Biomedical subjects

J F Marshall

Publications and source records attributed to J F Marshall.

At least 55 records · Page 3Linked to original sources

PubMed1998

Expression of alphav integrins and vitronectin receptor identity in breast cancer cells.

In the present study we have used fluorocytometry and immunoprecipitation to characterize the expression of alphav-containing integrins in a panel of eight human breast cancer cell lines and one normal human mammary epithelial line. We show that the classical vitronectin receptor alphavbeta3 is expressed in only one cell line (MDA-MB-231), whereas alphavbeta5 is expressed on all breast cancer cell lines and alphavbeta1 is expressed on the majority. Using adherence assays to purified ligands in the presence and absence of function-blocking monoclonal antibodies, we have demonstrated that alphavbeta5 mediates adhesion to vitronectin in the majority of these cells. In one cell line, ZR75-1, alphavbeta1 contributes significantly to adhesion to immobilized vitronectin. The formation of focal adhesions containing the alphav and beta1 subunits on vitronectin is also demonstrated by indirect immunofluorescence.

Breast Neoplasms↗

PubMed1998

The human hyaluronan receptor RHAMM is expressed as an intracellular protein in breast cancer cells.

The receptor for hyaluronan mediated motility (RHAMM) has been reported to mediate migration, transformation, and metastatic spread of murine fibroblasts. Here we describe the expression of two human RHAMM isoforms, which are generated by alternative splicing of the primary gene transcript, by a series of human breast carcinoma cell lines. A polyclonal antibody, raised against a bacterially expressed RHAMM fusion protein, detected an 85-90 kDa protein by western blot analysis. No correlation between the level of RHAMM mRNA and protein expression with known metastatic/malignant potential of the tumour cell lines was observed. Interestingly, we found that the antibody did not stain the cell surface but the cytoplasm of breast cancer cells. The intracellular localisation of RHAMM was confirmed by subcellular fractionation studies. RHAMM proteins were capable of binding to hyaluronan, but not to heparin or chondroitin sulphate, in an vitro binding assay. We also provide evidence that a potential hyaluronan-binding motif in the N terminus of the protein is not involved in the interaction of RHAMM with hyaluronan. Our findings lead us to conclude that RHAMM does not function as a conventional motility receptor for HA in human breast cancer cells and we suggest the term RHAMM be substituted by 'intracellular hyaluronic acid binding protein' (IHABP).

Alternative Splicing↗

PubMed1997

Activation status and function of the VLA-4 (alpha4beta1) integrin expressed on human melanoma cell lines.

We have examined the functional status of the VLA-4/alpha4beta1 integrin in a panel of human melanoma cell lines, focusing on the ability of cells expressing alpha4beta1 to mediate adhesion to the alpha4-specific ligands CS-1 peptide and VCAM-1. All melanoma cells expressing alpha4pbeta1 (8 of 10 lines examined) were capable of adhering to these specific ligands in adhesion assays, whereas 2 cell lines (HMB2 and VUP) which lacked surface alpha4 were unable to do so. Adherence of different melanoma cell lines to VCAM-1 was relatively uniform and not susceptible to upregulation with known integrin-activating factors, such as manganese ions, phorbol ester and activating monoclonal antibody (mAb) TS2/16. Cell adhesion to CS-1 peptide, however, varied according to cell surface receptor density and, in some cases, could be up-regulated by integrin-activating factors. Adhesion of SK23 cells to CS-1 peptide was increased by all 3 activating stimuli, whereas for all other melanoma cells an increase was obtained only by the use of TS2/16 mAb. Our data indicate not only an unusually low activation state of alpha4beta1 in SK23 cells but also heterogeneity in the activating capacity of the various stimuli. Moreover, a protein kinase C-dependent role in alpha4beta1 activity was suggested by adhesion assays carried out in the presence of the protein kinase C inhibitor calphostin C, which considerably reduced adhesion to CS-1 peptide.

Anti-Allergic Agents↗

PubMed1997

Modulation of human stromelysin 3 promoter activity and gene expression by human breast cancer cells.

The matrix-degrading enzyme family of matrix-metalloproteinases (MMPs) has been implicated in the process of tumour metastasis. Cellular protein and RNA localisation techniques have been used to show that, whilst several MMP genes are expressed in both cancer and stromal cells, stromelysin 3 is expressed only in stromal fibroblasts adjacent to cancer cells. Immunohistochemical and in situ hybridisation evidence suggests that neoplastic cells can stimulate stromal cell MMP production either in a paracrine fashion or by a cell-cell contact mechanism. Using 2 different lengths of the human stromelysin 3 (ST3) gene 5' flanking sequence cloned upstream of luciferase and CAT reporter genes, we now show that human breast cancer cells can directly activate the ST3 promoter. The putative response element in the ST3 promoter, which lies between 0.46 and 3.4 kb upstream of the transcription start site, is able to effect a 2- to 3-fold increase in downstream gene expression. We further show that this transcriptional up-regulation definitely occurs via a paracrine, and possibly via a cell-cell contact, mechanism. Confirmation that this ST3 promoter activation results in ST3 gene induction of a similar magnitude was shown using Northern blotting of stimulated fibroblasts. Our data provide further evidence that cancer cells can induce fibroblast MMP expression and help to explain the in vivo expression pattern of ST3 in breast cancer.

3T3 Cells↗

PubMed1997

Reduced eticlopride-induced Fos expression in caudate-putamen and globus pallidus after unilateral frontal cortex injury: relation to neglect.

Unilateral ablation of medial agranular cortex in rats results in neglect of contralateral stimuli and reductions in amphetamine-induced expression of the immediate early gene, c-fos, in both caudate-putamen and globus pallidus. Both unilateral neglect and the reductions in dopamine agonist induction of subcortical Fos immunoreactivity dissipate over a matter of weeks. Dopamine agonism induces Fos predominantly in striatonigral cells and in globus pallidus via striatopallidal disinhibition, whereas Fos is induced in striatopallidal cells by administration of antagonists of the D2 dopamine receptor subfamily. To examine more directly effects of cortical injury on striatopallidal function, induction of striatal Fos by the D2 antagonist eticlopride (0.1 mg/kg, s.c.) was examined in rats with medial agranular cortex ablation. In the same animals, eticlopride-induced Fos in globus pallidus was also examined. Five days after unilateral cortex injury, in rats showing neglect, the numbers of Fos immunoreactive nuclei induced by eticlopride were reduced by 50% in caudate-putamen and 25% in globus pallidus of the ipsilateral hemisphere. These lesion effects were restricted to dorsolateral caudate-putamen and dorsal pallidum. Three or more weeks after cortical injury, in rats recovered from neglect, eticlopride-induced Fos was normalized in caudate-putamen, but still decreased by 20% in globus pallidus. Along with previous findings, these results suggest that behavioral recovery from neglect produced by cortical injury may be at least partially mediated by normalizations of function of both striatopallidal and striatonigral neurons. In addition, the present findings suggest that normalization of function of pallidal cells activated by eticlopride is not necessary for behavioral recovery from frontal cortex ablation. Lingering reductions in excitatory cortico-subthalamo-pallidal input may be responsible for the longer-lasting dysfunctions of these pallidal cells.

Animals↗

PubMed1997

Differing influences of dopamine agonists and antagonists on Fos expression in identified populations of globus pallidus neurons.

Dopamine agonists increase the activity of globus pallidus neurons, as shown electrophysiologically and with Fos expression. More recently it has been shown that decreased D2 receptor activity also causes pallidal Fos expression. Similar responses occur in the striatum, where both dopamine agonists and D2 blockade induce Fos, although in separate neuronal populations (i.e. striatonigral and -pallidal). The present experiments investigate the possible differential dopaminergic regulation of Fos within pallidal neuronal populations, which were classified as parvalbumin-positive or -negative (with parvalbumin immunostaining), or as projecting to various target nuclei (with retrograde transport of FluoroGold iontophoresed into these nuclei). Rats with prior nigrostriatal lesions received saline, D1 agonist, or D2 agonist. Rats with no lesions received saline, combined D1/D2 agonists, or the D2 antagonist eticlopride. Two hours after drug injection, rats were perfused and their brains processed for double-labeling: either Fos staining with parvalbumin staining, or Fos or parvalbumin staining in FluoroGold-labeled sections. Overall, dopamine drug treatments induced more Fos in parvalbumin-negative than -positive cells. However, unlike dopamine agonists, eticlopride induced significant Fos only in the parvalbumin-negative cells. Dopamine agonist-induced Fos was found in pallidal neurons projecting to each of the target nuclei examined, in both normal and nigrostriatal-lesioned rats. Eticlopride-induced Fos occurred only in pallidal neurons projecting to the striatum, providing functional evidence for pallidostriatal cells without axon collaterals to other nuclei. It was also found that pallidostriatal neurons were distinguished from other projection populations by a relative lack of parvalbumin immunoreactivity. Pallidal cells respond heterogeneously to dopaminergic treatments based on their projection target and expression of parvalbumin. The smaller Fos responses in parvalbumin-containing cells may be due largely to the calcium buffering by the parvalbumin itself. Also, the pattern of Fos expression in pallidostriatal neurons suggests that dopamine regulates activity in these cells differently than in other projection populations.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗

PubMed1997

Integrin alpha5beta1-mediated adenovirus infection is enhanced by the integrin-activating antibody TS2/16.

Adenovirus internalization generally has been accepted to involve an interaction of the adenoviral penton base protein with alpha(v)beta3 and alpha(v)beta5 cell surface integrins. In this study we show that exposure of a panel of melanoma cells to the beta1-activating antibody TS2/16 rendered such cells more susceptible to adenovirus infection. This increase in adenoviral infectivity paralleled effects on cell adhesion, and both these characteristics were mediated, in part, by the alpha5beta1 integrin. These observations suggest that alpha5beta1 may act as an alternative adenovirus receptor and that integrin-activating strategies may improve the efficacy of recombinant adenoviruses as vectors for gene therapy.

Adenoviridae↗

PubMed1996

Intrastriatal AP5 differentially affects behaviors induced by local infusions of D1 vs. D2 dopamine agonists.

Using bilateral infusions into the rat striatum, the effects of the competitive NMDA receptor antagonist 2-amino-5-phosphopentanoic acid (AP5) on behaviors induced by the dopamine (DA) D1 receptor agonist SKF 82526 (fenoldopam) or the D2 receptor agonist quinpirole were determined. These effects were tested in DA-replete (intact) rats and in rats that were receiving injections of the monoamine-depleting drug reserpine. In both intact and reserpinized rats, fenoldopam induced significant sniffing. This effect was attenuated by simultaneous co-infusion of AP5 in the reserpinized rats. Quinpirole induced locomotion, sniffing, and oral behaviors, all of which were attenuated by AP5 co-infusion in the intact rats. In contrast, AP5 enhanced the quinpirole-induced sniffing of reserpinized rats. These findings suggest that distinct D1/glutamate and D2/glutamate relationships exist in the striatum, and that the nature of the latter is influenced by DA tone.

2-Amino-5-phosphonovalerate↗

PubMed1996

Chronic L-dopa alters striatal NMDA receptors in rats with dopaminergic injury.

To examine the effects of chronic L-dopa treatment on excitatory amino acid receptors, rats with unilateral nigrostriatal injury and intact controls were treated for 21 days with Sinemet (L-dopa+carbidopa) or vehicle followed by a 3-day washout period. Nigrostriatal damage induced by unilateral injection of 6-hydroxy-dopamine (6-OHDA) produced an extensive decline in [3H]mazindol binding in the ipsilateral caudate-putamen (CPu) and a small (7%) decline in [3H]glutamate binding to N-methyl-D-aspartate (NMDA) receptors in CPu. Sinemet treatment of 6-OHDA-injected rats further reduced the binding of [3H]glutamate to NMDA receptors. The greatest reductions (-34%) occurred in the denervated CPu, but moderate declines (-18% to -22%) were also observed in the CPu, nucleus accumbens and cingulate cortex of the intact hemisphere. Unexpectedly, chronic Sinemet treatment also caused a decrease in [3H]mazindol binding in both hemispheres of rats receiving unilateral 6-OHDA injections. L-Dopa/ carbidopa treatment did not affect [3H]glutamate or [3H]mazindol binding in neurologically intact rats.

Animals↗

PubMed1996

Identification by differential display of annexin-VI, a gene differentially expressed during melanoma progression.

To identify genes involved in the melanocyte to malignant melanoma conversion, we have applied differential display to the comparison of syngeneic murine B16F10 (metastatic melanoma) and Melan-a-immortalized melanocyte cell lines. Approximately 7000 bands were analyzed, revealing approximately 80 to be differentially displayed. Reverse Northern blotting and subsequent Northern blotting confirmed the reproducible differential expression of four transcripts. Three B16F10-specific bands encode novel genes or partially sequenced cDNAs of unknown function. One Melan-a-specific band was found to be identical to the 3' end region of the mouse Annexin VI mRNA and shown to have a reduced message expression in B16F10 relative to Melan-a. Differential expression was confirmed at the protein level with Western blotting using a rabbit polyclonal antiserum. Immunohistochemistry of human melanoma specimens with this antiserum revealed a decrease or loss of Annexin VI expression as melanomas progressed from a benign to a more malignant phenotype. Our results provide further evidence for a potential role of Annexin VI in tumor suppression.

Animals↗

PubMed1996

Cerebrocortical Fos expression following dopaminergic stimulation: D1/D2 synergism and its breakdown.

Using standard immunohistochemical techniques, we examined Fos expression in different areas and layers of cerebral cortex in rats following combined or separate stimulation of dopamine D1 and D2 receptors under normal conditions and following five days of reserpine (1 mg/kg/day), a treatment that causes a breakdown in requisite D1/D2 synergism. In normal animals, combined but not separate stimulation of D1 and D2 receptors elicited Fos expression in frontal and parietal, but not cingulate, cortex. Expression was highest in layer IV of primary somatosensory cortex; in frontal and secondary somatosensory cortex, Fos expression was lower and peaked in layer VI. Cortical Fos expression following amphetamine showed the same general pattern, and was blocked by either a selective D1 or D2 antagonist. Following reserpine treatment, stimulation of either D1 or D2 receptors gave rise to cortical Fos expression in patterns similar to each other and to combined D1/D2 stimulation in normal rats (except in frontal cortex in which separate D1 or D2 stimulation was unable to elicit Fos even following repeated reserpine treatment). The fact that cortical Fos expression was tightly associated with behavioral activation together with its laminar and areal distribution suggest that sensory input resulting from behavioral activation may be an important stimulus for this immediate-early gene response.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗

PubMed1996

Amphetamine-induced Fos expression in globus pallidus is altered by frontal cortex injury.

Functional recovery from cortical injury may result from subcortical compensatory processes. This study examined basal gangliar expression of the immediate early gene c-fos after unilateral medial agranular cortex (AGm) ablation. In the ipsilateral dorsal globus pallidus of rats demonstrating neglect of contralateral stimuli (sacrificed 5 days postinjury), the numbers of amphetamine-induced Fos-positive nuclei were reduced 37% compared to intact hemisphere values. These reductions were no longer apparent in recovered AGm-ablated rats (sacrificed 21 + days postinjury). These findings mirror in timing and direction the changes in Fos seen in dorsolateral striatum after AGm ablation.

Acoustic Stimulation↗

PubMed1996

Striatal and cortical NMDA receptors are altered by a neurotoxic regimen of methamphetamine.

Methamphetamine (m-AMPH) treatment produces long-lasting damage to striatal and cortical monoaminergic terminals and may also injure nonmonoaminergic cortical neurons. Evidence suggests that both dopamine (DA) and glutamate (GLU) play crucial roles in producing this damage. We used quantitative autoradiography to examine [3H]mazindol ([3H]MAZ) binding to striatal DA transporters and [3H]GLU binding to N-methyl-D-aspartate (NMDA) receptors in the striatum and cortex 1 week and 1 month after a neurotoxic regimen of m-AMPH. Rats received m-AMPH (4 mg/kg) or saline (SAL) (1 ml/kg) in four s.c. injections separated by 2 h intervals. One week after m-AMPH, the ventral and lateral sectors of the striatum showed the greatest decreases in both [3H]MAZ and [3H]GLU binding, while the nucleus accumbens (NA) showed no significant decreases. One month after m-AMPH, striatal [3H]MAZ binding was still significantly decreased, while NMDA receptor binding had recovered. Surprisingly, the parietal cortex showed a m-AMPH-induced increase in NMDA receptor binding in layers II/III and IV 1 week after m-AMPH and only in layers II/III 1 month after m-AMPH. The prefrontal cortex showed no m-AMPH-induced changes in NMDA receptor binding at either time point. This is the first demonstration that a regimen of m-AMPH that results in long-lasting damage to DA terminals can alter forebrain NMDA receptor binding. Thus, repeated m-AMPH treatments may produce changes in glutamatergic transmission in selected striatal and cortical regions.

Animals↗

PubMed1996

Frontal cortex ablation reversibly decreases striatal zif/268 and junB expression: temporal correspondence with sensory neglect and its spontaneous recovery.

This study's goal is to identify subcortical adaptations that may contribute to recovery of function following cortical injury. After unilateral aspiration of the medial agranular region of frontal cortex (AGm), rats demonstrate neglect of contralateral stimuli and recover within 3-4 weeks. Previous studies indicate that compensatory neural alterations involving dopamine (DA) occur following this cortical injury and that recovery from neglect produced by frontal injury is accompanied by normalization of glucose utilization within subcortical structures including the basal ganglia. The current study examined Zif and JunB, IEG protein products constitutively expressed in striatum, rendering it possible to investigate the effects of unilateral AGm ablation on striatal function during unstimulated as well as amphetamine-stimulated conditions. Five days after surgery, when contralateral neglect was still evident, the numbers of Zif-like or Jun-like immunoreactive (IR) nuclei in the ipsilateral striata of AGm-ablated rats were reduced. These lesion effects were similar for both constitutive and amphetamine-stimulated IEG expression and were restricted to the dorsolateral caudate-putamen, where excitatory input from AGm is most dense. In contrast, 3 or more weeks after AGm ablation, in rats demonstrating recovery, normal striatal Zif- and JunB-like immunoreactivity occurred. Thus, striatal zif/268 and junB expression is reduced 5 days after AGm injury in rats demonstrating neglect and normalized 3 or more weeks later in recovered rats. These findings indicate that adaptations involving the striatal medium spiny neuron, a site of convergence of cortical glutamatergic and nigral dopaminergic afferents, may contribute to behavioral recovery following neocortical injury.

Animals↗

PubMed1996

Functional and anatomical evidence for different dopamine dynamics in the core and shell of the nucleus accumbens in slices of rat brain.

The characteristics of dopamine (DA) uptake and release were compared in the core and shell of the nucleus accumbens (NAc). DA release was elicited from rat brain slices by local electrical stimulation, and its extracellular concentration was monitored with fast-scan cyclic voltammetry using Nafion-coated, carbon-fiber microelectrodes. The voltammetric results show that the values of DA release and uptake in the shell NAc are approximately one-third of those measured in the core region, and DA uptake in the shell was less sensitive than the core to inhibition by either cocaine or nomifensine. The density of [3H]mazindol binding sites in the NAc was examined by autoradiography and the shell was found to have an average of half the number of DA uptake sites measured in the core region. This combination of anatomical and functional results shows that DA neurotransmission in the shell NAc is distinct from that in the core region. These data are consistent with the view that multiple functional forms of the DA transporter, exhibiting disparate kinetics and pharmacology, exist in different brain regions that exhibit disparate kinetics and pharmacology. Different forms of the transporter, combined with different release kinetics and auto- and heteroreceptor activity, give a vast range of possibilities for regional variation in DA neurotransmission.

Animals↗

PubMed1996

The role of alpha v-integrins in tumour progression and metastasis.

Metastatic tumour spread is a pathologic process which can be described as altered cell growth associated with a series of adhesion/de-adhesion events which are coupled with regulated tissue degradation. Adhesion to, and migration through, the extracellular matrix (ECM) is necessary for the tumour invasion which is an important part of the metastatic process. Efficient proteolytic digestion of the molecules of the ECM appears to be facilitated by the localization of proteases at the cell surface-tissue interface (see also review by Stetler-Stevenson). Indeed, there is evidence from in-vitro studies to suggest that restriction of these enzymes to focal contact sites (areas of cell-substratum contact) may occur and that this sub-cellular juxtaposition of receptors and enzymes co-ordinates regulation of adhesion and proteolysis by the neoplastic cells. How such co-ordinated regulation is achieved and how this dynamic interplay is controlled during tumour development and progression are important areas of investigation.

Antigens, CD↗

PubMed1996

Unilateral frontal cortex ablation producing neglect causes time-dependent changes in striatal glutamate receptors.

This study's goal is to identify adaptations involving striatal glutamate (GLU) or dopamine (DA) receptors that may contribute to recovery of function following cortical injury. Unilateral aspiration of the medial agranular region of frontal cortex (AGm) in rats produces neglect of contralateral stimuli. Pharmacological and immunocytochemical studies suggest that glutamatergic and dopaminergic processes within striatum may contribute to spontaneous recovery from this neglect. This study examined by autoradiography radioligand binding to striatal GLU and DA receptor subfamilies in AGm-ablated rats surviving 5 days (unrecovered) or 3 or more weeks (recovered) postsurgery. Density of radioligand binding was quantified in striatal subregions by computerized image analysis. Compared to striatal binding densities in the intact hemisphere, [3H]kainate binding and [3H]GLU binding to NMDA receptors were decreased in the lesioned hemisphere of unrecovered AGm-ablated rats, but normalized (for kainate) or increased (for NMDA) in the lesioned hemisphere of recovered rats. Ablation of AGm did not affect [3H]AMPA binding or the binding of [3H]SCH23390, [3H]spiperone, or [3H]mazindol to dopaminergic D1 or D2 receptor subfamilies, or to DA uptake sites, respectively. The results suggest that a small percentage of NMDA and kainate receptors are located on corticostriatal axon terminals, and that over time an upregulation of striatal NMDA and/or kainate receptors may offset the loss of cortical glutamatergic input caused by cortical injury. These time-dependent alterations in GLU receptors may contribute to the recovery of function and normalizations of immediate early gene expression seen weeks after AGm ablation. Upregulation of striatal dopamine receptors was not evident, and thus is unlikely to mediate recovery from neglect produced by cortical injury.

Animals↗

PubMed1995

Intrastriatal DNQX induces rotation and pallidal Fos in the 6-OHDA model of Parkinson's disease.

The 6-hydroxydopamine rat model of Parkinson's disease was combined with intracerebral drug infusions to examine the influence of glutamate receptors on striatal output activity. When infused into the dopamine-denervated striatum, the AMPA-kainate receptor antagonist DNQX dose-dependently elicited contralateral rotation and ipsilateral Fos immunoreactivity (Fos-IR) in the globus pallidus, a target nucleus of striatal output. DNQX did not elicit rotation or Fos-IR in unlesioned or partially lesioned rats. In addition, the NMDA receptor antagonist AP-5 failed to induce rotation and had minimal effects on pallidal Fos-IR in lesioned rats. These results suggest a role for striatal AMPA-kainate receptors in the pathology and treatment of Parkinson's disease.

Animals↗