Adenosine, lidocaine and Mg2+ (ALM) fluid therapy attenuates systemic inflammation, platelet dysfunction and coagulopathy after non-compressible truncal hemorrhage

Abstract

Background: Systemic inflammation and coagulopathy are major drivers of injury progression following hemorrhagic trauma. Our aim was to examine the effect of small-volume 3% NaCl adenosine, lidocaine and Mg2+ (ALM) bolus and 0.9% NaCl/ALM 'drip' on inflammation and coagulation in a rat model of hemorrhagic shock.

Methods: Sprague-Dawley rats (429±4 g) were randomly assigned to: 1) shams, 2) no-treatment, 3) saline-controls, 4) ALM-therapy, and 5) Hextend®. Hemorrhage was induced in anesthetized-ventilated animals by liver resection (60% left lateral lobe and 50% medial lobe). After 15 min, a bolus of 3% NaCl ± ALM (0.7 ml/kg) was administered intravenously (Phase 1) followed 60 min later by 4 hour infusion of 0.9% NaCl ± ALM (0.5 ml/kg/hour) with 1-hour monitoring (Phase 2). Plasma cytokines were measured on Magpix® and coagulation using Stago/Rotational Thromboelastometry.

Results: After Phase 1, saline-controls, no-treatment and Hextend® groups showed significant falls in white and red cells, hemoglobin and hematocrit (up to 30%), whereas ALM animals had similar values to shams (9-15% losses). After Phase 2, these deficits in non-ALM groups were accompanied by profound systemic inflammation. In contrast, after Phase 1 ALM-treated animals had undetectable plasma levels of IL-1α and IL-1β, and IL-2, IL-6 and TNF-α were below baseline, and after Phase 2 they were less or similar to shams. Non-ALM groups (except shams) also lost their ability to aggregate platelets, had lower plasma fibrinogen levels, and were hypocoagulable. ALM-treated animals had 50-fold higher ADP-induced platelet aggregation, and 9.3-times higher collagen-induced aggregation compared to saline-controls, and had little or no coagulopathy with significantly higher fibrinogen shifting towards baseline. Hextend® had poor outcomes.

Conclusions: Small-volume ALM bolus/drip mounted a frontline defense against non-compressible traumatic hemorrhage by defending immune cell numbers, suppressing systemic inflammation, improving platelet aggregation and correcting coagulopathy. Saline-controls were equivalent to no-treatment. Possible mechanisms of ALM's immune-bolstering effect are discussed.

Fig 1. Schematic of the in vivo rat model of truncal bleeding and shock treated with two-phase resuscitation therapy.

Animals were anesthetized and mechanically ventilated for the entire experimental and monitoring period (~7 hr 15 min). Body temperature was allowed to drift after anesthesia.

Fig 2. Prothrombin time (sec), activated partial thromboplastin time (sec), and fibrinogen concentration (g/dL) after Phase 1 (A) and Phase 2 (B) resuscitation at baseline, and in shams, no treatment, saline controls, ALM therapy, and Hextend^® groups.

Values represent mean ± SEM. Baseline values were obtained from 8 healthy anaesthetized, ventilated rats with one femoral cut-down to obtain an arterial blood sample (see Methods). n = 8 for all groups after Phase 1 resuscitation and Baseline, Sham and ALM groups at Phase 2 resuscitation. n = 2 for Hextend^® group, n = 3 for No treatment group, and n = 5 for Saline controls at Phase 2 resuscitation due to early mortality. *p<0.05 compared to Baseline and ALM groups; ^#p<0.05 compared to Baseline; ^†p<0.05 compared to ALM group; ^¶p<0.05 compared to Baseline, Sham, and ALM therapy; ^‡p<0.05 compared to all groups except No Treatment group.

Fig 3. Platelet count (10⁹/L) (A) at baseline, after 60 min Phase 1 resuscitation, and 240 min and 300 min Phase 2 resuscitation, and ADP- (B) and collagen-induced (C) maximum platelet aggregation after Phase 2 resuscitation in shams, no-treatment, saline, ALM, and Hextend^® groups.

Values represent mean ± SEM. n = 8 for all groups. *p<0.05 compared to Sham and ALM groups; ^#p<0.05 compared to ALM group.

Fig 4. Temperature (°C) at baseline (Pre-bleed), and during Phase 1 and Phase 2 resuscitation in shams, no-treatment animals, saline controls, ALM therapy group, and Hextend^®-treated animals.

Values represent mean ± SEM. All values n = 8 except for groups with mortality. For No-Treatment group n≤7 from 30 min Phase 1; n = 0 from 150 min Phase 2. For Saline controls n≤6 from 45 min Phase 1. For Hextend^® group n≤7 from 15 min Phase 1 until 60 min Phase 2; n = 0 from 90 min Phase 2. ^†p<0.05 compared to all other groups.