Biomedical subjects
Publications and source records attributed to J Naessens.
Two monoclonal antibodies (mAbs) belong to this cluster: IL-A77 and IL-A165. They clustered closely in PC16c. The first mAb, an IgM, was raised against bovine activated B cells, the second was an IgG1 raised against bovine bone marrow cells.
gamma delta T cells in ruminants can be subdivided in two or more subpopulations on the basis of the expression of surface antigen WC1, which can exist in different isoforms. In this study, 18 monoclonal antibodies (mAbs) submitted to the Third International Workshop that were predicted to react with gamma delta TcR molecules were analysed and expression of their antigens was investigated on the different gamma delta T cell subpopulations. A set of control mAbs positive for TcR1 (86D), BoCD3 (MM1A), WC1 (B7A1, BAQ4A, CACTB32A, and BAQ89A) was included for comparative studies. Previous investigations demonstrated eight of the mAbs immunoprecipitated peptides with apparent M(r)s of 37 and/or 47 kDa, indicating they recognized determinants on the T cell receptor, TcR1. Two color flow cytometric analyses in the present study demonstrated the mAbs formed three groups; group 1, a set of mAbs that recognize TcR1 determinants expressed on all gamma delta T cells and groups 2 and 3, sets of mAbs that recognize TcR1 determinants on some gamma delta T cells: TcR1-N6 and TcR1-N7 respectively. mAbs from the latter groups define families of TcR1 molecules that express either one or both of the determinants. These antigenically distinct forms of TcR1 are expressed in equal proportion on the two gamma delta T cell populations that express one of the mutually exclusive isoforms of WC1, WC1-N3 and WC1-N4. The data indicate usage of the mAb-defined families of the gamma delta TcR is primarily restricted to the WC1+ subpopulation of gamma delta T cells. However, a small subpopulation of CD2+, WC1- gamma delta T cells expresses a form of TcR1 positive for the determinant TcR1-N6.
Monoclonal antibodies potentially specific for antigens expressed or upregulated on activated leukocytes were selected for further analysis from the panel submitted to the third international workshop on ruminant leukocyte antigens. The kinetics of expression of these activation antigens on resting peripheral mononuclear cells (PBMC) and PBMC stimulated with concanavalin A or staphylococcal superantigen SECI for 4, 24 or 96 h were compared, as well as their appearance on various subsets of cells. For some of them, a molecular mass could be determined after immunoprecipitation from radio-labeled, lectin-stimulated cells. Based on the results from the clustering, kinetic studies and biochemical data, evidence was gathered for assigning two additional mAbs to cluster BoCD25 (IL-2 receptor) and two mAbs to cluster BoCD71 (transferrin receptor). Four mAbs recognized an early activation antigen predominantly expressed on gamma delta T cells in short-term cultures. A number of other activation antigens were further characterized.
After the preliminary clustering of the whole workshop panel using flowcytometry data, we selected those clusters that were composed of mAbs that bound a broad range of cells. To obtain evidence that mAbs in each of these preliminary clusters detected the same antigen, they were tested for their capacity to compete with each other for binding to a target cell and the molecular weights of their antigens were estimated after immunoprecipitation. Most preliminary clusters in the non-lineage panel contained control mAbs that had been characterised in one of the previous workshops, and this study therefore increased the number of mAbs available to each of these non-lineage antigens. One new interesting cluster, BoWC14, was described which defined an antigen on myeloid cells and a subpopulation of CD4, CD8 and WC1 T lymphocytes (BT3/8.12 and IL-A155). An additional cluster, which did not fit into any other panel, contained mAbs specific for an antigen restricted to the erythroid lineage and received the nomenclature BoWC15 (ANA8, IL-A135, IL-A137, IL-A138 and IL-A160).
It has been reported that some breeds of cattle such as the N'Dama mount a more effective antibody response to the variable surface glycoprotein coat of trypanosomes and that this may contribute to their ability to control the infection. Thus we have investigated antibody responses to surface exposed epitopes of the variable surface glycoprotein in Trypanosoma congolense-infected N'Dama (trypanotolerant) and Boran (susceptible) cattle. Similar titres and isotypes were found in both N'Damas and Borans indicating that trypanotolerance is not associated with superior antibody-mediated destruction of trypanosomes. However, significant differences in antibody responses to cryptic VSG epitopes and non-trypanosome antigens were identified. Trypanosusceptible Boran cattle had low IgG1 responses to cryptic epitopes but high IgM responses to non-trypanosome antigens such as beta-galactosidase. In contrast the N'Dama cattle had significantly higher IgG1 responses to cryptic VSG epitopes and negligible responses to beta-galactosidase. These results indicate differences in the induction of anti-trypanosome immune responses between trypanotolerant and susceptible cattle infected with T. congolense.
Murine models have suggested that CD8 T cells might play a major parasite-promoting role in African trypanosomiasis. To assess the role of these cells in a natural host environment, we have depleted CD8 cells from Boran cattle in vivo and subsequently infected these animals with Trypanosoma congolense by tsetse fly challenge. Following administration of a mouse monoclonal anti-bovine CD8 antibody, we have been able to achieve a depletion of more than 99.9% in peripheral blood, spleen, prescapular lymph nodes, prefemoral lymph nodes, mesenteric lymph nodes and Peyer's patches. Depletion could be maintained over a 4-5-week infection period. Despite the almost total absence of CD8 cells, no effect whatsoever was observed on parasitemia. In addition, anemia, which is the main factor determining the mean survival time in cattle was not affected by the CD8 depletion.
Are there differences in the patient characteristics and clinical outcome for transaortic renal endarterectomy vs. bypass grafting when either technique is combined with infrarenal aortic replacement for occlusive or aneurysmal disease? Two common perceptions persist: (1) combined aortic and renal procedures have a high risk and (2) bypass is easier and safer than endarterectomy. To address these controversies we compared 52 consecutive patients undergoing concomitant aortic and renal reconstruction between 1987 and 1991: 26 with bypass and 26 with endarterectomy. Bypass patients were older (70 vs. 64 years, p = 0.001), had more extensive plaque extending into the distal renal artery and more severe baseline azotemia (creatinine = 2.6 vs 1.7 mg/dl, p = 0.01), more clinically evident coronary heart disease (89% vs. 56%, p = 0.001), and a greater need for nephrectomy of a small nonfunctional pressor kidney (23% vs. 0%) than endarterectomy patients. In contrast, endarterectomy patients more commonly required aortic replacement for occlusive disease than for an aortic aneurysm (endarterectomy: 65% vs. 35%; bypass: 19% vs 81%, p = 0.002) and tended to require more intraoperative technical revisions (12% vs. 4%) than bypass patients. Both groups, however, experienced no operative mortality, had similar cardiorespiratory morbidity, and achieved equal improvement in hypertension (69% vs. 65%). Bypass patients, who already had more severe preoperative azotemia than endarterectomy patients, showed less improvement in the creatinine level (Cr = 2.1 vs. 1.4 mg/dl, p = 0.01) and had greater need for late dialysis (30% vs. 4%, p = 0.01). Only one patient on dialysis had graft occlusion.(ABSTRACT TRUNCATED AT 250 WORDS)
To determine optimal management and outcome of infected extra-anatomic bypass grafts (EABG), we reviewed 28 patients (19 men and 9 women; mean age 70 years) treated over a 13-year period. Mean follow-up was 42 months. There were 16 axillofemoral (AF), 10 femorofemoral (FF), and two axillopopliteal (AP) grafts. Risk factors included previous prosthetic graft infection in 13 patients, enterocutaneous fistula in two, and mycotic aortic aneurysm in one. Initial management involved complete graft excision in 12 patients, partial graft excision in 10, and nonresectional therapy in six. Failure of nonresectional therapy and partial excision in three patients each required further operative intervention with graft excision. Reconstruction in patients eventually requiring graft excision (n = 25) entailed placement of a new prosthetic AF or AP graft in eight, an autogenous FF graft in five, combined prosthetic AF and autogenous FF bypass in two, autogenous iliofemoral bypass in one, obturator bypass in one, or no reconstruction in eight. Four autogenous FF reconstructions thrombosed immediately postoperatively, and three prosthetic reconstructions became infected. The mortality rate was 18% (FF = 20%, AF = 19%, AP = 0%). The amputation rate was 25% (AP = 100%, AF = 25%, FF = 10%) and was higher without arterial reconstruction (56% vs. 12%, p = 0.02). Two patients required hemipelvectomies and one had bilateral hip disarticulation. We conclude that EABG infections can be successfully treated but carry significant morbidity and mortality. Optimal management includes EABG resection and prompt revascularization, bearing in mind the risk of early thrombosis in autogenous grafts and reinfection in prosthetic grafts.
A set of monoclonal antibodies (MoAbs) to leucocyte antigens is an essential tool to identify different cell types and functional membrane molecules involved in immune responses. Since no MoAbs existed to bovine integrins, except against the beta 2 subfamily, we generated MoAbs to beta 3 integrin after the immunization of mice with bovine platelets. Two MoAbs, IL-A164 (IgG2a) and IL-A166 (IgG1), were selected that reacted specifically with bovine platelets and detected the same membrane molecule. The antigen was a heterodimer of two polypeptide chains of 122 kDa and 95 kDa as resolved by SDS-PAGE under reducing conditions. Although the Mr of the smaller subunit is identical to that of beta 2 integrin, preabsorption with an antibody to beta 2 (or CD18) did not remove the bovine antigen. Comparing the molecular masses of the two subunits in reduced and non-reduced forms showed a pattern that was similar to that of human GPIIb/IIIa (also called alpha IIb beta 3 or CD41a). Reduction of the bovine molecule increased the apparent Mr of the light chain from 76 kDa to 95 kDa, while the heavy subunit changed from 136 kDa to 122 kDa. As with human GPIIb, the decrease in Mr of the alpha-subunit is probably a result of a small disulphide-linked polypeptide, although no additional evidence for this was detected for the bovine integrin. Sequencing of the N-terminal amino acids of both bovine polypeptides showed identity of the bovine integrin with human GPIIb/IIIa.
Twenty-three large employers in the Minneapolis/St. Paul area formed the business health care action coalition group (BHCAC) to obtain medical care for their employees and families. Requirements of BHCAC included: 1) detailed reporting of outcomes of both preventive and standard health care, 2) development and successful implementation of practice guidelines, and 3) development of a functioning automated medical record. The coalition was contracted to meet these requirements. A nonprofit foundation--the Institute for Clinical Systems Integration (ICSI)--was formed and the data standards committee created. To ensure registration information transmitted by network would be attached to the correct patient's computer file; a standardized registration data set was agreed upon to be recorded in a uniform manner. This innovative change has allowed, encouraged, and required that clinical data be transferable electronically among organizations. The following are the ICSI data standard requirements for ICSI member organizations: the Health Level 7 (HL7) is to be used for data transmission [1]. Fields to be used include: 1) Social Security Number (SSN): The SSN was chosen as the patient identifier for the reasons listed in [2]. Record the official SSN assigned by the Social Security Administration; record a pseudo-SSN if a social security number is not available following the Veterans Administration (VA) Hospital algorithm. 2) Name: Use the American National Standards Institute (ANSI) Healthcare Informatics Standards Planning Panel (HISPP) and Message Standards Developers Subcommittee (MSDS) Standard for representation of a person's name [3]; completely record all of the person's legal name (including any punctuation, hyphenated and double last names); nicknames and appellations are to be recorded separately; salutations and suffixes are optional, but must be placed in separate computer fields from other name parts. 3) Gender: Allowable values include FEMALE, MALE, OTHER, UNKNOWN. 4) Date of Birth: Include the day, month, year (including century); permitted is approximate or estimated birth dates provided they are indicated as such. Use the ANSI HISPP MSDS standard for date representation [4]. 5) Address: Record at least one address; multiple addresses desirable identified by type or usage (i.E., home, business, office). Allow two or more lines for street, city, U.S. state or Canadian Province; identify by standard two-letter postal abbreviation--zip code (at least 5-digits, preferably 9) or postal code and country. 6) Home ID or Medical Record Number (MRN): Record if different than Social Security Number.
Alterations in the phenotype and function of cells isolated from bovine afferent lymph were studied following tsetse-transmitted Trypanosoma congolense infection. Little alteration was observed in the output of the CD2+ T cells in the lymph, and within this population the CD4:CD8 ratio remained relatively constant. By contrast, a marked decrease was observed in the output of gamma delta T cells over the first 7 days following infection. The number of B cells increased between 2 and 6 days post-infection, and thereafter returned to pre-infection values. Little change was observed within the afferent lymph veiled cell population. Examination of activation markers on the lymphocyte fraction of afferent lymph revealed a decrease in the number of cells expressing the Interleukin-2 receptor alpha-chain from Day 5 post-infection. At this time the expression of ACT 1, another early activation marker, was seen to increase. Afferent lymph cells collected pre-infection and on the first 4 days post-infection proliferated in response to stimulation with Concanavalin A in vitro. This response to mitogenic stimulation was completely abrogated from day five post-infection. However, these cells were not capable of suppressing the capacity of normal peripheral blood mononuclear cells to respond to mitogenic stimulus in co-culture assays. These studies suggest that although a degree of lymphocyte activation occurs in the afferent lymph following tsetse-transmitted infection with T. congolense, this may be sub-optimal owing to the immunosuppression which appears to operate at the level of the skin and the lymph nodes.
B cells from the peripheral blood and spleen of Trypanosoma congolense-infected cattle and from the peripheral blood of an uninfected cohort were analysed for ability to secrete antibody and for expression of surface antigens before and after in vitro culture with interleukin-2, lipopolysaccharide and pokeweed mitogen. Antibody-secreting cells (ASC) were only detected in lymphocytes from peripheral blood after in vitro stimulation. The frequency of ASC was greater in cultures of lymphocytes from infected cattle than from the uninfected cohort. The frequency of ASC was positively correlated with the number of B cells expressing the transferring receptor but not with the expression of the CD5 antigen.
The expression of major histocompatibility complex class II (MHC II) non-polymorphic antigens detected by four monoclonal antibodies was investigated in Theileria parva-infected and non-infected cloned lymphoid cell lines, bulk cultures, and in peripheral blood mononuclear cells (PBMC) and lymph node cells (LNC) of experimentally infected calves. Compared with non-infected cell lines, both immunofluorescence microscopy and flow cytofluorometry analysis of infected lines of alpha beta T-cell, gamma delta T-cell and B-cell origin revealed high expression of MHC II MHC molecules. After T. parva infection in vitro, three alloreactive T cell clones, three interleukin-2 (IL-2)-dependent cell lines and a concanavalin A (Con A)-stimulated bulk culture all had an increase both in the proportion of MHC II+ cells and in their mean fluorescence intensity. Radioimmunoprecipitation of class II molecules biosynthesized in infected and non-infected cells revealed that they were constitutively produced in infected cells, and were a slightly larger relative mass than the MHC II molecules of uninfected cells. In a study of the serial expression of MHC II antigens in PBMC and LNC of six calves inoculated with a lethal dose of T.parva, MHC II expression by non-parasitized cells peaked at Days 7 (LNC) or 9 (PBMC) following inoculation and, subsequently, MHC II non-expressing parasitized lymphocytes progressively outnumbered MHC II-expressing parasitized cells. In two calves studied in detail, MHC II expression in PBMC and LNC generally, and in T cells particularly, increased during the course of the disease. Finally, among LNC sorted for MHC II expression at 11 and 17 days after parasite inoculation, the proportion of parasitized cells increased markedly in MHC II non-expressing populations and was reduced or increased only slightly in MHC II-expressing populations. These findings indicate that: (1) enhanced MHC II antigen expression by parasitized lymphocytes may be important in the pathogenesis of the lymphoproliferation that characterizes T. parva infection; (2) the in vivo preponderance of MHC II non-expressing over MHC II-expressing T. parva-infected cells may reflect host-mediated destruction or antigenic modulation of parasitized MHC II-expressing cells.
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Three monoclonal antibodies (mAbs), P13 (#90), IL-A78 (#76) and IL-A79 (#77), defined a new cluster, called BoWC8. The antigen defined by these mAbs is a molecule of 160 kDa under reducing conditions, and is expressed on lymphocytes late (6 days) after activation. The molecule can be expressed on multiplying T cells, but not on B cells.
Monoclonal antibodies (MAbs) from three temporary clusters recognized antigens expressed on platelets and a wide variety of other tissues. To confirm the clustering of the mAbs, we biochemically characterized the antigens recognized by each cluster. TC9 (BoCD44) contained mAbs to a 95 kDa ruminant antigen, homologous to human CD44. TC11 (BoWC9) defined a 25 kDa antigen and TC7 (BoWC11) defined an antigen composed of two subunits of 49 and 79 kDa.
We analysed the surface phenotype of bovine bone marrow erythroid and myeloid progenitor cells with monoclonal antibodies (mAbs) from the Second Workshop. For the antibodies tested, no difference could be observed in burst-forming unit (erythroid) and colony-forming unit (erythroid) both are positive for BoCD44, BoWC9, MHC Class I, transferrin receptor and the p150/158 antigen detected by BT3/8.12, but neither express BoCD11a, BoCD45, BoWC5 or the antigen recognized by mAb Bo116. The myeloid progenitor cells, colony-forming unit (granulocyte/macrophage), can be discriminated from the erythroid progenitors by the absence of a transferrin receptor and the expression of BoCD11a and BoWC5 antigens. By selecting the right panel of mAbs, it should now be possible to enrich bone marrow cells for erythroid and/or myeloid progenitor cells.