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Biomedical subjects

H Hedman

Publications and source records attributed to H Hedman.

At least 19 recordsLinked to original sources
PubMed2003

LRIG1 and epidermal growth factor receptor in renal cell carcinoma: a quantitative RT--PCR and immunohistochemical analysis.

In all, 31 renal cell carcinomas (RCCs) were examined for expression of the potential tumour suppressor LRIG1 (formerly Lig-1) and the epidermal growth factor receptor (EGFR). Eight matched samples of uninvolved kidney cortex were also evaluated. Gene expression was examined by quantitative real-time RT-PCR. In the eight matched sample pairs (uninvolved kidney cortex and tumour), protein expression was examined by immunohistochemistry. Conventional (clear cell) tumours showed an expected upregulation of EGFR. LRIG1 expression was generally downregulated in conventional and papillary RCC but not in chromophobic RCC. The ratio between EGFR and LRIG1 was more than 2.5-fold higher in the eight tumours compared with matched uninvolved kidney cortex and was at least two-fold higher than the mean normal ratio in 21 of 31 samples analysed. The observed downregulation of LRIG1 and increased EGFR/LRIG1 ratios are consistent with LRIG1 being a suppressor of oncogenesis in RCC by counteracting the tumour-promoting properties of EGFR. Further studies are justified to elucidate the explicit role of LRIG1 in the oncogenesis of RCC.

Adult↗
PubMed2002

Rapid induction of long-lasting drug efflux activity in brain vascular endothelial cells but not malignant glioma following irradiation.

The influence of radiotherapy on malignant glioma multidrug resistance to chemotherapy was evaluated because patients with glioma often are treated with a combination of radiotherapy and chemotherapy. Multidrug resistance gene (MDR1, mdr1a, and mdr1b) transcripts were found in human and rat glioma cell lines. P-Glycoprotein (Pgp) was immunohistochemically detected in glioma cell lines and in the rat brain vascular endothelial cell line (RBE4). A multidrug resistance pump efflux activity assay demonstrated increased calcein efflux of RBE4 endothelial cells, but not glioma cells, 2 h after irradiation and still increased 14 d after irradiation. The increased efflux was equally inhibited by verapamil with or without irradiation. In the rat intracranial glioma model (BT4C), Pgp was demonstrated in capillary endothelial cells of the tumor tissue and surrounding normal brain, but not in tumor cells. The expression of gene transcripts or Pgp was not affected by irradiation. The results indicate that long-lasting verapamil-resistant drug efflux mechanisms are activated in brain endothelial cells after irradiation. The results might explain the poor efficacy of chemotherapy following radiotherapy and contribute to consideration of new treatment strategies in the management of malignant glioma.

ATP Binding Cassette Transporter, Subfamily B↗
PubMed2001

Cloning, characterization, and expression of human LIG1.

Growth factor receptors are frequently amplified and over-expressed in various human cancers. Recently, a Drosophila cell surface protein, Kekkon-1, was found to participate in an epidermal growth factor (EGF) driven negative feedback loop. Kekkon-1 is induced by EGF, binds to the EGF-receptor, and inhibits receptor-mediated signaling. Here, we have searched for human genes with homologies to Kekkon-1 and identified human LIG1. The gene is the human homologue of mouse Lig-1 and is located on chromosome band 3p14, a region frequently deleted in various human cancers. It is predicted to encode a transmembrane cell-surface protein with extracellular leucine-rich repeats and immunoglobulin-like domains. LIG1 mRNA was detected in all tissues analyzed. The highest and lowest relative expression levels were found in brain and spleen, respectively, and differed by more than 200-fold. Taken together, our data are compatible with a role for LIG1 as a growth and tumor suppressor in human tissues.

Amino Acid Sequence↗
PubMed2001

Distribution of estramustine-binding protein during postnatal development of rat brain.

Estramustine-binding protein (EMBP) is expressed in several types of brain tumors, such as astrocytoma, ependymoma, and meningioma. It binds the cytotoxic drug estramustine with high affinity and is suggested to cause accumulation of the drug in EMBP-expressing tumor cells. In this study, the spatial distribution of EMBP in normal rat brain was studied with immunohistochemistry. Brains from male and female rats of different ages were used. EMBP was found in the cytoplasm of ependymal cells, in the leptomeninges, mainly the arachnoid, and in scattered neurons. Moreover, staining was seen in nuclei of choroid plexus cells, in the granular cell layer in the cerebellum, and in a few scattered endothelial cells. The nuclear staining was more frequent in younger animals. No obvious difference in EMBP expression between male and female rats was observed. The expression of EMBP in rat brain was confirmed with nested RT-PCR. Future studies are justified to elucidate the role of EMBP-like proteins in CNS and in brain tumors.

Age Factors↗
PubMed2000

Estramustine-binding protein in malignant glioma in rat.

Estramustine is a chemotherapeutic drug, used in the treatment of prostatic carcinoma. In the prostate, it binds specifically to a 46 kDa glycoprotein called estramustine-binding protein (EMBP), which consists of three polypeptide components; C1, C2, and C3, each coded for by a specific gene. Expression of EMBP and binding of estramustine has also been detected in malignant glioma in both rats and humans. Elevated levels of this protein in astrocytoma have proved to correlate with poor prognosis. In the present work, expression of all three polypeptide components of EMBP was confirmed in an orthotopic rat glioma model with nested reverse transcriptase PCR and Western blot (molecular weights of 8, 10, and 12 kDa). Specific binding of estramustine with a Kd of 40 for male and 50 for female rats, and a total number of binding sites of 0.7 and 0.4 pmol/mg proteins for male and female rats respectively, was demonstrated with Scatchard plot analysis. These binding characteristics are similar to those of prostatic EMBP. Further studies to elucidate how EMBP expression affects the effect of estramustine treatment, and its putative prognostic value is of special clinical interest. The confirmation of BMBP expression in BT4C rat glioma demonstrates its suitability as a model system for such studies.

Animals↗
PubMed1999

Role of the leukocyte function antigen-1 conformational state in the process of human immunodeficiency virus type 1-mediated syncytium formation and virus infection.

Human immunodeficiency virus type 1 (HIV-1)-mediated syncytium formation is recognized as being highly dependent on intercellular adhesion molecule (ICAM)-1-leukocyte function-associated molecule 1 (LFA)-1 interaction, whereas the process of infection with cell-free virions is independent of such complementary interaction. Our group has recently demonstrated that an antibody-mediated induction of the high affinity state of LFA-1 for ICAM-1 renders target T cells more prone to HIV-1-dependent syncytium formation and infection by ICAM-1-bearing virions. To further substantiate these results, we made use of mutant cell lines expressing LFA-1 in either a low (parental HPB-ALL and HAmut) or a high affinity state for ICAM-1 (HAP4) and compared syncytium formation and virus infection. Cells expressing the activated form of LFA-1 were found to be more susceptible to HIV-1-induced syncytium formation and to infection by ICAM-1-bearing HIV-1 particles. The observed increase was solely due to the LFA-1 activation state because it was abrogated by anti-LFA-1 or anti-ICAM-1 antibodies and not due to variations in surface expression of LFA-1, CD4, or the chemokine coreceptor CXCR4. However, a linear relation was seen between the level of CXCR4 surface expression and susceptibility to syncytium formation/virus infection when ICAM-1-LFA-1 interaction was either absent (i.e., infection with ICAM-1-negative virions) or abrogated (treatment with anti-LFA-1 or anti-ICAM-1 antibodies). These results emphasize the important role of the LFA-1 activation state with respect to virus-induced syncytium formation and HIV-1 infection.

Cell Line↗
PubMed1997

Defective expression of beta 1-integrins in cells with constitutively active alpha L beta 2-integrins.

We have investigated a potential relationship between expression of beta 1-integrins and adhesiveness of the beta 2-integrin LFA-1 (alpha L beta 2, CD11a/CD18). By an approach of random mutagenesis and selection we established clones from the human acute lymphatic leukemia cell line HPB-ALL with (i) constitutively active LFA-1 and (ii) with no apparent integrin-beta 1 cell surface expression. Thirty-seven of 42 clones selected for activated LFA-1 were found to have lost apparent integrin-beta 1 expression. Conversely, 7 of 21 clones selected for lack of beta 1 expression were found to have activated LFA-1. Since this pointed toward a possible coupling between beta 1 expression and LFA-1 activity, we further analyzed at which level beta 1 expression was blocked. We focused on one clone, HAP4, with activated LFA-1 and no detectable beta 1 cell surface expression and found, surprisingly, that it expressed wild-type levels of beta 1 mRNA and, in Western blots of whole cell lysates, apparently normal levels of beta 1 protein. However, in addition to beta 1 of the expected molecular weight, HAP4 expressed a unique 48-kDa band recognized by the polyclonal anti-beta 1 antiserum. Immunoprecipitation experiments revealed that the epitope recognized by the anti-beta 1 antibody 4B4 was hidden or lost. The alpha 4-chain was found in its precursor form but it did not associate with any beta-chain, and it was not processed to its mature form. Instead alpha 4-chains were eventually degraded. Taken together this showed that beta 1-chains were produced but not properly processed in HAP4. From this we propose that HAP4 is deficient in a gene product required both for proper beta 1 folding and for repression of LFA-1 adhesiveness.

Clone Cells↗
PubMed1996

Generation of mutant cells with constitutively active beta 2-integrin LFA-1.

To elucidate the molecular mechanisms behind regulation of integrin adhesiveness, we have developed a system for the generation of mutant cell lines where the adhesive functions of specific molecules were modulated. The human lymphatic leukemia cell line HPB-ALL was chemically mutagenized and rare cells expressing constitutively active beta 2-integrin LFA-1 (alpha L beta 2, CD11a/CD18) were selected from the pool of mutagenized cells, by repeated cycles of binding to immobilized ICAM-1. The selected population showed 20-fold more binding to ICAM-1 compared to the starting population. A sub-clone of the selected population, termed HAP4, was further analyzed. In contrast to the original cell line, HAP4 had constitutively active LFA-1 and adherence to ICAM-1 was insensitive to inhibition by okadaic acid and cytochalasin B or to inhibition of ATP synthesis. The activation of LFA-1 appeared specific since the cells did not show a general increase in adhesiveness. No major aberration in cell surface carbohydrate expression was found and the expression of cell surface molecules including CD43, CD4, CD8, and MHC class I and II was normal. However, apparent cell surface expression of integrin beta 1 chain was lost and alpha 4 chain was greatly diminished. Our study demonstrates the feasibility of generating cells with mutated adhesive functions and the characterized cells will serve as tools to further dissect the mechanisms underlying integrin regulation.

CD18 Antigens↗
PubMed1996

Regulation of alpha 4 integrin avidity in human B cells: requirement for dephosphorylation events for high avidity VCAM-1 binding.

The authors compared the phosphorylation-dependent signal transduction pathways involved in the regulation of adhesiveness of integrin LFA-1 with that of alpha 4 integrins binding to VCAM-1. The authors developed an in vitro method to monitor changes in adhesiveness using a VCAM-1 fusion protein coupled to magnetic beads. For LFA-1, a similar method has previously been established using an ICAM-1 fusion protein. Binding of cells was monitored and found to be strictly integrin alpha 4 and VCAM-1 dependent. The serine/threonine phosphatase inhibitors okadaic acid and calyculin A were equally potent in inhibiting binding to VCAM-1 as to ICAM-1, and inhibition of protein phosphatase-1 (PP1) is proposed to be the important denominator. Similarly, the phorbol ester PDBu, potentially stimulating protein phosphatase-1 and staurosporine, an inhibitor of serine/threonine kinases, enhanced adhesion to VCAM-1 as has previously been shown for ICAM-1. A major difference was that a significant portion of the binding to VCAM-1 was not susceptible to inhibition by drugs while binding to ICAM-1 could be completely inhibited. We propose that the adhesiveness of the alpha 4 integrins for VCAM-1 and of LFA-1 for ICAM-1 is regulated by similar or identical protein kinases and phosphatases.

B-Lymphocytes↗
PubMed1995

The development and course of test reactions to gold sodium thiosulfate.

In our department, gold sodium thiosulfate has become the 2nd most common allergen in routinely patch tested dermatitis patients, with a rate around 10%. Test reactions to this compound often appear late, sometimes so late that active sensitization may be suspected. This study was performed to study the time course of the allergic reaction to gold sodium thiosulfate and to elucidate whether late test reactions mean active sensitization. 10 patients with contact allergy to gold sodium thiosulfate (0.5% pet.) were retested epicutaneously (e.c.) and intracutaneously (i.c.) with dilution series. The clinical course was followed for 2 months with initially short intervals, later more extended. During the entire study, 26 positive e.c. reactions were diagnosed. Within the 1st week, 17 (65%) were recorded. 12 reactions (46% of 26) were noted at the ordinary reading, 3 days after test application. After 10 days, another 9 reactions (35%) appeared. The patients with the latter reactions also had positive test reactions within the 1st week. After 2 months, 9 reactions remained. Out of 30 i.c. tests applied, 25 became positive within 1 week. 19 (76%) of these reactions changed in morphology from thin infiltrates to deep nodules. Another 4 nodules appeared in patients with previous negative i.c. tests. All 23 nodules remained after 2 months. E.c. and i.c. test reactions to gold sodium thiosulfate are long-lasting. Positive patch test reactions emerging after 10 days do not automatically imply active sensitization. To diagnose contact allergy to gold sodium thiosulfate, the ordinary reading at day 3 is insufficient; even reading at 1 week is insufficient and must be supplemented by a reading at 3 weeks. All the i.c. test reactions, however, appeared within 1 week and, in several, a dermal nodule was formed.

Allergens↗
PubMed1995

Evaluation of an organ-donor-card campaign in Sweden.

One of the aims of this study was to evaluate an information campaign carried out in three geographical areas of Sweden in the winter of 1992-93. The campaign was intended to increase public awareness of organ donation and to increase the signing of donor cards. Another objective was to test the effects of different kinds of information. These were: A) an extensive "package" of information including training of key groups, lecturing at meetings and exhibitions, and advertisements of donor cards: B) a brochure to households including two donor cards; and C) a combination of A and B. Yet another aim was to reassess public opinion on transplantation issues, which had been surveyed before in 1987, 1988, and 1990. Random samples of the population in three campaign areas and a control sample were surveyed before and after the campaign, altogether 5600 persons. The average response rate was 69% (1992) and 68% (1993). In the two areas where the brochure had been distributed to the households, the rate of donor card holders had more than doubled (from 3% and 5% to 13% and 12%). In the two areas where the brochure had not been distributed, the rate was unchanged (5%). In the "brochure areas" also a somewhat larger number of people had informed their relatives about their decisions, compared with people in the other areas. In all campaign areas considerably more people were aware of the cards than in the control area. No attitude changes could be shown in any area.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1994

Monocyte vesiculation is a possible mechanism for dissemination of membrane-associated procoagulant activities and adhesion molecules after stimulation by lipopolysaccharide.

Endotoxin-stimulated monocytes can elicit a dual procoagulant response. They express tissue factor and expose phosphatidylserine in the outer leaflet of the plasma membrane. Tissue factor, a membrane glycoprotein, is the cellular trigger of blood coagulation reactions. Phosphatidylserine is an essential anionic phospholipid for surface amplification of thrombin generation. In this study the distribution of these two procoagulant entities between activated monocytes and derived microparticles was assessed after stimulation by LPS. The presence of CD14, CD11a, and CD18, and possible associated adhesion potential were examined, particularly on microparticles. Tissue factor was evidenced by using a specific functional assay and flow cytometry. Phosphatidylserine exposure was monitored through its catalytic activity in a thrombin generation assay and by flow cytometry with the use of FITC-conjugated annexin V, a protein probe of anionic phospholipids. CD14, CD11a, and CD18 were detected by flow cytometry. The interaction of microparticle CD11a/CD18 with intracellular adhesion molecule-1 was demonstrated by using immobilized recombinant intracellular adhesion molecule-1 fusion protein. The major part of tissue factor and phosphatidylserine-dependent procoagulant activity was associated with microparticles after LPS stimulation. This was confirmed by flow cytometry. The presence of functional CD11a/CD18, and CD14 on microparticles testifies to an associated adhesion potential. These results show that membrane vesiculation could be responsible for dissemination of inducible monocyte procoagulant activities and suggest that derived microparticles could also participate in endothelium stimulation. This emphasizes the role of monocyte as a central element in the coupling between inflammation/infection and thrombosis.

Annexin A5↗
PubMed1992

Regulation of LFA-1 avidity in human B cells. Requirements for dephosphorylation events for high avidity ICAM-1 binding.

Regulation of the avidity of LFA-1 (CD11a/CD18, alpha L beta 2) for its ligand ICAM-1 (CD54) was studied in human B cells by evaluating the effects of a phorbol ester, anti-IgM antibodies, staurosporine, and okadaic acid. We monitored changes in LFA-1 avidity by quantifying binding of cells to an immobilized rICAM-1 fusion protein. In this assay, the protein kinase C-activating phorbol ester PDB and anti-IgM antibodies, as well as the protein kinase inhibitor, staurosporine, were able to induce LFA-1-dependent binding to ICAM-1. This demonstrates that the high avidity state of LFA-1 can be induced by a protein kinase C-dependent and by a protein kinase C-independent pathway. Furthermore, treatment of the cells with the protein phosphatase inhibitor, okadaic acid, inhibited binding to ICAM-1. Treatment with staurosporine before addition of okadaic acid not only induced enhanced binding of cells to ICAM-1, but also dramatically reduced the ability of okadaic acid to inhibit binding. These results suggest a critical role for a protein phosphatase in inducing the high avidity state of LFA-1 as well as a role for a protein kinase in inducing the low avidity state of LFA-1.

Alkaloids↗
PubMed1992

Stimulation of human naive and memory T helper cells with bacterial superantigen. Naive CD4+45RA+ T cells require a costimulatory signal mediated through the LFA-1/ICAM-1 pathway.

The role of the accessory molecule ICAM-1 in activation of subpopulations of human T cells was examined using the bacterial superantigen staphylococcal enterotoxin A (SEA) as a MHC class II and TCR-dependent polyclonal T cell activator. Human T cells responded with different sensitivity to SEA when presented on mouse accessory cells expressing a human transfected MHC class II gene product. Mouse L cells cotransfected with both MHC class II (DR2A or DR7) and ICAM-1-stimulated T cells at 100-fold lower concentrations of SEA as compared to the single transfected cells. mAb reacting with the CD11a, CD18, or ICAM-1 molecules efficiently inhibited T cell activation with the cotransfected HLA-DR2A/ICAM-1 cell but did not influence T cell activation with the HLA-DR2A single transfected cell. Analysis of the ICAM-1 requirement on CD4+ memory (CD4+45RO+) and naive (CD4+45RA+) T cells revealed that CD4+45RA+ naive Th cells were hyporesponsive to SEA-induced activation with the HLA-DR2A single transfectant. However, cotransfection of ICAM-1 enabled these cells to respond to low doses of SEA implicating that they are more dependent on accessory molecules than the CD4+45RO+ cells. rICAM-1 immobilized on a plastic surface, was able to strongly costimulate SEA-induced T cell activation with the HLA-DR2A single transfectant, suggesting that costimulatory signals mediated to the T cells through LFA-1 can be delivered physically separated from the TCR signal. CD4+45RO+ memory and CD4+45RA+ naive Th cells apparently differ in their capacities to be activated by SEA bound to HLA-DR. Although the TCR molecule densities are similar in these two subsets, costimulation with ICAM-1 is required for activation of the CD4+45RA+, but not the CD4+45RO+ T cell subset at 1 to 10,000 ng/ml concentrations of SEA. This observation indicates different activation thresholds of naive and memory Th cells when triggering the TCR over a wide dose interval of superantigen.

Antigens, Bacterial↗
PubMed1992

Physical separation of ICAM-1 binding cells.

Studies of interactions between specific molecules involved in adhesion between cells are often complicated since cells possess many different types of surface molecules influencing adhesion. To be able to further study interactions between the adhesive cell surface molecule LFA-1 and its ligand ICAM-1, we have produced an ICAM-1 fusion protein in which the extracellular part of ICAM-1 has been fused to a part of an IgG heavy chain. The fusion protein was produced by CHO cells and was easily purified in large quantities from the cell culture supernatant. The protein was coupled to magnetic beads and was shown to have binding characteristics similar to native ICAM-1. The coupled beads were used for magnetic isolation of ICAM-1 binding cells thereby providing a method for detecting changes in avidity for ICAM-1 and also for the analysis of ICAM-1 binding cells.

Amino Acid Sequence↗