Objective— To evaluate effects of polymyxin B sulfate
(PMB) on response of horses to endotoxin, using
an ex vivo model.
Animals—8 healthy horses.
Procedure—In a crossover design, 3 doses of PMB
(100, 1,000, and 10,000 U/kg of body weight) and
physiologic saline solution (control) were evaluated.
Prior to and for 24 hours after administration of PMB,
blood samples were collected into heparinized tubes
for use in 2 assays. For the endotoxin-induced tumor
necrosis factor (TNF) assay, blood samples were incubated
(37 C for 4 h) with 1 ng of Escherichia coli or
Salmonella Typhimurium endotoxin/ml of blood.
Plasma was harvested and assayed. For the residual
endotoxin activity assay, plasma was collected into
sterile endotoxin-free borosilicate tubes, diluted 1:10
with pyrogen-free water, and incubated for 10 minutes
at 70 C. Escherichia coli endotoxin (0.1 or 1
ng/ml of plasma) was added to the thawed samples
prior to performing the limulus ameobocyte lysate
assay. Serum creatinine concentrations were monitored
for 1 week.
Results—Compared with baseline values, PMB
caused a significant dose- and time- dependent
decrease in endotoxin-induced TNF activity.
Compared with baseline values, residual endotoxin
activity was significantly reduced after administration
of 10,000 U of PMB/kg. Compared with baseline values,
1,000 and 5,000 U of PMB/kg should inhibit 75%
of endotoxin-induced TNF activity for 3 and 12 hours,
respectively. Serum creatinine concentrations
remained within the reference range.
Conclusion and Clinical Relevance—Results of the
study suggest that PMB is a safe, effective inhibitor of
endotoxin-induced inflammation in healthy horses.
( Am J Vet Res 2001; 62:72–76)
Objective—To evaluate the effects of a standardized exercise test to exhaustion in horses on leukocyte function ex vivo.
Animals—6 Thoroughbred geldings.
Procedures—Blood samples were obtained from each horse before exercise; at exhaustion (termed failure); and at 2, 6, 24, 48, and 72 hours after exercise to evaluate hematologic changes, rate of leukocyte apoptosis, and leukocyte production of reactive oxygen species (ROS) ex vivo. To assess leukocyte function, leukocyte ROS production in response to stimulation with lipopolysaccharide, peptidoglycan, zymosan, and phorbol myristate acetate was evaluated. Apoptosis was evaluated via assessment of caspase activity in leukocyte lysates.
Results—In response to lipopolysaccharide, production of ROS by leukocytes was significantly increased at 2 hours and remained increased (albeit not significantly) at 6 hours after exercise, compared with the preexercise value. In the absence of any stimulus, leukocyte ROS production was significantly increased at 6 and 24 hours after exercise. In contrast, ROS production in response to phorbol myristate acetate was significantly decreased at 6, 24, and 72 hours after exercise. Leukocyte ROS production induced by zymosan or peptidoglycan was not altered by exercise. Leukocytosis was evident for 24 hours after exercise, and neutrophilia was detected during the first 6 hours. A significant increase in the rate of leukocyte apoptosis was detected at failure and 72 hours after exercise.
Conclusions and Clinical Relevance—Results indicated that strenuous exercise undertaken by horses causes alterations in innate immune system functions, some of which persist for as long as 72 hours after exercise.
Objective—To determine which antimicrobials that
are used to treat neonatal foals with septicemia attributable
to Escherichia coli will minimize endotoxin
release from bacteria and subsequent activity of
inflammatory mediators while maintaining bactericidal
Sample Population—Blood samples from 10 healthy
Procedure—Escherichia coli isolates A and B were
isolated from 2 septicemic foals, and minimal
inhibitory concentrations (MIC) were determined for
9 antimicrobials. Five of these antimicrobials were
tested in vitro at 2 and 20 times their respective
MIC. Whole blood or mononuclear cells grown in tissue-
culture media were incubated with 105 colonyforming
units of E coli and each antimicrobial or
saline (0.9% NaCl) solution. After 6 hours, number
of viable bacteria remaining was determined, and
supernatant was tested for endotoxin and tumor
Results—Testing in whole blood was compromised
by bactericidal effects of the blood itself. In mononuclear
cell suspensions, each antimicrobial significantly
reduced the number of viable bacteria to low or undetectable
amounts. Antimicrobials did not differ significantly
in efficacy of bacterial killing. Amikacin used
alone or in combination with ampicillin resulted in significantly
less endotoxin activity than did ampicillin,
imipenem, or ceftiofur alone. There was a correlation
between TNF-α and endotoxin activity.
Conclusions and Clinical Relevance—Aminoglycosides
appear less likely to induce endotoxemia
and TNF-α synthesis during bactericidal treatment of E
coli septicemia, compared with β-lactam antimicrobials.
Use of ampicillin, imipenem, or ceftiofur in the
treatment of septicemic neonatal foals should be
accompanied by appropriate treatment for endotoxemia.
(Am J Vet Res 2002;63:660–668)