Experimental strategies in the treatment of acute renal failure in sepsis - Chapter 7: Summary and conclusion

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Experimental strategies in the treatment of acute renal failure in sepsis

Johannes, T.

Publication date 2011

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Citation for published version (APA):

Johannes, T. (2011). Experimental strategies in the treatment of acute renal failure in sepsis.

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The development of acute renal failure (ARF) in sepsis is frequent, with a prevalence up to 40%. However, its pathogenesis remains only partially understood. To date, no specific renal protective or treatment modalities are available. Therefore the treatment of intensive care unit patients with severe acute kidney injury (AKI) often results in supportive therapy like costly renal-replacement. Renal tissue hypoxia may be a contributing factor in the progression of kidney failure. The mainstay of current conservative therapeutic interventions is aimed at increasing kidney perfusion and maintaining renal function by, for example, fluid expansive therapy. However, measures that enhance kidney perfusion have the potential to exacerbate kidney hypoxia by increasing oxygen consumption. Therefore, these clinical interventions may contribute to the development of AKI instead of preventing its progression. Because of this paradox, it is of utmost clinical importance to gain further insight into the patho-physiology of ARF and to find more specific therapeutic interventions aimed at reversing ARF and preventing the development of AKI.

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In this thesis we assumed that hypoxia and microcirculatory dysfunction play a role in the pathogenesis of septic renal failure and hypothesized that strategies aimed on preventing hypoxia and microcirculatory dysfunction result in maintained kidney function. To this end we established a rat model of endotoxin-induced ARF with signs of severe hemodynamic alterations like those seen in clinical practice. Early goal-directed therapy using fluid resuscitation to prevent hypoperfusion of vital organs is the standard therapy in sepsis. Therefore, fluid resuscitation was the primary therapeutic approach in all our studies. In addition to standard fluid resuscitation different treatment strategies were performed. All substances we tested are in clinical use and act on regulating vascular tone and/or secondarily influencing microcirculatory blood flow. Besides evaluating renal function by means of creatinine clearance we established a method that allowed comprehensive measurements of renal oxygenation. Using oxygen-dependent quenching of phosphorescence we were able to simultaneously measure cortical and outer-medullary microvascular PO2 (µPO2) and renal venous

PO2. Recovery of microvascular PO2 histograms from cortex and outer medulla allowed us to study

heterogeneity of intrarenal oxygenation.

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There is evidence that tissue hypoxia and microcirculatory dysfunction are contributors in the development of septic ARF. However, only a few studies looked at renal tissue oxygenation and kidney function in different animal models under septic conditions. As mentioned before, fluid resuscitation is an early therapeutic strategy in the treatment of septic shock, with the aim of restoring blood flow and oxygen delivery to vital organs. In Chapter 1 we tested the hypothesis that renal µPO2 and oxygen consumption (VO2,ren) are impaired during endotoxemia; that this effect is

associated with a diminished renal function; that fluid resuscitation with either colloids or crystalloids improves an impaired µPO2 and oxygen consumption and restores kidney function; and that colloids

are better at resuscitating than crystalloids in this context. In our model endotoxemia was accompanied by a reduction in renal blood flow and anuria, while the renal µPO2 and VO2,ren

remained relatively unchanged. Fluid resuscitation restored renal blood flow, renal oxygen delivery and kidney function to baseline values, and was associated with a redistribution of oxygen showing

Ringer's lactate increased the renal oxygen consumption. For the first time we demonstrated the presence of renal oxygen redistribution during fluid resuscitation, but from our results we concluded that the loss of kidney function during endotoxemia could not be explained by an oxygen deficiency. HES130/0.4 had no influence on the VO2,ren and restored renal function with the least increase in the

amount of renal work.

Based on the finding that no profound tissue hypoxia or decrease in microcirculatory PO2 could

be seen during endotoxemia we performed a second study. In Chapter 2 we hypothesized that heterogeneity of microcirculatory oxygen supply to demand in the kidney is obscured when looking at the average PO2 during endotoxemia. Like demonstrated before, the average PO2 remained

relatively unchanged during endotoxemia. Only after analysis of renal oxygen distribution we were able to demonstrate the appearance of microcirculatory hypoxic areas in the rat renal cortex. This finding led to a second series of experiments in which renal blood flow was mechanically reduced to values comparable to the flow reduction seen during endotoxemia. This second series of experiments should answer the question whether the cortical microcirculatory hypoxic areas were specific for endotoxemia and not simply a nonspecific phenomenon due to reduction of renal blood flow. The significant left shift in the cortical oxygen histogram toward hypoxia like seen during endotoxemia could not be observed in animals receiving mechanical reduction in renal blood flow. In a third group of animals we studied the effect of fluid resuscitation on the reversibility of microcirculatory hypoxic areas. In these animals the cortical microcirculatory hypoxic areas disappeared after resuscitation and renal function restored to 50% of baseline. In conclusion, endotoxemia was associated with the occurrence of cortical microcirculatory hypoxic areas that are not detected in the average PO2 measurement, proving the hypothesis of our study. These

observations suggest the involvement of hypoxia in the pathogenesis of endotoxemia-induced ARF. After demonstrating the occurrence of cortical microcirculatory hypoxic areas during endotoxemia we were searching for treatment strategies aimed on preventing microvascular hypoxia and maintaining renal function. Nitric oxide (NO) is an important molecule known to act on the renal microvascular tone. Therefore NO is consequently being involved in the regulation of intrarenal oxygen supply. Under septic conditions the main production of NO derives from iNOS, an enzyme that can be blocked by the clinically used glucocorticosteroid dexamethasone (DEX). In Chapter 3 of this thesis we tested the hypothesis that inhibition of iNOS by low-dose DEX would improve an impaired intrarenal oxygenation and kidney function. Two hours after development of endotoxemic shock one group of rats received a bolus of low-dose DEX (0.1 mg/kg) in addition to standard fluid resuscitation. In these animals, the renal iNOS mRNA expression was significantly suppressed 3 hours later. Furthermore low-dose dexamethasone prevented the appearance of cortical microcirculatory hypoxic areas, improved renal oxygen delivery, and significantly restored oxygen consumption. Besides a significant increase in systemic hemodynamic, DEX restored renal function and tubular sodium reabsorption to baseline values. In conclusion, the treatment of rats with low-dose dexamenthasone in addition to fluid resuscitation reversed endotoxin-induced renal failure associated by an improvement in intrarenal microvascular oxygenation. Therefore, low-dose DEX might have potential application in the prevention of septic acute renal failure.

Another class of substances known to modulate vascular tone are prostaglandins. In the kidney, prostaglandins uphold the balance between vasodilator and vasoconstrictor to maintain homeostasis and physiologic kidney function. Prostacyclin (PGI2) must be considered to play a potential renal

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hypothesis that exogenous prostacyclin would counterbalance an endotoxemia-induced intrarenal vasoconstriction and would therefore have beneficial effects on kidney function. Continuous infusion of the prostacyclin analogue iloprost (100 ng/kg/min) 2 hours after start of LPS-infusion in addition to fluid resuscitation resulted in the stabilization of hemodynamic parameters. All animals became anuric during endotoxemia. Only in animals receiving iloprost was creatinine clearance totally restored at the end of the experiment. Iloprost had no significant effects on average µPO2, but

prevented the occurrence of cortical microcirculatory hypoxic areas. The renal expression of iNOS mRNA was significantly increased in all animals receiving LPS after 5 hours. Only in animals receiving iloprost, iNOS mRNA expression was significantly suppressed in the inner medulla. In conclusion the prostacyclin analogue iloprost significantly restored kidney function of endotoxemic rats to baseline values. This beneficial effect of iloprost on renal function might be addressed to an improvement in intrarenal oxygenation.

In the last experimental chapter of this thesis we looked at the effects of a substance with no direct effects on the vascular tone of the renal microcirculation. Activated protein C (APC) has been shown to have beneficial effects on the inflammatory process and coagulation during sepsis. Inflammation and coagulopathy impair the microvasculature and therefore disturb oxygen transport to the tissue. The hypothesis of our study presented in Chapter 5 was that APC-treatment improves renal microvascular oxygenation and kidney function in endotoxin-induced acute renal failure in the rat. Two hours after LPS-bolus rats received in addition to fluid resuscitation continuous infusion of either 10 or 100 µg/kg/h of APC. Treatment with APC 100 prevented a further decline of mean arterial blood pressure and renal blood flow. APC, independently of concentration, had no significant effects on average µPO2, but prevented the occurrence of cortical microcirculatory hypoxic areas. All

animals receiving LPS had a significant decrease in creatinine clearance. Only in animals receiving APC 100, kidney function was significantly restored at the end of the experiment. In conclusion, APC 100 significantly restored renal function compared to standard fluid resuscitation in a rat model of endotoxemia. This was accompanied by protection against the occurrence of cortical microcirculatory hypoxic areas. Furthermore, this application best improved mean arterial blood pressure. Based on these results one could hypothesize that APC 100 infusion as adjuvant to standard fluid resuscitation might be useful as a renal protective strategy to preserve renal oxygenation and kidney function in the early stage of sepsis.

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In this thesis we present the result of different experimental therapeutic strategies to preserve kidney function in a rat model of endotoxemia. Therefore we established a model showing severe hemodynamic alterations and loss in kidney function a few hours after development of septic shock. Aim of our investigations was to create a setting representing a pathology like seen in ICU patients in septic shock. Our resuscitation strategies were also based on clinical standard, where fluid resuscitation to stabilize systemic hemodynamic parameters was the primary step. All different treatment strategies were therefore an addition to fluid resuscitation.

In our first study we could demonstrate a loss in kidney function during endotoxemia, which could be reverse by fluid resuscitation. Though the average µPO2 was only slightly affected during

endotoxemia we could show the appearance of cortical microcirculatory hypoxic areas as left shift in the cortical oxygen histogram. Furthermore we demonstrated a correlation between impaired microvascular oxygenation and kidney function proving the hypothesis of this thesis. Strategies

aiming on reversing microvascular hypoxia had beneficial effects on renal function. However, the proof of a causal correlation between renal hypoxia and functional consequences is still lacking. Further research is necessary to unveil a direct link between renal tissue hypoxia, critical oxygen delivery and renal function in the pathogenesis of acute renal failure in sepsis.

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Before trying to translate our findings to human septic renal failure, it is important to comment on our specific animal model. Although our model is akin to clinical conditions animal models can never be directly translated to human pathophysiology and the clinic setting. Rodent models of endotoxemia are frequently accompanied by a hypodynamic state, intrarenal vasoconstriction and decreased renal blood flow in contrast with the hyperdynamic response seen in large animal models or in human sepsis. For this reason, the results of animal studies have to be interpreted carefully in terms of clinical relevance.

Prior to test our renal protective treatment strategies in humans it is mandatory to check the reproducibility of our result in a large animal model for example in a porcine model of endotoxin-induced renal failure. If the outcome of such a study could demonstrate to have beneficial protective effect than a big step towards a randomized clinical trial would be made. Especially in terms of the fact that all the substances we tested are registered drugs in clinical use.

Due to clinical importance of finding renal protective strategies to prevent or treat septic acute renal failure the performance of such clinical trials would be of utmost interest.