| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1 Department of Pediatrics and 2 Department of Surgery The University of Texas Southwestern Medical Center, Dallas, Texas 75390
 |
ABSTRACT |
|---|
 TOP
 ABSTRACT
 INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Thermal trauma is
associated with cardiac myocyte apoptosis in vivo. To determine
whether cardiac myocyte apoptosis could be secondary to
burn-induced cytokines or inflammatory mediators, we investigated the
effects of tumor necrosis factor-
(TNF-) and burn plasma on a
murine cardiac myocyte cell line and primary culture myocytes. HL-1
cells were exposed to plasma isolated from burned or sham rats. Burn,
but not sham plasma, induced significant increases in caspase-3
activity and DNA fragmentation. Similar results were obtained in
primary culture rat myocytes. A dose-dependent increase in caspase-3
activity was observed when HL-1 cells were incubated with increasing
concentrations of TNF-. Even though TNF- increased
apoptosis, enzyme-linked immunosorbent assay detected no
TNF- in burn plasma. Burn plasma also failed to induce TNF- mRNA,
eliminating an autocrine mechanism of TNF- secretion and binding.
Also, treatment of burn plasma containing rhuTNFR:Fc failed to inhibit
apoptosis. To examine the possibility that endotoxin within
burn plasma might account for the apoptotic effect, burn plasma was
preincubated with rBPI21. Caspase-3 activity was reduced to
control levels. These data indicate that burn plasma induces apoptosis in cardiac myocytes via an endotoxin-dependent
mechanism and suggest that systemic inhibition of endotoxin may provide a therapeutic approach for treatment of burn-associated cardiac dysfunction.
tumor necrosis factor-; thermal trauma; caspase-3; enzyme-linked
immunosorbent assay
| |
INTRODUCTION |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
APOPTOSIS is an evolutionarily conserved process that permits the removal of diseased or damaged cells, including those that are infected, malignant, or developmentally redundant. The intrinsic cell-suicide program of apoptosis is characterized by the activation of caspases, which cleave a discrete set of proteins leading to disruption of DNA and nuclear structure followed by cellular disintegration into apoptotic bodies. The packaged remnants in apoptotic bodies are then phagocytosed by macrophages (32).
Although apoptosis is often evolutionarily adaptive, it may also be associated with the progression of human diseases. Cardiac myocyte apoptosis has been implicated in the pathogenesis of heart failure of diverse etiologies, including myocarditis (20), ischemia-reperfusion injury (13), chronic pressure overload (2, 3), congestive heart failure (28), and sepsis (27). Recently, Horton (14) demonstrated that myocyte apoptosis occurred in the ventricular myocardium of rats undergoing severe thermal trauma and was temporally correlated with the development of cardiac depression.
There are several potential explanations for the development of myocyte
apoptosis following thermal trauma. It is possible that a
decrement in coronary perfusion pressure, despite volume resuscitation,
may result in ischemic myocardial damage. Ischemia could be worsened by transient hypoxemia from acute lung injury and
increased lung permeability following burn trauma (19,
31). However, it is also possible that apoptosis is
secondary to cytokines present in the circulation following injury. A
prime candidate is tumor necrosis factor- (TNF-), which may be
increased after thermal trauma and which has been associated with the
disruption of endothelial integrity and the loss of cardiac function
(6, 11, 12). Presumably, TNF- might cause
apoptosis via activation of the intracellular death domain of
TNFR1 with subsequent triggering of the caspase cascade
(18).
The purpose of this study was to determine whether plasma obtained from thermally injured rats could induce apoptosis, and furthermore to identify which factor(s) present in plasma accounts for myocyte apoptosis in this model. The identification of a proapoptotic factor could provide a therapeutic target potentially capable of ameliorating cardiac failure following burn injury.
| |
METHODS |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Animals. Pathogen-free adult male Sprague-Dawley rats (325-350 g) were purchased from Harlan Laboratories, (Houston, TX). All animals were acclimated to their surroundings for 5 days before experimentation. The research protocol was conducted in accordance with the guidelines of the Institutional Review Board for Animal Research at The University of Texas Southwestern Medical Center and within the guidelines of the American Physiological Society and The National Institutes of Health.
Burn procedure and plasma isolation. Rats were deeply anesthetized with methoxyflurane and secured in a constructed template device. They were subjected to full-thickness dermal burns comprising 43 ± 1% of the total body surface area, as previously described (17). Sham-burned rats were subjected to an identical procedure except they were immersed in room temperature water. After immersion, the rats were immediately dried and placed in individual cages to recover from anesthesia. Burned rats did not display discomfort or pain, and all rats consumed food and water within 45 min of the burn procedure.
Plasma was harvested from either burned or sham-burned rats 4 h postinjury. The rats were deeply anesthetized with methoxyflurane, and cardiac puncture was used to collect blood into plasma separator tubes containing lithium heparin (Microtainer). The tubes were then spun at 200 g for 10 min. The plasma was aliquotted and stored at
80°C.
Cardiomyocyte isolation. To examine several aspects of burn-mediated cardiomyocyte dysfunction, hearts were collected at several times postburn; time-matched sham burns were included to provide appropriate controls. All animals received an intraperitoneal injection of heparin (2,000 units) 20-30 min before the animal was euthanized. Hearts were harvested and placed in a petri dish containing room temperature medium [in mM: 113 NaCl, 4.7 KCl, 0.6 KH2PO4, 0.6 Na2HPO4, 1.2 MgSO4, 12 NaHCO3, 10 KHCO3, 20 D-glucose; and 0.5× basal medium Eagle amino acids (40×, GIBCO-BRL 11130-051); 10 mM HEPES; 30 mM taurine; 2.0 mM carnitine; and 2.0 mM creatine, which was bubbled constantly with 95% O2-5% CO2]. The hearts were then cannulated via the aorta and perfused with heart medium at the rate of 12 ml/min for a total of 5 min in a nonrecirculating mode. Enzymatic digestion was initiated by perfusing the heart with a digestion solution; 60 ml of digestion solution was prepared by adding 50 mg of collagenase II (Worthington 4177, Lot No. MOB3771), 50 mg of bovine serum albumin (BSA), fraction V (GIBCO-BRL 11018-025), 0.5 ml trypsin (2.5%, 10×, GIBCO-BRL 15090-046), 7.5 µl CaCl2 (100 mM) to 34.5 ml of heart medium prepared as described above, and 15 ml 2,3-butane dionemonoxime (BDM) stock (40 mM). Enzymatic digestion was accomplished by recirculating this solution through the heart at a flow rate of 12 ml/min for 20 min. The temperature of the heat exchange perfusion apparatus was maintained such that all solutions perfusing the heart were constant at 37°C. At the end of the enzymatic digestion, the ventricles were removed and mechanically disassociated in 6 ml of enzymatic digestion solution plus 6 ml of 2× BDM/BSA solution [prepared by adding 3 g BSA, fraction V (GIBCO-BRL 11018-025) to 150 ml of BDM stock (40 mM)]. After mechanical disassociation by mincing with fine scissors, the tissue homogenate was filtered through a mesh filter into a conical tube; cells adhering to the filter were collected by washing with an additional 10-ml aliquot of 1× BDM-BSA solution that was prepared by combining 100 ml of BDM stock, 40 mM; 100 ml of heart medium prepared as described above, and 2 g of BSA, fraction V (GIBCO-BRL 11018-025). 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 1× BDM-BSA. The cells were then washed and pelleted further in BDM-BSA buffer with increasing increments of calcium (100, 200, 500 µM, and a final concentration of 1,000 µM). After the final pelleting step, the supernatant was removed, and the pellet was resuspended in minimum essential medium (MEM) [which was prepared by adding 10.8 g 1× MEM (Sigma M-1018), 1 g NaHCO3, 2.3 g HEPES, and 10 ml penicillin-streptomycin (100×, GIBCO-BRL 1540-122) with 950 MilliQ water; total volume is then adjusted to 1 liter. At the time of MEM preparation, the medium was bubbled with 95% O2-5% CO2 for 15 min, the pH was adjusted to 7.1 with 1 M NaOH, and 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 per milliliter, and viability was determined.
Cell culture.
The HL-1 cell line was obtained from Dr. William C. Claycomb, Louisiana
State University Medical Center. HL-1 cells are a cardiac muscle cell
line derived from the AT-1 subcutaneous mouse atrial cardiomyocyte
tumor lineage, which maintain the morphological, biochemical, and
electrophysiological phenotype of adult myocytes in culture
(5). HL-1 cells can be passaged serially and cultured as
adherent cells in T-25 flasks as described (5). We
estimate by terminal deoxynucleotidyl transferase (TdT)-mediated
dUTP nick-end labeling (TUNEL) analysis that ~8% of confluent
HL-1 cells undergo apoptosis without stimulation. The cells are
grown in Ex-Cell 320 medium part A and B (JRH Biosciences; Lenexa, KS),
10% fetal bovine plasma, 10 µg/ml insulin, 50 µg/ml endothelial
cell growth supplement, 1 µM retinoic acid, 10 µM norepinephrine,
100 U/ml penicillin, 100 µg/ml streptomycin, 250 pg/ml amphotericin,
and an additional 1× nonessential amino acids. Cells were stimulated with 1-50 ng/ml recombinant mouse TNF-. Cells were stimulated after reaching confluency for 24 h. Confluent HL-1 cells were treated with 50 ng/ml rhuTNFR:Fc (Immunex), a soluble TNF- receptor, which binds and neutralizes the bioactivity of TNF-. Confluent HL-1
cells were also treated with plasma isolated from either burned or
sham-burned rats, which had been incubated with 2 µg/ml of a
recombinant NH2-terminal fragment of bactericidal
permeability-increasing protein (rBPI21, XOMA; Berkeley,
CA). Incubation with rBPI21 occurred for 30 min before
exposure to HL-1 cells to neutralize any endotoxin present in the
plasma (25). All cells were removed from the flasks by
gentle scraping and agitation for analysis.
TUNEL assay.
Twenty microliters (1 × 106 cells/ml) of HL-1 cells
were harvested from six-well macrotiter plates by gentle agitation,
transferred to a ProbeonPlus slide (Fisher Scientific; Houston, TX),
and allowed to adhere for 1 h at room temperature. The slides were
then fixed in 10% formalin for 10 min at room temperature, washed in
phosphate-buffered saline (PBS; pH 7.4) twice for 5 min at room
temperature, postfixed in ethanol-acetic acid (2:1 vol/vol) for 5 min
at 20°C, and washed twice in PBS (pH 7.4) for 5 min at room
temperature. Cells were stained using in situ apoptosis
detection reagents (Intergen; Purchase, NY). To accomplish this, cells
were preincubated in equilibration buffer (13 µl/cm2)
with a plastic coverslip for 5 min at room temperature. Excess equilibration buffer was drained and TdT, diluted in reaction buffer
containing digoxigenin-labeled nucleotide, or for negative controls,
glass-distilled water in reaction buffer containing digoxigenin-labeled
nucleotide, was applied to the cells (11 µl/cm2) for 60 min at 37°C in a moist chamber. At the end of the incubation, one to
four slides were immersed in a Coplin jar with 35 ml of stop reagent
(Intergen), diluted 1:35 in glass-distilled water, and incubated for 10 min at 37°C with constant agitation. They were then washed three
times with PBS (pH 7.4) for 3 min at room temperature. Cells were
stained with antidigoxigenin-fluorescein in blocking solution for 30 min at room temperature in a dark, moist chamber (13 µl/cm2) and washed three times in PBS (pH 7.4) for 5 min
at room temperature. The slides were counterstained with propidium
iodide (3 µl/cm2) (Ventana Medical Systems; Tucson, AZ)
at room temperature in the dark and sealed with a glass coverslip
mount. Slides were stored at 20°C until examination.
Caspase-3 activity assay.
Caspase-3 activity was quantified by measuring a relative
synthetic peptide composed of aspartic acid, glutamic acid, valine, and
aspartic acid (DEVDase), or caspase, cleavage activity. The assay was carried out using the ApoAlert Caspase-3 Assay Kit
(Clontech). HL-1 cells were incubated in six-well plates to confluency
and were stimulated with TNF- or plasma as described in
RESULTS. Negative control cells were not stimulated.
Caspase activity was measured according to the manufacturer's
instruction. All results were calculated against a standard pNA
calibration curve. To confirm the correlation between protease activity
and signal detection, we also performed the control reaction of
incubating a TNF--induced sample with caspase-3 inhibitor (DEVD-fmk)
before adding substrate.
DNA-based ELISA.
Mono- and oligonucleosomes produced by endogenous endonucleases were
detected using mouse monoclonal antibodies. HL-1 cells were grown in
48-well plates until confluent and were stimulated with varying
concentrations of TNF- and/or 10% burn or sham-burned plasma for
24 h. The Cell Death Detection ELISA PLUS kit (Roche Diagnostics)
was used to assess apoptosis, and the manufacturer's directions were followed for the assay with the exception that after
lysis, 5 µl instead of 20 µl from the supernatant were transferred into the streptavidin-coated microtiter plate. All samples were done in
duplicate, and the absorbance values were averaged. Results were
calculated after subtracting the background value of the immunoassay
from the average of the absorbance values.
Mouse TNF- immunoassay.
After a 24-h exposure to plasma isolated from either burned or
sham-burned rats, HL-1 cells were collected into a 1.5-ml Eppendorf tube and were resuspended in 10 mM HEPES, pH 7.4, 2 mM EDTA,
0.1% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, 5 mM dithiothreitol, 1 mM phenylmethylsulfonyl fluoride, 10 µg/ml
pepstatin A, 10 µg/ml aprotinin, and 20 µg/ml leupeptin. After the
cells were lysed with a 25-g needle, they were then microcentrifuged for 20 min at 14,000 g. Fifty microliters of each resulting
supernatant were used for the enzyme-linked immunosorbent assay (ELISA)
assay to measure TNF-. The assay was performed as per the
manufacturer's instructions (Quantkine Murine, R&D Systems).
RT-PCR.
All reagents and primers were purchased from GIBCO-BRL. RNA (1.5 µg)
was reverse transcribed with the use of six units of SuperScript RTII,
12.5 ng/µl of oligo (dT)12-18, and 500 µM of each
dNTP in a solution containing 1 unit Rnasin solution, 5 µM
1,4-dithiothreitol, and 1× buffer (50 mM Tris · HCl, pH 8.3; 75 mM KCl, and 3 mM MgCl2) in a total volume of 20 µl.
The reaction was carried out at 37°C for 60 min, followed by 95°C
for 5 min to heat inactivate the reverse transcriptase. The primers for TNF- were 5' CCC GGT ACC CTC AGA TCA TCT TCT CAA AAT 3' and 5' TTC
TCC AGC TGG AAG ACT CC 3'.
amplification: one cycle of 94°C for
2 min, followed by 27 cycles of (94°C for 45 s, 63°C for
45 s, 72°C for 45 s) and then one cycle of 72°C for 5 min.
| |
RESULTS |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Plasma isolated from burned rats stimulates apoptosis in HL-1 and primary culture cardiac myocytes. Confluent HL-1 cardiac myocytes were exposed to medium containing 10% plasma harvested from either burned or sham-burned rats for a 24-h period. Plasma was harvested 4 h after burn wound based on caspase-3 activity assays and a DNA-based ELISA, which measured the number of nucleosomes containing single- or double-stranded DNA. Maximum HL-1 cell apoptosis was observed using plasma harvested 4 h postburn compared with 2, 8, 18, and 24 h after burn trauma (data not shown).
A significant 3.6-fold increase in caspase-3 activity was observed in HL-1 cells treated with plasma harvested from burned versus sham-burned animals (Fig. 1A). This increase was comparable to the apoptotic rise observed in confluent HL-1 cells treated with 20 ng/ml TNF- for 24 h. Consistent with
the caspase-3 data, a 3.4-fold increase in apoptosis was also
detected using the DNA-based ELISA (Fig. 1B).
|
|
concentrations were measured by ELISA in the primary culture
myocytes both at the time of harvest and 24 h after plasma stimulation. Less than 1 pg/ml of TNF- was detected at any time point in either the control, sham, or burn-stimulated cells.
In an independent experiment, the effect of burn plasma on HL-1 cells
was also examined using TUNEL. The results are shown in Fig.
3. As can be seen in Fig. 3, B
and D, when HL-1 cells are exposed to either TNF- as a
positive control or 10% burn plasma, apoptotic cells appear as
green bodies as a result of the TUNEL-FITC staining. In contrast, in
Fig. 3, A and C, when HL-1 cells are either
untouched as a control or exposed to 10% plasma isolated from
sham-burned rats, the cells appear red from the propidium iodide
staining. These results are consistent with the findings presented in
Fig. 1. We conclude that plasma harvested from burned, but not
sham-burned animals, is capable of causing apoptosis in HL-1
cells.
|
Burn plasma-induced apoptosis is TNF independent.
It has previously been demonstrated that TNF- causes
apoptosis in a variety of cell types, including myocytes
(27). We confirmed that TNF- also induced a
dose-related increase in apoptosis in HL-1 cardiac myocytes. A
sixfold increase in caspase-3 activity was observed between cells
treated with 1 ng/ml TNF- and 50 ng/ml TNF- (Fig.
4A). As an independent
confirmation of apoptosis, we also observed a sevenfold
increase in the enrichment of mono- and oligosomes (Fig.
4B), confirming the dose-dependent increase in
apoptosis using the ELISA assay. A fourfold increase in
TUNEL-positive cells was observed (data not shown) between control HL-1
cells and those incubated with 50 ng/ml TNF-. Taken together, these results confirm that exposure of HL-1 cardiac myocytes to TNF- results in apoptosis.
|
, we hypothesized that the apoptotic response observed was due to TNF-. However, this hypothesis was refuted by three independent groups of experiments. First, an ELISA
assay specific for mouse TNF- detected no measurable TNF- in
either burn or sham burn plasma (data not shown). To investigate the
possibility that TNF- was synthesized by HL-1 cells in response to
burn plasma, the TNF- ELISA was also performed on the culture medium
after the 24-h coincubation of burn plasma with HL-1 cells. Again, no
TNF- was detected in culture medium (data not shown). Despite the
lack of TNF- in the medium, it was theoretically possible that
TNF- might be secreted in small quantities by HL-1 cells and act in
autocrine fashion such that TNF- might not be detected in the
culture medium. To investigate this possibility, we performed RT-PCR to
sensitively detect TNF- mRNA in the stimulated HL-1 cells. No
TNF- mRNA was detected by RT-PCR (Fig.
5).
|
was not involved in HL-1 cell
apoptosis, we pretreated plasma with 50 ng/ml of rhuTNFR:Fc before incubation with HL-1 cells. Despite blockade of TNF-
bioactivity, no decrease in apoptosis was observed in samples
treated with rhuTNFR:Fc, as measured by both the caspase-3 assay and
the DNA-based ELISA. These experiments indicated that TNF- is not
responsible for cardiac myocyte apoptosis induced by burn
plasma (Fig. 6, A and
B).
|
Burn plasma-induced apoptosis is endotoxin dependent.
Because apoptosis was not caused by TNF-, we next
hypothesized that plasma endotoxin might be responsible for inducing
apoptosis. To determine whether endotoxin was capable of
inducing apoptosis in HL-1 cardiac myocytes, we incubated cells
with lipopolysaccharide (LPS, 10 ng/ml to 1 µg/ml) for 24 h. A significant increase in caspase activation was detected at
LPS concentrations higher than 100 ng/ml (Fig.
7A). The LPS-stimulated
increase in caspase activity could be alleviated by the addition of 2 µg/ml of a recombinant NH2-terminal bactericidal
permeability-increasing protein (rBPI21) to the LPS
containing medium for 30 min before exposure to HL-1 cells (Fig.
7A). To determine whether the LPS-induced apoptosis was secondary to increased TNF- secretion, we also examined caspase activity in HL-1 cells treated with 50 ng/ml of rhuTNFR:Fc. No significant decrease in caspase-3 activity was observed (data not
shown).
|
| |
DISCUSSION |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Myocyte apoptosis occurs in several cardiac diseases,
including myocarditis (20), ischemia-reperfusion
injury (13), chronic pressure overload (2,
3), congestive heart failure (28), and sepsis
(27). Recently, we demonstrated that myocyte
apoptosis occurred in the ventricular myocardium of rats
following severe thermal trauma and was temporally correlated with the
development of cardiac dysfunction (14). Here, we
demonstrate by multiple methods, that myocyte apoptosis
following thermal injury is mediated, at least in part, by factors in
burn plasma. Apoptosis was neither associated with the presence
of TNF- in burn plasma, nor the induction of TNF- in either HL-1
cells or primary culture myocytes exposed to burn plasma. However,
apoptosis was significantly reduced by inhibition of endotoxin
activity in burn plasma by rBPI21, a recombinant
NH2-terminal fragment of the bactericidal
permeability-increasing protein.
The ability of burn plasma to induce apoptosis was examined in HL-1 cells, a cardiac muscle cell line that maintains an ultrastructure similar to primary cardiac myocytes and maintains the ability to spontaneously contract while remaining in a mitotic state typical of normal in vivo immature mitotic cardiomyocytes (5). HL-1 cells represent a particularly useful model, because it has been recently shown that human cardiac myocytes are capable of entering mitosis and undergoing division after injuries such as myocardial infarction (1). We show that in HL-1 cells, caspase-3 activity was induced by burn plasma to levels at least three times greater than control levels. Caspase-3 activation confirms that at least one apoptotic signal transduction pathway has been activated. In addition to caspase signaling, we also demonstrated the presence of late-stage apoptotic cells via TUNEL staining and a DNA-based ELISA measuring the enrichment of mono- and oligosomes from the systematic breakdown of the nucleosomal DNA.
Whereas HL-1 cells represent a useful and relevant myocyte model, we further demonstrated that burn plasma was capable of stimulating apoptosis in primary culture rodent myocytes. Using the caspase-3 assay, a twofold increase was observed in myocytes that were exposed to plasma isolated from burned animals, confirming the HL-1 cells as a relevant model system.
Because TNF- is a potent inducer of cardiac myocyte
apoptosis (22) and because TNF- is frequently
induced by severe systemic insults, we fully expected that TNF-
would account for the apoptotic activity of burn plasma. This
hypothesis proved completely false. No TNF- was detected in burn
plasma, and no TNF- or TNF- mRNA was produced by HL-1 cells. This
lack of TNF- in burn plasma is not entirely unexpected, because
elevation of plasma TNF- was uncommon in multiple studies of severe
burn injury in humans (9, 10, 29).
Because TNF- was not directly involved in the pathogenesis of
apoptosis in this model, we investigated additional
possibilities. Burn trauma results in a multitude of cellular changes
that may trigger the apoptotic cascade, including increases in
intracellular calcium concentration, a rise in reactive oxygen
metabolites, and an increase in both local and systemic levels of
interleukin-1 (16, 21, 30). In addition, burn injury can
also lead to transient intestinal ischemia, contributing to a
disruption of the normal gut barrier function and translocation of
indigenous bacteria or bacterial products (7, 8, 15).
Because endotoxin infusion was recently associated with cardiac myocyte
apoptosis in a rat model (26), we hypothesized
that endotoxin present in the plasma of burned animals might also
account for cardiac apoptosis in the burn model.
rBPI21, a recombinant form of the neutrophil-derived protein binds with high affinity to the lipid A portion of LPS and inhibits all LPS bioactivity, including its ability to promote apoptosis (4). Bactericidal permeability-increasing protein is known to neutralize LPS activities in vitro and in vivo (24). When burn plasma was preincubated with rBPI21, the caspase-3 activity dropped 2.7 times compared with the activity observed in using untreated burn plasma. These results indicate that a significant portion of the observed apoptotic activity is due to endotoxin contained within the plasma collected from the burned animals. The observation that pretreatment of the cells with rBPI21 does not result in the abolition of all apoptosis, suggests the presence of other unidentified proapoptotic factors within the burn plasma. The source of endotoxin could be from the gut via translocation or from colonization of the burn wound itself. Whichever the specific origin, these data suggest that the systemic inhibition of endotoxin may provide a therapeutic approach for the treatment of burn-associated cardiac dysfunction.
| |
ACKNOWLEDGEMENTS |
|---|
This work was supported by the National Institute of General Medical Science Grant GM-21681-36.
| |
FOOTNOTES |
|---|
Address for reprint requests and other correspondence: B. P. Giroir, Children's Medical Center, 1935 Motor St., Dallas, TX 75235.
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.
10.1152/ajpheart.00393.2001
Received 10 May 2001; accepted in final form 6 November 2001.
| |
REFERENCES |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
1.
Beltrami, A,
Urbanek K,
Kajstura J,
Yan S,
Finato N,
Bussani R,
Nadal-Ginard B,
Silvestri F,
Leri A,
Beltrami A,
and
Anversa P.
Evidence that human myocytes divide after myocardial infarction.
N Engl J Med
344:
1750-1757,
2001.
2.
Bing, O.
Hypothesis: apoptosis may be a mechanism for the transition to heart failure with chronic pressure overolad.
J Mol Cell Cardiol
26:
943-948,
1994.
3.
Bromme, H,
and
Holtz J.
Apoptosis in the heart: when and why?
Mol Cell Biochem
163/164:
261-275,
1996.
4.
Casey, L.
Immunologic response to infection and its role in septic shock.
Crit Care Clin
16:
110-120,
2000.
5.
Claycomb, WC,
Lanson NAJ,
Stallworth BS,
Egeland DB,
Delcarpio JB,
Bahinski A,
and
Izzo NJJ
HL-1 cells: a cardiac muscle cell line that contracts and retains phenotypic characteristics of the adult cardiomyocyte.
Proc Natl Acad Sci USA
95:
2979-2984,
1998.
6.
Crum, RL,
Dominic W,
Hansbrough JF,
Shackford SR,
and
Brown MR.
Cardiovascular and neurohumoral responses following burn injury.
Arch Surg
125:
1065-1069,
1990.
7.
Cuevas, P,
Ishiyama M,
Koizumi S,
Woodruff P,
Kaufman A,
and
Fine J.
Role of endotoxemia of intestinal origin in early death from large burns.
Surg Gynecol Obestet
138:
725-730,
1974.
8.
Deitch EA and Bridges RM. Effect of stress and trauma on bacterial
translocation from the gut. J Surg Res 42, 1987.
9.
Drost, A,
Burleson D,
Cioffi W,
Jordan B,
Mason A,
and
Pruitt B.
Plasma cytokines following thermal injury and their relationship with patient mortality, burn size, and time postburn.
J Trauma
35:
335-339,
1993.
10.
Endo, S,
Inada K,
Yamada Y,
Kasai T,
Takakuwa T,
Nakae H,
Kikuchi M,
Hoshi S,
Suzuki M,
and
Yamashita H.
Plasma tumour necrosis factor- (TNF-) levels in patients with burns.
Burns
19:
124-127,
1993.
11.
Ferrari, R.
The role of TNF in cardiovascular disease.
Pharmacol Res
40:
97-105,
1999.
12.
Gelfand, JA,
Donelan M,
and
Burke JF.
Preferential activation and depletion of the alternative complement pathway by burn injury.
Ann Surg
198:
58-62,
1983.
13.
Gottlieb, R,
Burleson K,
Kloner R,
Babior B,
and
Engler R.
Reperfusion injury induces apoptosis in rabbit cardiomyocytes.
J Clin Invest
94:
1621-1628,
1994.
14.
Horton, J.
Cellular basis for burn-mediated cardiac dysfunction in adult rabbits.
Am J Physiol Heart Circ Physiol
271:
H2615-H2621,
1996.
15.
Horton, JW.
Bacterial translocation after burn injury: the contribution of ischemia and permiability changes.
Shock
1:
286-290,
1994.
16.
Horton, JW.
Oxygen free radicals contribute to postburn cardiac cell membrane dysfunction.
J Surg Res
61:
97-102,
1996.
17.
Horton, JW,
Baxter CR,
and
White JW.
Differences in cardiac responses to resuscitation from burn shock.
Surg Gynecol Obstet
168:
201-213,
1989.
18.
Idriss, HT,
and
Naismith JH.
TNF and the TNF receptor superfamily: structure-function relationship(s).
Microsc Res Tech
50:
184-195,
2000.
19.
Iglesias, J,
LaNoue J,
Rogers T,
Nguyen H,
and
Turnage R.
Physiological basis of pulmonary edema during intestinal reperfusion.
J Surg Res
80:
156-163,
1998.
20.
Kawano, H,
Okada R,
Kawano Y,
Sueyoshi N,
and
Shirai T.
Apoptosis in acute and chronic myocarditis.
Jpn Heart J
35:
745-750,
1994.
21.
Koshy, US,
Burton KP,
Le TH,
and
Horton JW.
Altered ionic calcium and cell motion in ventricular myocytes after cutaneous thermal injury.
J Surg Res
68:
133-138,
1997.
22.
Krown, KA,
Page MT,
Nguyen C,
Zechner D,
Gutierrez V,
Comstock KL,
Glembotski CC,
Quintana PJ,
and
Sabbadini RA.
Tumor necrosis factor alpha-induced apoptosis in cardiac myocytes. Involvement of the sphingolipid signaling cascade in cardiac cell death.
J Clin Invest
98:
2854-2865,
1996.
24.
Marra, M,
Wilde C,
Collins M,
Snable J,
Thornton M,
and
Scott R.
The role of bactericidal/permeability increasing protein as a natural inhibitor of bacterial endotoxin.
J Immunol
148:
532-537,
1992.
25.
Marra, M,
Wilde C,
and
Griffithe J.
Bactericidal/permeability-increasing protein has endotoxin-neutralizing activity.
J Immunol
144:
662-666,
1990.
26.
McDonald, T,
Grinman M,
Carthy C,
and
Walley K.
Endotoxin infusion in rats induces apoptotic and survival pathways in hearts.
Am J Physiol Heart Circ Physiol
279:
H2053-H2061,
2000.
27.
Meldrum, D.
Tumor necrosis factor in the heart.
Am J Physiol Regulatory Integrative Comp Physiol
274:
R577-R595,
1998.
28.
Olivetti, G,
Abbi R,
Quaini F,
Kajstura J,
Cheng W,
Nitahara J,
Quaini E,
Di Loreto C,
Beltrami C,
Krajewski S,
Reed J,
and
Anversa P.
Apoptosis in myocytes in end-stage heart failure.
N Engl J Med
336:
1131-1141,
1996.
29.
Vindenes, H,
Ulvestad E,
and
Bjerknes R.
Concentrations of cytokines in plasma of patients with large burns: their relation to time after injury, burn size, inflammatory variables, infection, and outcome.
Eur J Surg
164:
647-656,
1998.
30.
White DJ, Maass D, Sanders B, and Horton JW. The potential role of
of elevated intracellular calcium in the development of postburn
cardiac dysfunction. Surg Forum 46, 1995.
31.
Wright, J,
Nwariaku F,
Clark J,
Falck J,
Rogers T,
and
Turnage R.
Effect of diabetes mellitus on endotoxin-induced lung injury.
Arch Surg
134:
1354-1358,
1999.
32.
Zhang, L,
Xiao Y,
and
He J.
Cocaine and apoptosis in myocardial cells.
Anatom Rec (New Anat)
257:
208-216,
1999.
This article has been cited by other articles:
|
|
 |
|
  R. Matyal Newly Appreciated Pathophysiology of Ischemic Heart Disease in Women Mandates Changes in Perioperative Management: A Core Review Anesth. Analg., July 1, 2008; 107(1): 37 - 50. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Li, X. Jiao, L. Tao, H. Liu, Y. Cao, B. L. Lopez, T. A. Christopher, and X. L. Ma Tumor necrosis factor-{alpha} in mechanic trauma plasma mediates cardiomyocyte apoptosis Am J Physiol Heart Circ Physiol, September 1, 2007; 293(3): H1847 - H1852. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. L. Carlson, D. L. Maass, J. White, P. Sikes, and J. W. Horton Caspase inhibition reduces cardiac myocyte dyshomeostasis and improves cardiac contractile function after major burn injury J Appl Physiol, July 1, 2007; 103(1): 323 - 330. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Zhang, H.-Y. Wang, R. Bassel-Duby, D. L. Maass, W. E. Johnston, J. W. Horton, and W. Tao Role of interleukin-6 in cardiac inflammation and dysfunction after burn complicated by sepsis Am J Physiol Heart Circ Physiol, May 1, 2007; 292(5): H2408 - H2416. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Q. Zang, D. L. Maass, J. White, and J. W. Horton Cardiac mitochondrial damage and loss of ROS defense after burn injury: the beneficial effects of antioxidant therapy J Appl Physiol, January 1, 2007; 102(1): 103 - 112. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Carlson, D. L. Maass, D. J. White, J. Tan, and J. W. Horton Antioxidant vitamin therapy alters sepsis-related apoptotic myocardial activity and inflammatory responses Am J Physiol Heart Circ Physiol, December 1, 2006; 291(6): H2779 - H2789. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. L. Maass, J. White, B. Sanders, and J. W. Horton Role of cytosolic vs. mitochondrial Ca2+ accumulation in burn injury-related myocardial inflammation and function Am J Physiol Heart Circ Physiol, February 1, 2005; 288(2): H744 - H751. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. W. Horton, J. Tan, D. J. White, D. L. Maass, and J. A. Thomas Selective decontamination of the digestive tract attenuated the myocardial inflammation and dysfunction that occur with burn injury Am J Physiol Heart Circ Physiol, November 1, 2004; 287(5): H2241 - H2251. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. M. White, P. E. Constantin, and W. C. Claycomb Cardiac physiology at the cellular level: use of cultured HL-1 cardiomyocytes for studies of cardiac muscle cell structure and function Am J Physiol Heart Circ Physiol, March 1, 2004; 286(3): H823 - H829. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Visit Other APS Journals Online |
