Journal of Applied Physiology Email Content Delivery
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

J Appl Physiol 103: 948-958, 2007. First published June 21, 2007; doi:10.1152/japplphysiol.00606.2006
8750-7587/07 $8.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF) Free
Right arrow All Versions of this Article:
103/3/948    most recent
00606.2006v1
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Horton, J. W.
Right arrow Articles by Minei, J. P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Horton, J. W.
Right arrow Articles by Minei, J. P.

Bactericidal/permeability increasing protein attenuates the myocardial inflammation/dysfunction that occurs with burn complicated by subsequent infection

Jureta W. Horton, David L. Maass, D. Jean White, and Joseph P. Minei

Department of Surgery, University of Texas Southwestern Medical Center, Dallas, Texas

Submitted 31 May 2006 ; accepted in final form 18 June 2007


   ABSTRACT
TOP
 ABSTRACT
METHODS AND MATERIALS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Intubation and mechanical ventilation after burn contribute to pneumonia-related infection. Although postburn presence or absence of endotoxin has been described, inactivation of Toll-like receptor 4 signaling has been shown to improve postburn organ function, suggesting that LPS participates in burn-related susceptibility to infection. We hypothesized that bactericidal/permeability-increasing protein (rBPI) given postburn would attenuate myocardial inflammation/dysfunction associated with postburn septic challenge given 7 days postburn. Rats were given burn over 40% total body surface area, lactated Ringer 4 ml·kg–1·% burn–1; burns received either vehicle or rBPI, 1 mg·kg–1·h–1 for 48 h postburn. Postburn day 7, subgroups of burns and shams were given intratracheal Klebsiella pneumoniae, 4 x 106 CFU to produce burn complicated by sepsis; additional sham and burn subgroups received intratracheal vehicle to produce sham sepsis. Vehicle-treated groups: 1) sham burn + sham sepsis 2) sham burn + sepsis, 3) burn + sham sepsis, 4) burn + sepsis. rBPI-treated groups: 5) sham burn + sham sepsis, 6) sham burn + sepsis, 7) burn + sham sepsis, 8) burn + sepsis. Cardiomyocyte cytokine secretion and myocardial function were studied 24 h after septic challenge, postburn day 8. Pneumonia-related infection 8 days after vehicle-treated burn produced myocyte cytokine secretion (pg/ml), indicated by increased myocyte TNF-{alpha}, 549 ± 46; IL-1beta, 50 ± 8; IL-6, 286 ± 3 levels compared with levels in sham myocytes (TNF-{alpha}, 88 ± 11; IL-1beta, 7 ± 1; IL-6, 74 ± 10; P < 0.05). Contractile dysfunction was evident from lower left ventricular pressure ±dP/dt values in this group compared with sham. rBPI attenuated myocyte cytokine responses to septic challenge and improved contractile function, suggesting that burn-related mobilization of microbial-like products contribute to postburn susceptibility to infection.

TNF-{alpha}; IL-1beta; IL-6 cytokines; left ventricular function; primary cardiomyocytes; intratracheal Klebsiella pneumoniae challenge

ALTHOUGH THE MOLECULAR BASIS of burn-related cardiac dysfunction is complex and not completely understood, numerous studies have implicated proinflammatory mediators such TNF-{alpha}, IL-1beta, and IL-6 as well as oxygen-derived free radicals and leukocyte-derived products as mediators of postburn cardiac contraction and relaxation deficits (14, 23, 25, 26, 37, 39). Recent studies have suggested that signaling through the Toll-IL-1 pathway, a critical component of mammalian responses to endotoxin, plays a pivotal role in burn-related myocardial inflammatory cytokine responses (38) as well as cardiac contractile dysfunction (47). In this regard, the use of mice expressing either a mutant form of or no Toll-like receptor 4 (TLR4) was shown to abrogate postburn inflammatory cytokine responses, suggesting that signaling through TLR4 participates in burn-related complications such as downstream organ inflammation, dysfunction, and failure (38, 47).

Bactericidal/permeability-increasing protein (BPI) is a 55-kDa neutrophil-derived polypeptide that binds and neutralizes endotoxin activity (3, 20, 28, 29, 35, 48). Recombinant BPI (rBPI) has been applied as an immunotherapeutic agent in a number of injury and disease states including hindlimb ischemia-reperfusion injury (18), severe meningococcemia (13), bile duct ligation (31), endotoxemia (3, 9), and liver resection (50). Intravenous administration of rBPI in adult rats was shown to provide significant protection against the myocardial systolic and diastolic dysfunction that occurred 24 h postburn (47). Although investigators have shown low levels of endotoxin in the systemic circulation after burn injury, several investigators have proposed that in vivo injury activates a mechanism that increases sensitivity to even minute amounts of endotoxin, rendering the injured subject susceptible to infectious complications and development of multiple organ dysfunction (32, 41–43, 46). This hypothesis has been supported by the finding that burn trauma increased plasma levels of lipopolysaccharide binding protein (LBP)/CD-14, specific molecules that recognize and bind low levels of LPS (12, 44, 53). More recent studies using a Spectral Diagnostics endotoxin assay (Spectral Diagnostics, Toronto, ON, Canada) have shown a significant rise in circulating (blood) endotoxin levels after burn over greater than 15% total body surface area (TBSA), suggesting a potential role for this mediator in postburn signaling cascades (4). This present study examined the hypothesis that administration of rBPI after experimental burn over 40% TBSA would reduce the compartmental (myocardial) as well as systemic (plasma) proinflammatory cytokine response and myocardial contractile dysfunction that occur with pneumonia infectious challenge given 7 days after a previous burn injury (22, 49).


   METHODS AND MATERIALS
TOP
 ABSTRACT
 METHODS AND MATERIALS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Experimental model.   Adult Sprague-Dawley rats weighing 325–350 g were obtained from Harlan Laboratories (Houston, TX) and housed in the animal care facility. Commercial rat chow and tap water were available ad libitum and rats were maintained at a constant temperature with a 12:12-h light-dark cycle. All work was performed under a protocol that was approved by The University of Texas Southwestern Medical Center Institutional Animal Care and Research Advisory Committee, and the work conformed to all guidelines outlined in the Guide for the Care and Use of Laboratory Animals published by the American Physiological Society.

Catheter placement and burn procedure.   Rats were lightly anesthetized with isoflurane 18–20 h before the burn experiment, and body hair on the side, back, and neck was closely clipped. The neck region was treated with a surgical scrub, the left carotid artery was exposed, and a polyethylene catheter (PE-50) was inserted into the artery and advanced retrogradely to the level of the aortic arch. In addition, a polyethylene catheter (PE-50) placed in the right external jugular vein was used to administer fluids and drugs. All catheters were filled with heparinized saline, exteriorized, and secured at the nape of the neck. Eighteen hours after catheter placement, animals were deeply anesthetized (isoflurane) and secured in a constructed template device, and the surface of the skin exposed through the aperture in the template was immersed in 100°C water for 10 s on the back and upper sides. Use of the template produced well circumscribed full-thickness dermal burns over 40% of the TBSA (1). Exposure to this water temperature in adult rats destroys all underlying nerves and avoids injury to underlying organs. Sham burn rats were subjected to identical preparation, except that they were immersed in room temperature water to serve as controls. Immediately after immersion, rats were dried, returned to individual cages, and each external jugular catheter was connected to a swivel device (BSP99 syringe pump, Braintree Scientific, Braintree, MA) for fluid administration. Body temperature was measured with a rectal temperature probe (YSL-Tele thermometer, model 44TA, Yellow Springs Laboratory, Herefordshire, UK), and respiratory rate was monitored by counting respiratory movement. Systemic blood pressure was measured intermittently during the first 24 h postburn to determine adequacy of fluid resuscitation by using a Gould-Statham pressure transducer (model IDP23, Gould Instruments, Oxnard, CA) connected to a Grass medical recorder (model 7D polygraph, Grass Instruments, Quincy, MA). A Grass tachycardiograph (model 7P4F) was used to monitor heart rate. A Grass Poly VIEW data-acquisition system was used to convert acquired data into digital form. Blood pressure and heart rate were also measured 24 h after septic challenge (postburn day 8) in all experimental groups (see Table 2), and a blood sample was collected to measure acid-base balance, Na+/Ca2+ levels, and plasma cytokines.


View this table:
[in this window]
[in a new window]

  		Table 2. Hemodynamic and metabolic responses measured 24 h after septic challenge (postburn day 8)

 
Lactated Ringer resuscitation after burn injury.   Fluid resuscitation consisted of lactated Ringer solution, 4 ml·kg–1·% burn–1 with one-half of the calculated volume given intravenously during the first 8 h postburn and the remaining volume given over the next 16 h postburn. The total volume of Ringer given over the first 24 h postburn was 50–56 ml. Buprenorphine (0.5 mg/kg) was given every 12 h for the first 3 days of the postburn period. Burned rats did not display discomfort or pain, moved freely about the cage, and consumed food and water within 15 min after recovering from isoflurane anesthesia. In the sham burn animals, the external jugular vein was cannulated and lactated Ringer solution was given to maintain catheter patency (0.2 ml/kg h); sham burns also received identical regimens of analgesics (buprenorphine) throughout the study period.

Experimental groups.   Adult rats were randomly divided to receive either burn injury over 40% TBSA + fluid resuscitation or sham burn injury (handled in an identical manner but immersed in room temperature water). Minutes (5–10) after burn injury (or sham burn), subgroups of sham burn and burn rats were divided to receive either vehicle (0.175 ml/h of 0.005 M sodium citrate/0.15 M sodium chloride, groups 1, 2, 3, and 4) or rBPI (prepared in a solution of 0.005 M sodium citrate/0.15 M sodium chloride) as a continuous infusion given as 1 mg·kg–1·h–1 for 48 h (groups 5, 6, 7, and 8). A total of eight experimental groups were included and are described in Fig. 1. These included group 1, sham burn, intravenous vehicle, and no rBPI therapy followed on post-sham burn day 7 by sham septic challenge; group 2, sham burn, no rBPI therapy followed on post-sham burn day 7 by intratracheal Klebsiella pneumoniae to produce septic challenge in the absence of previous burn injury; group 3, burn over 40% TBSA given intravenous vehicle followed on postburn day 7 by intratracheal vehicle to produce burn + sham sepsis; in group 4, burn over 40% TBSA + intravenous vehicle was followed on postburn day 7 by intratracheal Klebsiella pneumoniae challenge to produce septic challenge complicated by previous burn injury. In groups 5–8, all animals received rBPI therapy to produce the following groups: group 5, sham burn + rBPI therapy for 48 h after sham burn followed on postburn day 7 by administration of intratracheal vehicle to produce sham burn + rBPI + sham septic challenge; group 6, sham burn + rBPI + septic challenge; group 7, burn + rBPI + sham septic challenge; and group 8, burn + rBPI + intratracheal Klebsiella pneumoniae sepsis. All rats were studied with regard to cytokine secretion, cardiac function, myocyte secretion, and calcium handling 24 h after the administration of intratracheal vehicle or intratracheal Klebsiella pneumoniae challenge and on postburn day 8 (Fig. 1).


Figure 1
View larger version (17K):
[in this window]
[in a new window]

  		Fig. 1. Experimental groups. rBPI, recombinant bactericidal/permeability-increasing protein.

 
The presence of infection in our rat model was determined from the presence of bacteremia (blood samples for bacterial culture were collected 24 h after septic challenge and blood was streaked onto blood agar plats, TSA II 5% sheep blood) and changes in body temperature as well as hemodynamic (arterial blood pressure and heart rate) and metabolic alterations (lactate, base excess). Rats were used to examine either myocardial function in vitro (Langendorff, n = 7–10 rats/group) or to prepare cardiac myocytes (n = 4–5 rats/group) to measure myocyte cytokine secretion and myocyte sodium and calcium handling (a total of 11–15 rats/experimental group).

Preparation of bacterial inoculum.   Klebsiella pneumoniae type 3 was obtained from the American Type Culture Collection (ATCC 6303, Rockville, MD) in lyophilized form. Bacteria were reconstituted and then passed through the cerebrospinal fluid of a rabbit to increase virulence; aliquots were prepared and stored at –80°C. Before each experiment, individual aliquots were thawed, inoculated into Muller Hinton broth with supplement C (Difco, Kansas City, MO), and incubated overnight at 37°C in the presence of 5% CO2. The broth was then centrifuged and the resultant pellet was washed twice with sterile endotoxin-free PBS to remove any impurities adherent to the bacteria. The bacteria were then resuspended in sterile endotoxin-free PBS, agitated, and then drawn up into sterile tuberculin syringes in 0.4 ml aliquots. Bacterial colony-forming units (CFU) were determined by plating 100 µl of the bacterial suspension onto blood agar plates in serial dilutions and incubating the plates overnight at 37°C. The number of viable bacterial inoculated into animals in either the pneumonia alone or in the burn + pneumonia groups was ~4 x 106 CFU.

Induction of sepsis.   Rats were anesthetized with isoflurane and placed in a supine position. The area over the trachea was prepped with a surgical scrub (povidine-iodine, Betadine). A midline incision was made over the trachea; the trachea was identified and isolated via blunt dissection. A 0.4-ml aliquot of either sterile endotoxin-free PBS or bacterial suspension (Klebsiella pneumoniae, 4 x 106 CFU) was injected directly into the trachea. After the wound was closed with surgical staples, the animals were placed on a 30° incline to ensure accumulation of the injected fluid into the lungs.

Cardiomyocyte isolation.   Adult rat cardiomyocytes were isolated as described previously (6). Briefly, the heart was removed and placed in a petri dish containing room temperature heart medium [113 mM NaCl; 4.7 mM KCl; 0.6 mM KH2PO4; 0.6 mM Na2HPO4; 1.2 mM MgSO4; 12 mM NaHCO3; 10 mM KHCO3; 20 mM D-glucose; 0.5x minimum essential medium (MEM) amino acids (50x, GIBCO/BRL 11130-051); 10 mM HEPES; 30 mM taurine; 2.0 mM carnitine; and 2.0 mM creatine]. Hearts were cannulated via the aorta and perfused with heart medium at a rate of 12 ml/min for a total of 5 min in a nonrecirculating mode. Enzymatic digestion was initiated by perfusing the heart with digestion solution that contained 34.5 ml of heart medium described above plus 50 mg of collagenase II (Worthington 4177, lot no. MOB3771), 50 mg BSA, fraction V (GIBCO/BRL 11018-025), 0.5 ml trypsin (2.5%, 10x, GIBCO/BRL 15090-046), 15 µM CaCl2. Enzymatic digestion was accomplished by recirculating this solution through the heart at a flow rate of 12 ml/min for 20 min. All solutions perfusing the heart were maintained at a constant temperature of 37°C. At the end of the enzymatic digestion, the ventricles were removed and mechanically disassociated in 6 ml of enzymatic digestion solution containing a 6-ml aliquot of 2x BSA solution (2 mg BSA, fraction V to 100 ml of heart media). After mechanical disassociation with fine forceps, the tissue homogenate was filtered through a mesh filter into a conical tube. The cells adhering to the filter were collected by washing with an additional 10 ml aliquot of 1x BSA solution (100 ml of heart medium described above, and 1 g of BSA, fraction V). Cells were then allowed to pellet in the conical tube for 10 min. The supernatant was removed and the pellet was resuspended in 10 ml of 1x BSA. The cells were washed and pelleted further in BSA buffer with increasing increments of calcium (100, 200, and 500 µM, to a final concentration of 1,000 µM). After the final pelleting step, the supernatant was removed, and the pellet was resuspended in MEM (prepared by adding 10.8 g 1x MEM, Sigma M-1018, 11.9 mM NaHCO3, 10 m HEPES, and 10 ml penicillin-streptomycin, 100x, GIBCO/BRL 1540-122 with 950 ml MilliQ water); total volume was adjusted to 1 liter. At the time of MEM preparation, the medium was bubbled with 95% O2-5% CO2 for 15 min and the pH adjusted to 7.1 with 1 M NaOH. The solution was then filter sterilized and stored at 4°C until use. At the final concentration of calcium, the cardiomyocyte cell number was calculated and myocyte viability was determined (24).

Cardiac myocyte cytokine response.   Myocytes were pipetted into microtiter plates at 5 x 104 cell·ml–1·well–1 (12-well cell culture cluster, Corning, Corning, NY) for 18 h (CO2 incubator at 37°C). Supernatants were collected to measure myocyte-secreted TNF-{alpha}, IL-1beta, IL-6, and IL-10 (rat ELISA, Endogen, Woburn, MA). Cell viability, cardiomyocyte cell number, and nonmyocyte cell number were determined for each preparation; less than 2% of the total cell number in a myocyte preparation was noncardiomyocytes regardless of experimental group (21).

Cardiomyocyte calcium and sodium measurements.   Cardiomyocytes were loaded with the calcium indicator fura 2-AM for 45 min or sodium-binding benzofurzan isophthalate for 1 h at room temperature in the dark. Myocytes were then suspended in 1.0 mM calcium containing MEM, washed to remove extracellular dye, and placed on a glass slide on the stage of a Nikon inverted microscope. The microscope was interfaced with Grooney optics for epi-illumination, a triocular head, phase optics, and x30 phase contrast objective and mechanical stage. The excitation illumination source (300 W compact Xenon arc illuminator) was equipped with a power supply. In addition, this InCyt Im 2 Fluorescence Imaging System (Intracellular Imaging, Cincinnati, OH) included an imaging workstation and Intel Pentium Pro200 MHZ-based personal computer. The computer-controlled filter changer allowed alternation between the 340 and 380 excitation wavelengths. Images were captured by monochrome charge-coupled device camera equipped with a TV relay lens. InCyt Im2 Image software allowed measurement of intracellular calcium and sodium concentrations from the ratio of the two fluorescent signals generated from the two excitation wavelengths (340 nm/380 nm); background was removed by the InCyt IM2 software. The calibration procedure included measuring fluorescence ratio with buffers containing different concentrations of either calcium or sodium. At each wavelength, the fluorescence emissions were collected for 1-min intervals, and the time between data collection was 1–2 min. Since quiescent or noncontracting myocytes were used in these studies, the calcium levels measured reflect diastolic levels (22, 49).

Isolated heart perfusion.   To examine cardiac contraction and relaxation, rats were then anticoagulated 24 h after burn trauma with sodium heparin (200 units, Elkins-Sinn, Cherry Hill, NJ) and cervically dislocated. The heart was rapidly removed and placed in a petri dish containing ice-cold (4°C) Krebs-Henseleit bicarbonate buffered solution (in mM: 118 NaCl, 4.7 KCl, 21 NaHCO3, 1.25 CaCl2, 1.2 MgSO4, 1.2 KH2PO4, and 11 glucose). All solutions were prepared each day with demineralized, deionized water and bubbled with 95% 02-5% CO2 (pH, 7.4; PO2, 550 Torr; PCO2, 38 Torr). A cannula placed in the ascending aorta was connected via glass tubing to a buffer-filled reservoir for perfusion of the coronary circulation at a constant flow rate. Hearts were suspended in a temperature-controlled chamber maintained at 38°C, and a constant-flow pump (Ismatec, model 7335-30, Cole-Parmer Instrument, Chicago, IL) was used to maintain perfusion of the coronary artery (ml/min) by retrograde perfusion of the aortic stump cannula. Coronary perfusion pressure was measured and effluent was collected to confirm coronary flow rate. Contractile function was assessed by measuring intraventricular pressure with a water-filled latex balloon attached to a polyethylene tube and threaded through the apex of the left ventricular (LV) chamber. Peak systolic left ventricular pressure (LVP) was measured with a Statham pressure transducer (model P23ID, Gould Instruments, Oxnard, CA) attached to the balloon cannula, and the rates of LVP rise (+dP/dt) and fall (–dP/dt) were obtained by using an electronic differentiator (model 7P20C, Grass Instruments) and recorded (Grass, model 7DWL8P). LV developed pressure was calculated from peak systolic LV pressure and LV end-diastolic pressure. Data from the Grass recorder was input to a Dell Pentium computer and a Grass PolyVIEW data-acquisition system was used to convert acquired data into digital form. Each heart was first studied at a constant coronary flow rate, constant LV end-diastolic volume, and constant heart rate to collect stabilization data (Table 3). Each heart was then subjected to incremental increases in LV volume (preload), and function was measured at each level of preload. Finally, each heart was again studied as perfusate calcium was incrementally increased, and function was measured at each perfusate calcium concentration. These interventions (increases in preload or perfusate calcium) allowed us to construct LV function curves presented in Figs. 4 and 5.


View this table:
[in this window]
[in a new window]

  		Table 3. Plasma cytokines measured 24 h after septic challenge (postburn day 8)

 

Figure 4
View larger version (20K):
[in this window]
[in a new window]

  		Fig. 4. Left ventricular developed pressure (LVP) and LVP rise and fall (±dP/dt) responses to incremental increases in LV volume (n = 7–9 rats/group). Experimental groups include group 1, sham burn + sham sepsis, no rBPI; group 5, sham burn + rBPI + sham sepsis; group 4, burn injury + septic challenge and no rBPI; and group 8, burn injury + rBPI therapy + septic challenge. Kleb, Klebsiella pneumoniae. Ventricular function was studied 24 h after septic or sham septic challenge. All values are means ± SE. *Significant difference from sham at P < 0.05 (ANOVA, Student-Newman-Keuls). +Significant rBPI related effects, for example, group 8 vs. group 4 at P < 0.05.

 

Figure 5
View larger version (17K):
[in this window]
[in a new window]

  		Fig. 5. LVP and ±dP/dt responses to incremental increases in perfusate calcium (n = 7–9 rats/group). Experimental groups include group 1, sham burn + sham sepsis, no rBPI; group 5, sham burn + rBPI + sham sepsis; group 4, burn injury + septic challenge and no rBPI; and group 8, burn injury + rBPI therapy + septic challenge. Ventricular function was studied 24 h after septic or sham septic challenge. All values are means ± SE. *Significant difference from sham at P < 0.05 (ANOVA, Student-Newman-Keuls). +Significant rBPI related effects, for example group 8 vs. group 4 at P < 0.05.

 
Statistical analysis.   All values are expressed as means ± SE. ANOVA was used to assess an overall difference among the groups. Levene's test for equality of variance was used to suggest the multiple comparison procedure to be used. If equality of variance was suggested, multiple comparison procedures were performed (Bonferroni); if inequality of variance was suggested, Tamhane multiple comparisons were performed. Probability values <0.05 were considered statistically significant (analysis was performed using SPSS for Windows, version 7.5.1). Mortality rates were evaluated by {chi}2 analysis.


   RESULTS
TOP
 ABSTRACT
 METHODS AND MATERIALS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Evidence of sepsis and mortality.   Bacteremia occurred 24 h after intratracheal administration of Klebsiella pneumoniae as indicated by the positive blood cultures in groups 2, 4, 6, and 8 (Table 1). These findings were paralleled by a significant fall in mean arterial blood pressure and metabolic acidosis (Table 1). There was no significant difference in the number of bacterial CFU either in whole blood or in bronchoalveolar lavage samples among the groups given septic challenge, regardless of rBPI therapy (Table 1). It was of interest that whereas the number of Klebsiella pneumoniae CFU per milliliter of blood was similar in groups 4 and 8 given burn + sepsis, mortality rate 24 h after intratracheal Klebsiella pneumoniae challenge in the absence of postburn rBPI therapy (group 4) was significantly higher (47%) compared with mortality observed after Klebsiella pneumoniae challenge in the presence of postburn rBPI therapy (group 8, mortality 0%, P < 0.05). All animals given burn in the absence of infection (groups 3 and 7) survived the 8-day experimental period (Table 1).


View this table:
[in this window]
[in a new window]

  		Table 1. Evidence of systemic infection and mortality 24 h after septic challenge (postburn day 8)

 
Hemodynamic and metabolic response to burn and sepsis.   Compared with shams (groups 1 and 5), mean arterial blood pressure was lower 24 h after septic challenge complicated by previous burn (groups 4 and 8) regardless of rBPI therapy and despite fluid resuscitation with lactated Ringer solution both after burn injury (Parkland formula) and after septic challenge (Table 2). Compared with shams, hematocrit measured 8 days after burn (groups 3, 4, 7, and 8) was lower, regardless of the presence or absence of postburn infection, likely owing to the hemodilutional effects of aggressive volume resuscitation from burn injury. Compared with values measured in sham burns (group 1 and 5), base excess was altered 8 days after burn injury in the absence of sepsis (groups 3 and 7), after sepsis in the absence of previous burn injury (groups 2 and 6), as well as after burn complicated by septic challenge (groups 4 and 8), regardless of rBPI therapy. Whole blood lactate levels were significantly elevated 8 days after burn in the absence of septic challenge and in the absence of rBPI therapy (group 3) and in burn + sepsis with no rBPI therapy (group 4) and compared with sham burn (group 1, P < 0.05). Administration of rBPI therapy after burn injury attenuated changes in lactate associated with either burn alone (group 7) and compared with group 3 or in burn + septic challenge (group 8) and compared with burn + septic challenge in the absence of rBPI therapy (group 4, P < 0.05). Thus administration of rBPI therapy for 48 h after the initial burn injury and prior to Klebsiella pneumoniae challenge (group 8) improved whole blood lactate compared with values measured in the group given burn injury and no rBPI therapy followed by septic challenge (group 4, P < 0.05).

Systemic and compartmental inflammatory cytokine responses.   In the absence of rBPI therapy, systemic pro- and anti-inflammatory responses were evident 24 h after septic challenge in the absence of previous burn injury (group 2) as well as 8 days after burn injury in the absence of septic challenge (group 3) and in burn injury complicated by septic challenge (group 4) as indicated by the rise in plasma TNF-{alpha}, IL-1beta, IL-6, and IL-10 (Table 3). Administration of rBPI therapy for 48 h after an initial burn injury and in the absence of a subsequent septic challenge (group 7) attenuated the systemic inflammatory response observed 8 days after burn and compared with group 3 (burn, no rBPI, and no septic challenge, P < 0.05). These differences again were indicated by lower plasma TNF-{alpha}, IL-1beta, IL-6, and IL-10 levels measured in group 7 and compared with values measured in group 3, P < 0.05. In addition, rBPI therapy in group 8 (burn complicated by sepsis) reduced plasma cytokine levels compared with values measured in burn + sepsis in the absence of rBPI therapy (group 4, P < 0.05).

Cardiomyocytes isolated from sham burn-sham septic animals (group 1) secreted small amounts of TNF-{alpha} (Fig. 2, top), IL-1beta (Fig. 2, bottom), IL-6 (Fig. 3, top), and IL-10 (Fig. 3, bottom); these pro- and anti-inflammatory responses were likely related to anesthesia and animal handling. Administration of rBPI therapy in sham burn + sham sepsis animals (group 5) attenuated TNF-{alpha}, IL-1beta, IL-6, and IL-10 secretion by cardiomyocytes compared with values measured in group 1 (sham burn + sham sepsis and no rBPI therapy, P < 0.05). Although we have shown previously that cardiomyocytes secrete copious amounts of TNF-{alpha}, IL-1beta, IL-6, and IL-10 over the first 24 h after burn over 40% TBSA, the data from the present study confirm that pro- and anti-inflammatory responses by cardiac myocytes persist 8 days after burn (group 3, burn + sham sepsis) compared with values measured in sham burn + sham sepsis group (group 1, P < 0.05). rBPI therapy in group 7 reduced cardiac myocyte cytokine secretion compared with that measured in group 3, P < 0.05. Septic challenge on postburn day 7 in the absence of previous burn injury (groups 2 and 6) had robust cytokine secretion by cardiac myocytes that was not altered by rBPI therapy. Administration of rBPI therapy for 48 h after the initial burn injury followed by Klebsiella pneumoniae challenge on postburn day 7 (group 8) was associated with significant attenuation of TNF-{alpha}, IL-1beta, IL-6, and IL-10 secretion by cardiac myocytes compared with values measured in burn + septic challenge in the absence of rBPI therapy (group 4, P < 0.05).


Figure 2
View larger version (19K):
[in this window]
[in a new window]

  		Fig. 2. TNF-{alpha} (top) and IL-1beta (bottom) secretion by cardiomyocytes from all experimental groups (n = 5–6 rats/group). All values are means ± SE. *Significant difference from sham at P < 0.05 (ANOVA, Student-Newman-Keuls). {dagger}Significant recombinant bactericidal/permeability-increasing protein (rBPI)-related effects, for example, group 7 vs. group 3, group 8 vs. group 4 at P < 0.05. #Significant difference in burn + sepsis (group 4) vs. burn alone (group 3) or sepsis alone (group 2) at P < 0.05.

 

Figure 3
View larger version (20K):
[in this window]
[in a new window]

  		Fig. 3. IL-6 (top) and IL-10 (bottom) secretion by cardiomyocytes from all experimental groups (n = 5–6 rats/group). All values are means ± SE. *Significant difference from sham at P < 0.05 (ANOVA, Student-Newman-Keuls). {dagger}Significant rBPI related effects, for example, group 7 vs. group 3, group 8 vs. group 4 at P < 0.05. #Significant difference in burn + sepsis (group 4) vs. sepsis alone (group 2) or burn alone (group 3) at P < 0.05.

 
Cardiac contractile function.   Cardiac function was studied in vitro during stabilization of the perfused heart at a constant preload, constant coronary flow rate, and constant heart rate (Table 4). Septic challenge 7 days after sham burn (groups 2 and 6) impaired LV performance in all animals, regardless of rBPI therapy, P < 0.05. LVP and maximal ±dP/dt (±dP/dtmax) measured 8 days after burn injury in the absence of septic challenge (groups 3 and 7) confirmed a significant recovery in myocardial contraction and relaxation performance compared with values previously reported in burns studies 24 h after burn over 40% TBSA and given conventional lactated Ringer fluid resuscitation (21, 23–26). Eight days after burn injury, there were no significant differences in LVP and ±dP/dt in groups 3 and 7 compared with values measured in shams (group 1) regardless of rBPI therapy. Septic challenge on postburn day 7 in rats given no rBPI therapy (group 4) produced significant myocardial contraction and relaxation deficits as indicated by a lower LVP and ±dP/dt in these groups compared with values measured in group 1, P < 0.05. However, LVP and ±dP/dt measured 24 h after septic challenge in the burns given rBPI therapy during the early postburn period (group 8) had significantly improved LV performance (P < 0.05) compared with values measured in rats given burn + septic challenge and no rBPI therapy (group 4, P < 0.05).


View this table:
[in this window]
[in a new window]

  		Table 4. LV function measured on 24 h after septic challenge (postburn day 8)

 
We then determined whether burn complicated by septic challenge altered cardiac responsiveness to inotropic challenge (groups 4 and 8) compared with responses observed in the respective sham burn + sham sepsis groups (groups 1 and 5). As seen in Fig. 4, hearts from animals given sham burn and sham sepsis and no rBPI (group 1) or sham burn + sham sepsis + rBPI (group 5) had similar systolic (+dP/dtmax) and diastolic (–dP/dtmax) responses to increases in LV volume, similar responses to incremental increases in perfusate calcium (Fig. 5), and similar responses to increases in coronary flow rate (Fig. 6) when studied on post-sham burn day 8. Hearts from burns given septic challenge and no rBPI therapy (group 4) had significant decreases in systolic performance as indicated by lower LVP as well as a reduced rate of maximal pressure generation (+dP/dtmax) compared with hearts from sham burned animals (group 1, P < 0.05). These hearts also had impaired diastolic function as indicated by the lower rates of LV relaxation (–dP/dtmax) compared with values measured in sham burns in group 1, P < 0.05. Ventricular defects in this group were evident from reduced responses to either increases in preload (Fig. 4), decreased responses to increases in perfusate calcium (Fig. 5), or decreased responses to incremental increases in coronary flow rate (Fig. 6). Systolic and diastolic function in burns treated initially with rBPI therapy for 48 h followed by septic challenge on postburn day 7 (group 8) were significantly improved over function measured in burns given no rBPI therapy + septic challenge (group 4, P < 0.05, ANOVA and Student-Newman-Keuls). This improved contractile performance was evident from improved LVP and ±dP/dt responses to increases in preload, perfusate calcium challenge, or increases in coronary flow rate.


Figure 6
View larger version (17K):
[in this window]
[in a new window]

  		Fig. 6. LVP and ±dP/dt responses to incremental increases in coronary flow rate (n = 8–9 rats/group). Experimental groups include group 1, sham burn + sham sepsis, no rBPI; group 5, sham burn + rBPI + sham sepsis; group 4, burn injury + septic challenge and no rBPI; and group 8, burn injury + rBPI therapy + septic challenge. Ventricular function was studied 24 h after septic or sham septic challenge. All values are means ± SE. *Significant difference from sham at P < 0.05 (ANOVA, Student-Newman-Keuls). +Significant rBPI related effects, for example, group 8 vs. group 4 at P < 0.05.

 
Cardiomyocyte calcium and sodium responses to burn injury and sepsis.   Since calcium is a vital effector of myocardial contractile dysfunction, we used fluorescent indicators to quantitate myocyte calcium and sodium levels. Septic challenge in the absence of previous burn injury (groups 2 and 6) produced a significant rise in cardiac myocyte calcium (Fig. 7, top) and sodium (Fig. 7, bottom) regardless of rBPI therapy. Burned animals given sham septic challenge in the absence of rBPI therapy and studied on postburn day 8 (group 3) had persistent elevations in cardiomyocyte calcium (Fig. 7, top) and sodium (Fig. 7, bottom). Similarly, septic challenge on postburn day 7 in burns (group 4) produced robust calcium and sodium accumulation by cardiomyocytes. Cardiomyocyte calcium and sodium loading was attenuated by rBPI therapy in burns given no septic challenge (group 7) as well as in rats given burn injury complicated by septic challenge (group 8).


Figure 7
View larger version (21K):
[in this window]
[in a new window]

  		Fig. 7. Cardiomyocyte calcium concentrations ([Ca2+]i, top) and sodium levels ([Na+]i, bottom) in all experimental groups (n = 5–6 rats/group). All values are means ± SE. *Significant difference from sham at P < 0.05 (ANOVA, Student-Newman-Keuls). {dagger}Significant rBPI related effects, for example, group 7 vs. group 3, group 8 vs. group 4 at P < 0.05.

 

   DISCUSSION
TOP
 ABSTRACT
 METHODS AND MATERIALS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
Bacterial sepsis, multiple organ failure, and increased mortality continue to be major problems in burn intensive care units. Loss of gut barrier function and bacteria and bacterial endotoxin translocation are thought to play a significant role in the pathophysiology of postburn sepsis (15, 40) and the presence of endotoxin or bacteria in peripheral organs or in whole blood have been described after major burn (4, 7, 11, 16, 27, 35). Elimination or neutralization of gram-negative bacteria is accomplished by BPI. In this present study, administration of rBPI during the first 48 h after scald burn over 40% TBSA attenuated inflammatory responses to a second insult that was accomplished postburn day 7 (Klebsiella pneumoniae challenge). These data are consistent with a previous report that rBPI exerts beneficial effects by neutralizing endotoxin released during the initial burn injury, as well as inhibiting burn-related induction of nitric oxide, TNF-{alpha}, and IL-6 production by several cell types (8, 48).

In our study, the BPI-related attenuation of inflammatory responses to a second septic challenge was associated with a significant improvement in myocardial contractile function and a reduction in infection-related mortality. Animals given rBPI during the early postburn period had significantly better LVP and ±dP/dt responses to incremental increases in LV volume as well as improved responses to increases in perfusate calcium. Improved organ function after postburn bacteremia in our study was consistent with a report by Giroir and colleagues (13), who described that administration of rBPI reduced clinically significant morbidities and improved outcome of children with meningococcemia. Previous reports have described that rBPI therapy attenuates cardiovascular depression that occurs in endotoxic shock (2, 33, 35, 36). Similarly, administration of recombinant rBPI has been shown to exert protective effects in numerous experimental models including gram-negative sepsis (2, 35), ischemia-reperfusion injury (18), gut mucosal injury (19) bile duct ligation (31), liver resection (5, 50), and hemorrhage associated with trauma (10). The beneficial effects of rBPI have been attributed to blockade of leukosequestration and attenuation of remote tissue injury (18), attenuation of gut mucosal injury that occurs after trauma and burn (19), improved microcirculatory function (5), amelioration of hypercoagulability (51), and inhibition of endotoxin stimulated production of inflammatory cytokines such as TNF-{alpha}, IL-6, and nitric oxide (8, 30, 52).

Although we propose that rBPI therapy exerted protective effects by interrupting the burn-related priming of the innate immune system, we also considered that rBPI may itself blunt the subsequent immune response to infectious challenge and have nothing to do with preventing the initial immune responses after burn. The inclusion of animals given septic challenge in the absence of previous burn injury and treated with either rBPI therapy or given intravenous vehicle addressed this issue. Administration of rBPI therapy for 48 h in the absence of burn injury followed by septic challenge on postburn day 7 (group 6) failed to alter the sepsis-related inflammatory cytokine responses shown to occur with vehicle-treated sepsis (group 2) and failed to improve LV function measured 24 h after septic challenge. Thus rBPI therapy did not protect against subsequent infectious challenge in the absence of an initial priming insult such as thermal injury. These data further support the hypothesis that burn injury primes the innate immune response such that a later pneumonia-related sepsis results in exaggerated proinflammatory cytokine responses and exaggerated myocardial dysfunction. In our study, rBPI administration after burn attenuated that initial burn-related priming of the innate immune response, lessening both the myocardial inflammation and impaired LV performance that were evident after burn complicated by septic challenge in the absence of rBPI intervention (group 8 vs. group 4).

It is clearly recognized that LPS recognition includes LPS-binding protein and CD14 and that LPS/LBP/CD14 signal through the TLR4 pathway, producing downstream phosphorylation of I{kappa}B, NF-{kappa}B nuclear translocation, and inflammatory cytokine secretion. Burn-related signaling through this pathway has been implicated by clinical and experimental studies that describe increased concentrations of LBP and CD14 in severely burned patients (12, 44, 53), burn-related I{kappa}B phosphorylation (47), and NF-{kappa}B nuclear translocation in experimental burn (6). In this regard, Fang and colleagues (12) described that thermal injury upregulated LPS binding protein/CD14 and TNF-{alpha} mRNA expression in several organs, whereas early treatment with rBPI attenuated the burn-related rise in hepatic and pulmonary endotoxin levels and decreased the number of bacteria translocating to mesenteric lymph nodes. Our laboratory's previous studies (47) showed that initiating rBPI within 4 h after burn injury abrogated systolic and diastolic dysfunction in the early postburn period (24 h), producing myocardial contraction and relaxation responses to increases in either LV volume, coronary flow rate, or perfusate calcium concentrations that were similar to responses observed in hearts from unburned animals. These previous studies suggested that endotoxin/CD14/Toll/IL-1 signaling contribute to downstream organ dysfunction after major burn injury.

This present study is the first, to our knowledge, to provide evidence that rBPI therapy in the early postburn period provides significant organ protection and attenuates the systemic and peripheral organ inflammatory cytokine response to a subsequent septic insult (Klebsiella pneumoniae challenge). Although previous studies have described low endotoxin levels in the systemic circulation after burn injury, Fang and colleagues (12) described that less than 0.1 µg per milliliter LPS was required to activate cells. Systemic endotoxin concentrations in septic patients and more recently in burn patients have been reported as high as 100 times this amount (4, 5, 34). It is likely that burn-related loss of gut barrier function and bacterial/endotoxin translocation produce systemic levels of LPS that are sufficient to enhance sensitivity to a second septic insult such as aspiration-related pneumonia.

In summary, gram-negative pneumonia-related septic challenge that occurred 7 days after vehicle-treated burns (no rBPI therapy) produced myocardial cytokine secretion and a rise in systemic cytokine levels, indicating both compartmentalized as well as systemic inflammation. This inflammatory response was paralleled by profound myocardial contraction and relaxation defects. Administration of recombinant rBPI in the immediate postburn period (for 48 h after burn) attenuated proinflammatory responses to a subsequent septic challenge and improved myocardial contractile performance. In addition, mortality rate associated with infection that occurred 8 days after a previous burn injury (47%) was ablated by the administration of rBPI therapy in the early postburn period. These data suggest that burn-related mobilization of microbial-like products contribute to postburn susceptibility to subsequent infectious complications. Therapeutic strategies that target bacterial products during the early postburn period appear to decrease susceptibility to subsequent septic challenge, improving organ function and decreasing mortality with a septic insult after burn injury.


   GRANTS
TOP
 ABSTRACT
 METHODS AND MATERIALS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
This research was funded by National Institute of General Medical Sciences Grant 5 P50 GM21681-40.


   FOOTNOTES
 

Address for reprint requests and other correspondence: J. W. Horton, Dept. of Surgery, Univ. of Texas Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75235-9136 (e-mail: jureta.horton{at}utsouthwestern.edu)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
TOP
 ABSTRACT
 METHODS AND MATERIALS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. Adams HR, Baxter CR, Parker JL, Senning R. Development of acute burn shock in unresuscitated guinea pigs. Circ Shock 8: 613–625, 1981.[Web of Science][Medline]
  2. Ammons WS, Kung AH. Recombinant amino terminal fragment of bactericidal/ permeability-increasing protein on endotoxin shock in conscious rabbits. Shock 2: 324–331, 1994.[Web of Science][Medline]
  3. Arditi M, Zhou J, Huang SH, Luckett PM, Marra MN, Kim KS. Bactericidal/permeability-increasing protein protects vascular endothelial cells from lipopolysaccharide-induced activation and injury. Infect Immun 62: 3930–3936, 1994.[Abstract/Free Full Text]
  4. Barber RC, Foster D, Bender J, Omer-Abdalla A, Lemaire S, Burris A, Purdue GF, Horton JW. Utility of the endotoxin activity assay (EAA) to assess endotoxemia in burn patients. 7th World Congress on Trauma, Shock, Inflammation and Sepsis, Munich, Germany, 2007.
  5. Boermeester MA, Houdijk AP, Straatsburg IH, van Noorden CJ, van Leeuwen PA. Organ blood flow after partial hepatectomy in rats: modification by endotoxin-neutralizing bactericidal/permeability-increasing protein (rBPI23). J Hepatol 31: 905–912, 1999.[CrossRef][Web of Science][Medline]
  6. Carlson D, White J, Maass DL, Nguyen RC, Giroir BP, Horton JW. I{kappa}B overexpression in cardiomyocytes prevents NF-{kappa}B activation and provides cardioprotection in trauma. Am J Physiol Heart Circ Physiol 284: H804–H814, 2003.[Abstract/Free Full Text]
  7. Chung DH, Herndon DN. Multiple converging mechanisms for postburn intestinal barrier dysfunction. Crit Care Med 32: 1803–1804, 2004.[CrossRef][Web of Science][Medline]
  8. Corradin SB, Heumann D, Gallay P, Smith J, Mauel J, Glauser MP. Bactericidal/permeability-increasing protein inhibits induction of macrophage nitric oxide production by lipopolysaccharide. J Infect Dis 169: 105–111, 1994.[Web of Science][Medline]
  9. De Winter RJ, von der Mohlen MA, Van Lieshout JJ, Wedel N, Nelson B, Friedmann N, Delemarre BJ, Van Deventer SJ. Recombinant endotoxin-binding protein (rBPI123) attenuates endotoxin-induced circulatory changes in humans. J Inflamm 45: 193–206, 1995.[Web of Science][Medline]
  10. Demetriades D, Smith JS, Jacobson LE, Moncure M, Minei J, Nelson BJ, Scannon PJ. Bactericidal/permeability-increasing protein (rBPI21) in patients with hemorrhage due to trauma: results of a multicenter phase II clinical trial. rBPI21 acute hemorrhagic trauma study group. J Trauma 46: 667–677, 1999.[Web of Science][Medline]
  11. Demling RH. The clinical relevance of defining the mechanism for altered gut permeability in a "two-hit" model of injury and infection. Crit Care Med 32: 2356–2357, 2004.[Web of Science][Medline]
  12. Fang WH, Yao YM, Shi ZG, Yu Y, Wu Y, Lu LR, Sheng ZY. Effects of recombinant bactericidal/permeability-increasing protein on endotoxin translocation and lipopolysaccharide-binding protein/CD14 expression in rats after thermal injury. Crit Care Med 29: 1452–1459, 2001.[CrossRef][Web of Science][Medline]
  13. Giroir BP, Scannon PJ, Levin M. Bactericidal/permeability-increasing protein-lessons learned from the phase III, randomized, clinical trial of rBPI21 for adjunctive treatment of children with severe meningococcemia. Crit Care Med 29: S130–S135, 2001.[CrossRef][Web of Science][Medline]
  14. Giroir BP. Bactericidal/permeability increasing protein: clinical development and potential pediatric applications. Pediatr Infect Dis J 20: 885–887, 2001.[CrossRef][Web of Science][Medline]
  15. Gosain A, Gamelli RL. Role of the gastrointestinal tract in burn sepsis. J Burn Care Rehabil 26: 85–91, 2005.[CrossRef][Web of Science][Medline]
  16. Han TH, Lee SY, Kwon JE, Kwak IS, Kim KM. The limited immunomodulatory effects of escharectomy on the kinetics of endotoxin, cytokines, and adhesion molecules in major burns. Mediators Inflamm 13: 241–246, 2004.[CrossRef][Web of Science][Medline]
  17. Hansbrough J, Tenenhaus M, Wilstrom T, Braide M, Rennekampff OH, Kiessig V, Bjursten JM. Effects of recombinant bactericidal/permeability-increasing protein (rBPI123) on neutrophil activity in burned rats. J Trauma 40: 886–892, 1996.[Web of Science][Medline]
  18. Harkin DW, Barros D'Sa AA, McCallion K, Hoper M, Halliday MI, Campbell FC. Bactericidal/permeability-increasing protein attenuated systemic inflammation and acute lung injury in porcine lower limb ischemia-reperfusion injury. Ann Surg 234: 233–244, 2001.[CrossRef][Web of Science][Medline]
  19. Harkin DW, D'Sa AA, Yassin MM, Hoper M, Halliday MI. Gut mucosal injury is attenuated by recombinant bactericidal/permeability-increasing protein in hind limb ischemia-reperfusion injury. Ann Vasc Surg 15: 326–331, 2001.[CrossRef][Web of Science][Medline]
  20. Horwitz AH, Ammons WS, Bauer RJ, Dedrick R, Nadel R, Williams RE, Liu PS. rBPI(10-193) is secreted by CHO cells and retains the activity of rBPI21. J Endotox Res 10: 97–106, 2004.[CrossRef][Web of Science]
  21. Horton JW, Maass DL, White DJ, Sanders B. Hypertonic saline dextran suppresses burn-related cytokine synthesis by cardiomyocytes. Am J Physiol Heart Circ Physiol 280: H1591–H1601, 2001.[Abstract/Free Full Text]
  22. Horton JW, Maass DL, White DJ, Sanders B. Myocardial inflammatory responses to sepsis complicated by previous burn injury. Surg Infect (Larchmt) 4: 363–378, 2003.[CrossRef][Medline]
  23. Horton JW, Mileski WJ, White DJ, Lipsky P. Monoclonal antibody to intercellular adhesion molecule-1 reduces cardiac contractile dysfunction after burn injury in rabbits. J Surg Res 64: 49–56, 1996.[CrossRef][Web of Science][Medline]
  24. Horton JW, White DJ, Maass DL. Gender-related differences in myocardial inflammatory and contractile responses to major burn trauma. Am J Physiol Heart Circ Physiol 286: H202–H213, 2004.[Abstract/Free Full Text]
  25. Horton JW, White DJ. Role of xanthine oxidase and leukocytes in postburn cardiac dysfunction. J Am Coll Surg 181: 129–137, 1995.[Web of Science][Medline]
  26. Horton JW, White DJ. U-74,500A inhibition of burn-induced cardiac dysfunction. J Surg Res 52: 251–257, 1992.[CrossRef][Web of Science][Medline]
  27. Horton JW. Bacterial translocation after burn injury: the contribution of ischemia and permeability changes. Shock 4: 286–290, 1994.
  28. Hubacek JA, Buchler C, Aslanidis C, Schmitz G. The genomic organization of the genes for human lipopolysaccharide binding protein (LBP) and bactericidal permeability increasing protein (BPI) is highly conserved. Biochem Biophys Res Commun 236: 427–430, 1997.[CrossRef][Web of Science][Medline]
  29. Jellema WT, Veerman DP, DeWinter RJ, Wesseling KH, Van Deventer SJH, Hack CE, Van Lieshout JJ. In vivo interaction of endotoxin and recombinant bactericidal/permeability-increasing protein (rBPI23): hemodynamic effects in a human endotoxemia model. J Lab Clin Med 140: 228–235, 2002.[CrossRef][Web of Science][Medline]
  30. Jin H, Yang R, Marsters S, Ashkenazi A, Bunting S, Marra MN, Scott RW, Baker JB. Protection against endotoxin shock by bactericidal/permeability-increasing protein in rats. J Clin Invest 95: 1947–1952, 1995.[Web of Science][Medline]
  31. Kimmings AN, Van Deventer SJ, Obertop H, Gouma DJ. Treatment with recombinant bactericidal/permeability-increasing protein to prevent endotoxin-induced mortality in bile duct-ligated rats. J Am Coll Surg 189: 374–379, 1999.[CrossRef][Web of Science][Medline]
  32. Koike K, Moore FA, Moore EE, Pogetti RS, Tuder RM, Banerjee A. Endotoxin after gut ischemia/reperfusion causes irreversible lung injury. J Surg Res 52: 656–662, 1992.[Web of Science][Medline]
  33. Koyama S, Shibamoto T, Ammons WS, Saeki Y. rBPI23 attenuates endotoxin-induced cardiovascular depression in awake rabbits. Shock 4: 74–78, 1995.[Web of Science][Medline]
  34. Lee JD, Kato K, Tobias PS, Kirkland TN, Ulevitch RJ. Transfection of CD14 in 70Z/3 cells dramatically enhances the sensitivity to complexes of lipopolysaccharide (LPS) and LPS binding protein. J Exp Med 175: 1697–1705, 1992.[Abstract/Free Full Text]
  35. Levy P, Elsbach P. Bactericidal/permeability-increasing protein in host defense and its efficacy in the treatment of bacterial sepsis. Curr Infect Dis Rep 3: 407–412, 2001.[Medline]
  36. Lin Y, Ammons WS, Leach WJ, Kung AH. Protective effects of a recombinant N-terminal fragment of bactericidal/permeability increasing protein on endotoxin shock in conscious rabbits. Shock 2: 324–331, 1994.[Web of Science][Medline]
  37. Maass DL, Hybki Peters D, White J, Horton JW. The time course of cardiac NF-{kappa}B activation and TNF-{alpha} secretion by cardiac myocytes after burn injury: contribution to burn-related cardiac contractile dysfunction. Shock 17: 293–299, 2002.[CrossRef][Web of Science][Medline]
  38. Maass DL, White DJ, Thomas JA, Horton JW. TLR4 mutation/deletion prevents postburn myocardial inflammation (Abstract). Shock 21: 53, 2004.
  39. Maass DL, White J, Horton JW. IL-6 and IL-1beta act synergistically with TNF-{alpha} to alter cardiac contractile function after burn trauma. Shock 18: 360–366, 2002.[CrossRef][Web of Science][Medline]
  40. Magnotti LJ, Deitch EA. Burns, bacterial translocation, gut barrier function, and failure. J Burn Care Rehabil 26: 383–391, 2005.[CrossRef][Web of Science][Medline]
  41. Mannick JA, Rodrick ML, Lederer JA. The immunologic response to injury. J Am Coll Surg 193: 237–244, 2001.[CrossRef][Web of Science][Medline]
  42. Mileski WJ, Winn RK, Harlan JM, Rice CL. Sensitivity to endotoxin in rabbits is increased after hemorrhagic shock. J Appl Physiol 73: 1146–1149, 1992.[Abstract/Free Full Text]
  43. Murphy TJ, Paterson HM, Kriynovich S, Zang Y, Kurt-Jones EA, Mannick JA, Lederer JA. Linking the "two-hit" response following injury to enhanced TLR4 reactivity. J Leukoc Biol 77: 16–23, 2005.[Abstract/Free Full Text]
  44. Myc A, Buck J, Gonin J, Reynolds B, Hammerling U, Emanuel D. The level of lipopolysaccharide-binding protein is significantly increased in plasma in patients with the systemic inflammatory response syndrome. Clin Diagn Lab Immunol 4: 113–116, 1997.[Medline]
  45. Peng X, Yan H, You Z, Wang P, Wang S. Effects of enteral supplementation with glutamine granules on intestinal mucosal barrier function in severe burned patients. Burns 30: 135–139, 2004.[CrossRef][Web of Science][Medline]
  46. Schwacha MG, Somers SD. Thermal injury induces macrophage hyperactivity through pertussis toxin-sensitive and -insensitive pathways. Shock 9: 249–255, 1998.[Web of Science][Medline]
  47. Thomas JA, Tsen MF, White DJ, Horton JW. TLR4 inactivation and rBPI(21) block burn-induced myocardial contractile dysfunction. Am J Physiol Heart Circ Physiol 283: H1645–H1655, 2002.[Abstract/Free Full Text]
  48. Weiss J, Elsbach P, Shu C, Castillo J, Grinna L, Horowitz A, Theofan G. Human bactericidal/permeability-increasing protein and a recombinant NH2-terminal fragment cause killing of serum-resistant gram-negative bacteria in whole blood and inhibit tumor necrosis factor release induced by the bacteria. J Clin Invest 90: 1122–1130, 1992.[Web of Science][Medline]
  49. White J, Thomas J, Maass DL, Horton JW. The cardiac effects of burn injury complicated by aspiration-pneumonia-induced sepsis. Am J Physiol Heart Circ Physiol 285: H47–H58, 2003.[Abstract/Free Full Text]
  50. Wiezer MJ, Meijer C, Sietses C, Prins HA, Cuesta MA, Beelen RH, Meijer S, van Leeuwen PA. Bactericidal/permeability-increasing protein preserves leukocyte functions after major liver resection. Ann Surg 232: 208–215, 2000.[CrossRef][Web of Science][Medline]
  51. Yamashita M. Bactericidal/permeability-increasing protein ameliorates hypercoagulability after hemorrhagic shock. Thromb Res 87: 323–329, 1997.[CrossRef][Web of Science][Medline]
  52. Yao YM, Bahrami S, Leichtfried G, Redl H, Schlag G. Pathogenesis of hemorrhage-induced bacteria/endotoxin in translocation in rats. Effects of recombinant bactericidal/permeability-increasing protein. Ann Surg 221: 398–405, 1995.[Web of Science][Medline]
  53. Yao YM, Yu Y, Lu LR. Increased soluble CD14 levels correlate with endotoxemia in critical patients with multiple organ failure. In: The Immune Consequences of Trauma. Shock and Sepsis: Mechanisms and Therapeutic Approaches, edited by Faist E. Bologna, Italy, Monduzzi Editore, 1997, p. 149–155.




This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF) Free
Right arrow All Versions of this Article:
103/3/948    most recent
00606.2006v1
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Horton, J. W.
Right arrow Articles by Minei, J. P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Horton, J. W.
Right arrow Articles by Minei, J. P.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2007 by the American Physiological Society.