Pentoxifylline enhances the protective effects of hypertonic saline solution on liver ischemia reperfusion injury through inhibition of oxidative stress

BACKGROUND：Liver ischemia reperfusion（IR）injury triggers a systemic inflammatory response and is the main cause of organ dysfunction and adverse postoperative outcomes after liver surgery.Pentoxifylline（PTX）and hypertonic saline solution（HTS）have been identified to have beneficial effects against IR injury.This study aimed to investigate if the addition of PTX to HTS is superior to HTS alone for the prevention of liver IR injury.METHODS： Male Wistar rats were allocated into three groups. Control rats underwent 60 minutes of partial liver ischemia, HTS rats were treated with 0.4 mL/kg of intravenous 7.5% NaCl 15 minutes before reperfusion, and HPTX group were treated with 7.5% NaC1 plus 25 mg/kg of PTX 15 minutes before reperfusion. Samples were collected after reperfusion for determination of ALT, AST, TNF-α, IL-6, IL-10, mitochondrial respiration, lipid peroxidation, pulmonary permeability and myeloperoxidase. RESULTS： HPTX significantly decreased TNF-α 30 minutes after reperfusion. HPTX and HTS significantly decreased ALT,AST, IL-6, mitochondrial dysfunction and pulmonary myelo- peroxidase 4 hours after reperfusion. Compared with HTS only, HPTX significantly decreased hepatic oxidative stress 4 hours after reperfusion and pulmonary permeability 4 and 12 hours after reperfusion. CONCLUSION： This study showed that PTX added the beneficial effects of HTS on liver IR injury through decreases of hepatic oxidative stress and pulmonary permeability.

RADIOFREQUENCY ABLATION USING HYPERTONIC SALINE SOLUTION INSTILLATION: AN EX VIVO STUDY

Objective To determine whether radiofrequency ablation using hypertonic saline solution instillation can increase the extent of thermally mediated coagulation in ex vivo pig liver tissue. Methods Fifty-six radiofrequency ablation lesions were produced in fresh ex vivo pig’s liver. According to different saline solutions, the lesions were divided into six groups: 25% acetic acid, 18% NaCl, 10% NaCl, 5% NaCl, 0.9% NaCl, and distilled water. After radiofrequency ablation, the lesions size and morphology were measured and compared.Results Using different instillation, the volume of coagulation necrosis was different: 25% acetic acid >18% NaCl >10% NaCl >5% NaCl >0.9% NaCl>distilled water. Conclusion Radiofrequency ablation using hypertonic saline solution instillation can increase the volume of radiofrequency ablation induced necrosis.

Effect of Osmotic Pressure on Mycoplasma hyopneumoniae Strain 168

[Objective] This study aimed to investigate the effect of hypertonic solution on the morphology,size,incubation time and cell viability of Mycoplasma hyopneumoniae（Mhp）.[Method] Mhp was stimulated by using NaCl hypertonic solution with a final concentration of 1.8% for 1,2,4,6 and 8 min,respectively.After staining and microscopic examination,the particle size and CCU（color change unit） were determined,and PCR identification was conducted.[Result] After treated with hypertonic solution for different time,Mhp was shrunken and turned round,the particle size was reduced,and the cell number rapidly decreased or even disappeared after 2 min.CCU was reduced by a gradient,and the observation time was extended for 1 d.The target band of Mhp could be amplified from samples after treated with hypertonic solution for different time.After treated with 1.8% NaCl solution for 1-6 min,Mhp changed in the morphology and size but still had viability.[Conclusion] This study provided reference data for exploring the entrance of Mhp into blood circulation after intramuscular injection and a new immunization route of swine Mycoplasma pneumonia vaccine.

Hypertonic saline solution resuscitation in hemorrhagic shock dogs

Objective: To find out the optimal concentration, infusion rate and dosage of saline for resuscitation.Methods: Forty five dogs were used to establish hypovolemic shock models. The dogs were resuscitated with saline of different concentrations and different dosages under different infusion rates, and the resuscitation results were compared. Results: The best concentration was 7.5 %, the best rate of infusion 20 ml/min (a volume equivalent to 15% of the shed blood) and the best dosage 5.71 ml/kg. The method was effective for resuscitation, the mean arterial pressure (MAP) could be elevated to 89% of the baseline, and this MAP could be kept for more than one hour. Conclusions: Using 7.5 % sodium chloride solution equivalent to 15% of the shed blood at an infusion rate of 20 ml/min can achieve a best resuscitation result.

Immunoprotective effects of HSH resuscitation on rats with hemorrhagic shock

Objective: To evaluate the immunoprotective effects of HSH resuscitation on rats with hemorrhagic shock. Methods: Controlled hemorrhagic shock was induced by blood withdrawal. The animals were divided into 3 groups: sham group (n=16), RL group (n=18), and HSH group (n=18). Then RL group and HSH group were divided into 3 subgroups. The rats were killed 1, 2 and 4 h after resuscitation. HE staining, transmission electron microscope and TUNEL staining were used to detect apoptosis of lymphocytes in the spleen. Apoptosis index (AI) of each group was calculated. Realtime quantitative RT-PCR was performed to record TNF-α mRNA copies in the spleen. Results: There was no difference in AI among HSH group and 1 h subgroup of RL and sham group(P>0.05); AI of the 2 and 4 h subgroups of RL group was higher than that of other groups (P<0.05); there was no significant difference in AI among HSH subgroups(P>0.05). The expression of TNF-α mRNA was higher in RL 1 h subgroup than the others (P<0.05),and there was no difference between HSH 4 h subgroup and sham group. Conclusion: HSH can down-regulate the expression of TNF-α mRNA in the spleen, reduce excessive apoptosis of lymphocytes in the spleen, and protect the immunofunction following hemorrhagic shock.

Effect of hypertonic saline solution on the left ventricular functions of isolated hearts from burned rats

Objective: To study the effect of hypertonic saline solution on the left ventricular functions of isolated hearts from burned rats. Methods: Thirty six Wistar rats were used and divided into 4 groups: (1) normal hearts perfused with isotonic Krebs Henseleit solution; (2) normal hearts perfused with Krebs Henseleit solution which contained 215 mmol/L Na +; (3) hearts of rats suffered from 25% TBSA third degree burn and perfused with isotonic Krebs Henseleit solution; (4) hearts of the burned rats perfused with Krebs Henseleit solution which contained 215 mmol/L Na +. The systolic and diastolic functions of the left ventricle were observed.Results: During perfusion, there were very short periods of decrease in heart systolic and diastolic functions at first, but they recovered very soon and even became stronger than normal both in the normal and burned rats. The systolic and diastolic functions of the hearts increased very significantly when the perfusion solution was changed to isotonic solution from the hypertonic solutions. The effect of the hypertonic saline solution on the ventricular systolic and diastolic improvements was stronger in the hearts of the burned rats than that in the normal hearts. Conclusions: Hypertonic saline solution can directly affect myocardium and significantly improve the ventricular systolic and diastolic functions, especially in the hearts of the burned rats.

Hypertonic saline resuscitation contributes to early accumulation of circulating myeloid-derived suppressor cells in a rat model of hemorrhagic shock

Background Hemorrhagic shock is usually associated with complicated immune and inflammatory responses, which are sometimes crucial for the prognosis. As regulators of the immune and inflammatory system; proliferation, migration, distribution and activation of myeloid-derived suppressor cells （MDSCs） are intimately linked to the inflammation cascade. Methods In a model of severe hemorrhagic shock, thirty-five rats were randomly divided into control, sham, normal saline resuscitation （NS）, hypertonic saline resuscitation （HTS）, and hydroxyethyl starch resuscitation （HES）, with seven in each group. MDSCs were analyzed by flow cytometric staining of CD11b/c＊Gra~ in peripheral blood mononuclear cells （PBMC）, spleen cell suspensions, and bone marrow nucleated cells （BMNC）. Simultaneously, the expressions of arginase-1 （ARG-1） and inducible nitric oxide synthase （iNOS） mRNA in MDSCs were evaluated by quantitative reverse transcription-polymerase chain reaction （qRT-PCR）. Results In the early stage after hemorrhagic shock, fluid resuscitation and emergency treatment, the MDSCs in the PBMC of NS, HTS and HES groups markedly increased, and MDSCs in BMNC of these groups decreased accordingly, significantly different to the control group. In hemorrhagic shock rats infused with HTS at the early resuscitation stage, MDSCs in PBMC increased about 2 and 4 folds, and MDSCs in BMNC decreased about 1.3 and 1.6 folds, as compared to the sham group respectively, with statistically significant difference. Furthermore, compared to the NS and HES groups, the MDSCs in PBMC of HTS group increased 1.6 and 1.8 folds with statistically significant differences; the MDSCs decrease in BMNC was not significant. However, there was no statistically significant difference in MDSCs of spleen among the five groups. In addition, compared to the control, sham, NS and HES groups, the ARG-1 and iNOS mRNA of MDSCs in PBMC, spleen and BMNC in the HTS group had the highest level of expression, but no statistically significant differences were noted. Conclusions In this model of rat with severe and controlled hemorrhagic shock, small volume resuscitation with HTS contributes to dramatically early migration and redistribution of MDSCs from bone marrow to peripheral circulation, compared to resuscitation with NS or HES.

Effect of hypertonic saline resuscitation on heme oxygenase-1 mRNA expression and apoptosis of the intestinal mucosa in a rat model of hemorrhagic shock

Background Massive blood loss due to trauma is the leading cause of death in trauma patients and military combatants. The fluid category of resuscitation for hypotensive trauma patients is open to debate. This study was conducted to investigate the early effects of hypertonic and isotonic saline solutions on heme oxygenase-1 （HO-1） mRNA expression and apoptosis in the intestinal mucosa of rats with hemorrhagic shock. Methods A model of severe hemorrhagic shock was established in 21 Sprague-Dawley rats. The rats were randomly divided into sham, normal saline resuscitation （NS）, and hypertonic saline resuscitation （HTS） groups, with 7 in each group. We assessed and compared the HO-1 mRNA expression and apoptosis in the small intestinal mucosa of rats after hemorrhagic shock and resuscitation using the SYBR Green I fluorescence quantitative reverse transcriptase polymerase chain reaction, fluorescein-iso-thiocyanate-annexin V/propidium iodide double staining, and flow cytometry. Results In the early stage of hemorrhagic shock and resuscitation, marked apoptosis occurred in the small intestinal mucosa from both the NS and HTS groups. The apoptotic rate in the NS group was higher than that in the HTS group （P 〈0.01）. Among the three groups, HO-1 mRNA mucosa from the HTS group had the highest level of expression; however, the differences were not significant. There was a significant negative correlation between HO-1 mRNA expression and apoptosis in the small intestinal mucosa from the NS and HTS groups after hemorrhagic shock and resuscitation. Conclusions In this rat model of severe hemorrhagic shock, HTS resuscitation with a small volume is more effective than NS resuscitation in reducing apoptosis of the intestinal mucosa. Further, HO-1 mRNA over-expression in the intestinal mucosa may be one of the molecular mechanisms of HTS in the resuscitation of hemorrhagic shock.

Variations of p38 MAPK and slCAM-1 with therapeutic effect of different resuscitation fluids on severe traumatic patients

Objective: To investigate the dynamic variation and action mechanism of sICAM-1 and p38 mitogen-activated protein kinases (MAPK) signal transduction in human severe trauma and resuscitation, as well as the effect of lactated Ringer's solution ( LR), 7.5% sodium chloride solution ( HS ) and 20% albumin injection ( ALB ) on the incidence and mortality of multiple organ dysfunction syndrome (MODS).Methods: Seventy-two severe trauma patients (ISS score 16-43) were divided into ISS≤25 and ISS＞25 groups (each group was subdivided into LR, HS and ALB groups). ELISA was used to measure the concentration of sICAM-1. Western blot was used to measure the expression of p38 MAPK.Results: Compared with LR group, the transfusion volume needed for maintaining systolic blood pressure ≥ 90 mm Hg was significantly decreased in HS and ALB groups (P＜0.05). Compared with the control group, the concentration of blood sICAM-1 and the expression of p38 MAPK was elevated from 4 to 48 hours after trauma in all experimental groups (P ＜ 0.05 -0.01 ). At 4, 12, and 24 hours, there was significant correlation between the expression of p38 MAPK and sICAM-1 (P ＜ 0. 01).Compared with LR group, sICAM-1 and p38 MAPK in HS and ALB groups were decreased (P ＜ 0.05). sICAM-1 and p38 MAPK were significantly higher in the group of ISS ＞25 than that of ISS ≤ 25 (P ＜ 0.05 ). MODS incidence and mortality were significantly higher in the group of ISS ＞ 25 than that of ISS ≤ 25 ( P ＜ 0.05 ). MODS incidence and mortality were lower in HS and ALB groups than LR group (P＜0.05).Conclusions: The up-regulation of polymorphonuclear neutrophil-endotheliocytes (PMN-EC) adhesion may be due to the increased sICAM-1 expression during severe trauma. The up-regulation of sICAM-1 expression is correlated with the activation of p38 MAPK. During severe trauma, the levels of sICAM-1 and p38 MAPK, as well as the incidence and mortality of MODS are lower when HS and ALB are used than single lactated LR solution is used.