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Biomedical subjects

M Hansson

Publications and source records attributed to M Hansson.

At least 91 records · Page 5Linked to original sources
PubMed1994

B- and T-cell responses in congenic mice to repeat sequences of the malaria antigen Pf332: effects of the number of repeats.

The Plasmodium falciparum antigen Pf332 comprises degenerated 11-amino-acid repeats with regularly spaced pairs of glutamic acid. Epitopes formed by such repeats are recognized by polyclonal and monoclonal antibodies that interfere with the life cycle of the blood stages of the malaria parasite. In order to study the immunogenicity of one such Pf332 repeat sequence (SVTEEIAEEDK), fusion proteins containing ZZ (two IgG binding domains of staphylococcal protein A) and dimers, trimers or tetramers of the malarial sequence were injected into mice. To analyse possible major histocompatibility complex class II restrictions of the immune response, mice of different H-2 haplotypes were used. A significant antibody response was elicited by administration of all the three fusion proteins in mice expressing the I-Ak allele (B10.BR, B10.A(2R) and B10.A(4R)) whereas B10 and C57BL/6 (H-2b) mice were low responders. In comparison, B10.D2 (H-2d) mice were low responders to fusion proteins with 2 or 3 repeats but responded well to the protein containing 4 repeats. Lymph node cells from B10.BR (H-2k) mice, primed in vivo with ZZ-fusion proteins containing either 2 or 4 repeats, proliferated in vitro in response to repeat sequences fused to ZZ or to an unrelated fusion partner, as well as to a synthetic peptide containing less than two repeats. In contrast, a response of lymph node cells from B10.D2 (H-2d) mice was only obtained when a fusion protein containing 4 repeats was used both for in vivo priming and in vitro restimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Single-step recovery of a secreted recombinant protein by expanded bed adsorption.

We have used an expanded bed adsorption procedure for efficient recovery of a recombinant fusion protein, directly from a crude fermentor broth without prior cell removal. The fusion protein was designed to have a relatively low isoelectric point (pI) to allow anionic exchange adsorption at pH 5.5 where most Escherichia coli host proteins are not adsorbed. The gene product was secreted to the culture medium of the E. coli host cells in high yields (550 mg/l). The separation of cells and the concentration and recovery of the fusion protein could therefore be achieved by a single unit operation. The yield after the expanded bed adsorption exceeded 90 percent. Furthermore, the significant volume reduction by the expanded bed adsorption, enabled efficient and straight-forward polishing of the product by a subsequent affinity chromatography step, for removal of contaminating DNA and pyrogenic compounds to levels acceptable for regulatory authorities. An overall yield exceeding 90 percent was maintained after the affinity chromatography polishing step. The procedure outlined here is suitable for large-scale bioprocesses and allows efficient removal of cells, host proteins, contaminating DNA and endotoxins.

Adsorption↗
PubMed1994

Bacillus subtilis HemY is a peripheral membrane protein essential for protoheme IX synthesis which can oxidize coproporphyrinogen III and protoporphyrinogen IX.

The hemY gene of the Bacillus subtilis hemEHY operon is essential for protoheme IX biosynthesis. Two previously isolated hemY mutations were sequenced. Both mutations are deletions affecting the hemY reading frame, and they cause the accumulation of coproporphyrinogen III or coproporphyrin III in the growth medium and the accumulation of trace amounts of other porphyrinogens or porphyrins intracellularly. HemY was found to be a 53-kDa peripheral membrane-bound protein. In agreement with recent findings by Dailey et al. (J. Biol. Chem. 269:813-815, 1994) B. subtilis HemY protein synthesized in Escherichia coli oxidized coproporphyrinogen III and protoporphyrinogen IX to coproporphyrin and protoporphyrin, respectively. The protein is not a general porphyrinogen oxidase since it did not oxidize uroporphyrinogen III. The apparent specificity constant, kcat/Km, for HemY was found to be about 12-fold higher with coproporphyrinogen III as a substrate compared with protoporphyrinogen IX as a substrate. The protoporphyrinogen IX oxidase activity is consistent with the function of HemY in a late step of protoheme IX biosynthesis, i.e., HemY catalyzes the penultimate step of the pathway. However, the efficient coproporphyrinogen III to coproporphyrin oxidase activity is unexplained in the current view of protoheme IX biosynthesis.

Amino Acid Sequence↗
PubMed1993

Cell-surface display of heterologous epitopes on Staphylococcus xylosus as a potential delivery system for oral vaccination.

A system has been developed for the surface expression of heterologous receptors on the cell surface of Staphylococcus xylosus. Gene fragments encoding peptides to be displayed on the cell surface can be assembled by applying a polymerization strategy based on the class-IIS restriction enzyme BspMI and thereafter subcloned into an Escherichia coli-staphylococci shuttle vector designed for targeting of produced fusion proteins to the outer cell surface of the Gram-positive host cell. A heterologous receptor was genetically assembled and expressed on the surface of S. xylosus where the separate regions could be independently probed in immunogold assays, using antisera reacting with different regions of the recombinant receptor. In addition, a receptor-specific humoral immune response could be elicited in mice by oral immunization with recombinant S. xylosus cells, suggesting that these type of Gram-positive bacteria might offer potential vehicles for oral vaccination.

Administration, Oral↗
PubMed1993

Murine thymocytes with ability to inhibit IL-2 production. II. Characterization of a subpopulation with regulatory function in the thymus.

Studies were performed to characterize the thymocyte subset responsible for the efficient inhibition of spleen cell interleukin-2 (IL-2) production. By different cell separation techniques, C-mediated cytotoxicity, immunoabsorbance, and cell sorting by flow cytometry, we have identified two phenotypically distinct subpopulations of thymocytes. One subset, belonging to the minor population (3-5%) of the CD4-CD8-, i.e., double-negative thymocytes, is defined as the subset from which the suppressive thymocytes are generated. After 28 hr of Con A stimulation, these cells undergo a phenotypical change in vitro and generate a population exerting the inhibitory effect. This latter subset inhibits 95-99% of the IL-2 produced by spleen cells and is characterized by expressing the CD8 antigen, high levels of HSA, low levels of CD3, and being IL-2R positive (HSA+CD4-CD8+CD3lowIL-2R+). Based on the experiments where stimulated CD4+CD8+, i.e., double-positive thymocytes, failed to suppress IL-2 production, we conclude that the CD8+ immature single-positive thymocytes are generated directly from the DN subset as an intermediate stage to the DP cells. When CD8(+)-stimulated thymocytes were enriched, the suppression was efficient even at thymocyte:spleen cell ratio of 0.01:1. It is suggested that this subpopulation of thymocytes may serve as a regulatory set of cells during critical stages of thymic maturation.

Animals↗
PubMed1993

Heme b (protoheme IX) is a precursor of heme a and heme d in Bacillus subtilis.

Bacillus subtilis can synthesise cytochromes containing a-, b-, c- and d-type heme. The biosynthetic pathways of these heme prosthetic groups were investigated by using strains blocked in uroporphyrinogen III synthesis from porphobilinogen or in heme b (protoheme IX) synthesis from uroporphyrinogen III. The results strongly suggest that heme a and heme d are both synthesised from heme b (protoheme IX). They also indicate that B. subtilis contains a novel ferrochelatase involved in the synthesis of siroheme.

Bacillus subtilis↗
PubMed1993

Plasmodium falciparum: the immune response in rabbits to the clustered asparagine-rich protein (CARP) after immunization in Freund's adjuvant or immunostimulating complexes (ISCOMs).

The Plasmodium falciparum clustered asparagine-rich protein (CARP) is a merozoite-associated antigen which contains approximately 30% asparagine. Analysis of the DNA sequences located 5' of the cloned 1.4-kb CARP gene in the P. falciparum genome suggests that this gene fragment may encode the complete CARP and that the gene product is a protein of M(r) 50,000. To analyze the immunogenicity of CARP, the gene was expressed as a fusion protein with staphylococcal protein A (SpA-CARP). Immunization of rabbits with SpA-CARP in Freund's complete adjuvant (FCA) resulted in a strong antibody response against CARP as measured in ELISA. This response was efficiently boosted and sustained over a long time while that induced by two immunizations with SpA-CARP in ISCOMs was weak and of shorter duration. In both instances, the antibody levels against CARP were further increased by a second booster injection consisting of either SpA-CARP or CARP fused to the serum albumin-binding region (BB) of streptococcal protein G (BB-CARP) in PBS, indicating that immunizations with SpA-CARP in FCA or ISCOMs had induced a CARP-specific immunological memory. Boosting with BB-CARP in PBS was more efficient than boosting with SpA-CARP in PBS. In all rabbits, the antibodies obtained after the booster with CARP in PBS were the most efficient inhibitors of merozoite invasion in vitro. The antisera reacted with the intracellular parasite in immunofluorescence and with a band of M(r) 50,000 in immunoblotting while several high-molecular-weight components as well as the one of M(r) 50,000 were immunoprecipitated. The specificity of the antibody responses varied between the different rabbits as indicated in ELISA, with short synthetic peptides representing different CARP sequences. Taken together, the results suggest that a previously cloned genomic DNA fragment may encode the complete P. falciparum blood-stage antigen CARP and that CARP is immunogenic in rabbits both when administered in FCA or ISCOMs.

Amino Acid Sequence↗
PubMed1993

Presence of immunoreactive atrial natriuretic peptide in nerve fibres and conduction cells in the conduction system of the bovine heart.

Previous findings of atrial (A-type) natriuretic peptide (ANP) in nervous tissue, such as the brain and the superior cervical ganglia, led us to investigate the possible occurrence of ANP in nervous tissue in the heart. The distribution of ANP in the bovine heart, particularly its conduction system, was examined by the use of immunohistochemical methods and an antiserum against alpha-hANP. ANP immunoreactivity was frequently detected in atrial myocytes and in the Purkinje fibres of the AV-bundle, and was sometimes seen in the Purkinje fibres of the bundle branches and their ramifications. On the other hand, ANP immunoreactivity was never seen in the conduction cells of SA- and AV-nodes. ANP immunoreactivity was also detected in small nerve-fibre varicosities, mainly in the AV-node and AV-bundle. Most of these varicosities were located in the proximity of the conduction cells, but some occurred close to fine blood vessels or in the walls of arterioles. These observations show for the first time that ANP immunoreactivity is present not only in atrial myocytes and conduction cells but also in nerve-fibre varicosities in the conduction system. The observations suggest that ANP may act as a neuromodulator and/or neurotransmitter in the conduction system.

Animals↗
PubMed1993

The colony stimulating factors.

Hematopoiesis is a dynamic process, which generate in the range of 10(9) cells/kg each day of erythroid and myeloid cells respectively. In vitro assays that were developed 20 years ago, have been used to define factors that can stimulate growth and differentiation of bone marrow (BM) derived progenitor cells. These growth factors for hematopoiesis were termed Colony Stimulating Factors (CSFs) since the assay system was to induce colonies. With the application of molecular biologic approaches, the genes encoding for these CSFs have been localized and cloned. Production of CSFs and other soluble signal substances (cytokines) as pure proteins have led to important insights into how hematopoiesis is regulated by a complex network made up by interactions between cells and cytokines. The availability of CSFs in clinically useful amounts has also led to clinical trials with new strategies for treating hematopoietic dysfunctions, congenital or acquired. Because others have recently reviewed clinical applications or basic science studies on the colony stimulating factors, we will summarize the two with focus on common features between the different CSFs.

Animals↗
PubMed1993

Immunogenicity in rabbits and monkeys of influenza ISCOMs conjugated with repeated sequences of the Plasmodium falciparum antigen Pf155/RESA.

Rabbits and monkeys were immunized with two fusion proteins, ZZ-M3 and ZZ-M5, coupled to pre-formed influenza virus membrane glycoprotein ISCOMs. The fusion proteins comprise two IgG-binding domains from staphylococcal protein A (ZZ) and repeated amino acid sequences from the C-terminal (M3) or central (M5) repeat regions of the Plasmodium falciparum antigen Pf155/RESA. The induced antibody responses were of long duration, could be efficiently boosted and were comparable to those obtained with Freund's Adjuvant. The produced antibodies reacted with M3, M5, protein A and the influenza glycoprotein, recognized Pf155/RESA and inhibited merozoite invasion in vitro. These results suggest that coupling of immunogens to pre-formed ISCOMs may be a basis for construction of multivalent subunit vaccines.

Amino Acid Sequence↗
PubMed1993

Activated natural killer cells suppress myelopoiesis in mice with severe combined immunodeficiency.

The in vivo effect of natural killer (NK) cell activation on autologous myelopoiesis was studied in an environment deficient of functional T and B cells. Administration of 3,6-bis[2-(Dimethylamino)-ethoxy]-9H-xanthen-9-one dihydrochloride) Tilorone) or recombinant interleukin-2 (rIL-2) to mice with severe combined immunodeficiency (C.B.-17 scid/scid) resulted in an increase in YAC-1 lysis by their splenocytes as well as bone marrow cells. Recombinant IL-2 furthermore led to a fivefold increase in the cellularity of the spleen. When assayed against human NK/lymphokine-activated killer (LAK) target, K562 cell line, the IL-2-activated mouse cells exhibited no cytotoxicity across the species barrier. Both agents induced a profound suppression of myelopoietic progenitor cells as measured in a 7-day granulocyte-macrophage colony forming cell (GM-CFC) assay. We conclude that the presence of neither functional T nor B cells is necessary for NK cells to mediate inhibition of myelopoiesis in the autologous host.

Animals↗
PubMed1993

Lipoprotein alterations in children with bacterial meningitis.

Abnormalities in serum lipids, including hypertriglyceridemia, are common during infectious disorders. However, the lipoprotein pattern during infections, particularly in children, has been investigated to only a limited extent. We have monitored alterations in serum lipoproteins in eight children with a severe bacterial infection (meningitis) by a quantitating method measuring cholesterol and triglycerides in each major class. The levels of triglycerides in serum and in low-density lipoproteins were markedly elevated during the infection, whereas the amount of cholesterol in high-density lipoproteins was decreased. The cholesterol to triglyceride ratio was decreased in low-, as well as in high-density lipoproteins. These lipoprotein abnormalities may, at least in part, be explained by a depressed lipolytic activity of lipoprotein lipase, the key enzyme for removal of triglycerides in man. Serum triglycerides and the levels of cholesterol in high-density lipoproteins, as well as the ratio between these parameters, may be used as indicators of inflammatory activity.

Child, Preschool↗
PubMed1993

Severity of alcohol dependence; rating by patient records.

A method to assess the severity of alcohol dependence by rating patient records was developed using the items of the 'alcohol abuse' subscale of the Swedish version of the Alcohol Use Inventory (AVI). The records of 37 alcoholics were rated and the scores were compared with the patients' self-report scores of the AVI and clinical assessments based on the structured clinical interview for DSM-III-R (SCID). The inter-rater reliability was rs = 0.88. The correlations between the record ratings and the alcoholics' self-ratings were rs = 0.77 for both raters and 0.55-0.57 between the record ratings and the total SCID-scores. The correlations between the self-ratings and SCID-scores were rs = 0.90. It is suggested that this method may facilitate record-based follow-up studies of alcoholics.

Adult↗
PubMed1993

Solid-phase gene assembly of constructs derived from the Plasmodium falciparum malaria blood-stage antigen Ag332.

A general method for solid-phase gene assembly on streptavidin-coated magnetic beads has been developed. The introduction of biotin in the 5'-end of the initiation oligonucleotide enables anchoring to the bead by means of the streptavidin-biotin interaction. The immobilization of one oligonucleotide enables controlled, stepwise annealing/ligation of successive 5'-phosphorylated oligonucleotides to rapidly build up predesigned gene constructs. In this report, we have assembled gene constructs of different lengths derived from the Plasmodium falciparum malaria blood-stage antigen Ag332. The encoded gene products were subsequently expressed in Escherichia coli using two parallel expression systems based on staphylococcal protein A and streptococcal protein G, respectively.

Amino Acid Sequence↗
PubMed1992

Diverse effect of cytokine treatment of tumor cells on specific versus non-specific cytotoxicity.

The effect of IFN-gamma and TNF-alpha treatment of an ovarian carcinoma line on the sensitivity to lysis by specific CTL clones and non-specific Tumor Associated Lymphocytes (TAL), isolated from the ascites fluid, was analyzed. The in vitro established TAL line displayed a non-specific lytic activity against the autologous tumor as well as against several allogeneic tumor lines. Pretreatment with IFN-gamma alone, or in combination with TNF-alpha, rendered the carcinoma line less susceptible to lysis by the autologous TAL line. Conversely, susceptibility to lysis by tumor specific T cell clones, isolated from the TAL line, was increased as a result of cytokine pretreatment. Several TCR-alpha/beta+, CD8+ T-cell clones showing a more specific pattern of lysis against the autologous tumor were isolated. Lysis of the autologous tumor by these clones involved the TCR-alpha/beta via a MHC-class I restricted mechanism dependent on the adhesion molecules ICAM-1 and LFA-3, as inferred from antibody blocking studies. The enhanced sensitivity to specific CTL clones seen after cytokine treatment may be related to the enhanced expression of ICAM-1 molecules on the ovarian carcinoma. These results have implications for cytokine based immunotherapy, where IFN-gamma may enhance the effects of tumor associated specific CTL while decreasing that of non-specific effector cells.

Ascitic Fluid↗
PubMed1992

OKT3-induced cytokine mRNA expression in human peripheral blood mononuclear cells measured by polymerase chain reaction.

The kinetic profile of cytokine gene expression in normal human peripheral mononuclear cells (MNC) activated by an anti-CD3 monoclonal antibody was studied. The presence or absence of 10 different cytokine mRNA were measured in a polymerase chain reaction (PCR) assisted mRNA amplification assay. After 2 h of stimulation the mRNA for interleukin-1 alpha (IL-1 alpha), interleukin-2 (IL-2), interleukin-3 (IL-3), tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) were detectable and remained present during the whole time period studied (22 h). Interleukin-6 (IL-6) and granulocyte macrophage colony stimulating factor (GM-CSF) were detected after 4 h, while interleukin-10 (IL-10) mRNA did not appear until after 7 h; they all remained expressed at 22 h. A transient expression of interleukin-4 (IL-4) mRNA was observed between 4 and 7 h of stimulation. No gene expression of granulocyte colony stimulating factor (G-CSF) was detected at any time. These results show that anti-CD3 stimulation of MNC leads to a rapid sequential induction of different cytokine mRNA, some with a very transient expression.

Base Sequence↗
PubMed1992

Murine thymocytes with ability to inhibit Il-2 production: I. Genetic differences between mouse strains and characterization of the model system.

An experimental system has been established to understand the poor interleukin-2 (Il-2) production by activated thymocytes. This model system is further characterized here and studies were done on the possible mechanism(s) involved. Thymocytes activated by Concanavalin-A (Con-A), or through the CD3 complex of the T-cell receptor (TCR), inhibit 95-99% of the Il-2 production by spleen cells, while thymocytes stimulated by rIl-2 or lipopolysaccharides (LPS) do not. The mechanism of inhibition is not due to production of soluble factors, consumption of available interleukin-1 (Il-1) or Il-2, but is dependent on cell-to-cell contact. Although cellular contact is needed, cytotoxicity is not involved. Prostaglandin production is not required for the generation or exertion of the inhibitory activity. Protein and DNA synthesis are necessary for exertion of the suppressive effect. We also demonstrate a genetic difference between different mouse strains in the ability to generate the inhibitory thymocytes. Activated Balb/c thymocytes inhibit spleen cells' Il-2 production in a non-MHC-restricted manner. Our studies demonstrate a regulatory capacity of activated thymocytes in vitro. This ability of the postnatal cells could be of relevance for understanding the latter events in T-cell education in the thymus and the role of Il-2 during this process.

Animals↗