Biomedical subjects

J H Phillips

Publications and source records attributed to J H Phillips.

At least 37 records · Page 2Linked to original sources

PubMed1999

An activating immunoreceptor complex formed by NKG2D and DAP10.

Many immune receptors are composed of separate ligand-binding and signal-transducing subunits. In natural killer (NK) and T cells, DAP10 was identified as a cell surface adaptor protein in an activating receptor complex with NKG2D, a receptor for the stress-inducible and tumor-associated major histocompatibility complex molecule MICA. Within the DAP10 cytoplasmic domain, an Src homology 2 (SH2) domain-binding site was capable of recruiting the p85 subunit of the phosphatidylinositol 3-kinase (PI 3-kinase), providing for NKG2D-dependent signal transduction. Thus, NKG2D-DAP10 receptor complexes may activate NK and T cell responses against MICA-bearing tumors.

Amino Acid Sequence↗

PubMed1999

Cutting edge: human 2B4, an activating NK cell receptor, recruits the protein tyrosine phosphatase SHP-2 and the adaptor signaling protein SAP.

The genetic defect in X-linked lymphoproliferative syndrome (XLP) is the Src homology 2 domain-containing protein SAP. SAP constitutively associates with the cell surface molecule, signaling lymphocytic activation molecule (SLAM), and competes with SH2-domain containing protein tyrosine phosphatase-2 (SHP-2) for recruitment to SLAM. SLAM exhibits homology with the mouse cell surface receptor 2B4. The human homologue of 2B4 has now been identified. It is recognized by the c1.7 mAb, a mAb capable of activating human NK cells. Human 2B4 became tyrosine phosphorylated following pervanadate-treatment of transfected cells and recruited SHP-2. SAP was also recruited to 2B4 in activated cells. Importantly, the 2B4-SAP interaction prevented the association between 2B4 and SHP-2. These results suggest that the phenotype of XLP may result from perturbed signaling not only through SLAM, but also other cell surface molecules that utilize SAP as a signaling adaptor protein.

Amino Acid Sequence↗

PubMed1999

Leukocyte-associated Ig-like receptor-1 functions as an inhibitory receptor on cytotoxic T cells.

Leukocyte associated Ig-like receptor-1 (LAIR-1) is a surface molecule expressed on human mononuclear leukocytes that functions as an inhibitory receptor on human NK cells. In addition to NK cells, LAIR-1 is expressed on T cells, B cells, macrophages, and dendritic cells. Most cells express two biochemically distinct forms of LAIR-1, which we now show are likely alternative splice variants of the same gene. Cross-linking of LAIR-1 on human T cell clones results in inhibition of cytotoxicity only in T cell clones that lack CD28 and are able to spontaneously lyse certain targets in vitro. Moreover, the cytolytic activity of freshly isolated T cells, which is thought to be mainly due to "effector" T cells, can be inhibited by anti-LAIR-1 mAb. Thus, LAIR-1 functions as an inhibitory receptor not only on NK cells, but also on human T cells. This indicates that LAIR-1 provides a mechanism of regulation of effector T cells and may play a role in the inhibition of unwanted bystander responses mediated by Ag-specific T cells.

Alternative Splicing↗

PubMed1999

Bioengineering of elastic cartilage with aggregated porcine and human auricular chondrocytes and hydrogels containing alginate, collagen, and kappa-elastin.

Transplantation of isolated chondrocytes has long been acknowledged as a potential method for rebuilding small defects in damaged or deformed cartilages. Recent advances in tissue engineering permit us to focus on production of larger amounts of cartilaginous tissue, such as might be needed for reconstructive surgery of the entire auricle. In this report we describe modification of the basic techniques that lead to production of a large amount of elastic cartilage originated from porcine and human isolated chondrocytes. Small fragments of auricular cartilage were harvested from children undergoing ear reconstruction for microtia or extirpation of preauricular tags and from ears of juvenile pigs. Enzymatically isolated elastic chondrocytes were then agitated in suspension to form the chondronlike aggregates, which were further embedded in molded hydrogel constructs made of alginate and type I collagen augmented with kappa-elastin. The constructs were then implanted in nude mice and harvested 4 and 12 weeks after heterotransplantation. The resulting neocartilage closely resembled native auricular cartilage at the gross, microscopic, and ultrastructural levels. Immunohistochemistry and electron microscopy additionally confirmed that the newly produced cartilage contained the major components of the elastic cartilage-specific matrix, including collagen type II, proteoglycans, and well-assembled elastic fibers.

Alginates↗

PubMed1999

Physical and functional association of LFA-1 with DNAM-1 adhesion molecule.

Whereas ligation of the DNAM-1 adhesion molecule triggers cytotoxicity mediated by normal NK and T cells, this function was defective in NK cell clones from leukocyte adhesion deficiency syndrome. However, genetic reconstitution of cell surface expression of LFA-1 restored the ability of DNAM-1 to initiate anti-DNAM-1 mAb-induced cytotoxicity, indicating a functional relationship between DNAM-1 and LFA-1. Further studies demonstrated that LFA-1 physically associates with DNAM-1 in NK cells and anti-CD3 mAb stimulated T cells, for which serine phosphorylation of DNAM-1 plays a critical role. In addition, cross-linking of LFA-1 induces tyrosine phosphorylation of DNAM-1, for which the Fyn protein tyrosine kinase is responsible. These results indicate that DNAM-1 is involved in the LFA-1-mediated intracellular signals.

Animals↗

PubMed1999

Mandibular reconstruction in children using the vascularized fibula.

In this study, the authors review their experience with vascularized fibula transfers for mandibular reconstruction in children. They outline the indications for such reconstruction, their method of contouring the fibula to accurately resemble the resected mandible, reconstruction of the temporomandibular joint, and the use of vascularized muscle for the management of associated soft-tissue deficiencies. Ten consecutive patients, 5 to 17 years of age and undergoing this procedure, were assessed from a medical, dental, radiographic, and photographic standpoint. The fibulae were elevated via a lateral approach, osteotomized, as required, and fixation was achieved with titanium miniplates and screws. All transfers survived, with viability confirmed by early postoperative bone scanning. Five patients had temporomandibular joint reconstruction, and five patients required simultaneous reconstruction of soft-tissue defects with associated vascularized muscle. The postoperative follow-up ranged from 3 to 30 months The occlusion of the remaining dentition, mandibular symmetry and projection, adequacy of lining and skin cover, and maximal mouth opening were reassessed. Occlusion was class 1 in all patients, and free-flap stability and function were in the normal range. However, soft-tissue contour was a problem. All wounds healed primarily without donor-site complications and with minimal recipient-site complications.

Adolescent↗

PubMed1999

Comparison of anthropometric and cephalometric measurements of the adult face.

To tabulate and assess quantitative differences between anthropometric and corresponding radiographic cephalometric measurements obtained from the same persons, 19 projective linear measurements were taken from the surface of the heads and faces of 41 patients with cleft lip, cleft palate, or both, all of whom were white North Americans aged 14 to 29 years. They underwent radiographic examination shortly afterward, and corresponding cephalometric measurements were obtained. Differences between the methods were assessed by the numeric differences between the mean values of concurrent measurements. Statistical difference was assessed by paired t test, Pearson product-moment correlations, and intraclass index for degree of agreement between findings. By numeric difference, 6 of 19 anthropometric measurements were similar to (within 2% of) those taken from radiographs. Paired t tests disclosed significant differences between 16 of 19 measurements. Half of the six surface measurements similar to their skeletal counterparts showed no statistical difference; the other half showed only moderately significant differences. Differences between the 13 "dissimilar" measurement pairs (differences > 2%) were highly significant. Good correlations were found in five of the six similar measurements, which may have an important prognostic value in understanding changes in the craniofacial measurements of the face. Knowledge of the correlations between all major measurements of the head and face on the surface and skeleton is essential for anticipating changes in the morphologic characteristics of the growing face.

Adolescent↗

PubMed1999

The effect on facial growth of pediatric mandibular fractures.

The incidence of facial fractures in the pediatric population is between 1.4% and 15% of all maxillofacial traumas. Forty-one percent of pediatric facial fractures involve the mandible. No study has commented on the incidence of mandibular fractures that go on to develop growth disturbances leading to asymmetry and malocclusion. A retrospective chart review was carried out that identified and followed 88 children who sustained mandibular fractures and presented to The Hospital for Sick Children in Toronto during the 13-year period from 1980 to 1993. Patient follow-up ranged from 2 to 15 years, and was performed via phone survey and medical/orthodontic chart review. Patients who required orthodontics and orthognathic surgery were identified. Results indicated that a pediatric mandibular fracture does not lead to a higher incidence of orthodontic intervention. Furthermore, children younger than 4 years or older than 12 years rarely require orthognathic surgery to correct facial growth disturbances following mandibular fractures. In contrast, 22% of children age 4 to 7 years, and 17% of children age 8 to 11 years required orthognathic surgery to correct facial growth disturbances following mandibular fractures. Condylar fractures were the most common site of mandibular fracture, and led to facial asymmetry most frequently.

Adolescent↗

PubMed1998

Identification of functional human splenic memory B cells by expression of CD148 and CD27.

Memory B cells isolated from human tonsils are characterized by an activated cell surface phenotype, localization to mucosal epithelium, expression of somatically mutated immunoglobulin (Ig) variable (V) region genes, and a preferential differentiation into plasma cells in vitro. In spleens of both humans and rodents, a subset of memory B cells is believed to reside in the marginal zone of the white pulp. Similar to tonsil-derived memory B cells, splenic marginal zone B cells can be distinguished from naive follicular B cells by a distinct cell surface phenotype and by the presence of somatic mutations in their Ig V region genes. Although differences exist between human naive and memory B cells, no cell surface molecules have been identified that positively identify all memory B cells. In this study, we have examined the expression of the receptor-type protein tyrosine phosphatase CD148 on human B cells. CD148(+) B cells present in human spleen exhibited characteristics typical of memory B cells. These included an activated phenotype, localization to the marginal zone, the expression of somatically mutated Ig V region genes, and the preferential differentiation into plasma cells. In contrast, CD148(-) B cells appeared to be naive B cells due to localization to the mantle zone, the expression of surface antigens typical of unstimulated B cells, and the expression of unmutated Ig V region genes. Interestingly, CD148(+) B cells also coexpressed CD27, whereas CD148(-) B cells were CD27(-). These results identify CD148 and CD27 as markers which positively identify memory B cells present in human spleen. Thus, assessing expression of these molecules may be a convenient way to monitor the development of memory B cell responses in immunocompromised individuals or in vaccine trials.

B-Lymphocyte Subsets↗

PubMed1998

Negative regulation of human T cell activation by the receptor-type protein tyrosine phosphatase CD148.

T cell activation represents a balance between positive and negative signals delivered via distinct cell surface molecules. Many cytoplasmic protein tyrosine phosphatases are involved in regulating cellular responses by antagonizing the action of protein tyrosine kinases. CD148 is a receptor-type protein tyrosine phosphatase expressed by all human mononuclear cells. We have investigated the effect of CD148 on TCR-mediated activation of human T cells. Overexpression of wild-type, but not a phosphatase-deficient, CD148 in Jurkat T cells inhibited TCR-mediated activation, evidenced by reduced expression of the early activation Ag CD69, inhibition of tyrosine phosphorylation of many intracellular proteins including the critical protein tyrosine kinase ZAP-70, and impairment of mitogen-activated protein kinase activation. Taken together, these results suggest that CD148 is an important phosphatase involved in negatively regulating the proximal signaling events during activation of Ag-specific T cells.

Gene Expression Regulation↗

PubMed1998

CD148: a receptor-type protein tyrosine phosphatase involved in the regulation of human T cell activation.

Following ligation of the TCR and costimulatory molecules such as CD28, T cells proliferate and secrete cytokines. Several other cell surface molecules have been identified that are capable of augmenting activation mediated via the TCR. These include CD2, CD27, CD40 ligand, and signaling lymphocytic activation molecule. Here, we have characterized the expression and function of CD148, a recently identified receptor-type protein tyrosine phosphatase. CD148 is expressed at low levels on resting T cells, but is up-regulated following in vitro activation. Cross-linking CD148 with immobilized anti-CD148 mAb induced vigorous proliferation of anti-CD3 mAb-activated, highly purified peripheral blood T cells in an IL-2-dependent, cyclosporin A-sensitive manner. This effect was greatest after 8 days of in vitro culture, suggesting that this molecule is involved in the latter stages of a T cell response. CD148-induced proliferation was significantly greater for CD8+ T cells than for CD4+ T cells. Thus, CD148 is a receptor-type protein tyrosine phosphatase involved in the activation of T lymphocytes.

Animals↗

PubMed1998

Protein kinase C is involved in the regulation of both signaling and adhesion mediated by DNAX accessory molecule-1 receptor.

DNAX accessory molecule-1 (DNAM-1) is a signal-transducing adhesion molecule involved in the cytolytic function mediated by CTL and NK cells. In the present study, we have investigated various perimeters of DNAM-1-mediated signaling and adhesion. Although adhesion of DNAM-1 to its ligand does not require divalent cations, protein synthesis, or RNA transcription, activation of protein kinase C (PKC) is required for DNAM-1 functioning. Furthermore, mutation of the putative PKC-binding site in the cytoplasmic domain of DNAM-1 (Ser329 to Phe329) prevents both ligand binding and PMA-induced phosphorylation of the DNAM-1 receptor. These results indicate that PKC phosphorylates Ser329 of DNAM-1 and plays a critical role for both DNAM-1 adhesion and signaling.

Amino Acid Sequence↗

PubMed1998

Ly-49D and Ly-49H associate with mouse DAP12 and form activating receptors.

Several members of the Ly-49 receptor family inhibit NK cell-mediated lysis of targets expressing appropriate MHC class I molecules. Ly-49D and Ly-49H, two Ly-49 molecules that lack immunoreceptor tyrosine-based inhibitory motifs (ITIM) in their cytoplasmic domains, associate with mouse DAP12, a molecule that possesses an immunoreceptor tyrosine-based activation motif (ITAM). Cotransfection of either Ly-49D or Ly-49H with DAP12 induces surface expression of both Ly-49 and DAP12. The Ly-49/DAP12 complex was coimmunoprecipitated from the transfected cells, demonstrating a physical association of DAP12 with Ly-49D or Ly-49H in the plasma membrane. Stimulation of transfectants with Abs recognizing either Ly-49D or Ly-49H results in cellular activation, as assessed by induction of tyrosine phosphorylation of multiple cellular substrates.

Adaptor Proteins, Signal Transducing↗

PubMed1998

Killer cell inhibitory receptors for MHC class I molecules regulate lysis of melanoma cells mediated by NK cells, gamma delta T cells, and antigen-specific CTL.

NK cells and T cells express killer cell inhibitory receptors (KIR) recognizing polymorphic MHC class I molecules. Although prior studies have established that MHC class I can protect normal and transformed hematopoietic cells from NK cell lysis, the role of MHC class I on the recognition of solid tumors has been controversial. In this study, we investigated whether interactions of KIR with their ligands on melanoma tumor cells could inhibit tumor cell lysis by NK and gamma delta T cell clones. Ligation of the NK cell receptor KIR3DL1 by HLA-Bw4 allotypes resulted in inhibition of cytotoxicity against HLA-B*4403-transfected melanomas as well as against melanomas endogenously expressing HLA-Bw4 allotypes. Similarly, interactions of KIR2DL2 or KIR2DL3 (KIR2DL2/3) with HLA-Cw3-related allotypes on melanomas resulted in decreased tumor cell lysis. We also investigated whether signaling via KIR affected melanoma recognition by CTL. Introduction of KIR3DL1 molecules into HLA-A*0201-restricted gp100-specific CTL resulted in inhibition of lysis of gp100+ melanomas co-expressing HLA-A*0201 and HLA-Bw4 allotypes. These results suggest that disrupting interactions of KIR with their ligands on tumor cells in vivo may enhance antitumor responses mediated by both innate and adaptive immune effector cells.

Animals↗

PubMed1998

Modulation of natural killer cell cytotoxicity in human cytomegalovirus infection: the role of endogenous class I major histocompatibility complex and a viral class I homolog.

Natural killer (NK) cells have been implicated in early immune responses against certain viruses, including cytomegalovirus (CMV). CMV causes downregulation of class I major histocompatibility complex (MHC) expression in infected cells; however, it has been proposed that a class I MHC homolog encoded by CMV, UL18, may act as a surrogate ligand to prevent NK cell lysis of CMV-infected cells. In this study, we examined the role of UL18 in NK cell recognition and lysis using fibroblasts infected with either wild-type or UL18 knockout CMV virus, and by using cell lines transfected with the UL18 gene. In both systems, the expression of UL18 resulted in the enhanced killing of target cells. We also show that the enhanced killing is due to both UL18-dependent and -independent mechanisms, and that the killer cell inhibitory receptors (KIRs) and CD94/NKG2A inhibitory receptors for MHC class I do not play a role in affecting susceptibility of CMV-infected fibroblasts to NK cell-mediated cytotoxicity.

Antigens, Viral↗

PubMed1998

HLA-E binds to natural killer cell receptors CD94/NKG2A, B and C.

The protein HLA-E is a non-classical major histocompatibility complex (MHC) molecule of limited sequence variability. Its expression on the cell surface is regulated by the binding of peptides derived from the signal sequence of some other MHC class I molecules. Here we report the identification of ligands for HLA-E. We constructed tetramers in which recombinant HLA-E and beta2-microglobulin were refolded with an MHC leader-sequence peptide, biotinylated, and conjugated to phycoerythrin-labelled Extravidin. This HLA-E tetramer bound to natural killer (NK) cells and a small subset of T cells from peripheral blood. On transfectants, the tetramer bound to the CD94/NKG2A, CD94/NKGK2B and CD94/NKG2C NK cell receptors, but did not bind to the immunoglobulin family of NK cell receptors (KIR). Surface expression of HLA-E was enough to protect target cells from lysis by CD94/NKG2A+ NK-cell clones. A subset of HLA class I alleles has been shown to inhibit killing by CD94/NKG2A+ NK-cell clones. Only the HLA alleles that possess a leader peptide capable of upregulating HLA-E surface expression confer resistance to NK-cell-mediated lysis, implying that their action is mediated by HLA-E, the predominant ligand for the NK cell inhibitory receptor CD94/NKG2A.

Animals↗

PubMed1998

Immunoreceptor DAP12 bearing a tyrosine-based activation motif is involved in activating NK cells.

Natural killer (NK) cells express cell-surface receptors of the immunoglobulin and C-type lectin superfamilies that recognize major histocompatibility complex (MHC) class I peptides and inhibit NK-cell-mediated cytotoxicity. These inhibitory receptors possess ITIM sequences (for immunoreceptor tyrosine-based inhibitory motifs) in their cytoplasmic domains that recruit SH2-domain-containing protein tyrosine phosphatases, resulting in inactivation of NK cells. Certain isoforms of these NK-cell receptors lack ITIM sequences and it has been proposed that these 'non-inhibitory' receptors may activate, rather than inhibit, NK cells. Here we show that DAP12, a disulphide-bonded homodimer containing an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain, non-covalently associates with membrane glycoproteins of the killer-cell inhibitory receptor (KIR) family without an ITIM in their cytoplasmic domain. Crosslinking of KIR-DAP12 complexes results in cellular activation, as demonstrated by tyrosine phosphorylation of cellular proteins and upregulation of early-activation antigens. Phosphorylated DAP12 peptides bind ZAP-70 and Syk protein tyrosine kinases, suggesting that the activation pathway is similar to that of the T- and B-cell antigen receptors.

Amino Acid Sequence↗

PubMed1998

Association of DAP12 with activating CD94/NKG2C NK cell receptors.

While the inhibitory NK cell receptors for MHC class I express immunoreceptor tyrosine-based inhibitory motifs that recruit intracellular tyrosine phosphatases and prevent NK cell effector function, the activating NK cell receptors lack intrinsic sequences required for cellular stimulation. CD94/NKG2C, an activating NK cell receptor of the C-type lectin superfamily that binds to HLA-E, noncovalently associates with DAP12, a membrane receptor containing an immunoreceptor tyrosine-based activating motif. Efficient expression of CD94/NKG2C on the cell surface requires the presence of DAP12, and charged residues in the transmembrane domains of DAP12 and NKG2C are necessary for this interaction. These results provide a molecular basis for the assembly of NK cell receptors for MHC class I involved in cellular activation and inhibition.

Adaptor Proteins, Signal Transducing↗