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D Boraschi

Publications and source records attributed to D Boraschi.

At least 37 records · Page 2Linked to original sources
PubMed1995

Single-cell analysis of macrophage chemotactic protein-1-regulated cytosolic Ca2+ increase in human adherent monocytes.

The increase in intracellular free Ca2+ ([Ca2+]i) associated with interaction of monocyte chemotactic protein-1 (MCP-1) and related chemokines beta with adherent human blood monocytes was investigated at the single-cell level. We used f-MLP as reference chemotactic agent. MCP-1 caused an increase in [Ca2+]i in individual adherent monocytes, with 95% of cells responding to the chemokine at 20 ng/mL. Response to MCP-1 was already detectable at 1 pg/mL, whereas at least 5 ng/mL were required for significant chemotactic response. The kinetics of the increase in [Ca2+]i were considerably different for MCP-1 compared with f-MLP. MCP-1 produced a slow increase of [Ca2+]i that reached a plateau in 5 to 7 minutes. On the other hand, the increase of [Ca2+]i induced by f-MLP appeared to be biphasic, with a fast phase peaking after 5 to 40 seconds followed by a slower wave. Blocking of Ca2+ channels by Ni2+ or Cd2+ and/or chelation of extracellular free Ca2+ considerably reduced but did not abolish response to MCP-1, had no effect on the first wave of [Ca2+]i induced by f-MLP, and completely abrogated the second, slower wave. Thapsigargin, which empties intracellular Ca2+ stores, inhibited f-MLP-induced [Ca2+]i increase but fully blocked the action of MCP-1 only when combined with Ni2+. Thus, increase of [Ca2+]i induced by MCP-1 is apparently due to independent opening of a channel and mobilization from intracellular stores, whereas f-MLP-induced mobilization of Ca2+ from stores causes subsequent opening of a channel. At variance with MCP-1, the related chemokine MCP-2 induced only a low increase of [Ca2+]i in about 40% of adherent monocytes. Inhibition of chemokine-induced increase of [Ca2+]i by cholera or pertussis toxin indicated that MCP-1 and MCP-2 activate monocytes through different intracellular pathways. These results demonstrate at the single-cell level that the mechanisms and dynamics of increased [Ca2+]i are considerably different for f-MLP and chemokines beta. In addition, the [Ca2+]i increase induced by the two related chemokines beta MCP-1 and MCP-2 appears to be differently regulated, suggesting interaction with distinct receptors.

Cadmium↗
PubMed1995

Identification of MIP-1 alpha/LD78 as a monocyte chemoattractant released by the HTLV-I-transformed cell line MT4.

It is known that the HTLV-I-transformed cell line MT4 releases chemotactic activity for monocytes spontaneously. The MT4 monocyte chemoattractant was purified to homogeneity and sequencing of 25 amino acids revealed identity with the C-C chemokine macrophage inflammatory protein-1 alpha (MIP-1 alpha/LD78). An anti-MIP-1 alpha/LD78 rabbit antiserum substantially inhibited chemotaxis of the MT4 chemoattractant. MT4 cells constitutively expressed MIP-1 alpha/LD78 but not the C-C chemokines MCP-1, RANTES, and MIP-1 beta/Act2 and the C-X-C chemokines IL-8, gro alpha, and gro beta. MT4-derived MIP-1 alpha/LD78 was active on monocytes but was a weak chemoattractant for polymorphonuclear leukocytes. Thus, MIP-1 alpha/LD78 is a major monocyte chemoattractant released by HTLV-I-transformed T cells. Expression of MIP-1 alpha/LD78, a leukocyte chemotactic and myelosuppressive molecule, may play an important role in the manifestations of HTLV-I-related diseases.

Amino Acid Sequence↗
PubMed1993

Type II interleukin-1 receptor is not expressed in cultured endothelial cells and is not involved in endothelial cell activation.

Interleukin-1 (IL-1) profoundly affects a number of functions of endothelial cells (EC). It was previously shown that EC express the type I 80-Kd IL-1 receptor (IL-1 RI). In this study we define the expression and functional significance of the type II IL-1R (IL-1 RII) in EC. Human umbilical vein EC did not express appreciable levels of IL-1 RII mRNA as assessed by Northern analysis or reverse transcription and polymerase chain reaction. Exposure to various cytokines (including IL-4, which augments IL-1 RII in neutrophils) failed to induce IL-1 RII mRNA. The binding of radiolabeled IL-1 beta to EC was blocked by antitype I (M4) but not by antitype II (M22) monoclonal antibodies (MoAbs). MoAbs directed against the IL-1 RI (M1 and M4) inhibited the induction of IL-6 and adhesion molecules in EC by IL-1, whereas an anti-IL-1 RII (M22) was inactive. The human IL-1 receptor antagonist (IL-1ra) preferentially interacts with IL-1 RI versus IL-1 RII in the mouse. IL-1 ra inhibited the response of mouse endothelial cells to IL-1. We conclude that EC selectively express the IL-1 RI and that this is involved in the response of this cell type to IL-1.

Animals↗
PubMed1993

Cytokines as vaccine adjuvants: interleukin 1 and its synthetic peptide 163-171.

The possibility of preventing infectious diseases by employing efficacious vaccine is rapidly growing as a consequence of the new technologies in recombinant DNA and protein chemistry. However, the increasing number of synthetic and recombinant antigens further stresses the role of appropriate adjuvants to ensure maximal vaccine activity and the protection of all vaccinees. Several approaches can be applied to develop safe and effective agents capable of enhancing specific immune responses which can then protect the host from the pathogen. Among others, the direct use as adjuvant of those cytokines which are induced in animals by the classical Freund's adjuvants has recently become a matter of investigation. In particular, interleukin 1 (IL-1) has been shown to possess adjuvant activity for a variety of infectious and tumour antigens. However, the numerous side effects associated with the proinflammatory action of IL-1 represent a serious disadvantage for its use as a vaccine adjuvant. It was therefore of great interest that a nonpeptide contained in the IL-1 beta sequence (residues 163-171 corresponding to the sequence VQGEESNDK) is devoid of all proinflammatory activities but maintains the immunostimulating activity of the whole IL-1 beta. Thus, peptide 163-171 was successfully employed in animals to potentiate the specific immune response against T-helper-dependent cellular antigens, T helper-independent polysaccharidic antigens and recombinant as well as synthetic antigenic preparations derived from human pathogens. Furthermore, IL-1 and peptide 163-171 have been successfully used in tumour vaccines in experimental systems. It can therefore be concluded that peptide 163-171 is potentially a good candidate as vaccine adjuvant for human use.

Adjuvants, Immunologic↗
PubMed1992

Human peritoneal mesothelial cells produce many cytokines (granulocyte colony-stimulating factor [CSF], granulocyte-monocyte-CSF, macrophage-CSF, interleukin-1 [IL-1], and IL-6) and are activated and stimulated to grow by IL-1.

To investigate the role of peritoneal mesothelial cells in regulating hematopoiesis, as well as inflammation, healing, and tissue regeneration processes, long-term cultures of peritoneal mesothelial cells from human endocavitarian fluids were established. The purity of the cell population was assessed by morphologic and immunocytochemical criteria. Five peritoneal mesothelial cell cultures were analyzed for cytokine expression. Macrophage colony-stimulating factor (M-CSF), granulocyte-CSF (G-CSF), interleukin-1 alpha (IL-1 alpha), IL-1 beta, and IL-6 transcripts were constantly but variably detected throughout the culture period, while granulocyte-monocyte-CSF (GM-CSF) expression started as the cell culture aged. No IL-2, IL-3, IL-4, IL-5, or IL-7 transcripts were detected in the same samples. Corresponding cytokine activities were detected in the supernatants of the cultures. Peritoneal mesothelial cells proliferated after the addition of exogenous IL-1 beta or IL-1 alpha, whereas the addition of recombinant GM-CSF, G-CSF, M-CSF, or IL-6 failed to trigger proliferation. IL-1 receptor type I transcripts were detected in peritoneal mesothelial cells. Moreover, IL-1 was able to upregulate the expression of the genes that code for G-CSF, GM-CSF, IL-1 alpha, and IL-1 beta in these cells. These data indicate that peritoneal mesothelial cells produce many cytokines and suggest that IL-1 is a regulatory molecule for peritoneal mesothelial cells.

Antigens, CD↗
PubMed1992

Binding and internalization of the 163-171 fragment of human IL-1 beta.

The mechanisms of cell association of the human interleukin (IL-1 beta) immunostimulatory fragment 163-171 have been studied. The fragment was able to associate abundantly to both IL-1R- and IL-1R+ cells. Binding was strictly temperature dependent, was not saturable and could be inhibited by excess amounts of unlabelled 163-171 peptide but not by IL-1 beta, suggesting that the 163-171 fragment is not an IL-1R-binding domain of IL-1 beta. The fragment is readily internalized by cells by a cytochalasin-insensitive mechanism and it localizes mainly in the cytoplasm. It is concluded that the active domain 163-171 of IL-1 beta can be taken up by cells through a receptor-independent, temperature-dependent mechanisms and that its ability to activate cellular functions is based on IL-1R-independent intracellular pathways.

Amino Acid Sequence↗
PubMed1992

Effects of intraperitoneal recombinant interleukin-1 beta in intraperitoneal human ovarian cancer xenograft models: comparison with the effects of tumour necrosis factor.

The effect of intraperitoneal (i.p.) injection of recombinant human interleukin-1 beta (rhIL-1 beta) was studied in three i.p. nude mouse xenograft models of human ovarian cancer (HU, OS, and LA). Intraperitoneal rhIL-1 beta administration led to a dose dependent replacement of peritoneal ascitic tumour with solid tumours attached to the peritoneum and intraabdominal viscera in two (HU and LA) out of the three xenograft models. In the third xenograft model (OS), low doses of rhIL-1 beta (10 ng day) promoted micrometastatic peritoneal implants of tumour, but higher doses of rhIL-1 beta (1 microgram day) had a marked antitumour effect. This was due to direct cytotoxicity for tumour cells and was not related to peritoneal neutrophil influx induced by rhIL-1 beta. Recombinant human TNF (rhTNF) also promoted tumour implantation in all three xenograft models, but its antitumour effects differed from rhIL-1 beta. TNF increased the survival of HU and LA bearing mice, but had no antitumour effect in the OS xenograft model. Analysis of peritoneal fluid and tumour xenografts showed that TNF induced murine IL-1 in the tumour bearing mice. The magnitude of IL-1 induction indicated that TNF induced IL-1 did not contribute significantly to its effects.

Adult↗
PubMed1991

Endothelial cells express the interleukin-1 receptor type I.

Interleukin-1 (IL-1) profoundly affects a number of functions of vascular cells. Two distinct IL-1 receptors (IL-1R) are expressed on different cell types: the 80 Kd IL-1RI on T cells and fibroblasts, and the 68 Kd IL-1RII on B cells and myelomonocytic cells. The presence and functionality of IL-1R on vascular cells has been investigated by using polyomatransformed mouse endothelial cell (EC) lines (sEnd.1 and tEnd.1). These cells expressed specific and saturable binding sites for IL-1 (1,273 sites per cell with kd 9.5 x 10(-11) mol/L for sEnd.1, and 771 sites per cell with kd 8.5 x 10(-11) mol/L for tEnd.1, with radioiodinated IL-1 alpha as ligand). Binding of IL-1 was also evident at single cell level by autoradiography. By cross-linking studies, the molecular weight of the IL-1 binding protein on EC was approximately 80 Kd. This was confirmed by the presence in EC of mRNA for the 80 Kd IL-1RI. The IL-1RI on EC was apparently functional, since EC responded to IL-1 with IL-6 mRNA expression and IL-6 bioactivity production. These results were extended to human EC and vascular smooth muscle cells, which were also found to express mRNA for IL-1RI.

Animals↗
PubMed1991

Binding of IL-1 beta to IL-1R type II at single cell level.

To gain information on the possible biologic role of IL-1R type II (IL-1RII), expression of the 68-kDa IL-1 binding protein on human lymphoblastoid B cells was investigated at single cell level. Binding of iodinated IL-1 beta was evaluated by autoradiography on cytosmears of IL-1RII positive B cell lines RAJI, the RAJI clone 1H7, and STS 25. Results obtained suggest an heterogeneity of IL-1RII expression within the B cell population, with only 5 to 16% of the cells able to bind IL-1 beta. Up-regulation of IL-1RII expression by dexamethasone, evident in conventional binding assays, was achieved through both increase in the number of IL-1 binding cells (14-30%) and augmentation of receptor density on positive cells, By combining autoradiography with immunocytochemical staining, it could be shown that about 80% of IL-1RII + cells were negative for Ki67, a nuclear antigen expressed from late G1 to M phase. Cell cycle dependent expression of IL-1RII was confirmed on cells enriched in different phases of the cell cycle by counterflow centrifugal elutriation. It is thus proposed that IL-1RII is associated to the cell cycle.

Antigens, CD↗
PubMed1991

Quantitation of biologically active IL-1 by a sensitive assay based on immobilized human IL-1 receptor type II (IL-1RII).

A rapid and sensitive solid-phase radioassay is described for the quantitative detection of human interleukin-1 (IL-1) based on its capability to bind the nitrocellulose-immobilized IL-1 receptor solubilized from plasma membranes of a subclone of the human B cell lymphoma Raji. The assay can detect human IL-1 beta levels as low as 1 X 10(-11) M, both in physiological buffers and in human plasma. Much lower sensitivity was observed for human IL-1 alpha (3.7 X 10(-9) M) and murine IL-1 beta (2 X 10(-9) M). This assay has the advantage to specifically detect only the correctly folded biologically active IL-1. Simple pretreatment procedure that selectively removes IL-1 beta from samples has been devised so that the ratio of the two IL-1s isoforms in the sample can be precisely determined. This assay represents a fast method for the simultaneous-testing of large numbers of biological samples.

Filtration↗
PubMed1991

Murine interferon-gamma/interleukin-1 fusion proteins used as antigens for the generation of hybridomas producing monoclonal anti-interleukin-1 antibodies.

In several biological systems interferon-gamma (IFN-gamma) and interleukin-1 (IL-1) act synergistically. We therefore examined whether it would be possible to construct IFN-gamma/IL-1 hybrid proteins that would be more active than the individual components. Hybrid proteins were examined that consisted of the amino-terminal 118 residues of mouse IFN-gamma and the 156 or 152 carboxyl-terminal residues of mouse IL-1 alpha or IL-1 beta, respectively. They were obtained by ligation of the respective coding sequences and expression of the fused genes under control of the PL promotor in Escherichia coli. Both the IFN-gamma/IL-1 alpha and the IFN-gamma/IL-1 beta fusion proteins were purified by affinity chromatography on an anti-IFN-gamma monoclonal antibody column. Analysis of biological activities showed that these fusion proteins were less active than the individual cytokines. Specific antiviral activity of the IFN-gamma/IL-1 beta hybrids was less than 0.1% that of IFN-gamma and D10.G4.1 T-cell proliferative (IL-1) activity amounted to 0.1% that of mouse IL-1. Affinity-purified preparations of the IFN-gamma/IL-1 alpha hybrid were found to contain variable proportions of a Mr 14,000 degradation product possessing IFN-gamma activity, whereas the undegraded Mr 30,000 fusion protein, while being devoid of detectable IFN-gamma activity, did possess IL-1 activity (1%). Serum from rats immunized with the IFN-gamma/IL-1 alpha hybrid contained high levels of IL-1 alpha-binding and -neutralizing antibodies and IFN-gamma-binding antibodies, but no detectable levels of IFN-gamma-neutralizing antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Comparison of human interleukin-1 beta and its 163-171 peptide in bone resorption and the immune response.

Human interleukin-1 beta (IL-1 beta) caused a dose- and time-dependent enhancement of the release of 45Ca from prelabeled mouse calvaria in organ culture. In addition, IL-1 beta dose-dependently stimulated the formation of prostaglandin E2 (PGE2) and 6-keto-PGF1 alpha in the calvarial bones. However, IL-1 beta-induced 45Ca release was only partially inhibited by blocking the PGE2 response with indomethacin, suggesting that enhanced PGE2 formation in response to IL-1 beta is not necessary to obtain a bone resorptive effect, but that prostaglandins potentiate the action of IL-1 beta. The synthetic nonapeptide VQGEESNDK, corresponding to the fragment 163-171 of human IL-1 beta, administered simultaneously with antigen (SRBC) to C3H/HeN male mice, induced a dose-dependent enhancement of specific antibody-producing cells in the spleen (PFC). The degree of PFC stimulation was comparable to that caused by native human IL-1 beta. In mouse bone cultures, neither 45Ca release nor prostanoid formation was stimulated by fragment 163-171. These data indicate that (1) IL-1 beta-induced stimulation of bone resorption is dissociable from IL-1 beta-induced increase of prostanoid biosynthesis and (2) the epitope of the IL-1 beta molecule involved in the immunostimulatory effects may be different from that involved in the stimulatory effects on bone resorption.

Amino Acid Sequence↗
PubMed1991

Cytokine-associated tissue injury and lethality in mice: a comparative study.

A comparative study was performed to examine the lethal effects of several cytokines injected into mice sensitized with actinomycin D (Act-D). Consistent with published data, human tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) (0.2-5 micrograms) caused the death of the animals within 8-12 hr after injection. Human interleukin-6 (IL-6) and interleukin-8 (IL-8) (0.6-6 micrograms) known to be induced by TNF-alpha did not show any lethal effects, indicating that TNF-alpha-associated lethality is not mediated by IL-6 or IL-8. Human tumor necrosis factor-beta (TNF-beta) (also called lymphotoxin), which shares structural and functional properties with TNF-alpha, was as potent as TNF-alpha in its lethal effects. Murine interferon-gamma (IFN-gamma) (0.04-5 micrograms) was also tested and showed no lethal effects in this model. In addition, a synthetic peptide corresponding to amino acid residues 163-171 of IL-1 beta, and which has been shown to lack the inflammatory effects of IL-1 beta, also caused no lethality among Act-D sensitized mice. The pretreatment of mice with IL-6, IL-8, or IFN-gamma had no protective effects on TNF-alpha or IL-1 beta-induced lethality in contrast to the protection observed by a pretreatment with TNF-alpha/IL-1 beta themselves or with endotoxin. Histopathologic data showed that severe tissue injury in vital organs is associated with the rapid lethality among sensitized mice.

Animals↗
PubMed1991

Differential inhibition of IL-1 beta activities and receptor binding by monoclonal antibodies mapping within a discrete region of the protein.

The importance of the region in position 148-192 for the biological activities and receptor-binding capacity of the human IL-1 beta protein has been assessed by the use of mAbs. Four mAbs have been used, which recognize different epitopes within the 148-192 region. None of the mAbs could inhibit binding of IL-1 beta to IL-1RI (expressed on T cells and fibroblasts), suggesting that the 148-192 region does not contain IL-1RI binding sites. Conversely, mAbs Vhp20 (recognizing the fragment 166-169) and BRhD2 (directed to an epitope in the sequence 177-186) recognize sites partially involved in binding to IL-1RII (expressed on B cells, macrophages, and PMN). Only mAbs BRhD2 and FIB 1 (which recognizes an epitope in the sequence 174-186) can inhibit IL-1 beta-induced thymocyte proliferation, whereas all four can inhibit the adjuvant capacity of IL-1 beta in vivo. It is concluded that the region 148-192 encompasses domains important for T cell activation but not for binding to the IL-1RI on T cells, others involved in immunostimulation in vivo, and others important for binding to IL-1RII, although not directly involved in it.

Amino Acid Sequence↗
PubMed1991

Immunohistochemical localization of IL-1 alpha and IL-1 beta in normal human placenta.

Many reports show that interleukin 1 (IL-1) is produced by mouse and human placenta but the cell type that is responsible for this production has yet to be identified. For this reason we attempted to localize IL-1 alpha and IL-1 beta directly on formalin-fixed paraffin-embedded normal human placentae at different stages of pregnancy using immunohistochemical techniques and specific antibodies. The results obtained show that both IL-1 forms are localized to villous syncytiotrophoblast and to extravillous trophoblast, while villous cytotrophoblast and cytotrophoblast columns are unreactive. A gradual decrease of reactivity was observed with increasing gestation age for both IL-1 forms, but the staining for IL-1 beta was in all sections higher than for IL-1 alpha. Although the physiological role of IL-1 in pregnancy has yet to be established, the presence of this cytokine in the cells facing maternal blood and tissues suggests a possible involvement in the immunoregulation of fetal acceptance.

Female↗
PubMed1991

Radiation protection and restoration by the synthetic 163-171 nonapeptide of human interleukin 1 beta.

We have previously reported that the synthetic nonapeptide VQGEESNDK, position 163-171 of human interleukin 1 (IL-1 beta), when injected in immunodepressed mice, shows immunorestorative activity similar to that of the whole protein, but with no IL-1-like inflammatory effects [Frasca et al., J. Immunol. 141, 2651-2655 (1988)]. In the present study we have compared the protective and restorative activities of the nonapeptide and human recombinant (hur) IL-1 beta on the survival of lethally irradiated mice. When mice were given a single injection of different doses of the nonapeptide or hurIL-1 beta 20 h before total-body irradiation, both molecules increased the percentage survival of mice exposed to 750 or 850 cGy, but not to 950 cGy. The nonapeptide, however, is less effective than hurIL-1 beta and displays a different dose-response relationship, suggesting that the two molecules act through different radioprotective pathways. When mice were injected with the nonapeptide or hurIL-1 beta immediately after exposure to 850 cGy, the percentage survival was also increased but restoration was lower than protection in both cases. The nonapeptide was also less effective than hurIL-1 beta in restoration, but the two molecules displayed a comparable dose-response relationship as if they shared similar mechanisms. These findings indicate that the 163-171 nonapeptide is able to protect from lethal radiation injury and to restore viability. The nonapeptide appears less effective than hurIL-1 beta but does not exhibit the IL-1-like side effects of the whole molecule.

Adjuvants, Immunologic↗
PubMed1990

Interleukin 1-induced augmentation of experimental metastases from a human melanoma in nude mice.

This study has examined the effect of the cytokine interleukin 1 (IL-1) on metastasis formation by the human melanoma A375M in nude mice. We have found that human recombinant IL-1 beta (a single injection greater than 0.01 micrograms per mouse i.v. given before tumor cells) induced an augmentation of experimental lung metastases from the A375M tumor cells in nude mice. This effect was rapidly induced and reversible within 24 h after IL-1 injection. A similar effect was induced by human recombinant IL-1 alpha and human recombinant tumor necrosis factor, but not by human recombinant interleukin 6. 5-[125I]odo-2'-deoxyuridine-radiolabeled A375M tumor cells injected i.v. remained at a higher level in the lungs of nude mice receiving IL-1 than in control mice. In addition, IL-1 injected 1 h, but not 24 h, after tumor cells enhanced lung colonization as well, thus suggesting an effect of IL-1 on the vascular transit of tumor cells. These findings may explain the observation of enhanced secondary localization of tumor cells at inflammatory sites and suggest that modulation of secondary spread should be carefully considered when assessing the ability of this cytokine to complement cytoreductive therapies.

Animals↗