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T Timonen

Publications and source records attributed to T Timonen.

At least 19 recordsLinked to original sources
PubMed1992

Migration of recombinant IL-2-activated T and natural killer cells in the intercellular space of human H-2 glioma spheroids in vitro. A study on adhesion molecules involved.

The migration of rIL-2-activated T and NK cells into the intercellular space of glioma tissue was studied using multicellular spheroids grown from the human H-2 glioblastoma cell line as targets. Lymphocytes of all analyzed subtypes migrated into the spheroids, but CD56+ cells were particularly migratory. Lymphocytes and the H-2 tissue expressed adhesion molecule subunits for the following potential cell-cell or cell-matrix interactions: alpha 3 beta 1 (VLA-3) to fibronectin, laminin, and collagen; alpha 4 beta 1 (VLA-4) and alpha 5 beta 1 (VLA-5) to fibronectin; alpha 6 beta 1 (VLA-6) to laminin; alpha 4 beta 1 to VCAM-1; alpha L beta 2 (Leu-CAMa/LFA-1) to CD54 (ICAM-1); CD44 to fibronectin, collagen, laminin, hyaluronate; CD2 to CD58 (LFA-3); and CD56 (N-CAM) to CD56. In the H-2 tissue, CD54 and VCAM-1 were expressed as a gradient. The expression of CD54 was weak in the peripheral zone and the expression was stronger in the quiescent deeper zone, whereas the distribution of VCAM-1 showed an inversed pattern. The low expression of CD54 was up-regulated along the frontier of migrating lymphocytes. The migration was almost totally prevented by the anti-CD18 (beta 2) mAb IB4 and TS1/18, and also strongly inhibited by the anti-CD54 mAb LB-2. Instead, mAb known to inhibit the binding of beta 1 integrins to fibronectin were not significantly inhibitory. However, a combination of the GPEILDVPST and GRGDS peptides, which compete for the binding of alpha 4 beta 1 and alpha 5 beta 1 to fibronectin and may also affect other adhesion systems, partially prevented migration.

Amino Acid Sequence↗
PubMed1991

Recognition of chromosome 6-associated target structures by human lymphokine-activated killer cells.

Different populations of unstimulated and IL-2-activated PBL were used in binding and killing assays against somatic mouse/human lymphocyte cell hybrids containing different human chromosomes. Unstimulated PBL effector cells showed low binding and killing activity to both cell hybrids and mouse parental cell lines. However, IL-2-activated killer (LAK) cells bound strongly to, and effectively killed, cell hybrids carrying human chromosome 6, but were inefficient in both assays to mouse parental cells and to cell hybrids not carrying human chromosome 6. These results show that human LAK cells but not endogenous NK cells bind and kill mouse/human lymphocyte hybrids containing human chromosome 6. We thus suggest that LAK cells recognize ligands encoded by genes on chromosome 6.

Chromosomes, Human, Pair 6↗
PubMed1991

A monoclonal antibody to the human leukocyte adhesion molecule intercellular adhesion molecule-2. Cellular distribution and molecular characterization of the antigen.

The DNA of the human leukocyte adhesion molecule intercellular adhesion molecule-2 (ICAM-2) was synthesized by the polymerase chain reaction, and a protein A-ICAM-2 fusion protein was expressed in Escherichia coli. The fusion protein was used as immunogen to obtain mAb to ICAM-2. The 6D5 antibody was selected by its reactivity in immunofluorescence with the endothelial cell line Eahy926. The antibody precipitated a 55,000 m.w. glycoprotein from radioactivity surface labeled cells and also reacted in Western blotting. As measured by immunofluorescence flow cytometry, the antibody reacted with lymphoblastoid B cells of normal origin, some Burkitt lymphoma cell lines, vascular endothelial cells, the endothelial cell line Eahy926, 4 and a subpopulation of Con A-stimulated blood mononuclear cells. The two Ig-like domains of ICAM-2 were separately expressed in E. coli, and the antibody was shown to react with the NH2-terminal domain. The antibody inhibited the CD11/CD18-dependent binding of HL-60 promyelocytic leukemia cells to transfected COS-1 cells.

Antibodies, Monoclonal↗
PubMed1991

The expression of human intercellular adhesion molecule-2 is refractory to inflammatory cytokines.

The beta 2-integrin CD11a/CD18 binds to the intercellular adhesion molecules (ICAM)-1 (CD54) and ICAM-2. ICAM-1 has a wide distribution, and its expression is up-regulated by various cytokines. In contrast, ICAM-2 has a more restricted distribution, and is mainly expressed on endothelial cells. In the present study we show that it is not induced by inflammatory cytokines or other treatments on any of several cells studied. Moreover, antibodies to the intercellular adhesion ligands were not able to block all CD11a/CD18-dependent adhesion, indicating the presence of additional CD11a/CD18 ligands.

Antigens, CD↗
PubMed1991

Role of CD16 (Fc receptor III) and interleukin-2 in the reactivation of natural killer cells after inhibitory target cell contact.

Contact with natural killer (NK)-resistant monolayer targets is an inhibitory signal to NK cells. In this study, we have analyzed the effect of such effector/target cell interactions on the CD16 (FcRIII) expression on lymphocytes and the role of CD16 and interleukin-2 (IL-2) in the reactivation of their cytolytic machinery. Coculturing peripheral blood mononuclear cells with NK-resistant monolayer cells did not change the percentage of CD 16-positive effector cells, although this treatment effectively inhibited their cytotoxicity against NK-sensitive targets. The inhibited effector cells partially regained their activity by incubating for 24 h in medium supplemented with 10% fetal calf serum (FCS), whereas human albumin-, newborn calf serum- or human AB serum-supplemented media had no reactivating effect. Monoclonal class IgG1, IgG2a and IgM anti-CD16 antibodies [Abs; 3G8, CLB-CD16 (CLB-FcR gr1) and Leu 11b], and normal rabbit IgG (NR-IgG) prevented the FCS-mediated reactivation of cytotoxicity, whereas nonreactive control Abs significantly enhanced it. The detection of the CD16 antigen by the monoclonal anti-CD16 Abs Leu 11a and Leu 11c was blocked by the above anti-CD16 Abs and NR-IgG, while the expression of other NK cell-associated surface molecules (CD2, CD56) remained unchanged. Mere blocking of CD16, using a short-term incubation with anti-CD16 Abs, had an insignificant effect on endogenous NK activity, suggesting that CD16 is involved in NK cell (re)activation rather than in the killing process itself. In the presence of IL-2, inactivated effector cells also regained their killing activity. The IL-2-induced reactivation was not inhibited by anti-CD16 Abs. The results suggest that FCS-derived factors and soluble nonreactive immunoglobulins enhance the NK activity of down-regulated effector cells via CD16, and that CD16 and IL-2 receptors represent alternative independent pathways of NK cell reactivation.

Antibodies, Monoclonal↗
PubMed1990

Participation of CD11a-c/CD18, CD2 and RGD-binding receptors in endogenous and interleukin-2-stimulated NK activity of CD3-negative large granular lymphocytes.

The effect of RGD-sequence-containing pentapeptides and monoclonal antibodies (MAbs) against the adhesion molecules CD11a-c/CD18, ICAM-1 (CD54) and CD2 on the binding and cytotoxicity of endogenous (freshly purified) and IL-2-stimulated CD3-negative NK cells has been studied. Antibody to CD18 exerted the most significant inhibition of adhesion and cytotoxicity of endogenous NK cells to MOLT-4 lymphoma cells, followed by antibodies to CD2 and CD54. Antibodies to either CD11a, CD11b or CD11c did not inhibit adhesion when used separately, whereas as a mixture their inhibitory capacity was as strong as that of anti-CD18. Antibodies against CD18, CD54 and CD2 exerted an additive effect on the inhibition of adhesion. Their combination eradicated the binding of endogenous NK cells. The RGD-containing peptide did not inhibit the binding or cytotoxicity of freshly purified NK cells to MOLT-4, whereas some inhibition was detected against the adenocarcinoma cell line COLO-205. According to FACS analysis, IL-2 increases the expression of CD2 and CD54 on NK cells. However, the relative contribution of the adhesion molecules to the NK cell binding did not change as a result of the stimulation with IL-2. The RGD-peptide substantially inhibited the binding of IL-2-stimulated killer cells to COLO, and the combination of this peptide with MAbs to CD18, CD54 and CD2 practically blocked the adhesion. Our results indicate that both CD11a-c/CD18- (involving the 3 heterodimers) and CD2-dependent adhesion pathways are used by LGL in endogenous and IL-2 stimulated natural killing. In addition, RGD-binding receptors are involved in adhesion to some target cells.

Antigen-Antibody Reactions↗
PubMed1990

Damage to multicellular human H-2 glioma spheroids incubated with LAK cells: an ultrastructural study.

Local brain tumor therapy using lymphokine-activated killer (LAK) cells and recombinant interleukin-2 (rIL-2) has not proved to be effective in preliminary clinical trials. One obstacle to effective use of this therapy is ignorance about the events that follow contact of the LAK cells with glioma tissue. We used multicellular spheroids grown from human glioma cell lines as targets to study, in vitro, the effect of LAK cells against three-dimensional glioma tissue. Here we describe the ultrastructural changes in spheroids of H-2 glioma cells incubated in pellets of LAK cells for up to 24 hours. In H-2 spheroids, cellular damage was not restricted to the effector cell-target cell (effector-target) contact; it extended farther, at least partly because of nonspecific changes in the spheroid micromilieu. Formation of cytoplasmic blebs, a characteristic effect of T cells, natural killer cells, and LAK cells on single target cells, also occurs in H-2 spheroids, and it is not limited to the effector-target contact area either. These findings suggest that LAK cells release membrane-damaging agents that remain active outside the effector-target area, in the micromilieu of H-2 spheroid tissue.

Cytotoxicity, Immunologic↗
PubMed1990

Leukocyte-cell adhesion: a molecular process fundamental in leukocyte physiology.

Leukocyte-cell adhesion is a form of physical contact characterized by fast (firm) stickiness between the cells. To analyze the biology and molecular basis of this process, an adhesion-specific assay was developed: the phorbol ester-induced aggregation of human lymphocytes. This rapid and antigen-independent intercellular adhesion requires cellular metabolism, an intact cytoskeleton and extracellular divalent cations, and is mediated by preformed cell-surface proteins referred to as CAMs. Phorbol ester also induces aggregation of monocytes and granulocytes, as well as adhesion of T lymphocytes to either B cells or monocytes and of the leukocytes to vascular endothelial cells. By using the adhesion-specific assay and blocking monoclonal antibodies, several CAMs have been identified, namely the Leu-CAM family (CD11a-c/CD18) and ICAM-1 (CD54). The Leu-CAM family is composed of Leu-CAMa (CD11a/CD18), Leu-CAMb (CD11b/CD18) and Leu-CAMc (CD11c/CD18), three glycoprotein heterodimers made of a common beta-chain and distinct alpha-chains. ICAM-1 is an adhesive ligand for Leu-CAMa. Expression and use of the various CAMs is selective in different types of leukocytes. The Leu-CAMs have been purified and partially characterized. CD18, whose gene is on human chromosome 21, contains 5-6 N-linked complex-type oligosaccharides, and CD11 binds Ca++. Another adhesion pathway is mediated by CD2 and CD58. CD2, a glycoprotein selectively expressed by T cells, is a receptor for CD58, a cell-surface adhesive ligand with broad tissue distribution. Antibodies to the latter CAMs do not block the phorbol ester-induced lymphocyte aggregation. Adhesion is involved in a large variety of leukocyte functions. Anti-Leu-CAM antibodies block induction of IL-2 production and lymphocyte proliferation. Lymphocyte-mediated cytotoxicity is also inhibited. Endogenous NK and LAK cells use Leu-CAMs, ICAM-1 and CD2, and sometimes RGD receptors, to bind and kill tumor cells. Endogenous compounds such as H2O2 and LTB4 also induce Leu-CAM-dependent adhesion in monocytoid cells and granulocytes, respectively, and degranulation of the latter cells is enhanced by the adhesion process. Homologous CAMs have been identified in rabbit and mouse. In in vivo studies in the former species, anti-Leu-CAM antibodies block adhesion of leukocytes to vascular endothelium and thereby their migration into extravascular tissues. The antibodies thus inhibit granulocyte accumulation and plasma leakage in inflammatory lesions, and induce lympho- and granulocytosis, indicating that cell-adhesion contributes to the distribution of leukocytes in the body.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1990

Interleukin-2 activated lymphocytes (LAK cells) as potential tumor tracers.

The homing characteristics and infiltrative capacity of interleukin-2 activated human peripheral blood lymphocytes, the lymphokine activated killer (LAK) cells, were studied. In vitro stimulated 111In-oxine labeled lymphocytes were injected into the hypogastric artery during hysterectomy, performed because of endometrial carcinoma. Scintigrams demonstrated clear homing of the lymphocytes into the area of the malignant tumor. No selective homing was detectable when labeled red blood cells were injected in a similar fashion. To analyze the infiltrative capacity of the activated lymphocytes, they were incubated in vitro with tumor spheroids grown from cultured glioma cell lines. As revealed by antibodies against the leukocyte common antigen and immunoperoxidase techniques, the activated lymphocytes infiltrated the three-dimensional tumor tissue slowly as a frontier. These results show that, in addition to their previously suggested potential role in cancer therapy, interleukin-2 activated lymphocytes may possibly also be useful as tumor tracers.

Female↗
PubMed1990

Cardiac involvement in bone marrow transplantation: electrocardiographic changes, arrhythmias, heart failure and autopsy findings.

In order to evaluate cardiac involvement in bone marrow transplantation (BMT) we reviewed serial electrocardiograms, chest X-rays and cardiac pathology at autopsy in 45 consecutive adult patients undergoing BMT for hematologic malignancies at our institution. All patients were pretreated with cyclophosphamide (CY, 120 mg/kg) and total body irradiation (TBI, 10-12 Gy). A total of 15 patients developed electrocardiographic ST-segment or T-wave changes and/or arrhythmias post-BMT; four of them also suffered from congestive heart failure. The arrhythmias included frequent ventricular extrasystoles (two patients), paroxysmal atrial fibrillation (one patient), repeated supraventricular tachycardia (one patient) and QT-prolongation with ventricular tachyarrhythmias (one patient). Twelve patients showed an early drop of the total QRS voltage sum exceeding 15% of the pretransplant reading. In a subgroup of five patients the voltage drop was associated with ST-segment or T-wave changes; three of them developed congestive heart failure. At autopsy in 15 patients, the heart weight was on the average 113% of predicted. Myocardial edema, fibrosis and cellular hypertrophy were the most common microscopic findings. Two patients had marantic endocarditis of the aortic valve. Thus, in our experience clinically significant heart involvement affects 5-10% of patients undergoing BMT after pretreatment with CY and TBI. While cardiac complications are generally not a major problem of BMT, they may have serious consequences for individual patients and should therefore be carefully watched for in the treatment of BMT recipients.

Adolescent↗
PubMed1989

Effect of LAK cells against three-dimensional tumor tissue. In vitro study using multi-cellular human glioma spheroids as targets.

The anti-tumor mechanisms of local LAK cell therapy are difficult to study in vivo. We describe a method to study in vitro the action of LAK cells against three-dimensional tumor tissue. Spherical cell aggregates (spheroids) grown from human glioma cell lines H-2 and U-251 were labeled with 51Cr and then incubated for up to 24 h with LAK cells. After the incubation, most spheroids were still macroscopically identifiable, and the measured reduction of volume did not correlate to the extent of damage. LAK cells infiltrated into spheroid tissue slowly as a frontier which explains why the specific 51Cr release was clearly slower from spheroids than corresponding single cell suspensions. The infiltrated area was at 1 to 2 h very thin but by 8 to 12 h consisted already of several cell layers. Most H-2 spheroids became totally infiltrated by 16 to 24 h whereas in U-251 spheroids the infiltration usually remained peripheral. In accordance with the different extent of infiltration, H-2 spheroids were clearly more sensitive to LAK cells than U-251 spheroids: at E/T ratio 10:1 the mean specific 51Cr release by 24 h was 63 and 36%, respectively. A single exposure to LAK cells released 51Cr from H-2 spheroids approximately 12 h but over 24 h from U-251 spheroids. The spheroid model can be used to study the infiltrative capacity and cytotoxicity of LAK cells against three-dimensional tumor tissue, and the method may help to find an optimal mode of local LAK cell therapy, i.e., proper combination of lymphokines and LAK cells, and proper timing of their administration.

Cell Line↗
PubMed1989

MAC technique (morphology antibody chromosomes) in phenotypic identification of proliferating NK and T cells in interleukin-2-stimulated lymphocyte cultures.

Human peripheral blood lymphocytes were activated with recombinant interleukin 2 (rIL-2) and cultured in fetal calf serum (FCS)- or human-AB-serum-supplemented media. Proliferative cells were identified by the MAC technique (morphology antibody chromosomes) which enables the immunoenzymatic identification of both mitotic and non-proliferating cells in unfractionated lymphocyte populations. The results indicate that the phenotype of more than 90% of proliferative lymphocytes can be characterized by using antibodies against T cells and NK cells. Substantial mitotic activity of CD4-positive (CD4+) T cells was observed only in FCS-supplemented cultures, whereas in serum-free or human-AB-serum-supplemented cultures mostly CD8+ T cells and NK cells proliferated. The proportion of NK cells among all mitotic cells varied between 14 and 32%. Interestingly, in unfractionated cultures approximately 13% NK cells entered mitoses in the presence of rIL-2, suggesting that the poor proliferative capacity of purified NK cells demonstrated previously may be due to the lack of accessory stimulatory signals. The proliferation of B cells was minimal in all experiments. The MAC technique is a useful addition to the techniques by which lymphocyte growth regulation is monitored.

Cells, Cultured↗
PubMed1988

CD11a-c/CD18 and GP84 (LB-2) adhesion molecules on human large granular lymphocytes and their participation in natural killing.

Effect of mAb against the CD11a-c, CD18, and GP84 adhesion molecules on the binding and cytotoxicity of human NK cells was studied. The target cells were K562, MOLT-4, Raji, and fresh uncultured autologous endometrial carcinoma cells. Antibodies against adhesion relevant epitopes of CD11a(TA-1/LFA-1), CD11b(Mol/OKM1/Mac1), or CD11c (Leu-M5) did not inhibit NK function. The mAb 60.3 against CD18, the common beta-chain associated to CD11a-c, strongly inhibited both the binding and cytotoxicity of large granular lymphocytes (LGL) against all the target cells tested. Also the antibody LB-2 against the GP84 adhesion molecule inhibited NK function to some degree. 60.3 and LB-2 antibodies exerted an additive effect in the inhibition of both binding and cytotoxicity. However, even this antibody combination did not completely block NK activity, suggesting a heterogeneity of adhesion structures in the NK system. According to both FACS analyses and immunoprecipitation studies, all the tested antibodies recognized either a subpopulation or all of LGL. On the other hand, antibodies against CD11b, CD11c, and LB-2 showed only marginal reactivity with highly purified LGL-free T cells.

Antibodies, Monoclonal↗
PubMed1988

Cyst fluid of glioma does not inhibit the killing action of lymphokine-activated killer (LAK) cells in vitro.

In theory, lymphokine-activated killer (LAK) cells offer a potential method to treat cerebral gliomas, especially low-grade gliomas. LAK cells would be administered by repeated injections straight into the cavity of a subtotally removed tumour. However, brain-tumour cyst fluid has been shown to be immunosuppressive in lymphocyte stimulation tests. Therefore we wanted to know whether the fluid would reduce the killing efficacy of LAK cells. Using a standard cytotoxity test based on 51Cr release, we compared in vitro the cytotoxity of LAK cells against K-562 tumour cells in brain-tumour cyst fluid, autologous serum and allogeneic serum. Five patients with cystic glioma and one with cystic meningioma were studied and no inhibition of cytotoxity of LAK cells was observed.

Adolescent↗
PubMed1988

Characteristics of soluble tumour-derived proteins that inhibit natural killer activity.

We have previously shown that various benign and malignant natural killer (NK)-resistant monolayer cells inhibit endogenous human NK activity, probably by reducing the secretion of cytotoxic factors from the effector cells. The nature of the molecules responsible for the inhibition has been unclear. In this study we show that phosphate-buffered saline (PBS) extracts of ovarian cystadenocarcinoma tissue and normal uterine smooth muscle strongly inhibit NK activity. Fractionation of tumour extracts by gel chromatography revealed major inhibitory activity in the Mr range 160,000-180,000, and other weaker inhibiting activities in the Mr ranges 50,000-70,000 and 20,000. The active material of Mr range 160,000-180,000 was adsorbed on anion exchange chromatography column at neutral pH and physiologic NaCl concentration, and it was eluted by 0.31-0.34 M NaCl. The inhibitory molecule was sensitive to proteolysis. No relation of this compound to immunoglobulins or trypsin and urokinase inhibitors was detected. The unfractionated extract inhibited NK activity apparently by the same mechanism as the monolayer target cells, i.e. by reducing the secretory capacity of effector cells. The data strongly suggest that the NK-inhibiting compounds described in this work are involved in the inactivation of NK cells by intact monolayer cells.

Cytotoxicity, Immunologic↗
PubMed1988

Influence of terbutaline on natural killer cell activity.

Studies on the effect of cAMP-inducing agents on NK activity have been contradictory. The aim of this study was to investigate the acute effects of beta-agonists on NK activity in vivo in 15 asthmatics and 3 healthy volunteers. Blood samples of NK activity were taken at regular intervals after placebo and after subcutaneous injection of 7 micrograms/kg of terbutaline. NK activity was measured by the standard 4-h Chromium51 release assay against the leukemic line K 562 at a 50:1 effector/target cell ratio. Compared with placebo, terbutaline induced within 30-60 min a significant increase in NK activity which lasted less than 2 h. Further studies are necessary to investigate the effect of long-term beta-agonist treatment on NK activity.

Adolescent↗