Hypertonic Saline Resuscitation Research Articles

Hypertonic saline resuscitation enhances blood pressure recovery and decreases organ injury following hemorrhage in acute alcohol intoxicated rodents

Acute alcohol intoxication (AAI) impairs the hemodynamic and arginine vasopressin (AVP) counter-regulation to hemorrhagic shock (HS) and lactated Ringer's solution (LR) fluid resuscitation (FR). The mechanism of AAI-induced suppression of AVP release in response to HS involves accentuated nitric oxide (NO) inhibitory tone. In contrast, AAI does not prevent AVP response to increased osmolarity produced by hypertonic saline (HTS) infusion. We hypothesized that FR with HTS during AAI would enhance AVP release by decreasing periventricular nucleus NO inhibitory tone, subsequently improving mean arterial blood pressure (MABP) and organ perfusion. Male Sprague-Dawley rats received a 15-hour alcohol infusion (2.5 g/kg + 0.3 g/kg/h) or dextrose (DEX) before HS (40 mm Hg × 60 minutes) and FR with HTS (7.5%, 4 ml/kg) or LR (2.4 × blood volume removed). Organ blood flow was determined, and brains were collected for NO content at 2 hours after FR. HTS improved MABP recovery in AAI (109 vs. 80 mm Hg) and DEX (114 vs. 83 mm Hg) animals compared with LR. This was associated with higher (>60%) circulating AVP levels at 2 hours after FR compared with those detected in LR animals in both groups. Neither AAI alone nor HS in DEX animals resuscitated with LR altered organ blood flow. In AAI animals, HS and FR with LR reduced blood flow to the liver (72%), small intestine (65%), and large intestine (67%) compared with shams. FR with HTS improved liver (threefold) and small intestine (twofold) blood flow compared with LR in AAI-HS animals. The enhanced MABP response to HTS was prevented by pretreatment with a systemic AVP V1a receptor antagonist. HTS decreased periventricular nucleus NO content in both groups 2 hours after FR. These results suggest that FR with HTS in AAI results in the removal of central NO inhibition of AVP, restoring AVP levels and improving MABP and organ perfusion in AAI-HS.

Research studies that have influenced practice of neuroanesthesiology in recent years: A literature review

Through evolving research, recent years have witnessed remarkable achievements in neuromonitoring and neuroanesthetic techniques, with a huge body of literature consisting of excellent studies in neuroanaesthesiology. However, little of this work appears to be directly important to clinical practice. Many controversies still exist in care of patients with neurologic injury. This review discusses studies of great clinical importance carried out in the last five years, which have the potential of influencing our current clinical practice and also attempts to define areas in need of further research. Relevant literature was obtained through multiple sources that included professional websites, medical journals and textbooks using key words “neuroanaesthesiology,” “traumatic brain injury,” “aneurysmal subarachnoid haemorrhage,” “carotid artery disease,” “brain protection,” “glycemic management” and “neurocritical care.” In head injured patients, administration of colloid and pre-hospital hypertonic saline resuscitation have not been found beneficial while use of multimodality monitoring, individualized optimal cerebral perfusion pressure therapy, tranexamic acid and decompressive craniectomy needs further evaluation. Studies are underway for establishing cerebroprotective potential of therapeutic hypothermia. Local anaesthesia provides better neurocognitive outcome in patients undergoing carotid endarterectomy compared with general anaesthesia. In patients with aneurysmal subarachnoid haemorrhage, induced hypertension alone is currently recommended for treating suspected cerebral vasospasm in place of triple H therapy. Till date, nimodipine is the only drug with proven efficacy in preventing cerebral vasospasm. In neurocritically ill patients, intensive insulin therapy results in substantial increase in hypoglycemic episodes and mortality rate, with current emphasis on minimizing glucose variability. Results of ongoing multicentric trials are likely to further improvise our practice.

The Cardioprotective Effect of Hypertonic Saline Is Associated with Inhibitory Effect on Macrophage Migration Inhibitory Factor in Sepsis

Sepsis can cause myocardial dysfunction, which contributes to the high mortality of sepsis. Hypertonic saline (HS) has been reported to increase myocardial contractility in sepsis. In the present study, mechanisms of action of HS resuscitation (4 mL of 7.5% NaCl per kilogram) on cardiac function have been evaluated in septic rats. HS was administered 1 h after LPS (10 mg/kg, i.v.) challenge. The mean arterial blood pressure significantly decreased 4 h after LPS challenge, and septic shock was observed at the end of experiment (6 h). Posttreatment with HS prevented hypotension caused by LPS and significantly improved cardiac function, evidenced by increases in left ventricular developed pressure, mean +dP/dt and −dP/dt. The amplitude of electrical-stimulated intracellular Ca2+ transient in isolated single cardiomyocytes was significantly reduced after 6 h LPS insult, which was recovered by HS. In addition, LPS resulted in significant increases in neutrophil myeloperoxidase activity, macrophage migration inhibitory factor (MIF), and NF-κB phospho-p65 protein levels in myocardium at 6 h, which were significantly attenuated by HS. In conclusion, HS improved myocardial contractility and prevented circulatory failure induced by endotoxemia, which may attribute to improvement of intracellular calcium handling process and inhibitory effects on neutrophil infiltration and MIF production in hearts.

The effectiveness of hypertonic saline and pentoxifyline (HSPTX) resuscitation in hemorrhagic shock or sepsis tissue injury; comparison with the LR, HES-HTS, LR-PTX treatmentsStatus: PendingCategory: Trauma/Disaster Med

The effectiveness of hypertonic saline and pentoxifyline (HSPTX) resuscitation in hemorrhagic shock or sepsis tissue injury; comparison with the LR, HES-HTS, LR-PTX treatmentsStatus: PendingCategory: Trauma/Disaster Med

Damage Control Immunoregulation: Is There a Role for Low-Volume Hypertonic Saline Resuscitation in Patients Managed with Damage Control Surgery?

Hypertonic saline (HTS) is beneficial in the treatment of head-injured patients as a result of its potent cytoprotective effects on various cell lines. We hypothesize that low-volume resuscitation with 3 per cent HTS, when used after damage control surgery (DCS), improves outcomes compared with standard resuscitation with isotonic crystalloid solution (ICS). This is a 4-year retrospective review from two Level I trauma centers. Patients included had 10 units or more of packed red blood cells during initial DCS. On arrival to the trauma intensive care unit (TICU), patients were resuscitated with low-volume 3 per cent HTS or with conventional ICS. A cohort analysis was performed comparing resuscitation strategies. Univariate analysis of continuous data was done with Student t test followed by multivariate analysis. Of 188 patients included, 76 were in the low-volume HTS group and 112 in the ICS group. Demographics were similar between the groups. Over the next 48 hours after DCS in HTS versus ISC groups, intravenous fluids were given: 1920 ± 455 mL versus 8400 ± 1200 mL (P < 0.0001); urine output was 4320 ± 480 mL versus 1940 ± 480 mL(P < 0.0001); mean TICU length of stay was 10 ± 8 versus 16 ± 15 days (P < 0.01); prevalence of acute respiratory distress syndrome was 4.0 versus 13.4 per cent (P = 0.02); sepsis was 6.6 versus 15.2 per cent (P = 0.06); multisystem organ failure was: 2.6 versus 16.1 per cent (P < 0.01); and 30-day mortality was 5.3 versus 15.2 per cent (P = 0.03). There was no difference for prevalence of renal failure at 5.3 versus 3.6 per cent (P = 0.58). Low-volume resuscitation with HTS administered after DCS on arrival to the TICU may have a protective effect on the polytrauma patient. We believe that this study demonstrates a role for low-volume resuscitation with HTS to improve outcomes in patients undergoing DCS.

Changes in lymph proteome induced by hemorrhagic shock

Damage-associated molecular patterns (DAMPs) released from host tissue after trauma and hemorrhagic shock (HS) have been shown to activate polymorphonuclear cells (PMNs) and lead to acute lung injury and systemic inflammatory response syndrome. The avenue by which DAMPs reach the circulation is unclear; however post-HS lymph has been shown to contain biologically active mediators. We therefore studied the time course of DAMP detection in systemic lymph and the effect of isotonic versus hypertonic resuscitation on DAMPs production and PMN activation in vitro. A canine HS/hind-limb lymph cannulation model was used. Animals were bled to a mean arterial pressure of 40 mm Hg and were resuscitated with shed blood plus equivalent amounts of Na+as either lactated Ringer's solution or 7.5% hypertonic saline solution (HSS). Lymph samples were collected at baseline, end-shock, and at various times after resuscitation. DAMPs were isolated from lymph samples and detected by Western blot for high-mobility group box 1 and mitochondrial DNA. Priming of naive PMNs was indexed by mitogen-associated protein kinase phosphorylation. Human pulmonary microvascular endothelial cell monolayers were established and exposed to the various lymph samples. Endothelial intracellular adhesion molecule expression, apoptosis, and monolayer permeability were determined. DAMPs were detected in lymph samples starting at the end of the shock period and peaking at 120 minutes after resuscitation. HSS resuscitation resulted in the highest levels of DAMPs detected in systemic lymph and plasma. PMN mitogen-associated protein kinase activation was noted during the resuscitation phase and peaked 120 minutes after resuscitation. Similar temporal changes in human pulmonary microvascular endothelial cell intracellular adhesion molecule expression and cellular injury were noted after shock with the greatest effect noted with the hypertonic saline resuscitation regimen. Lymph represents an important avenue for the delivery of DAMPs into the systemic circulation after HS. HSS lead to a significant increase in DAMPs production in the model. This finding may account for the conflicting data regarding the salutary effects of HSS resuscitation noted in clinical versus experimental shock studies. ).

The effectiveness of hypertonic saline and pentoxifylline (HTS–PTX) resuscitation in haemorrhagic shock and sepsis tissue injury: Comparison with LR, HES, and LR–PTX treatments

PurposeTo compare lung and liver injury and laboratory results in haemorrhagic shock and sepsis models treated with combinations of lactated Ringer's solution (LR), 7.5% hypertonic saline (HTS), hydroxyethyl starch (HES), and pentoxifylline (PTX). MethodsMale Sprague-Dawley rats (200–290g) were assigned randomly to one of four treatment groups (n=16 per group): (1) LR; (2) HES; (3) LR–PTX; and (4) HTS–PTX. Each group was subdivided into (1) haemorrhagic shock (n=8) and (2) sepsis (n=8) model groups. A venous catheter was used to inject resuscitation fluids, and an arterial catheter was used to withdraw blood and monitor mean arterial pressure (MAP). Lung and liver histology, bronchoalveolar lavage (BAL) fluid, and cytokine levels were evaluated. ResultsThe mean lung injury score was 1.7. At 24h after treatment, the total leucocyte count in the BAL fluid was significantly (p<0.05) higher with LR treatment (10×106±0.8) than with other treatments in the sepsis model groups (HES, 6×106±1.2; LR–PTX, 5×106±1.5; HTS–PTX, 5×106±0.6). The higher total leucocyte count after LR treatment was attributable to a greater increase in the number of neutrophils (17±1.5%) compared with increases after the other treatments (HES, 6±0.8%; LR–PTX, 10±1.3%; HTS–PTX, 5±0.4%). In the sepsis model groups, the total hepatic injury score was also significantly (p<0.05) higher with LR treatment (9.9±0.5) than with the other treatments (HES, 6.7±0.8; LR–PTX, 5.6±0.7; HTS–PTX, 3.1±0.9). This also occurred in the shock model (LR, 10.6±2.1; HES, 5.8±0.9; LR–PTX, 7.3±0.9; HTS–PTX, 3.5±0.9). As compared with LR treatment, HTS–PTX resuscitation resulted in a 49% decrease in TNF-α, 29% decrease in IL-1β, and 58% decrease in IL-6 in the shock model at 24h (p<0.05), and the respective decreases were 45, 24, and 35% in the sepsis model (p<0.05). ConclusionHTS–PTX was superior to HES, LR–PTX, and LR for treating shock and sepsis, and LR–PTX and HES gave better results than LR therapy alone.

Beneficial Effect of Hypertonic Saline Resuscitation in a Porcine Model of Severe Acute Pancreatitis

Hypertonic saline (HTS) solution resuscitation has been used in a variety of clinical settings. The aim of this study was to assess the effect of HTS resuscitation on hemodynamics, systemic oxygenation, and organ damage in a porcine model of severe acute pancreatitis. Eighteen anesthetized and mechanically ventilated pigs were divided into 3 groups: HTS group, lactated Ringer solution (LR) group, and sham-operated group. Severe acute pancreatitis was induced in the first 2 groups by injecting 5% sodium taurocholate into the pancreatic duct, and the investigation period was 12 hours. Hemodynamic parameters, urine output, oxygenation parameters, and serum parameters were recorded consecutively. Finally, histologic examinations of the kidney, intestine, pancreas, and lung were performed. In the HTS group, cardiac output decreased less significantly compared with the LR group. Furthermore, aspartate aminotransferase, creatinine, and lactate levels increased significantly in all animals with severe acute pancreatitis, but the increasing tendency was slower in the HTS group. Nevertheless, the histopathologic analysis revealed similar injuries of the kidney, intestine, pancreas, and lung between the HTS and LR groups. Early administration of HTS generally improves hemodynamics and peripheral oxygenation. Despite these normalized parameters, organ damage could not be diminished to a significant degree during observation.

Bolus hypertonic or normal saline resuscitation in gram-negative sepsis: systemic and regional haemodynamic effects in sheep

Bolus hypertonic or normal saline resuscitation in gram-negative sepsis: systemic and regional haemodynamic effects in sheep

Increased Neutrophil Adenosine A3 Receptor Expression Is Associated With Hemorrhagic Shock and Injury Severity in Trauma Patients

Hypertonic saline (HS) has been investigated as an immune modulator following hemorrhagic shock and sepsis. The polymorphonuclear neutrophil (PMN) response to HS is regulated by the release of ATP, which is converted to adenosine and activates adenosine receptors. Binding to A3 adenosine receptors promotes PMN activation, and inhibition of A3 receptors improves the efficacy of HS resuscitation. A3 receptor expression of PMNs has not been previously evaluated in injured patients. Whole blood was obtained from 10 healthy volunteers and 60 injured patients within 2 h of injury. Inclusion criteria were blunt or penetrating injury with evidence of hypovolemic shock (systolic blood pressure [SBP] ≤90 mmHg and base deficit ≥6 mEq/L or need for blood transfusion) or evidence of severe traumatic brain injury including initial Glasgow Coma Scale score of 8 or less or evidence of traumatic brain injury on head computed tomography scan (head Abbreviated Injury Score ≥3) or intubation in the field or emergency department. A3 receptor expression was assessed by flow cytometry. Polymorphonuclear neutrophils were also exposed to fMLP or HS (20-40 mM) in vitro. Clinical data were collected including admission physiology, injury severity (Injury Severity Score [ISS]), development of multiple organ failure, and survival. In normal volunteers, less than 1% of PMNs expressed A3 receptors on the cell surface. A3 receptor expression was significantly higher in injured patients, and the level of expression correlated with the severity of injury (ISS ≥25: A3 positive PMN 36.6% vs. ISS <25: 16.2%; P = 0.019) and degree of hypovolemic shock (SBP ≤90 mmHg: A3 positive PMN 43.8% vs. SBP>90 mmHg: 20.6%; P = 0.008). Stimulation with fMLP or HS increased A3 expression in normal volunteers, but only in patients with ISS of less than 25 or without hypovolemic shock. A3 receptor expression on the surface of PMNs is upregulated by injury, and increased expression levels are associated with greater injury severity and hypovolemic shock. Hypertonic saline increases A3 expression of PMNs from healthy volunteers and less severely injured patients.

Hypertonic Saline Resuscitation of Hemorrhagic Shock Does Not Decrease In Vivo Neutrophil Interactions With Endothelium in the Blood-Brain Microcirculation

Resuscitation of hemorrhagic shock with isotonic crystalloids has been shown to activate polymorphonuclear neutrophils (PMNs). Although hypertonic saline (HTS) can reduce PMN activation and interactions with endothelial cells (EC) in systemic microvascular beds, no data exist demonstrating that the same occurs in the unique blood-brain barrier microcirculation. We hypothesized that resuscitation of hemorrhagic shock with HTS would blunt brain in vivo PMN-EC interactions. Wistar rats (250-350 g) underwent craniotomy and placement of a window for live intravital viewing of pial vessels. Twenty animals were bled to a mean arterial pressure of 30 mm Hg to 35 mm Hg for 1 hour and resuscitated with shed blood and either 5% HTS (6 mL/kg) or Ringer's lactate (RL) (2× shed blood volume). Circulating rhodamine-6G-labeled PMN in pial venules were captured by videomicroscopy at baseline (preshock), end of the shock period, after resuscitation, and every 15 minutes to 30 minutes for 2 hours. Hemodynamics and arterial gases were monitored. Off-line footage analysis allowed comparisons of PMN-EC interactions between groups. Animals in both groups developed significant metabolic acidosis (p < 0.01) after hemorrhage, but postresuscitation blood pressures were similar at all time points. Crystalloid resuscitation volumes were 10× greater in RL than HTS animals (p < 0.001). For all time points, we did not observe the expected reduction in PMN rolling and adhesion in HTS animals, instead noted trends of consistently lower interactions in RL counterparts. In contradistinction to studies evaluating the systemic microcirculation, HTS may activate PMN-EC crosstalk in the blood-brain microcirculation. Further studies are needed to analyze whether this effect is due to the unique nature of the blood-brain interface.

The progress of perioperative fluid therapy Strategy

Background Adopting appropriate fluid therapy strategy for perioperative patients is essential to perioperative management, which could reduce perioperative complications, shorten hospitalization days and prognosis of postoperative patients. Objective In order to provide evidence for clinical selection of fluid therapy strategy, we reviewed the history and the progress of perioperative fluid therapy strategy and generate current best evidence of some controversial issues. Content We reviewed the history and the progress of perioperative fluid therapy strategy, including early goal directed therapy, the strategy of fast track surgery, acute high-capacity perfusion during anesthesia induction and small volume hypertonic saline resuscitation. We also summarized some controversial issues such as the selection of fluid for resuscitation and the security of artificial colloid. Trend To achieve the fundamental goal of improve the organ perfusion and oxygen delivery to protect organ function, future study are warranted to the technology and approach of fluid assessment and monitoring. It is urgent to explore new resuscitation fluid which has reliable effect with mild adverse reactions. Key words: Perioperative period; Fluid therapy; Crystal; Colloid

A3 Adenosine Receptor Inhibition Improves the Efficacy of Hypertonic Saline Resuscitation

We reported previously that hypertonic saline (HS) treatment can prevent or upregulate the function of polymorphonuclear neutrophils (PMNs) via A2a-type adenosine receptors or A3-type adenosine receptors (A3R), respectively. A3R translocate to the cell surface upon PMN stimulation, and thus, HS promotes PMN responses under conditions of delayed HS treatment. Here we investigated if inhibition of A3R improves the protective effects of HS resuscitation in a mouse sepsis model. We found that HS nearly triples extracellular adenosine concentrations in whole blood and that inhibition of A3R with the selective antagonist MRS-1191 dose dependently improves the inhibitory effect of HS. MRS-1191 at a concentration of 1 nM enhanced the inhibitory effect of HS and reduced stimulatory effects of delayed HS treatment. Using a mouse model of cecal ligation and puncture (CLP)-induced sepsis, we found that MRS-1191 reduces acute lung injury and PMN accumulation in lung tissue. Whereas delayed HS treatment (4 mL/kg of 7.5% NaCl) of mice 1 h after CLP aggravated PMN accumulation, lung tissue damage, and mortality 24 h after CLP, infusion of MRS-1191 (2 ng/kg body weight) combined with HS reduced these detrimental effects of delayed HS treatment. Our data thus show that A3 receptor antagonists can strengthen the beneficial effects of HS resuscitation by avoiding stimulatory adverse effects that result from delayed HS administration.

Hypertonic Saline Enhances Host Defense and Reduces Apoptosis in Burn Mice by Increasing Toll-Like Receptors

Hypertonic saline (HTS) is useful in the management of intracranial hypertension and shock patients. The aim of this study was to investigate whether HTS enhances host defense in burn mice through the increase of Toll-like receptors (TLRs) and nuclear factor κB (NF-κB) activation. C57BL/6, TLR4, C3H/HeN, and C3H/HeJ (nonfunctional TLR4 mutant) mice underwent burn and were given 10 mL/kg HTS (7.5% NaCl, 1.28 mol/L), 10 mL/kg saline (154 mmol/L), or 80 mL/kg saline (154 mmol/L) at 8 h after burn. At 24 h after burn, mesenteric lymph nodes were harvested and assayed for bacterial translocation (BT). Next, animals received i.p. Escherichia coli challenge, and bacterial clearance was measured. Finally, peritoneal cells were isolated for assay of bacterial killing activity, phagocytic activity, apoptotic ratio, NF-κB DNA binding activity, and expression of TLR4, MyD88, p-Akt, pp38, macrophage inflammatory protein 2, and Bcl-xL. Hypertonic saline decreased BT in C57BL/6 and C3H/HeN mice but not in TLR4 mutant mice. Also, HTS increased bacterial clearance and bacterial killing activity and decreased apoptotic ratio of peritoneal cells from C57BL/6 and C3H/HeN mice but not TLR4 or C3H/HeJ mice. Finally, HTS increased NF-κB activity and expression of TLR4, MyD88, p-Akt, pp38, macrophage inflammatory protein 2, and Bcl-xL in C57BL/6 but not in TLR4 mice. Hypertonic saline increases bacterial clearance and bacterial killing activity and decreases thermal injury-induced BT in wild-type but not in TLR4 mutant mice. Given that HTS induces NF-κB activity and TLR4, MyD88, and pp38 expression but decreases the apoptosis of inflammatory cells, we conclude that HTS resuscitation enhances host defense against bacterial challenge and reduces apoptosis of inflammatory cells in burn mice by increasing TLR4 expression and NF-κB activation.

Hypertonic sodium resuscitation after hemorrhage improves hemodynamic function by stimulating cardiac, but not renal, sympathetic nerve activity

Small volume hypertonic saline resuscitation can be beneficial for treating hemorrhagic shock, but the mechanism remains poorly defined. We investigated the effects of hemorrhagic resuscitation with hypertonic saline on cardiac (CSNA) and renal sympathetic nerve activity (RSNA) and the resulting cardiovascular consequences. Studies were performed on conscious sheep instrumented with cardiac (n=7) and renal (n=6) sympathetic nerve recording electrodes and a pulmonary artery flow probe. Hemorrhage (20 ml/kg over 20 min) caused hypotension and tachycardia followed by bradycardia, reduced cardiac output, and abolition of CSNA and RSNA. Resuscitation with intravenous hypertonic saline (1.2 mol/l at 2 ml/kg) caused rapid, dramatic increases in mean arterial pressure, heart rate, and CSNA, but had no effect on RSNA. In contrast, isotonic saline resuscitation (12 ml/kg) had a much delayed and smaller effect on CSNA, less effect on mean arterial pressure, no effect on heart rate, but stimulated RSNA, although the plasma volume expansion was similar. Intracarotid infusion of hypertonic saline (1 ml/min bilaterally, n=5) caused similar changes to intravenous administration, indicating a cerebral component to the effects of hypertonic saline. In further experiments, contractility (maximum change in pressure over time), heart rate, and cardiac output increased significantly more with intravenous hypertonic saline (2 ml/kg) than with Gelofusine (6 ml/kg) after hemorrhage; the effects of hypertonic saline were attenuated by the β-receptor antagonist propranolol (n=6). These results demonstrate a novel neural mechanism for the effects of hypertonic saline resuscitation, comprising cerebral stimulation of CSNA by sodium chloride to improve cardiac output by increasing cardiac contractility and rate and inhibition of RSNA.

24 Comparison of Normal Saline, Hypertonic Saline and Hypertonic Colloid Resuscitation Fluids in An Infant Animal Model of Hemorrhagic Hypovolemic Shock

Background: Few studies have analyzed the effect of resuscitation fluid composition in infant animal models of hemorrhagic-hypovolemic shock. Our hypothesis was that hypertonic and hyperoncotic fluid will achieve goal-directed hemodynamic endpoints with less volume infusion than normal saline. Methods: Prospective, randomized study in 34 two month-old piglets (9.9±2kg). Following anaesthesia, paralysis and mechanical ventilation, hypovolemia was induced by controlled 30 ml/kg bleed. After 30 min, pigs were randomized to receive: Normal Saline (S0.9%) 30 ml/kg, n=11, Hypertonic 3% Saline (S3%) 15 ml/kg n=12, or Albumin 5% plus Hypertonic 3% Saline (A5%-S3%) 15 ml/kg, n=11. Hemodynamic (heart rate (HR), mean arterial pressure (MAP), cardiac index (CI)), brain tissue oxygenation by near infrared spectroscopy (bStO2) and tissue perfusion (arterial lactate and gastric pHi) parameters were compared by ANOVA. Results: 30 min. after bleeding, high baseline HR: 192±37, and low MAP 72±19 mmHg, CI 2.8±1L/min/m2, bStO2: 44±8%, gastric pHi 7.1±0.2 and lactate 3.4 ±2.3 mmol/L were recorded with non significant differences between groups. 30 min after infusion there were non significant differences between groups for HR (S0.9%: 188±14, S3%: 184±14, A5%-S3%: 151±14 bpm); MAP (S0.9%: 80±7, S3%: 86±7, A5%-S3%: 87±7 mmHg); CI (S0.9%: 4.9±0.5, S3%: 5.1±0.4, A5%-S3%: 5.3±0.4 ml/min/m2); lactate (S0.9%: 2.8±0.7, S3%: 2.3±0.6, A5%-S3%: 2.5±0.6mmol/l); bStO2 (S0.9%: 43.9±2.2, S3%: 40.1±2.5, A5%-S3%: 45.7±2.3 %); and pHi (S0.9%: 7.07±0.6, S3%: 7.25±0.5, A5%-S3%: 7.23±0.5). Conclusion: In this model of pediatric hypovolemic shock, both hypertonic (S3%) and hyperoncotic (A5%- S3%) fluids achieved similar hemodynamic endpoints, requiring half of the volume compared to normal saline.

Hypertonic stress regulates T cell function via pannexin-1 hemichannels and P2X receptors

Hypertonic saline (HS) resuscitation increases T cell function and inhibits posttraumatic T cell anergy, which can reduce immunosuppression and sepsis in trauma patients. We have previously shown that HS induces the release of cellular ATP and enhances T cell function. However, the mechanism by which HS induces ATP release and the subsequent regulation of T cell function by ATP remain poorly understood. In the present study, we show that inhibition of the gap junction hemichannel pannexin-1 (Panx1) blocks ATP release in response to HS, and HS exposure triggers significant changes in the expression of all P2X-type ATP receptors in Jurkat T cells. Blocking or silencing of Panx1 or of P2X1, P2X4, or P2X7 receptors blunts HS-induced p38 MAPK activation and the stimulatory effects of HS on TCR/CD28-induced IL-2 gene transcription. Moreover, treatment with HS or agonists of P2X receptors overcomes T cell suppression induced by the anti-inflammatory cytokine IL-10. These findings indicate that Panx1 hemichannels facilitate ATP release in response to hypertonic stress and that P2X1, P2X4, and P2X7 receptor activation enhances T cell function. We conclude that HS and P2 receptor agonists promote T cell function and thus, could be used to improve T cell function in trauma patients.

Effects of Hypertonic Saline on CD4 +CD25 +Foxp3 + Regulatory T Cells After Hemorrhagic Shock in Relation to iNOS and Cytokines

Hemorrhagic shock and resuscitation induce immunosuppression. CD4(+)CD25(+)Foxp3(+) regulatory T Cells (Foxp3(+) Tregs), iNOS and cytokines may affect these severe conditions such as acute respiratory distress syndrome and multiple organ failure after hemorrhagic shock and resuscitation. Foxp3(+) Tregs have been described to be specific and play a key role in the control of the immune system. Immune condition may be restored by hypertonic saline resuscitation that inhibits pro-inflammatory effects of cytokine. Our aim was to investigate how hypertonic saline resuscitation affected Foxp3(+) Tregs after hemorrhagic shock and resuscitation in relation to iNOS and cytokines. Male C57BL6/J and B6.129P2- NOS2(tm1Lau)/J (iNOS gene knockout) mice were used in creating hemorrhagic shock model. Mice were divided into two groups, each according to the type of resuscitation. (1) Wild HS: resuscitation with hypertonic saline (4 mL/Kg of 7.5% NaCl) and the shed blood (SB); (2) wild 2LR: resuscitation with lactated Ringer's solution and the SB; (3) iNOS knockout HS: similarly resuscitated as wild HS; (4) iNOS knockout 2LR: similarly resuscitated as wild 2LR. Samples of thymus and spleen were harvested at 2, 6, 24, 48, and 72 h after resuscitation. CD4(+) T cells and Foxp3(+) Tregs were analyzed at 24, 48, and 72 h. At 2, 6, 24, and 48 h, plasma cytokines were assayed and expression of iNOS (NOS2) was also measured by immunofluorescence. NOS2 of HS and 2LR wild groups at 2 and 6h in spleen increased compared with the control group. At 6h, NOS2 in HS wild group was significantly lower than in 2LR wild group. Plasma levels of interleukin (IL)-6, TNF- α, MCP-1, and IL-10 increased at 2h. Both in wild type and iNOS knockout mice, hypertonic saline resuscitation decreased plasma IL-6, TNF-α, and MCP-1 levels at 2 h; CD4(+) T cells in spleen and thymus decreased at 24, 48, and 72 h, and Foxp3(+) Tregs in spleen at 48 h increased, however, hypertonic saline resuscitation did not affect the Foxp3(+). These results show that in early phase, the inflammatory cytokines in plasma might affect iNOS expression and cytokines. Further, this study showed that hypertonic saline resuscitation and suppression of iNOS might improve immunosuppressive reaction after hemorrhagic shock.

Effect of Hypertonic Saline Resuscitation on CD4+CD25+ Regulatory T Cells and γδ T Cells After Hemorrhagic Shock and Resuscitation in Relation to Apoptosis and iNOS

Hemorrhagic shock and resuscitation induce immunosuppression. CD4CD25 regulatory T cells and gammadeltaT cells may affect these immunosuppressive conditions. Hypertonic saline resuscitation reduces damage to organs and apoptosis and also restores immunosuppressive condition. We investigated how hypertonic saline resuscitation affected the induction of CD4CD25 regulatory T cells and gammadeltaT cells, and their apoptosis after hemorrhagic shock and resuscitation, and its relationship to inducible nitric oxide synthase (iNOS) (nitric oxide production). Male inbred C57BL6/J mice 8-week to 12-week-old as wild type and iNOS gene knock out (iNOS-/-), weighing 20 g to 35 g, were used. Hemorrhagic shock model of +/-40 mm Hg for 60 minutes was setup. Animals were randomly assigned to the following four resuscitation group: (1) wild HS: resuscitation with hypertonic saline (4 mL/Kg of 7.5% NaCl) and shed blood (SB), (2) wild 2LR: resuscitation with lactated Ringer's solution (two times the volume of the SB) and SB, (3) iNOS knockout HS, and (4) iNOS knockout 2LR. Untreated groups for wild and iNOS knockout mice were designated as control groups. Samples of thymus and spleen were harvested at 2 hours, 6 hours, 24 hours, and 48 hours after resuscitation. CD4CD25 regulatory T cells and gammadeltaT cells were analyzed using three-color flow cytometry. (1) gammadelta T cells increased earlier at 24 hours and CD4CD25 regulatory T cells increased later at 48 hours compared with controls in spleen of wild type (p < 0.01). (2) Hypertonic saline resuscitation suppressed gammadelta T cells compared with 2LR at 24 hours in iNOS knockout mice in spleen (p < 0.05). Hypertonic saline resuscitation increased apoptosis of CD4CD25 regulatory T cells at 48 hours in iNOS knockout mice in spleen (p < 0.01). (3) CD4CD25 regulatory T cells of iNOS knockout both in HS and 2LR groups at 48 hours decreased compared with wild type both in HS and 2LR groups in spleen (p < 0.01). (4) Apoptotic gammadelta T cells both in spleen and thymus in iNOS knockout mice at 48 hours increased compared with those in wild type (p < 0.05, respectively, except gammadelta T cells 2LR in spleen: p = 0.058). gammadelta T cells increased earlier at 24 hours, whereas CD4CD25 regulatory T cells increased later at 48 hours in spleen of wild type. Hypertonic saline was effective without the presence of iNOS, i.e., decreased gammadelta T cells at 24 hours and increased apoptosis of CD4CD25 regulatory T cells at 48 hours. CD4CD25 regulatory T cells at 48 hours without iNOS decreased compared with those of wild type. gammadelta T cells at 48 hours induced apoptosis under the condition without iNOS in spleen and thymus. iNOS worked as an accelerating factor for immunosuppressive condition, affected apoptosis, and immunoenhancing effect by hypertonic saline.

AFFERENT PATHWAYS INVOLVED IN CARDIOVASCULAR ADJUSTMENTS INDUCED BY HYPERTONIC SALINE RESUSCITATION IN RATS SUBMITTED TO HEMORRHAGIC SHOCK

The peripheral hyperosmolarity elicited by intravenous infusion of hypertonic saline (HS) can be beneficial in treating hemorrhagic shock. However, the neural mechanisms involved in this resuscitation remain unknown. The present study sought to determine the effects of selective baroreceptor denervation on arterial blood pressure response during HS resuscitation in rats submitted to hemorrhagic shock. Male Wistar rats (280-320 g) were anesthetized with thiopental sodium (40 mg/kg, i.v.), and the femoral artery and jugular vein were cannulated for MAP and heart rate recording and HS infusion (3 mol/L NaCl; 0.18 mL/100 g body weight, >2 min). Hemorrhagic shock was obtained by withdrawing blood over 30 min until a MAP of 60 mmHg was obtained. This level of MAP was maintained for a further 30 min through subsequent blood withdrawal or reinfusion. Next, animals were divided into selective aortic and/or carotid denervation or sham groups before infusing HS. Results showed that in the sham group (n = 12), HS infusion increased MAP to levels close to baseline (from 65 +/- 3 to 112 +/- 5 mmHg, 10 min after HS). In the aortic denervated group (n = 10), HS infusion also increased MAP (from 54 +/- 3 to 112 +/- 5 mmHg, 10 min after HS). In contrast, in the carotid denervation group (n = 8), the increase in MAP induced by HS infusion was abolished (from 53 +/- 3 to 73 +/- 12 mmHg, 10 min after HS). These results indicate that in hemorrhaged rats, HS infusion produces a pressor effect that is likely to be mediated through carotid rather than aortic baroreceptors.