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Biomedical subjects

J M Green

Publications and source records attributed to J M Green.

At least 19 recordsLinked to original sources
PubMed2000

Coordinate regulation of T cell activation by CD2 and CD28.

T cell activation requires co-engagement of the TCR with accessory and costimulatory molecules. However, the exact mechanism of costimulatory function is unknown. Mice lacking CD2 or CD28 show only mild deficits, demonstrating that neither protein is essential for T cell activation. In this paper we have generated mice lacking both CD2 and CD28. T cells from the double-deficient mice have a profound defect in activation by soluble anti-CD3 Ab and Ag, yet remain responsive to immobilized anti-CD3. This suggests that CD2 and CD28 may function together to facilitate interactions of the T cell and APC, allowing for efficient signal transduction through the TCR.

Animals↗
PubMed2000

Gender differences in sweat lactate.

Sweat rate may affect sweat lactate concentration. The current study examined potential gender differences in sweat lactate concentrations because of varying sweat rates. Males (n = 6) and females (n = 6) of similar age, percentage body fat, and maximal oxygen consumption (VO2max) completed constant load (CON) cycling (30 min--approximately 40% VO2max) and interval cycling (INT) (15 1-min intervals each separated by 1 min of rest) trials at 32 (1) degrees C wet bulb globe temperature (WBGT). Trials were preceded by 15 min of warm-up (0.5 kp, 60 rpms) and followed by 15 min of rest. Blood and sweat samples were collected at 15, 25, 35, 45, and 60 min during each trial. Total body water loss was used to calculate sweat rate. Blood lactate concentrations (CON approximately equal to 2 mmol.l-1, INT approximately equal to 6 mmol.l-1) and sweat lactate concentrations (CON and INT approximately equal to 12 mmol.l-1) were not significantly different (P > 0.05) at any time between genders for CON or INT. Overall sweat rates (ml.h-1) were not significantly different (P > 0.05) between trials but were significantly greater (P < or = 0.05) for males than for females for CON [779.7 (292.6) versus 450.3 (84.6) ml.h-1] and INT [798.0 (268.3) versus 503.0 (41.4) ml.h-1]. However, correcting for surface area diminished the difference [CON: 390.7 (134.4) versus 277.7 (44.4) ml.h-1, INT: 401.5 (124.1) versus 310.6 (23.4) ml.h-1 (P < or = 0.07)]. Estimated total lactate secretion was significantly greater (P < or = 0.05) in males for CON and INT. Results suggest that sweat rate differences do not affect sweat lactate concentrations between genders.

Adult↗
PubMed1999

Role of cholesterol in formation and function of a signaling complex involving alphavbeta3, integrin-associated protein (CD47), and heterotrimeric G proteins.

Integrin-associated protein (CD47) is a multiply membrane spanning member of the immunoglobulin superfamily that regulates some adhesion-dependent cell functions through formation of a complex with alphavbeta3 integrin and trimeric G proteins. Cholesterol is critical for the association of the three protein components of the supramolecular complex and for its signaling. The multiply membrane spanning domain of IAP is required for complex formation because it binds cholesterol. The supramolecular complex forms preferentially in glycosphingolipid-enriched membrane domains. Binding of mAb 10G2 to the IAP Ig domain, previously shown to be required for association with alphavbeta3, is affected by both the multiply membrane spanning domain and cholesterol. These data demonstrate that cholesterol is an essential component of the alphavbeta3/IAP/G protein signaling complex, presumably acting through an effect on IAP conformation.

Antibodies, Monoclonal↗
PubMed1999

Proline residues in CD28 and the Src homology (SH)3 domain of Lck are required for T cell costimulation.

The Src family tyrosine kinases Lck and Fyn are critical for signaling via the T cell receptor. However, the exact mechanism of their activation is unknown. Recent crystal structures of Src kinases suggest that an important mechanism of kinase activation is via engagement of the Src homology (SH)3 domain by proline-containing sequences. To test this hypothesis, we identified several T cell membrane proteins that contain potential SH3 ligands. Here we demonstrate that Lck and Fyn can be activated by proline motifs in the CD28 and CD2 proteins, respectively. Supporting a role for Lck in CD28 signaling, we demonstrate that CD28 signaling in both transformed and primary T cells requires Lck as well as proline residues in CD28. These data suggest that Lck plays an essential role in CD28 costimulation.

Alanine↗
PubMed1999

Structural requirements for CD43 function.

The regulation of T cell activation and adhesion by CD43 (leukosialin, sialophorin) has been thought to be mainly a function of the large size and negative charge of the extracellular domain of the protein. In this work, we demonstrate that the cytoplasmic tail is both necessary and sufficient for the negative regulatory effect of CD43 on cell-cell adhesion. Expression of mutant CD43 proteins in primary T cells from CD43-deficient mice demonstrated that the antiproliferative effect of CD43 is also dependent upon the cytoplasmic tail. In contrast, Ab-mediated costimulation through CD43 does not require the intracellular domain of CD43. These data demonstrate that CD43 primarily serves as a negative regulator of T cell activation and adhesion, and that this is mediated not exclusively by passive effects of the extracellular domain, but requires participation of the cytoplasmic tail, perhaps through interactions with the cytoskeleton, or alternatively, active regulation of intracellular signaling pathways.

Animals↗
PubMed1999

Cell spreading distinguishes the mechanism of augmentation of T cell activation by integrin-associated protein/CD47 and CD28.

Integrin-associated protein (IAP/CD47) is a 50 kDa transmembrane protein initially defined as a regulator of beta3 integrin-mediated functions in neutrophils. IAP also can synergize with the TCR in T cell activation independent of beta3 integrins. To analyze the mechanism for IAP synergy with TCR, we expressed in Jurkat cells a chimeric molecule, consisting of the CD16 extracellular domain, the CD7 transmembrane domain and the TCR zeta chain cytoplasmic tail (CD16-7-zeta), which on its own is unable to induce IL-2 production. Ligation of IAP acted in synergy with TCR to induce IL-2 transcription and synthesis, but failed to synergize with the signal generated by CD16-7-zeta, while CD28 was a potent co-stimulator with both TCR and CD16-7-zeta. The failure of IAP to activate Jurkat together with CD16-7-zeta correlated with a lack of c-Jun N-terminal kinase, but not extracellular-signal-regulated kinase activation. Jurkat adhesion to anti-IAP, but not anti-CD28, induced cell spreading and the same domains of IAP required for augmentation of T cell activation were required to induce cell spreading. IAP synergy with TCR signaling likely results from its ability to stimulate adhesion to a ligand-expressing surface or antigen-presenting cell (APC), rather than from initiation of a novel signaling cascade. We conclude that a major role for ligation of IAP in T cell activation is to enhance the efficiency of TCR signaling by causing T cells to spread on an APC or surface.

Antigens, CD↗
PubMed1999

The validity of ratings of perceived exertion for cross-modal regulation of swimming intensity.

BACKGROUND: This study examined the use of Borg's category Ratings of Perceived Exertion (RPE) scale for prescribing and self-regulating swimming intensity. Subjects were males and females (n = 19) ages nineteen to fifty-eight who regularly swam for fitness. EXPERIMENTAL DESIGNS: Subjects completed six trials. Each trial was separated by a minimum of forty-eight hours. Mean cycle ergometry heart rates at estimated RPE-overall 12 and 16 were compared to mean swimming heart rates at produced RPE-overall 12 and 16. Also, mean arm ergometry heart rates at estimated RPE-arms 12 and 16 were compared to mean swimming heart rates at produced RPE-arms 12 and 16. Cycling and arm ergometry anchor trials familiarized subjects with testing protocol and Borg's scale prior to estimation and production trials. Comparisons were made using a one-way ANOVA (alpha 0.05). RESULTS: Mean cycling heart rate at RPE-overall 16 was not significantly different from mean swimming heart rate at RPE-overall 16. Mean swimming heart rate was significantly greater than cycling heart rate at RPE-overall 12. Mean swimming heart rates at RPE-arms 12 and 16 were significantly greater than arm ergometry heart rates at RPE-arms 12 and 16. CONCLUSIONS: Results suggest that RPE-overall 16 may be useful in prescribing a higher exercise intensity for swimmers. However, adjustments from RPE-overall 12 are needed for establishing a lower intensity. Additionally, arm ergometry-based RPEs may require adjustments to be effective in prescribing and regulating swimming intensity. Results suggest that cycling and arm ergometry anchored RPE responses should be used with discretion when prescribing and regulating swimming intensity.

Adult↗
PubMed1998

Regulation of in vitro and in vivo T cell activation by CD43.

Accessory molecule interactions can be critical in determining the outcome of a T cell's encounter with antigen. Cell adhesion proteins may augment T cell responses by facilitating TCR engagement of the antigen-MHC complex, while co-stimulatory molecules may deliver distinct signals that modulate T cell responsiveness. CD43 (leukosialin, sialophorin) has been suggested to influence cell activation by steric hindrance based upon the large size and glycosylation of the protein, as well as the relative abundance of the protein on the cell surface. In this paper we examine both in vitro and in vivo T cell-dependent responses in CD43-deficient mice. We demonstrate that T cells from CD43-deficient mice are hyper-responsive following both in vivo and in vitro activation, and that this is observed in response to not only TCR-CD3-mediated stimulation, but also following receptor-independent activation. This data suggests that mechanisms other than non-specific steric hindrance are important in the regulation of T cell activation by CD43.

Animals↗
PubMed1998

Characterization of mutations that allow p-aminobenzoyl-glutamate utilization by Escherichia coli.

An Escherichia coli strain deficient in p-aminobenzoate synthesis was mutagenized, and derivatives were selected for growth on folic acid. Supplementation was shown to be due to p-aminobenzoyl-glutamate present as a breakdown product in commercial folic acid preparations. Two classes of mutations characterized by the minimum concentration of p-aminobenzoyl-glutamate that could support growth were obtained. Both classes of mutations were genetically and physically mapped to about 30 min on the E. coli chromosome. A cloned wild-type gene from this region, abgT (formerly ydaH) could confer a similar p-aminobenzoyl-glutamate utilization phenotype on the parental strain. Interruption of abgT on the plasmid or on the chromosome of the mutant strain resulted in a loss of the phenotype. abgT was the third gene in an apparent operon containing abgA, abgB, abgT, and possibly ogt and might be regulated by a divergently transcribed LysR-type regulator encoded by abgR. Two different single-base-pair mutations that gave rise to the p-aminobenzoyl-glutamate utilization phenotype lay in the abgR-abgA intercistronic region and appeared to allow the expression of abgT. The second class of mutation was due to a tandem duplication of abgB and abgT fused to fnr. The abgA and abgB gene products were homologous to one another and to a family of aminoacyl aminohydrolases. p-Aminobenzoyl-glutamate hydrolysis could be detected in extracts from several of the mutant strains, but intact abgA and abgB were not essential for p-aminobenzoyl-glutamate utilization when abgT was supplied in trans.

Amino Acid Sequence↗
PubMed1998

Role of epidermal growth factor receptor family members in growth and differentiation of breast carcinoma.

Members of the epidermal growth factor (EGF) family of tyrosine kinase receptors are involved in the regulation of cell growth and differentiation, and are found to be expressed in many types of cancers. Activation of these receptors can be elicited by multiple ligands, resulting in the formation of a spectrum of heterodimer complexes and a number of biological outcomes. A clear demonstration of biological activation by a single complex has been difficult to address because of the endogenous expression of HERs (human EGF-like receptors) in many cell lines. We have generated a collection of cell lines expressing all HERs alone or in all pairwise combinations in a clone of NIH 3T3 cells (3T3-7d) devoid of detectable EGF receptor family members. Transformation, as measured by growth in soft agar, only occurred in cells expressing two different HER family members. Transformation with activated Neu and the rate of in vivo tumour formation were also correlated with the expression of multiple HERs in the same cell. To further our understanding of the role of heterodimer signalling, we demonstrated that, within a breast carcinoma cell line, activation of HER-3 results in cellular differentiation, prolonged activation of extracellular-signal-related kinase 1 (ERK1) activity and an increase in p21CIP1/WAF1 nuclear staining. In contrast, activation of HER-4 is mitogenic, induces transient activation of ERK1 activity and decreases the nuclear staining of p21CIP1/WAF1. These differences in biochemical and biological responses are correlated with the contrasting abilities of HER-3 and HER-4 to be down-regulated from the cell surface. The cell-surface localization of HER-3 does not change in response to ligand, whereas activation of HER-4 results in a loss of cell-surface staining followed by accumulation into a perinuclear compartment.

3T3 Cells↗
PubMed1997

Role for a glycan phosphoinositol anchor in Fc gamma receptor synergy.

While many cell types express receptors for the Fc domain of IgG (Fc gamma R), only primate polymorphonuclear neutrophils (PMN) express an Fc gamma R linked to the membrane via a glycan phosphoinositol (GPI) anchor. Previous studies have demonstrated that this GPI-linked Fc gamma R (Fc gamma RIIIB) cooperates with the transmembrane Fc gamma R (Fc gamma RIIA) to mediate many of the functional effects of immune complex binding. To determine the role of the GPI anchor in Fc gamma receptor synergy, we have developed a model system in Jurkat T cells, which lack endogenously expressed Fc gamma receptors. Jurkat T cells were stably transfected with cDNA encoding Fc gamma RIIA and/or Fc gamma RIIIB. Cocrosslinking the two receptors produced a synergistic rise in intracytoplasmic calcium ([Ca2+]i) to levels not reached by stimulation of either Fc gamma RIIA or Fc gamma RIIIB alone. Synergy was achieved by prolonged entry of extracellular Ca2+. Cocrosslinking Fc gamma RIIA with CD59 or CD48, two other GPI-linked proteins on Jurkat T cells also led to a synergistic [Ca2+]i rise, as did crosslinking CD59 with Fc gamma RIIA on PMN, suggesting that interactions between the extracellular domains of the two Fc gamma receptors are not required for synergy. Replacement of the GPI anchor of Fc gamma RIIIB with a transmembrane anchor abolished synergy. In addition, tyrosine to phenylalanine substitutions in the immunoreceptor tyrosine-based activation motif (ITAM) of the Fc gamma RIIA cytoplasmic tail abolished synergy. While the ITAM of Fc gamma RIIA was required for the increase in [Ca2+]i, tyrosine phosphorylation of crosslinked Fc gamma RIIA was diminished when cocrosslinked with Fc gamma RIIIB. These data demonstrate that Fc gamma RIIA association with GPI-linked proteins facilitates Fc gamma R signal transduction and suggest that this may be a physiologically significant role for the unusual GPI-anchored Fc gamma R of human PMN.

Antigens, CD↗
PubMed1997

CD28 engagement and proinflammatory cytokines contribute to T cell expansion and long-term survival in vivo.

To mount a productive response to Ag, CD4+ T cells in mice must divide, differentiate, and survive at least until the Ag has been eliminated. It has been suggested that to accomplish this, T cells must receive two signals, one through their TCRs and a second through CD28. The second signal through CD28 has been thought to fulfill two roles, to stimulate T cell proliferation and to promote T cell survival. In this paper we confirm that CD28 engagement can contribute to vigorous T cell expansion in mice injected with superantigens. However, CD28 engagement does not protect T cells produced during a superantigen-specific proliferative response from undergoing subsequent deletion. Even if CD28 is bound, 4 days after superantigen exposure, the majority of T cells produced in response to superantigen exposure are eliminated in vivo. In contrast, this loss of superantigen-stimulated T cells can be prevented by the inflammatory stimuli created by injection of bacterial LPS. This protection does not require engagement of CD28 by its ligands, B7-1 and B7-2. These data suggest that productive T cell responses in mice involve a number of signals, including those initiated through TCR and CD28, which are primarily involved in the activation and expansion of T cells, and others delivered by proinflammatory cytokines that protect an activated T cell from subsequent deletion.

Abatacept↗
PubMed1997

Women's need for information before attending genetic counselling for familial breast or ovarian cancer: a questionnaire, interview, and observational study.

OBJECTIVES: To describe women's information needs prior to genetic counselling for familial breast or ovarian cancer. DESIGN: Prospective study including semistructured telephone interviews before genetic counselling, observations of consultations, completion of postal questionnaires, and face-to face interviews within two months of counselling. SUBJECTS: 46 women attending genetic counselling for familial breast or ovarian cancer. MAIN OUTCOME MEASURES: Subjects' understanding of process and content of genetic counselling before attending and attitudes about their preparation for the counselling session. RESULTS: Although all women interviewed before the clinic expected to discuss their risk of developing cancer and risk management options, there was evidence of a lack of knowledge about the process and content of genetic counselling, 17 (37%) women said they did not know what else would happen. Most women interviewed after counselling viewed it positively, but 26 (65%) felt they had been inadequately prepared and 11 (28%) felt that their lack of preparation meant that they could not be given an accurate estimation of their risk of cancer. CONCLUSIONS: Some women felt that they did not obtain optimum benefit from genetic counselling because they were inadequately prepared for it. We suggest that cancer family history clinics should provide women with written information about the process and content of genetic counselling before their clinic attendance.

Adult↗
PubMed1997

Preimplantation diagnosis and other reproductive options: attitudes of male and female carriers of recessive disorders.

This questionnaire-based study describes in detail the attitudes of 245 carriers of recessive disorders to preimplantation diagnosis (PID). These are compared to responses to other reproductive options for carriers, that is prenatal diagnosis (PND), adoption, donor insemination (DI) and egg donation. The positive attitudes of this sample to PID and corresponding negative attitudes to adoption, DI and egg donation are striking. Almost the entire sample indicated that they viewed early reassurance and the opportunity to avoid a termination of pregnancy as important advantages of PID. Disadvantageous aspects of PID were acknowledged, but did not result in many participants finding this approach unacceptable. The study revealed a number of differences between men and women, especially in their perceptions of the disadvantages of PID. Women saw the practical difficulties of PID as more important disadvantages than did the men, and were very much more concerned about the issues involving embryos. Men, conversely, rated the impact upon their partners as a more important disadvantage. These findings underline the importance of considering both sets of views. Despite support for PID, in an overall assessment it did not displace PND as the most useful reproductive option.

Adoption↗