical and morphological similarity.(ABSTRACT TRUNCATED AT 250 WORDS)

Alkaline Phosphatase↗
PubMed1991

The effect of a stiff spinal implant on the bone-mineral content of the lumbar spine in dogs.

The response of canine lumbar vertebral bone to the application of a stiff transpedicular screw-plate implant from the third to the fifth lumbar vertebra, without arthrodesis, was investigated. Five groups of six dogs each were studied: dogs that had not had an operation (control group); dogs that had had a sham operation, with survival periods of three and six months; and dogs that had received an implant, with survival periods of three and six months. The results were the same in specimens from the control group and the group that had had a sham operation. In the groups that had received an implant, dual-photon absorptiometry revealed an insignificant decrease in bone-mineral content at the bypassed fourth lumbar segment (17 per cent at three months and 12 per cent at six months). When the data for the three and six-month intervals were pooled, the mean decrease in bone-mineral content of 14 per cent was significant. Histomorphic study yielded similar results; the mean decrease in bone-mineral content for the pooled three and six-month specimens was significant (16 per cent). Losses were similar for the ventral and dorsal columns. Histomorphometric analysis also showed a significant (13 per cent) loss of bone-mineral content at the adjacent caudad (sixth lumbar) vertebra for the pooled three and six-month groups. The results of the methods of analysis of loss of bone-mineral content correlated strongly. Under these experimental conditions, a stiff spinal implant caused loss of bone-mineral content of the bypassed vertebral segment, although the loss was less than anticipated and did not increase between three and six months.

Absorptiometry, Photon↗