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Biomedical subjects

R C Kennedy

Publications and source records attributed to R C Kennedy.

At least 55 records · Page 3Linked to original sources
PubMed1996

Immunogenicity of group B Streptococcus type III polysaccharide-tetanus toxoid vaccine in baboons.

Maternal vaccination has been proposed as a rational approach for the prevention of neonatal group B streptococcal (GBS) disease. In this study, baboons were used as a nonhuman primate model to evaluate the immunogenicity of a GBS type III glycoconjugate vaccine. Type III-specific immunoglobulin G with opsonic activity was induced after vaccination with type III polysaccharide coupled to tetanus toxoid administered with an aluminum adjuvant. This suggests that baboons could be used in evaluating maternal transfer of GBS-specific antibodies by vaccination during pregnancy.

Animals↗
PubMed1996

Resistance of previously infected chimpanzees to successive challenges with a heterologous intraclade B strain of human immunodeficiency virus type 1.

To test whether the protective effects of attenuated simian immunodeficiency virus vaccines in macaques were applicable to the human immunodeficiency virus type 1 (HIV-1)-chimpanzee system, two groups of animals, previously infected with HIV-1(IIIB) or HIV-1(SF2) were each challenged with a heterologous clade B virus, HIV-1(DH12). Following challenge, the parameters measured included virus isolation (from plasma, peripheral blood mononuclear cells, and lymph node tissue); quantitative DNA PCR using primers capable of distinguishing HIV-1(IIIB), HIV-1(SF2), and HIV-1(DH12) from one another; and serologic assays to monitor changes in binding and neutralizing antibodies. In contrast to an HIV-1-naive chimpanzee that rapidly became infected following the inoculation of HIV-1(DH12), the two chimpanzees previously infected with HIV-1(IIIB) resisted repeated and escalating inoculations of HIV-1(DH12), as monitored by virus isolation and PCR. The two animals previously infected with HIV-1(SF2) became infected with HIV-1(DH12) but in contrast to the case with the HIV-1-naive chimpanzee, no cell-free viral RNA was detected in the plasma by the branched DNA procedure and levels of peripheral blood mononuclear cell-associated viral DNA were reduced 35- to 50-fold.

AIDS Vaccines↗
PubMed1995

Nucleic acid vaccination against virally induced tumors.

Plasmid DNA (pSV3-neo) encoding the large tumor antigen (T-ag) of simian virus 40 (SV40) was used to actively immunize mice to assess the induction of SV40 T-ag specific immunity. Mice were injected with naked DNA intramuscularly, and both the cellular and humoral immune responses were compared to those elicited in mice immunized with recombinant protein. Administration of recombinant SV40 T-ag elicited high titer antibodies reactive with SV40 T-ag, whereas inoculation with DNA failed to generate comparable levels of SV40 T-ag specific antibody. Conversely, antigen specific cytotoxic T lymphocyte activity was generated in mice immunized with pSV3-neo, but was not detected in mice immunized with the recombinant protein. Moreover, the cellular immunity generated by the injection of pSV3-neo DNA was protective against a lethal challenge with syngeneic SV40 transformed cells. Together, these data indicate that active immunization with genes encoding tumor specific antigens may be an efficacious strategy for the induction of cell-mediated mechanism(s) to prevent cancer.

Animals↗
PubMed1995

Induction of antibodies to the human immunodeficiency virus type 1 by immunization of baboons with immunoglobulin molecules carrying the principal neutralizing determinant of the envelope protein.

The hypervariable region 3 (V3) within the disulfide-bridged loop of the envelope protein of the human immunodeficiency virus type 1 (HIV-1) contains an amino acid sequence that was defined as a principal neutralizing determinant (PND). A 19-amino acid residue consensus sequence (designated V3C) predicted from the PND sequences of 245 isolates as well as a sequence from the PND of the WMJ2 HIV-1 isolate (designated V3M) were expressed on the variable region of murine-human immunoglobulin (Ig) chimeras that were designated Ig-V3C and Ig-V3M, respectively. The HIV-1 sequences on the Ig chimeras preserved their antigenicity and interacted with antibodies specific for peptides encompassing the V3C and V3M sequences. In baboons, Ig-V3C and Ig-V3M induced antibodies that bound V3C and V3M peptides as well as the glycoprotein gp120 envelope protein of HIV-1 MN isolate. In addition, the baboons' antisera were able to prevent infection of CD4 SupT1 susceptible T cells by HIV-1 MN. Finally, Ig-V3M chimeras were able to stimulate in vitro production of antibodies specific for the HIV-1 envelope-derived peptides by lymphocytes from HIV-1-infected human subjects.

AIDS Vaccines↗
PubMed1995

T cells and HIV-induced T cell syncytia exhibit the same motility cycle.

Ameboid cells ranging in complexity from Dictyostelium amebas to human polymorphonuclear leukocytes (PMNs) translocate in a cyclical fashion. Using computer-assisted motion analysis, we have analyzed the motility of human lymphocytes of the immortal SupT1 cell line and of a peripheral blood mononuclear cell population highly enriched for CD4-positive cells (CD4-enriched PBMCs) on four substrates--plastic, dehydrated rat tail collagen, hydrated rat tail collagen, and bovine aortic endothelium. In addition, we have analyzed the motility on these substrates of syncytia induced by human immunodeficiency virus (HIV) in cultures of both cell types. It is demonstrated that both SupT1 cells and CD4-enriched PBMCs exhibit a motility cycle with a period of 1.6 min that is independent of substrate, independent of average cell velocity, and similar to the periods of translocating Dictyostelium amebas and PMNs. More surprisingly, it is demonstrated that HIV-induced SupT1 and PBMC syncytia with volumes 10 to 100 times those of single cells exhibit the same motility cycle as their single-cell progenitors. These observations support the generality of the motility cycle in animal cells and, for the first time, demonstrate that the cycle is independent of cell size.

CD4 Antigens↗
PubMed1995

HIV-induced syncytia in peripheral blood cell cultures crawl by extending giant pseudopods.

It was previously demonstrated that HIV-induced syncytia of the immortalized T cell line SupT1 reorganize their cytoskeleton and form a spherical supernuclear complex, thus mimicking the organization, polarity, and morphology of a single SupT1 cell. Then, through extension of a single, giant pseudopod, these syncytia, which grow to more than 100 times the volume of a single SupT1 cell, translocate along a substratum. To verify that syncytium motility is not peculiar to the SupT1 cell line, we have analyzed the cytoskeletal organization and motile capabilities of HIV-induced syncytia formed in peripheral blood cell cultures containing more than 90% CD4-positive cells. The results demonstrate that although peripheral blood T cells differ from SupT1 cells in size and morphology, they are continuously motile and translocate along a substratum in a manner quite similar to that of SupT1 cells, and peripheral blood T cell syncytia induced by HIV-1LAI as well as two additional clinical isolates translocate by the extension of a giant anterior pseudopod in a fashion indistinguishable from that of HIV-induced SupT1 syncytia. Together, these results support the generalization that HIV-induced T cell syncytia are motile.

Adult↗
PubMed1995

Examination of lymphokines induced in mice following immunization with recombinant simian virus 40 large tumor antigen.

Baculovirus-derived recombinant simian virus 40 (SV40) large tumor antigen (T-Ag) was used to immunize BALB/c mice to examine the lymphokines produced following immunization. Specifically, we examined production of interleukin-2 (IL-2), IL-4, IL-5 and interferon gamma (IFN gamma) from immune lymphocytes cultured with decreasing concentrations of recombinant SV40 T-Ag. We identified elevated levels of IFN gamma and IL-2 by enzyme-linked immunosorbent assay and a murine CTLL-2 proliferation bioassay respectively. We were unable to detect either IL-4 or IL-5. These data indicate the previously reported tumor immunity induced by recombinant SV40 T-Ag immunization most likely reflects a TH1-like immune response based on the in vitro production of both IFN gamma and IL-2 by immune lymphocytes.

Animals↗
PubMed1995

Phagocytosis of individual CD4+ T cells by HIV-induced T cell syncytia.

Transmission electron microscopic analysis of HIV-induced syncytia of the CD4+ SupT1 cell line has revealed profiles of whole T cells in the syncytium cytoplasm. Serial sections demonstrate that these T cells are completely enveloped by a second membrane in the syncytium cytoplasm and represent phagosomes. Pycnosis of engulfed T cell nuclei, vacuolation of the cytoplasm of engulfed T cells, and the association of engulfed T cells with dense vesicular clusters in the syncytium cytoplasm support the conclusion that they represent phagosomes. In addition, transmission electron micrographs of the syncytium surface reveal giant pseudopodial extensions wrapping around T cells, in a fashion similar to bacterial and yeast phagocytosis by professional phagocytes. These results suggest that phagocytosis is a characteristic acquired during HIV-induced syncytium formation, and that it may represent an avenue of T cell death in addition to fusion in HIV-infected SupT1 cell cultures.

CD4-Positive T-Lymphocytes↗
PubMed1995

Presence of neutralizing antibodies to heterologous human immunodeficiency virus type 1 isolates in sera of infected individuals is not predictive of rate of disease progression.

These studies were undertaken to examine whether the presence of human immunodeficiency virus type 1 (HIV-1)-neutralizing antibodies in sera of infected individuals would alter the rate of disease progression. HIV-1-infected individuals (n = 87) were initially examined for neutralizing activity in vitro against both laboratory and tissue culture-adapted clinical heterologous HIV-1 isolates. The neutralizing activities of sera were determined by a 90% or greater reduction in HIV-1 p24 levels in vitro. In a cross-sectional analysis of all infected individuals, we observed that sera from asymptomatic individuals neutralized a significantly greater number of heterologous HIV-1 isolates than sera from symptomatic patients. Patients who could be followed up longitudinally (n = 24) were then studied to determine the impact of neutralizing antibodies on the rate of disease progression. We observed no significant difference between the numbers of HIV-1 isolates neutralized in vitro by sera from patients who remained clinically stable and by those from patients who progressed rapidly. Our data indicated that the presence or absence of neutralizing antibodies to heterologous HIV-1 isolates was not associated with the rate of disease progression.

Adult↗
PubMed1995

A chemically defined liquid medium that supports primary isolation of Rochalimaea (Bartonella) henselae from blood and tissue specimens.

Rochalimaea (Bartonella) henselae is a fastidious, slowly growing, gram-negative bacillus that is an etiologic agent of bacillary angiomatosis, cat scratch disease, and related syndromes. Accumulation of direct microbiologic evidence of the relationship between the organism and the syndromes compatible with cat scratch disease has been hindered by the difficulties in the primary isolation of the organism from infected tissue specimens. A chemically defined liquid medium was developed to support the growth of Rochalimaea species to facilitate study of the organism. This medium was also used successfully to isolate R. henselae from clinical specimens from infected patients and a domestic cat. Recovery of R. henselae in this was more successful than when recovery was attempted on solid agar. This cell-free, extract-free, defined medium additionally supported the growth of Rochalimaea quintana and Afipia felis.

Angiomatosis, Bacillary↗
PubMed1994

Immunization of BALB/c mice with recombinant simian virus 40 large tumor antigen induces antibody-dependent cell-mediated cytotoxicity against simian virus 40-transformed cells. An antibody-based mechanism for tumor immunity.

In this report, we describe an immune mechanism that seems to play a role in tumor immunity in BALB/c mice challenged with SV40-transformed cells. This immune mechanism was induced by immunization of mice with rSV40 large tumor Ag (T-Ag). After rSV40 T-Ag immunization, BALB/c mice were protected from a lethal tumor challenge with syngeneic SV40-transformed cells (mKSA). Specifically, we examined CTL activity, NK activity, complement-dependent cytotoxicity, and Ab-dependent cell-mediated cytotoxicity (ADCC) in rSV40 T-Ag-immunized mice that were protected from a subsequent lethal tumor challenge. Immune splenocytes from surviving animals that were subsequently primed in vitro with rSV40 T-Ag demonstrated little to no SV40 T-Ag-specific CTL activity. In addition, natural immunity, as assessed by NK lytic activity, was comparable to that of unimmunized animals. Similarly, minimal significant complement-dependent cytotoxicity of radiolabeled mKSA cells was demonstrated by SV40 T-Ag-specific serum Abs. However, SV40 T-Ag-specific Abs using peritoneal exudate cells as effectors, exhibited significant levels of ADCC against radiolabeled mKSA target cells. Together, these studies indicate that rSV40 T-Ag immunization elicited ADCC that may represent a potential humoral immune mechanism involved in the protection of BALB/c mice from a lethal challenge with syngeneic SV40-transformed cells.

Animals↗
PubMed1994

Folding patterns of immunoglobulin molecules identified by urea gradient electrophoresis.

The reversible denaturant-induced unfolding of immunoglobulin molecules has been analyzed by transverse urea gradient gel electrophoresis and the effects that urea-induced unfolding exerts on the functional properties associated with their variable region, i.e. antigen binding and idiotypic expression, have been determined by Western blot analysis. Results obtained from these experiments indicate that urea-induced unfolding of the immunoglobulin molecule is a highly cooperative reversible process that occurs through a two-state transition with no accumulation of intermediates. The unfolding transition has its midpoint at about 6.5 M urea and appears to be slow on the time scale of electrophoresis. Folding intermediates in rapid equilibrium with the unfolded state as well as molecular forms with different electrophoretic mobility can be detected during refolding reactions. Results from Western blot analysis confirm the highly cooperative reversible urea-induced unfolding of immunoglobulin molecules and demonstrate that the unfolding transition leads to disappearance of both antigen binding and idiotypic expression, whereas the ability to interact with antibodies directed to continuous epitopes of the variable region is preserved. After progressive removal of the denaturing agent, the variable region refolds into structures that regain the functional properties of the native conformation.

Animals↗
PubMed1994

SV40 large tumor antigen associated synthetic peptides define native antigenic determinants and induce protective tumor immunity in mice.

Synthetic peptides were utilized to define antigenic determinants on simian virus 40 (SV40) large tumor antigen (T-ag). Six synthetic peptides representing predicted B-cell epitopes on SV40 T-ag were used to immunize mice to compare the humoral immune responses and ascertain the ability of the peptide preparations to induce protective tumor immunity in vivo. Anti-peptide antibodies from BALB/c and C57BL/6 mice were examined for reactivity with SV40 T-ag by various immunologic assays. Antibodies from both strains to four of the peptides recognized recombinant SV40 T-ag by ELISA. However, T-ag recognition by anti-peptide antibodies differed when assessed by Western blot. Antibodies induced by the same four peptides in BALB/c mice recognized T-ag, whereas only three of the sex peptides induced antibodies in C57BL/6 mice capable of recognizing SV40 T-ag by Western blot. Flow cytometric analysis revealed that antibodies to peptides corresponding to T-ag amino acid residues 632-652 and 690-708 from BALB/c mice were able to recognize the surface of SV40 transformed cells, whereas five of the six peptides induced surface reactive antibodies in C57BL/6 mice. More important, peptides 632-652 and 690-708 elicited a protective immune response in BALB/c mice subsequently challenged with a lethal dose of syngeneic SV40 transformed cells. However, this tumor immunity was incomplete as only 50% of the mice survived the tumor challenge. These data indicate that antibodies induced by synthetic peptides corresponding to predicted B-cell epitopes on SV40 T-ag are capable of recognizing native and denatured determinants on T-ag. Furthermore, immune responses elicited by selected peptides partially protected BALB/c mice from a lethal tumor challenge.

Amino Acid Sequence↗
PubMed1994

Fine specificity of the murine immune response to SV40 large tumour antigen utilizing synthetic peptides that define selected epitopes.

Baculovirus-derived recombinant simian virus 40 large tumour antigen (SV40 T-ag) was used to immunize BALB/c, C57Bl/6 and CB6/F1 mice and their anti-SV40 T-ag antibody responses were examined for the ability to bind synthetic peptides representing six predicted B cell epitopes on SV40 T-ag. In C57Bl/6 mice, anti-SV40 T-ag antibodies failed to bind any of the six SV40 T-ag peptides. However, the antibody responses induced in both BALB/c and CB6/F1 mice recognized synthetic peptides corresponding to two distinct epitopes (amino acids 690-708 and 660-679, respectively) associated with the carboxyl-terminal half of SV40 T-ag. In addition, murine MoAbs (BALB/c) generated to native SV40 T-ag, and previously characterized as recognizing the carboxyl-terminus of SV40 T-ag by deletion mutant analysis, also bound the synthetic peptide (residues 690-708) defining the carboxyl-terminus of SV40 T-ag. These data indicate that the antibody responses induced in BALB/c and CB6/F1 mice by immunization with baculovirus-derived recombinant SV40 T-ag are capable of recognizing sequential carboxyl-terminal epitopes on SV40 T-ag defined by peptides 690-708 and 660-679, respectively. No statistically significant differences in anti-SV40 T-ag antibody titres were observed between the three inbred mouse strains. These data suggested that the fine specificities of the anti-SV40 T-ag responses as assessed by synthetic peptide binding were different in the three inbred strains of mice examined. Finally, in vivo tumour challenge studies comparing recombinant SV40 T-ag with the two carboxyl-terminus peptide epitopes indicated that some tumour immunity was induced in BALB/c, but not CB6/F1 mice, by immunization with peptide 690-708 conjugated to a carrier protein. These studies suggest that the carboxyl-terminal region of SV40 T-ag represents a continuous sequential epitope involved in both the antibody response to SV40 T-ag and tumour immunity in BALB/c mice.

Amino Acid Sequence↗
PubMed1994

Molecular characterization of immunoglobulin variable regions from murine monoclonal antibodies specific for simian virus 40 large tumour antigen.

In this report we compare the immunoglobulin variable (IgV) region function and structure relationships of murine monoclonal antibodies that recognize simian virus 40 (SV40) large tumour antigen (T ag). Comparison of monoclonal antibody V region function is based on SV40 T ag epitope recognition and idiotype (Id) expression. Structural comparisons are based on V region gene sequence determination. The data presented herein, suggests that a high degree of homology within both the V kappa and VH regions, along with minor differences within V kappa complementarity determining regions (CDR) may result in the detection of similar SV40 T ag epitopes by the monoclonal anti-SV40 T ag preparations. The expression of a cross-reactive Id also appears to be based on the high degree of homology within both V kappa and VH regions and depends on conformational interactions imparted by both regions.

Amino Acid Sequence↗
PubMed1994

Production and characterization of murine monoclonal antibodies specific for baboon IgG heavy and light chain epitopes.

We have generated a panel of murine monoclonal antibodies (MAbs) that recognize baboon IgG epitopes. The reactivity of the MAbs with IgG from other primate species was also examined. Specificity for IgG heavy (H) or light (L) chain epitopes was determined by Western blot analysis. The H chain-specific MAbs were analyzed for IgG subclass specificity and the L chain-specific MAbs for reactivity with baboon IgM and polymeric sIgA. Finally, an ELISA was developed to demonstrate the utility of the MAbs in analysis of humoral immune responses in baboons.

Animals↗