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Biomedical subjects

M Bennett

Publications and source records attributed to M Bennett.

At least 73 records · Page 4Linked to original sources
PubMed1995

Pleotropic effects of dietary DHEA.

We present data pertaining to some of the in vivo effects associated with dietary DHEA administration to mice and rats. Dietary DHEA leads to: (1) decrease in body weight gain; (2) relative increases in liver weight; (3) liver color change; (4) induction of hepatic peroxisomal enzymes; (5) proliferation of hepatic peroxisomes with increased cross-sectional area; (6) decreased hepatic mitochondrial cross-sectional area; (7) elevated levels of hepatic cytosolic malic enzyme; (8) slight decreases, significant decreases, or significant increases in serum triglyceride levels, depending on mouse strain; (9) increases in total serum cholesterol levels; (10) significant decreases in the hepatic rates of fatty acid synthesis; (11) significant increases in the hepatic rates of cholesterol synthesis; (12) decreases in both protein content and specific activity of hepatic mitochondrial carbamoyl phosphate synthetase-I without concomitant changes in serum urea nitrogen; (13) induction of glutathione S-transferase activity in liver; (14) decrease in hepatic endogenous protein phosphorylation; (15) increase in hepatic AMPase and GTPase activities; (16) formation of 5-androstene-3 beta,17 beta-diol as a major metabolite of DHEA by subcellular fractions of liver, which is reflected in serum and tissue levels; and (17) reduction in serum prolactin levels.

Animals↗
PubMed1995

Rapid rejection of H2k and H2k/b bone marrow cell grafts by CD8+ T cells and NK cells in irradiated mice.

Acute rejection of transplanted bone marrow cell (BMC) grafts can occur within 48 h in unsensitized, lethally irradiated mice, and NK cells have been implicated as the effector cells. Recently, we observed that both CD8+ TCR-alpha beta+ T and NK cells of irradiated mice could rapidly reject allogeneic lymph node cell grafts. In this study, we evaluated the ability of NK and CD8+ T cells to mediate rejection of H2k or H2k/b BMC grafts by pretreating groups of mice with depleting mAbs. H2k BMC were transplanted into syngeneic, B6 (H2b), BALB/c (H2d), NZB (H2d), and (NZB x B6)F1 (NZB6F1, H2d/b) hosts. Proliferation measured 5 days after cell transfer indicated that syngeneic, B6, and BALB/c hosts accepted H2k BMC grafts. However, CD8+ T cells from NZB and poly I:C-treated BALB/c hosts, and NK cells from poly I:C-treated NZB6F1, hosts, rejected H2k BMC grafts. In NZB6F1 hosts, there was an added effect of anti-TCR-alpha beta and anti-NK1.1 mAbs. It is possible that T and NK cells cooperate in rejecting H2k or H2k/b BMC grafts in certain hosts. Transplantation of H2k/b, but not H2k/d, BMC into similar recipients had the same fate as H2k BMC. Thus, certain CD8+ T cells may share a similar recognition system with NK cells.

Animals↗
PubMed1995

NK cell subsets in the regulation of murine hematopoiesis. I. 5E6+ NK cells promote hematopoietic growth in H-2d strain mice.

NK cells are able to reject bone marrow allografts in lethally irradiated mice. 5E6 is a marker expressed on a subset of NK cells that is responsible for the rejection of H-2d homozygous bone marrow cell (BMC) allografts. This suggests that the 5E6+ NK cell subset somehow recognizes and is deleterious for H-2d BMC. However, unlike Ly-49+ NK cells, 5E6+ cells are not deleted or even down-regulated in H-2d-homozygous mice. We wanted to determine, therefore, the role of the 5E6+ and 5E6- NK cell subsets in the normal physiologic regulation of hematopoiesis in H-2d strains of mice. Surprisingly, both in vivo depletion studies of normal mice and studies in which the subsets were purified and cultured with syngeneic BMC in vitro demonstrated that in H-2d mice, the 5E6+ subset of NK cells did not inhibit, but instead promoted, growth of H-2d BMC. Depletion of the 5E6+ subset also resulted in decreased marrow engraftment after syngeneic bone marrow transplantation in H-2d mice. Analysis of the cell culture supernatants of the purified subsets indicated that the functional effects of the subsets on hematopoiesis correlated with the relative amounts of hematopoietic growth-promoting cytokines produced by the NK cells. These results demonstrate that physiologically relevant subsets of NK cells exist that are involved in the homeostatic regulation of hematopoiesis and that they can be distinguished on the basis of 5E6 expression.

Animals↗
PubMed1995

Cloning and functional characteristics of murine large granular lymphocyte-1: a member of the Ly-49 gene family (Ly-49G2)

Large granular lymphocyte (LGL) 1 is a cell surface glycoprotein expressed on a subset (50%) of C57BL/6 natural killer (NK) cells. Immunoprecipitation experiments reveal that the LGL-1 protein exists as a disulfide-linked 40-kD homodimer. Functional studies of LGL-1+ cells indicate that selected H-2d target cells are not lysed efficiently by these interleukin (IL)-2-cultured NK cells. These findings suggested that LGL-1 may be a member of the Ly-49 gene family. Here we report the molecular cloning of the LGL-1 cDNA from a severe combined immunodeficient-adherent lymphokine-activated killer cell library transfected into Cos-7 cells and find LGL-1 to be homologous to the Ly-49 gene at both the nucleotide (85%) and amino acid levels (73%). Sequencing of our LGL-1 cDNA has revealed it to be nearly identical to the Ly-49G2 cDNA recently isolated by cross-hybridization with an Ly-49 probe. LGL-1 represents a type II transmembrane protein of 267 amino acids with its carboxyl end exposed extracellularly. The LGL-1 protein contains 11 highly conserved cysteine residues and a 25-amino acid transmembrane region. Southern blot analysis demonstrates that there are a number of homologous genes in mouse DNA that hybridize strongly to LGL-1. Northern analyses using poly A+ RNA from LGL-1+ NK cells indicate that LGL-1 is expressed as a 1.4 kb mRNA. Two-color flow cytometry analysis (FCA) of C57BL/6 splenic NK cells demonstrates that LGL-1 and Ly-49 label overlapping subsets of cells. FCA identifies four subsets of NK cells as defined by LGL-1 versus Ly-49 staining. We have sorted these individual subsets, expanded them in IL-2, and performed cytotoxicity experiments to determine their target cell profiles in relation to class I expression. Results of these studies are complex, but indicate that Ly-49 may not be the only molecule that recognizes class I as an inhibitory signal for cytotoxicity. LGL-1+ cells also fail to lyse several H-2d-expressing tumor targets and concanavalin A lymphoblasts from BALB/c but not C57BL/6 mice. This inhibition of lysis by LGL-1+ NK cells is negated by addition of monoclonal antibody (mAb) 4D11 that recognizes the LGL-1 protein. When mAbs to the class I molecules H-2Dd and H-2Ld (alpha 1 alpha 2 domains only) are added to cytotoxicity assays, LGL-1+ cells lyse H-2d targets very effectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Cloning and characterization of 5E6(Ly-49C), a receptor molecule expressed on a subset of murine natural killer cells.

5E6 is a cell surface molecule expressed on a subpopulation of murine natural killer (NK) cells that are involved in the specific rejection of H-2d or H-2f (hemopoietic histocompatibility determinant 2) bone marrow cell grafts. Here, we isolated and cloned the gene encoding 5E6 and determined the nucleotide sequence of the cDNA. 5E6 is nearly identical to Ly-49C; the deduced amino acid sequence reveals a polypeptide of 266 amino acids with a molecular weight of 31,284 that contains multiple cysteine residues to explain its disulfide-linked homodimer structure and five potential N-linked glycosylation sites. 5E6 is a type II integral membrane protein with an extracellular carbohydrate recognition domain characteristic of C-type (Ca(2+)-dependent) animal lectins. Chromosomal mapping indicates that 5E6 is located within the NK gene complex on chromosome 6. The sequence of 5E6 mRNA and the degree of glycosylation of 5E6 protein are under genetic control. Immunoprecipitation before removal of N-linked sugars reveals different size molecules. There are several nucleotide differences among BALB/c, B6, and NZB mRNAs; however, none of them would be expected to affect N-glycosylation. Of particular interest are two findings: (a) BALB/c, B6, and (BALB/c x B6)F1 5E6 reduced molecules are approximately 65, 54, and 54 kD, and (b) the cDNA sequence of (BALB/c x B6)F1 is identical to B6. Thus, there appears to be allelic exclusion of 5E6 expression that may be related to the ability of F1 hybrid mice to reject parental H-2d bone marrow cell grafts.

Amino Acid Sequence↗
PubMed1995

Bone marrow cell transplants involving donors and hosts with haplotypes derived from spretus mice.

Intra-H2 recombinant inbred mice derived from matings between B10 (H2b) and B10.SP2 (H2sp2) mice, with an H2 haplotype derived from Mus spretus, have been used to map genes at H2. Recombinants 10115, 10484, R40, 9347, and 9950 were used as donors or hosts in bone marrow cell (BMC) transplants in irradiated mice. From previous studies of Mus musculus mice, the antigens (Ag) on BMC appear to be inherited recessively. The mechanisms offered include codominant inheritance of transacting genes that regulate expression of BMC Ag (Hh hypothesis) and codominant inheritance of class I Ag motifs capable of sending "negative signals" to effector natural killer (NK) cells (missing self hypothesis). Our results indicate that stem cell donors that express the same class I Ag, but differ at genes between Bat2 and Tnfa in the H2-S/D interval, can differ in immunogenicity of transplanted stem cells. The structural gene for the H2sp2 Ag appears to map telomeric of Bat2 and is codominantly inherited. An H2b gene capable of inhibiting expression of the H2sp2 Ag (or contributing to class I motifs capable of inhibiting NK cell mediated lysis of H2sp2 BMC) maps in the Bat2/Tnfa gene segment, but requires homozygosity for this function and may require the H2-Db gene as well. Although H2sp2 hosts reject H2b BMC, hosts (10115, 10484, R40, and 9347 strain) that are H2b in the centromeric, and H2sp2 in the telomeric, portion of H2 accept H2b BMC grafts. These two observations have not been made with haplotypes entirely of Mus musculus origin. The data do not support the Hh hypothesis, and are consistent with the missing self hypothesis only if the gene (requiring homozygozity for function) in the Bat2/Tnfa region codes for a particular protein or peptide that associates with Db to generate a "protective motif."

Animals↗
PubMed1995

Hybrid and allogeneic resistance to T cell grafts mediated by murine NK and CD8+ T cells.

Lethally irradiated mice can reject H-2 allogeneic or parental strain stem cells in bone marrow cell (BMC) grafts within 48 h after transplantation. This rapid rejection of BMC grafts occurs without prior sensitization and is mediated by NK1.1+ NK cells. One hypothesis to account for the ability of host NK cells to mediate acute rejection of allogenic and parental stem cells is that these effector cells recognize hemopoietic histocompatibility (Hh-1) Ags on the donor stem cells. T cells present in the donor BMC can prevent NK cell-mediated rejection. However, lethally irradiated mice can also reject T cells present in lymph node cell preparations that respond to alloantigens of the host. T cell grafts from H-2k/Hh-1k, H-2r/Hh-1null, and H-2ia1/Hh-1null donors were rejected by CD8+ NK1.1- T cells. Ags other than Hh-1 Ags appeared to be recognized by these CD8+ T cells. In contrast, host NK cells rejected H-2d/Hh-1d T cell grafts, whereas both NK cells and CD8+ TCR-alpha beta + T cells rejected H-2b/Hh-1b T cell grafts. Therefore, both CD8+ TCR-alpha beta T cells and NK cells mediate allogeneic and hybrid resistance to T cell grafts.

Animals↗
PubMed1995

Transplantable NK cell progenitors in murine bone marrow.

Differentiation of NK cells from pluripotent hematopoietic stem cells is a poorly understood process. Although it is known that NK cells are bone marrow derived and dependent upon an intact bone marrow microenvironment for complete maturation, it is not known if they arise from an intermediate lymphoid stem cell or from progenitors exclusively committed to the NK lineage. To determine whether phenotypically distinct committed NK progenitor cells exist in murine bone marrow, we sorted cells capable of repopulating recipient mice with mature NK cells upon i.v. transfer. We identified a rare population of bone marrow cells with the phenotype Ly6+ Lin- c-kit+ CD43high Fall-3high TSA-1- AA4.1low Rh123high that is highly enriched for the ability to generate NK cells after transplantation. Although these cells are relatively depleted of Rh123low pluripotent stem cells, they are highly enriched for both lymphoid and myeloid repopulating ability. Thus, we have found no evidence to support the existence of a phenotypically distinct transplantable progenitor population in mouse bone marrow that is either exclusively committed to the NK cell lineage or exhibits the functional characteristics of a common lymphoid stem cell.

Animals↗
PubMed1995

Biochemical indices of vitamin B12 nutrition in pregnant patients with subnormal serum vitamin B12 levels.

To determine the significance of the commonly observed fall in serum vitamin B12 levels during pregnancy, serum levels of the B12 metabolites methylmalonic acid (MMA) and homocysteine (Hcy) were measured in a group of 50 pregnant patients with subnormal serum B12 (range 45-199 pg/ml) and the results compared with those of 25 pregnant controls (serum B12(208-580) pg/ml). Mean values for serum MMA and total Hcy in the subnormal B12 group were 445.4 nmol/L and 7.03 mumol/L, respectively, which were not significantly different from the mean MMA of 440.5 nmol/L and Hcy of 6.88 nmol/L in the controls. For the total group of patients, neither serum MMA nor serum Hcy levels correlated with serum B12. One-third of pregnant patients showed elevated serum MMA values, independent of B12 status. Significant elevation of serum Hcy was detected in only two patients, both with subnormal serum B12 and hematological evidence of B12 deficiency. We conclude that the usual fall in serum B12 concentration in pregnancy does not reflect B12 deficiency at the biochemical level. In establishing true B12 deficiency in pregnancy, the serum Hcy level (in the absence of folate deficiency) but not serum MMA, is of value.

Female↗
PubMed1995

Activation of murine epidermal gamma delta T cells through surface 2B4.

Dendritic epidermal T cells (DETC) are gamma delta T cells that normally reside in murine skin. They express on their surface the 2B4 molecule, a 66-kDa glycoprotein of the immunoglobulin gene superfamily thought to be associated with anti-tumor cytotoxicity by natural killer and lymphokine-activated killer cells. Here, we show that ligation of surface 2B4 transduces cell activation signals in DETC. Treatment with anti-2B4 monoclonal antibodies triggers the secretion of interferon-gamma and interleukin-2 by DETC lines, induces proliferation of resting DETC lines, amplifies anti-CD3-dependent proliferation of DETC freshly isolated from mouse skin; and up-regulates egr-1 and c-fos mRNA expression. These results indicate a unique pathway for DETC activation.

Animals↗
PubMed1995

Hybrid resistance: 'negative' and 'positive' signaling of murine natural killer cells.

Murine NK cells can reject allogenic or parental-strain bone marrow cells (BMC) in vivo and can lyse T lymphoblasts in vitro. The 'missing self' hypothesis states that absence or presence of 'negative signals' from target cell class I antigens (Ag) to NK receptors determines whether or not lysis occurs. Indeed, lysis of parental-strain blasts by purified F1 NK cell subsets occurred only in the presence of anti-receptor antibodies. Evidence for 'positive signaling' to NK cells by class I Ag includes rejection of D8 (Dd) transgene to B6) BMC by B6 hosts. The outcome of other BMC transplants contradict the missing self idea, because donors with identical class I Ag differ in compatibility with certain hosts. Perhaps class I Ag-NK cell receptor interactions dominate over other target-NK cell interactions. These interactions are usually 'negative' but can be 'positive'.

Animals↗
PubMed1995

Ambulatory blood pressure in pregnancy: comparison of the Spacelabs 90207 and Accutracker II monitors with intraarterial recordings.

OBJECTIVE: Our purpose was to compare systolic and diastolic blood pressures obtained with the Spacelabs 90207 (Spacelabs Medical Products, Dee Why, Australia) or Accutracker II (Suntech Medical Instruments, Melbourne, Australia) ambulatory blood pressure monitoring devices with intraarterial blood pressures in pregnant women. STUDY DESIGN: Direct (intraarterial) and resting blood pressures with the ambulatory blood pressure monitoring device were compared in 39 pregnant women (14 Accutracker II and 25 Spacelabs 90207). RESULTS: The Accutracker II device underestimated direct systolic pressure by -9 (-13, -3) mm Hg (median, interquartile range) (p = 0.028) but gave similar diastolic pressure. The Spacelabs 90207 device gave similar systolic pressures but overestimated direct diastolic pressure by 7 (2, 12) mm Hg (p < 0.001). Variability for systolic and diastolic blood pressures within subjects was similar with the two devices. Both received poor gradings by standards of the British Hypertension Society and did not meet criteria of the Association for the Advancement of Medical Instrumentation, with intraarterial recordings used as the reference. CONCLUSIONS: The Accutracker II device significantly underestimated resting direct systolic pressure, whereas the Spacelabs 90207 device significantly overestimated resting direct diastolic pressure in pregnant women. Although poor gradings were achieved for both devices when intraarterial pressures were used as the reference, this is similar to comparisons of routine mercury sphygmomanometry with intraarterial recordings and does not mean these devices are unsuitable for use in pregnancy.

Blood Pressure Monitoring, Ambulatory↗
PubMed1995

Comparison of the susceptibility of the red fox (Vulpes vulpes) to a vaccinia-rabies recombinant virus and to cowpox virus.

Sylvatic rabies can be efficiently controlled by vaccination of foxes with a vaccinia-rabies recombinant virus. However, the risk of recombination between the engineered vaccine virus and other orthopoxviruses endemic in wildlife, such as cowpox virus, still needs to be investigated. In this study, foxes inoculated orally and intradermally with cowpox virus were found to be not very susceptible to cowpox virus, which was isolated from only the oropharynx and tonsils, at low titre and for only five days after inoculation. Thus the risk of recombination between these viruses in foxes is very low.

Animals↗
PubMed1995

Serological evidence for the reservoir hosts of cowpox virus in British wildlife.

The reservoir host of cowpox virus in Western Europe is not known, but epidemiological evidence from human and feline infections indicates that the virus is probably endemic in small wild rodents. Therefore, serum and tissue samples were collected from a variety of wild British mammals and some birds, and tested for evidence of Orthopoxvirus infection. Antibody reacting with cowpox virus was detected in 9/44 (20%) bank voles (Clethrionomys glareolus), 8/24 (33%) field voles (Microtus agrestis), 17/86 (20%) wood mice (Apodemus sylvaticus) and 1/44 house mice (Mus musculus), but in no other animal species tested. Although virus was not isolated from any animal, this serological survey, together with other evidence, suggests that bank and field voles and wood mice are the main reservoir hosts of cowpox virus in Great Britain.

Animals↗
PubMed1995

Protection against Mycoplasma pulmonis infection by genetic vaccination.

The induction of an immune response against a foreign protein usually requires purification of that protein, which is injected into animals. The isolation of pure protein is time consuming and costly. Recently, a technique called biolistic transformation (biological ballistic system) microparticle injection, gene gun, or particle bombardment was developed. The basic idea is that DNA or biological material coated onto heavy tungsten or gold particles is shot into target cells or animals. We have vaccinated mice by introducing the gene (Mycoplasma pulmonis DNA or a specific fragment) encoding a protein recognized by a protective monoclonal antibody directly into the skin or muscle of mice by two methods: (i) using a hand-held form of the biolistic system that can propel DNA-coated gold microprojectiles (2 micrograms of DNA) directly into the skin; (ii) using a conventional intramuscular injection of the DNA (100 micrograms) into quadricep muscles of transfected mice. HeLa cells were transfected in vitro by the gene gun or by the liposomal delivery system. Indirect immuno-fluorescent antibody (IFA) assay of culture cells indicated that both methods could be successful. Production of antibody and cell-mediated immunity against M.pulmonis were monitored by assaying serum IFA and enzyme-linked immunosorbent assay (ELISA), and delayed type hypersensitivity. In addition, macrophage migration inhibition and lymphocyte transformation to antigen in spleen cells were also tested. Both delivery systems induced humoral and cellular immunity, and vaccinated the mice against infection. Genetic immunization by using the gene gun saves time, money, and labor; moreover, this general method is also applicable to gene therapy.

Animals↗
PubMed1995

2B4, a new member of the immunoglobulin gene superfamily, is expressed on murine dendritic epidermal T cells and plays a functional role in their killing of skin tumors.

Dendritic epidermal T cells (DETC), which are skin-specific members of the tissue-resident gamma delta T-cell family, are characterized by their potential to kill selected tumor targets by a non-major histocompatibility complex (MHC)-restricted mechanism. We have recently identified a new receptor molecule, 2B4, that appears to be associated with non-MHC-restricted recognition of tumor targets by natural killer cells. The purpose of this study was to determine whether DETC express 2B4 molecules, and, if so, to assess their functional roles in DETC-mediated killing of tumor targets. Short-term DETC lines as well as DETC freshly procured from skin expressed surface 2B4, as detected with a specific monoclonal antibody. Removal of interleukin (IL)-2 from DETC cultures caused substantial reduction in 2B4 expression levels as well as a reduction in cytotoxic capacity against YAC-1 targets in a standard 51Cr-release assay. Conversely, exposure to IL-2, but not to IL-7, elevated both 2B4 expression and cytotoxicity. To assess the functional roles played by surface 2B4, we pretreated DETC lines with anti-2B4 antibody and then tested for their killing potential. Anti-2B4, but not the control antibody, augmented their capacity to lyse YAC-1 targets (51Cr-release assays) and to disrupt the monolayers of Pam-212-transformed keratinocytes (visual assessment). Thus, we conclude that DETC express, in an IL-2-dependent manner, 2B4 molecules, which may play a unique role in the killing of skin-derived tumors.

Animals↗