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Biomedical subjects

J B McCarthy

Publications and source records attributed to J B McCarthy.

At least 55 records · Page 3Linked to original sources
PubMed1996

CD44 expression in benign and malignant nevomelanocytic lesions.

CD44 is an integral membrane glycoprotein that is a principal receptor for hyaluronan and plays a role in cell-extracellular matrix interactions. Recent studies of melanomas in mouse models have suggested that increased CD44 expression by these tumors may relate to metastatic potential. Immunohistochemical expression of CD44 (standard [s] and variant [v6]) in benign and malignant nevomelanocytic lesions was assessed in formalin-fixed, paraffin-embedded tissue and was correlated with histological parameters and prognostic factors. Cases included benign nevi (three junctional, four compound, five intradermal, five blue, six Spitz, one deep penetrating), architecturally disordered (dysplastic) nevi (three, and primary (22) and metastatic melanomas (eight). All of the benign lesions showed diffuse and essentially uniform membrane staining of CD44s in nevomelanocytic cells, regardless of lesion size, depth, or extent of dermal involvement. In contrast, semiquantitative analysis (0 to 3+) of the primary melanomas showed heterogeneous and decreased staining of CD44s, which inversely correlated with lesion size (-0.569) and depth of invasion (-0.622 and -0.617 for Breslow's depth and Clark's level, respectively). These results were significant at P < .05. CD44s expression in metastases paralleled that of their respective primaries. None of the benign nevomelanocytic lesions showed CD44v6 staining. In contrast, all of the malignant nevomelanocytic lesions showed cytoplasmic staining of the tumor cells. Pretreatment with chondroitinase did not alter CD44s staining. CD44s expression by immunohistochemical determination is uniform in benign nevomelanocytic lesions. Malignant melanomas show decreased, heterogeneous staining that inversely correlates with increasing size, depth, and level of invasion. CD44 expression may be a prognostic indicator in malignant melanomas. Tumor staining with anti-chondroitin sulfate monoclonal antibodies suggests that CD44s may be expressed as a chondroitin sulfate proteoglycan in primary melanomas.

Adult↗
PubMed1996

CD44/chondroitin sulfate proteoglycan and alpha 2 beta 1 integrin mediate human melanoma cell migration on type IV collagen and invasion of basement membranes.

Tumor cell invasion of basement membranes (BM) represents one of the critical steps in the metastatic process. Tumor cell recognition of individual BM matrix components may involve individual cell adhesion receptors, such as integrins or cell surface proteoglycans, or may involve a coordinate action of both types of receptors. In this study, we have focused on the identification of a cell surface CD44/chondroitin sulfate proteoglycan (CSPG) and alpha 2 beta 1 integrin on human melanoma cells that are both directly involved in the in vitro invasion of reconstituted BM via a type IV collagen-dependent mechanism. Interfering with cell surface expression of human melanoma CSPG with either p-nitro-phenyl-beta-D-xylopyranoside treatment or anti-CD44 monoclonal antibody (mAb) preincubation (mAb) preincubation inhibits melanoma cell invasion through reconstituted BM. These treatments also strongly inhibit melanoma cell migration on type IV collagen, however, they are ineffective at inhibiting cell adhesion to type IV collagen. Purified melanoma cell surface CD44/CSPG, or purified chondroitin sulfate, bind to type IV collagen affinity columns, consistent with a role for CD44/CSPG-type IV collagen interactions in mediating tumor cell invasion. In contrast, melanoma cell migration on laminin (LM) does not involve CD44/CSPG, nor does CD44/CSPG bind to LM, suggesting that CD44/CSPG-type IV collagen interactions are specific in nature. Additionally, anti-alpha 2 and anti-beta 1 integrin mAbs are capable of blocking melanoma cell invasion of reconstituted BM. Both of these anti-integrin mAbs inhibit melanoma cell adhesion and migration on type IV collagen, whereas only anti-beta 1 mAb inhibits cell adhesion to LM. Collectively, these results indicate that melanoma cell adhesion to type IV collagen is an important consideration in invasion of reconstituted BM in vitro, and suggest that CD44/CSPG and alpha 2 beta 1 integrin may collaborate to promote human melanoma cell adhesion, migration, and invasion in vivo.

Basement Membrane↗
PubMed1996

Neuronal protection from cerebral ischemia by synthetic fibronectin peptides to leukocyte adhesion molecules.

Leukocytes play an important role in the development of ischemia/reperfusion injury. Recent work in our laboratory has demonstrated that a mixture of synthetic fibronectin peptides to leukocyte adhesion molecules reduces ischemic brain damage after transient focal cerebral ischemia. The purpose of this study was to evaluate the efficacy of the individual peptides on leukocyte accumulation, infarct size, and neurological outcome in rats subjected to 1 h of cerebral ischemia and 48 h of reperfusion. Thirty-five animals were divided into five groups: transient ischemia without treatment (Group I), treatment with arginyl-glycyl-aspartic acid (RGD) peptide (Group II), connecting segment (CS)-1 peptide (Group III), fibronectin (FN)-C/H-V peptide (Group IV), and scrambled FN-C/H-V peptide (Group V). Groups III and IV showed a significant decrease in the degree of leukocyte infiltration in the lesion and in the infarct size (p < 0.05) when compared to Groups I, II, and V. The neurological grade of Groups III and IV was significantly better than in Groups I, II, and V at 48 h after reperfusion (p < 0.01). Thus, in addition to demonstrating the potential efficacy of synthetic peptides as therapeutic agents for ischemia-reperfusion, these results also offer new insights into the mechanisms of leukocyte arrest and recruitment in ischemia/reperfusion injury.

Animals↗
PubMed1996

Synthetic fibronectin peptides and ischemic brain injury after transient middle cerebral artery occlusion in rats.

Leukocytes play an important role in the development of ischemia-reperfusion injury. This study was conducted to ascertain whether synthetic peptides corresponding to the cell- and heparin-binding sequences of fibronectin that disturb leukocyte adhesion molecules were effective in neuronal protection after transient focal cerebral ischemia in rats. The authors evaluated the efficacy of peptides on infarction size, leukocyte infiltration in the ischemic tissue, and neurological outcome in rats subjected to 1 hour of cerebral ischemia and 48 hours of reperfusion. Twenty-one animals were divided into three groups: transient ischemia without treatment (Group I), transient ischemia with administration of vehicle (Group II), and transient ischemia with administration of fibronectin peptides (Group III). The mean myeloperoxidase activity (U/g wet wt) in the ischemic area was as follows: Group I, 0.19% +/- 0.05; Group II, 0.21% +/- 0.03; and Group III, 0.08% +/- 0.02. The mean size of the infarction as a percentage of the total hemispheric volume was as follows: Group I, 38.35% +/- 1.34%; Group II, 39.21% +/- 2.42%; and Group III, 25.81% +/- 4.87%. Group III showed a significant decrease in myeloperoxidase activity in the lesion and the infarction size was smaller when compared to Groups I and II (p < 0.05). The neurological grade in Group III was significantly better than in Groups I and II at 48 hours after reperfusion (p < 0.01). This study is the first to explore the therapeutic potential of synthetic fibronectin peptides in brain protection after transient focal ischemia, and the results also serve as a basis for studies of important cellular and molecular events that contribute to tissue damage.

Animals↗
PubMed1996

Reduction of brain injury using heparin to inhibit leukocyte accumulation in a rat model of transient focal cerebral ischemia. I. Protective mechanism.

Heparin has long been established as an anticoagulant. Although heparin has been demonstrated to reduce brain injury after ischemia and reperfusion, its mechanism of action remains unknown. Recent investigations reveal that it can modulate biological processes such as binding to adhesion receptors on endothelial cells and leukocytes. The authors hypothesized that heparin's protective effect is closely related to its antileukocyte adherence property. They evaluated the efficacy of sulfated polysaccharides (unfractionated heparin, low-molecular-weight heparin, heparan sulfate, chondroitin sulfate C, and dextran sulfate) on leukocyte accumulation, infarction size, and neurological outcome after transient focal cerebral ischemia in rats subjected to 1 hour of ischemia and 48 hours of reperfusion. Forty-nine animals were included in the study. The animals receiving unfractionated heparin or dextran sulfate showed a significant reduction in leukocyte accumulation, infarct size, and neurological dysfunction 48 hours after reperfusion (p < 0.05) when compared to untreated animals. The animals receiving unfractionated heparin also showed significantly better results than the animals receiving an equivalent anticoagulant dose of low-molecular-weight heparin. These data indicate that heparin's antileukocyte property plays a more important role than its anticoagulant ability in neuronal protection. The relative potency of the sulfated polysaccharides tested in leukocyte depletion was closely related to their degree of sulfation. Thus, in addition to demonstrating the potential efficacy of heparin as a therapeutic agent for ischemia and reperfusion injury by the prevention of leukocyte accumulation, the results also serve as a basis for studying important cellular and molecular events that contribute to tissue damage.

Animals↗
PubMed1996

Reduction of brain injury using heparin to inhibit leukocyte accumulation in a rat model of transient focal cerebral ischemia. II. Dose-response effect and the therapeutic window.

The administration of massive doses of heparin has been demonstrated to reduce reperfusion injury. The authors have found that heparin's antileukocyte adhesion property may play a more important role than its anticoagulant property in preventing ischemia and reperfusion injury. Although the administration of massive doses of heparin has been demonstrated to reduce brain injury after ischemia and reperfusion, the optimum dosage and timing for heparin administration remain unknown. The purpose of this study was to evaluate the dose-response effect and determine the time during which heparin must be administered to inhibit leukocyte accumulation, reduce infarct size, and improve neurological outcome in rats subjected to 1 hour of cerebral ischemia and 48 hours of reperfusion. Forty-nine animals were included in the study. The animals receiving commercial unfractionated heparin at a total dose of 2.67 to 4 mg/kg showed a significant inhibition of leukocyte accumulation, reduced infarct size, and lessened neurological dysfunction 48 hours after reperfusion (p < 0.05) when compared to untreated animals. The animals receiving unfractionated heparin within 3 hours after reperfusion also showed significantly better results than untreated animals. These data indicate that standard doses of heparin prevent reperfusion injury, and relatively late postischemic administration of heparin also is effective in brain protection. These findings may have therapeutic potential as an adjunct to thrombolytic therapy and possibly for other perfusion deficiencies with leukocyte-endothelial interaction. In view of these encouraging experimental findings, the clinical application of heparin administration after ischemia and reperfusion warrants serious consideration.

Animals↗
PubMed1995

A peptide model of basement membrane collagen alpha 1 (IV) 531-543 binds the alpha 3 beta 1 integrin.

Tumor cell adhesion to the triple-helical domain of basement membrane (type IV) collagen occurs at several different regions. Cellular recognition of the sequence spanning alpha 1(IV)531-543 has been proposed to be independent of triple-helical conformation (Miles, A. J., Skubitz, A. P. N., Furcht, L. T., and Fields, G. B. (1994) J. Biol. Chem. 269, 30939-30945). In the present study, integrin interactions with a peptide analog of the alpha 1(IV)-531-543 sequence have been analyzed. Tumor cell adhesion (melanoma, ovarian carcinoma) to the alpha 1(IV)531-543 chemically synthesized peptide was inhibited by a monoclonal antibody against the alpha 3 integrin subunit, and to a lesser extent by monoclonal antibodies against the beta 1 and alpha 2 integrin subunits. An anti-alpha 5 monoclonal antibody and normal mouse IgG were ineffective as inhibitors of tumor cell adhesion to the peptide. Two cell surface proteins of 120 and 150 kDa bound to an alpha 1(IV)531-543 peptide affinity column and were eluted with 20 mM EDTA. When the eluted proteins were incubated with monoclonal antibodies against either the alpha 3 or beta 1 integrin subunit, proteins corresponding in molecular weight to alpha 3 and beta 1 integrin subunits were precipitated. No proteins were immunoprecipated with monoclonal antibodies against the alpha 2 or alpha 5 integrin subunits. Thus, the alpha 3 beta 1 integrin from two tumor cell types has been shown to bind directly to the alpha 1 (IV)531-543 peptide. The alpha 1(IV)531-543 peptide is the first collagen-like sequence that has been shown to bind the alpha 3 beta 1 integrin.

Amino Acid Sequence↗
PubMed1995

Identification of a homologous heparin binding peptide sequence present in fibronectin and the 70 kDa family of heat-shock proteins.

This study was undertaken to characterize the potential heparin affinity of an amino-acid sequence within the 70 kDa heat-shock family of proteins (HSPs) that shares homology with a heparin-binding sequence present in the carboxy-terminus of fibronectin (FN), defined by the synthetic peptide, FN-C/H-II (KNNQKSEPLIGRKKT). To first define the heparin binding sequence within FN-C/H-II, solid phase binding assays were performed using overlapping, short (7 amino acids) synthetic peptides corresponding to the amino-acid sequence within FN-C/H-II. Only the sequence LIGRKKT bound [3H] heparin, and the LIGRKKT peptide blocked heparin binding to intact fibronectin by 47% (+/- 0.4, p < 0.001). A computer-generated homology search revealed that two members of the 70 kDa HSP family, HSP70 and HSC70, contain the sequences LIGRK and LIGRR, respectively. Examination of heparin binding using affinity chromatography indicated that while native HSC70 binds heparin, native HSP70 does not. Treatment of the heparin-unbound fraction with heat or urea led to enhanced HSP70 binding to heparin affinity columns. Soluble LIGRKKT peptide or anti-FN-C/H-II IgG also significantly inhibited heparin binding to HSC70 that had been purified by heparin affinity chromatography. Finally, Western blot analysis of HSC70 purified by heparin affinity chromatography demonstrated that polyclonal anti-FN-C/H-II IgG could recognize HSC70. These data demonstrate that LIGRK or LIGRR represent a a common heparin binding motif in fibronectin, HSP70, and HSC70, and are consistent with a proposed role for heparin or similar polyanionic structures in the function of the 70 kDa heat-shock proteins.

Amino Acid Sequence↗
PubMed1995

Spreading and focal contact formation of human melanoma cells in response to the stimulation of both melanoma-associated proteoglycan (NG2) and alpha 4 beta 1 integrin.

In this study, we evaluated the potential role for a specific melanoma-associated chondroitin sulfate proteoglycan core protein, termed NG2, to collaborate with alpha 4 beta 1 integrin in focal contact formation in human melanoma cells. Although melanoma cells adhered to substrata coated with either the alpha 4 beta 1 integrin binding fibronectin synthetic peptide CS1-OVA or anti-NG2 mAbs, no spreading or focal contact formation was observed on either substratum. However, melanoma cells spread and formed focal contacts on "chimeric substrata" coated with CS1-OVA and the anti-NG2 mAb, 9.2.27, indicating that engaging both adhesion receptors changes the adhesion phenotype of melanoma cells by reorganizing the cytoskeleton. The collaboration between the two receptors is specific to fibronectin, since cells adherent on substrata coated with low concentrations of either laminin and 9.2.27 or type IV collagen and 9.2.27 failed to spread, while cells adherent on low concentrations of fibronectin and 9.2.27 exhibited a fully spread morphology. Two selective tyrosine kinase inhibitors, genistein and herbimycin A, totally inhibited cell spreading on the substrata coated with CS1-OVA and 9.2.27, indicating that tyrosine kinase(s) is important for cell spreading and focal contact formation. When cells were cultured on substrata coated with CS1-OVA and 9.2.27, two proteins (M(r) 130,000 and 120,000) were tyrosine phosphorylated in a genistein- and herbimycin A-sensitive fashion. These proteins were not immunologically related to pp125FAK or alpha 4 beta 1 integrin. Importantly, when melanoma cells were cultured on substrata coated with CS1 and then stimulated with 9.2.27-conjugated microsphere beads, formation of focal contacts and stress fibers was also observed, indicating that NG2 can collaborate with alpha 4 beta 1 integrin when each receptor is engaged on distinct and separate substrata. These results demonstrate that NG2 acts as a coreceptor for spreading and focal contact formation in association with alpha 4 beta 1 integrin in melanoma cells and suggest a model in which the NG2 core protein communicates to alpha 4 beta 1 integrin by an inside-out signaling mechanism.

Antibodies, Monoclonal↗
PubMed1995

Functional involvement of sciatic nerve-derived versican- and decorin-like molecules and other chondroitin sulphate proteoglycans in ECM-mediated cell adhesion and neurite outgrowth.

We have previously described two proteoglycans from human sciatic nerve which are immunochemically related to the chondroitin sulphate proteoglycans versican and decorin. The chondroitin sulphate of the versican-like molecule and the core protein of the decorin-like molecule have been found previously to be up-regulated after lesioning the adult mouse sciatic nerve. To investigate if the versican- and decorin-like molecules are involved in cell-extracellular matrix interactions, we studied the effect of both molecules on cell adhesion. The versican- and decorin-like molecules, substrate-coated in a mixture with fibronectin, but not with laminin or collagen types I or IV, inhibited the adhesion of several cell lines, neonatal dorsal root ganglion neurons and Schwann cells. The inhibitory activity was concentration-dependent and mediated by the chondroitin sulphate. Furthermore, when different proteoglycans were incubated with fibronectin, only the versican- and decorin-like molecules and the chondroitin sulphate proteoglycan aggrecan, but not the heparan sulphate proteoglycan perlecan, were able to inhibit fibronectin-mediated cell adhesion. The versican- and decorin-like molecules, substrate-coated alone or in a mixture with fibronectin or laminin, were at most slightly inhibitory to neurite outgrowth from PC12 phaeochromocytoma cells and neonatal dorsal root ganglion neurons. In a solid-phase ligand-binding assay the versican- and decorin-like molecules interacted with fibronectin, but not with laminin or collagen types I and IV. Binding of the versican-like molecule to fibronectin and inhibition of cell adhesion by this molecule was mediated via the heparin and cell-binding domains of fibronectin. These observations suggest that binding of the two proteoglycans to fibronectin is involved in the modulation of adhesion of cells to fibronectin.

Animals↗
PubMed1995

Adolescent character formation and psychoanalytic theory.

The foremost implication of the Freudian theories of adolescence has been that the analyst enters into an alliance with the patient's developmental process. During sessions, stock is routinely taken of the adolescent patient's defenses and drive fixations. The interpersonal and object relations theories of adolescents' character formation varied with each other and also with the assumptions of classical Freudian metapsychology. Yet, Sullivan, Fairbairn, and Winnicott all stressed relatedness in character formation. They each urged that the attitudes of the actual person in significant relationships, as well as the internal representations of the self and the object, shaped the character tendencies of the child or adolescent. Sullivan was very outspoken about his belief that there could be no uniformly valid theory of character, because people are unique. However, for Sullivan, the needs for the validation of self-worth, and for freedom from anxiety, were universal stimuli for the increasing organization of character trends. In both interpersonal theory and object relations theories, dissociative processes were of paramount importance as defensive operations. Dissociation by the adolescent resulted in further instances of ego splitting (for Fairbairn), of the bad me (for Sullivan), and of the false self (for Winnicott). Fairbairn, and to some extent Winnicott, used the language of classical Freudian theory in order to shape an object relations theory of adolescent development. In spite of their theoretical differences, Sullivan, Fairbairn, and Winnicott spoke with a singular voice in dismissing the exclusive significance of libidinal fixations in character consolidation. I now wish to review Freud's case of Dora as an addendum to this short critical appraisal. The analysis of Dora readily lends itself to a discussion of the confluence of the psychoanalytical models' clinical theories. Dora's unfortunate experience in treatment offered a compelling example of the precariousness of adolescents' adjustment in the midst of developmental and family turmoil.

Adolescent↗
PubMed1995

Cooperative signaling by alpha 5 beta 1 and alpha 4 beta 1 integrins regulates metalloproteinase gene expression in fibroblasts adhering to fibronectin.

Rabbit synovial fibroblasts (RSF) express basal levels of the metalloproteinases (MMP) collagenase, stromelysin-1 and 92-kD gelatinase when plated on intact fibronectin (FN), but elevated levels when plated on either the central RGD-containing cell-binding region of FN (120FN) or antibody against the alpha 5 beta 1 integrin, suggesting that domains outside 120FN may suppress the induction of MMP (Werb, Z., P. M. Tremble, O. Behrendtsen, E. Crowley, and C.H. Damsky. 1989. J. Cell Biol. 109:877-889). We therefore attempted to reconstitute the basal signaling of intact FN by plating RSF on 120FN together with domains of FN outside this region. Large COOH-terminal fragments containing both the heparin-binding and HICS domains suppressed MMP when combined with 120FN. To map the active sequences, peptides from this region and larger fragments that did, or did not, include the CS-1 portion of IIICS were tested. Only CS-1 peptide, or larger fragments containing CS-1, suppressed MMP expression induced by 120FN. In contrast, peptide V from the heparin-binding region, shown previously to stimulate focal contact formation, further enhanced MMP expression by RSF when present on the substrate with 120FN. RSF expressed alpha 4 beta 1 integrin, the receptor for CS-1, and the anti-alpha 4 mAb blocked the ability of CS-1 to suppress MMP induction by 120FN. These results show that signals modulating MMP expression and focal contact assembly are regulated independently, and that cooperative signaling by alpha 5 beta 1 and alpha 4 beta 1 integrins plays a dominant role in regulating expression of these extracellular matrix-remodeling genes in response to FN. This work demonstrates directly the modular way in which information in the extracellular matrix is detected and processed by cell surface receptors.

Amino Acid Sequence↗
PubMed1995

Adherence of Streptococcus pneumoniae to immobilized fibronectin.

Adherence to extracellular matrix proteins, such as fibronectin, affords pathogens with a mechanism to invade injured epithelia. Streptococcus pneumoniae was found to adhere to immobilized fibronectin more avidly than other streptococci and staphylococci do. Binding was dose, time, and temperature dependent. Trypsin treatment of the bacteria resulted in decreased binding, suggesting that the bacterial adhesive component was a protein. Fragments of fibronectin generated by proteolysis or by expression of recombinant gene segments were compared for the ability to bind pneumococci and to compete against bacterial binding to immobilized fibronectin. Fragments from the carboxy-terminal heparin binding domain were consistently active, suggesting that this region contains the pneumococcal binding site, a region distinct from that supporting the attachment of most other bacteria.

Bacterial Adhesion↗
PubMed1995

Endothelial cell interactions with synthetic peptides from the carboxyl-terminal heparin-binding domains of fibronectin.

Fibronectin (FN) plays an important role in endothelial cell adhesion, spreading, and motility. Within FN, a number of functional domains have been identified, including the 33/66-kD carboxyl-terminal heparin-binding fragments, which support the adhesion of vascular endothelial cells. A number of synthetic peptides representing amino acid sequences within the 33/66-kD fragments have been shown to promote the adhesion, spreading, and migration of a variety of cell types. Our working hypothesis is that one or more of these sequences may also mediate vascular endothelial cell adhesion, spreading, and migration to the 33/66-kD fragments. In support of this hypothesis, we have demonstrated that endothelial cells from various sources adhered in a concentration-dependent manner to surfaces coated with FN, the 33/66-kD fragments, and synthetic peptides derived from the 33/66-kD fragments of FN. FN and the 33/66-kD fragments also promoted endothelial cell spreading and migration. Although each of the six synthetic peptides tested supported endothelial cell adhesion, only one of these peptides within the carboxyl-terminal heparin-binding domain (FN-C/H-V) promoted endothelial cell spreading and migration. Cell spreading on FN-C/H-V, as well as on FN and the 33/66-kD fragments, was associated with the formation of a well-developed actin cytoskeleton and the formation of focal contacts. FN-C/H-V (but not scrambled FN-C/H-V) inhibited cell spreading on FN and the 33/66-kD fragments in a concentration-dependent manner. FN-C/H-V had a modest effect on the adhesion of a clonal population of rat heart endothelial cells (RHE-1A) to the 33/66-kD fragments of FN and no effect on RHE-1A cell adhesion to FN. These findings suggest that peptide FN-C/H-V is unique among this group of peptides derived from the 33/66-kD heparin-binding fragments of FN in its ability to promote the adhesion, spreading, and migration of vascular endothelial cells and further suggest that the sequence defined by this peptide plays an important role in vascular endothelial cell interactions with the 33/66-kD fragments of FN.

Amino Acid Sequence↗
PubMed1995

Direct adhesion to bone marrow stroma via fibronectin receptors inhibits hematopoietic progenitor proliferation.

In long-term bone marrow cultures, stroma-adherent progenitors proliferate significantly less than nonadherent progenitors. Thus, close progenitor-stroma interactions may serve to regulate or restrict rather than promote hematopoietic progenitor proliferation. We hypothesized that signaling through adhesion receptors on hematopoietic cells may contribute to the inhibition of proliferation observed when progenitors are in contact with stroma. We demonstrate that progenitors cultured physically separated from stroma in a transwell proliferate significantly more than progenitors adherent to stroma. Furthermore, proliferation of colony forming cells (CFC) is reduced after specific adhesion to stroma, metabolically inactivated glutaraldehyde-fixed stroma, stromal-extracellular matrix, or the COOH-terminal heparin-binding domain of fibronectin. Nonspecific adhesion to poly-L-lysine fails to inhibit CFC proliferation. That the VLA-4 integrin is one of the receptors that transfers proliferation inhibitory signals was shown using blocking anti-alpha 4 monomeric F(ab) fragments. Furthermore, when synthetic peptides representing specific cell attachment sites within the heparin-binding domain of fibronectin were added to Dexter-type marrow cultures, significantly increased recovery and proliferation of CFC was observed, suggesting that these peptides disrupt adhesion-mediated proliferation inhibitory events. Thus, negative regulation of hematopoiesis may not only depend on the action of growth inhibitory cytokines but also on growth inhibitory signals resulting from direct adhesive interactions between progenitors and marrow stroma.

Amino Acid Sequence↗
PubMed1994

Characterization of soluble CD44 in the circulation of mice. Levels are affected by immune activity and tumor growth.

ELISA determinations revealed substantial concentrations (0.49 to 2.10 micrograms/ml) of soluble CD44 in murine serum, with some variation among normal mouse strains. At least three species of CD44 were identified by immunoprecipitation and SDS-PAGE analysis of serum. The most prominent was indistinguishable in mobility from that extracted from normal and transformed lymphocytes and was estimated in this way to be approximately 90 kDa. A similar estimate resulted from gel filtration under nondenaturing conditions, followed by ELISA. However, lymphocyte membrane-extracted and soluble CD44 had different mobilities after treatment with neuraminidase plus O-glycosidase, and the core protein of soluble CD44 might be 17 to 20 kDa smaller than that of CD44 on lymphocyte membranes. Furthermore, an Ab to cytoplasmic residues of CD44 failed to recognize soluble CD44 recovered from the circulation or in lymphoma culture supernatants. These observations would be consistent with cleavage of CD44 from cell surfaces; and protease inhibitors slowed the loss of CD44 from cultured lymphomas. Serum CD44 levels were significantly reduced in immunodeficient CD17.SCID and BALB/c.Xid mice, and elevated in tumor-bearing mice. Mild graft-vs-host (GVH) reactions also resulted in increased concentrations of CD44, as did autoimmune disease in BXSB and MRL/lpr strains of mice. Serum with high concentrations of CD44 partially blocked the binding of one ligand, hyaluronate, to CD44-bearing hybridoma cells. The degree of inhibition was positively correlated with CD44 concentration. These findings indicate that substantial quantities of CD44 can be released into the circulation by cleavage from cell surfaces and that this process is markedly influenced by immune system activity and tumor growth. The material seemed to be intact and potentially functional.

Amino Acid Sequence↗