Recognition that resuscitation with Ringers lactate (RL) potentiates trauma-hemorrhagic shock (T/HS)-induced organ injury and systemic inflammation has led to a search for improved initial fluid resuscitation regimens. However, one relatively neglected component in the search for new and novel resuscitation strategies is a determination of what fluid resuscitation therapy (i.e., control group) the new experimental regimen of interest should be tested against. Thus, we tested the effects of three commonly used resuscitation strategies on trauma-shock-induced gut and lung injury, as well as neutrophil activation and red blood cell (RBC) function.
Male Sprague Dawley rats were subjected to a laparotomy (trauma) and 90 minutes of sham shock (trauma-sham shock [T/SS]) or a laparotomy plus hemorrhagic shock (T/HS), followed by a reperfusion period of 3 hours. The T/HS groups were resuscitated either with their shed blood (SB), or half the SB and 1.5 times the SB volume as RL (SB/RL), or 3 times the SB volume as RL (3RL). The T/SS groups received either no resuscitation or RL at 1.5 times the SB volume of the T/HS rats. Gut injury was quantified by measuring intestinal permeability to flourescein dextran (FD-4), as well as by histologic analysis of the terminal ileum. Lung injury was assessed histologically and by the magnitude of neutrophil sequestration as reflected in myeloperoxidase levels. Neutrophil activation was measured by quantitating the level of CD11b expression using flow cytometry. RBC injury was analyzed by measuring the RBC deformability.
As compared with the T/SS groups, all three T/HS resuscitation regimens were associated with morphologic evidence of gut and lung injury, increased gut permeability, pulmonary leukosequestration, systemic neutrophil activation, and decreased RBC deformability (p < 0.05). However, the effect of the resuscitation regimens varied based on the tissues and cells tested. Morphologically, gut and lung injury as well as pulmonary neutrophil sequestration was worse in the 3RL T/HS group than the other two T/HS groups. As compared with the other two T/HS resuscitation regimens, resuscitation with the SB/RL combination was associated with less of an increase in gut permeability, systemic neutrophil activation, and RBC rigidification (p < 0.05).
The type of resuscitation regimen used influenced the extent of organ injury and cellular activation or dysfunction observed after T/HS with different resuscitation regimens showing varying effects depending on the cell or organ tested. Thus, when testing novel fluid resuscitation regimen, attention must be paid to the control resuscitation regimen used.
[Show abstract][Hide abstract] ABSTRACT: Abstract Hemorrhagic shock (HS) is the single most common cause of death in civilian and military personnel experiencing trauma (Alam & Rhee, 2007). Immediate resuscitation for HS can involve the administration of supplemental oxygen (O2) above ambient levels (0.21) using a non-rebreather face mask and high flow O2 of 10-15 L/min (Trauma, 2008). With supplementary O2, reactive oxygen species (ROS) may be increased leading to oxidative damage to deoxyribonucleic acid (DNA), proteins and lipid membranes (Rushing & Britt, 2008). During hyperoxia, ROS can cause a secondary oxidative injury to cells and tissues especially in the lungs and diaphragm. The optimal amount of O2 to be administered following HS is not clearly defined. The purpose of this dissertation was to investigate the optimal fraction of inspired oxygen (FIO2) to be administered following HS by determining the amount of hydrogen peroxide (H2O2) and apoptosis in the lungs and diaphragm. Previous animal studies have demonstrated that dopamine (DA) can scavenge free radicals or decrease ROS by increasing blood flow and can decrease apoptosis by activating β-2 adrenoreceptors (Communal, Singh, Sawyer, & Colucci, 1999; Patterson et al., 2004; J. D. Pierce, Goodyear-Bruch, Hall, & Clancy, 2006; J. D. Pierce, Goodyear-Bruch, Hall, Reed, & Clancy, 2008) Therefore, we conducted additional experiments to determine if DA with various FIO2s following HS reduces apoptosis in these tissues. Adult male Sprague-Dawley rats (n = 112) were anesthetized; a tracheostomy was performed and catheters were inserted in the carotid and femoral arteries. HS was elicited by withdrawing 40% of the rat's blood volume over 30 minutes. This was followed by the rat breathing one of the following FIO2 concentrations (0.21, 0.40, 0.60, and 1.00) without and with concurrent DA (10 mcg/kg/min) for 60 minutes. The animal was euthanized and the lungs and diaphragm excised and prepared for measurement of H2O2 and nuclear DNA damage (apoptosis). Hydrogen peroxide was quantified using dihydrofluorescein diacetate (Hfluor-DA) and laser scanning cytometry. Percent apoptosis was determined using differential dye up-take and fluorescent microscopy. The amount of lung and diaphragm H2O2 and percent apoptosis were greatest in the 0.21 and 1.00 FIO2 concentrations and the least amounts were observed in rats when using 0.40. Infusing DA significantly decreased H2O2 and apoptosis in both tissues at all FIO2's except 0.40. The lack of difference in rats receiving 0.40 with DA was because of the already reduced H2O2 and apoptosis values. In conclusion, an FIO2 of 0.40 was optimal for attenuation of lung and diaphragm H2O2 and apoptosis following HS. When greater FIO2s are necessary, adding DA to the resuscitation regimen may diminish ROS-induced cellular injury.
[Show abstract][Hide abstract] ABSTRACT: This study was designed to determine the effects of various resuscitation fluids on intestinal injuries after hemorrhagic shock and resuscitation (HS/R) and to determine the potential mechanisms. We induced HS by bleeding male Sprague-Dawley rats to a blood pressure of 30 to 40 mmHg for 60 min. Sixty minutes later, the rats were killed (HS group) or immediately resuscitated with L-isomer lactated Ringer's solution (HS + LR group), shed blood (HS + BL group), or hydroxyethyl starch (HS + HES group) to maintain the blood pressure to the original value during the 60-min resuscitation period. Three hour after resuscitation, bacterial translocation (BT), intestinal permeability, ileal levels of tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, malondialdehyde (MDA), oxidized and reduced glutathione (GSH and GSSG), myeloperoxidase (MPO) activity, nuclear factor (NF)-kappaB, activator protein (AP)-1 activation, and ileal microscopic and ultrastructural histological changes were measured. Another experiment was designed for survival study of 24 h. HES 130/0.4 solution was as effective as shed blood, required a small volume requirement to restore circulation, and significantly reduced HS/R-induced ileal villous morphological injuries with an anti-inflammatory effect, as reflected by a reduction of TNF-alpha, IL-6, MPO activity, and NF-kappaB activation. In addition, HES resuscitation also reduced intestinal permeability and BT and caused less oxidative stress as reflected by a reduction of MDA, GSSG/GSH and AP-1 activation along with restored GSH, whereas shed blood couldn't. No significant difference was observed in outcome among groups. HES 130/0.4 resuscitation prevents HS/R induced intestinal injury by modulating inflammatory response and preventing oxidative stress in a rat model of hemorrhagic shock. These physiological protective effects appear to be mediated by down-regulation of the transcription factor NF-kappaB and AP-1.
[Show abstract][Hide abstract] ABSTRACT: Flow and laser scanning cytometry are used extensively in research and clinical settings. These techniques provide clinicians and scientists information about cell functioning in a variety of health and disease states. An in-depth knowledge and understanding of cytometry techniques can enhance interpretation of current research findings. Our goal with this review is to reacquaint clinicians and scientists with information concerning differences between flow and laser scanning cytometry by comparing their capabilities and applications.
A Pubmed abstract search was conducted for articles on research, reviews and current texts relating to origins and use of flow and laser scanning cytometry. Attention was given to studies describing application of these techniques in the clinical setting.
Both techniques exploit interactions between the physical properties of light. Data are immediately and automatically acquired; they are distinctly different. Flow cytometry provides valuable rapid information about a wide variety of cellular or particle characteristics. This technique does not provide the scanned high resolution image analysis needed for investigators to localize areas of interest within the cell for quantification. Flow cytometry requires that the sample contain a large amount disaggregated, single, suspended cells. Laser scanning cytometry is slide-based and does not require as large of a sample. The tissue sample is affixed to a slide allowing repeated sample analyses. These cytometry techniques are used in the clinical setting to understand pathophysiological derangements associated with many diseases; cardiovascular disease, diabetes, acute lung injury, hemorrhagic shock, surgery, cancer and Alzheimer's disease.
Understanding the differences between FCM and LSCM can assist investigators in planning and design of their research or clinical testing. Researchers and clinicians optimize these technique capabilities with the cellular characteristics they wish to measure delineating molecular and cellular events occurring in health and disease. Discovery of mechanisms in cells using FCM and LSCM provide evidence needed to guide future treatment and interventions.
International Journal of Clinical Monitoring and Computing 08/2010; 24(4):251-9. DOI:10.1007/s10877-010-9242-4 · 1.99 Impact Factor
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