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Biomedical subjects

S A Prasad

Publications and source records attributed to S A Prasad.

15 recordsLinked to original sources
PubMed2001

Heterosubtypic immunity to influenza A virus in mice lacking IgA, all Ig, NKT cells, or gamma delta T cells.

The mechanisms of broad cross-protection to influenza viruses of different subtypes, termed heterosubtypic immunity, remain incompletely understood. We used knockout mouse strains to examine the potential for heterosubtypic immunity in mice lacking IgA, all Ig and B cells, NKT cells (CD1 knockout mice), or gamma(delta) T cells. Mice were immunized with live influenza A virus and compared with controls immunized with unrelated influenza B virus. IgA(-/-) mice survived full respiratory tract challenge with heterosubtypic virus that was lethal to controls. IgA(-/-) mice also cleared virus from the nasopharynx and lungs following heterosubtypic challenge limited to the upper respiratory tract, where IgA has been shown to play an important role. Ig(-/-) mice controlled the replication of heterosubtypic challenge virus in the lungs. Acute depletion of CD4+ or CD8+ T cell subsets abrogated this clearance of virus, thus indicating that both CD4+ and CD8+ T cells are required for protection in the absence of Ig. These results in Ig(-/-) mice indicate that CD4+ T cells can function by mechanisms other than providing help to B cells for the generation of Abs. Like wild-type mice, CD1(-/-) mice and gamma(delta) (-/-) mice survived lethal heterosubtypic challenge. Acute depletion of CD4+ and CD8+ cells abrogated heterosubtypic protection in gamma(delta) (-/-) mice, but not B6 controls, suggesting a contribution of gamma(delta) T cells. Our results demonstrate that the Ab and cellular subsets deficient in these knockout mice are not required for heterosubtypic protection, but each may play a role in a multifaceted response that as a whole is more effective than any of its parts.

Administration, Intranasal↗
PubMed2001

Cutting edge: recombinant adenoviruses induce CD8 T cell responses to an inserted protein whose expression is limited to nonimmune cells.

CD8 T cells (T(CD8+)) play a crucial role in immunity to viruses. Current understanding of activation of naive T cells entails Ag presentation by professional APCs (pAPCs). What happens, however, when viruses evolve to avoid infecting pAPCs? We have studied the consequences of this strategy by generating recombinant adenoviruses that express influenza A virus nucleoprotein under the control of tissue-specific promoters. We show that the immunogenicity of such viruses requires their delivery to organs capable of expressing nucleoprotein. This indicates that infection of pAPCs is not required for adenoviruses to elicit a T(CD8+) response, probably due to a cross-priming via pAPCs. While this bodes well for recombinant adenoviruses as vaccines, it dims their prospects as gene therapy vectors.

Adenoviridae↗
PubMed2001

Effect of intravenous antibiotics on exercise tolerance (3-min step test)) in cystic fibrosis.

Most children with cystic fibrosis (CF) feel better and display more energy after a course of intravenous antibiotics (IVABs), but this is not always reflected by a satisfactory improvement in lung function. We assessed the change in exercise tolerance after treatment with IVABs using the 3-min step test, and compared it with changes in spirometric lung function and arterial oxygen saturation (SaO(2)). Thirty-six children (mean age, 13.8 years) were enrolled from two tertiary CF centers during an inpatient stay for IVABs. After 10-14 days of treatment, there was a significant improvement in median FEV(1) from 43% to 57% of predicted values (P < 0.0001), and median FVC from 66% to 73% of predicted values (P < 0.0001), while median SaO(2) significantly increased from 95% to 96.5% (P < 0.05). This was accompanied by a reduction in resting heart rate (median 118 bpm to 109 bpm, P < 0.005) and subjective breathlessness at rest (median visual analogue score 2.2 to 0.8, P < 0.005). All outcomes of exercise tolerance were improved after IVABs. There was a reduction in maximum heart rate (median 156 bpm to 150 bpm, P < 0.05) and an increase in minimum SaO(2) (median 93.5% to 94.5%, P = 0.08) measured during the step test. There was also a reduction in subjective breathlessness (median visual analogue score of 5.5 to 4.2, P < 0.005) and objective breathlessness (median 15-count score of 3 to 2, P < 0.0001) measured immediately after the step test. Exercise testing was a useful outcome measure for monitoring effectiveness of inpatient therapy, and complemented spirometry and SaO(2) monitoring. The simple ward-based 3-min step test was found to be a particularly suitable method for measuring changes in exercise tolerance in children with CF.

Adolescent↗
PubMed2001

Exercise tolerance in children with cystic fibrosis undergoing lung transplantation assessment.

The aim of this study was to compare the 6-min walk test against the recently developed 3-min step test, as measures of exercise tolerance in children with moderate to severe cystic fibrosis (CF) lung disease referred for lung transplantation assessment. Twenty-eight children with CF (16 girls, 12 boys), with a mean age of 13.7 yrs (range 7.2-17.8 yrs) and mean forced expiratory volume in one second of 34% predicted (range 17%-67%) were recruited. All subjects performed both the 6-min walk and 3-min step-tests. Outcome measures were maximum rise in heart rate (HR), and maximum fall in arterial oxygen saturation (Sa,O2). There was no significant difference in resting HR or Sa,O2 prior to starting the two tests. Both step and walk tests produced significant rises in median HR (from 114-149 min(-1), p<0.0005, and 119-138 min(-1), p<0.0005, respectively) and significant falls in Sa,O2 (both from 94-92%, p<0.0005). The step test produced a significantly greater percentage rise in HR (30% versus 18%, p<0.0005) and a significantly greater percentage fall in Sa,O2 (4% versus 2%, p=0.002). Bland-Altman analysis gave wide 95% limits of agreement (10.7-29.3% for rise in HR, -2.14.6% for fall in Sa,O2). The step test was well tolerated. The 3-min step test produced a greater fall in Sa,O2 and a greater rise in HR than the 6-min walk test in children with moderate to severe CF lung disease. It may be of value when assessing a child's suitability for lung transplantation.

Adolescent↗
PubMed2000

Fifteen-count breathlessness score: an objective measure for children.

Breathlessness is an important symptom of respiratory disease and its quantification is useful, especially during exercise testing. However, measures of perceived breathlessness are not readily understood by children and are somewhat subjective. We studied an objective score: the 15-count breathlessness score, in which subjects take a deep breath and then count out loud to 15; the number of breaths taken to complete the count is the score. Fifty-four children with cystic fibrosis (CF) performed a standard 6-min walk and 3-min step test (30 steps/min for 3 min). The 15-count score was compared with the modified Borg scale after exercise. A further 45 children with CF and 33 healthy schoolchildren underwent an incremental step test (20, 30, and then 40 steps/min for 2 min each), using the 15-count score, then the Borg scale, and then a standard visual analogue score between increments. The 15-count score was significantly increased after both the walk and the step test (P < 0.0001), although the step test made children significantly more breathless than the walk test (P < 0.0001). At baseline, there were no differences in any of the breathlessness scores between the CF and normal children. After the full 6 min of the incremental step test, CF children were significantly more breathless than the normal children, as measured by 15-count (P < 0.0001), Borg (P < 0.0005), and visual analogue scores (P < 0.0005). All scores increased significantly as exercise intensity increased over time, but the slope estimates were significantly greater for CF patients than for normal children (P < 0.0005). The 15-count score has been evaluated as an objective measure of breathlessness. It is easy to explain and perform, and can be used by any child capable of counting fluently to 15 in any language. It is best used in conjunction with a subjective score, and either the Borg scale or a visual analogue score is appropriate.

Adolescent↗
PubMed1998

A step in the right direction: assessing exercise tolerance in cystic fibrosis.

Exercise tolerance may be reduced in patients with cystic fibrosis, but it is not always possible to predict this from standard lung function measurements. Formal exercise testing may, therefore, be necessary, and the test should be simple and readily available. We have developed a "3-minute step test" and compared it with the standard 6-minute walking test. Subjects stepped up and down a 15-cm-high single step at a rate of 30 steps per minute for 3 minutes. The effect of the step test on spirometry was tested first in 31 children with CF (mean age, 12.0 years), who had a mean (range) baseline forced expired volume in 1 second (FEV1) of 64% (18-94%) of predicted values. The step test was then compared with the standard 6-minute walk in a further 54 patients with cystic fibrosis (mean age, 12.5 years), with mean (range) baseline FEV1 of 61% (14-103%) of predicted values. Outcome measures were minimum arterial oxygen saturation (SaO2), maximum pulse rate, and the modified Borg dyspnea score. Post-step test spirometry showed mean (95% CI) changes of -1.1% (-6.0 + 3.9%) for forced vital capacity, of -1.6% (-4.2 + 1.1%) for FEV1, and +0.25% (-2.8 + 3.3%) for peak expiratory flow, although 5/31 children showed >15% drop in one or more parameters. The step and walk tests both produced significant changes (P < 0.0001) in all outcomes, with a mean (range) minimum SaO2 of 92% (75-98%) versus 92% (75-97%), a maximum pulse rate of 145 b.p.m. (116-189) versus 132 (100-161), and a Borg score of 2.5 (0-9) versus 1.0 (0-5), respectively. Comparison of the two tests showed that the step test increased breathlessness (mean change Borg score, 2.3 vs. 0.8; P < 0.0001) and pulse rate (mean change, 38% vs. 24%, P < 0.0001) significantly more than the walk, whereas the decrease in SaO2 was similar (mean change, -2.9% vs. -2.6%; P = 0.12). Some patients with a significant drop in SaO2 (>4%) would not have the decrease predicted from their baseline lung function. Reproducibility for the two tests was similar. The step test is quick, simple and portable, and is not dependent on patient motivation. Although the step test is more tiring, its effect on SaO2 is similar to the 6-minute walking test. It is a safe test that may prove to be a valuable measure of exercise tolerance in children with pulmonary disease, although longitudinal studies are now needed.

Adolescent↗
PubMed1998

Calnexin expression does not enhance the generation of MHC class I-peptide complexes.

We investigated the requirement for calnexin in the biogenesis of MHC class I molecules. Mutant human cells lacking calnexin were infected with recombinant vaccinia viruses encoding mouse MHC class I molecules, Kd, Kb, Kk, Dd, Db, and Ld. Flow cytometry indicated that each of the six MHC class I allomorphs was transported to the cell surface at similar rates in calnexin-deficient cells and transfectants expressing calnexin. For Kb and Kd, the calnexin-independent biogenesis occurred regardless of whether the MHC class I molecules contained human or mouse beta 2-microglobulin. Also addressed was the effect of calnexin on the surface expression of Kb molecules bearing the immunodominant peptide from ovalbumin (OVA257-264). This was detected with a recently described monoclonal antibody specific for the Kb/peptide complex. Calnexin expression had no significant effect on the formation of Kb/peptide complexes generated from full-length OVA, cytosolic OVA257-264, or endoplasmic reticulum-targeted OVA257-264, which was expressed in the presence of the herpes simplex virus ICP47 protein to ensure detection of TAP-independent peptide-MHC class I complexes. Complementary results were obtained with TAP-independent formation of Kd/ peptide complexes. These findings indicate that calnexin is not required for the efficient assembly of MHC class I molecules with TAP-dependent or independent peptides.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

Differential APC requirements of self- and nonself-reactive T cells and T cell hybridomas specific for retinal S-antigen.

Autoreactive T cell hybridomas specific for pathogenic peptides of retinal S-Ag require a novel radiosensitive APC activity for IL-2 secretion that is distinct from Ag presentation by MHC class II. Antigen-dependent IL-2 secretion by self-reactive hybridomas was much more efficient with splenic APC than with thymic APC, although both provided similar levels of hybridoma TCR occupancy as measured by activation-induced cell death. Furthermore, thymic APC did stimulate IL-2 secretion by a non-self reactive hybridoma. To test the hypothesis that this activity was provided by a distinct cell population, fractionated splenocytes were tested for their ability to present Ag to these hybridomas. The most potent Ag presentation for IL-2 secretion was found to segregate with low-density, B cell-enriched fractions while adherent cells, or purified T cells were unable to support IL-2 production. Together with previous results, the data show that antigen presentation leading to IL-2 secretion by these autoreactive T cell hybridomas requires activated B cells, whereas TCR occupancy can be provided by several APC subsets.

Amino Acid Sequence↗
PubMed1997

Antigen presentation in uveitis.

Experimental autoimmune uveoretinits (EAU) is not only a valuable model for human inflammatory eye diseases, it is also a useful system for studying many aspects of immunobiology. One such aspect is self/non-self discrimination, the ability of the immune system to tolerate self molecules while responding aggressively to foreign antigens. Our laboratory has used EAU to investigate the mechanisms of T cell tolerance to retinal S-antigen (S-Ag). Several mechanisms have been proposed to maintain T cell tolerance to autoantigens, including clonal deletion and clonal anergy. As immunisation with S-Ag or pathogenic peptides activates uveitogenic T cells, tolerance to this autoantigen cannot be due to clonal deletion. Nevertheless, tolerance acts to keep these existing autoreactive T cells in a naive, or innocuous state. Here we suggest a novel mechanism--low-affinity occupancy of the autoantigen-specific T cell receptor (TCR) by self-antigen--that may act in concert with the well-known mechanisms to maintain tolerance to S-Ag in the LEW rat. This model differs from clonal anergy in that the missing antigen-presenting cell (APC) activity is not a co-stimulatory function but a TCR co-ligand that increases the avidity of the interaction between the TCR and its peptide-major histocompatibility complex (MHC) ligand. In the absence of this co-ligand only partial signals are generated through the TCR, leading to incomplete T cell activation. This model was deduced from experiments with T cell lines and hybridomas specific for S-Ag, which showed that: (1) autoreactive T cells required a novel APC function, (2) this novel function was necessary to provide complete TCR engagement, and (3) activation of autoreactive T cells was restricted to specific APC.

Animals↗
PubMed1995

A radiosensitive APC activity dissociates IL-2 secretion and activation-induced cell death by autoreactive T cell hybridomas.

T cell hybridomas were generated from a LEW rat T cell line specific for the uveitogenic peptide bov-B1 of bovine retinal S-antigen. Using these autoreactive hybridomas, IL-2 production and activation-induced cell death (AICD) were dissociated as outcomes of activation. The self-reactive hybridomas secrete IL-2 and undergo AICD in response to antigen presented by non-irradiated syngeneic splenocytes, whereas antigen presentation by irradiated splenocytes induced only AICD. IL-2 production by a non-self reactive hybridoma was unaffected by irradiation of the APC. Pretreatment of the APC with phorbol ester or lipopolysaccharide and IL-4 protected their ability to induce IL-2 secretion after gamma-irradiation. Although the co-stimulation-blocking reagent CTLA-4-Ig mimicked the effect of gamma-irradiation by preventing IL-2 secretion but not AICD, B7 expression on the APC was not radiosensitive, nor did co-stimulation, provided 'in trans' with a B7-expressing third-party cell, reconstitute antigen-specific hybridoma IL-2 secretion in response to irradiated APC. In summary, the data show that IL-2 secretion and AICD of a self-reactive T cell hybridoma can be dissociated as consequences of TCR occupancy in the presence of a functional co-stimulatory signal. It is proposed that the signals producing these events are transduced through the TCR-CD3 complex alone and reflect the differential outcomes of high- and low-affinity interactions.

Amino Acid Sequence↗
PubMed1994

Corneal endothelial cells inhibit T cell proliferation by blocking IL-2 production.

We have reported that Ia+ LEW rat corneal endothelial (CE) cells could not induce Ag-specific proliferation of MHC compatible T cell lines. Instead, the presence of CE cell monolayers inhibited T cell proliferation assays. We now report on the effect of CE cells on IL-2 production, IL-2R expression, and IL-2 responsiveness of T cells. Ag-specific growth inhibition of the 1C3.4 T cell hybridoma was unaffected by the presence of CE cells but IL-2 production was suppressed by CE cells. Similar CE cell-mediated inhibition of IL-2 secretion was found with fresh splenic lymphocytes stimulated by Con A. Controls showed that neither CE cells nor their supernatant affected the CTLL cell response to rIL-2, allowing reliable measurement of IL-2 production. Further evidence that the block is at the level of IL-2 production was shown by the observation that IL-2R expression was up-regulated after TCR occupancy whether or not CE cells were present. Also, T cells expressing IL-2R were responsive to exogenous rIL-2 whether or not CE cells were present. CE cells themselves do not induce IL-2R expression or IL-2 responsiveness. Under normal culture conditions, the R1170 rat T cell line continually expresses a significant level of IL-2R and dies before IL-2R is down-regulated to background levels. However, in the presence of CE cells, these R1170 cells down-regulate IL-2R and lose IL-2 responsiveness while remaining viable.

Animals↗
PubMedDate not supplied

Finding evidence to support airway clearance techniques in cystic fibrosis.

Chest physiotherapy, aiming to clear bronchopulmonary secretions, has become a mainstay in the respiratory management of cystic fibrosis (CF). Early diagnosis and new therapeutic interventions have dramatically improved the outlook for patients with this disorder and it is no longer a disease of childhood. Along with these changes chest physiotherapy has also progressed, with the development of several treatment modalities that are more effective and can be performed by the patient without assistance. This allows older children and adults with CF to lead more normal and independent lifestyles. Despite this progress questions remain regarding the efficacy and consequences of airway clearance techniques, the scientific evidence available to support the selection of the most appropriate treatment modality and, not least, the problems associated with the treatment-related burden that is placed on patients and their families.

Adolescent↗