Biomedical subjects

R J MacKay

Publications and source records attributed to R J MacKay.

At least 37 records · Page 2Linked to original sources

PubMed1996

Effect of vaccination of ponies with A4 anti-idiotypic antibody on serum idiotype (1C9) and antilipid A concentration.

OBJECTIVE: To evaluate the humoral response of horses to vaccination, using a murine monoclonal anti-idiotypic antibody (A4) that shares an epitope with lipid A. DESIGN: Serum concentrations of antilipid A antibody and 1C9 (epitope on murine monoclonal antilipid A antibody) were measured serially during the period of vaccination with A4. ANIMALS: 6 clinically normal adult ponies. PROCEDURE: Ponies were inoculated IM 3 times at monthly intervals with A4. Two weeks after each inoculation, serum was obtained and was assayed by ELISA for antilipid A and 1C9 concentrations. Additional vaccinations were given to 2 of the ponies after a several-month rest period. RESULTS: There was significant increase in 1C9 concentration (P < 0.0001) during the period of vaccination and a trend toward increased antilipid A concentration. The latter effect was not significant (P = 0.055). Additional vaccinations produced further increase in serum 1C9 concentration; antilipid A concentration increased in one of these ponies but not the other. Maximal antilipid A concentration recorded in these ponies was approximately 6 times preimmune concentration and was comparable to that found in a commercial antiendotoxin antiserum. 1C9 also was detected in the commercial antiserum. CONCLUSIONS: A4 anti-idiotype vaccination of horses is safe and may be effective in eliciting an antibody response against endotoxin. The finding of 1C9 in a commercial antiendotoxin antiserum indicates that this idiotype may be part of the normal polyclonal antibody response of horses to endotoxin. CLINICAL RELEVANCE: It may be possible to use anti-idiotypic antibody vaccination in horses to induce protection against the effects of endotoxin.

Animals↗

PubMed1995

Septic cholangiohepatitis and cholangiocarcinoma in a horse.

Septic cholangiohepatitis was diagnosed in an 11-year-old Warmblood gelding with a history of intermittent colic and fever. Klebsiella pneumoniae, susceptible to gentamicin, was cultured from the biopsy specimen. However, treatment with gentamicin was unsuccessful, and histologic examination and bacteriologic culture of a biopsy specimen obtained 3 weeks later revealed progression of the hepatic inflammation and yielded growth of gentamicin-resistant K pneumoniae. At this time, several discrete hyperechoic structures, suggestive of biliary calculi, were seen ultrasonographically. A change in antibiotic treatment was associated with gradual resolution of clinical signs. Five months after initial examination, the horse had a sudden onset of severe right forelimb lameness. The horse responded to treatment with antibiotics and phenylbutazone, but lameness and fever that was unresponsive to treatment recurred 7 months later, and the horse was euthanatized. Necropsy revealed nodules throughout the liver and a mass associated with the right metacarpophalangeal joint. Histologic and immunohistochemical examination revealed carcinomatous infiltration of the liver and metacarpophalangeal joint. The tumor was probably of biliary origin. Carcinoma should be considered in cases of septic cholangiohepatitis unresponsive to antibiotic treatment.

Animals↗

PubMed1995

Sarcocystis falcatula from passerine and psittacine birds: synonymy with Sarcocystis neurona, agent of equine protozoal myeloencephalitis.

Equine protozoal myeloencephalitis (EPM) is a neurologic disease of horses caused by Sarcocystis neurona. The horse is a dead-end host for S. neurona and the definitive and intermediate hosts have not previously been identified. We hypothesized that S. neurona is actually Sarcocystis falcatula, a parasite that cycles in nature between Virginia opossums (Didelphis virginiana) and any of a variety of avian intermediate hosts. We extracted DNA from S. falcatula sarcocysts in the muscle of a brown-headed cowbird (Molothrus ater) and from schizonts in a fixed specimen of lung from a Moluccan cockatoo (Cacatua moluccensis). Three segments of the small subunit ribosomal RNA (SSURNA) gene, containing a total of 742 nucleotides, were amplified by the polymerase chain reaction, sequenced, and compared with the SSURNA sequence from two isolates of S. neurona. The S. falcatula sequence was identical to the sequence of the S. neurona isolate UCD-1 and differed in only 3 positions from isolate SN5. Recent evidence, also based on SSURNA sequences, implicates the opossum as the definitive host of S. neurona. Based on the SSURNA gene sequences S. falcatula and S. neurona are synonymous, thus the parasite cycles between opossums and birds maintaining a reservoir of the organism from which horses can be infected.

Animals↗

PubMed1995

Effects of tumor necrosis factor blockade on interleukin 6, lactate, thromboxane, and prostacyclin responses in miniature horses given endotoxin.

A monoclonal antibody (MAB) against equine tumor necrosis factor-alpha (Eq TNF) was used to investigate the role of TNF in cytokine, eicosanoid, and metabolic responses of Miniature Horses given endotoxin. Plasma concentrations of interleukin 6 (IL-6), lactate, thromboxane A2 metabolite, and prostacyclin metabolite (6-keto-PGF1 alpha) were measured in 10 Miniature Horses given 0.25 microgram of lipopolysaccharide (LPS; Escherichia coli O55:B5)/kg of body weight. Five horses were given Eq TNF MAB and 5 were given isotype-matched MAB as control. All horses were given 1.86 mg of antibody/kg by IV infusion, 5 minutes before LPS was given IV. Blood samples were taken 20 minutes before and at multiple intervals for 24 hours after LPS was given. Interleukin 6 bioactivity in plasma was measured, using IL-6-dependent cell line (B9). Eicosanoid activities were assessed by enzyme immunoassay, and plasma lactate concentration was determined enzymatically. Data were analyzed by ANOVA and Tukey's honest significant difference test for significant (P < 0.05) effect of treatment. Horses given Eq TNF MAB had significantly (P < 0.050) lower peak mean +/- SEM IL-6 (59 +/- 29 U/ml), lactate (16 +/- 2.00 mg/dl), and 6-keto-PGF1 alpha (254 +/- 79 pg/ml) values then did horses given control MAB (880 +/- 375 U/ml for IL-6; 26 +/- 0.04 mg/dl for lactate; and 985 +/- 290 pg/ml for 6-keto-PGF1 alpha). There was no effect of anti-TNF treatment on LPS-induced thromboxane A2 metabolite production. Tumor necrosis factor mediated IL-6, lactate, and prostacyclin responses, without affecting thromboxane production in horses given LPS.

6-Ketoprostaglandin F1 alpha↗

PubMed1995

Effect of treatment with a monoclonal antibody against equine tumor necrosis factor (TNF) on clinical, hematologic, and circulating TNF responses of miniature horses given endotoxin.

Tumor necrosis factor-alpha (TNF) is an important mediator of endotoxin-induced pathologic changes. To help define the role of TNF in equids with endotoxemia, the effects of pretreatment with a murine monoclonal antibody (MAB) against equine TNF were evaluated in Miniature Horses given endotoxin. Five horses were given TNF MAB at a dosage of 1.86 mg/kg of body weight, IV, and 5 were given control MAB. Five minutes later, lipopolysaccharide (LPS; Escherichia coli O55:B5), 0.25 microgram/kg, was given to all horses by bolus IV infusion. Clinical signs of disease were monitored at intervals up to 24 hours after LPS infusion, and blood was taken for determination of WBC count, PCV, plasma total protein concentration, plasma TNF activity, and serum MAB concentration. Reduction of plasma TNF activity in anti-TNF-treated horses was highly significant (P < 0.001), compared with that in control horses. Horses given TNF MAB had significantly improved clinical abnormality score (P < 0.010), lower heart rate (P < 0.001), and higher WBC count (P < 0.001), compared with horses given control MAB. Rectal temperature, respiratory rate, PCV, and plasma total protein concentration were not significantly different between groups. Serum MAB concentration peaked at 68 micrograms/ml 30 minutes after the end of antibody infusion in both groups. Neutralization of LPS-induced TNF activity reduced the hematologic and clinical responses of horses given LPS IV.

Analysis of Variance↗

PubMed1995

Effect of tumor necrosis factor antibody on synovial fluid cytokine activities in equine antebrachiocarpal joints injected with endotoxin.

Six horses received intra-articular injections of a mixture of 1 micrograms of endotoxin/5 mg of equine tumor necrosis factor (eqTNF) monoclonal antibody in 1 antebrachiocarpal joint and an equal volume (2 ml) of 1 micrograms of endotoxin/5 mg of control antibody in the opposite joint. Synovial fluid sample collection (1 ml) was accomplished by use of an indwelling, intra-articular catheter at postinjection hours (PIH) 0, 1, 1.5, 2, 5, and 8, and by arthrocentesis at PIH 24. Joint fluid samples were analyzed for nucleated cell count, protein concentration, and TNF, interleukin 6 (IL-6), IL-1, and IL-1-inhibitory activities. To monitor local inflammation, each carpus was graded semiquantitatively for swelling prior to each sample collection. Tumor necrosis factor, IL-1, or IL-1-inhibitory activity was not detected in any synovial fluid sample collected before endotoxin/antibody was administered. However, low IL-6 activity (< 100 U/ml) was found in 2 of 12 preinjection samples. In joints injected with endotoxin/control antibody mixture, maximal mean +/- SEM activities for TNF (1,019 +/- 310 U/ml), IL-1 (173 +/- 102 U/ml), and IL-6 (10.8 +/- 3.1 x 10(4) U/ml) were observed at PIH 2, 5, and 8, respectively. Tumor necrosis factor and IL-1 activities returned to baseline values by PIH 8 and 24, respectively; however, IL-6 activity remained high. Interleukin 1 inhibitory activity (27.4 +/- 2.25 IU/ml) was detected in all PIH-24 samples from control joints, but was not detected at any other time in control joints (limit of detection, 20 IU/ml). Tumor necrosis factor activity was not detected in any synovial fluid sample from joints treated with endotoxin/eqTNF antibody. In contrast, endotoxin IL-1 inhibitory activity (PIH 24) was higher in eqTNF antibody-treated joints (41.0 +/- 7.7 IU/ml) than in control joints, but the difference was not significant. Mean WBC count and protein concentration in control and treated joints were maximal at PIH 8. The curves for mean values of WBC count and total protein concentration were not significantly different in treated versus control joints. Swelling in each treated joint was either less than or the same as that in the opposite control joint at even, time in the initial 8 PIH. There was significant (P = 0.043) difference between treated and control joints at PIH 5 and 8. These results describe a profile of synovial fluid TNF, IL-1, IL-6 bioactivities, and IL-1-inhibitory activity during the initial 24 hours of synovitis induced by intra-articular administration of endotoxin in horses. Our eqTNF monoclonal antibody was effective in neutralizing TNF activity in synovial fluid when administered intra-articularly with endotoxin in horses. The induction of IL-1, IL-1 inhibitory activity IL-6, WBC, and total protein concentration responses are largely independent of TNF activity in synovial fluid of horses receiving endotoxin intra-articularly.

Animals↗

PubMed1994

Effects of polymyxin B and Salmonella typhimurium antiserum on horses given endotoxin intravenously.

Polymyxin B and an antiserum against an Re mutant Salmonella typhimurium were evaluated for protective effect in an equine model endotoxemia. Six 3- to 5-month-old foals were given endotoxin (0.25 micrograms/kg of body weight) IV after no pretreatment, or pretreatment with polymyxin B (6,000 U/kg, IV) or S typhimurium antiserum (1.5 ml/kg, IV). When given without pretreatment, endotoxin caused transient recumbency and increases in rectal temperature, and heart and respiratory rates. In addition, leukopenia and increases in circulating tumor necrosis factor (TNF) and interleukin 6 (IL-6) activities were detected. Compared with results obtained when endotoxin was given alone, pretreatment with polymyxin B resulted in significantly (P < 0.05) lower maximal plasma TNF and IL-6 activities, and significantly lower rectal temperature and respiratory rate. In contrast, compared with effects of endotoxin given without pretreatment, use of antiserum was associated with significantly (P < 0.05) higher respiratory rate, maximal plasma IL-6 activity, and total TNF response (as determined by areas under curves of plasma TNF vs time). These results indicate that polymyxin B may have potential as a treatment for equine endotoxemia. Salmonella typhimurium antiserum had no positive effect in this model, and, under certain conditions, may exacerbate the actions of endotoxin.

Animals↗

PubMed1993

The mouse macrophage activation-associated marker protein, p71/73, is an inducible prostaglandin endoperoxide synthase (cyclooxygenase).

The inducible protein p71/73 marks the response of mouse macrophages to one of several stimuli (e.g., bacterial lipopolysaccharide or poly I:C) that trigger the expression of cytolytic activity when these cells have previously been primed for tumor cell killing by interferon-gamma (IFN-gamma). The results reported here identify this marker protein as the inducible prostaglandin endoperoxide synthase (PES), TIS10/PES-2. Identification was based on four findings: (1) p71/73, like the TIS10/PES-2 protein, was associated with cellular membranes; (2) the sequence of amino acids in the NH2 terminus of both p71 and p73 was 96% identical to the predicted NH2-terminal sequence of the TIS10/PES-2 protein; (3) a polyclonal antiserum raised against the COOH-terminal region of the TIS10/PES-2 gene product recognized p71/73 in immunoblots; and (4) dexamethasone, which blocks induction of TIS10/PES-2 expression, inhibited the induction of both p71/73 synthesis and tumoricidal activity in macrophage. Several regulatory roles for this protein in the activation process are possible.

Amino Acid Sequence↗

PubMed1993

Effects of intra-articularly administered endotoxin on clinical signs of disease and synovial fluid tumor necrosis factor, interleukin 6, and prostaglandin E2 values in horses.

In each of 4 horses, sterile synovitis was induced by intra-articular injection of 3 micrograms of Escherichia coli endotoxin (lipopolysaccharide, LPS) into one antebrachiocarpal joint; an equal volume (2 ml) of phosphate-buffered saline solution (PBSS) was injected into the opposite, control carpus. Blood and 1.5 ml of synovial fluid were obtained at postinjection hours (PIH) 0, 2, 4, 8, 12, 18, 42, 66, and 144. Synovial fluid sample collection was accomplished by use of an indwelling, intra-articular catheter through PIH 12, and by arthrocentesis subsequently. Joint fluid samples were analyzed for cell counts, protein concentration, cytologic variables, and tumor necrosis factor (TNF), interleukin 6 (IL-6), and prostaglandin E2 (PGE2) values. Tumor necrosis factor and IL-6 activities and WBC count were also measured in blood. To monitor local inflammation, skin temperature of each carpus was imaged, using a thermographic scanner prior to each sample collection time. Horses had minimal systemic effects. Mean (+/- SEM) rectal temperature increased significantly to 39.02 +/- 0.15 C only at PIH 18 after intra-articular injection of LPS. One horse had signs of mild depression from PIH 7 to 18, but its vital signs did not change appreciably. Each horse had mild signs of discomfort in the LPS-injected limb from PIH 1 to 3 until PIH 8 to 10. Mean peak surface temperature of the LPS-injected carpi was significantly higher than that of control carpi from PIH 8 to 144 (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗

PubMed1992

Primary meningeal lymphoma in a horse.

Primary meningeal lymphoma was diagnosed in an 18-year-old Morgan gelding. The horse was examined because of a 3-day history of progressive ataxia and weakness. The gait abnormalities were worse on the left side, and the pelvic limbs were more affected than the thoracic limbs. Additional findings included signs of depression, miosis of the left pupil, ptosis of the left upper eyelid, and areas of muscle atrophy on the left side of the neck and over the dorsal aspect of the left scapula. Inflammatory changes were evident in the CSF. At necropsy, there was diffuse and irregular thickening of the dura mater along the entire spinal cord. Histologic examination revealed infiltration of the leptomeninges with neoplastic lymphocytes.

Animals↗

PubMed1992

Focal premature physeal closure (hyena disease) in calves.

Focal premature closure of long-bone physes was the cause of conformational abnormalities that affected about 1% of a herd of dairy replacement calves. Affected calves were noticed to "bunny hop" by 3 months of age, and by 6 months of age, they developed abnormal conformation characterized by short pelvic limbs. This condition resembled "hyena disease," which has been described in dairy calves in Europe and Japan. With the exception of the aforementioned abnormalities and femoropatellar joint distention, a group of 5 affected calves examined were clinically normal. At necropsy, focal to almost complete closure of physes was found in the humeri, tibias, and femurs. Cause was not established for the condition; however, it was discovered that the calves had been given supplemental vitamin A/D3 in amounts greater than 10 times those recommended.

Animals↗

PubMed1992

Anti-equine tumor necrosis factor (TNF) activity of antisera raised against human TNF-alpha and peptide segments of human TNF-alpha.

Antisera raised in rabbits against either purified recombinant-derived human tumor necrosis factor (TNF)- alpha (huTNF) or huTNF peptide-bovine thyroglobulin conjugates were evaluated for anti-equine TNF (eqTNF) activity. Binding and neutralizing anti-eqTNF activities were found in antisera raised against whole huTNF or against either of the peptides containing the N-terminal 15 amino acids of huTNF (huTNF[1-15] and huTNF[1-31]). Anti-eqTNF activity was not detected in antisera raised against huTNF[65-79], huTNF[98-111] or huTNF[124-141] peptides. The addition of excess huTNF[1-15] completely inhibited the ability of anti-huTNF[1-15] to bind eqTNF and reduced by approximately 25% the anti-eqTNF activity of an antiserum raised against whole huTNF. Nonconjugated huTNF[1-15] did not have eqTNF agonist or antagonist activity. Results were consistent with previous structural and functional data implicating the N-terminus of huTNF in receptor binding and indicate that the homologue of huTNF[1-15] on eqTNF may be a potentially important target for neutralizing anti-eqTNF antibodies.

Amino Acid Sequence↗

PubMed1992

Induction of the acute-phase cytokine, hepatocyte-stimulating factor/interleukin 6, in the circulation of horses treated with endotoxin.

Because hepatocyte-stimulating factor/interleukin 6 (IL-6) is the principal inducer of acute-phase protein synthesis in the liver, quantification of its activity in blood provides an early and sensitive assessment of the acute-phase response. Circulating IL-6 activity was monitored in 4 adult horses for 72 hours after IV administration of endotoxin. In 4 experiments performed at weekly intervals and in randomized order, each horse was given endotoxin--1,000 30, 1, and 0 ng/kg of body weight. Plasma IL-6 activity was quantified as the ability to promote growth of the IL-6-dependent B-cell hybridoma, B13.29 clone B9. Interleukin-6 activity (171 +/- 10.2 U/ml) was found in all pretreatment plasma samples and was significantly (P less than 0.05) increased above baseline from 2 to 12 hours after 1,000 ng of endotoxin/kg was given and at 3 hours after 30 ng of endotoxin/kg was given. After 1,000- or 30-ng/kgt dosage of endotoxin, peak plasma IL-6 activity (10,128 +/- 4,096 and 1,555 +/- 1,326 U/ml, respectively) was observed for 3 hours. The IL-6 response of endotoxin-treated horses began about 1 hour after tumor necrosis factor appeared in the circulation, and its course closely approximated the endotoxin-induced febrile reaction. Significant increase in plasma IL-6 activity was not detected in horses given 1 ng of endotoxin/kg or control buffer.

Acute-Phase Reaction↗

PubMed1992

Association between serum cytotoxicity and selected clinical variables in 240 horses admitted to a veterinary hospital.

A total of 378 serum samples from 240 hospitalized horses and 47 sera from healthy control horses were assayed for growth effects on actinomycin D-treated L929 cells. On average, patient and control sera stimulated cell growth; however, mean percentage of the relative growth index (RGI) of sera from clinical cases was significantly (P less than 0.001) lower than that of control sera. Approximately 35% of patient sera and 6% of control sera had tumor necrosis factor-like cytotoxic activity for L929 cells (ie, RGI less than 100%). Sera from horses with either peritoneal leakage of gastrointestinal tract contents or any bacterial infection were significantly (P less than 0.05) more cytotoxic than sera from horses that did not have these clinical factors. A clear tendency was evident for horses that had the highest serum cytotoxicity (RGI less than 75%) to also have clinical profiles suggestive of endotoxemia. Fever, leukopenia, diarrhea, and gastrointestinal tract leakage were significantly (P less than 0.05) overrepresented among these horses, compared with horses without serum cytotoxicity. Bacterial infections and abdominal surgeries were also increased in this group, but not significantly. Of the 14 horses with serum RGI less than 75%, 13 had some form of gastrointestinal tract disease and the other had gram-negative septicemia. Survival to discharge was significantly (P less than 0.05) lower among horses in the high-cytotoxicity group than among horses without serum cytotoxicity. Diarrhea and bacterial infections were the only clinical factors found more frequently in horses with low serum cytotoxicity than in horses without serum cytotoxicity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

Activation-associated marker proteins: peptide mapping and their expression in macrophage cell lines.

The activation-associated markers, p47b and p71/73, have been recognized as minor proteins in peritoneal and bone marrow culture-derived macrophages activated for tumor cell killing. Proteins with identical characteristics of migration on 2-dimensional gels and comparable Cleveland peptide maps are described here in macrophage cell lines that could be activated for tumor cell killing (J774A.1, RAW 264, UNC-2). Macrophage cell lines that could not be activated (P388D1 and PU5-1.8) did not express the markers. The expression of these markers in activatable macrophage cell lines strengthens their association with the activation process and provides a bulk source of the proteins for purification studies.

Animals↗

PubMed1991

Cytotoxic tumor necrosis factor activity produced by equine alveolar macrophages: preliminary characterization.

Blood monocytes and alveolar macrophages (AM) were harvested from foals (aged 46 days to 6 months) and cultured in either medium alone or medium containing 10 micrograms/ml bacterial lipopolysaccharide (LPS). After 24 h, culture supernates were collected and analyzed for cytotoxic activity on sensitized L929 cells. Both monocytes and AM that had been treated with LPS produced significantly more cytotoxic activity than the same cell type exposed to medium lacking LPS. LPS-treated macrophages secreted significantly more cytotoxic activity (120 +/- 17.8 U/ml) than did LPS-treated monocytes (47.3 +/- 17.0 U/ml); however, constitutive production of cytotoxin by monocytes was higher (16.7 +/- 4.1 versus 1.2 +/- 1.2 U/ml). The identification of the cytotoxin as tumor necrosis factor (TNF) was strongly suggested by its reactivity with a rabbit antiserum directed against the N-terminal 15 amino acids of human TNF. TNF secretion by AM increased in a dose-dependent manner between LPS concentrations of 0.0001 and 1 microgram/ml, then leveled off. Most of the cytotoxic TNF activity produced by AM was secreted within the first 8 h after initial contact with LPS. Macrophage supernatant TNF was stable over a pH range of 6-11, but lost activity when kept at a pH less than 6. Equine TNF also was destroyed by exposure for 1 h to temperatures more than 60 degrees C. TNF bioactivity was recovered as a single peak after crude macrophage supernate was subjected to analysis by either anion exchange or gel filtration chromatography (molecular weight approximately 56,000).

Animals↗

PubMed1991

Tumor necrosis factor activity in the circulation of horses given endotoxin.

Serum and plasma from horses injected with endotoxin was examined for cytotoxic activity. Each of the cell lines, L929 and WEHI 164 clone 13, was sensitive to the cytotoxic effects of equine serum; however, a precipitation artifact caused by the use of isopropanol in the WEHI assay limited the use of this assay to samples containing less than 2 mg of protein/ml. In foals treated with a sublethal IV bolus of 5 micrograms of lipopolysaccharide (LPS)/kg and in adult horses given a low-dose continuous infusion of LPS (30 ng/kg/h for 4 hours), cytotoxic activity was detected in all serum or plasma samples taken between 30 minutes and 4 hours after LPS infusion began. In horses given either continuous or bolus LPS infusions, circulating cytotoxic activity peaked at 1 to 2 hours before decreasing sharply. The onset of pyrexia after LPS infusion coincided with the appearance of circulating cytotoxic activity, but the temperature remained high, even after cytotoxic activity disappeared. Treatment of horses with flunixin meglumine (1 mg/kg) appeared to blunt the pyrexic effect of low-dose continuous LPS infusion, but had no significant effect on circulating cytotoxic activity. Incubation of serum samples with an antibody raised against a portion of human tumor necrosis factor (TNF) resulted in the removal of greater than 90% of serum cytotoxicity, suggesting strongly that the cytotoxic activity was attributable to TNF. These findings are consistent with the hypothesis that TNF is an early acting mediator of the effects of endotoxin in the horse.

Animals↗

PubMed1989

Macrophage activation-associated proteins. Characterization of stimuli and conditions needed for expression of proteins 47b, 71/73, and 120.

The expression of cellular proteins was analyzed by two-dimensional gel electrophoresis during and after exposure of mouse macrophages to either mouse rIFN-gamma or natural MuIFN-beta sufficient to prime macrophages for tumor cell killing. The reversible inhibitor of protein synthesis, cycloheximide (CY), was included in some experiments during exposure to IFN. While it was present, CY suppressed protein synthesis by greater than 90%, but did not affect priming for tumor cell killing that was induced by either kind of IFN, as measured in cytotoxicity assays. Further analysis showed that, after CY and IFN were removed, protein synthesis recovered fully within 1 h. p47b, a protein that has been associated closely with the induction of the primed state in mouse macrophages, was then substantially expressed despite no new stimulation by IFN. Thus, macrophages in which protein synthesis had been reversibly inhibited delayed full processing of a signal delivered by IFN, until after protein synthesis had resumed. Such a delay in processing may explain how macrophages subsequently became activated, despite treatment with CY. The expression of the protein doublet, p71/73, was induced, regardless of which of three dissimilar agents (LPS, heat killed Listeria monocytogenes, poly I:C) was used to trigger the expression of cytolytic activity by primed macrophages. Therefore, the likelihood was increased that p71/73, expressed with p47b, is a valid phenotypic marker for fully activated, cytolytic macrophages. By contrast, p120, another protein that has been proposed as a marker of full activation in peritoneal macrophages, was expressed by bone marrow culture-derived macrophages regardless of whether or not they were cytolytic for tumor cells. It cannot be regarded as a reliable marker of macrophage activation in all circumstances, therefore.

Animals↗