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Biomedical subjects

J A Griffin

Publications and source records attributed to J A Griffin.

At least 19 recordsLinked to original sources
PubMed1991

Chimeric human immunodeficiency virus type 1/type 2 reverse transcriptases display reversed sensitivity to nonnucleoside analog inhibitors.

Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT), an important therapeutic target in the treatment of AIDS, is effectively inhibited by a class of nonnucleoside analog compounds that includes nevirapine (BI-RG-587) and tetrahydroimidazo[4,5,1-jk]-[1,4]benzodiazepin-2(1H)-one and -thione. We show that both tyrosine residues at positions 181 and 188 flanking the putative catalytic site of HIV-1 RT are required for sensitivity of the enzyme to these compounds. HIV-2 RT, which does not have tyrosines at these positions, is resistant to these nonnucleoside analog inhibitors. Substitution of the HIV-2 RT amino acid residues at position 181 or 188 into HIV-1 RT results in an enzyme that is resistant to these compounds while retaining sensitivity to 3'-azido-2',3'-dideoxythymidine triphosphate. HIV-2 RT substituted with amino acids 176-190 from HIV-1 RT acquires sensitivity to these nonnucleoside analog inhibitors.

Affinity Labels↗
PubMed1990

Induction of RNA-stabilized DNA conformers by transcription of an immunoglobulin switch region.

A deletion DNA rearrangement is associated with immunoglobulin class switching from IgM to IgG, IgA or IgE This recombination occurs in immunoglobulin switch regions, which are complex, highly repetitive regions of DNA. As switch regions become transcriptionally active just before switch recombination, analysis of the behaviour of these sequences during transcription could elucidate the mechanism of switch recombination. Here, we report that transcription of a supercoiled plasmid containing the murine IgA switch region (S alpha) leads to a loss of superhelical turns. The resulting series of less supercoiled plasmids is stabilized by RNA-DNA hybrids formed by the nascent RNA transcripts, which remain base-paired with their DNA templates.

Base Composition↗
PubMed1990

Evaluation of image quality in individual films of double film packets.

A total of 47 double film packets were exposed of a radiographic standard (step-wedge), a clinical model (dried skull), and a clinical situation (endodontic therapy). Dentists were employed as expert observers to compare detail and definition between front and back films within each packet. Front films (those closest to the source) had significantly superior image quality compared to back films (those further from the source). In vitro radiographic trials were significantly similar to in vivo radiographic results.

Chi-Square Distribution↗
PubMed1990

Periapical biopsy report: an analysis of over a 10-year period.

Biopsy reports of specimens from the apices of 1659 teeth physically present at the time the biopsy was performed were reviewed. The following information was recorded from each report: (a) source of specimen, (b) sex of patient, (c) age of patient, (d) location of tooth associated with lesion, and (e) diagnoses reported. Data were recorded and analyzed by chi-square analysis to note significant differences. This study found that 52% of the lesions were granulomas, 42% cysts, 2% periapical scars, and 4% other disorders. No differences were found between males and females in regard to age and location of lesions. Overall, the most common location for lesions was the maxillary anterior, followed by maxillary posterior, mandibular posterior, and finally the mandibular anterior jaw. This also was the case for all age ranges except 60 to 69 yr of age where lesions in the maxillary posterior area were most common. More granulomas were detected in all areas except in the mandibular posterior area where cysts were more common.

Adult↗
PubMed1989

Pre-pre-B cells containing only cytoplasmic B220 can be induced by dendritic cells and T cells to differentiate in vitro.

We have derived from spleens of nude mice early B lineage cells that were phenotypically compatible with a pre-pre-B cell stage of differentiation. Although these cells containing large basophilic granules had the B lymphocyte antigen B220, in the cytoplasm, they had no surface B220, no cytoplasmic or surface immunoglobulin heavy or light chains, no surface Thy-1, and no surface Ia. In addition, they appeared to have little or no heavy chain gene rearrangements, including the D to J that occurs on both chromosomes prior to the VH rearrangement that forms the code for the C mu heavy chain polypeptide. Cells at even this early stage of differentiation could be induced by DC-T to express B220 on the surface and to synthesize and then to secrete immunoglobulins. These phenotypic changes were associated with a morphologic change in the cells to a lymphoblastoid appearance. Different patterns of immunoglobulin secretion resulted when pre-pre-B cells were cocultivated with DC-T from different tissues; SP DC-T induced the secretion of only IgM, PP DC-T induced the secretion of IgM as well as IgG and IgA. The early inductive event(s) appeared to occur during cell-cell contact in aggregates of the inducing DC-T and the pre-pre-B cells.

Animals↗
PubMed1989

Many different levels of maturation and commitment are present in nontransformed cells identified serologically as pre-B cells.

The pre-B cell stage of B lymphocyte development is characterized by the presence of immunoglobulin heavy chains of the IgM isotype in the cytoplasm and no other heavy or light chains in the cytoplasm or on the surface. We established several cell lines that were identical in their serologically defined pre-B cell phenotypes and in their dependence upon interleukin 3 for growth, but which differed in their levels of cytoplasmic RNA from immunoglobulin constant region genes, in their rates of differentiation in vitro, and in the isotype profile of the antibodies that they secreted upon differentiation. The two cell lines that we have analyzed in detail, PF1 and PF1C, both contained RNA from the C mu and C delta heavy chain genes and from both the C kappa and C lambda light chain genes, even though they were not producing detectable polypeptide products from the C delta, C kappa, or C lambda genes. However, PF1C had higher levels of C gamma and C alpha RNA transcripts and differentiated in vitro under the influence of dendritic cells and T lymphocytes (DC-T) more rapidly than did PF1. If the DC-T were derived from spleens, all cell lines secreted only IgM. However, under the influence of DC-T from Peyer's patches PF1C secreted predominantly IgM, PF1 secreted primarily IgA, and a third line, PF3, secreted primarily IgG. Therefore, within the population of cells described as pre-B cells on the basis of their immunoglobulin gene polypeptide products, there are subpopulations that probably represent different levels of maturation and different levels of commitment to particular pathways of B lymphocyte development.

Animals↗
PubMed1988

VH gene expression by nontransformed pre-B cells upon differentiation in vitro.

Mice have more than 1000 VH gene segments, and each pre-B cell must choose a single one for rearrangement to encode the V portion of the antibody H chain. Presumably, all or most of the functional VH gene segments must be chosen by the population of B lymphocytes if the organism is to express the diversity that is observed in the immune system. Control of the selection of a VH gene segment for expression is not understood. We have found that the members of the VH gene family closest to the constant genes, the 7183 family, are transcribed in a manner that is specific for the stage of B cell development after pre-B cells derived from spleens of 6- to 8-wk-old nude mice are induced to differentiate in vitro by a mixture of dendritic cells and mitogen-activated T lymphocytes (DC-T). DC-T from spleens and lymph nodes induce transient high levels of synthesis of RNA from the 7183 VH family, whereas DC-T from Peyer's patches of mice of the same age as those from which spleen and lymph node DC-T were prepared did not induce the expression of RNA from that gene family. Spleen and Peyer's patch DC-T induce secretion of similar total amounts of antibody. Therefore, the RNA synthesis from members of at least one VH gene family is specific both for the lymphoid tissue in which B cell differentiation occurs and for the developmental stage of the B lymphoid cells.

Animals↗
PubMed1988

Non-B right-handed DNA conformations of homopurine.homopyrimidine sequences in the murine immunoglobulin C alpha switch region.

The switch region of IgA immunoglobulin in mice cloned into a recombinant plasmid contains a supercoil-dependent S1 nuclease hypersensitive site, indicative of a non-B-DNA secondary structure. This site maps to the (AGGAG)28 direct repeat (DR2) of the alpha switch region and appears at a negative superhelical density of greater than 0.02. Studies with P1 nuclease and bromoacetaldehyde indicate that this structure is also present at neutral pH. S1 nuclease sensitivity is retained for the shorter repeat (AGGAG)6GA in a recombinant plasmid but is not seen for the repeat (CTGAG)6, corresponding to the DR1 repeat of the alpha switch region, or in a sequence corresponding to a portion of the consensus sequence which contains a short stretch of alternating pyrine-pyrimidine residues. Fine mapping of the (AGGAG)6GA and flanking sequences with dimethyl sulfate, bromoacetaldehyde, osmium tetroxide, and diethyl pyrocarbonate reveals an asymmetric pattern of modification dependent on both pH and supercoiling. Two-dimensional gel electrophoresis at low pH shows the relaxation of 3 superhelical turns on formation of this structure by the (AGGAG)6GA repeat. These results are most consistent with the formation of an intramolecular triple-strand.

Animals↗
PubMed1987

A novel pre-B cell precursor: phenotypic characterization and differentiation induction by dendritic cell-T cell mixtures.

We have derived from spleens of nude mice early B lineage cell lines that are dependent upon factor(s) in supernatants of the WEHI-3B cell line. The cells are phenotypically at the pre-pre-B cell stage of differentiation. These cells are large and have basophilic granules in the cytoplasm. They are negative for cytoplasmic and surface immunoglobulin heavy or light chain, surface B220, surface Thy 1.2 and surface Ia but are positive for cytoplasmic B220. These cells at this early stage of maturation can be induced by mixtures of DC-T cells to express B220 on their surfaces and later to synthesize and ultimately to secrete immunoglobulin. These events are associated with morphologic changes in the cells to a lymphoblastoid appearance. Different pattern of immunoglobulin secretion were induced by DC-T from different tissues. The inductive event appears to occur during cell contact of pre-pre-B cells with the inducing DC-T cell mixture, but it does not appear to require IL-3. These data indicate that DC-T cell mixtures can provide signals for B cell differentiation at pre-pre-B cell stage as well as pre-B and mature B cell stage.

Animals↗
PubMed1987

Immunoadsorbent purification of carcinoembryonic antigen using a monoclonal antibody: a direct comparison with a conventional method.

Carcinoembryonic antigen (CEA) has been extracted and purified by two different methods from a single aqueous homogenate of liver metastases from a primary adenocarcinoma of the colon, thus enabling direct comparison to be made between a conventional technique using gel filtration and a monoclonal antibody immunoadsorbent method. Yield, immunoreactivity and purity have been determined in both cases by direct weighing, enzyme-linked immunoadsorbent assay, radioimmunoassay and SDS-polyacrylamide gel electrophoresis. Reactivity of the antigen with monoclonal and polyclonal anti-CEA antibodies recognising different epitopes was analysed by Western blotting. There appears to be no significant difference in immunoreactivity or purity by these criteria, but the immunoadsorbent method gave a higher yield of CEA for far less expenditure of time and effort. A variant of CEA with a lower molecular weight was also identified in both preparations.

Antibodies, Monoclonal↗
PubMed1986

VH gene expression is restricted in anti-IgG antibodies from MRL autoimmune mice.

Antibodies directed against IgG and DNA are found in the sera of autoimmune MRL/Mp lpr/lpr mice. Little is known of the molecular mechanisms underlying expression of such autoantibodies. We have investigated the binding diversity and pattern of VH gene expression in a panel of murine anti-IgG antibodies. We constructed eight hybridoma clones secreting IgM antibodies that bound to mouse IgG by using spleen cells from MRL/Mp lpr/lpr mice varying in age from 4 to 15 wk; one clone was derived from a 32-wk-old MRL +/+ mouse. The monoclonal IgM products exhibited varying binding specificities for intact mouse IgG, fragments of mouse IgG [Fc, Fab, (Fab')2], and heterologous IgG. Two of these antibodies crossreacted with B and/or Z DNA. Probes from seven of eight identified mouse VH gene families (7183, S107, Q52, J558, J606, 36-60, and 3609) were hybridized under high-stringency conditions with cytoplasmic RNA blots from each clone. Six clones hybridized only with the probe from the five-member 36-60 family. The remaining three clones crosshybridized with the 36-60 probe and the probe from the 60 member J558 family, perhaps reflecting somatic mutation from the original germline VH gene resulting in recognition by a probe from another family, in addition to the probe from the original germline family. Our results indicate that spontaneous MRL lpr/lpr anti-IgG antibodies are encoded predominantly by the 36-60 VH gene family and imply a nonrandom selection of this VH gene family in the production of these antibodies.

Animals↗
PubMed1986

Different pathways of differentiation of pre-B cell lines are induced by dendritic cells and T cells from different lymphoid tissues.

Nontransformed pre-B cells were induced to differentiate in vitro along several different but predictable pathways with only dendritic cells (DC) and concanavalin A-stimulated T lymphocytes. DC-T from spleen induced secretion of only IgM, whereas DC-T from Peyer's patches induced high levels of IgA and intermediate levels of IgM and IgG. Both the isotype of antibody secreted and the extent of pre-B cell differentiation were determined by the lymphoid tissue source of DC, not of T cells. The pre-B cells synthesizing detectable levels of only the IgM heavy chain had cytoplasmic RNA transcripts from both light chain constant region genes and from the entire length of the heavy chain constant locus. In cells secreting IgM there were deletions in the DNA flanking the Cmu coding region.

Animals↗
PubMed1985

Diversity in the germline antibody repertoire. Molecular evolution of the T15 VN gene family.

The T15 heavy chain variable region (VH) gene family in BALB/c mice includes four elements each greater than 88% homologous with the other. One of these elements, V1, encodes virtually all of the VH regions in BALB/c antiphosphorylcholine antibodies, while another element, V3, is a pseudogene and cannot be transcribed or translated. We have examined the structural features of this VH gene family in other mouse strains and, in particular, have cloned and sequenced the alleles of these gene segments present in B10.P mice. Each of the four B10.P sequences can be matched with its allelic counterpart in BALB/c mice. This represents the first successful analysis of allelism in antibody variable region gene segments. The V1B10.P allele, like its BALB/c counterpart, encodes most of the known phosphorylcholine binding heavy chains from C37BL/6 mice. Similarly, the V3B10.P gene segment is a pseudogene like V3BALB, although only two of four abnormalities present in the BALB/c allele are also present in the B10.P allele. Careful analysis of the specific substitutions observed in the T15 VH gene family suggests that environmental selection for functional combining regions contributes significantly to the pattern of variation in the germline antibody repertoire. In addition, evidence is presented supporting frequent gene conversion events in the divergence of antibody genes.

Alleles↗
PubMed1985

Immunoglobulin gene rearrangements and deletions in human Epstein-Barr virus-transformed cell lines producing different IgG and IgA subclasses.

During differentiation B lymphocytes may switch from the expression of surface IgM to the synthesis of IgG, IgA, or IgE isotypes by using a different heavy chain constant region (CH) gene. The molecular mechanisms by which switching occurs remain controversial. Rearrangements and deletions of CH genes 5' to the expressed gene have often been observed in the mouse and, more recently, in human cells that have switched isotypes. We have used human JH, C micro, C gamma, and C alpha probes to examine the extent of the deletions and rearrangements in clones of Epstein-Barr virus-transformed human cells that produce IgG1, IgG3, IgG4, or IgA1. Though deletions of CH genes 5' to the expressed CH gene were consistently observed, the rearrangement process appeared to be highly variable for the nonproductive CH gene locus: deletion or persistence of 5' CH genes, combinations of deletion and duplication of 5' genes, and deletions extending to 3' CH genes. Our results reveal an unexpected lack of specificity in the DNA deletions in cells that have undergone isotype switching.

B-Lymphocytes↗
PubMed1985

Recombinant DNA--potential for gene therapy.

Certainly, if progress in recombinant DNA technology continues at its present rate, we have every reason to expect many more major breakthroughs in the diagnosis and treatment of human disease. Much of the progress from which we already benefit is not the increased understanding of just the molecular basis of genetic disease, but also of the molecular mechanisms of viral, bacterial, and parasite pathogenicity. Exploiting cloned antigens from pathogens to make vaccines is an ever expanding approach to preventive medicine. The other realm from which we already benefit is that in which we have bacteria produce large quantities of product from normal cloned genes, such as insulin, for treatment of patients deficient in that gene product. Although we cannot expect to eliminate some, or even treat all, genetic disease within the foreseeable future, it is quite clear that research in the area of genetic engineering has vast potential for improving the conditions of mankind.

Bacteriophages↗