Biomedical subjects

G V Alm

Publications and source records attributed to G V Alm.

At least 19 recordsLinked to original sources

PubMed1992

A sensitive immunoassay for porcine interferon-alpha.

Two murine monoclonal antibodies (mAbs) directed against different epitopes on recombinant porcine interferon-alpha (IFN-alpha) were selected and used to construct a two-site ELISA. This ELISA, when performed in a one-step version, detected about 0.5 units ml-1 of IFN-alpha and showed similar sensitivity but better precision than a cytopathic effect inhibition bioassay. Estimates of IFN-alpha in tissue culture medium by the two assays correlated well. In contrast, one or several factors in porcine serum reduced the sensitivity of the ELISA. Measurements of IFN-alpha in porcine serum was, however, possible in a two-step version of the ELISA, with a sensitivity of about 1 unit IFN-alpha ml-1. Results of ELISA and bioassay agreed, except that the ELISA possibly produced false positive results in two out of a total of 91 sera negative in the bioassay. In addition, one of 23 sera positive in the bioassay was negative in the ELISA.

Animals↗

PubMed1992

Characterization of anti-interferon-alpha antibodies appearing during recombinant interferon-alpha 2a treatment.

Patients with malignant midgut carcinoid tumours received recombinant interferon-alpha 2a (rIFN-alpha 2a) or rIFN-alpha 2a and chemotherapy (streptozocin and doxorubicin) for 6 months, and then rIFN-alpha 2a alone. Antibodies, mainly of IgG type, binding to rIFN-alpha 2a developed in nine of 22 patients (41%), as determined by immunoassay. In seven patients, antibodies also neutralized the biologic (anti-viral) activity of rIFN-alpha 2a. Anti-IFN-alpha 2a antibodies were equally frequent in both sexes and treatment groups, but were not observed in those patients (n = 8) that had previously received other types of IFN. Antibodies appeared after a median of 6 months of rIFN-alpha 2a treatment and had a median duration of 6 months. The anti-IFN-alpha 2a antibody titres declined with time with no obvious relation to change of therapy, also during continued IFN-alpha 2a treatment. High titres of neutralizing antibodies appeared to impair anti-tumoural effects in individual potential responders. Anti-IFN-alpha 2a antibodies further examined in six patients bound to native IFN-alpha subtypes present in both allogenic and autologous leucocyte IFN-alpha. Such autoantibodies neutralized the biologic activity of autologous IFN-alpha in two patients, and in a third were partially neutralizing.

Aged↗

PubMed1992

Interleukin-4 down-regulates Sendai virus-induced production of interferon-alpha and -beta in human peripheral blood monocytes in vitro.

Recombinant interleukin-4 (rIL-4) caused a 65-70% reduction of the interferon-alpha (IFN-alpha) and -beta production induced by Sendai virus (SV) in human peripheral blood monocytes in vitro. Significant inhibition was seen at concentrations as low as 0.1 ng/ml. The rIL-4 also reduced levels of IFN-alpha and -beta mRNA by 60% and 67%, respectively, as well as the frequency of IFN-alpha and -beta mRNA-containing cells by 65% and 54%, respectively. The frequency of IFN-alpha/beta mRNA-containing cells was inhibited by rIL-4 throughout the whole course of induction by SV. IL-4 caused a shift of the grain count distribution towards less heavily labelled cells, suggesting an inhibitory effect of rIL-4 on most IFN-alpha/beta mRNA-containing cells. Antibodies to rIL-4 did not influence the normal IFN-alpha/beta response induced by SV, but abolished the inhibitory effect of the rIL-4. When rIL-4 was added to cells 4 h after start of stimulation by SV, at which time much mRNA has accumulated but little IFN-alpha/beta has been secreted, no inhibition of the IFN-alpha/beta production by rIL-4 was seen. IFN-gamma had only a minor reversing effect on the rIL-4 inhibition, but if cells were precultivated in medium with or without IFN-gamma for 6 h before SV induction, rIL-4 paradoxically enhanced the IFN-alpha/beta response. Our results suggest that rIL-4 inhibits an early step of IFN-alpha/beta induction in monocytes, at the level either of transcription of IFN-alpha/beta genes or of the processing or stability of mRNA. The IL-4 effects may however depend on the state of activation of the monocytes.

Actins↗

PubMed1992

Interferon-alpha but not -beta genes require de novo protein synthesis for efficient expression in human monocytes.

Monocytes produce interferon-alpha (IFN)-alpha) and -beta when human peripheral blood mononuclear cells (PBMCs) are stimulated in vitro by Sendai virus (SV). We found that about 70% of the IFN-producing cells (IPCs) expressed both IFN-alpha and -beta mRNA; the rest expressed only IFN-beta mRNA. In the presence of the protein synthesis inhibitor cycloheximide (CHX), the frequency of IFN-alpha mRNA-containing cells, measured after 6h, was decreased by 85-90%. Results of nuclear run-on transcription assays showed that CHX inhibited IFN-alpha gene expression. The frequency of IFN-beta mRNA-containing cells was not reduced by CHX. Actually, a threefold increase was observed at the lower CHX concentrations. Studies on the kinetics of IFN-alpha/beta mRNA induction showed that CHX accelerated the appearance of IFN-beta mRNA-containing cells, increased IFN-beta mRNA levels, and delayed the normally occurring post-inductional decrease of IFN-beta mRNA. Unexpectedly, an initially normal or even accelerated IFN-alpha mRNA response was seen in the presence of CHX during the first 3-4 h after SV stimulation. This occurred in a small proportion of the potential IPCs. However, CHX prevented the subsequent marked increase of IFN-alpha mRNA levels. Preincubation of PBMCs for 6 h in conditioned medium (CM) containing IFN and other cytokines prevented the CHX-mediated inhibition of IFN-alpha mRNA. Without preincubation this was not seen. The preincubation in CM caused an accelerated appearance of IFN-alpha mRNA, resembling that of IFN-beta mRNA. The results suggest that IFN-alpha and -beta genes are differentially regulated in the same monocytes, the former requiring de novo synthesis of intracellular protein(s) for efficient expression.

Blotting, Northern↗

PubMed1991

Phorbol ester-mediated inhibition of IFN-alpha/beta gene transcription in blood mononuclear leukocytes.

The phorbol ester PMA efficiently inhibits the in vitro IFN-alpha and -beta responses in human blood monocytes induced by Sendai virus and in natural IFN-producing cells induced by glutaraldehyde-fixed herpes simplex virus-infected human amnion (WISH) cells. This PMA-mediated inhibition of IFN-alpha/beta secretion is correlated with considerably reduced levels of IFN-alpha/beta mRNA, as determined by Northern blot analysis. Nuclear run-on assays further show that, at least in monocytes, PMA inhibits transcription of the IFN-alpha and -beta genes. The synthetic diacylglycerol 1-oleoyl-2-acetylglycerol has the same effects as PMA, and the protein kinase inhibitor staurosporine prevents the PMA-mediated inhibition of IFN-alpha/beta expression. These results suggest a role for protein kinase C in the inhibition of IFN-alpha/beta responses. The PMA does not inhibit the accumulation of IFN-beta mRNA in monocytes stimulated by Sendai virus in the presence of cycloheximide, indicating that the inhibitory action of the phorbol ester requires de novo protein synthesis.

Alkaloids↗

PubMed1991

Autoimmunity after alpha-interferon therapy for malignant carcinoid tumors.

OBJECTIVE: To determine the incidence of autoantibodies and autoimmune disease and their influence on therapeutic results during alpha-interferon treatment in patients with malignant midgut carcinoid tumors. DESIGN: Consecutive sample of patients. SETTING: University hospital. PATIENTS: One hundred thirty-five patients (70 women, 65 men; median age, 59 years) with biopsy-proven tumors, liver metastases, and no autoimmune disease. INTERVENTIONS: Leukocyte alpha-interferon (n = 88) or alpha-interferon 2b (n = 47) three times a week. MAIN OUTCOME MEASURES: Signs and symptoms of autoimmune disease or development of autoantibodies to thyroid antigens, nuclear antigens, or gastric parietal cells. Tumor responses were determined by reduced liver metastases or reduced urinary 5-hydroxyindole acetic acid excretion, or both. RESULTS: Twenty-five patients (19%) developed the following autoimmune disorders after a median of 9 months of therapy: thyroid disease (n = 18), systemic lupus erythematosus (n = 1), pernicious anemia (n = 4), and vasculitis (n = 2). Antibodies to microsomal thyroid antigen or thyroglobulin were detected in 16 patients before therapy and in another 11 patients during therapy. Antinuclear antibodies were detected in 16 patients before and in another 19 patients during therapy. Clinical thyroid disease developed in more than 60% of patients who had or developed thyroid antibodies but in only 7% of initially autoantibody-negative patients. Autoimmunity did not correlate with objective tumor response. CONCLUSION: Patients with malignant carcinoid tumors may develop autoimmune disease during alpha-interferon therapy, especially when autoantibodies are present. They should therefore be monitored for autoimmunity, which does not appear, however, to influence tumor responses.

Adult↗

PubMed1991

The induction of interferon-alpha and interferon-beta mRNA in human natural interferon-producing blood leukocytes requires de novo protein synthesis.

The induction of interferon-alpha (IFN-alpha) and IFN-beta mRNA in natural IFN producing (NIP) cells in cultures of human peripheral blood mononuclear cells (PBMCs), stimulated by glutaraldehyde-fixed Herpes simplex virus type 1 (HSV)-infected WISH cells, was studied. The protein synthesis inhibitor cycloheximide (CHX) totally prevented the appearance of both IFN-alpha and IFN-beta mRNA, also in cultures supplemented with a conditioned medium (CM) assumed to contain soluble factors necessary for the IFN induction. However, when PBMCs were preincubated for 4 h in medium supplemented with fetal bovine serum (FBS) with or without addition of CM, the subsequent induction of IFN-alpha/beta mRNA became partially resistant to CHX. In serum-free medium containing interleukin-3 (IL-3) or granulocyte-macrophage colony-stimulating factor (GM-CSF), the early induction of IFN-alpha mRNA became resistant to CHX, and, in contrast to FBS and CM supplemented medium, this was observed also without a preincubation of the PBMCs. In contrast, IL-1, IL-2, IL-4, IL-6, tumor necrosis factor-alpha (TNF-alpha), IFN-alpha, or IFN-gamma had no such effects. Our results suggests that de novo synthesis of proteins normally is required for the induction of IFN-alpha/beta mRNA. Such proteins might be cytokines, possibly CSFs, which in turn also may require protein synthesis for their actions. In contrast, the actual triggering signal provided by the HSV-inducer is independent of protein synthesis.

Culture Media↗

PubMed1991

Interferons and the colony-stimulating factors IL-3 and GM-CSF enhance the IFN-alpha response in human blood leucocytes induced by herpes simplex virus.

Human peripheral blood mononuclear cells (PBMC) were stimulated to produce interferon-alpha (IFN-alpha) by glutaraldehyde-fixed Herpes simplex virus type 1 (HSV)-infected WISH amnion cells in vitro. Different cytokines were included during the stimulation and tested for their ability to enhance the IFN-alpha response which occurs in the natural IFN-alpha producing (NIP) leucocytes. The total production of IFN-alpha and the numbers of IFN-alpha producing cells (IPCs) were increased by interleukin-3 (IL-3) or granulocyte-macrophage colony-stimulating factor (GM-CSF). Their most marked effect was to reduce the time required for induction of the IPC by the HSV-infected cells, thereby causing both an earlier peak of IPC numbers and secretion of IFN-alpha. Addition of IFN-alpha 2b did not alter the kinetics of the IFN-alpha response in the same way as the two CSFs, but instead generally increased the IPC numbers and the production of IFN-alpha. The IL-3 and GM-CSF, especially in combination with IFN-alpha, had the most pronounced enhancing effects on IPC numbers when PBMC were induced at low cell concentrations. The cytokines IL-1, IL-2, IL-4, IL-6 or tumour necrosis factor-alpha (TNF-alpha) had no detectable effects on the IFN-alpha response. The results suggest that cytokines such as the CSFs and IFNs may be involved in the regulation of NIP cell functions.

Cells, Cultured↗

PubMed1991

Flow cytometric analysis of natural interferon-alpha producing cells.

Herpes simplex virus-infected cells induce high interferon-alpha (IFN-alpha) production in infrequent cells among peripheral blood mononuclear cells (PBMC), designated natural IFN-alpha producing (NIP) cells. The properties of such NIP cells were compared with defined populations of leucocytes by means of flow cytometric analysis and sorting. The NIP cells are characterized as a discrete population of cells with high forward and low to intermediate orthogonal light scattering, similar to that of early progenitors of myeloid and lymphoid cells. However, they appear to lack the stem cell-associated molecule CD34. Furthermore, NIP cells cannot be localized to the myeloid line of cell differentiation, because they do not express the CD33, CD13, CD11b, CD15 or CD14 antigens. Neither do they express CD10 and CD19 antigens which are present in all stages of B-cell differentiation plasma cells excepted, nor CD7 antigens expressed on early T cells. In combination with previous results, our data support the view that the NIP cell is a unique and distinct cell type in peripheral blood, possibly with a physiological role in the defence against certain viral infections.

Antigens, CD↗

PubMed1991

Autoimmune phenomena in patients with malignant carcinoid tumors during interferon-alpha treatment.

Several previous reports suggest an association between treatment of patients with interferon-alpha (IFN-alpha) and development of autoantibodies and autoimmune disease. We here summarize the experience from a group of 135 patients with midgut carcinoid tumors treated with natural leukocyte IFN-alpha or recombinant IFN-alpha (rIFN-alpha). An unusual high incidence of antimicrosomal antibodies (MsAb) or anti-thyroglobulin antibodies (TgAb) and thyroid disease manifested as hyperthyroidism, hypothyroidism or a biphasic Hashimoto-like disease was seen, with female predominance. The incidence of antinuclear antibodies (ANA) was also increased, but equally in both sexes. Antibodies to parietal cells were found in 5 cases and 4 patients with pernicious anemia were detected. Two patients developed vasculitis of leukocytoclastic type and one a syndrome resembling systemic lupus erythematosus. Some patients treated with rIFN-alpha develop anti-IFN antibodies. Such antibodies may also be autoantibodies reacting with autologous IFN-alpha. They can neutralize the biologic activity of administrated IFN preparation and cause therapeutic failure. The implications of the various autoimmune manifestations during IFN-alpha treatment are discussed.

Autoantibodies↗

PubMed1990

A distinct population of nonphagocytic and low level CD4+ null lymphocytes produce IFN-alpha after stimulation by herpes simplex virus-infected cells.

Human PBMC were stimulated for 6 h in vitro by HSV or Sendai virus (SV) and analyzed by flow cytometry. IFN-alpha producing cells (IPC) were identified through their content of IFN-alpha mRNA by in situ hybridization using a 35S-labeled IFN-alpha 2 cRNA probe. The IPC induced by HSV-infected WISH cells lacked capacity to adhere to and phagocytose latex particles. The induction of IFN-alpha by free infectious SV occurring in monocytes was abolished by phagocytosis of latex particles present in the cultures during the induction period. Such latex particles actually enhanced the IFN-alpha response induced by glutaraldehyde-fixed HSV- or SV-infected WISH cells or by free intact HSV. The HSV-induced IPC did not express the CD14 Ag expressed on monocytes. Cell sorting was performed on HSV-induced PBMC labeled with phycoerythrin-conjugated anti-CD3 and FITC-conjugated anti-CD4 mAb. A small population consisting of 1.4% of all PBMC, which was CD3- but expressed low but significant levels of CD4, contained the majority of the IPC with a 50-fold increase of their frequency. This cell population had a forward- and right-angle light scatter different from typical monocytes/macrophages. The results therefore further delineate IPC among PBMC into monocytes, being stimulated by viruses such as SV. Another distinct population of infrequent but highly efficient IPC, tentatively designated natural IFN-alpha producing cells, is activated by stimuli such as HSV.

Antibodies, Monoclonal↗

PubMed1990

Infrequent but efficient interferon-alpha-producing human mononuclear leukocytes induced by herpes simplex virus in vitro studied by immuno-plaque and limiting dilution assays.

Human blood mononuclear leukocytes (PBMCs) producing interferon-alpha (IFN-alpha) after stimulation by herpes simplex virus type 1 (HSV) in vitro were identified by a filter immuno-plaque assay. Individual IFN-alpha-producing cells (IPCs) yielded between 0.5 and 2 units IFN-alpha, sufficient to protect cultures of MDBK cells against a viral challenge. Therefore, their frequency could be determined by a limiting dilution assay as well as by an immuno-plaque assay. Similar estimates of between 2 and 55 IPCs per 10(4) PBMCs at the peak of the IFN-alpha response were obtained by the two methods. IPCs were first detected 3 h after stimulation by HSV; their number peaked at 8 h and then declined. IPC frequencies were influenced by the concentrations of HSV and PBMCs during induction, but the quantity of IFN-alpha produced per IPC was relatively constant. The relation between the numbers of IPCs and PBMCs was linear at high PBMC concentrations, whereas at low PBMC concentrations fewer IPCs than expected were detected. The response could be fully restored by adding a combination of filler cells (Namalwa or U937 cells) and conditioned medium from 6-h HSV-induced PBMC cultures. Our results suggest that HSV induces an IFN-alpha response in a relatively rare population of efficient IPCs by complex mechanisms, which may involve cell cooperation and/or production of soluble factors.

Cells, Cultured↗

PubMed1990

Possible induction of systemic lupus erythematosus by interferon-alpha treatment in a patient with a malignant carcinoid tumour.

Interferon-alpha (IFN-alpha) is currently used in the treatment of various malignant tumours. Development of different autoimmune disorders has been reported in some patients during IFN-alpha therapy. Systemic lupus erythematosus (SLE) after treatment with IFN-alpha has not been described, although a majority of SLE patients have demonstrable serum levels of IFN-alpha, which correlate with disease activity and have been suggested to be of pathogenetic significance. In this paper we describe a patient with a malignant carcinoid tumour who developed a SLE-like syndrome during treatment with leucocyte IFN-alpha. The patient developed myalgia and low grade arthritis in multiple joints together with a high titre of antinuclear antibodies (ANA) and anti-dsDNA antibodies. After the treatment was stopped, the symptoms subsided although a moderate ANA titre persisted. However, the tumour continued to regress despite cessation of IFN-alpha therapy. During a short course with recombinant IFN-alpha the syndrome relapsed, supporting the concept that the SLE syndrome was precipitated by IFN-alpha. A connection between IFN-alpha treatment, the induced autoimmune disorder and regression of the carcinoid tumour is suggested.

Adult↗

PubMed1990

Deficient herpes simplex virus-induced interferon-alpha production by blood leukocytes of preterm and term newborn infants.

The ability of peripheral blood mononuclear cells (PBMC) from newborn infants, gestational age 24-42 wk, to produce interferon-alpha (IFN-alpha) on the first day after birth was studied in vitro. Human amnion cells (WISH) coated with herpes simplex virus type I and fixed by glutaraldehyde were used as IFN-alpha inducers. Individual IFN-alpha producing cells (IPC) among PBMC were determined by an immunoplaque assay. The frequency of IPC was low in all premature (less than or equal to 36 wk) infants (median 0.3 IPC/10(4) PBMC, range 0.0-2.6), and significantly higher (median 2.0 IPC/10(4) PBMC, range 0.0-16.4) in term infants (greater than 37 wk). The frequencies were lower in both groups of infants than in adults (7.3 IPC/10(4) PBMC, range 2.0-23.7). When a conditioned medium from cultures of herpes simplex virus type I-stimulated PBMC from adults was added to the IFN induction cultures, the frequencies of IPC increased in PBMC from both preterm and term infants, and in the latter group did not differ significantly from adult levels. The median production of IFN-alpha per IPC was 1.1 U (range 0.0-2.8) in premature infants, 1.0 U (range 0.0-8.8) in term infants and 3.2 U (range 1.5-8.0) in adults. When concentrations of PBMC in the cultures [corrected] were decreased, a decline of IPC frequencies occurred. This decline was more marked and started at higher PBMC concentrations in infants than in adults, and was prevented by addition of conditioned medium from herpes simplex virus type I-stimulated cultures of PBMC from adults.(ABSTRACT TRUNCATED AT 250 WORDS)

Humans↗

PubMed1989

Induction of mRNA for HLA-DR beta in human keratinocytes cocultured with interferon-gamma.

Explanted human keratinocytes exposed in vitro to recombinant interferon-gamma (IFN-gamma) were investigated for the appearance of mRNA for HLA-DR. Using in situ hybridization with a (35S)UTP-labelled HLA-DR beta cRNA probe, mRNA-positive cells were detected already within 6 h with maximal numbers of positive cells as well as the amount of mRNA per cell after 48 h. The corresponding protein HLA-DR, as analysed by immunoperoxidase staining, was detected on 20%-40% of the cells after 24 h and on almost all cells within 48 h. The expression of HLA-DQ and -DP antigens were always exceeded by that of HLA-DR. Whereas an increase in the concentration of IFN-gamma above 50 U/ml did not affect the maximal level of HLA-DR reactive cells, there was a fourfold increase in the frequency of cells reactive with HLA-DQ and a twofold increase for HLA-DP when the IFN-gamma concentration was raised from 50 to 500 U/ml. When IFN-gamma was withdrawn from the cultures, HLA-DR mRNA and protein synthesis ceased--indicating the continuous need for IFN-gamma to maintain the HLA-DR synthesis in keratinocytes.

DNA↗

PubMed1989

Appearance of interferon-alpha in serum and signs of reduced immune function in pigs after transport and installation in a fattening farm.

Pigs were transported from several breeding facilities at the age of 10-12 weeks and regrouped in a fattening farm, specialized in breeding pigs for subsequent slaughter. Blood samples were obtained from the animals just before transport and daily for 17 days after installation in the fattening farm. On each occasion a group of ten animals (170 animals in total) was sampled. The levels of interferon-alpha (IFN-alpha) in serum were measured as antiviral activity in a cytopathic effect inhibition assay. Beginning at day 4 after installation, a significant proportion of sera contained IFN-alpha, with the highest incidence of IFN-alpha positive animals (25%) and IFN-alpha titers on days 5-10. This indicates a high frequency of viral infections in the animals. The in vitro ability of peripheral blood mononuclear leukocytes (PBMCs) to produce IFN-alpha after stimulation by glutaraldehyde-fixed pseudorabies virus-infected PK15 cells and their proliferative response to the T-cell mitogen leukoagglutinin (LA) was also monitored. There was a significant, but moderate decrease in the ability of PBMCs to produce IFN-alpha during the observation period. In contrast, the response to the mitogen LA decreased markedly during the first 5 days, and thereafter remained at the same low level. The proliferative response to LA was significantly lower for PBMCs from serum of IFN-alpha-positive than from IFN-alpha-negative animals. These impaired PBMC responses could indicate a stress-induced immune depression, possibly contributing to the high incidence of viral infections.

Animal Husbandry↗

PubMed1989

Characterization of the blood mononuclear leucocytes producing alpha interferon after stimulation with herpes simplex virus in vitro, by means of combined immunohistochemical staining and in situ RNA-RNA hybridization.

A procedure is described for combined immunohistochemical staining and in situ RNA-RNA hybridization of human peripheral blood mononuclear leucocytes (PBMC). These cells were first stimulated in vitro by herpes simplex virus (HSV)-infected fibroblasts and after 6 h fixed and stained with a panel of antibodies against differentiation antigens. Alpha interferon (IFN-alpha)-producing cells (IPC) were identified by in situ hybridization by means of a 35S-labelled IFN-alpha 2 cRNA probe. The IPC were infrequent, one in 200-5000 PBMC, but heavily labelled with the cRNA probe. They lacked antigens typical of T and B lymphocytes, and were also essentially negative for the Leu-M5 antigen, present on a majority of monocytes. However, 50% of IPC expressed OKM5 antigens, corresponding to the thrombospondin receptor. The IPC lacked the antigens present on null lymphocytes detected by OKT16, but most of them expressed HLA-DR, -DP and -DQ antigens. The IPC may represent a small subpopulation in the monocyte/macrophage lineage, resembling cells described as antigen presenting and stimulators of autologous mixed lymphocyte reactions. Alternatively, they constitute a subpopulation among the null lymphocytes.

Antibodies, Monoclonal↗

PubMed1989

Interferons.

The interesting and complex biology of the interferon (IFN) system is briefly reviewed. Some encouraging results from the application of IFNs to therapy of human disease, as well as therapeutic problems are also pointed out. It is likely that increased knowledge of the functions of the IFN system will facilitate development of more efficient therapeutic strategies, involving e.g., mobilization of the patient's own ability to produce IFN, new combination therapies and circumvention of side effects such as development of antibodies neutralizing IFNs and autoimmunity.

Humans↗