Biomedical subjects

M D Kramer

Publications and source records attributed to M D Kramer.

At least 109 records · Page 6Linked to original sources

PubMed1991

A mouse model for Borrelia burgdorferi infection: approach to a vaccine against Lyme disease.

Lyme disease is the most common vector-transmitted illness in the USA and Europe. The pathogen, Borrelia burgdorferi, causes a complex spectrum of disease that affects the skin, joints, nervous system and heart. Patients in the early stages of the disease can often be successfully treated with antibiotics but this becomes less reliable as the disease progresses. The specific immune responses that are detectable in patients with Lyme disease are not sufficient to protect against illness; a vaccine against the infectious agent is, therefore, desirable. In this article Markus M. Simon and colleagues present a mouse model of Borrelia burgdorferi infection. Their work suggests that the outer surface protein A (OspA) of Borrelia burgdorferi may be a suitable vaccine candidate.

Animals↗

PubMed1991

Establishment of an enzyme-linked immuno-sorbent assay for urinary trypsin inhibitor by using a monoclonal antibody.

Monoclonal antibodies against inter-alpha-trypsin-inhibitor (ITI) were produced. One clone showing specificity for urinary trypsin inhibitor (UTI), a proteolytic fragment of ITI, which is excreted into urine, was selected for the establishment of an enzyme-linked immuno-sorbent assay (ELISA). The ELISA for the quantification of UTI was shown to work reproducibly in the range between 0.5 and 10 ng UTI/ml urine. Urines of several patients suffering from different lung diseases were screened for UTI using the established ELISA. Highest UTI levels were found in the urine of patients with lung empyema. A more moderate increase was observed in patients suffering from lung tuberculosis and from secondary and primary lung tumors.

Alpha-Globulins↗

PubMed1991

Antigenic domains of the HIV-1 vif protein as recognized by human sera and murine monoclonal antibodies.

To analyze the vif antibody response in individuals infected with the human immunodeficiency virus type 1 (HIV-1) and to determine antigenic epitopes on the vif protein, 104 HIV-1+ sera were screened for reactivity with a recombinant vif protein; 30 (28.8%) of these sera recognized the recombinant vif protein in immunoblot and were employed, together with 17 HIV-1/vif-negative control sera, in an enzyme immunoassay (EIA)-based epitope scanning assay with 183 overlapping decapeptides that covered the complete amino acid sequence of the HIV-1 vif protein (strain BH10). Of the 30 HIV-1/vif+ sera, 87% reacted with decapeptides comprising the two following epitopes: IEWRKKRY (vif amino acids 87-94) or DRWNKPQ (vif amino acids 172-178). The two epitopes were 89% and 100% conserved among different HIV-1 strains and their antigenicity could be confirmed by computer-assisted predictions of vif antigenic determinants. All the sera reactive with recombinant vif protein and with vif peptides originated from patients in CDC stages III or IV. Two murine anti-vif monoclonal antibodies reacted only with the seven C-terminal amino acids of the vif protein (SHTMNGH), which were not recognized by any of the human sera. Our results may be useful for further studies of vif seroreactivity and for the production of anti-vif mono- or polyclonal antibodies using vif peptides.

Amino Acid Sequence↗

PubMed1991

Recombinant outer surface protein a from Borrelia burgdorferi induces antibodies protective against spirochetal infection in mice.

The outer surface protein A (OspA) of Borrelia burgdorferi was isolated in its native form from strains ZS7 and B31 and as a recombinant protein from strain ZS7. Amino acid sequence analysis of internal peptides of native OspA (strain ZS7) revealed identity with the sequence deduced from the OspA gene. Repeated immunization of C57BL/6 and C.B-17 mice with any of the three OspA structures resulted in the generation of monospecific hyperimmune sera reactive with both native and recombinant OspA. Upon transfer of immune sera specific for either native OspA (strain B31) or recombinant OspA (strain ZS7) but not of those reactive with the recombinant 41-kDa flagellin-associated antigen, severe combined immunodeficient (SCID) mice were completely protected against infection with strain ZS7. The finding that monoclonal antibodies to OspA and to OspB but not to non-outer surface spirochetal structures such as flagellin, p20, p65, and p70 conferred protection in SCID mice makes OspA (and possibly OspB) a promising candidate vaccine against Lyme disease.

Amino Acid Sequence↗

PubMed1991

In vivo primed mouse T cells selectively express T cell-specific serine proteinase-1 and the proteinase-like molecules granzyme B and C.

Previous studies have shown that mouse CD8+ T lymphocyte clones (TLC) produce T cell-specific serine proteinase-1 (MTSP-1) as well as a family of six homologous molecules, termed granzymes B - G, which are structurally related to serine proteinases. Of these proteins, only MTSP-1 has been studied in detail. It has been shown to occur in the majority of CD8+ and a fraction of CD4+ T effector cells in vivo and in vitro and has demonstrable enzyme activity in these cells. The presence of the other serine proteinase-like molecules in T cells is less well defined. We have now analyzed the expression of mRNA species specific for granzymes B - G in activated T cell populations using the sensitive polymerase chain reaction which allows the detection of mRNA species from as little as 2 pg of total cytoplasmic RNA. We demonstrate that MTSP-1 and all six serine proteinase-like transcripts are expressed in a panel of four CD8+ and six CD4+ long-term-cultured TLC, though at greatly differing concentrations. In contrast, in vivo primed T cells of both phenotypes, CD4+ and CD8+, and in vitro activated T cells derived from short-term cultures only express mRNA species specific for MTSP-1 and CCP1 and little of those for CCP2, but no transcripts for granzymes D - G. These findings argue against the participation of granzymes D - G in T cell-mediated functions in vivo.

Animals↗

PubMed1991

Immunogold labeling of perforin and serine esterases in granulated metrial gland cells.

Cytotoxic T lymphocytes (CTL) and natural killer (NK) cells are cytolytic lymphocytes known to produce a pore-forming protein, named perforin or cytolysin, that lyses target cells by creating large pores on the target plasma membrane. Besides perforin, the granules of CTL and NK cells contain a family of serine esterases. Perforin has also been localized in granulated metrial gland (GMG) cells of the murine embryo implantation site by light microscopic immunostaining. Ultrastructural immunogold labeling with antibodies against perforin and a serine esterase (MTSP 1 or granzyme A) shows that GMG cells contain both perforin and serine esterases in the fine granular matrix of their granules. Perforin has been located in all of the granules, whereas gold particles corresponding to serine esterases have been found in most of the granules. Results from the double immunogold technique indicate that perforin and serine esterases colocalize to most of the same granules in GMG cells. This study supports the view that GMG cells are related to cytolytic lymphocytes.

Animals↗

PubMed1991

Maternal smoking and placenta previa.

We conducted a case-control study of the relation between smoking and placenta previa, using Washington State birth certificate data from 1984 through 1987. The study population was comprised of live, singleton births to women whose pregnancies were complicated by placenta previa (N = 598) and randomly selected controls (N = 2,422) from the same time period. We used logistic regression to estimate odds ratios (OR) and their 95% confidence intervals (CI). Maternal smoking approximately doubled the risk of placenta previa after adjustment for the confounding effect of maternal age (OR = 2.1, 95% CI: 1.7-2.5).

Adult↗

PubMed1991

Expression of T-cell-associated serine proteinase 1 during murine Leishmania major infection correlates with susceptibility to disease.

The expression of T-cell-associated serine proteinase 1 (MTSP-1) in vivo during Leishmania major infection was analyzed in genetically resistant C57BL/6 mice and in genetically susceptible BALB/c mice. Using a monoclonal antibody as well as an RNA probe specific for MTSP-1 to stain tissue sections, we found T cells expressing MTSP-1 in skin lesions and spleens of mice of both strains. In skin lesions, MTSP-1-positive T cells could be detected as early as 3 days after infection. Most importantly, the frequency of T cells expressing MTSP-1 was significantly higher in susceptible BALB/c mice than in resistant C57BL/6 mice. These findings suggest that MTSP-1 is associated with disease-promoting T cells and that it may be an effector molecule involved in the pathogenesis of cutaneous leishmaniasis.

Animals↗

PubMed1991

Myositis in mice inoculated with Borrelia burgdorferi.

The authors describe the appearance of myositis in immunocompetent and immunodeficient mice after subcutaneous inoculation with Borrelia burgdorferi by histology and immunohistology. Experimental infection of mice 1) causes inflammation of striated but not smooth muscles, 2) affects the entire musculoskeletal system, and 3) is characterized by perivascular and interfacicular infiltration of mononuclear leukocytes in the striated muscle leading to necrosis as well as disruption of muscle fibers. The lesions found in striated muscle specimens were most pronounced in immunodeficient (SCID), less severe in T-cell-deficient nu/nu (BALB/c, C57BL/6) and marginal to moderate or almost not present in immunocompetent AKR/N and C.B-17 mice, respectively.

Animals↗

PubMed1991

Activation of the plasminogen activator system in a keratinocyte cell line (HaCaT) by alkyl sulfates.

A human keratinocyte cell line (HaCaT) was analysed for plasminogen activator activity. By immuno-capture and immuno-inhibition studies we obtained evidence that HaCaT cells synthesize and secrete urokinase-type plasminogen activator (uPA) and tissue-type PA (tPA). Zymographic analysis by using fibrin agar indicator gels indicated that part of the PA activity in culture supernatants of the HaCaT line is complexed with putative PA inhibitors (PAI). Alkyl sulfates of different chain length were found to have a strong stimulatory effect on PA activity in HaCaT culture supernatants. Our findings are discussed in view of the known skin-irritating effects of alkyl sulfates in vivo.

Cell Line↗

PubMed1991

[Borrelia burgdorferi infection. II. Aspects of basic research, new approaches to diagnosis and therapy].

In this review article, current thinking about the infection caused by Borrelia burgdorferi is presented. The problems associated with diagnosis and treatment of Borrelia burgdorferi infection are pointed out. A series of monoclonal antibodies recognizing Borrelia burgdorferi-associated antigens is presented. In collaborative research projects these antibodies have been used for molecular biological (cloning and sequencing of Borrelia burgdorferi antigens) and immunological (protection experiments by passive transfer of monoclonal antibodies in an animal model) experiments. Taken together, these studies disclosed new aspects for diagnosis and therapy of the Borrelia burgdorferi infection. In this second part of the review article the above-mentioned molecular biological and protection experiments are summarized.

Amino Acid Sequence↗

PubMed1991

Mouse T-cell associated serine proteinase 1 degrades collagen type IV: a structural basis for the migration of lymphocytes through vascular basement membranes.

We show that CD8+ T-lymphocyte lines perferentially attach to collagen type IV and that mouse T-cell specific serine proteinase 1 (MTSP-1) preferentially degrades native basement membrane collagen type IV. In contrast, the interstitial collagen types I, II, III, V and VI appear not to be affected. The data reveal that MTSP-1 predominantly cleaves the alpha 2(IV) chain, which is found in the native triple helical structure of type IV collagen in a ratio of alpha 1(IV): alpha 2(IV) = 2:1 into small peptides. The cleavage of the alpha 2(IV) chain within the native collagen type IV molecules most likely results not only in a destabilization of single molecules but of the entire collagenous basement membrane scaffold at the site of MTSP-1 secretion.

Animals↗

PubMed1991

A mouse model for Borrelia burgdorferi infection: pathogenesis, immune response and protection.

Viable Borrelia burgdorferi (B. burgdorferi) organisms induce a chronic infection associated with arthritis, carditis and hepatitis in severe combined immunodeficiency (scid) mice but not in most of the adult mice from the various immunocompetent inbred strains tested. Furthermore, we have found that experimental inoculation of normal mice with B. burgdorferi organisms leads to the generation of antibodies and T cells specific for various spirochetal antigens including the outer surface proteins A and B (OspA, OspB) as well as flagellin. The assumption of a protective role of the immune response during B. burgdorferi infection in mice is supported by our recent findings that passively transferred B. burgdorferi-specific immune mouse sera as well as monoclonal antibodies to OspA are able to prevent the development of the disease in scid mice. We show now that purified OspA protein both in its native and recombinant form is immunogenic and that the antibodies generated are able to confer protection to scid mice against B. burgdorferi infection.

Animals↗

PubMed1990

Measurement of free human leukocyte elastase and human leukocyte elastase/alpha 1 proteinase inhibitor complexes by an enzyme-linked immunosorbent assay.

We report on an ELISA procedure for the quantitative analysis of total human leukocyte (PMN)-elastase, i.e., the simultaneous determination of (i) free (non-complexed) PMN-elastase and of (ii) PMN-elastase that is complexed to alpha 1 proteinase inhibitor. Simultaneous detection of both forms of PMN-elastase was achieved using a monoclonal antibody that recognizes a PMN-elastase-specific epitope present on both the uncomplexed and the complexed form of the enzyme. The test system described is reliable, easy to perform and permits the determination of total PMN-elastase in complex biological fluids such as plasma or seminal fluid. It is to be expected that this test system will be useful for investigations of human PMN-elastase in biological specimens obtained from both normal and pathological conditions.

Antibodies, Monoclonal↗

PubMed1990

Quantification of free alpha 2-macroglobulin and alpha 2-macroglobulin-protease complexes by a novel ELISA system based on streptococcal alpha 2-macroglobulin receptors.

An ELISA test system has been developed for the quantification of the two distinct forms of the proteinase inhibitor alpha 2-macroglobulin (alpha 2M): (i) free alpha 2M (functionally active), which is the electrophoretically slow form (alpha 2 MS), and (ii) the alpha 2 M-proteinase complex (functionally inactive), which is the electrophoretically fast form of alpha 2 M (alpha 2 MF). Discrimination between the two types of alpha 2 M was achieved using extracts of the two independent streptococcal strains, M1 and Sc1, which express receptors for alpha 2 MS and alpha 2 MF, respectively, in combination with a monoclonal antibody specific for alpha 2 M. The assay system described is easy and reliable and permits quantitation of alpha 2 MS and alpha 2 MF in complex biological samples such as plasma and cutaneous suction blister fluid.

Blister↗

PubMed1990

Activation of pro-urokinase by the human T cell-associated serine proteinase HuTSP-1.

The human T cell-associated serine proteinase-1 (HuTSP-1) is expressed by activated T lymphocytes and is exocytosed upon their interaction with target cells. Here, we report that HuTSP-1 is able to convert single-chain human pro-urokinase into the active two-chain enzyme. Time-dependent activation by HuTSP-1 of recombinant human pro-urokinase as well as natural pro-urokinase derived from human melanoma cells was demonstrated in a chromogenic assay specific for active urokinase type plasminogen activator and in immunoblotting experiments revealing the conversion of single-chain into two-chain urokinase. Control experiments excluded plasmin as the activating agent. These data suggest a novel pathway for plasmin generation during T cell-mediated processes such as immune responses and extravasation of immune cells.

Enzyme Activation↗

PubMed1990

Lyme carditis in immunodeficient mice during experimental infection of Borrelia burgdorferi.

Recently, we described the severe combined immunodeficiency (scid) mouse as a laboratory model for B. burgdorferi infection. Scid mice inoculated with the virulent low-passage tick isolate Borrelia burgdorferi ZS7 developed a severe pancarditis involving endocardium, myocardium and epicardium in the absence of functional B- or T-cells. Soon after inoculation perivascular infiltration was observed, later diffuse infiltration of the interstitium of the subendocardial and subepicardial areas was seen. The infiltrate was mainly mononuclear and predominantly composed of Mac-1+ cells. Concomitantly, fibroblast proliferation and augmented collagen deposition occurred in the interstitium. This was associated with the presence of B. burgdorferi organisms. The histopathological and ultrastructural findings observed in scid mice resemble those observed in human Lyme carditis. The data emphasize the suitability of the scid mouse as a model in which to study the role of the immune system in the pathogenesis of Lyme carditis.

Animals↗

PubMed1990

Demographic changes in coronary artery bypass surgery and its effect on mortality and morbidity.

Over the past 4-5 years, possibly with the advent of percutaneous transluminal coronary angioplasty (PTCA), there has been a changing patient population for coronary artery bypass surgery (CABS) with a gradual increase in the operative mortality. In an attempt to analyze the changing demographics in patients undergoing CABS and its effect on operative mortality, we analyzed data from 5536 consecutive patients undergoing isolated CABS. There was 4151 patients less than 70 years of age and 1385 patients greater than 70 years. Reoperative CABS procedures were performed in 385 patients, and CABS for post infarction unstable angina pectoris was performed in 578 patients. During the same time period, 2910 patients underwent PTCA. The mean age of bypass patients was 68.5 years with 38% being 70 years or older. The left ventricular ejection fraction in patients undergoing CABS averaged 38%. The average number of bypasses performed was 3.1. In comparison, patients presenting for PTCA were younger (average age 55), had normal ejection fractions (average 55%) and were predominantly treated for single or double vessel disease. The hospital mortality for elective CABS in patients less than 70 years of age was 1.8%, for reoperative CABS 3.6%, for post infarction unstable angina pectoris 4%, and for patients greater than 70 years 8%, for a combined operative mortality of 4.8%. These data suggest that because of the increasing number of elderly patients (greater than 70 years of age), and the increasing number of reoperative CABS cases and acute myocardial infarction patients with unstable angina pectoris presenting for CABS, the operative mortality will continue to rise.

Adult↗