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Biomedical subjects

P D King

Publications and source records attributed to P D King.

At least 19 recordsLinked to original sources
PubMed2001

A transcription function for the T cell-specific adapter (TSAd) protein in T cells: critical role of the TSAd Src homology 2 domain.

T cell-specific adapter (TSAd) protein is an Src homology 2 (SH2) domain-containing adapter molecule implicated in T cell receptor for antigen (TCR)-mediated interleukin 2 (IL-2) secretion in T cells. Here, we demonstrate that a substantial fraction of TSAd is found in the T cell nucleus. Nuclear import of TSAd is an active process that depends on TSAd SH2 domain recognition of a phosphotyrosine-containing ligand. Importantly, we show that TSAd can act as a potent transcriptional activator in T cells. Furthermore, the TSAd SH2 domain appears to be essential for this transcription-activating function independent of its role in nuclear import. Biochemical analyses suggest that a single TSAd SH2 domain ligand of 95-100 kD may be involved in these processes. Consistent with a role as a transcription activator, cotransfection of TSAd with an IL-2 promoter-reporter gene construct results in a considerable upregulation of IL-2 promoter activity. Further, we show that this augmentation requires a functional TSAd SH2 domain. However, TSAd does not appear to modulate the activity of the major recognized IL-2 gene transcription factors, nuclear factor kappaB (NF-kappaB), nuclear factor of activated T cells (NFAT), or activator protein 1 (AP-1). These findings point to the function of TSAd as a novel transcription-regulatory protein in T cells and illustrate the importance of the TSAd SH2 domain in this role.

Active Transport, Cell Nucleus↗
PubMed2001

Negative-feedback regulation of CD28 costimulation by a novel mitogen-activated protein kinase phosphatase, MKP6.

TCR and CD28 costimulatory receptor-cooperative induction of T cell IL-2 secretion is dependent upon activation of mitogen-activated protein (MAP) kinases. Using yeast-hybrid technology, we cloned a novel CD28 cytoplasmic tail (CD28 CYT) interacting protein, MAP kinase phosphatase-6 (MKP6), which we demonstrate inactivates MAP kinases. Several lines of evidence indicate that MKP6 plays an important functional role in CD28 costimulatory signaling. First, in human peripheral blood T cells (PBT), expression of MKP6 is strongly up-regulated by CD28 costimulation. Second, transfer of dominant-negative MKP6 to PBT with the use of retroviruses primes PBT for the secretion of substantially larger quantities of IL-2, specifically in response to CD28 costimulation. A similar enhancement of IL-2 secretion is observed neither in response to TCR plus CD2 costimulatory receptor engagement nor in response to other mitogenic stimuli such as phorbol ester and ionomycin. Furthermore, this hypersensitivity to CD28 costimulation is associated with CD28-mediated hyperactivation of MAP kinases. Third, a retroviral transduced chimeric receptor with a CD28 CYT that is specifically unable to bind MKP6 costimulates considerably larger quantities of IL-2 from PBT than a similar transduced chimeric receptor that contains a wild-type CD28 CYT. Taken together, these results suggest that MKP6 functions as a novel negative-feedback regulator of CD28 costimulatory signaling that controls the activation of MAP kinases.

Adult↗
PubMed2001

Hepatotoxicity of chemotherapy.

After assessment of tumor histology, the next important factor to consider in the selection of a chemotherapy regime is organ function. Patients who are to receive chemotherapy require careful assessment of liver function prior to treatment to determine which drugs may not be appropriate, and which drug doses should be modified. Following therapy abnormalities of liver function tests may be due to the therapy rather than to progressive disease, and this distinction is of critical importance. Furthermore, not all abnormalities in liver function are due to the tumor or its treatment, and other processes, such as hepatitis, must be kept in mind. This article reviews the hepatic toxicity of chemotherapeutic agents, and suggests dose modifications based upon liver function abnormalities. Emphasis is placed on agents known to be hepatotoxic, and those agents with hepatic metabolism.

Antineoplastic Agents↗
PubMed2000

T cell-specific adapter protein inhibits T cell activation by modulating Lck activity.

We previously reported the isolation of a cDNA encoding a T cell-specific adapter protein (TSAd). Its amino acid sequence contains an SH2 domain, tyrosines in protein binding motifs, and proline-rich regions. In this report we show that expression of TSAd is induced in normal peripheral blood T cells stimulated with anti-CD3 mAbs or anti-CD3 plus anti-CD28 mAbs. Overexpression of TSAd in Jurkat T cells interfered with TCR-mediated signaling by down-modulating anti-CD3/PMA-induced IL-2 promoter activity and anti-CD3 induced Ca2+ mobilization. The TCR-induced tyrosine phosphorylation of phospholipase C-gamma1, SH2-domain-containing leukocyte-specific phosphoprotein of 76kDa, and linker for activation of T cells was also reduced. Furthermore, TSAd inhibited Zap-70 recruitment to the CD3zeta-chains in a dose-dependent manner. Consistent with this, Lck kinase activity was reduced 3- to 4-fold in COS-7 cells transfected with both TSAd and Lck, indicating a regulatory effect of TSAd on Lck. In conclusion, our data strongly suggest an inhibitory role for TSAd in proximal T cell activation.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Usefulness of a pediatric colonoscope for colonoscopy in adults.

BACKGROUND: There are few published data on how different types of colonoscopes affect success in reaching the cecum and patient comfort. We examined the feasibility of using a pediatric colonoscope for routine colonoscopy in adults and investigated whether there were subgroups of patients in whom use of this instrument was preferable. METHODS: One-hundred fifty adults undergoing outpatient colonoscopy were randomized to colonoscopy with a standard colonoscope (Olympus CF-100L) or with a pediatric colonoscope (Olympus PCF-100). All procedures were performed by a faculty endoscopist and timed by an independent observer. After examinations, the endoscopist graded procedure difficulty and patients were given a questionnaire that assessed their experience. RESULTS: The adult (n = 77) and pediatric (n = 73) colonoscope groups were comparable in all outcomes measured, including success in reaching the cecum (91% vs. 93%, p = 0.61), mean time to reach the cecum (11.4 vs. 9.7 min, p = 0.07), mean total procedure time (21.8 vs. 21.9 min, p = 0.95), mean meperidine dose (55 vs. 52 mg, p = 0.17); median midazolam dose (2.0 mg in both groups, p = 0.10), the endoscopists' perception of procedure difficulty, and patient comfort scales. Of the 7 patients in whom colonoscopy with the adult colonoscope was unsuccessful, the cecum was reached in 4 by switching to a pediatric colonoscope (all women, 3 of whom had prior hysterectomy). In the 5 patients in whom colonoscopy with the pediatric colonoscope was unsuccessful, the cecum was reached in 1 by switching to an adult colonoscope. Including the cases in which the cecum was reached by switching to the alternative colonoscope, the overall frequency of cecal intubation was 143 of 150 (95%). Subgroup analysis disclosed no difference between the 2 groups in outcomes when gender, presence of diverticulosis, and patient size were considered. Colonoscopy with the pediatric colonoscope was more successful than with the adult instrument in reaching the cecum in women with prior hysterectomy (11 of 12 [92%] vs. 15 of 21 [71%]); however, the numbers in each group were relatively small and the difference was not significant (p = 0.22). CONCLUSIONS: The pediatric colonoscope is suitable for routine colonoscopy in adults. It is also useful in patients in whom colonoscopy with the adult colonoscope is unsuccessful in reaching the cecum (particularly in women). Additional study is needed to see if the pediatric colonoscope is actually superior to the adult colonoscopy for routine colonoscopy in women with prior hysterectomy.

Adult↗
PubMed1999

Benefit of intravenous antispasmodic (hyoscyamine sulfate) as premedication for colonoscopy.

BACKGROUND: We sought to determine whether premedication for colonoscopy with intravenous hyoscyamine sulfate (Levsin) was helpful from the standpoint of the colonoscopist and the patient. METHODS: One hundred sixteen adult patients were randomized to receive either 0.5 mg hyoscyamine sulfate intravenous (n = 57) or placebo (n = 59). After administration of study drug, patients were given meperidine and midazolam. Parameters measured included the time required to reach the cecum, total procedure time, and the endoscopist's perception of the adequacy of sedation, difficulty of insertion, and amount of colonic spasm on insertion and withdrawal. Patients were given a postprocedure questionnaire assessing their experience. RESULTS: In patients receiving hyoscyamine, there was a shorter cecal intubation time (median 9.2 vs. 12.9 minutes; p = 0. 01), shorter total colonoscopy time (median 20.5 vs. 25.0 minutes; p = 0.01), better patient sedation (p = 0.02), easier colonic insertion (p = 0.001), and less spasm on insertion (p = 0.01). No difference was found in the amount of spasm during withdrawal or the total dosages of meperidine or midazolam used. Patients receiving hyoscyamine sulfate reported being more comfortable during their procedures ( p < 0.001) and were more willing to repeat colonoscopy in the future (p = 0.0001). The only adverse effect seen during the study was a 27% incidence of sinus tachycardia that occurred in patients receiving hyoscyamine. CONCLUSIONS: Premedication with intravenous hyoscyamine sulfate was beneficial in terms of the time required for cecal intubation, total procedure time, adequacy of sedation, and scales of patient comfort. However, the high frequency of sinus tachycardia seen with the dose used in our study, which was extremely rapid in two patients, indicates the need for further study before the drug can be recommended as a routine premedication for colonoscopy.

Adolescent↗
PubMed1999

Patient attitudes toward undergoing colonoscopy without sedation.

OBJECTIVE: The vast majority of patients undergoing colonoscopy in the United States are given sedation. There are a number of potential advantages to performing colonoscopy without sedation. We sought to determine the attitude of patients toward unsedated colonoscopy in our three practice settings (a university medical center, a cancer center, and a Veterans Affairs medical center), and to see if there were factors that predicted willingness to try it. METHODS: Four-hundred thirty-four adult patients undergoing outpatient colonoscopy completed questionnaires before and after their procedures providing demographic information and assessing willingness to undergo colonoscopy without sedation. Patients were routinely given meperidine and midazolam for their procedures unless they specifically requested that they be unsedated (10 patients). RESULTS: Only 16.9% of our patients were willing to undergo colonoscopy on their preprocedure questionnaire. Willingness increased modestly on the postprocedure questionnaire to 22.6% (p = 0.01). Logistic regression analysis disclosed that male gender, having a college degree, low anxiety based on preprocedure anxiety scales, and lower doses of sedative drugs used during colonoscopy were the best predictors of willingness to undergo colonoscopy without sedation in the future. CONCLUSIONS: Only about a fifth of patients undergoing colonoscopy in our three practice settings expressed a willingness to try colonoscopy unsedated. Male gender, higher levels of education, and low anxiety scores on simple scales of preprocedure anxiety may help to predict willingness. Efforts to substantially increase the frequency of patients willing to undergo colonoscopy without sedation will likely require increased patient counseling and education.

Adult↗
PubMed1998

LCK-phosphorylated human killer cell-inhibitory receptors recruit and activate phosphatidylinositol 3-kinase.

HLA-specific killer cell inhibitory receptors (KIR) are thought to impede natural killer (NK) and T cell activation programs through recruitment of the SH2 domain-containing tyrosine phosphatases, SHP-1 and SHP-2, to their cytoplasmic tails (CYT). To identify other SH2 domain-containing proteins that bind KIR CYT, we used the recently described yeast two-bait interaction trap and a modified version of this system, both of which permit tyrosine phosphorylation of bait proteins. Using these systems, we show that KIR CYT, once phosphorylated by the src-family tyrosine kinase LCK, additionally bind the p85alpha regulatory subunit of phosphatidylinositol (PI) 3-kinase. Furthermore, we show that in an NK cell line, NK3.3, cross-linking of KIR results in recruitment of p85alpha to KIR and activation of PI 3-kinase lipid kinase activity. One consequence of KIR coupling to PI 3-kinase is downstream activation of the antiapoptotic protein kinase AKT. Therefore, in addition to providing negative signals, KIR may also contribute positive signals for NK and T cell growth and/or survival.

Cell Line↗
PubMed1998

Hepatitis C virus infection in non-Hodgkin's lymphoma.

Hepatitis C virus (HCV) infection is associated with immune-complex mediated disorders, including Type II mixed cryoglobulinaemia. Mixed cryoglobulinaemia is itself a low-grade B-cell lymphoproliferative disorder which may progress to non-Hodgkin's lymphoma (NHL). Studies from Europe and Asia have found a prevalence of hepatitis C virus infection in non-Hodgkin's lymphoma patients as high as 34%. Other viral infections are also associated with non-Hodgkin's lymphoma. We speculated that non-Hodgkin's lymphoma patients in the midwestern USA would have an increased prevalence of hepatitis C infection. We tested 73 patients with NHL and 20 controls with Hodgkin's disease for anti-HCV antibodies by EIA-2. Only 1/73 patients and no control subject was positive for anti-HCV. The anti-HCV positive patient had no identifiable risk factors for hepatitis C, and ALT was persistently normal. HCV-RNA testing by RT-PCR was negative. Thus, none of 73 non-Hodgkin's lymphoma patients could be confirmed to have hepatitis C infection. In a second part of the study, of 438 patients with HCV infection followed an average of 28.1 months, only one patient developed non-Hodgkin's lymphoma. We conclude that in our population, non-Hodgkin's lymphoma is not associated with hepatitis C virus infection. Based on these results and review of the literature, there are marked regional differences in the prevalence of hepatitis C infection in Non-Hodgkin's lymphoma.

Adult↗
PubMed1998

CD2-mediated activation of the Tec-family tyrosine kinase ITK is controlled by proline-rich stretch-4 of the CD2 cytoplasmic tail.

Ligation of the CD2 co-stimulatory receptor on human T lymphocytes induces tyrosine phosphorylation and activation of the Tec-family tyrosine kinase, ITK. To examine whether any of several proline-rich (PR) stretches of the CD2 cytoplasmic tail are necessary for ITK activation we introduced wild-type and mutated versions of rat CD2, each missing at least one PR stretch of the tail, into human Jurkat T leukemia cells. The influence of cytoplasmic tail mutations was then studied following stimulation of transfectants with the rat CD2 mAb pair, OX54/OX55. As predicted, wild-type rat CD2 was able to activate ITK in Jurkat cells. In addition, a truncation mutant, lacking the most membrane-distal PR stretch, PR6, was able to activate ITK. By contrast, all other studied truncation mutants, each of which is missing at least PR4-PR6, were unable to induce ITK activation. Of deletion mutants, deletion of the membrane-proximal PR stretches, PR1-PR3, did not impair rat CD2-mediated ITK activation. However, additional deletion of PR4 from a tail missing PR1 and PR2, deletion of PR2 and PR4, and deletion of PR4 alone from rat CD2 abrogated an ability to activate ITK. Thus, these results identify PR4 as an element of the CD2 tail that is required for activation of ITK. Furthermore, we show that, unlike wild-type rat CD2, PR4-deleted rat CD2 is unable to induce IL-2 secretion from Jurkat cells. This is consistent with the view that PR4-mediated activation of ITK is important for downstream signaling events induced by CD2 co-stimulation.

Animals↗
PubMed1997

Risk of mother-to-infant transmission of HIV-1 is not reduced in CCR5/delta32ccr5 heterozygotes.

To determine if the 32-bp deletion of the chemokine receptor CCR5 (delta32ccr5) protects against mother-to-infant transmission of HIV-1, specimens from all uninfected and infected children who were perinatally exposed to HIV-1 and observed since 1988 and whose mothers did not take zidovudine were assessed for delta32ccr5. The CCR5 genotype was determined using polymerase chain reaction (PCR) for 122 subjects, of whom 73 were HIV-1 infected and 49 were perinatally exposed but uninfected; 70% and 71%, respectively, were Caucasian. Eleven of 73 (15%) infected children and 4 of 49 (8%) exposed uninfected children were CCR5/delta32ccr5 heterozygotes (p = 0.40). Among subjects who had at least one Caucasian parent or grandparent, 11 of 51 (22%) HIV-1-infected persons and 4 of 35 (11%) uninfected persons were heterozygotes. None were homozygous for the delta32ccr5 allele. The estimated relative risk for mother-to-infant HIV-1 transmission in heterozygotes was 2.0. Furthermore, the 95% confidence interval (0.6, 7.3) suggested that it is unlikely that the true relative risk was <0.6. Thus, the infant CCR5/delta32ccr5 heterozygous genotype was not associated with a diminished risk of perinatally acquired HIV-1 infection.

Alleles↗
PubMed1997

Intercellular adhesion molecule-1 deficiency protects MRL/MpJ-Fas(lpr) mice from early lethality.

MRL/MpJ-Fas(lpr) (Fas(lpr)) mice develop a rapidly fatal form of systemic autoimmune disease characterized by glomerulonephritis and vasculitis similar to severe cases of systemic lupus erythematosus in humans. To evaluate the requirement for intercellular adhesion molecule-1 (ICAM-1) in the pathogenesis of tissue injury in this model, we created ICAM-1-deficient MRL/MpJ-Fas(lpr) (ICAM-1/Fas(lpr)) mice. ICAM-1 deficiency resulted in a striking improvement in the survival of Fas(lpr) mice (median +/- SEM survival of Fas(lpr) = 26 +/- 1.7 vs ICAM-1/Fas(lpr) = 47 +/- 2.4 wk, p < 0.0001) and the increased survival was associated with delayed elevations of blood urea nitrogen levels in the ICAM-1/Fas(lpr) mice. Histologic examination of the ICAM-1/Fas(lpr) mice revealed an overall reduction in glomerular disease and a significant reduction in vasculitis in the kidney, lung, skin, and salivary glands when compared with Fas(lpr). These findings indicate that ICAM-1 plays a major role in development of glomerular and vascular injury in Fas(lpr) mice.

Animals↗
PubMed1997

Effect of pregnancy and zidovudine therapy on viral load in HIV-1-infected women.

The objective of this study was to determine the effect of pregnancy and zidovudine (ZDV) on viral load in HIV-1 infected women. A prospective nonrandomized cohort study was conducted at a university medical center and affiliated clinic and included 44 HIV-1-seropositive pregnant women seen between June 1991 and September 1995. Twenty-three women initiated ZDV therapy during their pregnancy. Seventeen women did not take antiretrovirals, and four women took ZDV prior to and throughout pregnancy. HIV-1 viral load as determined by quantitative peripheral blood mononuclear cell (PBMC) culture and quantitative plasma RNA levels was measured at various times during pregnancy and in the postpartum period. HIV-1 load, by both infectivity and RNA levels, was relatively low and remained stable during pregnancy and through 6 weeks post partum. Initiation of ZDV therapy during pregnancy did not result in a significant decrease in viral load at delivery when controlling for the effect of pregnancy. In those women who received ZDV therapy only during pregnancy, there was a trend toward an increase in viral load measured by PBMC infectivity 6 months post partum compared with the levels before the initiation of ZDV. Mother-to-child transmission of HIV-1 occurred in one of 27 (4%) ZDV-treated women and in two of 16 (12.5%) untreated women. Among HIV-1-infected pregnant women with low viral levels, HIV-1 plasma RNA and infectivity remained stable during and after gestation. Although these results are based on a relatively small number of women and should be considered preliminary, the lack of significant ZDV-associated diminution in viral levels suggests that the protective effect of ZDV on the mother-to-child transmission of HIV-1 may not be due to the reduction in maternal viral levels but, by inference, may be due to the prevention of HIV-1 reverse transcription in the newborn.

Anti-HIV Agents↗
PubMed1997

Analysis of CD28 cytoplasmic tail tyrosine residues as regulators and substrates for the protein tyrosine kinases, EMT and LCK.

The CD28 cell surface receptor provides an important costimulatory signal for T cells necessary for their response to Ag. Early events in CD28 signaling include recruitment and activation of phosphatidylinositol 3-kinase (PI3-kinase) and activation of the protein tyrosine kinases (PTKs), LCK and EMT. Recruitment and activation of PI3-kinase is known to be dependent upon phosphorylation of tyrosine 173 of the CD28 cytoplasmic tail contained within a YMNM motif. By contrast, little is known of which residues of the CD28 tail, including tyrosines, are required for the activation of PTKs. To address this we studied the ability of truncation mutants and tyrosine to phenylalanine substitution mutants of the CD28 cytoplasmic tail to activate LCK and EMT in Jurkat T leukemia cells. Our results indicate that 1) activation of EMT is partially dependent upon tyrosine 173 of the CD28 tail, although it does not require PI3-kinase activation; 2) activation of LCK is independent of CD28 cytoplasmic tail tyrosine residues; and 3) elements sufficient for the activation of both kinases are contained within the first half of the tail. In addition we studied the CD28 tail as a substrate for both PTKs in in vitro kinase assays. We demonstrate that EMT can phosphorylate all four tyrosines of the CD28 tail, in contrast to LCK, which phosphorylates only tyrosine 173. Together with evidence that in vivo, tyrosines other than tyrosine 173 become phosphorylated following CD28 stimulation, this finding suggests that, like LCK, one function of EMT during CD28 signaling is phosphorylation of the receptor.

CD28 Antigens↗
PubMed1997

Effects of isradipine, an L-type calcium channel blocker on permanent and transient focal cerebral ischemia in spontaneously hypertensive rats.

Permanent or transient focal cerebral ischemia was induced in spontaneously hypertensive rats (SHR) using the intraluminal filament method. Successful occlusion of the middle cerebral artery (MCA) was achieved using 4/O filaments (terminal diameter 0.20-0.25 mm) coated with poly-L-lysine. The L-type calcium channel blocker isradipine (2.5 mg/kg) administered subcutaneously 30 min following permanent MCA occlusion significantly reduced the volume of ischemic brain damage in the cerebral hemisphere (25%; P = 0.0001), cerebral cortex (18%; P = 0.0034), and caudate nucleus (33%; P = 0.0002) when assessed at 24 h post-MCA occlusion. Isradipine did not affect the functional deficit (measured using a subjective neurological scoring system) induced by MCA occlusion. In SHR undergoing transient (2 h) MCA occlusion isradipine administered 30 min post-MCA occlusion produced a significant reduction (47%; P = 0.001) in hemispheric infarct volume, whereas isradipine administered at the onset of reperfusion did not confer any significant neuroprotection. No change in functional deficit was seen with isradipine with either dosing paradigm at 24 h post-MCA occlusion. These results demonstrate that the intraluminal filament method of MCA occlusion can be used in the SHR strain and also substantiates the neuroprotective efficacy of isradipine in SHR models of permanent and transient focal cerebral ischemia.

Animals↗