Background
Acute liver failure (ALF) and sepsis share some conformity, e.g. coagulopathy, shock and a high mortality. As a combination of both, the sepsis induced ALF is the lethal duo. Despite of intensive research and improvement of treatment, patients in septic shock as well as patients with ALF show a survival rate of less than 50% [
1,
2].
The liver and the intestine play a pivotal role in sepsis by modulating immune response due to release and filtering cytokines, bacterial fragments and vasomodulating substances. Hepatic hypoperfusion and microcirculation disturbances lead to contracted sinusoids, swollen sinusoidal endothelial cells and activated Kupffer cells. The hepatic response to this stimulus is a release of toxic, pro-inflammatory and vasoactive substances, which trigger the vicious circle of multi organ failure (MOF) and death [
3,
4]. The hepatic histopathological changes prominent after septic shock may be due to impaired microcirculation and therefore cellular hypoxia [
5]. Hepatic perfusion is clinically difficult to determine and can’t be predicted by systemic hemodynamic parameters. However cardiac output (CO) showed a direct correlation to total hepatic flow [
6] but hepatic microcirculation remained heterogeneous with regional flow deficits.
During septic shock in pigs superior mesenteric artery and hepatic microcirculation flow decreased about 50% and hydroxyethylstarch administration improved splanchnic blood flow but not microvascular liver perfusion [
4]. In addition repeated administration of hydroxyethylstarch can aggravate portal hypertension, liver failure and sepsis in patients with chronic liver disease [
7]. On the other side, Catre and colleagues have demonstrated protective effects of HES 130/0.4 on hepatic reperfusion damage [
8]. However volume resuscitation is a cornerstone in the treatment of sepsis and shock. The volume replacement strategies are different between countries and are even different between ICUs in the same hospital [
9]. Crystalloids (CL) and natural or artificial colloids (COL) are commonly used to restore fluid balance.
In the last decades controversy results were published and now the German sepsis guidelines 2010 recommended the use of CL in sepsis. Despite of numerous clinical studies there is no evidence for the safety use of third generation HES 130/0.4 in critical ill patients [
10]. In contrast there are increasing results of severe side effects of synthetic colloids to kidney function and mortality [
11‐
14]. However little is known about the impact of different CL and COL solutions on the hepatic and intestinal microcirculation in sepsis. Therefore we designed this study to investigate the direct influence of different solutions on liver and intestine microcirculation and function in a rodent sepsis model.
Discussion
Volume resuscitation is a pivotal therapy in sepsis and septic shock, but the level of evidence-based recommendation of the type of intravenous fluid is still low. Furthermore the required quantity of volume to stabilize haemodynamics remains a challenge by the complex interaction of endothelial-, epithelial barrier disruption, cytokine storm, septic cardiomyopathy, hypo- or hyperthermia, cavity edema, coagulation disorders with consecutive bleeding and co-morbidity [
23].
In patients suffering from endothelial barrier disruption like in sepsis, the estimated volume effect of colloids is less than expected. For example, in the Scandinavian Starch for Severe Sepsis/Septic Shock (6S) trial, patients in the HES group received almost the same amount of volume when compared with patients in the RA group to achieve hemodynamic stabilisation [
13]. Silva et al. have shown recently in rats with CLP-induced sepsis that an infusion of 6 ml/100gBW/h with gelatine resulted in hypervolemia [
21]. The right colloid to crystalloid ratio is still an open question, and we therefore decided to use a 1:1 ratio to investigate the same amount of different infusion solutions. Furthermore our fluid regime comprises two sections: 1) 0.5 ml/100gBW/h NaCl: to cover the normal fluid intake of rats and 2) additional we administered 1.0 ml/100gBW/h of the different solutions to compensate fluid loss of surgical procedures and sepsis (Figure
2). Singelton et al. showed a 40% mortality rate in the first 24 h and a significant elevated IL-6 level, when 25% of the coecum was ligated and punctured twice [
15]. We established this model in our lab to achieve severe sepsis and a reduction of lethality by sufficient analgesia and volume resuscitation. The aim of the protocol was to get a fast (within 24 h) onset of beneficial and/or adverse side effects of fluid solutions in septic rats, and not a “patient” adapted goal directed fluid resuscitation trial. The degradation of starch molecules or the metabolic state of rats are two of numerous differences between rodents and humans and conclusions from animal models to clinical practice should be done with caution. Most notably, our rodent model, investigating 24 hours after the CLP procedure, reflects the early stage of severe sepsis. Patients in the clinical setting are often treated in the later phases of severe sepsis.
However, the pathomechanism of septic liver failure is poorly understood and it is meanwhile known, that macrohemodynamic parameters do not reflect microcirculation and microcirculation by itself differs extensively between different organs, even when systemic macrohemodynamic steady state values were achieved. Little is known about the impact of different fluid solutions on different intestinal organs, and there is still an ongoing discussion about the ideal fluid solution in sepsis. The scepticism about the benefit of synthetic colloids in volume therapy is increasing [
10,
12,
24]. Colloid solutions should have a greater volume effect than crystalloids. Indeed, the colloid group had a significantly increased cardiac index, but only the HES solution resulted in an increased stroke volume index (Table
1). Despite of significantly reduced Hb levels, HES showed improved oxygen delivery, but other macrohemodynamic parameters, such as HR, MAP and CVP, did not show any differences between the septic groups (Table
1). Furthermore HES infused rats had a decreased peripheral resistance, and in line only HES showed dilatation of the intestine vessels. Additional HES improved volumetric flow of the liver sinusoids compared to Gel, NaCL and HES had no decreased microcirculation and number of capillaries under flow (CUF) in the liver compared to sham (Figure
4). Interestingly only gelatine revealed impaired gut microcirculation, whereas HES showed no such effect. RA revealed increased number of CUF and did not impair volumetric flow of the gut. The different patterns of liver and intestine microcirculation in the septic groups underline, that different organ react differently to sepsis and volume resuscitation. Taken together, 6% HES 130/0.4 revealed a higher volume effect, increased DO
2-I and showed improvement of liver microcirculation and RA might be benefical for the gut and liver microcirculation. But the improvements in the starch group did not correlate with survival. All septic animals showed the clinical signs of severe sepsis and revealed an overall mortality rate of 22%. HES, Gel and NaCL infused animals revealed mortality around 30%, whereas the RA group survived entirely. We have shown previously, that HES and Gel impair kidney function in rodent sepsis [
12]. COL solutions resulted in functional and histological impairments when compared with CL solutions. However, the effects of the different solutions on the kidney may be causative for the mortality differences between the groups.
RA was the only balanced solution in this trial, but clinical data about the influence of balanced solutions on mortality of patients remains heterogeneous. It has been stated recently by Guidet et al., that balanced crystalloid solutions do not show any effect on mortality of patients when compared with isotonic saline [
25]. In our study, animals treated with NaCl and Gel showed severe acidosis. HES treated animals, with NaCl as a part of the HES solution and saline as baseline infusion showed no deranged acid base status. However the acid–base equilibrium or the presence of a dilutional-hyperchloraemic acidosis was not the focus of this survey.
Gelatine, which is often described as safe and harmless, developed acidosis, which may be due to reduced Hb following anaerobic metabolism, and additionally Gel group showed severe hypoglycaemia which may increase lactate and aggravated metabolic acidosis. The acidosis may be the reason for the coagulopathy, but NaCl treated animal revealed also reduced pH-level and had no derangements in the coagulation parameters. However, Gelatine significantly impaired coagulation, represented by elevated PTT, INR, decreased platelet count, as well as clinical signs of coagulopathy (hematothorax, intraabdominal bleeding). This may be caused by a direct effect of gelatine on the coagulation system and on the liver coagulation factor synthesis. It is well known that artificial plasma expanders may influence the coagulation system more than crystalloids in-vitro and in-vivo beyond the effect of dilution [
26]. In addition, colloids could be stored by the reticulo endothelial system (RES) and may lead to impaired phagocytic activity particularly in septic shock [
27]. Reduced phagocytic properties of the RES lead to decreased elimination of activated fibronectin, which can be stimulated by collagenous structures such as gelatine. Thus the reduction of elimination might be another explanation for the activated coagulation system by gelatine in septic rats, which may lead to dissiminated intravascular coagulopathy.
Vlahos et al. demonstrated that CLP impaired hepatocytes exocrine synthesis, and incubation with colloids (albumin or HES) had a direct and dose depended impact on albumin and urea production [
28]. Furthermore CLP pre-treated hepatocytes developed severe morphological cell injury due to colloids despite metabolic activity in vitro. Thus significantly increased level of ALT and elevated AST in Gel group may be explained by acute liver damage. However, despite of decreased microcirculation, decreased sinusoidal diameter and impaired liver function histopathology investigation did not show any differences between the groups. This may due to the short investigation period of 24 h. In opposite to synthetical colloids RA showed no coagulopathy or liver derangements.
It has been shown in the literature, that HES exhibits an anti-inflammatory effect in different organs and species. The main differences with our experiment are the amount and incubation time of HES. Lv et al.showed an anti-inflammatory response of HES compared to CL, but they compared 6.4 ml/100gBW crystalloids up to 1,6 ml/100gBW HES within one hour infusion regime in rats [
29]. Thus the CL group might be hyperinfused and the consecutive hypervolemia increased cytokine levels itself. However, this infusion strategy (COL to CL ratio of 1:4) was the golden standard. We could show, that 24 h high colloid exposition lead to a significant increase of pro-inflammatory cytokines such as TNF-α, IL-1β and IL-6 in HES group.
Conclusion
The balanced crystalloid infusion RA could stabilize micro- and macrohemodynamic, had least side effects and all animals survived when used in septic rats. 6% HES 130/0.4 improved cardiac output, DO2-I and microcirculation of the liver, but showed elevated IL-1β, IL-6 and TNF-α levels and a mortality rate of 33%. Gelatine 4% revealed severe coagulopathy, hypoglycaemia, elevated liver enzymes, impaired microcirculation and a 29% lethality. Taken together, in our rodent model of CLP-induced severe sepsis, crystalloid infusion revealed best results in mortality and microcirculation when compared with colloid infusion.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
MAS contributed to animal preparation, performance of experimental work, analysis of mechanical and histological data, statistical analysis, and writing of the manuscript. JTI contributed to animal preparation, performance of experimental work, preliminary data analysis, and drafting of the manuscript. TS contributed to performance of experimental work, analysis of mechanical and histological data and statistical analysis. JB contributed to analysis of data and drafting of the manuscript. JS contributed to preliminary data analysis, and drafting of the manuscript. NS contributed to performance of experimental work and analysis of data. NR contributed to experimental design and writing of the manuscript. OE contributed to the experimental design. CW contributed to experimental design, supervision of experimental work, statistical analysis, writing of the manuscript, and supervision and overview of entire project. All Authors read and approved the final manuscript.