Characterization of tumor binding by the IC-21 macrophage cell line.

The purpose of this study was to determine if the SV40-transformed murine macrophage cell line IC-21 is a suitable model to study the selective high avidity binding of tumor cells by subpopulations of activated macrophages. IC-21 macrophages bound P815, RBL5, and EL-4 murine tumor cells with high avidity, as measured by the inverted centrifugation method. Tumor binding by IC-21 macrophages was competitively inhibited by crude membrane vesicles prepared from tumor cells but not by cell membranes prepared from nontransformed splenic leukocytes, suggesting that this process was mediated by tumor-specific binding sites. IC-21 macrophages and primary cultures of pyran copolymer-elicited peritoneal macrophages demonstrated similar tumor binding avidity, kinetics, saturability, and metabolic requirements for optimal high avidity tumor binding. However, compared with primary cultures of pyran copolymer-elicited peritoneal macrophages, IC-21 macrophages bound 4-fold more tumor cells and were more homogeneous for tumor binding capability. Finally, one third of maximal tumor cell binding by IC-21 macrophages was completed within 5 min of contact with tumor, suggesting that IC-21 macrophages constitutively expressed some high avidity tumor binding sites. Their stable and homogeneous capability for binding tumor cells and their ease of growth make the IC-21 macrophage cell line a potentially valuable model for elucidating the molecular mechanisms responsible for selective high avidity tumor binding by subpopulations of activated macrophages.

Oleic acid stimulates rapid translocation of cholinephosphate cytidylyltransferase in type II cells.

Activity of cholinephosphate cytidylyltransferase, the rate-limiting enzyme in phosphatidylcholine synthesis, increases rapidly in the lung after birth predominantly due to an increase in membrane-associated activity. Although there is strong evidence that enzyme translocation is a major regulatory mechanism in other cells, this mechanism has not been conclusively demonstrated in intact alveolar type II cells. In this study, we show that oleic acid stimulates rapid translocation of cytidylyltransferase activity and protein from cytosol to microsomes in both primary cultured fetal and adult type II cells and MLE12 cells, a cell line derived from murine distal respiratory epithelial cells. Shifts in subcellular distribution occurred within 5 min of exposure to 200 microM oleic acid. The magnitude of the increases in microsomal enzyme activity and immunoreactive protein levels was several-fold greater in d21 fetal cells than adult type II cells. Oleic acid-induced translocation was confirmed in in vitro translocation experiments. After incubating MLE12 cell postmitochondrial supernatants at 37 degrees C with oleic acid and separation of enzyme isoforms on glycerol density gradients, enzyme activity was decreased in gradient fractions corresponding to both cytosolic isoforms and microsomal activity increased 7.9-fold compared to the distribution of enzyme activity in postmitochondrial supernatants incubated at 4 degrees C without oleic acid. The increase in microsomal activity was associated with an increased incorporation of [14C]oleic acid in the membrane free fatty acid fraction. Developmental changes in type II cell membrane lipid composition may induce the rapid translocation/activation of cytidylyltransferase in the lung after birth.

A double-blind clinical trial in weight control. Use of fenfluramine and phentermine alone and in combination.

We performed a double-blind, controlled clinical trial comparing phentermine resin (30 mg in the morning), fenfluramine hydrochloride (20 mg three times a day), and a combination of phentermine resin (15 mg in the morning) and fenfluramine hydrochloride (30 mg before the evening meal), and placebo. We combined low doses of the two drugs to maintain efficacy while diminishing adverse effects. Eighty-one people with simple obesity (130% to 180% of ideal body weight) participated. Individualized diets were prescribed and discussed again during the 24-week study period. Weight loss in those receiving the combination (8.4 +/- 1.1 kg; mean +/- SEM) was significantly greater than in those receiving placebo (4.4 +/- 0.9 kg; Scheff é's test) and equivalent to that of those receiving fenfluramine (7.5 +/- 1.2 kg) or phentermine (10.0 +/- 1.2 kg) alone. Adverse effects were less frequent with the combination regimen than with other active treatments. Thirty-seven participants dropped out of the study, 18 for reasons related to drug treatment. Combining fenfluramine and phentermine capitalized on their pharmacodynamic differences, resulting in equivalent weight loss, fewer adverse effects, and better appetite control.

Determinants of response to anorexiants.

We analyzed data from an 81-patient clinical trial of anorexiant medication, searching for the predictors of response. In the trial we assigned treatments to participants by minimization, a process that decreases differences between treatment groups. Based on the literature, the investigators' experience, and the pharmacologic properties of the medications, we selected 25 factors for use in the minimization process. Retrospective examination by contingency-table analysis indicated that the most important predictors of weight loss in this study were weight loss during the 3-wk diet-only run-in period (chi 2, p less than 0.001), physician estimation of patients motivation (chi 2, P less than 0.003), participant eating habits (nighttime "binge" eaters responded best; chi 2, P less than 0.003), adherence to treatment (chi 2, P less than 0.01), and type of treatment (chi 2, P = 0.05). When multiple regression analysis was applied, several other factors aided in explaining the variance in the weight loss results. For example, duration of obesity was inversely related to weight loss and weight loss during the study increased with family income. Depending on the treatment used, anxiety and depression visual analogue scores also explained some of the variance. Failure to account for factors such as these in treatment assignment may account for the low discriminant power of clinical trials of anorexiants.

Tiopinac in rheumatoid arthritis: a three-phase dose-ranging, efficacy, and aspirin-withdrawal protocol.

We designed a clinical trial to obtain dose-ranging, efficacy, and aspirin-withdrawal data on tiopinac in patients with rheumatoid arthritis. To accomplish this without exposing the patients to risk of disease exacerbation and avoiding type II error, we used a 3-phase protocol adding tiopinac to current therapy. The 3 phases-open-label dose ranging, double-blind tiopinac versus placebo, and aspirin withdrawal-began after a single-blind run-in period. The manufacturer withdrew tiopinac from investigation because of toxicity at higher doses, but with only 13 patients we found that tiopinac (up to 300 mg/day) decreased walking time, painful joints, and morning stiffness and increased grip strength (p less than 0.05). Both the global evaluation by the investigators and patient ratings of their activity showed superiority of tiopinac (tiopinac: 5 better, 1 worse; placebo: 1 better, 6 worse; p = 0.028 by Fisher's exact test). Complete aspirin withdrawal could be accomplished in only 3 patients, although in 10 of 13 the dose could be reduced 50% of baseline or less. The 3-phase protocol indicated effectiveness, a dose range, and partial aspirin replacement with minimal patient risk.

Platelet alpha-adrenergic receptors in obesity: alteration with weight loss.

We studied platelet alpha-adrenergic receptor concentration and function in 19 subjects with simple obesity participating in a double-blind, controlled clinical trial of diet and anorexiants (phentermine, fenfluramine, or a combination of the two) or placebo. From wk 1 to wk 8, weight loss for the group as a whole was 4.9 +/- 0.7 kg (mean +/- SE). Concomitant with this weight loss, the platelet alpha-adrenergic receptor concentration rose from 85.7 +/- 5.8 to 113 +/- 5.8 fmol/mg protein. This increase moved the values for the obese subjects toward or beyond values in lean controls (100 +/- 10.5 fmol/mg protein). The response in the different treatment groups was similar. The receptor concentration increase was accompanied by a corresponding increase in alpha-adrenergic receptor-mediated platelet aggregation. For individual subjects the extent of weight loss over time generally correlated with percent receptor change. Altered adrenergic sensitivity occurring in obese subjects who are losing weight may have important implications in relation to external (therapeutic or inadvertent) administration of catecholamines.

The role of fever in the infected host.

Sepsis is a highly lethal clinical syndrome characterized by a systemic inflammatory response to infection. Fever, a non-specific acute-phase response, has been associated with improved survival and shortened disease duration in non-life-threatening infections. However, the influence of fever and the effects of antipyresis in patients with sepsis has not been prospectively studied in humans. This paper reviews the state of our knowledge concerning the biological effects of fever in infected hosts and the influence of fever and antipyretic therapy on survival during sepsis in experimental models and in man.

IFN-gamma inhibits lipopolysaccharide-induced interleukin-1 beta in primary murine macrophages via a Stat1-dependent pathway.

Interleukin-1 (IL-1) plays an important role in host defenses against microbial pathogens. Excessive production of this cytokine, however, may be responsible in part for the lethality observed during sepsis. Our studies show that interferon-gamma (IFN-gamma) downregulates lipopolysaccharide (LPS)-induced interleukin-1beta (IL-1beta) transcription in primary macrophages. This phenomenon does not occur in splenocytes or bone marrow-derived macrophages from signal transducer and activator of transcription (Stat1)-deficient mice, suggesting that Stat1, a transcription factor involved in IFN signaling, plays a critical role in this process. Moreover, nitric oxide (NO) was also involved in the downregulation of LPS-induced IL-1 by IFN, as addition of the inducible nitric oxide synthase (iNOS) inhibitor L-N(6)-(1-iminoethyl)lysine (NIL) negated the effect. Kinetic analysis of IL-1 and IFN levels in LPS-treated mice in vivo suggests that IFN-mediated inhibition of IL-1 might be an important negative feedback mechanism for limiting IL-1 generation in vivo.

Effects of hypothermia and hyperthermia on cytokine production by cultured human mononuclear phagocytes from adults and newborns.

We have shown previously that febrile range temperatures modify cytokine production by adult macrophages. In this study, we compared the effects of moderate hyperthermia and hypothermia on the kinetics of lipopolysaccharide (LPS)-induced cytokine expression in monocytes and macrophages of newborns and adults. During culture at 40 degrees C, the initial rates of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) secretion were preserved, but the duration of secretion was shorter than the duration at 37 degrees C. TNF-alpha and IL1-beta concentrations in 24-h 40 degrees C culture supernatants were reduced 18%-50%. IL-6 concentration in 24-h 40 degrees C cultures was reduced 26%-29% in all cells except adult macrophages. At 32 degrees C, changes in early (2 h) and sustained (24 h) cytokine expression were reversed compared with those caused by hyperthermia. Culturing adult macrophages at 32 degrees C blunted early secretion of TNF-alpha and IL-6 by 69% and 65%, respectively, and increased TNF-alpha concentration at 24 h by 48% compared with levels at 37 degrees C. In adult monocytes cultured at 32 degrees C, early IL-6 and IL-1 beta secretion was decreased 64% and 51%, respectively. We speculate that the burst/suppression cytokine profile at febrile temperatures might enhance early activation of host defenses and prevent prolonged exposure to potentially cytotoxic cytokines. Hypothermia, on the other hand, may worsen outcome in infections by delaying and prolonging cytokine production.

Potential role of interleukin-1 in the development of bronchopulmonary dysplasia.

Increased activities of inflammatory mediators unopposed by their inhibitors contribute to chronic lung injury and impaired healing in BPD. The deleterious effects of IL-1 beta, a cytokine involved in inflammation and host defense, are blocked by IL-1 receptor antagonist (IL-1Ra). We proposed that an imbalance of IL-1 beta and its inhibitors may contribute to the development of BPD. To determine the relative antigen concentrations of IL-1 beta and IL-1Ra and functional IL-1 activity in lung lavage of infants at risk for BPD, lung lavage was serially obtained from 1 to 28 days from 17 infants with evolving BPD, 13 infants with self-limited RDS, and 6 controls ventilated for nonpulmonary reasons. Overall, there was a high correlation between IL-1 beta antigen concentration and IL-1 activity (r = 0.82, p = 0.0001). There were no significant differences among the groups for lung lavage variables on day 1. However, in infants who developed BPD, IL-1 beta antigen concentration and IL-1 activity increased 16- and 61-fold, respectively, during the first week. IL-1Ra remained relatively unchanged during the first month. IL-1 beta/IL-1Ra antigen ratio was significantly higher on days 5 (median 0.024) and 7 (median 0.025) compared with day 1 (median 0.004), p < 0.05. These results suggest that a relative imbalance of IL-1 beta and IL-1Ra may contribute to prolonged inflammation in BPD.

Effects of assay medium composition on macrophage-mediated tumor cell binding and lysis.

Activated macrophages express selective, high avidity tumor binding and cytotoxicity for tumor targets. The reported macrophage-mediated tumor binding and killing activities vary considerably among studies. Whereas, some of these studies utilized identical tumor target cells and similar sources of macrophages, the composition of the assay media used by different groups varied with respect to: type of medium (Eagle's minimal essential medium (EMEM), RPMI-1640, Dulbecco's modified Eagle's medium with 1.0 g/l glucose (DMEMLG), or with 4.5 g/l, glucose (DMEMHG)), concentration of serum (0-20%), and the addition of certain reagents (amphotericin B, pyruvate, and Hepes). The purpose of this study was to evaluate the effects of varying these parameters of medium composition on macrophage-mediated high (HA) and low avidity tumor binding and cytolysis. Tumor cytolysis was measured with an 18 h 51Cr release assay using peritoneal macrophages from C3H/HeN mice primed in vivo with pyran copolymer and further stimulated in vitro with lipopolysaccharide (10 ng/ml). P815 tumor cells were used as targets at a 10:1 effector:target ratio. Binding of targets to macrophages was determined by a method utilizing inverted centrifugation. Under optimal conditions, 2 x 10(5) macrophages bound as many as 17,600 +/- 3565 tumor cells and caused up to 50.4 +/- 3.6% cytolysis. Assays performed in DMEMHG compared with the other three media tested resulted in HA tumor binding and cytolysis which were decreased by up to 42.5% (P less than 0.05) and 64.3% (P less than 0.01), respectively, compared with the three other types of medium. The addition of pyruvate (1 mM) to EMEM with 5% fetal calf serum (FCS) stimulated 98.8% (P less than 0.01) and 50.6% (P less than 0.1) increases in tumor binding and cytolysis, respectively compared with EMEM/5% FCS alone, while Hepes (25 mM) stimulated 58.3% (P less than 0.01) and 37.5% (P less than 0.1) increases in these activities. Amphotericin B (2.5 micrograms/ml) completely abrogated tumor cytolysis, but it caused no change in tumor binding. Serum produced variable effects on macrophage-mediated tumor killing. Five of six lots of FCS inhibited tumor lysis, by 16 to 98% (32.6 +/- 28.6%; mean +/- SD). However, the same lot of FCS which inhibited cytolysis by 98% enhanced HA binding by 152% (P less than 0.05). Finally, several commercially available serumless medium preparations supported macrophage-mediated tumor binding; however, none of the serumless media tested supported macrophage-mediated tumor cytolysis. We conclude that common differences in assay medium composition can markedly alter macrophage-mediated tumor cell binding and cytolysis.

Renal infarction due to renal artery dysplasia with dissection. Report of a case in a normotensive patient.

A previously healthy 44-year-old man with well-documented normotension had a sudden onset of left flank pain and delayed onset of constitutional symptoms, hematuria, and elevations of lactic dehydrogenase, serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, and creatinine levels. Angiography revealed unilateral renal artery fibromuscular dysplasia with dissection and infarction. In the year since, he has remained well and normotensive without therapy. The literature is reviewed.

Fever and the heat shock response: distinct, partially overlapping processes.

The heat shock response is an ancient and highly conserved process that is essential for surviving environmental stresses, including extremes of temperature. Fever is a more recently evolved response, during which organisms temporarily subject themselves to thermal stress in the face of infections. We review studies showing that fever is beneficial in the infected host. We show that core temperatures achieved during fever can activate the heat shock response and discuss some of the biochemical consequences of such an effect. We present data suggesting 4 possible mechanisms by which fever might confer protection: (1) directly killing or inhibiting growth of pathogens; (2) inducing cytoprotective heat shock proteins (Hsps) in host cells; (3) inducing expression of pathogen Hsps, an activator of host defenses; and (4) modifying and orchestrating host defenses. Two of these mechanisms directly involve the heat shock response. We describe how heat shock factor-1, the predominant heat-induced transcriptional enhancer not only activates transcription of Hsps but also regulates expression of pivotal cytokines and early response genes. The relationship between fever and the heat shock response is an illuminating example of how a more recently evolved response might exploit preexisting biochemical pathways for a new function.

Bacterial endotoxin is an active component of cigarette smoke.

BACKGROUND: Chronic bronchitis in cigarette smokers shares many clinical and histologic features with environmental lung diseases attributed to bacterial endotoxin (lipopolysaccharide [LPS]) inhalation. Experimental LPS inhalation mimics many of the acute effects of cigarette smoke in the lower airway. Therefore, we reasoned that LPS may be a biologically active component of cigarette smoke. DESIGN: The Limulus amebocyte lysate (LAL) assay was used to measure LPS in the tobacco and filter tip components of unsmoked 1R4F experimental cigarettes and commercially available "light" cigarettes, as well as in mainstream (MS) and sidestream (SS) smoke particles generated with an automated smoking machine and collected on ventilator mainflow filters. SETTING AND PARTICIPANTS: Blood LPS activity and plasma cytokine concentrations were measured in groups of healthy smokers and nonsmokers who reported to the walk-in clinic at the Baltimore VA Medical Center for unrelated complaints. MEASUREMENTS: Blood LPS levels were measured by LAL assay and plasma levels of tumor necrosis factor-alpha (TNF-alpha), interleukin 6 (IL-6), soluble TNF receptors I and II (sTNFR I and sTNFR II) were measured by enzyme-linked immunosorbent assay. RESULTS: Bioactive LPS was detected in both the tobacco portion (1R4F, 17.8+/-1.0 microg/cigarette; light, 26.8+/-7.3 microg/cigarette [mean+/-SE]) and filter tips (1R4F, 0.67+/-0.55 microg/cigarette; light, 0.70+/-0.39 microg/cigarette) of cigarettes. Bioactive LPS was also detected in both MS (1R4F, 120+/-64 ng/cigarette; light: 45.3+/-16 ng/cigarette) and SS smoke (1R4F, 18+/-1.5 ng/cigarette; light: 75+/-49 ng/cigarette). Although systemic absorption of inhaled LPS may occur, we failed to detect any differences between nonsmokers and smokers in median blood LPS levels (median values, 66.75 and 72.1 pg/mL, respectively; p = 0.55) or plasma concentrations of TNF-alpha (0 vs 0 pg/mL, respectively; p = 0.71), sTNFR I(1,469 vs 1,576 pg/mL, respectively), sTNFR II (2,011 vs 3,110 pg/mL, respectively), or IL-6 (8.8 vs 0 pg/mL, respectively; p = 0.20). CONCLUSIONS: Smoking one pack of cigarettes per day delivers a dose of respirable LPS that is comparable to the levels of LPS associated with adverse health effects in cotton textile workers. Thus, we suggest that the bioactive LPS in cigarette smoke may contribute to the pathogenesis of chronic bronchitis that develops in susceptible cigarette smokers.

Elevated tissue levels of interleukin-1 beta and tumor necrosis factor-alpha in vulvar vestibulitis.

OBJECTIVE: To compare levels of two inflammatory cytokines, interleukin-1 beta(IL-1 beta) and tumor necrosis factor-alpha(TNF-alpha), in selected regions of the vulva, vestibule, and vagina in women with vulvar vestibulitis and in asymptomatic controls. METHODS: Selective samplings of surgical specimens from 12 women undergoing perineoplasty for vulvar vestibulitis and ten pain-free subjects undergoing posterior vaginal repair were prepared into tissue homogenates and analyzed for concentrations of IL-1 beta and TNF-alpha. Interleukin-1 beta and TNF-alpha concentrations were measured by sandwich enzyme-linked immunosorbent assay. The results were reported after adjustment for total tissue protein concentration. RESULTS: Median tissue levels of IL-1 beta and TNF-alpha were elevated 2.3-fold and 1.8-fold, respectively, in women with vulvar vestibulitis relative to pain-free women. Median IL-1 beta tissue levels were increased significantly from 1.3 pg/mg to 3.0 pg/mg total protein in women with vulvar vestibulitis compared to pain-free women. Median TNF-alpha tissue levels were increased from 83 pg/mg to 148 pg/mg total protein in women with vulvar vestibulitis compared to pain-free women. Analysis by selected anatomic site of women with vulvar vestibulitis revealed a significant 2.2-fold higher median level of TNF alpha at the vulvar site compared to the vestibule. CONCLUSION: Concentrations of IL-1 beta and TNF-alpha were elevated in women with vulvar vestibulitis relative to those in asymptomatic controls. This elevation in inflammatory cytokines with vulvar vestibulitis varied according to anatomic site and was, paradoxically, lowest in the area of highest hyperalgesia, the vulvar vestibule. Inflammatory cytokine elevation may contribute to the pathophysiology of mucocutaneous hyperalgesia.

Exposure to febrile temperature modifies endothelial cell response to tumor necrosis factor-alpha.

Fever is an important regulator of inflammation that modifies expression and bioactivity of cytokines, including tumor necrosis factor (TNF)-alpha. Pulmonary vascular endothelium is an important target of TNF-alpha during the systemic inflammatory response. In this study, we analyzed the effect of a febrile range temperature (39.5 degrees C) on TNF-alpha-stimulated changes in endothelial barrier function, capacity for neutrophil binding and transendothelial migration (TEM), and cytokine secretion in human pulmonary artery endothelial cells (EC). Permeability for [(14)C]BSA tracer was increased by treatment with TNF-alpha, and this effect was augmented by incubating EC at 39.5 degrees C. Treating EC with 2. 5 U/ml TNF-alpha stimulated an increase in subsequent neutrophil adherence and TEM. Incubating EC at 39.5 degrees C caused a 30% increase in TEM but did not modify the enhancement of neutrophil adherence or TEM by TNF-alpha treatment. Analysis of cytokine expression in EC cultures exposed to TNF-alpha at either 37 degrees or 39.5 degrees C revealed three patterns of temperature and TNF-alpha responsiveness. Granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin (IL)-8 were not detectable in untreated EC but were increased after TNF-alpha exposure, and this increase was enhanced at 39.5 degrees C. IL-6 expression was also increased with TNF-alpha exposure, but IL-6 expression was lower in 39.5 degrees C EC cultures. Transforming growth factor-beta(1) was constitutively expressed, and its expression was not influenced either by TNF-alpha or exposure to 39.5 degrees C. These data demonstrate that clinically relevant shifts in body temperature might cause important changes in the effects of proinflammatory cytokines on the endothelium.

Concurrent expression of procoagulant and plasminogen activator activities by rabbit alveolar macrophages in vitro: opposite modulating effects of prostaglandin E2.

We examined the effects of arachidonic acid metabolites on the simultaneous expression of procoagulant (PC) and plasminogen activator (PA) activities by rabbit alveolar macrophages. Incubation with lymphocyte-conditioned medium (LCM) caused a significant increase in cell-associated PC activity. Co-treatment with indomethacin (1 microM) reduced this augmentation in PC activity by 33% (p less than 0.05). In contrast, indomethacin caused a 42% increase in PA activity released into incubation medium (p less than .05). Both effects of indomethacin were reversed by the addition of PGE2 in concentrations as low as 1 nM. Addition of 100 nM PGE2 to these cells caused an increase in PC activity 2.7-fold greater than that achieved by LCM alone, while PGE2 suppressed released PA activity by 62%. PGE2 and indomethacin had similar but less pronounced effects on phorbol myristate acetate-treated cells. These effects of PGE2 could be duplicated by PGE1, but not by any other arachidonic acid metabolite (PGF2 alpha, PGI2, PGD2, ddPGF2 alpha, LTB4, or LTC4). While PGE2 increases intracellular levels of cAMP, the observed effects on PC and PA activities could not be reproduced fully by treatment with dibutyryl cAMP. We conclude that PGE2 amplifies the augmentation of PC activity by stimulated alveolar macrophages while concurrently inhibiting expression of plasminogen activator. This suggests that PGE2 may be a significant mediator in regulating the highly interactive processes of inflammation and coagulation/fibrinolysis.

Dipyridamole stimulates urokinase production and suppresses procoagulant activity of rabbit alveolar macrophages: a possible mechanism of antithrombotic action.

Dipyridamole, an inhibitor of platelet aggregation, has been shown to have beneficial effects in disorders characterized by extravascular fibrin deposition. Mononuclear phagocytes are present in extravascular sites and are capable of expressing both plasminogen activator and procoagulant activities, which suggests these cells play a central role in extravascular fibrin turnover. We therefore sought to determine whether dipyridamole affects the expression of plasminogen activator and procoagulant activities by rabbit alveolar macrophages cultured in vitro. We found that dipyridamole (10 to 100 mumol/L) caused increases in both cell-associated and released plasminogen activator activity, which reached levels of 240% (P less than .05) and 543% (P less than .01) of controls, respectively. In contrast, dipyridamole decreased the cell-associated procoagulant activity of alveolar macrophages to as little as 21.3% of controls (P less than .01). Similar effects were seen in cells cotreated with lymphokines. The procoagulant activity expressed by these cells functioned as a tissue thromboplastin. The plasminogen activator of control and treated cells was a urokinase as determined by molecular weight characteristics (50 kilodaltons) and by antibody neutralization profiles using polyclonal antibodies against human urokinase and tissue plasminogen activator. These effects of dipyridamole could not be duplicated by structurally dissimilar agents sharing some of the pharmacological actions of dipyridamole; however, two pyrimidopyrimidine compounds structurally similar to dipyridamole effectively mimicked the effects on both procoagulant and plasminogen activator activities. We conclude that dipyridamole may have antithrombotic effects by directly modulating the role of mononuclear phagocytes in fibrin turnover. Thus, dipyridamole may be useful in situations where extravascular fibrin deposition is important to the pathogenesis of tissue injury and repair.

Benzodiazepine prescribing in a family medicine center.

We monitored benzodiazepine prescribing in a family medicine center for two years. A total of 1,886 prescriptions were written for four benzodiazepines in the following order of frequency: diazepam, chlordiazepoxide, oxazepam, clorazepate. The most frequent diagnostic indications were anxiety neurosis, hysterical neurosis, and vertebral column disorders. Most benzodiazepine recipients were women, but for alcohol abuse more prescriptions were written for men. More than half of the patients were between the ages of 25 and 44 years. Although daily doses were reduced for elderly patients, the course of therapy was often longer. Female physicians wrote fewer prescriptions for men than for women. More prescriptions for benzodiazepines were written during the summer months than during the rest of the year, and more were written on weekdays than on the weekend. Benzodiazepine recipients were given more prescriptions for other drugs than were the rest of the patients in the practice.