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Biomedical subjects

P D Griffiths

Publications and source records attributed to P D Griffiths.

At least 253 records · Page 14Linked to original sources
PubMed1988

Comparative sensitivity of three methods for the diagnosis of cytomegalovirus lung infection.

Conventional cell culture (CCC), an immunofluorescent assay for the rapid detection of CMV-induced early antigens (DEAFF) in infected fibroblasts and an immunofluorescent assay for the detection of CMV-infected cells obtained directly from clinical material were compared prospectively for their ability to detect CMV in bronchoalveolar lavage (BAL) material. CMV was detected by at least one method in 47 of 139 BALs (33.8%). The mean sensitivities of each of the three assays were 93% for DEAFF, 46% for CCC and 22% for the direct method. The mean time to diagnosis was 24 h, 15 days and 4 h for each of the methods, respectively. The use of monoclonal antibodies in the DEAFF test on BAL specimens provided a simple and rapid method for the diagnosis of CMV lung infection and was shown to be more reliable than conventional culture methods in achieving a diagnosis from BAL specimens.

Acquired Immunodeficiency Syndrome↗
PubMed1988

Sweat tests and flucloxacillin.

Sweat tests were carried out on 14 patients with cystic fibrosis and 14 controls when on no antibiotics and when taking oral flucloxacillin. No significant differences in results were obtained for either group whether on or off antibiotics. Oral flucloxacillin does not affect sweat electrolytes and is not a contraindication to sweat testing.

Adolescent↗
PubMed1988

Cytomegalovirus and beta 2 microglobulin in urine specimens. Reciprocal interference in their detection is responsible for artifactually high levels of urinary beta 2 microglobulin in infected transplant recipients.

We have previously shown that the binding of host beta 2 microglobulin (beta 2 m) by cytomegalovirus (CMV) in body fluids masks the viral antigenic determinants preventing its detection by CMV-specific monoclonal antibodies. We now report that the recognition of CMV-bound beta 2m by anti-beta 2m-specific antibodies in assays for beta 2m, results in erroneously high readings for beta 2m levels in urine specimens containing CMV. Urinary beta 2m levels have previously been reported to be elevated in patients with CMV infection--however, when virion bound beta 2m was removed by ultracentrifugation of urine specimens, the levels of free beta 2m were not found to be elevated in these patients. Since CMV is frequently excreted by transplant recipients and acquired immunodeficiency syndrome patients, our data suggest that measurements of urinary beta 2m levels in such patients are unreliable unless the urine specimens are confirmed to be free of CMV before analysis.

Animals↗
PubMed1988

Humoral immune responses within the lung of bone marrow transplant recipients studied by bronchoalveolar lavage.

We investigated 20 bone marrow transplant recipients with pneumonitis using bronchoalveolar lavage (BAL) to assess the humoral immune response in the lung. We measured the levels of total IgG, IgM and IgA in bronchoalveolar lavage fluid and serum, and albumin measurements were used to correct for simple diffusion of immunoglobulins from serum into the lung. We found evidence for the local production of immunoglobulins G, M and A in 15 patients. This was independent of the cause of the pneumonitis. We also found that, although seven patients who recovered from their pulmonary problem had evidence of considerable local production of immunoglobulin, eight patients who died were also producing immunoglobulins in the lung. Death due to pneumonitis in BMT recipients cannot, therefore, be ascribed to a failure of the local humoral immune response.

Adolescent↗
PubMed1988

Simplified quantification of urinary protein excretion in children.

The protein/creatinine ratio (Up/Uc) measured in 71 early morning urine samples (EMU) correlated closely with timed overnight urine (ONU) protein excretion rates (r = 0.96). The relationship was linear throughout the entire range of normal and abnormal protein excretion, an ONU rate of 1 mg/h/m2 body surface area being proportional to an EMU Up/Uc of 5 mg/mmol. Using the Coomassie Blue dye-binding method the upper limit of Up/Uc in 377 apparently healthy children and adolescents aged 3-19 years was shown to be 20 mg/mmol. Albumin/creatinine ratios (Ua/Uc) were also determined in the 377 healthy subjects, yielding a normal working upper limit of 3 mg/mmol. Although in normal individuals studied longitudinally the day-to-day variation of both Up/Uc and Ua/Uc was appreciable, all measurements remained within the cross-sectional normal range. While the determination of Ua/Uc has a role in the study of "microproteinuria", it is comparatively costly for routine use. The measurement of the EMU Up/Uc avoids errors and difficulties associated with timed urine collection, simplifies sample handling by the laboratory and is inexpensive. In clinical practice this is the method of choice for the quantification of proteinuria in patients with renal disease.

Adolescent↗
PubMed1987

Is cytomegalovirus interstitial pneumonitis in transplant recipients an immunopathological condition?

The conventional explanation for the high fatality rate due to cytomegalovirus (CMV) pneumonitis among allogeneic transplant recipients is that immunosuppression renders the host unable to control replication of this opportunistic agent. However, evidence from studies in man and the murine model of CMV show that virus replication in the lung is unrelated to the development of pathological effects, and that a host immune response is required for the induction of pneumonitis. Thus the hypothesis is that limited CMV replication in the lungs leads to display of a virus-coded protein, which is recognised by host T-cells, and that the pneumonitis is due to an uncontrolled accumulation and recruitment of such cells in the lungs. The reason why CMV is found in the lungs of patients with the acquired immunodeficiency syndrome (AIDS) without producing pneumonitis is probably because these patients cannot mount the pathogenic T-cell response. According to the hypothesis stated here, if the immune capabilities of AIDS patients can be restored, life-threatening CMV pneumonitis may develop.

Acquired Immunodeficiency Syndrome↗
PubMed1987

Immune donors can protect marrow-transplant recipients from severe cytomegalovirus infections.

To study the importance of transferred immunity against cytomegalovirus (CMV) in allogeneic, HLA-matched, T-cell-depleted bone-marrow transplantation, the incidence, severity, and outcome of CMV infections were studied in 40 CMV-seropositive recipients in relation to the donors' immunity against CMV. There was no significant difference in the incidence of CMV infections between recipients of seropositive (n = 27) and seronegative (n = 13) marrow. However, the incidence of CMV pneumonitis (8/13 compared with 4/27; p less than 0.001) and the mortality attributable to CMV infection (6/13 compared with 1/27, p less than 0.01) were significantly greater in the group with seronegative donors than in those with seropositive donors. Multivariate regression analysis showed that recipients of seronegative marrow had a fifteen-fold greater risk of CMV pneumonitis and a fifty-fold increase in risk of a fatal CMV infection than recipients of seropositive marrow. Thus, after T-cell depletion CMV-seropositive marrow protects seropositive recipients against severe CMV infections; whenever possible, therefore, such recipients should be given marrow from seropositive donors. Ultimately, active immunisation of CMV-seronegative donors might help to protect seropositive recipients of T-cell-depleted marrow transplants against severe CMV infections.

Adult↗
PubMed1987

Molecular biology and immunology of cytomegalovirus.

The application of modern biochemical techniques has led to a rapid improvement in our knowledge of the molecular biology of CMV. Several coding regions of the DNA genome have been identified with certainty and major virus-coded proteins have been given provisional names. The cascade expression of the CMV genome has been shown to be controlled by mechanisms similar to those found in other herpes viruses, together with novel post-transcriptional controls which remain to be defined. The control of CMV replication by the host involves both non-specific and specific defence mechanisms. The induction of natural killer cells and interferon early after CMV infection appears to be the most important aspects of the non-specific host defence against the virus. The cell-mediated immune response, in particular the generation of Tc cells against CMV early antigens, is probably the most important facet of the specific immune defence against CMV. When intact these defence mechanisms appear to be efficient in restricting viral replication; however, when such immunity is compromised, the balance rapidly swings in favour of the virus. As our understanding of the interaction between the host and the virus increases, it may be possible to redress the balance in such cases in favour of the host.

Animals↗
PubMed1987

Radioimmunoassay for the detection of IgG antibodies to herpes simplex virus and its use as a prognostic indicator of HSV excretion in transplant recipients.

We report the development of a solid-phase radioimmunoassay for the detection of IgG antibodies to herpes simplex virus (HSV), using a mouse monoclonal antibody specific for the Fc portion of human IgG as the radiolabelled detecting antibody. The binding ratio to test antigen at a single serum dilution, 1:100, correlated significantly with the endpoint titre by radioimmunoassay and with neutralising antibody titre. When compared to neutralisation the radioimmunoassay had a sensitivity of 100% and a specificity of 93%. We believe that the anomalous results are not false positives but represent an increased sensitivity of the radioimmunoassay. We found, by either radioimmunoassay or neutralisation, that high levels of antibody prior to transplantation were associated with a significantly increased risk of HSV excretion post-transplantation in both renal and bone marrow transplant recipients. Thus the radioimmunoassay is a sensitive, specific, and rapid test that can be used as a prognostic indicator of HSV excretion in transplant recipients.

Antibodies, Viral↗
PubMed1987

Use of monoclonal antibodies for the diagnosis of cytomegalovirus infection by the detection of early antigen fluorescent foci (DEAFF) in cell culture.

A pool of seven monoclonal antibodies, each reactive with cytomegalovirus (CMV) early antigens, was used in an indirect immunofluorescence method for the rapid detection of CMV-infected fibroblasts following inoculation with clinical specimens. A total of 1,639 specimens were examined, and the results were compared with those of conventional isolation procedures. The detection of CMV by early antigen fluorescent foci (DEAFF) was found to be comparable, both in terms of specificity and sensitivity, to that of conventional cell culture. Its great advantage, however, is the rapidity with which results are achieved. Thus, results were available from the DEAFF test within 24 hours of receipt of the specimens as compared to a mean of 16 days for cell culture. This single rapid assay for the detection of CMV in clinical samples may be performed by any laboratory familiar with cell culture techniques and in our hands is the preferred diagnostic method for CMV.

Antibodies, Monoclonal↗
PubMed1987

Regeneration of humoral immunity to herpes simplex virus following T-cell-depleted allogeneic bone marrow transplantation.

We investigated the adoptive transfer of immunity to herpes simplex virus (HSV) in 61 recipients of T-cell-depleted marrow allografts. Up to 3 months after bone marrow transplantation (BMT), high titres of HSV antibody (Ab) are passively acquired from blood products. This antibody has a half-life of 40.3 days, so that determination of recipient immunity to HSV cannot be made during the early post-transplant period. By 4 months after the allograft, seronegative recipients of seronegative donors had near-background levels of HSV Ab. Seropositive recipients, whether they were excreting virus or not, had high titres of HSV Ab regardless of donor status. Surprisingly, the seronegative recipients of HSV seropositive donors remained seronegative, a result at variance with observations made in the recipients allografts from which T-cells had not been removed. Thus, T-cell depletion modifies the adoptive transfer of humoral immunity to HSV, and an immune donor alone no longer ensures continued antibody production in the recipient.

Antibodies, Viral↗
PubMed1987

Cytomegalovirus strain AD169 binds beta 2 microglobulin in vitro after release from cells.

We previously reported that a host protein, beta 2 microglobulin (beta 2m) inhibited the detection of human cytomegalovirus (CMV) in urine specimens by enzyme immunoassay and postulated that beta 2m bound to the virus particle and masked the viral antigenic determinants. We report here that CMV strain AD169 grown in cell culture bound human beta 2m when this protein was added to cell culture fluids or when the virus was added to urine. Such binding was not seen with herpes simplex virus. CMV could also bind bovine beta 2m from foetal calf serum in cell culture fluids. The use of radiolabelled beta 2m in other experiments showed that CMV bound beta 2m after release from cells and that the bound beta 2m did not represent acquisition of class I HLA molecules during budding from host cell membranes. Immunoprecipitation studies showed that beta 2m was bound by two viral envelope proteins beta 2m BP1 (beta 2m-binding protein 1) and beta 2m BP2 of molecular masses 36,000 and 65,000 daltons respectively. beta 2m could not bind to separated viral proteins under reducing or non-reducing conditions. We propose that interaction of these two proteins on the viral surface is required to enable CMV to bind beta 2m.

Cells, Cultured↗
PubMed1987

Cytomegalovirus in urine specimens has host beta 2 microglobulin bound to the viral envelope: a mechanism of evading the host immune response?

We have previously reported that human cytomegalovirus (CMV) from urine specimens cannot be captured onto a solid phase by CMV-specific monoclonal antibodies that can capture CMV grown in vitro. We report here that CMV exists in vivo in body fluids such as urine as beta 2 microglobulin (beta 2 m)-coated particles. We have demonstrated the presence of beta 2m on CMV purified directly from urine by Western blotting and have shown that the beta 2m was associated with the viral envelope. Urinary CMV could be specifically bound by an affinity column comprising a monoclonal antibody specific for beta 2m bound to Sepharose. The beta 2m-coated urinary CMV could not be neutralized by hyperimmune globulin, human immune sera or murine monoclonal antibodies that could neutralize CMV grown in cell culture. We conclude that the binding of beta 2m by CMV masks the important antigenic sites necessary for neutralization which are recognized by man's immune response. We propose that CMV has evolved this mechanism of coating itself in a host protein as a mechanism of evading the host immune response and facilitating transmission between individuals.

Antibodies, Monoclonal↗
PubMed1987

Beta 2 microglobulin enhances the infectivity of cytomegalovirus and when bound to the virus enables class I HLA molecules to be used as a virus receptor.

We have previously demonstrated that human cytomegalovirus (CMV) binds the host protein beta 2 microglobulin (beta 2m) from body fluids or from cell culture media. In this report we have examined the effect of the beta 2m on viral infectivity. We have shown that the addition of human purified beta 2m, or a fraction of foetal calf serum corresponding to bovine beta 2m, to culture medium increased the amount of infectious extracellular CMV, compared to that from cells grown in serum-free medium. Metabolic labelling experiments demonstrated that this effect was not due to an increase in the amount of extracellular virus but to an increase in the infectivity of the virus present in extracellular fluids. We concluded that the binding of beta 2m by CMV increased its infectivity. We have shown that CMV and beta 2m compete for binding sites on fibroblasts. As the main binding site on cells for beta 2m is the class I HLA heavy chain we compared the binding of CMV to the Raji and Daudi cell lines which express or lack expression of class I HLA molecules. The binding of radiolabelled beta 2m-coated CMV was significantly higher to Raji cells than to Daudi cells. Furthermore, CMV could compete with beta 2m for binding to Raji cells, although the reverse was not true. These results demonstrate that CMV can use class I HLA molecules as a virus receptor. We propose that when coated with beta 2m, CMV has the capacity to displace beta 2m from the class I HLA heavy chain-beta 2m dimer on the cell surface and bind to cells. The fact that beta 2m enhances infectivity suggests that such binding leads to productive infection of cells.

Cell Line↗