Pyruvate Ringer Research Articles

Pyruvate alleviates lipid peroxidation and multiple-organ dysfunction in rats with hemorrhagic shock

ObjectivePyruvate can reduce lipid peroxidation, which plays a critical role in organ injury, in various models. However, it is not fully understood if this inhibition occurs in resuscitation of hemorrhagic shock (HS). This study examines effects of pyruvate Ringer solution (PR) in this respect in rats. MethodsRats, subjected to 45% blood loss, were randomly allocated to the 3 groups (n = 18): HS with no fluid resuscitation (group NR), HS resuscitated with lactated Ringer solution (LR) (group LR), and HS resuscitated with PR (group PR). Mean arterial pressure, plasma levels of thiobarbituric acid reactive substances (TBARS), and superoxide dismutase were measured at various time points until 360 minutes after hemorrhage. Visceral organs were harvested at the end for evaluations of the TBARS, antioxidant enzyme, and tissue water content. Other 54 rats with identical procedures without sampling were documented for 24-hour survival rates (n = 18) after fluid resuscitation. ResultsPyruvate Ringer solution significantly increased mean arterial pressure and decreased blood TBARS levels after lethal HS. It also reduced TBARS concentrations and glutathione peroxidase activities but significantly enhanced glutathione reductase activities in most organs and greatly improved the ratios of reduced glutathione over oxidized glutathione in various organs in group PR, compared to group LR. Furthermore, PR significantly improved various organ function and water contents relative to LR. Group PR showed a more than 2-fold higher 24-hour survival rate of group LR. ConclusionsPyruvate Ringer solution alleviated organ edema and injury and prompted survival partially through inhibition of lipid peroxidation in various organs in severe HS rats.

The effects of sodium pyruvate Ringer solution on hemodynamic and organ functions during shock stage in dogs with a 50% total body surface area full-thickness burn

To compare the effect of intravenous resuscitation with sodium pyruvate (Pyr) Ringer solution against lactated Ringer solution on hemodynamic and organ functions during shock stage in dogs with burn. 28 Beagle dogs were subjected to 50% total body surface area (TBSA) burn, and they were divided into three groups: burn injury without fluid resuscitation (NR, n=8), Ringer lactate solution (RL, n=10), and Pyr Ringer solution (RP, n=10). They were given intravenous fluid resuscitation according to Parkland formula 30 minutes after burn. The hemodynamics, organ functions and mortality were observed in conscious state before burn injury, and 2, 6, 8, 12, 24 hours after burn injury. Within 24 hours after burn, all the dogs in NR group died, and those in RL and RP groups were all alive. At 2 hours after burn, the mean arterial pressure (MAP), cardiac index (CI), dp/dt max of left ventricular contractility were significantly reduced in NR, RL and RP groups compared with those before injury (MAP: 45.33±7.78 mmHg vs. 141.67±5.98 mmHg, 91.33±10.25 mmHg vs. 142.33±6.16 mmHg, 98.67±9.54 mmHg vs. 142.83±5.47 mmHg; CI: 8.17±0.83 mL×s(-1)×m(-2) vs. 48.34±3.33 mL×s(-1)×m(-2), 16.84±2.17 mL×s(-1)×m(-2) vs. 47.34±1.67 mL×s(-1)×m(-2), 19.00±1.50 mL×s(-1)×m(-2) vs. 47.34±1.33 mL×s(-1)×m(-2); dp/dt max: 426.83±51.91 mmHg/s vs. 1 372.50±39.61 mmHg/s, 594.00±88.23 mmHg/s vs. 1 363.83±44.92 mmHg/s, 645.00±66.82 mmHg/s vs. 1 395.83±19.49 mmHg/s, all P<0.05), and the systemic vascular resistance (SVR) and alanine transaminase (ALT), creatinine (Cr), serum MB isoenzyme of creatine kinase (CK-MB), diamine oxidase (DAO) were significantly higher (SVR: 1 322.50±36.37 kPa×s×L(-1) vs. 281.45±8.84 kPa×s×L(-1), 777.50±41.84 kPa×s×L(-1) vs. 289.72±6.70 kPa×s×L(-1), 571.40±40.01 kPa×s×L(-1) vs. 286.27±8.66 kPa×s×L(-1); ALT: 89.50±4.11 U/L vs. 40.57±3.63 U/L, 89.25±4.88 U/L vs. 37.92±2.62 U/L, 86.30±5.61 U/L vs. 38.47±3.50 U/L; Cr: 75.62±4.61 μmol/L vs. 41.58±2.78 μmol/L, 77.00±5.92 μmol/L vs. 46.55±3.17 μmol/L, 74.13±2.56 μmol/L vs. 45.65±1.83 μmol/L; CK-MB: 13.122±0.282 kU/L vs. 1.557±0.009 kU/L, 8.885±0.272 kU/L vs. 1.497±0.009 kU/L, 8.692±0.180 kU/L vs. 1.490±0.005 kU/L; DAO: 2.26±0.14 kU/L vs. 0.25±0.02 kU/L, 1.50±0.07 kU/L vs. 0.25±0.01 kU/L, 1.37±0.07 kU/L vs. 0.25±0.02 kU/L, all P<0.05). All parameters in NR group kept on worsening till death, while hemodynamic and organ functions of two intravenous resuscitation groups were gradually improved, CI, SVR and DAO in RP group were significantly superior to those of RL group from 2 hours on after burn (all P<0.05), and dp/dt max and CK-MB in RP group were significantly better than those of RL group from 6 hours on after burn (dp/dt max: 1,082.33±63.59 mmHg/s vs. 1,018.60±47.36 mmHg/s, CK-MB: 7,898.70±255.74 U/L vs. 8,438.70±442.00 U/L, all P<0.05), and MAP was significantly better than that of RL group at 6 hours (124.67±9.39 mmHg vs. 114.33±9.16 mmHg, P<0.05), and Cr was significantly better than that of RL group from 24 hours on after burn (53.42±4.99 μmol/L vs. 60.77±3.11 μmol/L, P<0.05). The Pyr Ringer solution was superior to the Ringer lactate solution in improving hemodynamic and organ functions for intravenous resuscitation in dogs with 50%TBSA full thickness burn.

Pyruvate Ringer's Solution Corrects Lactic Acidosis and Prolongs Survival during Hemorrhagic Shock in Rats

BackgroundLactic acidosis is a life-threatening complication of hemorrhagic shock. There is no ideal therapy for it in the clinical setting. ObjectiveThis study was designed to investigate whether pyruvate Ringer's solution could treat hypoxic lactic acidosis associated with lethal hemorrhagic shock in rats. MethodsA total of 54 rats were subjected to hemorrhagic shock with mean arterial pressure (MAP) of 40 mm Hg for 1 h. They were then randomly divided into three groups (n = 18 each): Group N had no fluid resuscitation; Group L received lactated Ringer's solution infusion; and Group P received pyruvate Ringer's solution infusion. The survival rate was investigated after 24 h. In addition, a second set of 54 rats was selected for blood sampling, with identical methods for shock and resuscitation being followed, to determine arterial pH, blood gas analysis, lactate, pyruvate, and organs' enzyme activities at various time points. The MAP was monitored for 6 h in both populations. ResultsPyruvate Ringer's solution significantly increased the survival rate of rats subjected to fatal shock and receiving pyruvate Ringer's solution (Group P) by 1.5 times the survival rate in Group L at 24 h after fluid resuscitation (55.6% vs. 22.2%, respectively; p < 0.05). Pyruvate infusion maintained a higher MAP and fully corrected severe acidosis 1 h after resuscitation in comparison to the lactated infusion, and markedly decreased blood lactate levels and the lactate-to-pyruvate ratio 4 h after resuscitation. It also significantly improved serum markers of organ dysfunction. ConclusionPyruvate Ringer's solution efficiently treated hypoxic lactic acidosis and significantly increased the survival rate in rats with lethal hemorrhagic shock. Pyruvate Ringer's solution is potentially applicable to clinical resuscitation in humans.

Pyruvate-Enriched Resuscitation Protects Hindlimb Muscle From Ischemia–Reperfusion Injury

Tourniquets impose ischemia on distal musculature. Resuscitation with pyruvate, an energy substrate and antioxidant, may ameliorate muscle ischemia-reperfusion injury. After goats were exsanguinated to lower mean arterial pressure to 48 mmHg, femoral vessels were occluded for 90 minutes to impose hindlimb ischemia. Lactate Ringer's (LR) or pyruvate Ringer's (PR) solution was infused from 30 minutes ischemia until 30 minutes reperfusion. Pro- and antiapoptotic proteins and injury markers were measured in gastrocnemius at 4 hours reperfusion. Pro-oxidant NADPH oxidase activity and nitrotyrosine content, a footprint of nitrosative stress, doubled, and poly (ADP-ribose) polymerase cleavage, an early apoptotic event, increased 80% in LR-resuscitated vs. sham muscle, but PR prevented these increases. Antiapoptotic Bcl-X(L) content fell in LR-treated vs. sham and PR-treated muscle. Water content increased in LR- but not PR-resuscitated muscle. LR resuscitation imposed oxido-nitrosative stress and initiated proapoptotic mechanisms, while PR blunted these harmful consequences of muscle ischemia-reperfusion.

Histone deacetylase as therapeutic target in a rodent model of hemorrhagic shock: Effect of different resuscitation strategies on lung and liver

DNA transcription is regulated in part by acetylation of nuclear histones, controlled by 2 groups of enzymes: histone deacetylases (HDAC) and histone acetyl transferases (HAT). We have shown previously that hemorrhage and resuscitation are associated with HDAC/HAT imbalance, which influences the acetylation status of cardiac histones. The goals of this study were to determine whether: (1) resuscitation after hemorrhage affects histone acetylation in a fluid- and organ-specific fashion; and (2) administration of HDAC inhibitors influences histone acetylation and subsequent gene expression. In the first experiment, rats (n = 6/group) were subjected to volume-controlled hemorrhage and resuscitated with: (1) racemic lactated Ringer's (DL-LR); (2) L-lactated Ringer's (L-LR); (3) 7.5% hypertonic saline (HTS); (4) ketone Ringer's (KR); or (5) pyruvate Ringer's (PR). Control groups included: (6) no hemorrhage (Sham); and (7) hemorrhage with no resuscitation (NR). In the second experiment (n = 5/group), 3 HDAC inhibitors, valproic acid (VPA), trichostatin A (TSA), and suberoylanilide hydroxamic acid (SAHA), were added to normal saline and used as fluid for resuscitation. At the end of resuscitation, lung and liver tissues were subjected to subcellular protein fractionation and Western blotting to analyze histone acetylation. In addition, cDNA microarrays and RT-PCR were used to measure expression of selected genes. Hemorrhage did not change the level of histone acetylation in lungs, whereas resuscitation predominantly hyperacetylated histones. An analysis of histone acetylation on 10 lysine sites showed that L-LR, HTS, and KR resuscitation caused the largest number of changes (7, 6, and 6 respectively). SAHA hyperacetylated 7 sites in liver and affected expression of 57 genes (44 up, 13 down). Resuscitation with various fluids, as well as infusion of pharmacologic HDAC inhibitors affects histone acetylation in a fluid- and organ-specific fashion, even when administered post-insult for a limited period of time. Uniquely affected genes are associated with metabolism, cellular growth, proliferation, differentiation, transformation, and cellular signaling.

Differential effect of resuscitation on Toll-like receptors in a model of hemorrhagic shock without a septic challenge

It has been shown that the inflammatory response and cellular damage after hemorrhagic shock are influenced by resuscitation strategies. Toll-like receptors (TLRs) play an important role in signal transduction in inflammatory conditions. However, alterations in TLR expression following hemorrhagic shock and resuscitation have not been well documented. This study was conducted to measure the impact of different resuscitation strategies on TLR expression and downstream signaling in key organs. Sprague Dawley rats (n=38) were subjected to a severe volume-controlled hemorrhage protocol. After 75 min of shock, they were resuscitated over 45 min as follows: (1) lactated Ringer's (LR, 81 ml/kg), (2) ketone Ringer's (KR, 81 ml/kg), (3) 7.5% hypertonic saline (HTS, 9.7 ml/kg), (4) 6% hetastarch (HEX, 27 ml/kg), (5) pyruvate Ringer's (PR, 81 ml/kg). Sham hemorrhage (NH) and no resuscitation (NR) groups served as controls. The KR and PR solutions were identical to LR except for equimolar substitution of racemic lactate with beta hydroxybutyrate and sodium pyruvate, respectively. At the end of resuscitation, the expression of TLRs (types 1-10), and cytokines (IL-10, IL-1beta and TNF-alpha) were measured in the lung and spleen using RT-PCR. Levels of phosphorylated and total IkB-alpha and NF-kappaB were detected by Western blotting. The systemic and lung protein levels of TNF-alpha were measured using ELISA and immunohistochemistry. Expression of TLRs in the lung was affected more than in the spleen by hemorrhagic shock and resuscitation. In the lung, hemorrhage increased TLR-2, -3 and -6 (but not TLR-4) mRNA expression, with an up-regulation of the ratio of phosphor-NF-kappaBp65 and total NF-kappaBp65, NF-kappaBp65 activation, and enhanced systemic and tissue TNF-alpha protein levels. Post-resuscitation, TLR mRNA profile and subsequent downstream proteins in the lung and spleen were affected by the choice of resuscitation strategy. Hemorrhagic shock activates TLR signaling in lung, but not the spleen, probably through an up-regulation of TLR gene expression, and activation of NF-kappaB pathway. Resuscitation modulates this response in a fluid- and tissue-specific fashion.

Identification of Expression Patterns Associated with Hemorrhage and Resuscitation: Integrated Approach to Data Analysis

Although transcriptional profiling is a well-established technique, its application to systematic studying of various biological phenomena is still limited because of problems with high-volume data analysis and interpretation. This research project's objective was to create a comprehensive summary of changes in gene expression after hemorrhagic shock (HS), reliant and impartial of multiple variables, such as resuscitation treatments, organ analyzed, and time after impact. Rat model of severe (40% total blood loss) HS was employed. Hemorrhagic shock was treated with 6 different resuscitation strategies: (1) racemic lactated Ringer's (DL-LR); (2) L-lactated Ringer's (L-LR); (3) ketone Ringer's (KR); (4) pyruvate Ringer's (PR); (5) 6% hetastarch (Hex); (6) 7.5% hypertonic saline (HTS). Nonresuscitated and nonhemorrhaged rats served as controls. Ketone and pyruvate Ringer solutions were identical to the lactated Ringer's solution except for equimolar substitution of lactate with beta-hydroxybutyrate and sodium pyruvate, respectively. Total RNA from liver, lung, and spleen was isolated immediately (0 hour) and 24 hour postresuscitation. Each organ, time point and treatment was profiled using individual cDNA array (1,200 genes), to produce 183 separate data files. Methods of analysis included one-way and unbalanced factorial ANOVA, Sokal-Michener average linkage clustering and contextual mapping. : Unresuscitated HS produced the highest number (56) of upregulated expressions in spleen and lungs. HEX and HTS affected mostly pulmonary genes (22 and 9). Fourteen genes changed in response to combination of all three factors: treatment, organ, and time. Eighteen genes were identified as treatment-specific. Fifteen genes adjusted expression 24 hour post-treatment. The largest number of genes with altered expression (168) responded differently in all three organs. In this study 15 gene clusters were pinpointed. Contextual mapping identified novel and confirmed known pathways contributing to hemorrhage/resuscitation. We have reliably identified genes and pathways that are affected by HS and are responsive to resuscitation. Gene expression in various organs is affected differentially by HS, which can be further modulated by the choice of resuscitation strategy.

Cardiac histones are substrates of histone deacetylase activity in hemorrhagic shock and resuscitation

DNA transcription is regulated, in part, by acetylation of nuclear histones that are controlled by 2 groups of enzymes: histone deacetylases (HDAC) and histone acetyl transferases (HAT). Whether an imbalance in HDAC/HAT system plays a role in hemorrhage/resuscitation is unknown. The goals of this study were to determine whether hemorrhage results in deacetylation of cardiac histones and whether this can be corrected through the application of different resuscitation strategies or specific HDAC inhibitors. In the first experiment, rats (n = 6 per group) were subjected to volume-controlled hemorrhage and resuscitated with racemic lactated Ringer's solution, L-lactated Ringer's solution, 7.5% hypertonic saline solution, ketone Ringer's solution, and pyruvate Ringer's solution. Control groups included no hemorrhage (sham) and hemorrhage with no resuscitation. In the second experiment (n = 5 per group), 3 HDAC inhibitors (valproic acid, trichostatin A, and suberoylanilide hydroxamic acid) were added to saline solution resuscitation. Heart tissue was collected at the end of resuscitation. Isolated subcellular protein fractions were used in Western blotting to analyze the patterns of total protein acetylation and histone acetylation specifically. HDAC and HAT activity was measured in tissue extracts. Hemorrhage led to partial histone deacetylation. Resuscitation resulted in protein hyperacetylation in nuclear fractions only. A detailed analysis of histones (on 10 acetylation sites) revealed that ketone Ringer's solution hyperacetylated histones H2B, H3, and H4. The addition of suberoylanilide hydroxamic acid hyperacetylated histones more effectively than other resuscitation strategies, presumably by direct inhibition of HDAC activity. Hemorrhage/resuscitation is associated with HDAC/HAT activity misbalance, and the acetylation status of cardiac histones is influenced by the choice of resuscitation strategy. Shock-induced changes can be reversed through the infusion of pharmacologic HDAC inhibitor, even when it is administered after the insult for a limited period of time.

Hepatic Apoptosis after Hemorrhagic Shock in Rats Can Be Reduced Through Modifications of Conventional Ringer’s Solution

Resuscitation with racemic lactated Ringer's solution induces cellular apoptosis. This study was conducted to determine if the elimination of D-lactate isomer would attenuate apoptosis in the liver, and to investigate the possible mechanisms. Sprague Dawley rats (n=30, 5 per group) were subjected to modified volume-controlled hemorrhage and randomized to the following groups: no hemorrhage (sham); no resuscitation (NR); resuscitation with racemic lactated Ringer's (DL-LR); L-isomer LR (L-LR); ketone (beta-hydroxybuturate) Ringer's (KR); or pyruvate Ringer's (PR). Animals were sacrificed 2 hours later and expressions of proapoptotic proteins (BAD), antiapoptotic (bcl-2) proteins, and poly-ADP ribose polymerase (PARP) cleavage in liver were analyzed by Western blotting. Contribution of the phosphatidylinositol 3-kinase/serine/threonine kinase (PI3k/Akt) pathway was assessed by measuring total and phosphorylated PI3K, Akt, BAD, and endothelial nitric oxide synthase (eNOS) proteins. The terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) assay was used to detect the apoptotic cells. Liver ATP levels were measured using a luciferase reaction assay. Hemorrhage significantly decreased the hepatic ATP level and resuscitation improved it, but it returned to normal only in the L-isomer lactated Ringer's and ketone Ringer's groups. Expression of proapoptotic proteins was significantly lower in the pyruvate Ringer's and ketone Ringer's groups; L-isomer lactated Ringer's and pyruvate Ringer's resuscitation significantly increased bcl-2 expression. Poly-ADP ribose polymerase fragmentation and total number of apoptotic cells were significantly increased in the racemic lactated Ringer's group. There was no significant induction of Akt activity or changes in phosphorylated BAD, Akt, or eNOS levels. Resuscitation with racemic lactated Ringer's induces hepatic apoptosis, which is decreased if the D-isomer of lactate is eliminated. Apoptosis is reduced even more when lactate is substituted with beta-hydroxybutyrate or pyruvate. The beneficial effects are not through improvements in the energy status or activation of the PI3K/Akt pathway.

Energy Substrate-Supplemented Resuscitation Affects Brain Monocarboxylate Transporter Levels and Gliosis in a Rat Model of Hemorrhagic Shock

Monocarboxylate (MC)-supplemented resuscitation has been shown to attenuate cellular injury after hemorrhagic shock. However, little is known about its effect on the central nervous system. The brain can use MCs such as lactate, pyruvate, and beta-hydroxybutyrate as energy substrates. The transit of MCs into the central nervous system is facilitated by the monocarboxylate transporters (MCTs), and their blockage can exacerbate neuronal damage. We examined the expression of MCT1 and markers specific for activation of astroglia and microglia in the brains of rats subjected to hemorrhagic shock and resuscitation. The hypothesis was that resuscitation with MC-based fluids would be accompanied by MCT1 up-regulation and glial response. Rats (n = 30) were subjected to volume-controlled hemorrhage. Test groups included: sham, no resuscitation, resuscitation with normal saline, resuscitation with racemic lactated Ringer's solution, resuscitation with pyruvate Ringer's solution, and resuscitation with beta-hydroxybutyrate-containing ketone Ringer's solution. Plasma levels of MC were measured serially. The brains were investigated using GFAP, CD11b, CD43, MCT1, and GLUT1 immunohistochemistry. Rats resuscitated with MC-containing fluids had increased levels of MCT1 in brain endothelial cells and neuropil compared with sham rats. Enhanced staining was localized to the choroid plexus, astrocytic end feet, and white matter structures. None of the resuscitation treatment induced astrocytic hyperplasia, and pyruvate Ringer's solution and ketone Ringer's solution resuscitation led to hypertrophy of astrocytes. In hemorrhagic shock, resuscitation with MC-based fluids increased brain MCT1 level and led to activation of astrocytes. Enhanced MC trafficking could be an essential route for energy supply to neurons under adverse circulatory conditions.

Ketone and pyruvate Ringer's solutions decrease pulmonary apoptosis in a rat model of severe hemorrhagic shock and resuscitation

Background. Resuscitation fluids containing β-hydroxybutyrate (BHB) have been shown to decrease cellular injury after hemorrhagic shock and resuscitation through an unknown mechanism. We tested whether this effect was related to BHB-induced metabolic modulations. Methods. Male Sprague Dawley rats (n=30) were subjected to volume-controlled hemorrhage (27 mL/kg during 10 minutes followed by 75 minutes of shock during which another 8 mL/kg of blood was withdrawn). Experimental groups included the following: (1) sham, (2) no resuscitation (NR), (3) racemic lactated Ringer's (DL-LR) solution, (4) LR containing L-isomer only (L-LR), (5) ketone Ringer's solution with lactate substituted by BHB (KR), and (6) pyruvate Ringer's solution with lactate substituted by pyruvate (PR). The resuscitation fluids were infused during 45 minutes simultaneously with additional hemorrhage of 8 mL/kg. Hemodynamic and physiologic parameters and the plasma levels of BHB were serially measured. The animals were killed 2 hours after resuscitation, and tissues were frozen instantaneously for cellular adenylate extraction and adenosine triphosphate (ATP) and adenosine diphosphate analysis. Pulmonary apoptosis was studied using Western blotting, immunohistochemistry, and reverse transcriptase-polymerase chain reaction. Expression of enzymes involved in ketogenesis and ketolysis was analyzed by reverse transcriptase-polymerase chain reaction. Results. NR and resuscitation with DL-LR increased the expression of apoptotic markers, whereas resuscitation with KR and PR significantly decreased the expression of apoptotic markers in rat lungs. Resuscitation with KR was followed by a profound increase in plasma BHB levels; however, the expression levels of ketolytic enzymes were essentially unaffected. KR infusion did not induce significant improvements in tissue ATP levels. Conclusion. Resuscitation with KR and PR protects against pulmonary apoptosis without improving tissue ATP content. Therefore, metabolic modulation is unlikely to be the major mechanism by which BHB exerts its protective effects during reperfusion. (Surgery 2003;134:267-74.)