CD16, the low affinity Fc gamma receptor III for IgG (Fc gamma RIII), exists as a polypeptide-anchored form (Fc gamma RIIIA or CD16A) in human natural killer cells and macrophages and as a glycosylphosphatidylinositol-anchored form (Fc gamma RIIIB or CD16B) in neutrophils. CD16A requires association of the gamma subunit of Fc epsilon RI or the zeta subunit of the TCR-CD3 complex for cell surface expression. The CD16B is polymorphic and the two alleles are termed NA1 and NA2. In this study, CD16A and the two alleles of CD16B have been expressed in Chinese hamster ovary (CHO) cells and their ligand binding and phagocytic properties analyzed. The two allelic forms of CD16B showed a similar affinity toward human IgG1. However, the NA1 allele showed approximately 2-fold higher affinity for the IgG3 than the NA2 allele. Although all three forms of CD16 efficiently bound rabbit IgG-coated erythrocytes (EA), only CD16A coexpressed with the gamma subunit phagocytosed EA. The phagocytosis mediated by CD16A expressed on CHO cells was independent of divalent cations but dependent on intact microfilaments. CHO cells expressing CD16A-gamma and CD16A-zeta chimeras also phagocytosed EA. The phagocytosis was specifically inhibited by tyrphostin-23, a tyrosine kinase inhibitor. In summary, our results demonstrate that glycosylphosphatidylinositol-anchored CD16B alleles differ from CD16A in their ability to mediate phagocytosis. Furthermore, since studies with other Fc gamma Rs have shown that CHO cells lack the phagocytic pathway mediated by the cytoplasmic domain of Fc gamma Rs, the phagocytosis of EA by CHO cells stably transfected with CD16A and CD16A-subunit chimera provides an ideal system to dissect the phagocytic signaling pathways mediated by these Fc gamma R-associated subunits.

Animals↗

PubMed1995

Fas-activated serine/threonine kinase (FAST) phosphorylates TIA-1 during Fas-mediated apoptosis.

We have identified a serine/threonine kinase that is rapidly activated during Fas-mediated apoptosis. Fas-activated serine/threonine kinase (FAST) is phosphorylated on serine and threonine residues in Jurkat cells. In response to Fas ligation, it is rapidly dephosphorylated and concomitantly activated to phosphorylate TIA-1, a nuclear RNA-binding protein that has been implicated as an effector of apoptosis. Phosphorylation of TIA-1 precedes the onset of DNA fragmentation, suggesting a role in signaling downstream events in the apoptotic program. Our results introduce Fast and TIA-1 as components of a molecular cascade involved in signaling Fas-mediated apoptosis.

Amino Acid Sequence↗

PubMed1995

A phase II trial of human recombinant interleukin-2 administered as a 4-day continuous infusion for children with refractory neuroblastoma, non-Hodgkin's lymphoma, sarcoma, renal cell carcinoma, and malignant melanoma. A Childrens Cancer Group study.

BACKGROUND: Recombinant human Interleukin-2 (IL-2) has been effective at inducing measurable antitumor responses in adults with renal cell carcinoma and melanoma. It also is being tested as adjuvant therapy for patients with acute myeloid leukemia after autologous bone marrow transplantation. METHODS: The authors tested IL-2 in a pediatric Phase II trial using a regimen that has antitumor effects in adults and was proven to be tolerated acceptably in a prior Phase I pediatric trial. Thirty-eight patients were entered into this study of whom 36 received IL-2 and were evaluable (20 with sarcoma, 9 with neuroblastoma, 5 with renal cell carcinoma, and 1 each with melanoma and lymphoma). RESULTS: Interleukin-2 dose modifications were based on tolerance and toxicity, such that 46% of these patients received a 50% increase in IL-2 dose during the second week, and 81% of those receiving the elevated dose continued receiving this dose level during the third week of treatment. A single patient with renal cell carcinoma had a complete response that was maintained; the remaining 35 patients did not show objective evidence of tumor response sufficient to qualify as either a complete response or a partial response. CONCLUSIONS: Absolute lymphocyte counts were indicative of the immunostimulatory effect of this IL-2 regimen as observed for adults, with a median 7.2-fold increase. Nevertheless, despite immune activation, a sufficient number of patients were evaluated, indicating that IL-2 does not have measurable antitumor effects in children with large refractory sarcomas or neuroblastomas, whereas one of five children with renal cell carcinoma had a complete response, consistent with the 10-20% response rate observed in adults.

Adolescent↗

PubMed1995

Alterations in the signal-transducing molecules of T cells and NK cells in colorectal tumor-infiltrating, gut mucosal and peripheral lymphocytes: correlation with the stage of the disease.

T cells from mice bearing an experimental colon carcinoma, and from patients with colorectal and renal carcinomas, have atypical T-cell receptors (TCR). In the present study, further characterization of modulations in CD3- and CD16-associated zeta chain in peripheral blood lymphocytes (PBL) and tumor-infiltrating lymphocytes (TIL) from colorectal carcinomas was performed. Relative to PBL, the percentage of natural killer (NK) cells among fresh TIL was reduced, while a higher proportion of T cells expressing HLA-DR was found. As previously reported, we found significantly reduced levels of the CD3- and CD16-associated zeta chain in TIL and, to a lesser extent, also in patients' PBL. Levels of zeta chain in T and NK cells from non-cancerous colorectal tissue from patients were lower than in PBL but higher than in TIL, with a direct relationship between levels of this signal-transducing molecule and the distance from the tumor. In addition, zeta levels correlated with the Dukes' stage of the disease, since PBL from patients with lymph-node involvement or distant organ metastases (Dukes' stages C and D) had significantly less CD3 zeta than patients with localized disease (stages A and B). Patients' T cells also had decreased levels of cell-surface and cytoplasmic CD3 epsilon. We also observed reduced levels of the TCR accessory molecules CD4 and CD8, mainly on TIL but to a lesser extent also on patients' PBL. Biochemical analysis of anti-CD3 epsilon-immunoprecipitated TCR complexes demonstrated that the CD3 complex was not associated with the zeta chain, either on TIL or on PBL or on lymphocytes from non-cancerous colon tissue, suggesting a defect in the assembly of the TCR complex. Following several days of in vitro culture with recombinant interleukin-2 and phytohemagglutinin, anti-CD3 or anti-CD2 monoclonal antibodies (MAbs), levels of CD3 zeta chain as well as of cell surface CD3 epsilon were normalized. Our findings suggest an abnormal expression as well as assembly of several different signal-transducing molecules of T cells and NK cells, which correlate with the stage of the disease in patients with colorectal carcinomas.

Adult↗

PubMed1995

Interchangeability of conjugated Haemophilus influenzae type b vaccines in infants.

OBJECTIVE: To evaluate the safety and immunogenicity of two Haemophilus influenzae type b (Hib) conjugate vaccines when administered in serial combination. These vaccines consisted of Hib capsular polysaccharide polyribosyl-ribitol phosphate (PRP) conjugated to the meningococcal outer membrane protein (OMP) complex (PRP-OMP) and H influenzae oligosaccharide conjugated to a mutant toxin (CRM197) isolated from Corynebacterium diphtheriae (HbOC). DESIGN: Randomized, double-blind, clinical trial evaluating five Hib vaccination regimens. SETTING: Vaccine Treatment and Evaluation Units and affiliated private pediatric practices at Saint Louis (Mo) University, Vanderbilt University, Nashville, Tenn, and Baylor College of Medicine, Houston, Tex. PATIENTS: A total of 497 healthy 2-month-old infants scheduled to receive routine immunization. INTERVENTION: Participants received either PRP-OMP or HbOC given as recommended by the manufacturer, PRP-OMP at 2 and 6 months, HbOC at 2 months, then PRP-OMP at 4 and 6 months, or PRP-OMP at 2 months and then HbOC at 4 and 6 months. Unconjugated PRP was given at 15 months to evaluate priming. RESULTS: Geometric mean antibody concentrations differed significantly among the groups following the second and third immunizations of the primary series and following booster immunization with unconjugated PRP. On each occasion, the groups receiving serial combinations of PRP-OMP and HbOC achieved mean antibody concentrations that equalled or exceeded those of the groups receiving a single product. Adverse reactions did not vary by group. CONCLUSIONS: The studied sequential combinations of Hib vaccines were safe and at least as immunogenic as either vaccine alone.

Antibodies, Bacterial↗

PubMed1995

The RNA-binding protein TIAR is translocated from the nucleus to the cytoplasm during Fas-mediated apoptotic cell death.

We have determined the structure, intracellular localization, and tissue distribution of TIAR, a TIA-1-related RNA-binding protein. Two related isoforms of TIAR, migrating at 42 and 50 kDa, are expressed in primate cells. Unlike TIA-1, which is found in the granules of cytotoxic lymphocytes, TIAR is concentrated in the nucleus of hematopoietic and nonhematopoietic cells. Because TIAR can trigger DNA fragmentation in permeabilized thymocytes, it is a candidate effector of apoptotic cell death. Consistent with this possibility, we have found that the expression and intracellular localization of TIAR change dramatically during Fas-mediated apoptosis. TIAR moves from the nucleus to the cytoplasm within 30 min of Fas ligation. Redistribution of TIAR precedes the onset of DNA fragmentation and is not a nonspecific consequence of nuclear disintegration. Cytoplasmic redistribution of TIAR is not observed during cellular activation triggered by mitogens such as concanavalin A or phytohemagglutinin. Our results suggest that cytoplasmic redistribution of TIAR may be a general feature of the apoptotic program.

Antibodies, Monoclonal↗

PubMed1995

BOLD based functional MRI at 4 Tesla includes a capillary bed contribution: echo-planar imaging correlates with previous optical imaging using intrinsic signals.

We show that the EPI time course in functional MR imaging at 4 Tesla displays a multiphasic response in response to photic stimulation. Focal areas of gray matter display an initial negative change in signal intensity that reaches a maximum of 1% about 2 s after the onset of photic stimulation. This component then changes sign, reaching a positive maximum about 5 s after the onset of the stimulus. Other areas, including those where draining veins are visible, show only a positive signal change, reaching a maximum of about 6% after the onset of the visual stimulus. These time constants are in very good agreement with published data obtained with intrinsic optical mapping techniques, where a deoxygenation phase has been shown to occur in functionally specific cortical columns, followed by an increase in blood volume which is more distributed in nature. Thus, we believe there is strong evidence that a sensitivity to capillary oxygenation state is present in high S/N functional MR images obtained with EPI at 4 T.

Artifacts↗

PubMed1995

A prospective randomized comparison of an attached silver-impregnated cuff to prevent central venous catheter-associated infection.

The VitaCuff catheter, a specialized central venous catheter (CVC) with an attached silver-impregnated cuff, is designed to permit percutaneous placement and prolonged venous access. A prospective randomized study was undertaken comparing the VitaCuff with standard triple lumen catheters to determine if the VitaCuff reduces infection during extended use. All consenting patients underwent percutaneous placement of subclavian lines. By study design, control and VitaCuff catheters could remain in site for up to 7 and 14 days, respectively. Cultures were obtained from the preinsertion skin site, and upon removal, from the skin, hubs, infusates, CVC tip, and cuff. Statistical methods included chi 2, the Student t test, and the log-rank test on Kaplan-Meier estimates. Of 133 patients completing this study, 64 patients (48.1%) underwent VitaCuff placement and 69 patients (51.8%) served as controls. In 124 patients (93.2%), the indication for catheter placement was for perioperative care. Overall, 67 patients (50.4%) required central venous access > 7 days, necessitating > or = 1 additional line in 29 patients (21.8%). The incidence of pneumothorax per patient from the initial central line insertion was 4/104 (3.85%), significantly lower than the 4/29 (13.8%) incidence during secondary catheter placement (P = 0.046). Culture results upon catheter removal demonstrated a reduction in colonization of skin sites and hubs for the VitaCuff patients, but not for catheter tips or infusates. Regardless of the type of catheter used, colonization was dependent upon duration of insertion. The incidence of catheter-related sepsis was 6.8%, and did not differ significantly between the study groups. Multiple CVC insertions increase the incidence of pneumothorax. Because VitaCuff catheters permit extended access up to 14 days without increasing the incidence of sepsis, we recommend their use in patients who require prolonged CVC access.

Catheterization, Central Venous↗

PubMed1995

TIA-1: structural and functional studies on a new class of cytolytic effector molecule.

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Amino Acid Sequence↗

PubMed1995

Mineral concentration gradients in rat femoral diaphyses measured by X-ray microtomography.

The bone mineral concentrations of five rat femora were measured as a function of distance from the distal metaphysis by quantitative X-ray microtomography (XMT) at a resolution of approximately 23 x 23 x 15 microns3. Assuming the mineral phase of bone to be hydroxyapatite, Ca10 (PO4)6(OH)2, the mean cortical mineral concentration (CM) per transverse section was found to range from 1.33 to 1.47 g cm-3. Detectable variations in the bone mineral concentration between sections of femora from different animals could not be attributed to the age when the particular animal was sacrificed. An increase in CM with distance, L, from the distal growth plate was observed and a saturating exponential equation, CM = a - be-alpha L, was used to describe the changes. Each section of bone tissue was considered as a population of elementary volumes of bone (EVB) and L was related to the age of the EVB (TEVB). A simple model for the mineralization process of an EVB was then proposed. Each newly formed EVB accumulated mineral rapidly to give an initial mineral concentration of approximately 1.3 g cm-3 (parameter a-b). Their mineral concentrations then increased asymptotically to approximately 1.5 g cm-3 (parameter a) with a time constant of approximately 330 days. This slow maturation process is attributed to Ostwald ripening of the bone crystals with further crystal growth using ions from the extracellular fluid.

Analysis of Variance↗