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Biomedical subjects

M B Nelson

Publications and source records attributed to M B Nelson.

At least 19 recordsLinked to original sources
PubMed2001

IgG-recognizing shed tumor-associated antigens can promote tumor invasion and metastasis.

Tumors secreting glycoproteins that act as tumor-associated antigens have been described as highly invasive and metastatic. In this study, the consequences of the humoral immune response (HIR) against these antigens were investigated. Using an in vitro model of tumor cell invasion, results indicated that the invasiveness of tumor cells secreting antigenic secreted/shed tumor glycoproteins (STGP) increases in the presence of specific anti-STGP IgG, polymorphonuclear cells and monocytes. This in vitro model showed that the coincidental presence in the matrix of both STGP and specific anti-STGP IgG increases the local release of IL-1beta, IL-6 and vascular endothelial growth factor (VEGF) by stromal cells, but not by tumor cells. Using an in vivo model, the experiments show that immune-competent mice develop an anti-tumor HIR with anti-STGP IgG production. In this model, tumor growth was increased in parallel with the serum concentration of specific anti-STGP IgG. In athymic nude (nu/nu)-beige mice the same trend was observed, suggesting a T-cell-independent tumor-promoting effect induced by anti-STGP IgG. Tumor histology showed intense infiltration of IgG-positive plasma cells and lymphocytes. A severe combined immunodeficient-beige mouse-based in vivo model of tumors, experimentally infiltrated with monoclonal IgG plasmocytoma cells, showed that only specific anti-STGP-IgG-secreting cells could exacerbate tumor invasion, angiogenesis and metastasis. These results suggest that tumors shedding/secreting antigenic STGP can induce a host IgG immune response that can promote invasion and metastasis by inducing tumor infiltrating stromal cells to release proinflammatory cytokines and VEGF.

Animals↗
PubMed2001

In vivo imaging of light-emitting probes.

In vivo imaging of cells tagged with light-emitting probes, such as firefly luciferase or fluorescent proteins, is a powerful technology that enables a wide range of biological studies in small research animals. Reporters with emission in the red to infrared (>600 nm) are preferred due to the low absorption in tissue at these wavelengths. Modeling of photon diffusion through tissue indicates that bioluminescent cell counts as low as a few hundred can be detected subcutaneously, while approximately 10(6) cells are required to detect signals at approximately 2 cm depth in tissue. Signal-to-noise estimates show that cooled back-thinned integrating charge coupled devices (CCDs) are preferred to image-intensified CCDs for this application, mainly due to their high quantum efficiency (approximately 85%) at wavelengths >600 nm where tissue absorption is low. Instrumentation for in vivo imaging developed at Xenogen is described and several examples of images of mice with bioluminescent cells are presented.

Animals↗
PubMed2000

B lymphocyte pathology in human colorectal cancer. Experimental and clinical therapeutic effects of partial B cell depletion.

Accumulating data are showing that the humoral immune response against tumors could favor tumor progression. However, no B lymphocyte pathology has been reported in cancer. Using anti-IgM Ab we nonspecifically depleted B cells in tumor-bearing mice, a treatment that resulted in significant reduction of tumor burden. We analyzed the B lymphocyte phenotype of abdominal lymph nodes and peripheral blood from advanced colon cancer patients by flow cytometry, and compared the B cell phenotype with that found in samples from normal donors. In both lymph nodes and peripheral blood of cancer patients, abnormal populations of B lymphocytes appeared that express an increased CD21 and/or sTn antigens on their cell surface. All patients showed a reduction of CD19+ cells. In a limited clinical test, we analyzed the effects of a partial B cell depletion with Rituximab. The treated patients did not develop any side-effects; the CD21-hyperpositive lymphocytes were reduced, but the proportion of sTn-positive lymphocytes remained unaffected. Apparent reduction of the tumor burden was reported in 50% of the patients when the treatment was ended.

Animals↗
PubMed1999

Promotion of tumor invasion by cooperation of granulocytes and macrophages activated by anti-tumor antibodies.

We investigated the potential role of anti-tumor antibodies and tumor antigens in the formation of immune complexes which promote matrix degradation and angiogenesis. B-cell deficient or B-cell depleted mice showed a reduction in tumor invasion and metastasis. In vitro invasion assays and in vivo models of metastasis showed that anti-sTn antibodies and sTn tumor antigens form complexes which induce granulocytes and macrophages together to mediate tumor invasion and metastasis by processes including extracellular matrix degradation and angiogenesis. These results suggest the existence of a tumor promoting role of a B-cell immune response induced by shed tumor associated antigens of solid, nonlymphoid tumors.

Animals↗
PubMed1999

The R-type pyocin of Pseudomonas aeruginosa C is a bacteriophage tail-like particle that contains single-stranded DNA.

Pseudomonas aeruginosa R-type pyocin particles have been described as bacteriocins that resemble bacteriophage tail-like structures. Because of their unusual structure, we reexamined whether they contained nucleic acids. Our data indicated that pyocin particles isolated from P. aeruginosa C (pyocin C) contain DNA. Probes generated from this DNA by the random-primer extension method hybridized to distinct bands in restriction endonuclease-digested P. aeruginosa C genomic DNA. These probes also hybridized to genomic DNA from 6 of 18 P. aeruginosa strains that produced R-type pyocins. Asymmetric PCR, complementary oligonucleotide hybridization, and electron microscopy indicated that pyocin C particles contained closed circular single-stranded DNA, approximately 4.0 kb in length. Examination of total intracellular DNA from mitomycin C-induced cultures revealed the presence of two extrachromosomal DNA molecules, a double-stranded molecule and a single-stranded molecule, which hybridized to pyocin DNA. Sequence analysis of 7,480 nucleotides of P. aeruginosa C chromosomal DNA containing the pyocin DNA indicated the presence of pyocin open reading frames with similarities to open reading frames from filamentous phages and cryptic phage elements. We did not observe any similarities to known phage structural proteins or previously characterized pseudomonal prt genes expressing R-type pyocin structural proteins. These studies demonstrate that pyocin particles from P. aeruginosa C are defective phages that contain a novel closed circular single-stranded DNA and that this DNA was derived from the chromosome of P. aeruginosa C.

Amino Acid Sequence↗
PubMed1999

Young adult survivors of childhood cancer: attending to emerging medical and psychosocial needs.

As survival rates for childhood cancer have increased during the past three decades, a significant population of young adult survivors has emerged. Medical late effects of particular concern to this population, including reproductive issues, osteoporosis, cardiotoxicity, hepatitis C, and second malignancies are discussed. Educational, occupational, insurance, and other significant psychosocial sequelae are addressed. Models for medical and psychosocial follow-up with this population are described, and a psycho-educational intervention model, implemented by members of the Long-Term Information, Follow-up, and Evaluation (LIFE) team at Children's Hospital Los Angeles, is presented for consideration. Future research and clinical challenges are discussed.

Adult↗
PubMed1998

First results for resetting the antitumor immune response by immune corrective surgery in colon cancer.

BACKGROUND: A critical step for cancer recurrence is the failure of the cellular immune response. It is suspected that chronic humoral immune responses against some tumor-associated antigens (TAA) can contribute to that failure. METHODS: In this study, we tested the ability of an immune corrective surgical procedure to prevent recurrences of colon cancer in stages I, II, and III. Radiolabeled anti-TAG antibodies injected intravenously become concentrated on TAG-72 immune complexes presented by follicular dendritic cells, which are responsible for the persistent humoral response against TAG-72 TAA. Using a hand-held gamma probe, we can intraoperatively detect and remove lymph nodes involved in TAG-72 presentation. By removing these lymph nodes, together with the tumor tissue, presentation and source of TAG-72 are drastically reduced. RESULTS: The impact of this TAA suppression on the tumor recurrence process is analyzed in a sample of 24 patients. The immune corrective surgical procedure did not increase morbidity. Five years after surgery the following were disease free: 5 of 5 stage I, 6 of 6 stage II, and 10 of 13 stage III. The global survival of this group was 87.5%. Compared with the standard surgical treatment of colon cancer (58% survival for the same stages), this surgical immune corrective procedure introduces a statistically significant improvement of 29% (P <0.001). CONCLUSIONS: The surgical removal of lymph nodes involved in the persistent humoral immune response against TAA has an important beneficial impact on colon cancer treatment.

Antibodies, Neoplasm↗
PubMed1992

Outer membrane protein P6 of Haemophilus influenzae binds to its own gene.

Outer membrane protein P6 is an important antigen expressed on the surface of all strains of Haemophilus influenzae. The predicted amino acid sequence of P6 contains a region of alpha helices that shares sequence identity with a family of helix-turn-helix DNA-binding proteins. A search for sequence-specific binding sites that resemble an operator region within the gene revealed a sequence with striking homology to the consensus operator sequence for lambda Cro and repressor. To test the hypothesis that P6 binds its own gene, purified P6 on nitrocellulose was probed with plasmid DNA containing the P6 gene. P6 bound the P6 gene in this Southwestern blot assay. To further test the observation, gel shift analysis was performed. Gel shift assays using a P6-specific monoclonal antibody demonstrated that P6 in crude cell extracts binds to the region of the gene containing the putative binding site. Competition with a synthetic oligonucleotide corresponding to the putative binding site inhibited binding of P6 to the P6 gene on nitrocellulose and in the gel shift assay. In addition, this oligonucleotide bound directly to P6 on nitrocellulose. Finally, DNase footprinting confirmed that P6 bound specifically to the same region of the P6 gene. These results indicate that P6 binds to a sequence-specific site within its own gene, suggesting that P6 regulates its own expression. This represents the first example of a Gram-negative outer membrane protein binding to its own gene and has potentially important implications as a mechanism for regulation of expression of outer membrane antigens.

Amino Acid Sequence↗
PubMed1992

Mutations in the principal neutralization determinant of human immunodeficiency virus type 1 affect syncytium formation, virus infectivity, growth kinetics, and neutralization.

The principal neutralization determinant (PND) of human immunodeficiency virus type 1 envelope glycoprotein gp120 contains a conserved GPG sequence. The effects of a 29-amino-acid deletion of most of the PND, a 3-amino-acid deletion in the GPG sequence, and 16 single-amino-acid substitutions in the GPG sequence were determined in a transient expression assay. All mutant envelope glycoproteins were expressed at levels comparable to that of the wild-type envelope, and mutations in the GPG sequence did not affect processing to gp120 or, except for the 29-amino-acid deletion, binding to CD4. Of all of the mutants, only the GHG and GFG mutants induced formation of syncytia similar in size and number to those induced by the wild-type envelope. When the envelope expression level was increased 10-fold or more, several additional mutants (APG, GAG, GSG, GQG, GVG, and GPF) also induced syncytium formation. Transfection with infectious proviral molecular clones containing the GHG, GFG, APG, GAG, GSG, or GPF mutations induced production of viral particles; however, only the GPG, GHG, and GFG viruses produced active infections in CD4-bearing cells. Furthermore, whereas the wild-type virus was efficiently neutralized by PND polyclonal and monoclonal antibodies, the GHG- and GFG-containing viruses were not. These results show that mutations in the GPG sequence found within the PND do not affect envelope expression and do not significantly affect CD4 binding or production of viral particles but that they do affect the ability of the envelope to induce syncytia and those of the viral particles to infect CD4 cells and be neutralized by PND antibodies.

Amino Acid Sequence↗
PubMed1991

Molecular conservation of the P6 outer membrane protein among strains of Haemophilus influenzae: analysis of antigenic determinants, gene sequences, and restriction fragment length polymorphisms.

Infections caused by Haemophilus influenzae are a major worldwide health problem. In particular, nontypeable strains of H. influenzae are a common cause of otitis media in infants and children. A vaccine to prevent these infections would result in the prevention of substantial morbidity and cost savings. A problem in identifying an appropriate vaccine antigen has been the enormous antigenic heterogeneity among nontypeable strains of H. influenzae. The present study was undertaken to characterize the conservation of the P6 outer membrane protein (approximately 16,000 daltons) among strains of H. influenzae. A total of 20 type b strains and 20 nontypeable strains of diverse geographic and clinical origins was studied. Three approaches were taken. (i) Antigenic determinants recognized by monoclonal and polyclonal antibodies were present on P6 in all 40 strains tested. The molecular weight of P6 was identical in all strains. (ii) Comparison of the DNA sequences of the P6 genes from three epidemiologically and serologically unrelated strains demonstrated 100% homology at the amino acid level and 97 to 99% homology at the nucleotide level. (iii) Restriction fragment length polymorphism analysis demonstrated that the P6 gene and flanking sequences were highly conserved among all strains. These three independent series of experiments indicated that the P6 protein is highly conserved among strains of H. influenzae. P6 should receive serious consideration for inclusion in a vaccine to prevent infections caused by nontypeable H. influenzae.

Amino Acid Sequence↗
PubMed1988

Cloning and sequencing of Haemophilus influenzae outer membrane protein P6.

P6, a 16,600-dalton protein present in the outer membranes of both typeable and nontypeable strains, may be an important antigen in immunity to Haemophilus influenzae. The gene encoding P6 of a nontypeable strain of H. influenzae was cloned by using bacteriophage lambda gt11. Four recombinant phages were detected by screening plaques with monoclonal antibodies and a polyclonal antiserum. One recombinant phage, clone O, produced a full-length gene product which was expressed at a high yield. The DNA insert contained within this phage was cloned into the plasmid vector pUC18 to create the recombinant plasmid pBUD1. An Escherichia coli transformant containing this plasmid produced a protein which had an apparent molecular weight identical to that of H. influenzae P6, as determined by Western blot (immunoblot) analyses. Expression of the P6 polypeptide by both clone 0 and the transformant was independent of induction of the lac operon by isopropyl-beta-D-thiogalactopyranoside, suggesting that transcription was from the promoter of the P6 gene. Immunoelectron microscopy using a monoclonal antibody with specificity for a P6 surface epitope detected the presence of P6 on the surface of the transformant. The insert in pBUD1 was cut down in size to approximately 800 base pairs. The resultant plasmid, pBUD5, also coded for a full-length gene product. DNA sequence analysis revealed that the P6 gene contains transcriptional and translational sequences resembling those recognized in E. coli and a signal sequence characteristic of procaryotic membrane proteins. In addition, the carboxy terminus of this signal sequence shares homology with a common sequence found in bacterial lipoproteins, suggesting that P6 is a lipoprotein. Posttranslational proteolytic cleavage of the signal sequence would result in a protein composed of 134 amino acids.

Amino Acid Sequence↗
PubMed1986

Antigenic characterization of the P6 protein of nontypable Haemophilus influenzae.

The purpose of this study was to characterize the degree of antigenic heterogeneity or conservation of a 16,600-dalton outer membrane protein (P6) among strains of nontypable Haemophilus influenzae. Immunization of rabbits with P6 isolated from individual strains resulted in antibody to P6 of all 25 strains tested. The titers of antibody in the sera were similar among the strains. Whole organisms of two strains were used to immunize rabbits, and antibodies were produced to P6 of all strains tested. Monoclonal antibodies developed to P6 from mice immunized with whole cells of three different strains recognized determinants on P6 of all 25 strains tested. Finally, pooled normal human serum contained antibodies to P6 of all 25 strains assayed. These studies indicate that P6 is a highly conserved antigen on the outer membrane of nontypable H. influenzae.

Animals↗
PubMed1986

Identification of a 16,600-dalton outer membrane protein on nontypeable Haemophilus influenzae as a target for human serum bactericidal antibody.

A 16,600-D outer membrane protein is present in all strains of Haemophilus influenzae and antibodies to this protein are present in human serum. This study was designed to assess the role of this outer membrane protein (P6) in nontypeable H. influenzae as a target for human serum bactericidal antibody. P6 was isolated and coupled to an affinity column. Depleting normal human serum of antibodies to P6 by affinity chromatography resulted in reduced bactericidal activity of that serum for nontypeable H. influenzae. Immunopurified antibodies to P6 from human serum were bactericidal. Finally, preincubation of bacteria with a monoclonal antibody that recognizes a surface epitope on P6, inhibited human serum bactericidal killing. Taken together, these experiments establish that P6 is a target for human bactericidal antibodies. This observation provides evidence that P6 plays a potentially important role in human immunity to infection by nontypeable H. influenzae.

Antibodies, Bacterial↗
PubMed1985

Identification of a specific epitope of Haemophilus influenzae on a 16,600-dalton outer membrane protein.

A mouse monoclonal antibody that recognizes an epitope on a 16,600-dalton outer membrane protein was developed to nontypable Haemophilus influenzae. This epitope was present on all 115 isolates of H. influenzae tested, including typable and nontypable strains. Screening of 89 strains of other bacteria demonstrated that this epitope is a highly specific marker for H. influenzae because the epitope was absent in virtually all other bacterial species tested. Western blot assays were performed with two normal human serum samples and convalescent-phase serum from an adult with bacteremia due to nontypable H. influenzae. Antibody to the 16,600-dalton outer membrane protein was present in all three human serum samples.

Animals↗