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H F Dullens

Publications and source records attributed to H F Dullens.

At least 37 records · Page 2Linked to original sources
PubMed1993

Transfer of tumor immunity by both CD4+ and CD8+ tumor infiltrating T lymphocytes activated in vivo by IL-2 therapy of tumor bearing mice.

DBA/2 mice were inoculated i.p. with syngeneic SL2 lymphoma or P815 mastocytoma on day 0 and treated i.p. with 20,000 units IL-2/day on day 10-14. This treatment is curative for 70-90% of the tumor bearing mice. Peritoneal cells and/or spleen cells were isolated from responding mice at the last day of IL-2 therapy. The in vivo antitumor activity of these cells was tested in Winn Assays (i.p.) and by adoptive transfer (i.v.) into mice injected s.c. with tumor previously. Peritoneal exudate cells isolated on day 14 (PEC14) from mice cured of SL2 tumor were highly effective in Winn Assays. Up to 5 x 10(7) SL2 cells could be eliminated in naive mice when injected i.p. together with 2 x 10(7) PEC14. Adoptive transfer (i.v.) of PEC14, without the addition of IL-2, into mice s.c. injected with SL2 tumor cells 1 or 3 days earlier, also prevented outgrowth of the tumor cells. T cells isolated from the spleens were less effective. Only at E:T ratios of 100:1 and 10:1 was tumor outgrowth inhibited. The adoptive transfer of PEC14 resulted in a long lasting immunity of the recipient mice. Furthermore, it was shown that depletion of both the CD8+ and CD4+ T cells from the suspensions used in the transfer studies, resulted in a significant decrease of tumor inhibition. However, the effect was not abrogated completely. PEC14 isolated from IL-2-treated DBA/2 mice cured of P815 tumor, protected naive mice against P815 tumor at E:T ratio 20:1. Adoptive transfer (i.v.) of these PEC14 into mice bearing s.c. P815 did not have an antitumor effect. In conclusion, low dose i.p. IL-2 therapy predominantly induces locally both CD4+ and CD8+ T cells with a strong antitumor activity in vivo. The potency of the IL-2-induced immunity seems related to the type of tumor used.

Animals↗
PubMed1993

Interleukin-2-containing liposomes: interaction of interleukin-2 with liposomal bilayers and preliminary studies on application in cancer vaccines.

Interleukin-2 (IL-2) incorporation in liposomes was studied under different conditions. Information was obtained on the mechanism of interaction of glycosylated recombinant IL-2 with liposomal bilayers. This information was utilized to formulate liposomes with high levels of incorporated IL-2. Multilamellar vesicles were prepared by hydration of a lipid film with an IL-2 solution. The incorporation efficiency, measured with a bioassay after forced release of IL-2 from the vesicles, was strongly dependent on the charge of the liposomes and the pH and ionic strength of the hydration medium. Negatively charged liposomes composed of phosphatidylcholine/phosphatidylglycerol (9:1) and prepared with IL-2 dissolved in 10 mM NaAc/270 mM glycerol, 0.1% BSA, pH 5, showed the highest incorporation efficiency (81%) among the investigated preparations. This type of liposome was selected for further study. Electrostatics play a crucial role in the process of IL-2 association with this type of liposome. Initial studies concerning induction of protective tumor immunity by immunization with reconstituted membranes with muramyl tripeptide phosphatidylethanolamine indicate that coinjection of IL-2-containing liposomes provided a significant enhancement of the immune response.

Animals↗
PubMed1993

Cell surface glycoconjugates as possible target structures for human natural killer cells: evidence against the involvement of glycolipids and N-linked carbohydrate chains.

Natural killer (NK) cells can spontaneously kill various malignant cells, but the susceptibility towards NK cells differs greatly among different types of tumour cells. The molecules, which are recognized by NK cells, have not yet been identified, but there is ample evidence that target cell surface glycoconjugates are involved in the interaction with NK cells. In this report, we show that the recognition of K562 target cells by human NK cells depends on the presence of protein-bound determinants, implying that glycolipids are not the primary target structures on K562 cells. The NK susceptibility of K562 cells was not altered by enzymic removal of various cell surface carbohydrates or oligosaccharides, mostly related to N-linked carbohydrate chains. Treatment of K562 cells with 1-deoxynojirimycin and 1-deoxymannojirimycin, inhibitors of N-glycan processing, resulted in drastic alterations in the carbohydrate phenotype of the cell surface, as could be shown by flow cytometric analysis of the lectin-binding properties of the cells. Despite these clear changes in N-glycosylation, the NK susceptibility of K562 cells remained unaffected. Summarizing, the results described in this report show that potential target structures for NK cells are protein bound, but the involvement of a specific (N-linked) carbohydrate determinant in the interaction between NK cells and target cells could not be established.

1-Deoxynojirimycin↗
PubMed1993

Involvement of the omental lymphoid organ in the induction of peritoneal immunity against tumor cells.

The omental lymphoid organ (OLO) is a part of the greater omentum composed of vascularized spots containing lymphocytes, plasma cells and macrophages located in a regular pattern between fat cells. To gain insight in the involvement of the OLO in the induction of immunity against tumor cells in the peritoneal cavity, we studied the penetration of tumor cells into the OLO after intraperitoneal, subcutaneous and intravenous injection. Furthermore we analyzed the tumoricidal activity of macrophages isolated from the OLO. Our results indicate that the OLO is only infiltrated by tumor cells directly from the peritoneal cavity, but not by subcutaneously or intravenously injected tumor cells unless they have reached the peritoneal cavity. Bromodeoxy-uridine labelled tumor cells can be detected in the OLO within 10 min after i.p. injection. The penetration is facilitated by the induction of fenestrations between the mesothelial cells (lining the OLO) after intraperitoneal injection of the tumor cells. These fenestrations can also be seen after nonspecific stimulation of the peritoneal cavity. Macrophages isolated from the OLO of mice immunized against syngeneic as well as allogeneic tumor cells express a significant cytotoxicity, which (at least in the syngeneic situation) precedes the cytotoxicity of the macrophages isolated from the peritoneal cavity. In conclusion, our data support the hypothesis that immune reactions against intraperitoneally injected tumor cells are initiated in the OLO leading to 'peritoneal immunity' against these tumor cells.

Animals↗
PubMed1992

Reconstituted membranes of tumour cells (proteoliposomes) induce specific protection to murine lymphoma cells.

Antigens presented on cell membranes or on liposomes are usually more immunogenic than antigens in soluble form, this being one of the reasons for the weak immunogenicity of extracted tumour-associated transplantation antigens (TATA). The main objective of this study is to solubilize TATA from tumour cells and to present them on a membrane-like structure to the immune system. Crude tumour cell membranes of SL2 lymphosarcoma cells (a spontaneously arising, weakly immunogenic tumour) were solubilized with octylglucoside or sodium deoxycholate, and reconstituted membranes (proteoliposomes) were prepared by detergent removal. Mice immunized s.c. with reconstituted membranes were protected against an i.p. challenge with tumour cells. Although octylglucoside solubilized only 41% of the membrane proteins, the reconstituted membranes were as immunoprotective as crude membranes. (Glyco)proteins were probably the major membrane components in the reconstituted membranes that induce immunoprotection, as mice immunized with preparations constituted of (glyco)lipids from SL2 cells could not reject SL2 cells. If Freund's complete adjuvant was used with the first immunization injection, no potentiation of the elicited immune responses was observed. Besides the membrane TATA, SL2 cells contained an apparently non-membrane-bound TATA, which was found in the cytoplasm. It is concluded that detergent solubilization of membranes and subsequent preparation of reconstituted membranes can be used to obtain membrane tumour-associated antigens that retain activity for induction of protective tumour immunity. The major advantage of this method is that membrane proteins are solubilized and are subsequently presented on a membrane-like structure that resembles the tumour cell membrane. On theoretical and practical grounds it provides a promising alternative for whole-cell vaccines.

Animals↗
PubMed1992

Effector cells of low-dose IL-2 immunotherapy in tumor bearing mice: tumor cell killing by CD8+ cytotoxic T lymphocytes and macrophages.

The presence and cytotoxicity of tumor infiltrating cells is described in mice during effective immunotherapy with interleukin 2 (IL-2). DBA/2 mice were inoculated i.p. with 2 x 10(4) tumor cells on day 0 and treated with daily i.p. injections with 20,000 units IL-2 on days 10-14. Mice bearing a large syngeneic i.p. tumor burden (SL2 lymphoma, P815 mastocytoma, L5178Y lymphoma, or L1210 lymphoma) could be cured by i.p. immunotherapy with these low doses of IL-2. In the peritoneal cavity of these mice an infiltrate of mononuclear cells was present. Similar numbers of lymphocytes (10(6)-10(7)) and macrophages (+/- 10(7)) were present in control tumor bearing mice and IL-2 treated tumor bearing mice. The ratio of CD4+/CD8+ T lymphocytes in the peritoneal cavity of mice rejecting the SL2 tumor was smaller than 0.5, whereas this ratio is about 2 in naive mice. In the spleens of IL-2 treated tumor bearing mice only a minor decrease of CD4+/CD8+ ratio was observed from 2.1-2.4 to 0.9-1.9. T cells isolated from the peritoneal cavity of mice inoculated with SL2 tumor cells and treated with IL-2, were highly cytotoxic to SL2 cells: at E:T ratio 2:1 the cytotoxicity index was 37 +/- 3. This cytotoxicity was specific and mediated by CD8+ T lymphocytes. Macrophages that were present in the peritoneal cavity of mice treated with IL-2 were also highly cytotoxic. The C.I. of these cells was 63-76% at E:T ratio 1:1. Cytotoxic macrophages were also present in untreated tumor bearing mice. The i.p. injections of IL-2 (20,000 units/day) caused a four-fold increase in the local NK-activity in the peritoneal cavity in naive mice. These IL-2 injections did not generate LAK-activity in vivo. Specificity of the in vivo tumor rejection was tested by injection SL 2 i.p. on day 0 and P815 i.p. on day 10, or vice versa, followed by IL-2 treatment. Only the tumor cells that were injected on day 0 were rejected. These in vivo experiments point to specific tumor rejection. In conclusion, both cytotoxic macrophages and CTL's are present in a sufficient number and with sufficient cytotoxicity to explain the killing of tumor cells in the peritoneal cavity. The CTL-activity seems of decisive importance for tumor rejection as this is induced by IL-2.

Animals↗
PubMed1991

Intratumoral low-dose interleukin-2 induces rejection of distant solid tumour.

This study shows that local tumor treatment with low-dose recombinant interleukin-2 (IL-2) can mediate rejection of a large distant solid tumour. When SL2 lymphoma cells were injected intraperitoneally (i.p.) in syngeneic DBA/2 mice on day 0.70% of these mice were cured by daily i.p. injections with 20,000 units IL-2 on days 10-14. After injecting mice with SL2 both i.p. and subcutaneously (s.c.) on the flank. 50% of the mice treated i.p. with low-dose IL-2 rejected both the i.p. tumour and the large distant s.c. tumour. In contrast, i.p. IL-2 treatment on days 10-14 cured fewer than 10% of the mice bearing only a s.c. SL2 tumour. The described IL-2 immunotherapy also caused systemic tumour rejection in mice bearing both ascitic and solid P815 mastocytoma. Thus it was shown that low-dose IL-2 can induce systemic tumour rejection, when injected at a site of tumour growth. Interleukin-2-induced rejection of s.c. SL2 tumour was highly specific, as mice that were rejecting i.p. and solid s.c. SL2 lymphoma did not reject solid P815 mastocytoma, which was injected s.c. simultaneously on the other flank. Furthermore, solid s.c. tumours consisting of mixtures of SL2 and P815 were not rejected in mice that rejected i.p. SL2 or P815. We conclude that intratumoral injections of low-dose IL-2 can enhance an ongoing weak immune reaction against the tumour resulting in systemic tumour rejection.

Animals↗
PubMed1991

Lymphokine-activated killer-cell-mediated killing of WiDr colon carcinoma cells is inhibited by K562 erythroleukemia cells.

Lymphokine activated killer (LAK) activity was induced in human peripheral mononuclear blood cells by human recombinant interleukin-2. Monocytes were required for optimal rapid proliferation of cells with LAK activity. They had no influence on the expression of tumoricidal activity by the LAK cells. The effector cells killed K562 erythroleukemia cells and WiDr colon cells differently, i.e. contact areas with WiDr cells were limited, whereas the contact areas between effector cells and K562 cells were much longer. Using mixtures of hot and cold target cells it was shown that effector cells preferably bind with K562 cells, impeding the binding of WiDr cells. Differences in expression of cytotoxicity of LAK cells against WiDr and K562 cells respectively was also observed after culturing the LAK cells for a relatively longer period. Cytotoxicity against WiDr was maximal at 3-16 days after starting LAK cell generation, whereas cytotoxicity against K562 was kept constantly high for at least 21 days. The addition of biological response modifiers [PHA and anti-CD3 antibody (OKT3)] during the LAK cell induction also had different effects on the expression of LAK activity against WiDr and K562 cells. Whereas PHA, in combination with rIL-2 had no significant influence on the cytotoxicity against WiDr cells, the cytotoxicity against K562 was significantly inhibited. Addition of anti-CD3 antibody diminished the cytotoxicity against WiDr target cells and had no influence on the cytotoxicity against K562 cells.

Carcinoma↗
PubMed1991

Acid ethanol fixation and polyester wax embedding combines preservation of antigenic determinants with good morphology and enables simultaneous bromodeoxyuridine (BrdU) labeling.

Lymphoid tissue, and/or isolated peripheral mononuclear blood cells were fixed in acid ethanol and embedded in polyester wax (melting point 37 C). The excellent cytomorphology obtained allowed distinguishing different types of individual lymphoid and nonlymphoid cells. Furthermore, this procedure was satisfactory in the immunophenotyping of histiocytes, endothelial, mesenchymal, epithelial cells, different (sub-) types of lymphocytes and also of lymphokine activated killer (LAK) cells. The staining patterns obtained with the different poly- and monoclonal antibodies on polyester wax sections were not only analogous to those obtained on frozen sections, but cells which had incorporated bromodeoxyuridine could be double labeled with specific antiserum.

Bromodeoxyuridine↗
PubMed1991

Parathymic lymph nodes during growth and rejection of intraperitoneally inoculated tumor cells.

The omental lymphoid organ (OLO) is a part of the greater omentum composed of vascularized milky spots situated between fat cells and containing lymphocytes, plasma cells and macrophages. We analysed the disappearance of intraperitoneally injected tumor cells from the peritoneal cavity and their infiltration into and disappearance from the OLO and the parathymic lymph nodes (PTLN) that drain the peritoneal cavity. After intraperitoneal inoculation of irradiated syngeneic tumor cells, they were visible in the OLO within 24 h. After 3 days, no tumor cells were seen anymore, but there were many macrophages that had phagocytosed tumor cells. After intraperitoneal inoculation of nonirradiated syngeneic tumor cells, a solid growing tumor mass developed in the OLO. The PTLN were also invaded by these nonirradiated tumor cells within 24 h as transfer of cell suspensions of these lymph nodes into naive mice leads to the death of the recipient mice due to tumor growth. However, from day 6 onwards, after tumor inoculation, these lymph nodes contained no tumor cells, despite progressive tumor growth intraperitoneally and in the liver and lungs. In allogeneic mice, tumor cells were rejected in the OLO after 7-10 days. Simultaneously, the number of lymphocytes and macrophages increased and a plasma cell reaction developed. The PTLN did not have tumor-inducing potency at any stage after intraperitoneal tumor injection. This study suggests, that the PTLN are more effective in the (local) eradication of tumor cells than the OLO. Still, considering its immunological potential, the OLO might be important as 'inducer' of immunological reactions in the peritoneal cavity.

Animals↗
PubMed1990

Mechanisms of tumour rejection in the murine DBA/2-SL2 concomitant immunity system.

The mechanisms of rejection of secondary tumours were studied in concomitant immunity using transplants of standard-size solid SL2 tumours in syngeneic DBA/2 mice. In such a system primary tumor implants are not rejected, in contrast to secondary tumour implants. The second tumour was mainly rejected 2-4 days after implantation. Both primary tumour and secondary tumour implants (which are rejected) contained hardly any lymphocyte infiltrate, whereas, 2-4 days after implantation they contained 40%-50% macrophages, which were cytotoxic in vitro. Transfer (s.c.) of these tumours to naive mice showed that cellular infiltrates in the secondary implants did not always cause tumour rejection. Serum collected on day 4 after implantation of the secondary tumour was cytotoxic to SL2 tumour cells in vitro, whereas serum from mice with only primary implants was not cytotoxic on day 4 after implantation. Preliminary characterization of this cytotoxic factor showed that it was heat-labile, as cytotoxicity disappeared after 30 min at 56 degrees C, the molecular mass of the factor was higher than 100 kDa, and it was not IgG. We hypothesize that secondary tumours in the DBA/2-SL2 concomitant immunity system are rejected mainly between 2 and 4 days after implantation as a result of the combined action of cytotoxic serum and the presence of 40%-50% cytotoxic macrophages. The primary tumour is not rejected at 2-4 days after implantation as there is no cytotoxic factor.

Animals↗
PubMed1989

Immunotherapy of mice with a large burden of disseminated lymphoma with low-dose interleukin 2.

Successful immunotherapy with recombinant interleukin 2 (rIL-2) of mice bearing a large burden of lymphokine-activated killer-resistant disseminated SL2 lymphoma is described. When mice were challenged i.p. with 2 x 10(4) SL2 cells on day 0 and treated with daily i.p. injections of 5,000 units rIL-2 on days 3-7, no therapeutic effect was observed. However after treatment with daily IL-2 injections on day 10-14, 25% of the mice survived. Ten days after this tumor challenge more than 10(8) SL2 cells were present growing as ascitic tumor. On day 10, SL2 cells were also present as solid tumor in the greater omentum and as metastases in lungs and liver. Surviving mice were able to reject a second challenge with SL2 cells given on day 60. A second challenge with P815, another DBA/2 tumor, resulted in death of the mice due to tumor development. This finding is of particular importance as the SL2 cells are resistant to lymphokine-activated killer activity. Thus local (i.p.) injection of low dose rIL-2 can cause the systemic rejection of advanced and metastasized cancer. Our data indicate that IL-2 can strongly enhance a specific immune reaction against tumor cells.

Animals↗
PubMed1989

Initial immunochemical characterization of specific macrophage-arming factor.

This paper describes the initial immunochemical characterization of specific macrophage-arming factor (SMAF). SMAF is an antigen-specific factor that is released by (sensitized) T lymphocytes after contact with the specific antigen. It renders macrophages specifically cytotoxic. The specificity is dependent on the tumor-mouse combination. In allogeneic systems the specificity is H-2-directed, whereas in the syngeneic systems the specificity is tumor-specific. SMAF has a molecular mass of 65-85 kDa (established by gel filtration). By affinity chromatography SMAF could not be adsorbed with anti-(kappa + lambda light chain) immunoglobulins or anti-IgG from SMAF-containing supernatants. SMAF could be adsorbed with the monoclonal antibody 14-30 (directed against specific T-cell factors), and could be eluted from columns containing the latter. Furthermore, SMAF could also be adsorbed with and eluted from affinity chromatography columns to which specific tumor cell membranes or KCl extracts of these tumor cell membranes were coupled. Other tumor cell membranes could not adsorb SMAF. Together these data show that SMAF is not an antibody but a T-cell factor with an antigen-specific recognition site.

Animals↗
PubMed1989

Production of specific macrophage-arming factor precedes cytotoxic T lymphocyte activity in vivo during tumor rejection.

Recently we published a hypothesis on the immunological events occurring during tumor rejection. One of the implications of this hypothesis is that specific macrophage-arming factor (SMAF) is produced early during the initiation of the immune response, whereas the "classical" cell-mediated immune response components, such as cytotoxic T lymphocytes (CTL), are produced later, that is, during the amplifier-effector phase. In this paper we establish the kinetics of the induction of (a) lymphocytes producing SMAF and (b) CTL. Groups of DBA/2 mice were injected i.p. once, twice or three times with irradiated and/or non-irradiated syngeneic SL2 tumor cells, the injections being given at intervals of 10 days. After each of these injections the production of SMAF and the expression of CTL activity were established. The results showed that in the peritoneal cavity SMAF-producing lymphocytes appeared earlier than cytotoxic lymphocytes (CTL). In addition, it was shown (a) that SMAF does not interfere with the in vitro cytotoxicity expressed by CTL and (b) that in addition to CTL memory cells, SMAF-producing memory cells were also induced after injection of syngeneic tumor cells. These data support the hypothesis that SMAF is involved in the early phase of the cellular immune response against tumors, whereas CTL are induced later.

Animals↗
PubMed1988

Lack of correlation between in vivo peroxidatic activity patterns and tumoricidal activity in vitro of peritoneal macrophages after intraperitoneal immunization with tumor cells.

Immunization of C57BL mice with one inoculum of 10(7) DBA/2-derived SL2 lymphosarcoma cells resulted in a +/- 20-fold increase in the total number of peritoneal cells. The number of macrophages showed a 10-fold increase from 3 x 10(6) (control mice) to 3.4 x 10(7) cells at day 8 after immunization. Within this macrophage population, four different cell types, based on the ultrastructural peroxidatic activity patterns, could be distinguished: exudate macrophages, resident macrophages, resident-exudate macrophages and peroxidatic-activity-negative macrophages. The number of exudate macrophages significantly increased in the peritoneal cavity after immunization: at day 8 after immunization, a peak value of 10(7) cells was observed. At the same time, there were 2.2 x 10(7) peroxidase-activity-negative macrophages present (representing the control value x 50). Significant in vitro tumoricidal activity of the isolated macrophages could not be measured until 8 days after immunization. At that time, a cytotoxicity index of 68 was reached. After immunization of the C57BL mice with 3 injections with allogeneic SL2 cells, there were no dramatic changes in the number of peritoneal cells after the last immunization. Only immediately after the last immunization was a minor increase in peroxidatic-activity-negative macrophages seen. But already at 5 days after the last immunization, the composition of the peritoneal suspension was similar to that of non-immunized mice with predominantly resident macrophages. The cytotoxicity of the peritoneal macrophages from hyperimmunized mice was constantly high during 1-15 days after the last immunization (cytotoxicity index ranged from 66-72). In order to study which type(s) of macrophage(s) (resident, exudate, resident-exudate or peroxidatic-activity-negative) is/are responsible for the cytotoxicity measured in vitro, peritoneal cell suspensions (obtained after immunization) were fractionated according to their affinity to wheat germ agglutinin (WGA) coupled to Sepharose columns. Comparison of the values of cytotoxicity measured before and after separation into "subtypes" of the macrophages revealed that the expression of cytotoxicity is not correlated with any of the "sub-types", especially when the peroxidatic activity pattern is is taken as a criterion.

Animals↗
PubMed1987

Spontaneous tumor rejection is not always due to a complete cellular rejection.

In this paper we have studied whether there is a correlation between efficacy of hosts to reject tumor cells spontaneously and the induction of cytotoxic macrophages in these recipients due to injection (immunization) with tumor cells. Results show that there is no cause-effect relationship between macrophage cytotoxicity and tumor rejection in vivo. This lack of cause-effect relationship is greatly influenced by differences in immunogenicity between the various tumor cell lines as well as by the release of tumor-factors by some of these tumors.

Animals↗
PubMed1986

Tumor growth stimulatory macrophages induced by a subclinical bacterial infection in vivo.

Nonelicited peritoneal macrophages obtained from normal mice from our animal house unexpectedly expressed a strong tumor growth stimulatory effect in vitro. Macrophages expressing this stimulatory effect had an aberrant morphology compared to the morphology of normal macrophages as observed by electron microscopy. The results of immunization of these affected mice with tumour cells led to the usual lymphocyte sensitization. No external symptoms were observed, and the mice looked healthy. Treatment of the affected macrophage donors with antibiotics resulted in the abolishment of the tumor growth stimulatory effect by the macrophages. Thus, this tumor cell growth stimulation by macrophages was probably due to a subclinical infection of the mice.

Animals↗