Pathophysiology of Hemorrhage as It Relates to the Warfighter

Abstract

Saving lives of wounded military warfighters often depends on the ability to resolve or mitigate the pathophysiology of hemorrhage, specifically diminished oxygen delivery to vital organs that leads to multiorgan failure and death. However, caring for hemorrhaging patients on the battlefield presents unique challenges that extend beyond applying a tourniquet and giving a blood transfusion, especially when battlefield care must be provided for a prolonged period. This review describes these challenges and potential strategies for treating hemorrhage on the battlefield in a prolonged casualty care situation.

Introduction

Traumatic injury occurs in both combat and civilian populations; it is the leading cause of death in the United States for people aged 1–44 (1). Many traumatic injuries result in hemorrhage. In the recent wars in Iraq and Afghanistan, the inability to control hemorrhage was the leading cause of potentially survivable death (2, 3), as it has been throughout history. Since 1990, United States forces have been engaged in counterinsurgency conflicts that produce relatively few casualties, who were easily evacuated to a surgical capability for definitive surgical care within a matter of minutes to hours. Indeed, Defense Secretary Robert Gates promulgated a 2009 policy that required prehospital evacuation of critically injured casualties in ≤60 min (“the Golden Hour”), producing a median transport time of 43 min and a significant decrease in mortality (4, 5). Additionally, these wars saw the use of advanced life-saving interventions, such as more liberal use of tourniquets to arrest severe extremity bleeding, blood products for resuscitation, hemostatic dressings, and improved training in combat casualty care (6). Together, these policies, tools, and education produced the lowest case fatality rates in history, despite increasing injury severity scores throughout the conflicts (7). Lessons learned have been adopted into civilian trauma practice as well.

Casualty care during the recent conflicts also benefited from the ability to expend tremendous resources to save severely injured patients. For example, it was not uncommon to massively transfuse (>10 units) blood into hemorrhaging patients before and during surgical procedures (6). But what if war reverts to those of the past, with advanced weaponry, mass casualties, limited resources, and the inability to freely move them from the battlefield? This is the issue posed as the United States and its allies contemplate the possibility of conflicts against near-peer adversaries that are expected to produce greater numbers of casualties (100s to 1,000s per day) but without air superiority facilitating evacuation (FIGURE 1). In current planning, the expectation is that future conflicts will be fought simultaneously in cyberspace, from the air and sea, and with ground large-scale combat operations (8). Scenarios in which a combat medic must provide prolonged casualty care with only the resources that s/he can carry have occurred (e.g., Black Hawk Down scenario); these are expected to increase in future conflicts but with many more casualties. In essence, the prolonged care scenarios envisioned are a return to those seen in the World Wars.

FIGURE 1.

A comparison of casualty care management challenges during recent conflicts and future near-peer conflicts The table at bottom represents effects of these situational challenges on the development of pathophysiology.

Although we were successful in treating hemorrhagic shock in the conflicts of the 1990s to 2021 by relying on rapid evacuation and ample resources, treatment must evolve further to maintain recent case fatality rates under future conditions. However, the physiology underlying the development of hemorrhagic shock is unchanged; studying its pathophysiology will yield insights into future treatments that could be used to maintain patients even under conditions of delayed surgical repair. This review focuses on the pathophysiology of hemorrhage in a prolonged care scenario, with the assumption that initial hemorrhage control has been achieved but the medic lacks resources to provide full resuscitation. The intent here is to provide an overview of the multifactorial nature of the pathophysiology following hemorrhage; each subsection is worthy of its own detailed review, and references are provided to such reviews.

Hemorrhage Progression

There are four commonly accepted stages of hemorrhage, from minor blood loss to the development of shock, a condition in which tissue perfusion is too low to provide adequate tissue oxygenation and maintain normal metabolic functions (TABLE 1; Ref. 9). Class 1 and class 2 stages represent conditions of compensated blood loss because autonomic and neurohumoral compensatory responses maintain hemodynamic stability. During this period of mild to moderate hemorrhage, vital organ perfusion and oxygenation are preserved. Class 3 is considered decompensated shock because compensatory mechanisms are no longer effective to maintain vital organ oxygenation. Class 4 is the most severe level of hemorrhagic shock and is associated with impending cardiovascular collapse, vital organ injury, and death.

Table 1.

Stages of hemorrhage

Class Designation	Blood Volume Lost, mL	% of Total Blood Volume Lost	MAP/SP	Pulse Pressure	HR	RR	Urinary Output	Perfusion in Vital Organs
1	<750	<15	―	―	―	―	―	Compensated
2	750–1,500	15–30	―↓	↓	↑	↑	―↓	Compensated
3	1,500–2,000	30–40	↓	↓	↑	↑	↓	Starting decompensation
4	>2,000	>40	↓↓	↓↓	↑↓	↑↓	↓↓	Decompensated

HR, heart rate; MAP, mean arterial pressure; RR, respiratory rate; SP, systolic pressure. Table adapted from Ref. 9 with permission.

The onset of hemorrhage produces an immediate decrease in blood pressure that stimulates an important compensatory mechanism, the arterial baroreflex. Baroreflex-mediated autonomic mechanisms inhibit the cardioinhibitory center, resulting in sympathetic nervous system activation and parasympathetic inhibition; this produces an increase in peripheral vascular resistance and an increase in heart rate (FIGURE 2; Refs. 10, 11). Sympathetic activation also produces constriction of major capacitance vessels, resulting in increased venous return (12). In addition, baroreflex activation of the sympathetic nervous system stimulates the adrenal gland to secrete epinephrine and norepinephrine to augment vasoconstriction and increases in heart rate. Baroreflex activation also stimulates the release of various neuroendocrine hormones such as renin from the kidney to activate the renin-angiotensin system and the secretion of vasopressin from the hypothalamus to mediate vasoconstriction and increase intravascular volume (11, 13–16). Other neuroendocrine hormones released during hemorrhage include aldosterone (17, 18) and neuropeptide Y (15), which also contribute to the maintenance of vascular volume and organ perfusion. These compensatory baroreflex responses and regional autoregulation act to maintain blood pressure and blood flow to vital organs (heart and brain) despite blood volume loss. Although respiratory compensatory responses to hemorrhage and the mechanisms that regulate these responses are not well understood, measurements in conscious rabbits revealed that during the early stage of hemorrhage arterial partial pressure of carbon dioxide ($\mathrm{P}{\mathrm{a}}_{\mathrm{C}{\mathrm{O}}_2}$) decreases but respiration rate does not change (19).

FIGURE 2.

Flow diagram of early compensation of vital organ perfusion after hemorrhage Activation is represented by solid black lines, and inhibition is represented by dashed red lines. Therapeutic interventions and their targets are shown in green. CO, cardiac output; MAP, mean arterial pressure; SNS, sympathetic nerve system; TPR, total peripheral resistance.

Capillary fluid shift is the major mechanism for volume expansion in early hemorrhage before resuscitation or reestablishment of water balance via the renin-angiotensin system. The first step of capillary fluid shift is rapid movement of protein-free fluid from the interstitium into capillaries. The second step is a slow shift of proteins from tissue into the circulation to support plasma oncotic pressure (20). However, the driving force for movement of protein depends on movement of fluid from cells to the interstitium, which may be a “bottleneck” in severe or prolonged hemorrhage because of ATP depletion and cellular edema. Establishment of an improved osmotic gradient across pericapillary interstitial spaces that facilitates this fluid shift may therefore be a therapeutic target.

The stress response and rapid release of catecholamines alter systemic and local metabolism, with the end result of providing more fuel (glucose) to tissues. Stress hyperglycemia can be detected within minutes after injury because of inhibited insulin secretion and elevated glycogenolysis and glucagon release or cortisol-mediated gluconeogenesis (21–23). Elevated glucose can have both deleterious and beneficial effects during the early stages of hemorrhage; therefore, treatments to prevent early stress hyperglycemia must be considered with caution. For example, stress hyperglycemia can increase endothelial damage, endoplasmic reticulum stress, acidosis, inflammatory responses, and reactive oxygen species (ROS) (23–25). Meanwhile, stress hyperglycemia may facilitate catabolism shift from peripheral tissue to the vital organs, such as the brain and the heart, the two organs most vulnerable to ischemia. Attempts to interfere with this exceedingly complex multisystem adaptive response may be harmful (26).

With continued or more severe hemorrhage, stroke volume and cardiac output continue to decrease, physiological mechanisms that attempt to maintain homeostasis eventually fail, resulting in a dramatic and sudden decrease in total peripheral resistance, mean arterial pressure, and heart rate (10, 11, 27). If severe and prolonged, hemorrhage results in a mismatch between tissue oxygen delivery ($\dot{\text{D}}{\mathrm{O}}_2$) and oxygen consumption ($\dot{\text{V}}{\mathrm{O}}_2$). When the body does not have an adequate oxygen supply, cells rely on anaerobic glycolysis, which results in increased levels of blood lactate and decreases in blood pH resulting in metabolic acidosis. The increases in blood hydrogen ion and carbon dioxide levels stimulate the chemoreceptor reflex, resulting in an activation of central respiratory centers to compensate for the metabolic acidosis (28). Ultimately, the presence of continued tissue hypoxia and metabolic acidosis produces progressive cellular deterioration and cell death and multiorgan failure (29). Eventually, with prolongation of this uncompensated phase of blood loss, irreversible hemorrhagic shock occurs, which is fatal (30). For a more in-depth review of compensatory responses to hemorrhage, see Ref. 31.

Mitochondrial and Cellular Function

Decreased $\dot{\text{D}}{\mathrm{O}}_2$ stalls forward electron flux through the mitochondrial electron transport chain, leading to the loss of proton pumping and the loss of inner membrane potential, which halts ATP synthesis (FIGURE 3). If the inner membrane potential rises high enough during hypoxia, ATP synthase reverses, hydrolyzing ATP generated via anaerobic mechanisms to regenerate membrane potential; estimates suggest that during hypoxia at least 35% and as much as 90% of all anaerobically generated ATP is hydrolyzed by the reversed ATP synthase (32–37).

FIGURE 3.

Forward and reversed electron transport in shock The forward flow of electrons through the electron transport chain (orange arrows) is decreased by shock, which causes a loss of O₂ delivery and a loss of ATP synthesis, leading to reverse electron transport (red arrows) at the flavin mononucleotide (FMN) and Qi site of complex I and the Qo site of complex III. These reversed electrons generate reactive oxygen species (ROS), which can be both destructive (complex I) and adaptive (complex III). Potential therapeutic interventions and their targets are suppressors of site 1Q electron leak (S1QEL) and Szeto-Schiller peptides. HIF-1α, hypoxia-inducible factor 1 alpha. Figure modified from Sabiston Textbook of Surgery (21st ed.) (162) with permission.

As forward electron flux through the electron transport chain slows, demand for high-energy electrons from NADH falls, leading to NADH accumulation (37, 38), while succinate accumulates from forward tricarboxylic acid (TCA) cycle activity (39–41). Because the NADH-to-NAD+ ratio controls the redox state of complex I’s flavin mononucleotide (FMN) site, the hypoxic accumulation of NADH favors the generation of ROS at complex I via slip of electrons from the FMN site onto oxygen (42). Similarly, reoxygenation causes the rapid oxidation of accumulated succinate, which drives ROS generation through complex I via reversed electron transport (43).

Cellular ATP depletion is therefore the trigger of ischemic injury (FIGURE 4). ATPase inactivation reduces sodium and calcium efflux and limits the reuptake of calcium by the endoplasmic reticulum, thereby producing increases in both intracellular calcium and sodium. In addition, accumulation of hydrogen ions secondary to anaerobic metabolism further exacerbates these processes by decreasing sodium-potassium-ATPase, enhancing sodium/hydrogen exchanges, and thus reversing transport of sodium/calcium. Calcium-mediated opening of mitochondrial permeability transition pores (mPTPs) and initiation of apoptosis cascades further dampen ATP production and lead to cell death (44–46).

FIGURE 4.

Flow diagram of ischemia-induced cell death Therapeutic interventions and their targets are shown in green. Ca, calcium; ER, endoplasmic reticulum; H, hydrogen; IL-1, interleukin-1; K, potassium; mPTP, mitochondrial permeability transition pore; Na, sodium; NCX, sodium/calcium exchanger; NOX; nicotinamide adenine dinucleotide phosphate oxidases; ROS, reactive oxygen species; SNA, sympathetic nerve activation; TNF-α, tumor necrosis factor-alpha; VDAC, voltage-dependent anion channel.

During ischemia, mitochondria are an early source of ROS due to generation of superoxide mainly from complex I, translocation of P66shc (a protein regulator of cellular redox state and apoptosis) to the outer membrane, and deactivation of antioxidative systems. Knockout mice missing P66shc exhibit enhanced resistance to oxidative stress and ischemic injury, suggesting P66shc as a potential therapeutic target (47). With progressive hypoxia and ATP depletion, generation of superoxide from the hypoxanthine-xanthine-uric acid pathway becomes dominant (44, 48, 49). In addition, stress hyperglycemia and immune cells via nicotinamide adenine dinucleotide phosphate oxidase/myeloperoxidase pathways have been shown to be important sources of free radicals.

Regulated Cell Death

Into this century, cell death was generally grouped into apoptotic (programmed) and necrotic (uncontrolled) (50), but we have come to recognize numerous types of nonapoptotic cell death that operate via regulated pathways. There are several excellent reviews on known forms of regulated cell death (RCD) (51–53). Certain pathways have greater battlefield relevance by relation to ischemia-reperfusion injury with attendant low-energy states and oxidative stress. Briefly, these are pyroptosis (caspase-1-mediated oncotic/inflammatory cell death) (54); necroptosis [damage-associated molecular patterns (DAMPs)-triggered RIPK complex leads to MLKL necrosome] (55, 56); parthanatos (hyperactivation of PARP1, increased mitochondrial apoptosis-inducible factor) (57); and mPTP-driven regulated necrosis (caspase-independent, p53-triggered mPTP opening results in oncotic rupture of mitochondria) (58, 59). Intervening against only one type of RCD is insufficient, as cells under stress will shunt to another form of cell death if able, and the inhibitor of one type can trigger another (52, 60, 61). Given the speed with which RCD pathways can be initiated (within minutes), timely intervention may extend tissue viability and prevent unnecessary cell death with resultant distant organ and immune dysfunction via DAMPs. Knowledge regarding the granular, tissue-specific proteomic and transcriptomic changes occurring in the setting of traumatic hemorrhagic shock is therefore needed to inform early treatment, reduce RCD, and prevent detrimental sequelae.

The Challenge of Traumatic Hemorrhage and Organ Failure

Ultimately, cellular dysfunction and death eventuate in damage to critical organs. In addition to organ dysfunction produced by ischemia, there is often the concomitant presence of tissue damage such as penetrating or blunt soft tissue injury, bone fracture, or traumatic brain injury. There is evidence that trauma affects cardiovascular and metabolic responses to hemorrhage and thus further exacerbates ischemic injury (62–64). First, traumatic injury and related nociceptive signals act to shunt blood away from metabolically active organs to relatively inactive tissues such as skeletal muscle after hemorrhage (62, 65). In fact, traumatic injury has been associated with decreased tolerance to hemorrhage, with nociception signaling as a possible mechanism (62, 66). Second, metabolic derangements, such as pyruvate and lactate accumulation and TCA cycle dysfunction, have been found in rats sustaining both trauma and hemorrhage (67). Pathophysiological changes induced by trauma not only further decrease tissue oxygen and energy supply but may also simultaneously increase the need for $\dot{\text{D}}{\mathrm{O}}_2$ and increase systemic metabolic rate, thus synergistically exacerbating ischemia in metabolically active organs. Inflammatory responses following trauma can also increase $\dot{\text{V}}{\mathrm{O}}_2$ and ATP depletion (68, 69). Furthermore, direct toxic effects of myoglobin, potassium, and other toxins released from injured muscle have been identified (70–72). Although these effects of traumatic injury are known, many animal models of hemorrhagic shock still do not combine hemorrhage with tissue damage. It should also be noted that, in addition to hemorrhage and traumatic injury effects, treatments such as prolonged tourniquet use on an extremity can also promote the release of such toxins and produce detrimental systemic effects upon tourniquet release.

Preserving organ function during prolonged care scenarios in warfighters with trauma and hemorrhage is therefore a critical goal for military research. Neurohumoral compensatory responses preferentially direct cardiac output to the brain and heart, which are critical organs requiring oxygen delivery, while “sacrificing” other organs. For example, although more resilient to ischemia than the brain and heart, the kidney is vulnerable to prolonged traumatic shock, and acute kidney injury (AKI) has long been a life-threatening complication to battlefield injuries. During World War II, mortality associated with AKI approached 100%; the development and application of dialysis techniques in the Korean War reduced mortality in patients with AKI to 60% (73, 74). Although there is a trend toward a reduction in overall rates of AKI due to less severe injury because of body armor wear and improved tactical field care in the current theaters, the overall mortality caused by AKI is still high, with any decrement in renal function significantly increasing morbidity and mortality (75, 76). Indeed, AKI occurred in current theaters despite the majority of evacuations occurring in <60 min (5); the frequency of AKI and dysfunction of other organs is expected to increase in delayed evacuation scenarios. Acute decreases in renal perfusion pressure due to hypotensive hemorrhage may overwhelm renal autoregulation and suppress glomerular filtration rate (i.e., prerenal injury), but this is reversible should renal perfusion be restored. However, prolonged ischemia may eventuate in acute tubular necrosis (intrarenal injury), in which tubules are damaged and autoregulation is lost (77). While AKI is one consequence of traumatic hemorrhage, dysfunction of other organs and the system as a whole (e.g., multiorgan failure) may also be a consequence of severe traumatic injury. Unfortunately, there is currently no preventive strategy for AKI and other sequelae of traumatic shock during delayed evacuation to a hospital setting. Current research efforts focus on preventing or delaying the cellular consequences of hemorrhage in an effort to protect vital organs for a prolonged period.

Another example of organ failure during hemorrhage is “blood failure.” Blood cells, circulating plasma components, and the vascular endothelium are in constant contact with one another, each contributing to the maintenance of homeostasis for the others. Accordingly, the blood and endothelium can be seen as a single organ system, as prone to failure during hemorrhage as any other (78). Taking this view, blood failure can be defined as “low tissue oxygen delivery, endotheliopathy, platelet dysfunction, and coagulopathy” (79). Hypoxia acts to disrupt metabolic function in endothelial cells just as in other cells, with the result that the endothelial barrier is compromised and becomes “leaky.” Importantly, normal endothelial regulation of blood flow at the microcirculatory level by various anticoagulant mechanisms is disrupted, the endothelial glycocalyx is degraded, and inflammatory cells are activated (79, 80).

In the blood failure concept, traumatic coagulopathy (i.e., the reduced ability to form clots) is a consequence of widespread cellular failure due to hypoxia, especially of platelets and the vascular endothelium (78). This failure of hemostasis is thought to be associated with both injury severity as well as poor outcomes after trauma (81); it is therefore of interest both for diagnostic potential and as a therapeutic target. Trauma patients can present with hyperfibrinolysis (enhanced destruction of formed clots), depletions in coagulation factors, as well as the activation of protein C, all of which contribute to coagulopathy and poor outcomes (82–84). Platelets become dysfunctional, further decreasing the ability to form clots. Therefore, the constellation of patient presentations and potential etiologies makes the coagulopathy of trauma very challenging to study and even more difficult to treat (85). Hypothermia and metabolic disturbances further amplify clotting dysfunction by decreasing platelet function and coagulation enzyme activity, leading to the well-known “lethal triad” of trauma: acidosis, hypothermia, and coagulopathy (86). Antifibrinolytic therapy with intravenous tranexamic acid (TXA; FIGURE 2) improves survival after trauma if given acutely (87) but worsens mortality if given >3 h after injury (88); TXA is now given routinely if administration is possible within 3 h of injury. In addition to its antifibrinolytic activation, TXA has beneficial effects on endothelial barrier function and reduces tissue edema (79) as well as other beneficial effects to include reducing inflammatory processes and protecting gut barrier function during ischemia-reperfusion injury (89–92).

Potential Treatment Strategies

Inherent in the discussion above is that there are a variety of physiological deficits in the hemorrhaging patient. However, the two most critical deficits are 1) the loss of blood itself leading to hemodynamic instability and 2) a decrease in oxygen delivery to vital organs. So the most effective treatment is fluid resuscitation to increase intravascular volume, thereby restoring blood pressure, and the optimal resuscitation fluid would also have the ability to carry oxygen. As discussed below, whole blood is currently considered the optimal resuscitative fluid as it replaces both volume and oxygen-carrying capacity. However, the logistics of blood delivery far-forward (e.g., the need for refrigeration, weight) may be daunting in future battlefield scenarios, and thus there is renewed interest in development of low-volume pharmacological adjuncts (“antishock drugs”) with the aim of altering the host response to the hemorrhagic insult to delay and/or prevent the onset of ischemic injury. For prehospital care, putative therapies with lower weight and size, a long shelf-life, and stability in variable environmental temperatures are desired. To this end, new pharmacological adjuncts are being considered that will further improve oxygen delivery and utilization, stabilize mitochondrial function, and/or prevent RCD. TABLE 2 provides a sample list of therapeutics currently under investigation, most already FDA approved and available for “off-label” use.

Table 2.

Pharmacological agents that have demonstrated beneficial effects after hemorrhage

Drug	Mechanism of Action	Posthemorrhage Survival Time	Posthemorrhage Survival Proportion	Model Species	References
Tranexamic acid	Inhibits plasmin, suppresses DAMP release	+	+	Rat, human	(87, 139–142)
Polyethylene glycol-20k	Improves microcirculatory perfusion and $\dot{\text{D}}{\mathrm{O}}_2$	+	?	Rat, pig	(113–115)
Trans-sodium crocetinate	Enhances oxygen diffusion into tissues	?	+	Rat	(121)
Valproic acid	Histone deacetylase inhibitor; mitochondrial stabilization	+	+	Rat, pig	(143–149)
Cyclosporine A	Calcineurin inhibitor; mitochondrial stabilization	+	+	Rat	(150)
Centhaquine	α2β-Adrenergic receptors	+	+	Rat, pig, human	(107–110)
Naloxone, naltrexone	µ-, δ-Receptor antagonist	+	+	Dog, horse, primate, human	(151–156)
Melatonin	NF-κB inhibitor, antioxidant	+	+	Rat, pig	(135, 157, 158)
β-Hydroxy-butyrate	Ketone body to provide cellular fuel	+	+	Rat, pig	(135, 157, 159)
Minocycline/doxycycline	PARP/caspase/MMP inhibition	+	+	Mouse, rat	(160, 161)

DAMP, damage-associated molecular patterns; $\dot{\text{D}}{\mathrm{O}}_2$, oxygen delivery; MMP, matrix metalloproteinase; NF-κB, nuclear factor kappa B; PARP, poly(ADP-ribose) polymerase.

Resuscitation: Restoration of Fluid Volume and Blood Pressure

At the start of the recent conflicts, the standard for resuscitation of hemorrhaging trauma patients was prehospital infusion of crystalloid (e.g., lactated Ringer’s, saline) or colloid (e.g., Hextend) solutions, followed by administration of blood products upon hospital arrival. However, it became evident that prehospital infusion of these fluids produced detrimental effects, including hemodilution, endothelial dysfunction, and coagulopathy (93). By 2006, Army physicians recognized the superiority of fresh whole blood transfusion to conventional blood component therapy (94, 95), as whole blood both increases intravascular volume and provides hemoglobin for oxygen delivery (FIGURE 2). This was not new, as whole blood transfusions were used for treatment of military casualties in both World Wars, the Korean War, and the Vietnam War (93, 96). Transfusion of whole blood at or close to the point of injury, when available, has become the preferred prehospital resuscitation fluid for military forces, with Special Operations forces carrying both whole blood on combat missions and equipment to facilitate a “walking blood bank” (97). Because of the superiority of whole blood as a prehospital resuscitation fluid (98), the military experience has now been transferred to prehospital civilian care, with whole blood being carried by first responders in some United States cities (99).

In future conflicts, however, the need for blood during mass casualty scenarios will outstrip the supply of blood and the resources to collect blood from others, which is particularly problematic if donors also must continue the fight. One potential solution for trauma patients has been the development of artificial blood substitutes, including hemoglobin-based oxygen carriers; none are yet FDA approved. Another potential solution is to provide plasma, which contains clotting factors and is protective of endothelial function (FIGURE 2). Two recent civilian trials have been performed using thawed plasma in addition to standard of care during prehospital care, with somewhat conflicting results (100, 101); in one study, prehospital administration of plasma during helicopter transport resulted in decreased 30-day mortality (100), whereas in the other there was no survival benefit to the administration of plasma during ground ambulance transport (101). However, post hoc analysis of the combined data set demonstrated improved survival when transport times were >20 min (102). Although use of thawed plasma presents logistical challenges, freeze-dried (lyophilized) plasma has efficacy and physiological responses equivalent to thawed plasma (103). Again, this is not new, as freeze-dried plasma was used by the United States military during World War II but was halted after hepatitis outbreaks. Freeze-dried plasma products are currently in use by French, German, Norwegian, and Israeli forces; the United States military has had access to the French product since 2018 under the FDA’s expanded access program. Currently, the United States military and the Department of Health and Human Services are funding developmental efforts for freeze-dried plasma (104).

Another approach to maintain blood pressure is to pharmacologically produce vasoconstriction. Although general vasoconstriction with norepinephrine is counterproductive, recent studies suggest that selective venoconstriction may be helpful. Centhaquine, a promising resuscitative agent, acts by stimulating alpha_2β-adrenergic receptors in the veins to cause constriction, which results in an increase in venous return, cardiac output, and mean arterial pressure in animal models of hemorrhage (FIGURE 2; Refs. 105–107). It has also been found to prevent AKI and restore renal blood flow in a rat hemorrhage model (108). Centhaquine has completed phase III clinical trials in hypovolemic shock patients and has been demonstrated to increase pulse pressure, reduce blood lactate levels, and improve base deficit associated with a reduction in multiple organ dysfunction and 28-day all-cause mortality (109, 110).

Oxygen Delivery Enhancement

Enhancing oxygen-carrying capability has long been considered a therapeutic strategy. However, although tissue perfusion is markedly impaired after hemorrhage, the decrease in arterial Po₂ is usually minor (111). In fact, reduction in tissue microcirculatory perfusion is a major contributor to impaired tissue oxygen supply (111, 112). Therefore, compounds able to improve microcirculatory perfusion, such as polyethylene glycol-20k (FIGURE 2), may be more effective than oxygen carriers in maintaining organ oxygenation. Polyethylene glycol-20k molecules can enter the interstitial space and cause the movement of isotonic fluid from interstitial and intracellular spaces into the capillaries by osmotic forces (113–115), Polyethylene glycol-20k thus establishes effective gradients across cellular/interstitial/vascular spaces, prevents cell edema, and facilitates water movement from cell to capillary to produce volume expansion in the circulation and improve organ perfusion during hemorrhage (114, 115).

Facilitating oxygen diffusion into tissues is another potential strategy to maintain tissue oxygenation under ischemic conditions. The rate of oxygen diffusion from red blood cells into surrounding tissues is determined by the pressure gradient of oxygen and the resistance to oxygen transport, with plasma providing a great deal of this resistance (116, 117). Trans-sodium crocetinate is an oxygen diffusion-enhancing compound that increases hydrogen bonding among water molecules, thus enhancing the rate and distance of oxygen diffusion into tissues (FIGURE 2; Refs. 118, 119). Trans-sodium crocetinate has been used in clinical trials to enhance oxygen diffusion into hypoxic cancer cells (120). Previous studies have explored its therapeutic potential for treating hemorrhagic shock (121, 122).

Mitochondrial Stabilization

As discussed above, ROS generation is detrimental during ischemia, but ROS are also generated on reperfusion and reoxygenation of tissues. ROS both degrade mitochondrial function and serve as critical components of the cellular response to hypoxia; mitochondrial ROS may be required to stabilize the master-regulatory transcription factor hypoxia-inducible factor 1 alpha (HIF-1α) (123, 124). Interestingly, newly identified site-specific mitochondrial complex I (125) and III (126) inhibitors have shown that ROS generated at different sites in the electron transport chain have different impacts on cellular responses to hypoxia and reoxygenation (127). ROS from the matrix-facing quinone site on complex I appear to be destructive, whereas ROS from the intermembrane space-facing quinone site of complex III are critically required for stabilizing HIF-1α (127–129). The prevention of ROS generation at complex I without preventing ROS generation at complex III, such as suppressors of site 1Q electron leak (S1QEL; FIGURE 3), could therefore represent an appealing strategy for preserving mitochondrial function without preventing critical signaling mechanisms (127). Additionally, the preservation of cardiolipin in its native (i.e., multiply unsaturated) conformation with Szeto–Schiller peptides represents a promising approach to mitochondrial stabilization during stress (FIGURE 3; Ref. 130).

Prevention of RCD

Casualties suffering from traumatic shock have mere minutes before they begin to circulate DAMPs and exhibit early steps of RCD pathways (131); accordingly, favorable pharmacokinetics and route of administration become critical as the target tissues may have the most compromised circulatory supply. Ideally, such a therapeutic could be given at point of injury as an adjunct to resuscitation, delaying or preventing the onset of RCD, resultant DAMP release, and subsequent end organ injury. If targeting RCD, a combination of agents will likely be required, as preventing one RCD pathway typically leads to shunting toward another (51). Pharmacological agents proposed to ameliorate RCD include valproic acid (FIGURE 4), which helps to reduce apoptosis and is the subject of a current clinical trial to reduce AKI in liver transplant patients (https://clinicaltrials.gov/ct2/show/NCT04531592). Likewise, cyclosporine A (FIGURE 4) acts to stabilize mitochondria and prevent RCD after ischemia (132, 133). Although valproic acid and cyclosporine A seem to work for many types of apoptosis, it remains to be seen whether these potential treatments are a good option for reversing hemorrhagic shock, as these therapeutics rely on many time- and energy-consuming processes such as cellular entry, receptor binding, transcription, and protein translation and synthesis. Under the ischemic conditions following hemorrhage, there may not be enough cellular energy to allow these treatments to work.

The ideal of a rapidly induced hibernation state in humans has been explored, for obvious reasons; i.e., hibernating organisms survive with a greatly reduced metabolic rate, then rebound without loss of tissue or function (134–136). Unfortunately, the process of entering a proper hibernating/torpor state generally takes place over weeks, but perhaps could be compressed to days, or even hours; agents successful in rapidly inducing torpor states with small animals, however, have generally failed in larger-animal studies (137). Although the hemorrhaging casualty has only minutes before catastrophic changes begin to occur at the cellular level, possible clues may be taken from how hibernating organisms manage extreme metabolic deprivation. The Defense Advanced Research Projects Agency (DARPA) initiated the Biostasis program in an attempt to extend the Golden Hour by developing novel pharmacological approaches to slow metabolic processes, thereby resulting in a metabolic arrest that might protect functional capacity until definitive medical care is available (138).

Conclusions

Although the nature of war may change, the physiology underlying traumatic hemorrhage does not. However, the potential return to large-scale combat operations presents renewed challenges to a military medical system with historically low case fatality rates in recent wars; these may, in fact, represent an aberration in the history of warfare. Potential solutions to face these challenges must be based both on a greater understanding of the underlying physiology of hemorrhage and on the logistical realities of providing care in these scenarios. The relatively new understanding of the blood-endothelium as an organ that fails in hemorrhagic shock provides new opportunities for potential intervention. However, the rapid time course of RCD activation must also be taken into account when contemplating interventions. Undoubtedly, there is no one “silver bullet,” but ultimate resuscitation strategies will require combinations of oxygen-delivering solutions to increase blood volume and adjuncts to make the host tissue more tolerant to ischemia.