Methyl isobutyl ketone-induced hepatocellular carcinogenesis in B6C3F1 mice: A constitutive androstane receptor (CAR)-mediated mode of action.

In a National Toxicology Program (NTP) chronic inhalation study with methyl isobutyl ketone (MIBK), increases in hepatocellular adenomas and hepatocellular adenomas and carcinomas (combined) were observed in male and female B6C3F1 mice at 1800 ppm. A DNA reactive Mode-of-Action (MOA) for this liver tumor response is not supported by the evidence as MIBK and its major metabolites lack genotoxicity in both in vitro and in vivo studies. Constitutive androstane receptor (CAR) nuclear receptor-mediated activation has been hypothesized as the MOA for MIBK-induced mouse liver tumorigenesis. To further investigate the MOA for MIBK-induced murine liver tumors, male and female B6C3F1, C57BL/6, and CAR/PXR Knockout (KO) mice were exposed to either 0 or 1800 ppm MIBK for 6 h/day, 5 days/week for a total of 10 days. On day 1, mice were implanted with osmotic mini-pumps containing 5-Bromo-2-deoxyuridine (BrdU) 1 h following exposure and humanely euthanized 1-3 h following the final exposure. B6C3F1 and C57BL/6 mice had statistically significant increases in liver weights compared to controls that corresponded with hepatocellular hypertrophy and increased mitotic figures. Hepatocellular proliferation data indicated induction of S-phase DNA synthesis in B6C3F1 and C57BL/6 mice exposed to 1800 ppm MIBK compared to control, and no increase was observed in MIBK exposed CAR/PXR KO mice. Liver gene expression changes indicated a maximally-induced Cyp2b10 (CAR-associated) transcript and a slight increase in Cyp3a11(PXR-associated) transcript in B6C3F1 and C57BL/6 mice exposed to 1800 ppm MIBK compared to controls, but not in Cyp1a1 (AhR-associated) or Cyp4a10 (PPAR-α-associated) transcripts. CAR/PXR KO mice exposed to 1800 ppm MIBK showed no evidence of activation of AhR, CAR, PXR or PPAR-α nuclear receptors via their associated transcripts. MIBK induced hepatic effects are consistent with a phenobarbital-like MOA where the initiating events are activation of the CAR and PXR nuclear receptors and resultant hepatocellular proliferation leading to rodent liver tumors.

Abnormal mobility of neonatal polymorphonuclear leukocytes. Relationship to impaired redistribution of surface adhesion sites by chemotactic factor or colchicine.

To determine the mechanism(s) of diminished, stimulated, and directed migration of neonatal (N) polymorphonuclear leukocytes (PMN), chemotactic factor (CF) sensory and PMN effector functions were studied in healthy N and adult or maternal controls (C). N PMN demonstrated high affinity binding for N-formyl-methionyl-leucyl-[3H]phenylalanine (fMLP), which was saturable between 40 and 100 nM as observed with C PMN. The kinetics of binding and the characteristics of dissociation of binding by N PMN were equivalent to control PMN. Both "threshold" and "peak" concentrations (1 and 10 nM, respectively) of fMLP effected comparable PMN chemiluminescence among neonates and controls. An equivalent threshold concentration (0.05 nM) of fMLP effected N and C PMN shape change in suspension, and a maximally effective concentration (5 nM) induced comparable bipolar configuration, although uropod formation was only 38 +/- 8% of N PMN, compared with 73 +/- 11% of C PMN (P less than 0.01). Striking abnormalities of N PMN adherence were identified: mean +/- SD base-line (unstimulated) N adherence values (39 +/- 8%) were equal to C (38 +/- 9%), but diminished increments in response to single CF stimuli were noted among N (fMLP: 42 +/- 7% (N), 70 +/- 11% (C); C5a: 41 +/- 6% (N), 68 +/- 6% (C); BCF: 41 +/- 6% (N), 63 +/- 9% (C), P less than 0.01 for each CF). On sequential exposure to increasing concentrations of CF N PMN failed to demonstrate expected decreased adherence values; sequential stimuli with fMLP (0.1 nM, 10 nM) or C5a (8 microgram protein/ml, 32 microgram protein/ml) effected mean +/- 1 SD values of 51 +/- 9% (N), 30 +/- 9% (C), and 34 +/- 10 (N), 48 +/- 14% (C), respectively. As demonstrated with a latex bead-binding technique, N PMN failed to redistribute adhesion sites to the cell's tail under the same experimental conditions; in 21 N samples studied, restricted unipolar binding occurred in 33 +/- 8% (fMLP) or 37 +/- 7% (C5a) of PMN in contrast to C values of 70% (fMLP), or 71% (C5a), P less than 0.001. Similar findings were observed when PMN were preincubated with colchicine (25 microgram/ml); expected diminished adherence scores (compared with base-line values) were demonstrated with C PMN but not with N cells, P less than 0.01. Additionally colchicine-induced redistribution of adhesion sites as was observed with C samples (72 +/- 14% unipolar binding) was significantly (P less than 0.001) less among N PMN (31 +/- 11% unipolar binding). These investigations indicate that CF sensory mechanisms of N PMN are normal, compared with healthy adult or maternal controls. Diminished stimulated locomotion of the N PMN may be functionally related to reduced modulation of cell adhesiveness by chemotactic stimulation.

Recruitment of CD11b/CD18 to the neutrophil surface and adherence-dependent cell locomotion.

Chemotactic stimulation of neutrophils results in translocation of CD11b/CD18 (Mac-1) from intracellular storage pools to the cell surface. Though results from several laboratories indicate that the newly arrived surface Mac-1 is not involved in the adherence induced by the initial stimulus, the present study addresses the hypothesis that this Mac-1 plays a role in subsequent adherence-dependent functions. The response of human neutrophils to changing concentrations of a chemotactic stimulus was evaluated by determining the amount of newly arrived surface Mac-1, and Mac-1-dependent adhesion and locomotion. Small step-wise increases in the concentration of f-Met-Leu-Phe (FMLP) resulted in proportional stepwise increases in surface Mac-1 that plateaued within 2-4 min. This newly arrived Mac-1 supported adhesion to protein-coated surfaces only when the cells were exposed to an additional increase in the FMLP stimulus level. Adherence-dependent cellular locomotion was evaluated in chambers that allowed rapid changes in the stimulus concentration. Repeated small increments in the stimulus level at 200-s intervals resulted in significantly longer migration paths than a single-step increase in the stimulus. The results support the hypothesis that small increments in the chemotactic stimulus bring Mac-1 to the cell surface, and this newly mobilized Mac-1 is available for adherence-dependent locomotion with subsequent increases in the concentration of the stimulus.

Recognition of an endothelial determinant for CD 18-dependent human neutrophil adherence and transendothelial migration.

Human neutrophil (PMN) attachment to human umbilical vein endothelial cells (HUVEC) was evaluated in vitro using two MAbs, R6-5-D6 and RR1/1, that recognize intercellular adhesion molecule-1 (ICAM-1), and one MAb, TS1/18, that recognizes CD18. Pretreatment of the HUVEC with anti-ICAM-1 MAbs produced greater than 50% inhibition of attachment to HUVEC, and IL-1 (0.5 U/ml)- or lipopolysaccharide (LPS) (10 ng/ml)-stimulated HUVEC, and greater than 99% inhibition of f-Met-Leu-Phe (0.5 nM) enhanced adherence. Anti-ICAM-1 MAbs also inhibited by greater than 85% the transendothelial migration induced by 4-h IL-1 (0.5 U/ml) and LPS (10 ng/ml) activation of the HUVEC. That these effects involved a CD18-dependent mechanism is supported by the following results: pretreatment of PMN with TS1/18 produced the same degree of inhibition of attachment and migration as seen with R6-5-D6. In addition, the use of both MAbs together did not further increase the inhibition of cell attachment to stimulated HUVEC. The attachment of PMN from patients with CD18 deficiency to stimulated HUVEC was not reduced by R6-5-D6, and both R6-5-D6 and TS1/18 revealed the same time course for appearance and disappearance of an adherence component on stimulated HUVEC not blocked by either MAb. These results demonstrate that attachment and transendothelial migration of PMN in vitro depend substantially on both CD18 on the PMN and ICAM-1 on the endothelial cell.

Abnormalities of polymorphonuclear leukocyte function associated with a heritable deficiency of high molecular weight surface glycoproteins (GP138): common relationship to diminished cell adherence.

Investigations of polymorphonuclear leukocyte (PMN) function were performed in a 5-yr-old white female with delayed umbilical cord separation, impaired pus formation, and a severe defect of PMN chemotaxis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated an almost total deficiency of a high molecular weight glycoprotein(s) (GP138) in the granule and membrane fractions of the patient's cells, and NaB3H4-galactose oxidase labeling demonstrated the absence of a major glycoprotein complex on the surface of her PMNs. Monoclonal antibodies (MAb) were employed in flow cytometry experiments to demonstrate that two previously characterized glycoproteins (Mo1 and LFA1) were undetectable on the surface of the patient's PMNs and monocytes. Immunoprecipitation of 125I-labeled patient cells with subunit specific MAbs confirmed that the alpha-subunits of Mo1 (155 kD) and LFA1 (177 kD) and their common beta-subunit (94 kD) were totally deficient. Functional analyses of patient PMNs demonstrated severe impairment of adherence- and adhesion-dependent cell functions including spreading, aggregation, orientation in chemotactic gradients, antibody-dependent cellular cytotoxicity, and phagocytosis of particles (Oil-Red-0-paraffin, zymosan) selectively opsonized with C3-derived ligands. Patient PMNs demonstrated a normal capacity to rosette with IgG or C3b-coated sheep erythrocytes, but rosette formation with C3bi-coated erythrocytes was profoundly diminished. Adhesion-independent functions including shape change, N-formyl-methionyl-leucyl-3H-phenylalanine binding, and O-2 generation or secretion elicited by soluble stimuli were normal. Membrane fluidity, surface charge, and microtubule assembly were also normal. These findings provide new evidence that critical PMN surface glycoproteins are required to facilitate multiple adhesion-dependent cellular functions of the inflammatory response.

Interleukin-8 gene induction in the myocardium after ischemia and reperfusion in vivo.

Neutrophil adhesion and direct cytotoxicity for cardiac myocytes require chemotactic stimulation and are dependent upon CD18-ICAM-1 binding. To characterize the potential role of IL-8 in this interaction, canine IL-8 cDNA was cloned and the mature recombinant protein expressed in Escherichia coli BL21 cells. Recombinant canine IL-8 markedly increased adhesion of neutrophils to isolated canine cardiac myocytes. This adhesion resulted in direct cytotoxicity for cardiac myocytes. Both processes were specifically blocked by antibodies directed against CD18 and IL-8. In vivo, after 1 h of coronary occlusion, IL-8 mRNA was markedly and consistently induced in reperfused segments of myocardium. IL-8 mRNA was not induced in control (normally perfused) myocardial segments. Minimal amounts of IL-8 mRNA were detected after 3 or 4 h of ischemia without reperfusion. Highest levels of induction were evident in the most ischemic myocardial segments. IL-8 mRNA peaked in the first 3 h of reperfusion and persisted at high levels beyond 24 h. IL-8 staining was present in the inflammatory infiltrate near the border between necrotic and viable myocardium, as well as in small veins in the same area. These findings provide the first direct evidence for regulation of IL-8 in ischemic and reperfused canine myocardium and support the hypothesis that IL-8 participates in neutrophil-mediated myocardial injury.

Monoclonal antibody targeting of liposomes to mouse lung in vivo.

A monoclonal antibody (MoAb), 273-34A, specifically binds to an epitope expressed almost exclusively on capillary endothelial cells of the lung. Within 15 min after i.v. injection, approximately 80 to 85% of the injected radioiodinated MoAb 273-34A accumulates in the lung. Approximately 90 to 95% of the recovered dose is found in the lung for up to 1 week postinjection. Ratios of MoAb 273-34A to a nonspecific, irrelevant MoAb 135-14 are 250 to 285 times higher in the lung than in the serum. When 273-34A was coupled with palmitic acid and incorporated into liposomes, the amount of 125I-labeled liposomes recovered per g of tissue was 12 times higher in the lung than in the liver at 15 min postinjection, and 22 times higher at 5 h postinjection. At 24 h postinjection the amount of liposomes per gram of lung tissue was still 6.0 times the amount per gram of liver tissue. Liposomes conjugated to MoAb 273-34A locate in the lung 20 and 15 times better than do liposomes conjugated to the nonspecific MoAb 135-14 at 15 min and 24 h postinjection, respectively. The results indicate that this immunoliposome system could be used as a model for enhanced drug delivery to the lung. The potential use for delivering anticancer drugs for therapy of lung tumors is discussed.

Leukocyte activation with platelet adhesion after coronary angioplasty: a mechanism for recurrent disease?

OBJECTIVES: The purpose of this pilot study was to determine whether leukocyte activation occurs, whether leukocyte-platelet complexes develop and whether there is any association between these findings and clinical outcome after coronary angioplasty. BACKGROUND: Increased expression of CD11b on monocytes and neutrophils promotes their adhesion to endothelial cells, extracellular matrix and smooth muscle cells. Thrombin-activated platelets adhere to monocytes and neutrophils through P-selectin. These cell complexes may affect the inflammatory process and, thus, the outcome of coronary angioplasty. METHODS: During elective single-vessel coronary angioplasty in 11 men, samples were obtained for flow cytometric detection of CD11b, as well as the percent of leukocytes with adherent platelets and the intensity of bound platelet fluorescence (number of platelets/leukocyte). RESULTS: After angioplasty, there was an increase in CD11b (monocytes: p = 0.001, neutrophils: p = 0.02) and leukocytes with adherent platelets (p = 0.02). During follow-up, five patients remained in stable condition and six had subsequent clinical events: restenosis and progression of disease requiring coronary artery bypass grafting (n = 3), myocardial infarction involving the dilated artery (n = 1) and unstable angina (n = 2). Values for leukocyte CD11b expression, the percent of leukocytes with adherent platelets and the intensity of bound platelet fluorescence were higher both before and after angioplasty in the six patients experiencing clinical events. CONCLUSIONS: Despite standard aspirin and heparin therapy, leukocyte activation with platelet adherence occurs after coronary angioplasty. The magnitude of leukocyte activation and platelet adherence appears to be higher in patients experiencing late clinical events.

Chimeric 7E3 Fab (ReoPro) decreases detectable CD11b on neutrophils from patients undergoing coronary angioplasty.

OBJECTIVES: The purpose of this study was to monitor the effects of chimeric 7E3 Fab (ReoPro) on leukocyte and platelet activation and interaction during coronary angioplasty. BACKGROUND: Increased expression of CD11b on monocytes and neutrophils promotes their adhesion to endothelial cells, extracellular matrix and smooth muscle cells. Thrombin-activated platelets adhere via P-selectin to monocytes and neutrophils. These cell interactions may affect the outcome of coronary angioplasty. METHODS: During coronary angioplasty, venous blood was obtained for flow cytometric detection of leukocyte CD11b; platelet CD41a, CD61a and CD62P; the percentage of leukocytes with adherent platelets and the intensity of bound platelet fluorescence. RESULTS: Leukocyte CD11b expression increased after angioplasty in control patients (neutrophils 171+/-25 to 255+/-31 mean fluorescence intensity [MFI, mean+/-SEM], n=25, p < 0.0001; monocytes 200+/-40 to 248+/-36 MFI, n=17, p < 0.05) and decreased in the patients selected to receive chimeric 7E3 Fab (neutrophils 146+/-30 to 82+/-22 MFI, n=25, p < 0.0001; monocytes 256+/- 53 to 160+/-38 MFI, n= 17, p < 0.05). Neutrophil CD11b decreased after in vitro incubation of whole blood with chimeric 7E3 Fab (n=5, p=0.01), but fMLP-induced increases in CD11b were not prevented. The CD11b expression was unchanged and increased with fMLP stimulation after in vitro incubation of isolated neutrophils with chimeric 7E3 Fab. Direct-labeled chimeric 7E3 Fab was not detected bound to neutrophils in whole blood or isolated cells using flow cytometric techniques. Adhesion of isolated neutrophils to protein-coated glass was not prevented by in vitro incubation with chimeric 7E3 Fab. Platelet activation increased after angioplasty in control patients (CD62P 8.9+/-0.8 to 12.3+/-1.2 MFI, n=25, p < 0.05; CD41a 382+/-25 to 454+/-26 MFI, n=25, p < 0.05, CD61a 436+/-52 to 529+/-58 MFI, n=11, p < 0.05); it did not increase in the patients selected to receive chimeric 7E3 Fab (CD62P 13.2+/-1.0 to 9.0+/-0.9 MFI, n=25, p < 0.05; CD61a 398+/-32 to 410+/-38 MFI, n=7, p=NS). Leukocytes with adherent platelets tended to increase in the control group of patients and decrease after the procedure in patients selected to receive chimeric 7E3 Fab; individual and procedure-related variability were marked. CONCLUSIONS: Despite standard aspirin and heparin therapy, leukocyte and platelet activation with platelet adherence to leukocytes occurs after coronary angioplasty. Although chimeric 7E3 Fab does not bind to leukocytes directly, it influences CD11b expression in whole blood. Modulation of platelet and leukocyte activation and interaction by chimeric 7E3 Fab may contribute to an improved outcome after coronary angioplasty.

Impaired motility of neonatal PMN leukocytes: relationship to abnormalities of cell orientation and assembly of microtubules in chemotactic gradients.

To allow a further understanding of the pathogenesis of impaired stimulated locomotion by polymorphonuclear leukocytes (PMNs) in human neonates, we studied cellular orientation by neonatal PMNs in response to well-defined chemotactic gradients (Zigmond orientation chambers) and characterized the cytoplasmic microtubule (MT) complex of neonatal PMNs during cell orientation and movement. PMN suspensions obtained from 52 neonates demonstrated a diminished capacity to undergo orientation at all time intervals after exposure to gradients of N-formyl-methionyl-leucyl phenylalanine (f-Met-Leu-Phe) or C5a. Among responding (orienting) neonatal PMNs observed, only 70% (f-Met-Leu-Phe) or 59% (C5a) oriented accurately (toward chemotactic gradients) as compared to values of 96% (f-Met-Leu-Phe) or 92% (C5a) for adult controls. Furthermore, neonatal PMNs failed to alter their direction of orientation/migration when chemotactic gradients were reversed. Similar abnormalities were observed when 10-fold gradients of f-Met-Leu-Phe were employed over a concentration range between 10(-7) and 10(-11) M. Employing tubulin immunofluorescence, the cytoplasmic MT complex of-neonatal PMNs was assessed prior to and after cell exposure to uniform concentrations or gradients of chemotactic factors (CFs). MT assembly by neonatal PMNs studied under these experimental conditions was significantly diminished. Neonatal cell suspensions demonstrated 26 +/- 5 (f-Met-Leu-Phe) or 27 +/- 6 (C5a) MT/cell as compared to respective values of 36 +/- 6 or 35 +/- 5 for adult suspensions (P less than .001). MT lengths of neonatal PMNs increased from 6.7 +/- 1 micron (PBS) to 7.5 +/- 1 micron (f-Met-Leu-Phe) or 7.3 +/- 1 micron (C5a) as compared to values of 6.5 +/- 1 micron (PBS), 11.1 +/- 1 micron (f-Met-Leu-Phe), and 10.9 +/- 1 micron (C5a) for adult PMNs exposed to gradients or uniform concentrations of CFs (P less than .01 for both f-Met-Leu-Phe and C5a). Thus, the polymerized tubulin mass product of chemotactically stimulated neonatal PMNs (202 micron) was significantly (P less than .001) diminished as compared to adult PMNs (360 micron). As shown by a [3H]colchicine binding assay, impaired MT assembly could not be attributed to diminished cytoplasmic tubulin content of neonatal PMNs, which was comparable to adult PMNs.

Elevated blood lead levels associated with illegally distilled alcohol.

Whiskey produced in illegal stills (ie, "moonshine") remains an important and underappreciated source of lead toxicity in some rural counties of the Southeast. From March 5 through October 26, 1991, eight adult patients with elevated blood lead levels were identified at a rural county hospital in Alabama and were reported to the Alabama Department of Public Health notifiable disease surveillance system. A case-patient was defined as any person 17 years of age or more who presented to the hospital from January 1, 1990, through December 31, 1991, and had a blood lead level of 0.72 mumol/L or more (15 micrograms/dL or more). To identify cases and potential sources of lead exposure, we reviewed medical and laboratory records from the hospital, interviewed patients with elevated blood lead levels, and determined the lead content of moonshine samples. Nine patients met the case definition, including one patient who was not reported to the state. Patients ranged in age from 28 to 62 years; blood lead values ranged from 0.77 to 12.50 mumol/L (16 to 259 micrograms/dL). The most frequent signs of possible lead toxicity included seizures (six), microcytic anemia (five), and encephalopathy (two); one patient died. The only identified source of lead exposure for the nine patients was moonshine ingestion. Moonshine samples available from local stills contained sufficient amounts of lead (340 to 4600 mumol/L) to result in the observed blood lead levels. This investigation emphasizes the adverse health effects and ongoing public health impact of moonshine ingestion.

Comparative sensitivities of radioimmunoassay, crossoverelectrophoresis, and agar gel immunodiffusion for HBAg detection.

The relative sensitivities of radioimmunoassay, crossoverelectrophoresis, and agar gel immunodiffusion for detecting HBAg in unconcentrated versus concentrated serum samples are compared and discussed. The sensitivities of the various immunologic methods were enhanced severalfold in concentrated serum samples. This is a fundamental consideration in designing a screening program for detecting HBAg and anti-HBAg antibody in hospital patients and donors. The authors propose that serum samples that have marginal reactions or low counts (RIA) be concentrated with Lyphogel or MiniconTM-125 concentrator and retested before test results are reported. Specificity of positive results was studied by neutralization tests with human serum containing antibodies to HBAg.

Variations in temperature and oxygen content do not alter levels of thiobarbiturate reactive material in human breast tumour cells (ZR-75-1) incubated with gamma-linolenic acid.

The mechanism by which tumour cells may be killed in vitro by exogenous polyunsaturated fatty acids may involve lipid peroxidation. Gamma-linolenic acid caused a dose and time-dependent reduction in ZR-75-1 cell growth. However, altering either the incubator temperature (35, 37 and 39 degrees C) or the oxygen content (16, 21 and 26%) had little effect on either the growth of cells in the presence of gamma-linolenic acid or on thiobarbiturate reactive material levels over a 7 day period. Thus, small changes in cell culture conditions do not affect 18:3n-6 cytotoxicity or markers of lipid peroxidation.

Effects of macrocyclic trichothecene congeners on the viability and mitogenesis of murine splenic lymphocytes.

The effects of several congeners of the macrocyclic class of trichothecene mycotoxins on murine splenic cells in vitro were investigated. The mycotoxins were roritoxin B, myrotoxin B, roridins A, D and E, baccharinoids B4, B5 and B12, 16-hydroxyverrucarin A, and verrucarins A and J. Lymphocytes from CD-1 mice were cultured with each of the mycotoxins for 48 h to assess cytotoxicity. The maximum effect of various trichothecenes produced on cells occurred at concentrations ranging from 10(-6) to 10(-4) M. Mycotoxins had no effect at concentrations ranging from 10(-12) to 10(-7) M. The mitogenic stimulants concanavalin A, lipopolysaccharide, phytohemagglutinin, and pokeweed mitogen were added to splenic lymphocyte cultures along with varying concentrations of selected mycotoxins. Blastogenesis was inhibited at concentrations 2-5 orders of magnitude lower than those which produced lethality on resting lymphocytes.

Effect of prepuberal castration on porcine bulbospongiosus muscle.

The histochemical profile and ultrastructural properties of the bulbospongiosus muscle (BSM) fibers from 5-6 month old boars and barrows (castrated at 7 days of age), and intact week old piglets were compared. Based on myosin ATPase, preincubated at pH 4.2, BSM of boars contained predominately intermediately staining fibers, whereas BSM of barrows and piglets had a mixture of staining intensities. Fibers from boar BSM stained intensely for SDH, with subsarcolemmal and diffuse location of reaction product. Staining intensity for SDH was variable in BSM from barrows and piglets, with diffuse location of reaction product. The BSM of boars and barrows contained predominately dark fibers when stained for glycogen and phosphorylase, and the fibers were low in stored lipids. While the fibers were smaller in barrow as compared to boar BSM, ultrastructural differences between boar and barrow BSM were not detectable.

Assessment of pentachlorophenol toxicity in newborn calves: clinicopathology and tissue residues.

Newborn Holstein bull calves were fed either analytical pentachlorophenol (aPCP) or technical pentachlorophenol (tPCP) for 6 wk to establish and compare the clinical and pathologic manifestations of toxicity. Four groups of three calves/group were each fed either 1 or 10 mg X (kg body weight)-1 X d-1 of either aPCP or tPCP. A fifth group served as control. Dosages of both PCP preparations were normalized to contain equal concentrations of PCP. Toxic effects were observed only at the 10 mg/kg dose in the tPCP-treated calves. These effects included decreased body weight gain, anorexia, decreased serum protein concentration, elevated serum gamma glutamyl transferase, and decreased triiodothyronine (T3) and thyroxine (T4) concentrations. Histologic lesions included cortical atrophy in the thymus and squamous metaplasia and hyperkeratous changes in the Meibomian gland of the eyelid. Thyroid function, which was assessed in vivo by measuring the rate of T3 and T4 production over 4 h after thyrotropin-releasing hormone (TRH)-challenge, was not impaired suggesting an extrathyroidal site of toxic action. Although serum chemistry indicators were suggestive of hepatic injury there were no discernable lesions. Organ weight analyses were inconclusive but there was a tendency toward enlargement of liver, kidneys and thyroid and decreased weight of lungs, spleen and thymus. A toxic effect clearly related to PCP and not its contaminants was depressed active transport of p-aminohippurate measured in kidney slices in vitro. Steady state concentrations of PCP in serum were about 40 and 90 ppm for the 1 and 10 mg/kg groups, respectively. Concentrations of PCP among the major organs were comparable.

Anatomic, physiologic, and pharmacologic properties of the feline bulbospongiosus muscle.

Histologic examination was used to determine the anatomic organization of the feline bulbospongiosus muscle (BSM). Bilaterally symmetrical cranial, middle, and caudal muscle fiber groups were identified. The caudal group was the largest of the 3 groups. Mean cross-sectional diameter of nonfixed BSM fibers was 79 micron. Muscle spindles were not observed in serial histologic sections. The innervation of the BSM was provided by the deep perineal nerve. Contractile properties of the BSM were determined in vitro, using direct muscle stimulation. Mean time-to-peak tension values were 71.4 ms and 62.7 ms at 30 C and 37 C, respectively, and were longer than that for the feline periurethral striated muscle and external anal sphincter muscle, but slightly shorter than that for the slow-twitch soleus muscle. Stimulation frequency at peak tetanic tension was 40 Hz, twitch-to-tetanus ratio was 0.30, and stimulus interval at peak tension for double response was 44 ms. Electrophysiologic properties examined were resting potentials and miniature end-plate potentials (MEPP). Mean resting potential was 68 mV. The MEPP had low mean amplitude (0.57 mV), fast mean rise time (0.99 ms), and low maximal frequency (0.06 Hz). Only focally recordable MEPP were observed. Pharmacologic properties were investigated by alternating direct and indirect stimulation. The latter was accomplished by electric field stimulation of intramuscular nerve fibers. A high sensitivity of the BSM to d-tubocurarine blockade was present. The results of this study support classification of the BSM as a slow-twitch muscle.

Molecular definition of the bovine granulocytopathy syndrome: identification of deficiency of the Mac-1 (CD11b/CD18) glycoprotein.

Leukocytosis (34,600 WBC/microliter of blood) was detected in an apparently healthy 7-day-old Holstein heifer. Analysis of blood samples obtained over the next 41 days revealed chronic progressive neutrophilia, which peaked at greater than 85% neutrophils and exceeded 100,000 WBC/microliter. In vitro assessment of isolated blood neutrophils obtained from the heifer at 38 and 45 days of age revealed selected functional abnormalities. Endocytosis of immunoglobulin-opsonized Staphylococcus aureus and killing of this test organism by the calf's neutrophils were significantly diminished, as were phagocytosis-associated superoxide generation, chemiluminescence activity, and myeloperoxidase-catalyzed iodination. Diminished H2O2 elaboration by the calf's neutrophils was evident during ingestion of opsonized zymosan or on exposure to phorbol myristate acetate. Extracellular release (secretion) of elastase during ingestion of zymosan was also diminished, although total cell content of elastase was normal, compared with that of neutrophils from age-matched calves, and granular or other morphologic abnormalities of the calf's neutrophils were not evident by ultrastructural examination. Abnormalities of random migration were inconsistently detected, and normal or high degree of antibody-dependent cytotoxicity or natural killing by the calf's neutrophils was observed. Similar in vitro assessment of neutrophils obtained from the calf's dam revealed no functional abnormalities. The calf died at 48 days of age, with persistent fever and chronic diarrhea, despite administration of antibiotics. Histologic examination at necropsy revealed large numbers of intravascular neutrophils in most tissues, including massive neutrophil sequestration in spleen. However, a striking lack of extravascular neutrophils was evident in inflamed submucosa adjacent to intestinal ulcers heavily contaminated with enteric microorganisms. Bone marrow examination revealed diffuse myeloid hyperplasia, but no other abnormalities.

Juvenile cellulitis in dogs: 15 cases (1979-1988).

The records of 15 dogs diagnosed as having juvenile cellulitis (juvenile pyoderma, puppy strangles) were evaluated for clinical, laboratory, and therapeutic results. Mandibular lymphadenopathy was observed in 14 dogs, and was not associated with skin lesions in 5 dogs. Edema, pustules, papules, or crusts were noticed periorally, periocularly, on the chin or muzzle, or in the ears of those dogs with skin lesions. Eight dogs were lethargic; fever and anorexia were inconsistent findings. Four dogs had signs of pain on manipulation of their joints. Complete blood counts revealed leukocytosis with neutrophilia in 4 dogs, and normocytic, normochromic anemia in 6 dogs. Three dogs had suppurative lymphadenitis with many neutrophils. Cytology of the aspirate of pustules or abscesses in 6 dogs revealed many neutrophils without bacteria. Coagulase-positive Staphylococcus spp were isolated from draining lesions in 2 dogs. Intact abscesses and lymph nodes were negative for bacterial growth in 4 dogs. Three of these dogs were being administered antibiotics at the time of bacterial culturing. Cytology of the aspirates of joints in 3 of the 4 dogs with joint pain revealed suppurative arthritis with no bacteria, and the aspirates were negative for bacterial growth on culturing, although all 3 dogs were being administered antibiotics at the time of culturing. Of 12 dogs initially treated with antibiotics, only 4 (33%) responded favorably; the other 8 dogs were then given antibiotics and corticosteroids. Three dogs were initially given antibiotics and corticosteroids. All dogs treated concurrently with antibiotics and corticosteroids responded favorably. One of these dogs had a relapse after treatment was discontinued. The concurrent arthritis in 4 of the dogs resolved with treatment of the juvenile cellulitis and did not redevelop once the medication was discontinued. Concurrent treatment with antibiotics (cephalosporins) and prednisone (2.2 mg/kg of body weight/day) was the most consistently effective treatment in the dogs in this study.