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J Bernardo

Publications and source records attributed to J Bernardo.

At least 19 recordsLinked to original sources
PubMed1996

Rifampin preventive therapy for tuberculosis in Boston's homeless.

An epidemic of isoniazid (INH)- and streptomycin (SM)-resistant tuberculosis began among Boston's homeless population in 1984. Individuals with skin test conversions who agreed to preventive therapy received either INH, rifampin, or a combination of INH and rifampin. A total of 204 individuals with documented tuberculin skin test conversions who did not have active tuberculosis at the time of the clinical evaluation for their positive skin test were eligible for preventive therapy. Data on type and length of preventive therapy were obtained from the Tuberculosis Clinic and the Boston Tuberculosis Registry records at Boston City Hospital. The individuals were followed for development of active tuberculosis. Six of 71 (8.6%) individuals who received no therapy, 3 of 38 (7.9%) in the INH group, and none in the rifampin or rifampin plus INH groups (49 and 37 persons, respectively) developed active tuberculosis. Patients in the rifampin group were significantly less likely to develop tuberculosis than patients in the no therapy group (p = 0.04; odds ratio [OR] = 0.00, 95% confidence interval [CI] = 0.00-0.91). Treatment with any rifampin-containing preventive therapy (rifampin or rifampin plus INH) was effective (p < 0.01 ) in preventing development of active disease. The three INH failures were with organisms that were resistant to INH.

Adult↗
PubMed1995

Neutrophil functional responses depend on immune complex valency.

Ligand-induced cross-linking of Fc gamma receptors (Fc gamma R) on neutrophils plays a significant role in their stimulation, shown here by contrasting the responses induced by low valency immune complexes (LICs) and high valency immune complexes (HICs) and by cross-linking LICs in situ (L/Ab) after their addition to the cells. Multiparameter flow cytometry was used to measure immune complex (IC)-elicited changes in cytoplasmic Ca2+ concentration and initiation of the oxidative burst simultaneously in the same cell and to correlate these with Fc gamma R occupancy. We have previously shown that subpopulations of neutrophils respond maximally to subsaturating concentrations of HIC; saturating dosages stimulate the entire population. This discrepancy was not due to differences in receptor occupancy. The magnitude of the transient Ca2+ increase was independent of the dose of HIC but depended on the dose when an LIC was used. As shown here, L/Ab cross-linking elicited Ca2+ responses similar to those observed in HIC-stimulated cells. In contrast, LIC elicited only minimal intracellular delta pH and no oxidative burst or membrane potential changes at all unless Fc gamma R was cross-linked, accomplished by HIC or by L/Ab. However, azurophilic degranulation, as determined by elastase release, was not observed in cells stimulated by the in situ cross-linking method, whereas the HIC preparation triggered azurophilic degranulation. Thus, some Fc gamma R-mediated neutrophil effector functions such as azurophilic degranulation and oxidative burst initiation have an absolute requirement for Fc gamma R cross-linking, whereas signaling functions such as changes in membrane potential, intracellular pH, and intracellular Ca2+ concentration can occur, albeit more slowly and to a lesser extent, if single Fc gamma R are occupied.

Amiloride↗
PubMed1995

Role of the Fc gamma R subclasses Fc gamma RII and Fc gamma RIII in the activation of human neutrophils by low and high valency immune complexes.

Two Fc gamma receptor (Fc gamma R) subclasses on human neutrophils, Fc gamma RII and Fc gamma RIII, activate different cellular functions. To examine the involvement of each receptor subtype in polymorphonuclear leukocyte activation, Fab and F(ab')2 fragments of subclass-specific monoclonal antibodies ([mAbs] mAb IV.3 against Fc gamma RII and mAb 3G8 against Fc gamma RIII, respectively) were used to block the binding of low valency immune complexes (LICs) and high valency immune complexes (HICs). Flow cytometry then permitted the simultaneous quantitation of antibody and ligand binding, the elicited intracellular Ca2+ concentration (delta[Ca2+]int), initiation of the oxidative burst, and/or the phospholipase A activation in the same cell. We have previously demonstrated that subsaturating dosages of HIC bind uniformly to all the cells but elicit an "all-or-none" (i.e., dose independent) maximal delta[Ca2+]int in a dose-dependent subpopulation of the cells. In contrast, both the proportion of cells responding and the magnitude of the delta[Ca2+]int transient depend on the subsaturating dose of LIC, even though it too binds uniformly to all the cells, nonresponding as well as responding. These earlier findings have here been extended by single cell flow cytometric analysis to demonstrate that F(ab')2 Fc gamma RIII is the major Fc gamma R involved in HIC binding (and [Ca2+]int mobilization), as well as in oxidative burst and phospholipase A activation. In contrast, both receptor subclasses must be available for LIC-elicited delta[Ca2+]int, as blockage by either of the mAb Fab or F(ab')2 fragments abrogates this response, even though LIC binding to the receptors is not decreased. Furthermore, LIC elicited little oxidative burst activity and failed to activate phospholipase A but cross-linking to achieve multivalency, previously shown to induce [Ca2+]int and oxidative burst responses, elicited phospholipase A activity via Fc gamma RIII. Fc gamma RII's role appears to be modulation of the small, late Ca2+ influx observed at > 1 min, whereas Fc gamma RIII modulates all the earlier larger events. Thus, simultaneous observation of receptor identity, receptor occupancy, and consequent activation parameters in the same cell by flow cytometry permits use to demonstrate that Fc gamma RII is necessary for the small signal transduction elicited by LIC; it plays a relatively small role in polymorphonuclear leukocyte stimulation by HIC. Fc gamma RIII is the main receptor responsible for immune complex-elicited polymorphonuclear leukocyte responses; its efficacy is greatly enhanced when the receptors are cross-linked, either by preequilibrated multivalent complexes or by in situ cross-linking of bound LIC with excess antibody.

Antibodies, Monoclonal↗
PubMed1994

Cytokine binding to CD4+ inflammatory cells: implications for asthma.

While LCF is present in BAL early after antigen challenge, we know little about its other potential effects beyond CD4+ T cell, monocyte, and eosinophil chemotaxis and monocyte and CD4+ T cell activation. The work described here focuses on the hypothesis that the secreted protein products of T cells participate in the airway inflammatory process that underlies human asthma, and in particular that LCF could play an early role because of the unusual responsiveness of LCF-producing T to histamine. To date, most studies have addressed the measurement of cytokines derived from CD4+ T cells (e.g., IL-2, IL-3, IL-4, IL-5, and GM-CSF) in the airways of asthmatics, and attempted to correlate the presence of protein or mRNA with the complexion of the inflammatory infiltrate. These studies have been based upon the reports that there are increased numbers of CD4+ T cells in the airways of asthmatics, and that the presence of eosinophils might correlate with the secretion of TH2-type cytokines like IL-3, -4, and -5. Using this information as a background, our work has approached the problem in an entirely different way. We have focused our attention on the early events in antigen-induced asthma that are responsible for CD4+ cell accumulation in the lung, including CD4+ T cells, eosinophils, and monocytes. We have attempted to identify mechanisms by which mast cell mediators, in particular histamine, might play a role in the secretion of chemotactic lymphokines that are selective for CD4+ cells by using CD4 itself as a chemotactic factor receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Chemotactic peptide-induced cytoplasmic pH changes in incubated human monocytes.

Stimulation of phagocytic leukocytes with chemotactic factors results in transient acidification, followed by alkalinization of the cytosol. Human monocytes are known to alter their functional responses to the chemotactic peptide N-formylmethionyl-leucyl-phenylalanine (fMLP) in a complex fashion as they mature in vitro to macrophages. To examine the evolution of the cytoplasmic pH (pHi) response of monocytes to fMLP as they mature into macrophages, we incubated cells for 0, 24, 48, and 96 h (Medium-199 + 10% fetal bovine serum; 37 degrees C) and examined pHi using the fluorescent probe 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF; 1 microM) and a Perkin-Elmer 650/10 spectrofluorimeter (lambda em = 530 nm, lambda ex = 500, 450 nm) as previously described. The resting pHi of fresh (0 h) monocytes was 7.07 +/- 0.16 (SD) and was unchanged after incubation for 24, 48, or 96 h (7.09, 7.11, 7.05, respectively). Cells exhibited an fMLP dose-dependent cytoplasmic acidification, with maximal delta pHi occurring 30-60 s after exposure to 10(-7) M fMLP. The response to fMLP did not change with the duration of incubation and, as with neutrophils, cytoplasmic realkalinization was blocked by dimethylamiloride (20 microM). Incubation with 2-deoxyglucose (10 min, 5 mM), sufficient to inhibit by more than 90% the formyl peptide-stimulated superoxide generation by monocytes, slowed fMLP-induced acidification and abrogated the alkalinization. In addition, monocytes isolated from the blood of a patient with X-linked chronic granulomatous disease (CGD) underwent fMLP-induced acidification that was unmasked further by coincubation with dimethylamiloride, in a manner quantitatively similar to that of normal monocytes, despite the inability of the CGD cells to produce superoxide. The chemotactic factor-induced cytoplasmic pH responses of monocytes/macrophages remained constant as the cells matured in vitro and exhibited a dimethylamiloride-independent acidification and dependent alkalinization, as did the response in neutrophils. The cytoplasmic acidification of these cells thus did not correlate with the cells' production of superoxide and with the concomitant hexose monophosphate shunt activation, as has been suggested for other leukocyte types.

Calcium↗
PubMed1993

Evaluation of pulmonary systemic blood flow using ECG gated acquisition.

We propose a functional parametric analysis method using ECG-gated 99mTc-labeled red blood cell (RBC) imaging for detection and characterization of periodic variations in local blood activity in the lungs during cardiac cycle. We validated in animal experiments that such count variations correlate with cyclical pulmonary blood flow and may be used for evaluation of systemic-to-pulmonary shunts. Clinical studies were performed in 48 patients. After labeling the RBC pool with 99mTc, ECG-gated gamma camera images of both lung fields were acquired and processed to obtain Fourier transforms of time/activity functions in selected regions. The first harmonic parametric images of amplitude and phase were derived. There was an excellent correlation (r = 0.92) between activity variations and pulsatile flow measured by our method with that obtained by the thermodilution method in dog experiments (n = 10) after implantation of a systemic-to-pulmonary shunt. Patient studies showed the technique to be sensitive in detecting and quantifying abnormal systemic-to-pulmonary blood flow. Lung pulsatile flow can thus be noninvasively measured from functional parametric phase and amplitude images; the technique may be useful for detecting and quantifying abnormal systemic-to-pulmonary blood flow in man.

Animals↗
PubMed1992

Flow cytometric kinetic measurements of neutrophil phospholipase A activation.

The interrelationships between activation of phospholipases and neutrophil stimulus-induced Ca2+ responses remain unclear. We report here that immune complexes activate a phosphatidylcholine-specific phospholipase A in a neutrophil only after the cytoplasmic Ca2+ transient has been initiated in the same cell, while chemotactic peptide activation does not proceed via such a phospholipase A-mediated mechanism. Measurements of [Ca2+] changes and of phosphatidylcholine-specific phospholipase A activity were made by flow cytometry, using Indo-1 for Ca2+ indication, and a new fluorescent probe, bis-BODIPY-phosphatidylcholine, localized in the inner leaflet of the plasma membrane, to measure phospholipase A activation. Both 100 nM formyl-methionyl-leucyl-phenylalanine (with or without cytochalasin B) and 60 micrograms/ml insoluble immune complexes elicited cytoplasmic Ca2+ transients, but only insoluble immune complexes stimulated phospholipase A activation in a subpopulation of cells exhibiting an elevation of [Ca2+]in. Phospholipase A activation followed the Ca2+ transient, starting, in each cell, after [Ca2+]in had begun to decrease as Ca2+ redistributed in the activated cell. The products of this phospholipase activation were confirmed by thin layer chromatography. We conclude that neutrophils respond to immune complexes with an elevated cytoplasmic Ca(2+)-requiring phosphatidylcholine-specific phospholipase A activation and to chemotactic peptides by a different mechanism.

Boron Compounds↗
PubMed1991

Tuberculosis: a disease of the 1990s.

In the United States, the decades preceding the 1980s were characterized by a decline in the incidence of tuberculosis. More recently, the trend has undergone a significant reversal: Case rates have been increasing by 3% to 6% annually. In 1990, more than 25,700 cases were reported to the Centers for Disease Control. In a sense, tuberculosis is adapting to the '90s. The recent increase in its incidence tends to affect populations with identifiable characteristics. Among the most important of these groups are the populations at high risk for infection by the human immunodeficiency virus. The increase is also fueled by cases in populations that are medically underserved, including foreign-born persons from high-prevalence countries, persons with low incomes, and persons living in long-term-care facilities--especially persons with previous tuberculosis infection. Thus, factors such as homelessness, chronic alcohol or drug abuse, malnutrition, and crowded living conditions continue to favor development and transmission of disease. The increase in the incidence of tuberculosis appears to be greatest when subpopulations in such circumstances are also at high risk for HIV infection. Complex issues in the diagnosis and treatment of tuberculosis arise from these epidemiologic patterns. HIV infection is associated with unusual presentations of tuberculosis. Thus, the clinician must maintain a high index of suspicion for the disease in the setting of HIV infection or risk of the infection. The populations at greatest risk are likely to be mistrustful of the medical system, making the long-term administration of potentially toxic chemotherapy more difficult than it already is. Chronic substance abuse may complicate compliance and add further difficulties to the monitoring of chemotherapy. At the same time, the monitoring becomes even more important in the physician's effort to minimize adverse effects of the medications. Outbreaks of drug-resistant disease have recently occurred, complicating the selection of drugs and affecting the duration of treatment. Despite all of these problems, it is essential to establish a diagnosis and initiate treatment rapidly, both to arrest the disease process and to limit its transmission. Since Mycobacterium tuberculosis is spread to uninfected persons in aerosols generated by coughing or sneezing, the infectiousness of a patient with active disease can be related, at least in part, to the number of organisms seen on sputum smears. Initiation of therapy is followed by a rapid decline in infectivity.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Mechanisms of mastoparan-stimulated surfactant secretion from isolated pulmonary alveolar type 2 cells.

Mastoparan, a tetradecapeptide component of wasp venom, is a potent activator of secretion in a variety of cell types, and has been shown to activate purified G-proteins reconstituted into phospholipid vesicles with a preferential activation of Gi over Gs (Higashijima, T., Uzu, S., Nakajima, T., and Ross, E. R. (1988) J. Biol. Chem. 263, 6491-6494). To identify the biochemical activities of mastoparan in a cellular system, we characterized the effects of mastoparan on signal transduction pathways in rat pulmonary alveolar type 2 epithelial cells, which synthesize and secrete pulmonary surfactant. Mastoparan inhibited adenylylcyclase activity in a manner that was dose-dependent (IC50 = 30 microM), but sensitive to neither guanine nucleotide nor pertussis toxin (PT). Mastoparan induced a PT-sensitive increase in cellular inositol trisphosphate and a rapid rise in cytosolic calcium released from intracellular stores; the time to onset of the calcium rise, but neither the rate nor the amplitude of the rise, were PT-sensitive. Mastoparan also caused a dose- (EC50 = 16 microM) and time-dependent activation of arachidonic acid release that was completely insensitive to pretreatment with PT. Secretion of pulmonary surfactant was increased by mastoparan approximately 8-fold over constitutive levels at 1 h with an EC50 = 20 microM, and mastoparan-stimulated secretion was partially sensitive to PT at late time points and to inhibitors of arachidonic acid metabolism, but not to the protein kinase C inhibitor H7. These findings are consistent with the activation of Gi proteins in type 2 cells by mastoparan, although the lack of predicted triphosphoguanine nucleotide and PT sensitivity for some activities indicates that mastoparan does not act in a manner strictly analogous to liganded receptors or that some activities are not mediated by activation of Gi. While mastoparan is a potent secretagogue in several cell types, its secretory activity appears to have only a limited dependence on the activation of Gi proteins in type 2 cells.

Animals↗
PubMed1991

Effect of salt supplementation on amphotericin B nephrotoxicity.

It has been suggested that salt loading protects against amphotericin B-induced nephrotoxicity. The influence of saline loading on the nephrotoxic response to amphotericin B (50 mg/dose given i.v. over 4 hr 3 X/week for 10 weeks) was assessed in two groups of ten patients each who were diagnosed with mucocutaneous leishmaniasis. Patients were randomized to receive either 1 liter of 0.9% saline or 1 liter of 5% dextrose in water, administered i.v. over one hour in a double-blinded manner, directly prior to amphotericin B administration. Renal function was monitored on a weekly basis two days after the last dose of amphotericin B. Baseline characteristics were similar in both groups except for a slightly higher serum creatinine concentration (Cr) in the saline group (0.8 +/- 0.05 vs. 0.6 +/- 0.04 mg/dl). Baseline sodium (Na) excretion was relatively high (262 +/- 23 mmol/day in the dextrose group and 224 +/- 17 mmol/day in the saline group). None of the patients sustained an increase in Cr to values greater than 1.7 mg/dl. Although mean Cr remained within normal, there was a significant difference between the two groups over the ten week period, with the dextrose group sustaining a significant increase in Cr and the saline group remaining unchanged. Serum potassium (K) levels fell in both groups necessitating oral K supplementation. The saline group required significantly greater amounts of K supplementation to maintain a normal serum K. Amphotericin B caused a rapid reduction in the acidification ability of the kidney in response to an ammonium chloride load. Under these conditions, the saline group had a poorer ability to acidify the urine.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1990

Simultaneous flow cytometric measurements of cytoplasmic Ca++ and membrane potential changes upon FMLP exposure as HL-60 cells mature into granulocytes: using [Ca++]in as an indicator of granulocyte maturity.

Treatment of human leukemic HL-60 cells with N,N-dimethylformamide (DMF) induces them to mature until they reach granulocytoid morphology 3-6 d later. We have reported a maturation-dependent ability of these cells to respond to phorbol myristate acetate (PMA), as evaluated by membrane depolarization and by oxidative burst product formation (Newburger et al.: J. Biol. Chem. 259,3771, 1984). More recently we have attempted to develop techniques for simultaneous evaluation of these parameters during HL-60 cell maturation. Here, we compare the cytoplasmic [Ca++] and membrane potential changes elicited by the chemotactic peptide fMLP via simultaneous measurement of individual cells in a fluorescence-activated cell sorter (FACS), as done previously for mature granulocytes (Lazzari et al.: J. Biol. Chem. 261,9710, 1986). The stimulus-induced [Ca++]in changes are detected with the fluorescent probe Indo-1 and reproducibly increase in magnitude for a subpopulation of cells as the cells mature into granulocytes. Ca++ responsiveness to formyl peptide is restricted to a subpopulation of HL-60 granulocytes which expresses receptors for chemotactic peptide and consistently increases in magnitude (in response to the same concentration of agonist) with maturation. In contrast, there is less consistency in the direction or magnitude of membrane potential changes elicited under the same circumstances from the same maturing HL-60 cells.

Calcium↗
PubMed1990

Lymphocyte recruitment to the lung.

Lymphocyte recruitment in lymphoid tissues and inflammatory sites occurs in response to two events. The first is adherence of lymphocytes to specialized molecules expressed on the surface of appropriately stimulated vascular endothelial cells known as vascular addressins. The interaction occurs via specialized lymphocyte surface molecules known as homing receptors. There is considerable diversity among these molecules. At least three, and possibly four, different addressin-homing receptor pairs exist, regulating entry into peripheral lymph nodes, gut lymphoid tissue, BALT and intrathoracic lymphoid tissue, and inflamed synovium. Vascular addressins are expressed by specialized endothelial cells known as HEV. HEV are not found in normal lung parenchyma but may be induced to appear during an immune response. The mechanism for induction of HEV is unknown, although it may involve the action of inflammatory cytokines. It is not known whether separate endothelial cells exist with a propensity to develop into HEV or if any endothelial cells will develop into HEV if stimulated in the proper manner. Other accessory, lymphocyte-endothelium adhesion molecule pairs have been described, including LFA-1-ICAM-1 and CD4-HLA-DR. These molecules are induced by exposure of the endothelium to inflammatory cytokines, chiefly IFN-gamma. Thus, local humoral influences present during inflammation can alter the possibility of lymphocyte traffic through the endothelium by regulating the presence of lymphocyte adherence molecules. These processes have been documented to occur in the lung in normal homeostasis (e.g., BALT) and in disease (e.g., immunization with SRBC). After adherence, lymphocytes exit the circulation via amoeboid motility. This motility can be altered and enhanced through chemoattractant substances that act via surface receptors. The biochemical basis of cell motility is not entirely clear but appears to involve a link between the second messengers of receptor signaling and changes in the cytoskeleton, particularly actin filaments and microtubules. Like fibroblasts and smooth muscle cells, lymphocytes appear to respond to a number of "mitoattractants," substances that cause cell cycle entry and/or progression as well as enhanced motility. This relationship illustrates the integral relationship between cell motility and proliferation and suggests that the process of cell recruitment might also prime the recruitment cells to become activated to proliferate and perform effector function. Studies of lymphocyte-mediated lung disease confirm that antigen-specific as well as antigen-nonspecific lymphocytes are selectively recruited to the lung from the circulation during an inflammatory reaction in the lung.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1990

Tuberculosis infection in urban adolescents: results of a school-based testing program.

In 1986-87 a pilot tuberculosis (TB) skin testing program was introduced for seventh and tenth grade students in the Boston (Massachusetts) public schools. The 8.9 percent tuberculin positivity rate in tenth grade students was significantly higher than the 5.1 percent rate found in seventh graders. A majority of those who were skin test positive were born outside the United States. These results suggest that tuberculin testing in an urban school setting may identify a significant number of candidates for TB preventive therapy, particularly among tenth grade students and those who are foreign-born.

Adolescent↗
PubMed1989

Monoclonal antibody to SER immune suppressor detects polymeric forms of haptoglobin.

A series of monoclonal antibodies has been developed which is directed to a serum immunosuppressive factor, known as suppressive E-receptor factor (SER). SER, purified from the body fluids of cancer patients, is a polymeric form of haptoglobin, which is 100-1,000 times more potent an immunosuppressor than normal plasma haptoglobin and is immunochemically analogous to the neonatal form. Unlike the neonatal haptoglobin found in cord blood, SER, however, does not contain bound-hemoglobin. One group of monoclonal antibodies described in this study detects the polymeric forms of haptoglobin (SER) under non-denaturing conditions, but fails to recognize SER under the denaturing conditions of SDS-PAGE. A second group of monoclonal antibodies reacts only with the alpha subunit of haptoglobin but not with the beta subunit; in contrast, the commercially prepared polyclonal antisera to haptoglobin react with both the alpha and beta subunit. The average level of SER in normal human plasma (n = 19) was 1.0 micrograms/ml, regardless of age or sex. Since macrophages appear to secrete SER but do not synthesize haptoglobin, SER may represent an oxidized form of plasma haptoglobin generated from macrophages activated during an inflammatory response. These studies suggest that SER may be a negative feed-back regulator of immune response produced by activated macrophages.

Adult↗
PubMed1988

Time dependence of transmembrane potential changes and intracellular calcium flux in stimulated human monocytes.

An important characteristic of the functional differentiation of the blood monocyte is the development of its capacity to recognize and respond to stimuli. This ability is mediated to a large extent by specific receptor glycoproteins located on the cell surface. Stimulation of mononuclear phagocytes via these receptors results in a rapid rise in intracellular Ca++ concentration, accompanied or followed by a change in membrane potential, generation of oxidative products, degranulation, and effector functions such as phagocytosis, aggregation, or locomotion. While the development of these characteristics is difficult to characterize in vivo, several investigators have demonstrated in vitro changes in these cells that correlate with the development of effector function. To examine the mechanisms of specific membrane-stimulus interactions of monocytes as they differentiate into macrophage-like cells, we studied the responses of human monocytes and of monocytes incubated in serum-containing medium for up to 96 hr to the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP). Freshly isolated monocytes exhibited little change in transmembrane potential following stimulation with an optimal concentration of peptide and underwent a significant increase only after 48 hr in culture. While constant resting intracellular Ca++ concentrations were maintained during the culture period, intracellular Ca++ levels following fMLP stimulation increased with with incubation in serum, for up to 96 hr. In contrast, fMLP-induced respiratory burst activity increased from 0 to 24 hr in culture; it remained elevated at 48 hr but declined again by 96 hr. Incubation of the cells for 24 hr increased their random (unstimulated) motility in modified Boyden chambers but did not alter the cells' directed (chemotactic) response to fMLP in comparison to the response of freshly isolated monocytes. Peptide binding to the cells did not increase during the incubation period, indicating that an increase in receptor number or in affinity for fMLP was not responsible for the enhanced responsiveness to fMLP as incubation time increased. These studies indicate that incubation of monocytes in serum-containing medium leads to a complex, altered series of responses to fMLP that correlate with the differentiation of the original monocytes in vitro and may relate to the in vivo differentiation of monocytes to macrophages.

Calcium↗
PubMed1987

Lymphokine activation of T4+ T lymphocytes and monocytes.

The function of the T4 antigen, a marker for a differentiated T cell subset, is not well understood. Our previous observation that a chemoattractant human lymphokine, lymphocyte chemoattractant factor (LCF), which selectively induces motile responses in unactivated T4+ lymphocytes, led us to investigate whether LCF could also induce T4+ cell activation. Because LCF acts selectively on T4+ cells, we next determined whether the T4 antigen has a function in this LCF-induced cellular activation. A T4+ lymphocyte migratory response is induced by divalent anti-T4 antibody, but not by corresponding Fab fragments of the same antibody. Fab fragments of anti-T4 antibody, but not Fab fragments of anti-T3 antibody, block the migratory effect of both LCF and divalent anti-T4. Furthermore, LCF but not divalent anti-T4, evokes the expression of interleukin 2 (IL 2) receptors and HLA-DR antigen on T4+ lymphocytes in 24 hr. These effects are quantitatively similar to those observed by anti-T3 antibody activation. LCF-induced IL 2 receptor expression is blocked by co-incubation with anti-T4 antibody and anti-T4 Fab fragments, whereas anti-T3 activation is not inhibitable by anti-T4 Fab fragments. Because cultured monocytes express the T4 antigen, we investigated the action of LCF on cultured monocyte migration and HLA-DR expression. Induction of monocyte migration by LCF and anti-T4 antibody increases proportionally as T4 antigen expression increases in vitro. This enhanced migration is inhibitable by anti-T4 Fab fragments. Monocyte activation, as measured by augmented HLA-DR expression 24 hr after incubation with LCF, but not anti-T4 antibody, is quantitatively similar to the effects of interferon-gamma. Augmented HLA-DR expression is blocked by anti-T4 Fab fragments but not by antibody to interferon-gamma. These studies indicate that LCF interacts with T4+ lymphocytes and monocytes to induce migration and cellular activation.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1986

Production of lymphocyte chemokinetic activity by stimulated alveolar macrophages.

While the presence of a lymphocytic parenchymal infiltrate is characteristic of several lung diseases, the mechanisms responsible for the focal accumulation of lymphocytes within the lungs remain unclear. Since alveolar macrophages secrete several substances that affect lymphocyte function, we examined supernatants of stimulated, cultured guinea pig alveolar macrophages for their ability to alter lymphocyte motility. Guinea pigs were immunized by footpad injection of ovalbumin (OVA) emulsified in complete Freund's adjuvant. Fourteen days later, alveolar macrophages were obtained by bronchial lavage or teasing the lung parenchyma, enriched by adherence to plastic, and incubated for 3 and 24 hours in culture medium alone or medium containing either latex beads, OVA, or human serum albumin (HSA). Conditioned medium was harvested and assayed for chemoattractant activity against rat splenic lymphocytes in modified Boyden chambers. Regardless of stimulus, there was no evidence of enhanced lymphocyte motility above control values in supernatants harvested at 3 hours. At 24 hours, alveolar macrophages from OVA-sensitized guinea pigs stimulated with latex or OVA generated significant amounts of lymphocyte migration stimulating activity (LCA) (250 +/- 25 and 247 +/- 24 percent of control migration, respectively) compared to cells incubated in medium alone or with HSA (162 +/- 23 and 147 +/- 14 percent, respectively). Antigen recognition appears to be related to the presence of cytophilic anti-OVA antibody on the surfaces of alveolar macrophages of sensitized guinea pigs. LCA is resistant to neuraminidase, chymotrypsin, and heating to 56 degrees C, and was chemokinetic for T-lymphocytes. it elutes from Sephadex G-100 in two regions: one at approximately 67,000 d, and a second at approximately 15,000 d. These studies indicate that following systemic immunization, the guinea pig alveolar macrophage can react to specific antigen or phagocytosis of inert particulates by secreting a chemokinetic factor for T-lymphocytes, and may play a role in the pathogenesis of some types of antigen-induced lung disease.

Animals↗