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Biomedical subjects

A L Scott

Publications and source records attributed to A L Scott.

At least 37 records · Page 2Linked to original sources
PubMed1998

Kinetics and cross-species comparisons of host antibody responses to lone star ticks and American dog ticks (Acari: Ixodidae).

Understanding of the animal antibody response to tick salivary gland proteins is necessary to identify candidate antibodies that may have use as species- and feeding-duration-specific biomarkers of tick exposure in humans. The kinetics of the humoral immune response of rabbits to challenge feeding by 2 tick species [Amblyomma americanum (L.) and Dermacentor variabilis (Say)] was characterized by both enzyme-linked immunosorbent assay and immunoblot. Western blot analysis revealed that rabbits produced antibodies against both A. americanum and D. variabilis tick salivary gland antigens, with molecular weights ranging from 12.2 to 125 kDa; the antibody response against the saliva of both tick species possessed both unique and common aspects. The presence of antibody against low-molecular-mass (< 20 kDa) salivary gland antigens of A. americanum may be specific for A. americanum exposure. Antibodies directed against D. variabilis salivary gland antigens of 111, 86.3, and 85 kDa may be specific for D. variabilis exposure. The data suggest that host antibodies directed against specific tick salivary gland proteins might have use as species-specific biologic markers of tick exposure.

Animals↗
PubMed1998

Filarial nematode parasites secrete a homologue of the human cytokine macrophage migration inhibitory factor.

Filarial nematode parasites establish long-term chronic infections in the context of an antiparasite immunity that is strongly biased toward a Th2 response. The mechanisms that lead to this Th2 bias toward filarial antigens are not clear, but one possibility is that the parasites produce molecules that have the capacity to proactively modify their immunological environment. Here we report that filarial parasites of humans secrete a homologue of the human proinflammatory cytokine macrophage migration inhibitory factor (MIF) that has the capability of modifying the activity of human monocytes/macrophages. A cDNA clone isolated from a Brugia malayi infective-stage larva expression library encoded a 12.5-kDa protein product (Bm-MIF) with 42% identity to human and murine MIF. MIF homologues were also found to be expressed in the related filarial species Wuchereria bancrofti and Onchocerca volvulus. Bm-mif was transcribed by adult and larval parasites, and the protein product was found in somatic extracts and in the parasite's excretory-secretory products. Immunohistocytochemistry revealed that Bm-MIF was localized to cells of the hypodermis/lateral chord, the uterine wall, and larvae developing in utero. Unexpectedly, the activities of recombinant Bm-MIF and human MIF on human monocytes/macrophages were found to be similar. When placed with monocytes/macrophages in a cell migration assay, Bm-MIF inhibited random migration. When placed away from cells, Bm-MIF induced an increase in monocyte/macrophage migration that was specifically inhibited by neutralizing anti-Bm-MIF antibodies. Bm-MIF is the first demonstration that helminth parasites produce cytokine homologues that have the potential to modify host immune responses to promote parasite survival.

Amino Acid Sequence↗
PubMed1998

Antibody to a cDNA-derived calreticulin protein from Amblyomma americanum as a biomarker of tick exposure in humans.

The antibody responses of human and animal hosts were studied to determine the utility of antibody against recombinant tick calreticulin (rTC), a cDNA-derived protein isolated from salivary glands of Amblyomma americanum L., as a biologic marker of tick exposure. Rabbits fed upon by either A. americanum or Dermacentor variabilis Say developed significant anti-rTC antibody responses, as measured by both ELISA and immunoblot assay. In contrast, gerbils exposed to Aedes aegypti did not develop anti-rTC antibodies, as measured by ELISA or immunoblot assay. The utility of the assay was next evaluated in humans at high risk for tick exposure. During April through September 1990, 192 military personnel who originated from either Fort Chaffee, Arkansas or Fort Wainwright, Alaska were studied during maneuvers in tick infested areas at Fort Chaffee. Study subjects completed a questionnaire and had pre- and post-maneuvers serum specimens analyzed for antibodies to rTC. In adjusted analysis (controlling for age, fort of origin, attached tick during maneuvers, and bed netting use), the use of bed netting and home station were associated with post-maneuvers anti-rTC antibody seropositivity by ELISA. Subjects from Fort Wainwright were more likely to be seropositive for anti-rTC antibody (adjusted odds ratio = 5.3, 95% confidence interval [CI] = 1.1-25.6). Personnel who did not report the use of bed netting were more likely to be anti-rTC seropositive (adjusted odds ratio = 6.8, 95% CI = 1.4-32.4). Immunoblot assays showed that humans had specific anti-rTC antibody responses. The animal experiments demonstrate that hosts exposed to naturally feeding ticks develop anti-rTC antibodies. The data also indicate that hosts exposed to Ae. aegypti saliva may not develop antibodies against rTC. Observations in tick-exposed humans support the hypothesis that anti-rTC antibody seropositivity is a biologic marker of tick exposure.

Adult↗
PubMed1997

Cyclic food restriction, insulin and mammary cell proliferation in the rat.

We reported recently that weight cycling significantly increased the incidence of mammary cancer in virgin female rats that were pretreated with N-methyl-N-nitrosourea. The present study investigated the effect of weight cycling on mammary epithelial cell proliferation and its relationship to changes in plasma insulin, estrogen, progesterone and urinary corticosterone in 30 female virgin Sprague-Dawley rats. Animals were fed a modified AIN-76A diet containing 24.6% corn oil by weight. Weight-cycled (WC) rats were food restricted daily by either 33% or 50% of non-restricted controls for 1 week followed by 3 weeks compensatory refeeding and weight recovery over 18 weeks or 4.5 weight cycles. WC rats consumed 6-10% less food than controls (P = 0.01) but showed a 71-89% greater efficiency of food utilization for growth (P < 0.0001) than controls. There were no differences in total weight gain during treatment. Mammary lobuloalveolar and ductal cell proliferation of WC rats, measured by 5-bromo-2'deoxyuridine labelling, increased in a dose-response fashion, P = 0.03, P = 0.06 respectively in comparison to controls. Energy and substrate utilization measured by indirect calorimetry indicated WC animals expended less energy (P = 0.005) and utilized less glucose (P = 0.0001) and protein (P = 0.006) during restriction, and less lipid during recovery (P = 0.05) than controls. There were no significant differences in hormone levels between groups. Multiple regression analysis with plasma insulin, estrogen, progesterone and urinary corticosterone as independent variables (r = 0.947, r2 = 0.897, P = 0.003) showed that plasma insulin was the only significant predictor (P < 0.01) of mammary cell proliferation. In accord with this observation, tyrosine-phosphorylated activation of insulin receptor substrate-1, detected by immunoprecipitation and Western immunoblot analysis in mammary tumors of WC rats from our previous study, was 3-5 times greater than in non-restricted controls (P < 0.01). Present findings suggest that weight cycling in rats increases risk of breast cancer development via insulin stimulated mammary cell proliferation.

Animals↗
PubMed1996

Genes expressed in Brugia malayi infective third stage larvae.

We have used a tag sequencing approach to survey genes expressed in the third stage infective larvae of the human filarial nematode parasite Brugia malayi. RNA was isolated from late vector-stage L3 larvae after days 9 or 10 of infection in mosquitos, and converted to cDNA by reverse transcriptase. Double-stranded cDNA was produced by either conventional methods (non-SL cDNA library) or by PCR using the nematode spliced leader (SLI) and oligo(dT) primers (SL cDNA library). Two clone libraries (one from SL and one from non-SL cDNAs) were constructed in lambda ZapII. A set of these full-length clones was selected and 596 inserts were sequenced from the 5' end. We have identified 364 B. malayi genes (the majority of which are new) that encode housekeeping proteins, structural proteins, proteins of immediate immunological or drug-discovery interest as well as a large class of novel sequences which may prove to have significant involvement in host invasion. Extensive, genome-wide approaches to the analysis of larval gene expression are now possible for B. malayi. We present several examples of this approach.

Amino Acid Sequence↗
PubMed1996

Nematode sperm.

Parasitic nematode infections remain a major public health problem in many parts of the world. Because most of the current strategies aimed at controlling parasitic nematode infections have met with only limited success, it may be time to consider alternative approaches. An aspect of nematode biology that has drawn little attention as a target for control is the reproductive process. Although there are numerous facets of the overall reproductive biology of nematodes that hold potential as targets for intervention, Alan Scott here focuses on the male reproductive system, and outlines some of the known unique processes and characteristics of sperm formation and sperm function that could be exploited to block fertilization.

Journal Article↗
PubMed1996

Salivary gland changes and host antibody responses associated with feeding of male lone star ticks (Acari:Ixodidae).

The goal of this study was to demonstrate that male lone star ticks, Amblyomma americanum (L.), actively feed on rabbits during attachment and that the host is capable of mounting an immune response against male salivary gland proteins. During attachment, it was shown that male ticks salivary glands hypertrophy. An enzyme-linked immunosorbent assay was used to detect rabbit serum proteins in the midgut of previously attached male ticks. SDS-polyacrylamide gel electrophoresis showed that the proteins expressed in the male tick salivary gland changed during feeding, with several new proteins in the 15- to 50-kDa range synthesized during attachment. Rabbits mounted a detectable antibody response against male tick salivary gland proteins after 2 sequential feedings of male ticks. The antibodies were directed against a spectrum of male salivary gland proteins ranging from 18 to 160 kDa. Several of these proteins were not recognized by antibodies directed against female tick saliva proteins, and thus may be specific male salivary gland components. This evidence indicates that male A. americanum ticks actively feed during attachment, that their salivary gland proteins change during feeding, and that male salivary gland proteins are immunogenic.

Animals↗
PubMed1996

Muscle actin gene from Aedes aegypti (Diptera:Culicidae).

A recombinant phagemid containing a 1,240-bp insert encoding an actin was isolated from a yellow fever mosquito, Aedes aegypti (L.), complementary DNA library. This insert (pBS-Act35) contained an open reading frame of 822 bp whose deduced amino acid sequence exhibited > 95% homology with the carboxyl terminal 274 amino acids of Drosophila melanogaster Meigen and silkworm, Bombyx mori (L.), actin genes. Reverse transcriptase-polymerase chain reaction was used to clone and determine the sequence of the additional 306 nucleotides that comprise the 5' end of the gene. The coding nucleotide sequence of the whole gene (designated Aeact-1) exhibited between 81 and 89% homology with coding sequences of D. melanogaster and B. mori actin genes, and its deduced amino acid sequence exhibited > 95% homology with those genes. The highest similarity of Aeact-1 gene at the amino acid sequence level was with B. mori and D. melanogaster muscle actins. Southern blot analysis indicated that the Aedes genome contains at least 5 actin-related sequences.

Actins↗
PubMed1996

Cyclic food restriction alters substrate utilization and abolishes protection from mammary carcinogenesis female rats.

We tested the hypothesis that cyclic food restriction abolishes protection against mammary carcinogenesis. Virgin female Sprague Dawley rats (n = 159) were injected intraperitoneally with 25 mg/kg n-methyl-n-nitrosourea at 50 d of age. Eleven days later, rats were given free access to a 24.6 g fat/100 g AIN-76A diet (ad lib-c), fed in two meals (me-c), or fed in two meals restricted in weight by 33% for 1 wk followed by 3 wk of compensatory refeeding (me-r) for 18 wk or 4.5 restriction cycles. Energy and substrate utilization of 15 rats from each group was measured by indirect calorimetry. The me-r rats ate and weighed less (P < 0.0001), had a greater efficiency of food utilization (P < 0.01), and had a 12% higher incidence of mammary cancer (P < 0.0001) than ad lib-c rats after adjusting for the effect of final body weight. Resting metabolic rate was not different among groups, but me-r rats used less glucose during restriction and more glucose and less lipid for energy during body weight recovery than me-c rats (P < 0.0001). Increased energy efficiency and the shift in utilization of glucose and fatty acids followed closely the effects of cyclic food restriction and meal feeding on mammary carcinogenesis.

Adipose Tissue↗
PubMed1996

Immunoglobulin G subclass responses of children during infection with Onchocerca volvulus.

To characterize the patterns of immunoglobulin G (IgG) subclass and IgE reactivity during the early stages of onchocerciasis, sera were collected from 224 children (age groups, 2 to 5, 6 to 10, and 11 to 15 years) residing in a region of Sierra Leone where Onchocerca volvulus is endemic, and these samples were tested by enzyme-linked immunosorbent assay for their reactivity to adult antigens (OvAg) and against four recombinant proteins (OV11, OV27, OV29, and OV16). Over 88% of the samples contained detectable levels of anti-OvAg IgG. In samples from microfilaria (MF)-positive children, IgG4 responses were significantly elevated and constituted on average 39, 35 and 28% of the total IgG responses for the age groups of 2 to 5, 6 to 10, and 11 to 15 years, respectively. For MF-negative individuals, the mean contributions of IgG4 to the total IgG response were 11% (2 to 5 years), 27% (6 to 10 years), and 56% (11 to 15 years). OvAg-specific IgE was detectable in the sera from both MF-negative and MF-positive individuals. To increase the specificity of the response, recombinant antigens OV11, OV27, and OV29 were tested individually or as a cocktail. Nearly 50% of the MF-negative children and 85% of the MF-positive children had detectable levels of IgG against at least one of the recombinant antigens. Only a small portion of the IgG against the recombinant peptides was IgG4. The prevalence of IgG against OV16 in samples from MF-negative children was 51%, and that for MF-positive children was 75%. The general profile of the humoral immune responses mounted by both MF-positive and a large percentage of the MF-negative children during the initial phases of infection with O. volvulus is similar to the profile reported for adults harboring chronic O. volvulus infections. These results suggest that very quickly after infection, the interactions between parasite and host result in an immunological environment that may contribute to the maintenance of a long-term, chronic infection.

Adolescent↗
PubMed1996

Vitamin A deficiency enhances ozone-induced lung injury.

The present study determined the effects of vitamin A (vA) deficiency on the responses to ozone (O3) challenges in two inbred strains of mice that are differentially susceptible to O3-induced lung inflammation. Susceptible C57BL/6J (B6) and resistant C3H/HeJ (C3) dams at 2 wk gestation were fed test diets containing either 0 or 10 micrograms retinol/g diet. In mice that were maintained on vA-sufficient (vA+) diet, lung and liver tissue concentrations of vA and retinyl palmitate (RP) were significantly (P<0.05) lower in the B6 strain compared with C3, as measured by high-performance liquid chromatography techniques. vA and RP levels were significantly (P<0.05) reduced in lung and liver tissues of 8-wk old B6 and C3 mice that were maintained on a vA deficient (vA-) diet. vA+ and vA- mice of both strains were exposed to air or 0.3 ppm O3/72 h, and lung injury was assessed by differential cell count and total protein concentration in bronchoalveolar lavage (BAL) returns. O3 exposure caused significantly (P<0.05) greater increases in inflammatory cells and a total protein in BAL returns of vA+ B6 mice than vA+ C3 mice. vA deficiency significantly (P<0.05) enhanced O3-induced increases in polymorphonuclear leukocytes in C3 mice and epithelial cells loss in both strains. Compared with vA+ mice, lung permeability was also significantly (P<0.05) enhanced in vA- mice of both strains exposed to O3. vA replacement partially reversed the O3-induced lung injury that was enhanced by vA- diet. Results indicate that vA may have an important role in the pathogenesis of O3-induced lung injury in differentially susceptible inbred strains of mice.

Analysis of Variance↗
PubMed1995

Nucleoside diphosphate kinase from the parasitic nematode Brugia malayi.

Using a reverse transcription-polymerase chain reaction (RT-PCR) procedure that exploited the presence of a conserved 22-nucleotide spliced leader (SL) sequence that is trans-spliced to the 5' end of nematode transcripts, a novel Brugia malayi (Bm) infective-stage SL cDNA expression library was constructed and characterized. The library was immunoscreened with rabbit anti-infective-stage antibodies (Ab) and an immunodominant clone, BmG4-7, was identified and characterized. BmG4-7 contained a full-length cDNA that had significant sequence similarity to nucleoside diphosphate kinase (NDK)-encoding sequences reported from a number of species, including Drosophila melanogaster and humans. BmNDK was found to be constitutively transcribed during all stages of parasite development. An anti-BmNDK Ab was used to immunostain a Western blot of extracts from adult and larval parasites. The Ab specifically recognized a 17.5-kDa molecule in all of the parasite extracts. Molecular modeling of the BmNDK showed several regions surrounding the conserved catalytic site that may be important in the design of drugs specific for the disruption of NTP synthesis in filarial parasites.

Amino Acid Sequence↗
PubMed1995

Brugia malayi: retinoic acid uptake and localization.

Retinoids play an important role in development and growth in a wide range of cells and tissues. The vitamin A status of the host appears to be a factor in the development of certain parasites, suggesting that vitamin A is an essential micronutrient for the proper growth and differentiation of parasites. To determine if exogenous retinoic acid is taken up by filarial parasites, adult male and female Brugia malayi were exposed to [3H] retinoic acid. Results indicated that retinoic acid was taken up by adult parasites in a specific and dose-dependent manner. In tissue sections of [3H] retinoic acid-labeled B. malayi, label was shown to localize to the cellular and not the acellular portions of the worms with high density of label associated with both early and late embryonic forms. The specific uptake and distribution of retinoic acid in B. malayi suggest that it may play a role in parasite growth and development.

Animals↗
PubMed1995

Molecular cloning of a gene expressed during early embryonic development in Onchocerca volvulus.

Little attention has been paid to the reproductive biology of filarial nematode parasites as a possible target for immunological or chemotherapeutic intervention. An interruption of the reproductive process would, in addition to breaking the cycle of transmission, reduce the morbidity associated with certain filarial infections. As part of our efforts to define molecules that have important functions during filarial embryogenesis, antibodies against embryo-associated proteins were used to identify a 6308-bp cDNA sequence (ovt1) from an Onchocerca volvulus cDNA expression library. The ovt1 cDNA contained an open reading frame that coded for 2022 amino acids. The deduced amino acid sequence was highly hydrophilic, alpha-helical in nature and included two leucine zipper domains. OVT1 also contained a single Arg-Gly-Asp (RGD) site. The results of Southern blot analyses demonstrated that an ovt1-like gene occurs in a number of different species of filarial nematodes. In situ hybridization experiments to identify tissues that contain ovt1 transcripts showed that ovt1 was transcribed at high levels in the late morula/early blastocyst stage of embryonic development. Transcripts for ovt1 were also detected in O. volvulus larvae and in the hypodermal cells of adult parasites. Two fragments of ovt1 were expressed as fusion proteins and the fusion proteins were used to produce antibodies in rabbits. Both antibodies recognized a native protein with an apparent molecular mass of 230 kDa in extracts from gravid female O. volvulus. In addition, the antibodies reacted with a restricted number of lower-molecular mass bands which may represent the products of post-transcriptional or post-translational processing. The predicted coiled-coil structure and the sites of transcription suggest that OVT1 may be a component of the extracellular matrix.

Amino Acid Sequence↗
PubMed1995

Molecular cloning of a serine proteinase inhibitor from Brugia malayi.

The antigens produced by the infective-stage larvae of filarial parasites are potentially important targets for a protective immune response. A major impediment to studies on the biochemistry and molecular biology of antigens from infective larvae is a lack of parasite material. By employing a reverse transcription PCR-based strategy which exploited the presence of a conserved 22-nucleotide spliced leader sequence present at the 5' end of a proportion of nematode transcripts, spliced leader-containing cDNAs were amplified from the late-vector-stage larvae of the filarial nematode Brugia malayi. A major 1.4-kb PCR product was cloned into pBluescript. One of the PCR cDNA clones (BmY8) contained a 1,287-bp insert that encoded the first member of the serine proteinase inhibitor (serpin) superfamily to be described from nematodes. Reverse transcription PCR analysis of RNA isolated from different developmental stages of the parasite showed that transcription of the B. malayi serpin (Bmserpin) begins between days 8 and 9 of larval development within the insect vector and continues through to the adult and microfilarial stages. In immunoblot analyses of B. malayi somatic extracts, the native protein was estimated to have a molecular weight of 44,000. In immunoblots using excretory-secretory products from infective- and fourth-stage larvae, a single band with an estimated molecular weight of 75,000 was detected. A quantitative analysis of somatic extracts demonstrated that infective-stage larvae contained 10- to 16-fold-more Bmserpin than adults or microfilariae. Bmserpin was immunogenic in gerbils and was recognized strongly by sera from immunized animals. Bmserpin, which has the potential for modifying host defense responses, may play an important role in parasite survival during the early phase of vertebrate-stage development.

Amino Acid Sequence↗