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M D Grounds

Publications and source records attributed to M D Grounds.

At least 37 records · Page 2Linked to original sources
PubMed1997

Macrophages and dendritic cells in normal and regenerating murine skeletal muscle.

Mononuclear phagocytes and MHC class II+ dendritic cells (DC) were identified in frozen sections of skeletal muscle using a panel of pan-specific antimacrophage (MOMA-2, SER-4, Mac-1, F4/80), anti-major histocompatibility complex (MHC) class II (M5/114) and anti-DC (NLDC-145, N418, M342) monoclonal antibodies. Uninjured and regenerating skeletal muscle were investigated in SJL/J and BALB/c mice, strains with known differences in muscle regenerative capacity. Resident tissue macrophages and MHC class II+ DC were present within uninjured mouse muscle. A subpopulation of DC were positive for the pan-DC markers, N418 and M342, and negative for the lymphoid DC marker NLDC-145. Following crush injury, the macrophage population increased by day 2, became marked by day 3, and had decreased by day 6. In contrast, the number of MHC class II+ cells around the injury site increased steadily after injury and remained high at day 6. The numbers of macrophages and DC detected by immunohistochemical staining were consistently higher in SJL/J than BALB/c muscles. This study confirms that macrophages are a significant component of normal murine skeletal muscle and that these cells increase dramatically after injury. Furthermore the data also reveal for the first time that DC are present in normal skeletal muscle and that MHC class II+ cells, including DC, increase after injury. The presence of DC in muscle has important implications for the understanding of the immunobiology of muscle and immune-mediated processes such as the host versus graft responses following muscle transplants and autoimmune diseases affecting this tissue.

Animals↗
PubMed1997

The host environment determines strain-specific differences in the timing of skeletal muscle regeneration: cross-transplantation studies between SJL/J and BALB/c mice.

The difference in the timing of the regeneration process of skeletal muscle between SJL/J and BALB/c mice was investigated using grafts of whole skeletal muscle (both autografts and allografts). Histological, autoradiographic and immunohistochemical techniques were used in the investigation. Infiltration of leucocytes into autografts, numbers of desmin-positive myogenic cells and myotube formation were all more advanced in the SJL/J compared with BALB/c mice. Furthermore, autoradiographic evidence showed that myoblasts in the SJL/J autografts were synthesising DNA 12 h earlier than myoblasts in BALB/c autografts. In allografts, where SJL/J host mice received BALB/c grafts, and vice versa, leucocyte infiltration and myotube formation occurred earlier in the BALB/c muscles grafted into SJL/J hosts, than in the reverse situation with BALB/c hosts. The results show that, at least for whole muscle grafts, it is the host environment which determines the speed and outcome of the regenerative process.

Animals↗
PubMed1997

In vitro assessment of the biological activity of basic fibroblast growth factor released from various polymers and biomatrices.

The kinetics of controlled release of basic fibroblast growth factor (bFGF) from polymers (sutures, polycarbonate, Hydron, and Elvax), biopolymers (alginate), and biomatrices (lens capsules), and conditions for storage of bFGF (temperature, plastic type, heparin) were evaluated in vitro. Tissue culture proliferation bioassays with 3T3 fibroblasts, showed that only lens capsules with bFGF had a sustained release of bFGF for up to three weeks. The other materials released all of the 'bound' bFGF with two hours or produced an inflammatory response in vivo. Therefore, the lens tissue had the most potential for controlled long-term delivery of bFGF in vivo. These studies emphasise the importance of in vitro analysis of release kinetics of growth factors from a range of materials as a basis for potential in vivo applications.

3T3 Cells↗
PubMed1996

Enhancement of neovascularization in regenerating skeletal muscle by the sustained release of erucamide from a polymer matrix.

The angiogenic agent erucamide (cis-13-docosenamide), incorporated into a polymeric biomaterial (Elvax 40P, a copolymer of ethylene and vinyl acetate), was used to determine whether angiogenesis can be increased in the regenerating skeletal muscle, and whether the enhanced revascularization improves the new muscle formation. The angiogenic nature of this lipid was confirmed in a rat cornea-micropocket assay, prior to insertion of small strips of the polymer containing either 3 micrograms, 300 micrograms erucamide or only polymer as a control into the mid-region of crush-injured tibialis anterior (TA) muscles of forty-five adult male BALB/c mice. All TA muscles were sampled ten days after injury and analyzed morphometrically. Statistical analyses of the mean blood vessel area density in lesions from twelve perfused TA muscles (three from each of the erucamide-treated or control group), revealed a dose-dependent angiogenic effect of erucamide: a dosage of 3 micrograms increased mean blood vessel area density to 5.1% compared to 2.0% in controls, due to numerous large caliber, thin-walled vessels, whereas the mean vessel area density in both the 30-micrograms (3.5%) and 300-micrograms (1.5%) doses were similar to controls. However, at all three doses tested, erucamide did not significantly alter the degree of new muscle formation, connective tissue deposition, or removal of necrotic debris.

Animals↗
PubMed1996

The exogenous administration of basic fibroblast growth factor to regenerating skeletal muscle in mice does not enhance the process of regeneration.

The effects, in vivo, of the exogenous administration of bFGF on myogenesis of regenerating skeletal muscle was assessed either morphometrically or autoradiographically in three separate models of muscle injury in mice: crush-injured, denervated, and dystrophic (mdx) muscles. The bFGF was administered at various doses and different time schedules, sometimes in combination with heparin, into injured tibialis anterior muscles of mice. Delivery of the bFGF was either by direct intramuscular injection or by the sustained release from 888polymers (Hydron or Elvax) implanted into the muscles. The bioactivity of bFGF was confirmed in vitro by measuring its ability to stimulate the proliferation of BALB/c-3T3 fibroblasts and muscle precursor cell lines. The ability of bFGF to stimulate angiogenesis in vivo was confirmed by the implantation of controlled-release polymers containing bFGF into the normally avascular cornea of rats. No measurable effect of bFGF was seen in any of the models of skeletal muscle injury under these experimental conditions, indicating that the availability of biologically active bFGF is not a limiting factor in the regeneration of skeletal muscle following injury.

Animals↗
PubMed1995

The time course of basic fibroblast growth factor expression in crush-injured skeletal muscles of SJL/J and BALB/c mice.

Staining for basic fibroblast growth factor (bFGF), a potent mitogen, was examined in muscle recovering from a crush injury and compared between two mouse strains with distinctly different capacities for muscle regeneration to determine if bFGF staining and the steps and outcome of repair were related. Immunofluorescence studies on intact and crushed tibialis anterior muscle were carried out at 0, 1, 3, 6, and 12 h postcrush in SJL/J mice, and at 1, 2, 3, 5, 7, and 11 days after injury in both SJL/J and BALB/c mice (n = 2-4). Disrupted fibers showed increased sarcoplasmic staining for bFGF as little as 3-6 h after injury prior to infiltration with intensely fluorescent mononuclear cells (at 12-24 h). Fiber bFGF was maintained in SJL/J muscles for 2 days, but was lower in most damaged fibers of BALB/c muscles at the same time. Surviving stumps of crushed fibers, once sealed, exhibited sarcoplasmic extensions, some of which stained intensely for bFGF. These processes appeared to connect adjacent fiber stumps, and many were noted in association with aligned mononuclear cells (presumptive myoblasts) at the site of new myotube formation. In representative sections there were more bFGF-positive mononuclear cells present in SJL/J than BALB/c muscles. Intense bFGF localization marked newly regenerating myotubes in both SJL/J and BALB/c muscles, and such myotubes were more frequent and larger in SJL/J muscles. More bFGF was present in regenerating muscles of SJL/J compared with BALB/c mice (with respect to damaged myofibers, mononuclear cells, and myotubes) and this correlates with the superior new muscle formation seen in SJL/J mice. These studies support the idea of a positive relation between bFGF in damaged fibers, the bFGF-positive mononuclear cells, and the speed and success of muscle regeneration.

Animals↗
PubMed1995

Extracellular matrix, growth factors, genetics: their influence on cell proliferation and myotube formation in primary cultures of adult mouse skeletal muscle.

Cell proliferation and myotube formation in response to growth factors on various extracellular matrices (ECM) were investigated in primary skeletal muscle cultures from adult SJL/J and BALB/c mice. There was no difference between the rates of proliferation from primary cultures of SJL/J and Balb/c mice measured at 48 h in response to a range of concentrations of PDGF-AA, -AB, -BB, TGF beta 1, or LIF (added at 24 h). SJL/J primary cultures were more responsive to bFGF (which was the most potent mitogen) than were BALB/c cultures. Comparison of dose response curves to bFGF and TGF beta 1 grown on gelatin or Matrigel showed that the nature of the ECM did not have a significant affect. More myotubes formed at 4 days in SJL/J than in parallel BALB/c cultures on gelatin or Matrigel (P < 0.05). On gelatin more myotubes with 4 or more nuclei were formed in cultures from SJL/J than BALB/c muscles (P < 0.05); however, on Matrigel these myotubes occurred with similar frequency. Myotube formation examined in BALB/c muscle cultures grown on collagen i.v., entactin-free laminin, and fibronectin showed that none of these ECM components alone supported large myotube formation (4 or more nuclei) as well as did Matrigel, although fibronectin was as effective as Matrigel with respect to the total number of myotubes formed. Parallel experiments carried out using the myogenic H-2Kb(27) cell line showed similar effects with the exception of laminin which enhanced large myotube formation and desmin expression in the H-2Kb(27) but not in the primary muscle cultures. The greater sensitivity in mitogenic response to bFGF and the more extensive myotube formation seen in SJL/J compared with BALB/c cultures in vitro reflects the superior capacity for muscle regeneration of SJL/J mice in vivo.

Analysis of Variance↗
PubMed1995

Retarded myogenic cell replication in regenerating skeletal muscles of old mice: an autoradiographic study in young and old BALBc and SJL/J mice.

The patterns of skeletal muscle precursor cell replication after crush injury were compared by the use of autoradiographic techniques, in young (4-week-old) and old (39-week-old) BALBc and SJL/J mice. Similar comparisons were made between cut and crush lesions in old BALBc muscle. Muscle precursor cell replication commenced at 18-24 h after injury in both young and old muscles from both strains of mice. In young BALBc muscle the peak of myogenic activity at 60 h was 36 h earlier than in old mice. SJL/J muscle responded more rapidly than did BALBc: in young SJL/J the peak myogenic activity was at 46 h (14 h earlier than in young BALBc muscle), and in old SJL/J muscle the peak activity at 72 h was 24 h earlier than in old BALBc muscle. In all mice (both young and old) myogenic cell replication was substantially reduced by 120 h after injury. A comparison of the timing of muscle precursor cell replication in cut and crush lesions in old BALBc mice revealed a more rapid response in the cut lesion; this difference between the lesions is comparable with data from identical lesions in 6-8-week-old BALBc mice (McGeachie and Grounds 1987). However, the peak of myogenic replication in the older mice in the present study was some 26-36 h later than in the younger 6-8-week-old mice. These experiments show that, whilst muscle precursor cell replication commences at approximately the same time (about 24 h) after injury in young and old mice, the peak level of activity is delayed by some 24-36 h in old mice. In addition, the SJL/J mouse strain responds more rapidly and prolifically to muscle injury than does the BALBc strain.

Aging↗
PubMed1995

The genotype of bone marrow-derived inflammatory cells does not account for differences in skeletal muscle regeneration between SJL/J and BALB/c mice.

This study determined whether the genotype of bone marrow-derived inflammatory cells contributes to the more pronounced leukocytic exudation and extensive new muscle formation seen in SJL/J compared with BALB/c mice after a crush-injury (Mitchell et al. 1992). Female SJL/J mice were whole-body irradiated and reconstituted with male bone marrow from the BALB/c strain, and irradiated BALB/c females reconstituted with male SJL/J bone marrow. The mice were allowed to recover for 3 weeks and the tibialis anterior muscle (in a leg which had been protected from irradiation) was injured by crushing. At 3 and 10 days after injury the extent of necrotic debris, mononuclear leukocytic infiltration and new muscle formation was assessed in the muscles. The SJL/J mice reconstituted with BALB/c bone marrow showed extensive mononuclear leukocytic infiltration and clearance of necrotic debris when compared with BALB/c mice reconstituted with SJL/J bone marrow, and these strain-specific differences mirrored those seen with control bone marrow reconstituted hosts and non-irradiated hosts. The results show that the genotype of the bone marrow-derived macrophages is not responsible for the superior regeneration of crush-injured skeletal muscle in SJL/J mice, and it appears that factors intrinsic to the muscle tissue may be of central importance.

Animals↗
PubMed1994

Intrinsic differences in MyoD and myogenin expression between primary cultures of SJL/J and BALB/C skeletal muscle.

The time course of expression of the skeletal muscle-specific regulatory genes MyoD and myogenin was studied in primary cultures of skeletal muscle from adult SJL/J and BALB/c mice. In situ detection of expression with MyoD and myogenin riboprobes and myogenin antibody showed that the onset of expression of these genes occurred earlier in cells from SJL/J mice. Probe-positive cells and myotubes were also more frequent in cultures from SJL/J mice than in BALB/c. The onset of expression of MyoD and myogenin was delayed in cultured cells relative to the time course seen following injury in vivo. Myogenin protein was demonstrated in replicating cells and all myogenin-positive cells expressed desmin. The observed strain-specific difference infers a greater intrinsic myogenicity of cells in SJL/J muscle in vitro and reflects the superior capacity for new muscle formation previously reported in SJL/J mice in vivo.

Animals↗
PubMed1993

The role of macrophages in skeletal muscle regeneration with particular reference to chemotaxis.

The results from this investigation suggest that chemotactic factor(s) from damaged myofibers attract polymorphonuclear leukocytes (PML) and macrophages to the site of injury, while exudate macrophages but not PML induce a strong positive chemotactic response in myogenic cells. The AB and BB isoforms of platelet-derived growth factor (PDGF), transforming growth factor-beta (TGF-beta), basic fibroblast growth factor (bFGF), and leukemia inhibitory factor (LIF) (all of these are secreted by macrophages) were also shown to be chemoattractants for muscle precursor cells (MPC). The AA isoform of PDGF did not appear to have any such chemotactic effect on MPC. Macrophages were also shown, under tissue culture conditions, to stimulate the proliferation of MPC. This mitogenic effect was similarly demonstrated for the BB isoform of PDGF, bFGF, and TGF-beta but not for the AA or AB isoforms of PDGF nor for LIF. The results indicate that macrophages play a pivotal role in the orchestration of muscle fiber reconstitution.

Animals↗
PubMed1993

Age-related changes in replication of myogenic cells in mdx mice: quantitative autoradiographic studies.

Cell replication in muscle was measured by tritiated thymidine (3H-TdR) incorporation and autoradiography, in mdx mice from 2-44 weeks of age. Pre-mitotic labelling (within 1 h of 3H-TdR injection) was determined in 16 mice aged from 15 to 300 days. In 30 further mdx mice, one leg was irradiated 1 h after 3H-TdR injection to block DNA synthesis. Post-mitotic labelling was measured in both legs 10-15 days later. Between 20 and 60 days of age a very high proportion (up to 2%) of muscle (satellite cell) nuclei were replicating pre-mitotically; from 80-300 days cell replication was detectable but at much lower levels. Centrally placed nuclei within muscle fibres appeared at 24 days, increased rapidly to 50% by 50-100 days, declining thereafter to 25% at 300 days. In post-mitotic samples, labelled myotubes and labelled peripheral muscle nuclei (satellite cell nuclei and myonuclei) appeared at 28 days and were present in the mdx muscles through to 310 days, indicating continued cell replication and muscle regeneration. Myogenic cell replication was both retarded and inhibited by irradiation. These data demonstrate that muscle cell replication in mdx mice commences at about 3 weeks of age, is maximal at 4-8 weeks, but continues at lower levels until at least 44 weeks.

Aging↗
PubMed1993

The migration and intermixing of donor and host glia on nitrocellulose polymers implanted into cortical lesion cavities in adult mice and rats.

The fate of neonatal glia (mostly glial fibrillary acidic protein-positive astrocytes), cultured on nitrocellulose papers and implanted into cortical lesion cavities, was examined in adult mice and rats. In mice, a Y-chromosome-specific probe and in situ hybridization techniques were used to identify male cells. Male-female grafts allowed visualization of donor glia and their behaviour after transplantation; female-male grafts allowed an analysis of how host cells responded to the presence of the implants. There was substantial intermixing of cells, with many donor glia migrating away from the implants and host cells migrating onto both sides of the nitrocellulose paper. In rats, donor glia were labelled with fluorescein-conjugated latex microspheres prior to transplantation on nitrocellulose polymers. The rat data were broadly consistent with those obtained from the mouse; moreover, immunohistochemical studies in rats suggested that the majority of host cells migrating onto the previously cell-coated papers were astrocytes. In a number of studies, glia-coated polymers have been used in an attempt to promote the regrowth of axons across lesion sites in the brain and spinal cord. The present work suggests that both transplanted and host glia may influence the regenerative growth seen in such implants.

Animals↗
PubMed1993

Fusion of myogenic cells to the newly sealed region of damaged myofibres in skeletal muscle regeneration.

In regenerating skeletal muscle, sarcoplasmic extensions containing variable numbers of nuclei, widely referred to as 'buds' or 'stumps', are formed at the ends of damaged myofibres. In this paper we investigated whether the nuclei seen in the buds results from fusion of myogenic cells or from migration of myonuclei to the sealed ends of damaged myofibres in murine muscle regenerating after crush injury. The fusion of mononuclear and multinucleate myogenic cells to the buds was demonstrated by transmission electron microscopy. In order to elucidate the frequency and kinetics of cytoplasmic continuity between myotubes and sealed myofibres, we labelled the damaged myofibres with carbocyanine dye DiI (which inserts into the lipid bilayer and travels down continuous membranes) and the samples were then examined by confocal scanning microscopy. This technique showed that there was little fusion between myotubes and myofibres during the first 6 days after crush injury, but significant fusion had occurred by the tenth day especially at the newly sealed region of the damaged myofibre. A scheme for the repair of damaged skeletal muscle is presented.

Animals↗
PubMed1993

Studies on the structure of complement C3 and the stability of C3 derived phagocytic ligands C3b/iC3b in SJL/J and BALB/c mice.

Female SJL/J mice are more susceptible to development of experimental autoimmune myositis than most other mouse strains. Since complement has been implicated in the pathogenesis of inflammatory muscle disease in humans, quantitative and qualitative studies of complement C3 were undertaken in SJL/J and BALB/c mice to determine whether complement may influence disease susceptibility in SJL/J mice. In accordance with previous studies, mature male and female BALB/c mice were shown to have similar serum C3 concentrations. However, differences were found between mature male and female SJL/J mice. Male SJL/J mice have significantly higher serum C3 concentrations than SJL/J females and both sexes of BALB/c mice suggesting that serum C3 concentration may be variably influenced by sex in some mouse strains. Qualitatively, SJL/J mice were shown to have a different allotypic form of C3 (C3F) compared to the common electrophoretically slow form (C3S) found in BALB/c mice and most other mouse strains. Furthermore, studies on the decay rate of C3 revealed that C3b/iC3b fragments are converted to C3c/d at a faster rate in sera from female SJL/J mice compared to female BALB/c mice. Because removal and solubility of immune complexes is influenced by complement C3, it is possible that the more rapid decay of the phagocytic ligands C3b/iC3b may account for the increased susceptibility to development of autoimmune disease in female SJL/J mice.

Animals↗
PubMed1993

Comparison of basic fibroblast growth factor in X-linked dystrophin-deficient myopathies of human, dog and mouse.

Binding of polyclonal antibodies specific for bFGF was examined in tissue sections of myopathic and normal muscles from humans, dogs and mice. The proposal tested was that differences in the amount or distribution of bFGF in muscles of the 3 species, might correlate with the limited muscle regeneration seen in humans and dogs afflicted with x-linked muscular dystrophy, in contrast with the sustained new muscle formation in mdx mice with the homologous myopathy. There was a striking difference between the species in the binding of bFGF antibodies to extracellular matrix, particularly at the periphery of myofibres; binding was pronounced in mouse but weak or absent in human and dog muscle. Binding to muscle nuclei and sarcoplasm was also stronger in mice than in humans and dogs, and in all species was more pronounced in foetal than adult muscle. Increased binding of bFGF antibodies was seen in damaged and regenerating muscle cells in all myopathic specimens where these were present. This was associated with the regenerative process rather than with myopathy, as a similar pattern of bFGF expression was seen in mouse muscle regenerating after experimental crush injury. The higher extracellular staining for bFGF around the periphery of mouse myofibres correlated with the successful muscle regeneration in dystrophic mice. Results suggest that bFGF at the fibre periphery might stimulate a local increase in the numbers of muscle precursor cells which can respond to injury in the mdx mouse.

Adolescent↗