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Biomedical subjects

M D McKee

Publications and source records attributed to M D McKee.

At least 73 records · Page 4Linked to original sources
PubMed1997

Bone acidic glycoprotein-75 self-associates to form macromolecular complexes in vitro and in vivo with the potential to sequester phosphate ions.

Monoclonal antibody HTP IV-#1 specifically recognizes a complexation-dependent neoepitope on bone acidic glycoprotein-75 (BAG-75) and a Mr = 50 kDa fragment. Complexes of BAG-75 exist in situ, as shown by immunofluorescent staining of the primary spongiosa of rat tibial metaphysis and osteosarcoma cell micromass cultures with monoclonal antibody HTP IV-#1. Incorporation of BAG-75 into complexes by newborn growth plate and calvarial tissues was confirmed with a second, anti-BAG-75 peptide antibody (#503). Newly synthesized BAG-75 immunoprecipitated from mineralizing explant cultures of bone was present entirely in large macromolecular complexes, while immunoprecipitates from monolayer cultures of osteoblastic cells were previously shown to contain only monomeric Mr = 75 kDa BAG-75 and a 50 kDa fragment. Purified BAG-75 self-associated in vitro to form large spherical aggregate structures composed of a meshwork of 10 nm diameter fibrils. These structures have the capacity to sequester large amounts of phosphate ions as evidenced by X-ray microanalysis and by the fact that purified BAG-75 preparations, even after extensive dialysis against water, retained phosphate ions in concentrations more than 1,000-fold higher than can be accounted for by exchange calculations or by electrostatic binding. The ultrastructural distribution of immunogold-labeled BAG-75 in the primary spongiosa underlying the rat growth plate is distinct from that for other acidic phosphoproteins, osteopontin and bone sialoprotein. We conclude that BAG-75 self-associates in vitro and in vivo into microfibrillar complexes which are specifically recognized by monoclonal antibody HTP IV-#1. This propensity to self-associate into macromolecular complexes is not shared with acidic phosphoproteins osteopontin and bone sialoprotein. We hypothesize that an extracellular electronegative network of macromolecular BAG-75 complexes could serve an organizational role in forming bone or as a barrier restricting local diffusion of phosphate ions.

Animals↗
PubMed1997

Spontaneous calcification of arteries and cartilage in mice lacking matrix GLA protein.

Calcification of the extracellular matrix (ECM) can be physiological or pathological. Physiological calcification occurs in bone when the soft ECM is converted into a rigid material capable of sustaining mechanical force; pathological calcification can occur in arteries and cartilage and other soft tissues. No molecular determinant regulating ECM calcification has yet been identified. A candidate molecule is matrix GLA protein (Mgp), a mineral-binding ECM protein synthesized by vascular smooth-muscle cells and chondrocytes, two cell types that produce an uncalcified ECM. Mice that lack Mgp develop to term but die within two months as a result of arterial calcification which leads to blood-vessel rupture. Chondrocytes that elaborate a typical cartilage matrix can be seen in the affected arteries. Mgp-deficient mice additionally exhibit inappropriate calcification of various cartilages, including the growth plate, which eventually leads to short stature, osteopenia and fractures. These results indicate that ECM calcification must be actively inhibited in soft tissues. To our knowledge, Mgp is the first inhibitor of calcification of arteries and cartilage to be characterized in vivo.

Animals↗
PubMed1997

The relative importance of intramedullary, intracortical, and extraosseous soft-tissue blood flow to the repair of devascularized canine tibial cortex.

Previous studies have not quantified the relative contributions of intracortical and extracortical perfusion to cortical porosity and new bone formation. The current study was performed to determine the relative importance of intramedullary, intracortical, and extraosseous soft-tissue blood flow and type of tissue to the repair of devascularized canine tibial cortex. A 2.5-cm segment of tibia between two standardized osteotomies was devascularized. The segment was replaced anatomically and stabilized with a plate. The animals were divided randomly into two groups: skin coverage (N = 8) and muscle coverage (N = 8). Thirty-one days postoperatively, cerium141 microspheres were injected, prior to sacrifice, to measure blood flow. Extraosseous soft-tissue perfusion was the same in the skin coverage and muscle flap coverage groups. There was no relationship between intramedullary or extraosseous soft-tissue flow and depth of new bone formation and cortical porosity. Intracortical blood flow was directly related to depth of new bone formation (p = 0.0006) and cortical porosity (outer cortex, p = 0.001; inner cortex, p = 0.0001). These findings indicate that the cortical repair process is linked to the restoration of perfusion and that muscle coverage, rather than the quantity of blood flow, determines the extent of cortical repair.

Animals↗
PubMed1997

Nonunion of distal radial fractures associated with distal ulnar shaft fractures: a report of four cases.

Although complications following distal radial fractures are common, nonunion is rare. We report four cases of radial nonunion following distal radial fractures that occurred in elderly women after low-energy falls. The distinctive feature of these injuries was the presence of an associated distal ulnar shaft fracture. All patients had initial closed reduction and cast immobilization, which was unsuccessful. Reconstruction was complicated by deformity, disuse osteoporosis, radiocarpal stiffness, and distal radioulnar joint disruption. Wrist arthrodesis improved function and decreased pain and deformity in three patients. Metaphyseal distal radial fractures associated with distal ulnar shaft fractures represent an unstable injury pattern, and closed reduction and casting may not be adequate treatment. Once established nonunion is present, wrist arthrodesis is a useful salvage procedure.

Aged↗
PubMed1997

The effect of a femoral fracture on concomitant closed head injury in patients with multiple injuries.

OBJECTIVE: We sought to determine the effect of a femoral shaft fracture, and its treatment by early intramedullary nailing, on the neurologic outcome of patients with multiple injuries with a concomitant head injury. DESIGN: Retrospective, case-control design using a prospectively gathered trauma data base. MATERIALS AND METHODS: We identified 46 patients with multiple injuries (mean Injury Severity Score [ISS] = 33.2) with closed head injuries (mean Glasgow Coma Scale [GCS] score = 7.8) and femur fractures, and matched as controls 99 patients with multiple injuries with head injuries but without femur fractures for age, sex, mechanism of injury, ISS (mean ISS = 34.0), and GCS (mean GCS score = 8.0). Follow-up parameters examined included early mortality, length of hospital or intensive-care unit stay, neuropsychological testing, and level of neurologic disability. RESULTS: There were no significant differences in the demographics or injury parameters between the study and control groups. There were no significant differences between the two groups in terms of early mortality (study group, 28%; control group, 27%; p = not significant), length of hospital/intensive-care unit stay (study group, 17.5/6.9 days; control group, 18.0/6.3 days; p = not significant), level of neurologic disability, or results of cognitive testing. CONCLUSION: Our study suggests that a femoral fracture in a patient with a concomitant head injury does not increase mortality or neurologic disability, and supports the continued early intramedullary nailing of femoral fractures for these patients.

Adolescent↗
PubMed1996

Osteopontin at mineralized tissue interfaces in bone, teeth, and osseointegrated implants: ultrastructural distribution and implications for mineralized tissue formation, turnover, and repair.

Currently available data describing the gene expression and regulation, secretion, distribution, and protein chemistry of osteopontin (OPN) all are consistent with the notions of this protein functioning as an inhibitor of mineralization and/or as a mediator of cell-matrix and matrix-matrix/mineral adhesion (cohesion) during the formation, turnover, and repair of normal and pathological mineralized tissues. The properties and overall integrity of mineralized tissues are in part dictated by the nature of their interfaces--sites where organic and inorganic components of the extracellular matrix interact to provide biomechanical strength, regulate mineral ion homeostasis, and influence cellular events involved in mineralized tissue modeling, remodeling, and repair. High-resolution, colloidal-gold immunocytochemistry has been used to characterize the proteinaceous composition of these interfaces and to establish that the phosphorylated sialoprotein, OPN, is a major component found at these sites where it accumulates as a dense, planar "coating" of organic material termed either a cement line or a lamina limitans. Structural/functional features of OPN predict an ability of this protein to regulate calcification in the matrix proper of mineralized tissues and to participate, more specifically, in cell-matrix and matrix-matrix/mineral adhesion in laminae limitantes and cement lines, respectively. From the ultrastructural immunocytochemical data presented herein for OPN illustrating the cellular expression and extracellular matrix distribution of this protein, it is demonstrated that the production of OPN is one of the earliest, and latest, secretory activities of the osteoblast lineage and that this activity manifests itself morphologically as a cement line or a lamina limitans, respectively, at bone matrix interfaces. In laminae limitantes at bone surfaces, OPN appears to be involved in osteoclast adhesion and possibly haptotaxis. An OPN-containing cement line is also present at hard tissue interfaces in rat tooth, against osseointegrated titanium and hydroxyapatite implants and at the margins of surgically created bone defects--and there may influence biological adhesion in a manner similar to that proposed for normal bone. It is suggested, therefore, that in addition to its potential for influencing cell adhesion/dynamics in bones and teeth, OPN in cement lines may act as an interfacial adhesion promoter between apposing substrates, therein maintaining the overall integrity of bone during the bone remodeling sequence and "bonding" dissimilar tissues (or biocompatible materials) together in biological composites such as teeth and osseointegrated implants.

Animals↗
PubMed1996

Ultrastructural characterization and immunolocalization of osteopontin in rat calvarial osteoblast primary cultures.

As part of ongoing studies aimed at clarifying the early events of bone matrix deposition and mineralization, we have characterized primary osteoblast cultures using ultrastructural and immunocytochemical methods. Osteogenic cells were isolated by sequential enzymatic digestion of newborn rat (2-4-day-old) calvariae and grown for periods of 7 to 28 days on polystyrene, Thermanox plastic, or sputtered titanium. Bone-like nodules, showing a stratified organization of cells and collagen, were examined by scanning and transmission electron microscopy, and further characterized for mineral by backscattered electron imaging and X-ray microanalysis. Colloidal gold immunocytochemistry was used to examine the distribution of osteopontin in these nodules. Cells at the surface of the nodules were rounded, while those within the nodules generally appeared more flattened. Both cell types, particularly at early culture intervals, exhibited well-developed protein synthetic organelles. Collagen fibrils were present between the cell layers and some individual fibrils appeared mineralized. Aggregates of needle-shaped crystallites were sometimes apposed to the cell surface, frequently within invaginated regions of the cell membrane, while other mineralized masses of various sizes were present within the collagenous scaffolding. The periphery of the mineralized masses was often delimited by an electron-dense, lamina limitans-like layer. Focal accumulations and/or a more complete layer of afibrillar, mineralized organic matrix were sometimes observed at the interface between the cells and the surface of the culture dish. Osteopontin was immunodetected over the afibrillar and collagenous mineralized matrix throughout the cultures and, in some cases, labeling was concentrated over the peripheral, electron-dense material delimiting the mineralized masses. In conclusion, these data indicate that calvaria-derived osteoblasts produce an extracellular matrix with structural and compositional similarities to bone. Although not a regular observation, the accumulation of osteopontin on the surface of the culture substrate and at the periphery of masses of mineralized matrix may be analogous to what takes place in vivo at naturally occurring bone interfaces.

Animals↗
PubMed1996

Extracellular matrix in tooth cementum and mantle dentin: localization of osteopontin and other noncollagenous proteins, plasma proteins, and glycoconjugates by electron microscopy.

BACKGROUND: Noncollagenous proteins (NCPs) are considered to have multiple functions related to the formation, turnover, and repair of the collagen-based mineralized tissues. Collectively, they comprise a class of generally acidic, mineral-binding proteins showing extensive posttranslational modifications, including glycosylation, phosphorylation, and sulfation. METHODS. We have used colloidal-gold immunocytochemistry and lectin-gold cytochemistry, together with transmission electron microscopy, to examine the organic matrix composition of tooth cementum and the subjacent mantle dentin in rodent molar teeth. Molars were processed for immunocytochemistry using antibodies against osteopontin (OPN), osteocalcin (OC), bone sialoprotein (BSP), bone acidic glycoprotein-75 (BAG-75), albumin (ALB), and alpha 2HS-glycoprotein (alpha 2HS-GP), or for glycoconjugate cytochemistry using lectin-gold complexes. RESULTS: Ultrastructurally, at the advancing root edge in developing molars, OPN and BSP initially were associated with small calcification foci in the mantle dentin. With progressing mineralization, OC and alpha 2HS-GP appeared diffusely distributed throughout the calcified mantle dentin, and diminished as a gradient toward the circumpulpal dentin. Immediately following disruption of Hertwig's epithelial root sheath, cementum deposition commenced at the root surface occasionally with the appearance of a cement line rich in OPN. Cementum matrix proper contained abundant OPN, BSP, OC, and alpha 2HS-GP, but no or little BAG-75 or ALB. Protein immunolabeling, as well as lectin labeling for beta-D-galactose and N-acetyl-neuraminic acid and/or N-glycolyl-neuraminic acid, both being prominent sugars of certain NCPs, was primarily concentrated between, and at the surface of, collagen fibrils in acellular extrinsic fiber cementum. OPN, BSP, OC, and alpha 2HS-GP were also prominent components of cellular cementum and of Sharpey's fibers. In cellular cementum, laminae limitantes sometimes present delimiting cementocyte lacunae and cell process-containing canaliculi were also rich in OPN. Along the root surface, occasional cementoblasts exhibited intracellular labeling for OPN over the Golgi apparatus and secretory granules. CONCLUSIONS: We have identified OPN, BSP, OC, and alpha 2HS-GP as being prominent organic constituents of both mantle dentin and acellular and cellular cementum, and, have elucidated the details of their distribution at the ultrastructural level. The temporal appearance and spatial distribution of these organic moieties in the teeth root are similar to those seen during bone formation and are consistent with proposals that certain NCPs may be involved in regulating calcification and/or participating in cell-matrix and matrix-matrix/mineral adhesion events.

Animals↗
PubMed1996

Secretion of Osteopontin by macrophages and its accumulation at tissue surfaces during wound healing in mineralized tissues: a potential requirement for macrophage adhesion and phagocytosis.

Osteopontin (OPN), a noncollagenous, extracellular matrix sialoprotein found at relatively high levels in both normal and pathological mineralized tissues, is expressed by tissue-specific cells in bone, calcified cartilage, and teeth. On the other hand, a hallmark of OPN expression in pathologically mineralizing tissue, and in other soft tissues experiencing a more generalized type of necrotic injury, is the production of OPN by macrophages at the lesion site. In the present study, we have localized OPN and other noncollagenous proteins by ultrastructural colloidal-gold immunocytochemistry using a rat model in which mineralized tissue defects are surgically created in mandibular bone and teeth. The healing response was examined by immunocytochemistry and transmission electron microscopy at 10 min, 3 days and 7 days post-surgery using antibodies against OPN, bone sialoprotein, osteocalcin, bone acidic glycoprotein-75, fibronectin, and amelogenin. Whereas most of these proteins were characteristically distributed within their respective extracellular matrices as described previously, OPN was additionally observed to accumulate as a lamina limitans at surgically exposed bone and tooth surfaces, as well as at the surface of particulate, mineralized tissue debris. Intracellular labeling of the Golgi apparatus and secretory granules of macrophages at the lesion site demonstrated that OPN production by macrophages was a prominent secretory event of the inflammatory response during wound healing in mineralized tissues. Pseudopodal and lamellipodal cytoplasmic extensions of macrophages were observed in direct contact with the OPN-containing lamina limitans at these surfaces. Particulate, calcified debris internalized by macrophages also displayed a prominent surface "coating" of OPN. In conclusion, our interpretation of the present data is that OPN secreted by macrophages may serve as a macrophage adhesion protein, and where concentrated at the surface of small particulate, mineralized tissue debris, may act as an opsonin, thereby facilitating cell adhesion and phagocytosis by macrophages, a process likely mediated by integrin-binding, signal transduction, and cytoskeletal restructuring.

Amelogenin↗
PubMed1996

Immunocytochemical localization of Tamm-Horsfall protein in the kidneys of normal and nephrolithic rats.

Studies using in vitro systems have indicated that Tamm-Horsfall protein (THP) can interact with calcium oxalate (CaOx) crystals during kidney stone formation. However, information regarding the nature of its participation in this process remains controversial and unclear. In order to better understand the putative interaction of THP and crystals in vivo, we compared the localization of THP in normal rats and in chronic and semi-acute rat models of nephrolithiasis. In these rats, CaOx crystal deposits were induced in the kidneys by administering ethylene glycol (EG) in drinking water. The formation of CaOx mono- and dihydrate aggregates in the urine was confirmed by scanning electron microscopy. Immunohistochemical localization, as well as protein A-gold labeling at the ultrastructural level, demonstrated that in addition to its normal distribution, THP specifically associated with the renal crystal deposits. The THP-containing, organic matrix-like material consisted of a fine, fibrillar meshwork surrounding individual crystals and their aggregates. In addition, THP also appeared in the papilla, where it is normally absent, concurrent with the appearance of crystal deposits in the kidneys. These observations indicate that in nephrolithic rats the normal localization of THP is altered. Such an alteration may indicate an important physiological event related to crystal aggregation and kidney stone formation.

Acute Disease↗
PubMed1996

Management of humeral nonunion after the failure of locking intramedullary nails.

We reviewed 21 cases of humeral nonunion following the failure of "locking" humeral nails. The nails had been inserted as the primary operative procedure following humeral fracture in fifteen cases or after the failure of closed treatment in six cases. Reconstruction after the failure of these implants was complicated by poor bone stock and difficulty in achieving union. Although technically difficult, open reduction and internal fixation with plating and bone grafting (successful in nine of nine cases) was more consistent than exchange nailing (successful in four of 10 cases) in achieving union (p = 0.01). Two patients refused further surgical intervention. The degree of bone loss associated with a loose nail, the lack of success of exchange nailing, and the insertion site morbidity associated with humeral nail removal differentiate these nonunions from similar lower extremity problems. The degree of bone loss following failed locking nailing of the humerus is a major concern, and exchange nailing alone may not be an acceptable option to deal with this problem.

Adolescent↗