Biomedical subjects

L B Lachman

Publications and source records attributed to L B Lachman.

At least 37 records · Page 2Linked to original sources

PubMed1989

Detection and characterization of interleukin-1 in human dental pulps.

Pulp tissue was obtained from clinically intact third molars and premolars, and from teeth affected by caries and periodontal disease. After incubation and homogenization, supernatants were centrifuged, sampled and assayed for lymphocyte activating factor-like activity using the thymidine-incorporated D10.G4.1 cell-line assay. Significantly higher levels of mitogenic activity were found in the pulp supernatants of symptomatic carious teeth, and in those of third molars with attendant pericoronitis, than in pulps from symptomatic and asymptomatic periodontally affected teeth and asymptomatic third molars and premolars. As both interleukin-1 (IL-1) and interleukin-2 (IL-2) may be involved in the proliferation of D10.G4.1 cells, supernatants from carious pulps were further tested for IL-2 activity using murine natural killer (NK) cells. Those that had elicited a proliferative response of D10.G4.1 cells did not enhance NK proliferation, indicating that the factor responsible for clonal replication of D10.G4.1 cells was IL-1. To confirm the presence of IL-1, immunohistochemistry using a monoclonal antibody to IL-1 was performed on frozen and paraffin-embedded sections of pulps from each group. IL-1 was immunolocalized within cells in pulps from the caries symptomatic group. The cell type producing IL-1 was further characterized as an alpha-naphthyl-acetate-esterase-positive macrophage within the connective tissue stroma of pulps from the caries symptomatic group. This novel demonstration of IL-1 and IL-1 producing cells in human dental pulp indicates the involvement of this mediator of inflammation in dental disease.

Cells, Cultured↗

PubMed1989

Stimulation of glycosaminoglycan synthesis in cultured human dermal fibroblasts by interleukin 1. Induction of hyaluronic acid synthesis by natural and recombinant interleukin 1s and synthetic interleukin 1 beta peptide 163-171.

Hyaluronic acid (HA) is believed to play a critical role in wound healing and in morphogenesis. Factors controlling the production of HA by fibroblasts in normal and pathological states are not completely understood. In this report we have observed that natural human interleukin (IL-1)1 beta and human recombinant (hrIL)-1 alpha and beta are potent stimulators of HA production by fibroblasts in vitro. Hyaluronic acid is the major species of glycosaminoglycan (GAG) stimulated by IL-1 in fibroblasts. PGE2 does not appear to be involved directly in this IL-1 effect on fibroblasts, but stimulation of HA production by IL-1 is dependent on protein synthesis. The synthetic human IL-1 beta peptide 163-171 (Val-Gln-Gly-Glu-Glu-Ser-Asn-Asp-Lys), which has been previously shown to stimulate thymocyte proliferation but not fibroblast PGE2 production, is also able to stimulate fibroblast HA production. The synthesis and secretion of IL-1 by mononuclear phagocytes at sites of inflammation and immune reactions in vivo could potentially serve as a signal for fibroblasts to synthesize HA, which in turn could serve to facilitate and modulate reparative and immune processes by virtue of its ability to alter cell-cell, cell matrix, and cell-membrane receptor interactions.

Cells, Cultured↗

PubMed1989

Heterogeneity of interleukin 1 production in cultured Reed-Sternberg cell lines HDLM-1, HDLM-1d, and KM-H2.

Interleukin 1 (IL-1) is known for its role in modulating the immune response and is required for initiation of lymphocyte proliferation by means of increased IL-2 production by lymphocytes. Previously, the expression of IL-1 in H-RS cells in tissue sections was shown by using immunoperoxidase staining. For further confirmation, the production of IL-1 in cultured cells of the H-RS cell lines HDLM-1, HDLM-1d, and KM-H2 was examined by using a murine D10.G4.1 T cell proliferation bioassay and Northern blot hybridization with specific IL-1 cDNA probes. It was confirmed that two types of H-RS cells, HDLM-1 and KM-H2, can secrete IL-1, especially after treatment with phorbol ester. The amounts of IL-1 in H-RS cell culture medium ranged from approximately 0.5 to 2.5 ng/ml. The major IL-1 secreted by HDLM-1 cells was IL-1 alpha, and by KM-H2 cells was IL-1 beta. HDLM-1d cells did not produce IL-1. This finding indicates the heterogeneity of IL-1 production in H-RS cells. Such heterogeneity may apply to H-RS cells in vivo, based on the variable IL-1 staining of these cells in lymphoid tissues.

Animals↗

PubMed1988

Effect of recombinant granulocyte/macrophage colony-stimulating factor on human monocyte activity in vitro and following intravenous administration.

The purpose of these studies was to examine the antitumor properties of blood monocytes from patients undergoing a phase I trial with recombinant granulocyte/macrophage colony-stimulating factor (rGM-CSF). Peripheral blood monocytes from 7 patients receiving various doses of rGM-CSF by continuous infusion were isolated prior to therapy and at various times during the 2-wk infusion. Monocytes/cubic centimeter of blood increased in a dose-dependent fashion in patients receiving rGM-CSF. However, activation of monocyte-mediated tumorilytic properties was seen in only 1 of 7 patients. rGM-CSF administration also did not stimulate interleukin-1 or tumor necrosis factor production by blood monocytes. The failure to detect monocyte-mediated tumoricidal activation was not secondary to an inherent "defect" in the patients' monocytes because in vitro incubation with lipopolysaccharide alone or human recombinant gamma-interferon plus muramyl dipeptide resulted in monocyte-mediated cytotoxicity and in the secretion of interleukin-1 and tumor necrosis factor. rGM-CSF in vitro also did not stimulate the tumoricidal function of normal monocytes or the secretion of interleukin-1 or tumor necrosis factor. We concluded that systemic administration of rGM-CSF did not result in routine activation of monocyte-mediated cytotoxicity but did result in a dose-dependent rise in the number of circulating monocytes. Because these monocytes could be activated in vitro, we propose that, in an adjuvant therapy setting, rGM-CSF be combined with other activating agents to increase the pool of potential killer cells.

Cells, Cultured↗

PubMed1988

Plasma membrane-associated tumor necrosis factor. A non-integral membrane protein possibly bound to its own receptor.

Purified plasma membranes from LPS-activated human blood monocytes produced significant lysis and growth inhibition of the TNF-sensitive L929 murine fibroblast cell line. Unactivated human monocyte plasma membranes did not display either activity. Anti-TNF serum specifically inhibited the anti-tumor activity of activated monocyte membranes whereas anti-IL-1 serum or non-specific rabbit serum decreased neither the lysis nor growth inhibition of L929 cells. Membrane-associated TNF did not behave as an integral protein as it could be eluted from the plasma membranes by either high salt or low pH treatment. Plasma membranes cleared of membrane-associated TNF by high salt treatment were able to bind TNF, and this binding was specifically inhibited by preincubation of rTNF with specific anti-TNF serum. Western blot analysis of plasma membranes showed a membrane-associated TNF with a m. w. of approximately 17 kDa present only in the activated monocytes. When the plasma membranes were preincubated with the cross-linker agent dissuccinimidyl suberate, Western blot analysis revealed the presence of a TNF-binding protein with a Mr of approximately 102 kDa. These studies indicate that unlike IL-1, membrane-associated TNF is not an integral membrane protein and that TNF may be associated with the monocyte membrane by occupying the TNF R.

Cell Membrane↗

PubMed1988

Interleukin-1 injected into mammalian brain stimulates astrogliosis and neovascularization.

Interleukin-1 (IL-1), a protein produced by mononuclear phagocytes, helps to initiate the inflammatory response through its action upon a diverse population of cells. Recently this immunomodulator has been detected at sites of traumatized brain. As reported here, recombinant forms of IL-1 injected into the cerebral cortex of adult rats elicit not only astrogliosis but also new blood vessel growth. These responses are typical of brain injury and suggest that IL-1-secreting inflammatory cells may mediate wound healing in the CNS.

Animals↗

PubMed1988

Interleukin-1 is an astroglial growth factor in the developing brain.

The immunomodulator interleukin-1 (IL-1) is found to be an astroglial growth factor during development of the mammalian brain. In vitro studies indicate that ameboid microglia, a class of brain mononuclear phagocytes, are the likely source of IL-1. Examination of different brain regions during development shows IL-1 production only after the appearance of ameboid microglia. These observations suggest that brain mononuclear phagocytes secrete growth factors that regulate normal growth and development of the CNS.

Animals↗

PubMed1988

Cytotoxic liposomes: membrane interleukin 1 presented in multilamellar vesicles.

Paraformaldehyde-fixed lipopolysaccharide (LPS)-activated human monocytes produced significant lysis of the human melanoma cell line A375. The cytotoxic activity was retained following treatment of the fixed monocytes with anti-tumor necrosis factor (anti-TNF) antibodies but was specifically inhibited by a mixture of anti-TNF and anti-interleukin 1 (anti-IL 1) antibodies. A375 cells were also killed by plasma membranes purified from LPS-activated human blood monocytes. This activity was specifically inhibited by anti-IL 1 alpha antibodies, but only partially inhibited by anti-IL 1 beta antibodies. CHAPS detergent-extracted plasma-membrane IL 1 in its soluble form or associated with lyophilized liposomes was also able to kill A375 cells, and this antitumor activity was inhibited by anti-IL 1 antibodies. These results suggest that membrane IL 1, primarily IL 1 alpha, was cytotoxic for the A375 cells. CKS-17, a peptide synthesized with homology to a highly conserved region of the immunosuppressive retroviral envelope protein P15E, when covalently bound to BSA partially inhibited the IL 1 activities of tumor cell cytotoxicity and T-cell clone proliferation, displayed by purified plasma membranes, detergent-extracted membrane IL 1, or membrane IL 1 associated with liposomes. These findings indicate that cytotoxic membrane IL 1 can be solubilized by detergent, bound to the surface of liposomes, and specifically inhibited by anti-IL 1 antibodies or the immunosuppressive peptide CKS-17.

Cell Membrane↗

PubMed1987

A synthetic peptide homologous to the envelope proteins of retroviruses inhibits monocyte-mediated killing by inactivating interleukin 1.

The synthetic peptide CKS-17 has homology to a highly conserved region of the immunosuppressive retroviral envelope protein P15E, to envelope proteins of HTLV I, II, III, and to that encoded by an endogeneous C-type human retroviral DNA. CKS-17 inhibits the immune response of lymphocytes and the respiratory burst of human monocytes. Because P15E-related antigens are present in human malignant cell lines and cancerous effusions, we sought to determine the effect of CKS-17 on monocyte-mediated tumor cell lysis. Lysis of A375 tumor cells by lymphokine or lipopolysaccharide-activated human monocytes was inhibited by 10 microM CKS-17 (control, 79%; CKS-17-treated, 19%). Another synthesized peptide, CS-2, which has partial homology to CKS-17, failed to block monocyte-mediated killing. Thus, the inhibition by CKS-17 appeared to be specific. Because interleukin 1 (IL-1) is a cytocidal factor produced by activated monocytes, we also investigated the effect of CKS-17 on IL-1 production by monocytes and on direct IL-1-mediated cytotoxicity. CKS-17 did not block production or secretion of IL-1 by lipopolysaccharide- or interferon-gamma-activated monocytes. However, the direct cytocidal activity of both recombinant IL-1 alpha and IL-1 beta against A375 tumor cells was blocked by CKS-17. The cytotoxic activity of IL-1 was inhibited by CKS-17 if (a) IL-1 was preincubated with CKS-17 for 1 hr at 37 degrees C or (b) the A375 cells were incubated with CKS-17 for 1 hr prior to the addition of IL-1. CKS-17 also blocked IL-1-induced proliferation of murine thymocytes, the D10 T cell line, and an IL-1-responsive astrocytoma cell line. These data suggest that CKS-17 may be a potent inhibitor of IL-1.

Cytotoxicity, Immunologic↗

PubMed1987

Interleukin 1 release by human monocytes treated with liposome-encapsulated lipopolysaccharide.

Liposome therapy offers a unique method of delivering antifungals, antiprotozoans, and macrophage-activating factors directly to reticuloendothelial cells. Although liposomes are entering clinical trials, their effect on Il 1 synthesis and release has yet to be determined. The beneficial, as well as possible pathological, effects of liposome therapy may be due to IL 1 released by reticuloendothelial cells. This study examined the effects of multilamellar phospholipid vesicles on the synthesis and release of IL 1 from human peripheral blood monocytes. Liposomes consisting of phosphatidylcholine and phosphatidylserine in molar ratio of 7:3 did not stimulate IL 1 release, and did not diminish the level of IL 1 release when monocytes were subsequently stimulated with LPS or Staphylococcus aureus. Surprisingly, LPS encapsulated in liposomes failed to stimulate IL 1 release from monocytes. This defect was limited to the release of IL 1, because monocytes stimulated with liposomes containing LPS had control levels of intracellular (cytosolic) and membrane IL 1. By contrast, LPS associated with lyophilized liposomes induced the secretion of IL 1 and behaved similarly to free LPS. These findings indicate that LPS activation of monocytes may involve not only the synthesis of IL 1 but activation of the secretory mechanism for IL 1, and that these two events could be independent, that the density of LPS molecules at the surface of the liposome may influence the degree of monocyte activation as measured by intracellular IL 1 synthesis, membrane accumulation, and secretion into the medium, and that intracellular lipids may not interfere with the secretory mechanism, because liposomes made with phospholipids differing in acyl chain length or the headgroup behaved identically.

Cell Membrane↗

PubMed1987

Subcellular localization of human monocyte interleukin 1: evidence for an inactive precursor molecule and a possible mechanism for IL 1 release.

IL 1 activity, as assayed by the proliferation of responsive mouse thymocytes and a human astrocytoma cell line, was detected on the membrane of 1% paraformaldehyde-fixed activated human monocytes. Resting, unactivated monocytes did not display IL 1 activity. Maximum induction of membrane IL 1 was obtained from monocytes treated with polyclonal activators, such as LPS or Staphylococcus aureus, whereas adherence was a weak inducer of membrane IL 1. Isolated cell compartments as plasma membranes, crude lysosomes, and crude cytosol from activated human monocytes expressed significant IL 1 activity, whereas the endoplasmic reticulum showed no IL 1 activity. Exposure to trypsin of either fixed, activated human monocytes or cell compartments from unfixed monocytes, revealed biologically active IL 1 in the membrane, crude lysosome, and crude cytosol, but not in the endoplasmic reticulum. The IL 1 activity in the purified cytosol, prepared by extraction with digitonin, was considerably increased by the trypsin treatment, whereas the increase in IL 1 activity within crude lysosomes and plasma membranes was less. The cell compartments from nonactivated monocytes did not express active IL 1 and trypsin treatment revealed no active IL 1, suggesting the absence of a pool of the trypsin-sensitive form of IL 1. The data confirm the presence of membrane-bound IL 1 in activated human monocytes and indicate that an inactive precursor molecule can be found in the cytosol of such cells. Furthermore, the absence of IL 1 activity either in its active form or as the inactive precursor in the endoplasmic reticulum suggests that IL 1 is not a conventionally secreted protein. Because IL 1 was found in the cytosol as a precursor and in the lysosomal fractions in an active form, these data suggest that after the synthesis and processing of the cytosolic precursor, the 17-kda IL 1, is released via lysosomal vesicles.

Cell Membrane↗

PubMed1987

Liposomes expressing IL 1 biological activity.

We determined the activity of IL 1, obtained from various human monocyte subcellular compartments, when associated with liposomes. Soluble IL 1, bound to the outer surface of lyophilized liposomes, stimulated responsive target cells. However, this activity was not preserved when soluble IL 1 was incorporated into the inner chambers of classical liposomes. In contrast, monocyte plasma membranes that exhibited IL 1 activity had the same level of activity when presented on lyophilized liposomes and when incorporated inside the classical liposomes. However, monocyte plasma membranes bound to the outer surface of the liposomes exhibited greater activity than the monocyte membrane IL 1 itself in its soluble form. This suggests that membrane IL 1 is an integral membrane protein, readily integrated into the lipid bilayers. Like soluble IL 1, the expression of IL 1 activity present in the cytosol of activated monocytes was decreased by incorporation into liposomes, but was high and active when presented on lyophilized liposomes. The best artificial cell reconstitution was obtained with lyophilized liposomes in association with monocyte cytosol and plasma membranes. When an unactivated monocyte compartment was mixed with one from an activated monocyte, the signal was equal to that of the activated cell compartment alone. The IL 1 activity of activated cell fractions associated with lyophilized liposomes was determined, and an increase of IL 1 activity for both plasma membranes and cytosol was observed, whereas a decrease of the signal was obtained for the lysosomal compartment. Endoplasmic reticulum showed no IL 1 activity, even after trypsin treatment. The highest activity after trypsin treatment was recovered in the cytosol associated with lyophilized liposomes, suggesting that molecules obtained after this treatment were able to bind tightly to the lipid bilayers.

Cell Compartmentation↗

PubMed1987

Growth-promoting effect of recombinant interleukin 1 and tumor necrosis factor for a human astrocytoma cell line.

IL 1 can exhibit dichotomous effects in the sense that it is cytotoxic for certain cells, although growth promoting for other cells. Because IL 1 is growth promoting for astrocytes, but cytotoxic for melanoma cells, the current investigation evaluated the effect of IL 1 upon astrocytomas. The human astrocytoma U373 was found to incorporate [3H]thymidine after exposure to recombinant human IL 1 alpha and IL 1 beta and murine IL 1 alpha. Surprisingly, U373 also incorporated [3H]thymidine after exposure to recombinant TNF. The response of the U373 to IL 1 may be used as a simple and sensitive assay for IL 1.

Astrocytoma↗

PubMed1987

B-cell line-derived interleukin 1 is cytotoxic for melanoma cells and promotes the proliferation of an astrocytoma cell line.

We have previously described a unique molecular species of Interleukin 1 spontaneously produced by a human EBV-transformed B-cell line. This IL 1 shares a number of biological activities with monocytic IL 1 although its NH2-terminal amino-acid sequence is different from both IL 1 alpha and beta. In the present study we have investigated the activity of this B-cell IL 1 in two recently reported new assays for interleukin 1. We found that this B-cell IL 1 is able to promote the proliferation of the human astrocytoma cell line U373 in a dose dependent manner. We also describe that B-cell IL 1 is directly cytotoxic for a melanoma cell line, A375, but not for the tumor necrosis factor target cell, the murine transformed fibroblast line L929. These studies should prove useful in analysing structure-function relationships of the various IL 1 species, when the primary sequence of B-cell IL 1 becomes available.

Animals↗

PubMed1986

Interleukin 1 of the central nervous system is produced by ameboid microglia.

By screening specific populations of rat brain cells, we found that ameboid microglia secrete an 18 kD peptide with IL-1 biological activity. The IL-1 activity released by microglia was found to be identical to rat macrophage IL-1 on fractionation by gel filtration and high pressure liquid anion-exchange chromatography, and it was neutralized by an antiserum specific for murine IL-1. When added to astroglia grown in culture, microglial IL-1 increased the cell number of five- to sevenfold, and increased astroglial incorporation of [3H]thymidine by three- to fivefold. We propose that the proliferation of astroglia in specific brain regions may be regulated by the signaled release of IL-1 from activated microglial cells.

Animals↗

PubMed1986

Natural and recombinant human interleukin 1-beta is cytotoxic for human melanoma cells.

Human peripheral blood monocytes stimulated with LPS were found to release an activity that was cytotoxic for the A375 melanoma. Biochemical and immunological characterization of the activity indicated that IL 1-beta was the cytotoxic agent. Human recombinant IL 1-beta, purified to homogeneity, was directly cytotoxic for A375. Tumor necrosis factor, also released by activated monocytes, was not cytotoxic for the A375 melanoma.

Antineoplastic Agents↗

PubMed1986

Monocyte-conditioned medium, interleukin-1, and tumor necrosis factor stimulate the acute phase response in human hepatoma cells in vitro.

Human hepatoma cells mimic the acute phase response after treatment with monocyte-conditioned medium. Levels of secreted fibrinogen, alpha-1 acid glycoprotein, C-reactive protein, haptoglobin, and the third component of complement were elevated compared with control levels after 48 h of incubation with conditioned supernatant medium from an enriched fraction of normal peripheral monocytes. Albumin levels declined and alpha-1 antitrypsin remained unchanged. Levels of specific mRNA were measured by hybridization to slot blots and Northern blots and changed in correspondence with protein alterations. Interleukin-1 and tumor necrosis factor stimulated the third component of complement, but did not elevate any other member of the acute phase group and were therefore only partially active in this system. The identification of an in vitro model of the human acute phase response will permit analysis of the molecular basis for coordinate regulation of this group of facultative genes.

C-Reactive Protein↗

PubMed1986

Catecholamine-induced suppression of interleukin-1 production.

Recent evidence indicates that stress can suppress immune responses and thus increase the severity of viral and neoplastic diseases. Although, the mechanisms for stress-induced modulation of immunologic competence are unclear, neuroendocrine hormones are thought to be involved. A direct suppressive effect could result from the action of neuroendocrine hormones on lymphokine and monokine release. The central role of interleukin-1 (IL-1) in regulating cellular immune responses to infection and neoplasms stimulated the present study evaluating the effects of neuroendocrine hormones on IL-1 production. Norepinephrine and epinephrine inhibited the capacity of gamma interferon and lipopolysaccharide to stimulate IL-1 production from mouse peritoneal macrophages. Moreover, when intracellular and extracellular levels of IL-1 were quantitated, the studies demonstrated a catecholamine-mediated block in IL-1 synthesis without effect on its release. We also observed that exogenous cyclic AMP (cAMP) administered to mouse macrophages suppressed IL-1 production. This, coupled with the capacity of norepinephrine and epinephrine to enhance intracellular cAMP levels in macrophages, strongly suggested that the catecholamine-induced suppression of IL-1 production may be mediated by elevated intracellular cAMP levels. These findings demonstrate that selected stress-related neuroendocrine hormones can modulate IL-1 production by macrophages and further support the hypothesis that alteration of macrophage function by neuropeptides and neurohormones is a significant feature of stress-induced enhancement of viral and neoplastic disease.

Animals↗