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Biomedical subjects

T H The

Publications and source records attributed to T H The.

At least 127 records · Page 7Linked to original sources
PubMed1990

Symptomatic cytomegalovirus (CMV) infections identified by image cytometry and other parameters for CMV infection.

Thirty-eight renal transplant recipients were followed during the first 3 months after transplantation. Once weekly, cultures of urine and buffy coat for cytomegalovirus (CMV) were taken and an immunocytochemical assay for immediate early antigens of CMV (IEA assay) was performed. Thirty patients had evidence of a CMV infection and 11 had a symptomatic CMV infection. All symptomatic patients had one or more positive urine cultures or a positive IEA assay. However, 15 patients with positive urine cultures and 12 patients with a positive IEA assay lacked any signs of symptomatic CMV disease. Moreover, 6 out of 15 patients with positive buffy coat cultures for CMV did not have symptomatic CMV disease. Using a computerized system to quantify IEA-positive granulocytes, we show that the absolute number of positive cells per million correlates very well with the occurrence of symptomatic CMV disease.

Antibodies, Viral↗
PubMed1990

Autoantibodies against myeloid lysosomal enzymes in crescentic glomerulonephritis.

To investigate the possible association of crescentic glomerulonephritis (CGN) with autoantibodies to myeloid lysosomal enzymes, we tested sera from 35 consecutive patients with CGN without diagnostic immunohistological findings in their renal biopsy for the presence of antineutrophil cytoplasmic antibodies directed against a 29 kD antigen from azurophilic granules (29 kD-ANCA), against myeloperoxidase (MPO-ANCA) and against elastase (elastase-ANCA), using antigen-catching ELISAs with well-defined monoclonal antibodies. 29 kD-ANCA were present in the sera of all nine patients with CGN as part of biopsy-proven Wegner's granulomatosis (WG), of ten patients with CGN and clinically suspected WG, and of two patients with idiopathic CGN. Sera from the remaining patients with clinically suspected WG (N = 5) or idiopathic CGN (N = 6) were negative for 29 kD-ANCA, but invariably positive for MPO-ANCA. Neither of these antibodies could be detected in sera from patients with CGN of infectious origin (N = 3), different forms of CGN (N = 7), other renal lesions (N = 34), or normal controls (N = 52). None of the sera tested were positive for elastase-ANCA. Our results indicate that both vasculitis-associated CGN and idiopathic CGN are associated with autoantibodies against myeloid lysosomal enzymes. This finding places these disorders within one spectrum of diseases.

Autoantibodies↗
PubMed1990

Quantitative measurement of cytomegalovirus-specific IgG and IgM antibodies in relation to cytomegalovirus antigenaemia and disease activity in kidney recipients with an active cytomegalovirus infection.

In a longitudinal investigation 103 kidney recipients were studied with respect to the development of cytomegalovirus (CMV) specific antibodies of the IgG and IgM class, in relation to the detection of CMV antigenaemia (immediate early antigen, IEA), in weekly obtained blood samples during the first 3 months after transplantation. In 15 out of 49 (31%) seronegative patients a primary infection occurred, which was characterized by a quick rise in IgM antibody followed by a slower production of IgG antibody, high maximum numbers of IEA+ cells, and a CMV syndrome in 11 patients. In 35 out of 54 (65%) seropositive patients a secondary infection occurred. After a post-operative fall in the IgG antibody, which was also found in patients without an active infection and which was accompanied by a similar drop in serum albumin and IgG, a second dip in IgG antibody was found 6 days before the first IEA+ leucocyte appeared in the blood. This was followed by a significant increase, indicative of an active immune response in consequence of the infection, 18 days later. In 31 of these 35 patients an IgM response was found. This could be ascribed to the presence of rheumatoid factor activity in 20 of them. Eight patients who showed a transient rise in IgG antibody between the two dips could be distinguished from the remaining ones by a lower maximum number of IEA+ cells and less severe disease symptoms. The described results suggest that (i) an adequate humoral immune system may prevent symptomatic CMV disease in secondary infections; and (ii) CMV-specific antibodies may be removed from the circulation by antigens present in infected tissues before CMV antigenaemia becomes detectable.

Antibodies, Viral↗
PubMed1990

Early detection of active cytomegalovirus (CMV) infection after heart and kidney transplantation by testing for immediate early antigenemia and influence of cellular immunity on the occurrence of CMV infection.

To determine the incidence of active cytomegalovirus (CMV) infection after organ transplantation and its relationship with the immune system, 55 renal and 14 cardiac transplant recipients were closely monitored for active CMV infection (expression of CMV immediate early antigen in granulocytes--antigenemia--and positive cultures) and immune parameters. All 19 CMV-seronegative recipients with seronegative donors remained seronegative, showing that no CMV transmission occurred by leukocyte-depleted blood products. Primary CMV infection occurred in 4 of 11 (36%) patients with positive donors and was symptomatic in 1 (9%) patient. Active CMV infection was found in 29 of 39 (74%) seropositive patients and was symptomatic in 3 (8%) patients. CMV antigenemia was always the first indication of active CMV infection (antigenemia, on average, at day 45 +/- 15; immunoglobulin G rise at day 71 +/- 36; and positive cultures at day 70 +/- 17). Cellular immunity, as measured by lymphocyte proliferation (LPT), proved to be of importance in the prevention of active CMV infection, as 14 of 15 patients with negative LPT obtained active CMV infections with antigenemia and positive cultures, whereas 1 of 10 patients with positive LPT did so (P less than 0.0001).

Antibodies, Monoclonal↗
PubMed1989

LAMA tumor in the rat as an experimental model for pre-B-cell leukemia.

A late pre-B-cell leukemia model in the rat, the LAMA tumor, is described. A mouse monoclonal antibody (HIS30) was developed against LAMA cells. HIS30 reacts with a membrane antigen in tumor tissue, whereas its reactivity with normal tissues is limited to the zona glomerulosa of the adrenal cortex and to the adrenal medulla. HIS30 was used for both the immunohistological detection of tumor cells in tissue sections and the immunolocalization of tumor cells in vivo. To enable in vitro studies with the LAMA model, an in vitro growing cell line (LAMA-K1) was established from the LAMA tumor. LAMA-K1 is immunophenotypically similar to the original tumor. Two tumor transplantation models were characterized. In the first model LAMA was implanted s.c., and local tumor growth occurred at the injection site, which was then followed by lymphatogenic and subsequently hematogenic tumor spread. In the second model i.v. transplantation caused direct hematogenic tumor dissemination. In both models early dissemination was especially prominent to the bone marrow, spleen, and liver. Later in the disease most visceral organs became involved, and partial paralysis of the animal was observed in the end stage of the disease. In combination with HIS30, the LAMA pre-B-cell tumor offers a model for both the investigation of in vivo transplanted tumor cells and for the in vivo detection of tumor cells by HIS30 in LAMA tumor-bearing rats.

Animals↗
PubMed1989

Determination of human antibodies to specific antigens of cytomegalovirus using an antigen capture immunoassay.

The antigen capture immunoassay which is described herein is based on the binding of specific antigens of cytomegalovirus (CMV) by monoclonal antibodies bound to a solid phase. The specificity of the binding was demonstrated by the analysis of antigens labelled with [35S]methionine and captured by the bound monoclonal antibodies. These specific antigens are recognized in turn by specific anti-cytomegalovirus antibodies in human sera. The immunoassay permits quantitation of these specific anti-cytomegalovirus antibodies and should facilitate both qualitative and quantitative comparisons of the antibodies against specific CMV antigens in different individuals.

Antibodies, Monoclonal↗
PubMed1989

Detection and isolation of antigen-specific B cells by the fluorescence activated cell sorter (FACS).

A method is described for the isolation of antigen-specific B cells from immunized and subsequently boosted mice. Antigen-specific B cells were stained by incubation with fluorescein isothiocyanate (FITC)-labelled antigen and then detected and isolated in a fluorescence activated cell sorter (FACS). Ovalbumin (OVA) and Helix pomatia haemocyanin (HPH) were used as antigens in this procedure, yielding relative amounts of antigen-FITC-binding lymphocytes of 0.9 +/- 0.4% and 3.5 +/- 3.1%. The FITC-positive cells were visible as distinct cell populations in the FACS-generated histograms. All antigen-FITC-binding cells were B cells, as shown by double staining with phycoerythrin-conjugated anti-mouse Ig In addition, as tested in a spot-ELISA, the sorted, antigen-FITC-binding cell population contained almost the entire population of antigen-specific antibody-producing B cells. However, sorting had a negative influence on the antibody production capability of the sorted cells. Through washing of isolated spleen cells in the procedure before labelling with antigen-FITC proved to be essential for the specific detection of antigen-specific B cells, since staining without prior washing resulted in antigen-FITC binding to all B cells. This 'nonspecific' staining phenomenon was caused by the presence of antibodies, specific for the immunizing/boosting antigen, which were also present in the spleen cell suspension. These antibodies formed immune complexes with antigen-FITC and bound to Fc receptors present on all B cells, interfering in this way with any specific binding of antigen-FITC to sIg on the B cells.

Animals↗
PubMed1989

Association between active Wegener's granulomatosis and anticytoplasmic antibodies.

Autoantibodies reacting with the cytoplasm of granulocytes and monocytes (anticytoplasmic antibodies [ACPAs]) were found in 42 of 45 patients with active Wegener's granulomatosis (WG) (sensitivity, 93%). Specificity was tested in selected groups of patients with closely related diseases. Of 58 patients without a diagnosis of WG, 2 had ACPAs (specificity, 97%). The significance of ACPA titration for assessing or predicting disease activity was evaluated in a 16-month prospective study of 35 patients with WG. Seventeen relapses were observed and all were preceded by a significant rise of the ACPA titer. Anticytoplasmic antibodies are a specific and sensitive marker for active WG; a rising titer is a sensitive marker for the development of a relapse.

Adult↗
PubMed1989

INCA: software for consort 30 analysis of flow cytometric calcium determinations.

The INCA program converts Consort 30-generated fluorescence list mode data collected from Indo-1-stained cells to absolute intracellular calcium concentrations (nM Ca2+i). The calcium data are plotted vs. time, allowing the user to analyze the fractions of cells responding to a given stimulus. Converted files can be restored to disk after replacing FL1 and FL2 with time and calcium, respectively, for future analysis.

B-Lymphocytes↗
PubMed1989

An improved immunocytochemical method for the detection of human cytomegalovirus antigens in peripheral blood leucocytes.

A recently described immunoperoxidase method for the detection of nuclear human cytomegalovirus (HCMV) immediate early antigen (IEA) directly on peripheral blood leucocytes suffers from the drawback that the antigen is vulnerable to endogenous peroxidase inactivation procedures. To solve this problem a procedure is developed in which endogenous peroxidase is inactivated after binding and immobilization of the primary antibody with 4% formaldehyde. In combination with this procedure, three types of inactivation were investigated: glucose/glucose oxidase, hydrochloric acid and methanol/H2O2. Of these three, the first gives optimal results, especially in combination with methanol/acetic acid (20/1 v/v) as the primary fixative. This procedure results in preparations which allow for a more objective evaluation and enable automated examination using bright field microscopy. As a second improvement we developed a simple adherence method in order to diminish the risk of infection for the laboratory staff during processing of unknown blood samples. The protocol described shows great clinical potential for the diagnosis of HCMV infections.

Antigens, Viral↗
PubMed1989

Cytomegalovirus infection after organ transplantation: an update with special emphasis on renal transplantation.

Cytomegalovirus infections are still the most important infectious complications after organ transplantation. Besides historical notes this review will deal with new aspects concerning the epidemiology of the CMV, diagnostic modalities of CMV infection, the delicate counterbalance between the immune system and the CMV, as well as the symptomatology of this infection. Furthermore, aspects like prophylaxis and new, promising therapeutic regimes for treatment of infection will be dealt with. Although this update is applicable for all types of solid organ transplantation, emphasis will be on renal transplantation.

Animals↗
PubMed1989

Complement activation during an active cytomegalovirus infection after renal transplantation: due to circulating immune complexes or alternative pathway activation?

In 32 patients with a renal allograft, serial determinations after transplantation were made of C3d, the stable conversion product of the complement factor C3, as well as serial measurements of the anaphylatoxin C3a des arg. Furthermore, serial determinations were made on the presence of circulating immune complexes using three different assays (C1q binding assay, polyethylene glycol precipitation test, and indirect granulocyte phagocytosis test). Twenty patients were studied during an active cytomegalovirus (CMV) infection, and 12 patients were studied during allograft rejection or during stable phase after renal transplantation. In 12 patients with a CMV infection serial measurements were made of AP50 (alternative pathway of complement). During an active CMV infection elevated C3d as well as elevated C3a des arg levels were found and not in the control group (P less than 0.01). In 8 out of the 12 patients tested, with CMV infection, a decreased hemolytic activity of the alternative pathway (AP50) was found, together with the elevated levels of C3d and C3a des arg. Serum C4 levels were normal or high during CMV infection. Furthermore, circulating immune complexes were found to be positive in 15 out of the 20 patients with a CMV infection (both primary and secondary infections), and in 2 out of 12 patients of the control group. The complement activation found in the CMV group was not related to the presence of circulating immune complex-like material, since complement activation was present in advance of the appearance of the immune complexes, suggesting that complement activation was not due to classical pathway activation by those complexes. We conclude that our data are consistent with complement activation and the formation of biologically active peptides like C3a des arg in patients with an active CMV infection. The decreased hemolytic activity of the alternative pathway (AP50) together with the normal or high C4 levels suggest involvement of the alternative pathway, although further studies of the alternative pathway of C are warranted to confirm this hypothesis.

Adolescent↗
PubMed1989

Cytomegalovirus antigenemia as a useful marker of symptomatic cytomegalovirus infection after renal transplantation--a report of 130 consecutive patients.

In earlier work we demonstrated that CMV immediate early antigens can be detected in peripheral blood leukocytes of patients with active CMV infection. We now report a comparison of the antigenemia assay and an anti-CMV ELISA in a prospective longitudinal study of 130 renal transplant recipients who were monitored for active CMV infection during the first 3 months after transplantation. Active CMV infection developed in 56 patients. The antigenemia assay had a sensitivity of 89% and a specificity of 93% in the diagnosis of active CMV infection; for the ELISA these figures were 95 and 100%, respectively. In 22 of the 56 patients a CMV syndrome occurred. Antigenemia was demonstrated in all 22 patients while an antibody response occurred in 21 of them. The antigenemia assay became positive 8 +/- 7 days before the onset of symptoms while the antibody response was observed 4 +/- 9 days after the onset of symptoms. The pattern of antigenemia was helpful for monitoring the course of the infection. The maximum level of antigenemia was significantly higher and its duration significantly longer in symptomatic than asymptomatic infection. We conclude that CMV antigenemia is a sensitive, specific, and early marker of CMV infection. The antigenemia assay is of great value in monitoring patients with a high risk of CMV infection.

Adult↗
PubMed1989

Characterization of membrane antigens on human cytomegalovirus-infected fibroblasts recognized by human antibodies.

The antigens on the surface of human cytomegalovirus (HCMV)-infected fibroblasts which are recognized by human HCMV antibody-positive sera were characterized. Three HCMV-induced polypeptides, with apparent molecular masses of 53 to 63, 94, and 94 to 120 kilodaltons, were precipitated from 125I-surface-labeled cell extracts with different sera obtained from healthy individuals. Renal transplant recipients who were suffering from active HCMV infections recognized the same set of antigens. By the use of monoclonal antibodies, these antigens were identified as polypeptides belonging to the gcI and gcIII families of HCMV glycoproteins.

Antibodies, Monoclonal↗
PubMed1989

Detection of cytomegalovirus antigens and DNA in tissues fixed in formaldehyde.

Immunohistochemical techniques with monoclonal antibodies against cytomegalovirus (CMV) immediate early (IEA) and early antigens (EA), and in situ hybridisation, were used to detect CMV infection in routinely obtained, formaldehyde fixed and paraffin wax embedded tissues taken from bone marrow transplant patients, who had died form interstitial pneumonia. To improve the rates of detection of CMV-IEA and EA the wax embedded material was pretreated with 0.4% pepsin/HCl at 37 degrees C for 30 minutes. This pretreatment was also advantageous for in situ hybridisation. In the patients with histological evidence of CMV infection or positive viral culture from the lung tissue, or both, viral proteins and nucleic acids were detected in lung, as well as in other organs. Immunohistochemical techniques proved superior in heavily infected but necrotic tissues. In control patients (patients who had died from interstitial pneumonia without any evidence of CMV, or with no interstitial pneumonia at all) in situ hybridisation showed no positive signal, while immunohistochemical techniques showed only a few positive cells in lung tissue of one of nine patients. In addition to CMV-DNA analysis, formaldehyde-fixed, paraffin wax embedded tissue is amenable to immunohistochemical analysis with CMV monoclonal antibodies.

Antigens, Viral↗