anatomy and physiology of the hepatic circulation · anatomy and physiology of the hepatic...

Anatomy and Physiology of the Hepatic Circulation

Kerstin Abshagen, Angela Kuhla, Berit Genz and Brigitte Vollmar*Institute for Experimental Surgery, University Medicine Rostock, Rostock, Germany

Abstract

The liver with its complex functions in biosynthesis, metabolism, clearance, and host defense playsa central role in the physiology of the human body. Hepatic homeostasis is tightly dependent onadequate perfusion and microcirculation. Hereby, the liver presents with unique features, such as thedual hepatic arterial and portal venous blood supply and the fenestrated sinusoids, guaranteeing thesupply of the parenchymal tissue with oxygen and nutrients and the clearance of toxicants andforeign bodies from the bloodstream. With the introduction, development, and refinement of in vivoimaging techniques, sophisticated analyses contributed markedly to our current understanding of theregulation of hepatic blood flow and microvascular perfusion in both health and disease. Thischapter will address the physiology of the hepatic macro- and microcirculation, thereby highlightingthe dual blood supply of the liver with the intimate relationship between the two vascular systemsand the regulation of sinusoidal perfusion.

Glossary of Terms

CBS Cystathionine-beta-synthetaseCO Carbon monoxideCSE Cystathionine-gamma-lyaseeNOS Endothelial nitric oxide synthaseET EndothelinH2S Hydrogen sulfideHABR Hepatic arterial buffer responseHO Heme oxygenaseHSCs Hepatic stellate cellsiNOS Inducible nitric oxide synthaseKCs Kupffer cellsNO Nitric oxideNOS Nitric oxide synthaseSECs Sinusoidal endothelial cellsaSMA Alpha smooth muscle actin

*Email: [email protected]

PanVascular MedicineDOI 10.1007/978-3-642-37393-0_142-1# Springer-Verlag Berlin Heidelberg 2014

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Dual Blood Supply of the Liver and Hepatic Macrohemodynamics

With 2.5 % of the body weight, the liver is the largest organ in the body. Hepatic blood supplyamounts to approximately 25 % of the cardiac output and is provided by two inflows, the portal veinand the hepatic artery, entering the liver at its hilus together with the hepatic bile duct, lymphatics,and nerves. The portal vein drains the splanchnic organs and carries nutrient-rich but partlydeoxygenated blood. Nevertheless, more than 50% of the hepatic oxygen requirements are providedby portal venous blood due to the fact that the portal vein contributes to the liver’s blood supply withapproximately 75–80 % of its total inflow. The remaining 20–25 % of the total liver inflow isdelivered by the hepatic artery, originating from the celiac trunk and delivering well-oxygenatedblood (Rappaport 1980; Vollmar et al. 1992; Vollmar and Menger 2009). The total hepatic bloodflow ranges between 800 and 1,200 ml/min, which is equivalent to a total hepatic perfusion ofapproximately 100 ml/min per 100 g liver wet weight (Greenway and Stark 1971).

Hepatic oxygenation depends almost equally on both large afferent vessels. As in any other arteryof the body, oxygen saturation of the hepatic artery usually exceeds 95 %. Oxygen saturation ofportal blood during the fasting state ranges up to 85 %, being greater than that of other systemicveins, and, however, substantially drops after food ingestion. It is generally accepted that 50% of theoxygen requirements of the liver are provided by portal venous blood and the other half derive fromthe hepatic artery (Vollmar and Menger 2009). If oxygen demand is increased, the liver simplyextracts more oxygen from the blood in order to maintain oxygen uptake. Oxygen consumption bythe liver accounts for 20 % of the total body oxygen consumption.

The liver is interposed in an arterial high-pressure and a venous low-pressure system (Vollmar andMenger 2009). The valveless portal vein is a low-pressure/low-resistance circuit, while the hepaticartery supplies the liver with arterial blood via a high-pressure/high-resistance system (Greenwayand Stark 1971). The mean pressure in the hepatic artery is similar to that in the aorta, while portalvein pressure has been reported to range between 6 and 10 mmHg in man when determined by directcannulation (Balfour et al. 1954) or by splenic puncture (Atkinson and Sherlock 1954). In contrast tothe hepatic artery, being a vessel of resistance, the portal and hepatic veins are vessels of capacitance.Portal pressure depends primarily on the degree of constriction or dilatation of the mesenteric andsplanchnic arterioles and on intrahepatic resistance. Both afferent systems merge at the sinusoidalbed, where the pressure is estimated to be slightly, i.e., approximately 2–4 mmHg, above that in thesmallest collecting veins or the inferior vena cava (Eipel et al. 2010b).

Though only limited data exist, it appears that hepatic blood volume ranges from 25 to30 ml/100 g liver weight, accounting for 10–15 % of the total blood volume (Lautt 1977a).Estimations of hepatic blood volume are highly variable, as indirect calculations of hepatic bloodvolume from red blood cell content of the liver and arterial hematocrit are inaccurate and hepaticvenous pressure largely influences hepatic blood volume (Greenway and Stark 1971). Further, roughestimation suggests that more than 40 % of the hepatic blood is held in large capacitance vessels(portal vein, hepatic artery, and hepatic veins), while the sinusoids accommodate up to 60% as smallvessel content (Greenway and Stark 1971). Of note is the high compliance of the hepatic vascularbed, calculated as the change in blood volume per unit change in venous pressure (Lautt andGreenway 1976). In the cat, the hepatic blood volume increased in response to elevated venouspressure and was found doubled when hepatic venous pressure was elevated to 9.4 mmHg (Lautt andGreenway 1976). Hepatic blood volume may expand considerably in cardiac failure and, in turn,serves as an important blood reservoir in case of bleeding episodes, compensating up to 25 % of thehemorrhage by immediate expelling of blood from the capacitance vessels (Lautt 2007).


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Regulation of the Hepatic Macrocirculation

The Hepatic Arterial Buffer Response Under Physiologic ConditionsThe liver is the only organ within the human body being dually supplied. This unique aspectcharacterizes the hepatic macrocirculation and distinctly determines the regulation and distributionof blood flow. To subserve the hepatic role as a regulator of blood levels of nutrients and hormones,the task of the hepatic artery is to maintain total blood inflow and thereby hepatic clearance as steadyas possible (Lautt 1977b, 1980). This is achieved by the “hepatic arterial buffer response” (HABR),representing the close relationship between the hepatic artery and the portal vein in that the hepaticartery can produce compensatory flow changes in response to changes in portal venous inflow.Burton-Opitz observed an increase in hepatic arterial blood flow upon reduced portal venous inflowas early as 1911 (Burton-Opitz 1911), a mechanism which was termed HABR seventy years later in1981 byWayne Lautt (1981). Intrinsically, the hepatic artery dilates if portal flow decreases, and thehepatic artery constricts if portal flow increases (Lautt et al. 1990; Eipel et al. 2010b) (Fig. 1). Thisinteraction between the hepatic arterial and portal venous blood flow could be confirmed in patientsby simultaneous measurement using transit time ultrasonic volume flowmetry (Jakab et al. 1995)and by applying a hybrid OD-3Dmodeling method to simulate blood flow for the liver after a virtualright lobe hepatectomy method (Ho et al. 2012). The HABR is not primarily regulated by themetabolic demand and activity of the liver mass, because a hemodilution-induced reduction ofoxygen content in the inflow and outflow vessels or a massive increase of oxygen demand does notcause hepatic arterial vasodilation (Lautt 1983; Bredfeldt et al. 1985; Lautt and Greenway 1987).Instead, HABR mainly serves for the maintenance of the total liver inflow in case of portal venous

Hepatic blood flow under physiological conditionsPortal hyperperfusion results inHABR-induced hepatic artery constriction

Surgical reduction of portal venous inflowresults in HABR-induced hepatic artery dilation

Drug-induced hepatic artery dilation

HA

PVBF-HABF-ratio: 2.5

a b

c

dPV

extended hepatectomy ortransplantation of small-for-size grafts

IVC

HA

PVBF-HABF-ratio: 29

PVBF-HABF-ratio: 22

PV

HA

PVIVC

IVC

HAPV

IVC

Fig. 1 Hepatic hemodynamics in normal and reduced-size livers. (a) Preoperative hepatic blood flow in a donor liver orbefore extended hepatectomy representing a normal portal vein blood flow (PVBF)-hepatic artery blood flow (HABF)ratio of 2.5. (b) As a consequence of portal hyperperfusion, hepatic arterial buffer response (HABR) leads to hepaticarterial hypoperfusion of the reduced-size liver that is characterized by a dramatically increased PVBF/HABF ratio of29. (c) Surgical reduction of the portal venous inflow, for example, by splenectomy or hepatic artery ligation, leads toHABR-induced dilation of the hepatic artery and results in a reduced PVBF/HABF ratio of 22. (d) Possible effects ofpharmacological interventions to preserve hepatic artery supply. PVBF/HABF ratios are adopted from Troisi and deHemptinne 2003. HA: Hepatic artery; IVC: Inferior vena cava; PV: Portal vein (with permission of World JGastroenterol from Ref. (Eipel et al. 2010b))


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blood flow changes. The increase in hepatic arterial blood flow is capable of buffering approxi-mately 25–60% of the decreased portal flow (Lautt 1983; Lautt et al. 1985) (Fig. 1). In contrast to thehepatic artery that exhibits HABR and autoregulation (myogenic constrictive response to rise inpressure), the portal vein cannot control its blood flow, being simply the sum of outflows of theextrahepatic splanchnic organs. Thus, there is no reciprocity of the HABR, i.e., alterations ofthe hepatic arterial perfusion do not induce compensatory changes of the portal vascular flow(Lautt 1981) or resistance (Legare and Lautt 1987).

Up to now, the understanding is that the communication between the hepatic artery and the portalvein is mainly driven by vasoactive mediators. Volatile substances, like nitric oxide (NO) (Mathieet al. 1991; Ayuse et al. 2010), carbon monoxide (CO) (Hoetzel et al. 2008), and hydrogen sulfide(H2S) (Siebert et al. 2008), have been invoked to contribute to this regulatory mechanism. However,they have not been extensively studied in this context yet or have not been confirmed in their actionto mediate the HABR (Grund et al. 1997). On the contrary, adenosine has been repeatedly advocatedas the putative mediator (Lautt et al. 1985; Vollmar and Menger 2009). If the portal blood flow isreduced, adenosine accumulates in or less adenosine is washed away from the space of Mall thatsurrounds the hepatic arterial resistance vessels and portal venules and is contained within a limitingplate that separates this space from other fluid compartments (“adenosine washout hypothesis”;Lautt et al. 1985) (Fig. 2). Elevated adenosine concentrations lead to a dilation of the hepatic arterywith a subsequent increase of hepatic arterial flow. In contrast to potential mechanisms, includingneural and myogenic control, which are considered unlikely (Lautt 1981, 1983) or even disproved(Mathie et al. 1980; Alexander et al. 1989), there is mounting evidence that adenosine largelymediates the HABR. Firstly, adenosine produces dilation of the hepatic artery (Lautt et al. 1985;Lautt and Legare 1985). Secondly, portal venous application of adenosine increases hepatic arterialblood flow. Thirdly, the adenosine receptor antagonists 3-isobutyl-1-methylxanthine (Lauttet al. 1985) and 8-phenyltheophylline (Lautt and Legare 1985; Richter et al. 2001b; Browseet al. 2003) reduce the HABR, while dipyridamole, an adenosine uptake antagonist, potentiates

Fig. 2 Schematic structure of the hepatic lobule. The hexagonal liver lobule represents the smallest functional unit ofthe liver. The six portal triads in the corners of the lobule harbour portal veins (blue), hepatic arterioles (red) and bileductules (green). Upon entry into the lobule, the portal venous and arterial blood sources merge and flow from theperiphery along the sinusoids towards the central vein. However, the bile as well as the lymph flow retrogradely insidethe bile duct and the space of Disse respectively towards the portal field. The space between the stroma of the portal triadand the limiting plate of hepatocytes is called space of Mall and is thought to be the site of adenosine production.Adenosine mediates the HABR in that the concentration of adenosine is regulated by washout into the portal vein and thehepatic artery. If portal blood flow is reduced, less adenosine is washed away from the space ofMall, and the elevation inadenosine levels leads to dilation of the hepatic artery with a subsequent increase in hepatic arterial flow (“adenosinewashout hypothesis”) (Lautt et al. 1985)


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the magnitude of the HABR (Lautt et al. 1985). Adenosine also activates hepatic sensory nerves tocause reflex renal fluid retention, thus increasing the circulating blood volume and maintaining thecardiac output and portal flow (Lautt 2007). However, besides adenosine and probably othervasoactive substances (NO, CO, H2S), sensory innervation and sensory neuropeptides may alsobe involved, as HABR has been demonstrated to be partially mediated by the activation of capsaicin-sensitive sensory fibers (Biernat et al. 2005).

The Hepatic Arterial Buffer Response Under Pathologic ConditionsHABR plays a fundamental role in liver homeostasis not only under physiologic conditions,including the prenatal status (Ebbing et al. 2008), but also under pathologic conditions, such ascarbon dioxide pneumoperitoneum-induced abdominal hypertension and endotoxic shock whichablate the HABR (Ayuse et al. 1995; Gundersen et al. 1996; Richter et al. 2001a). It has been shownthat the NO donor sodium nitroprusside reverses the negative effects on hepatic arterial flow inducedby endotoxin and the nonselective NO synthase inhibitor NG-nitro-L-arginine methyl ester(Gundersen et al. 1996), while at the same time endotoxin-induced inhibition of HABR is reportedto be NO independent (Ayuse et al. 1995). Astonishingly, HABR is maintained in cirrhotic livers(M€ucke et al. 2000; Richter et al. 2000; Aoki et al. 2005). Increased flow hindrance of the cirrhoticliver due to a rise in sinusoidal resistance and capillarization of the hepatic microvasculature causesa decrease in portal venous inflow and oxygen supply (Gupta et al. 1998; Richter et al. 2000). Theresultant hypoxic tissue may release adenosine and mediate vasodilation with a compensatoryincrease in hepatic arterial blood flow by the adenosine A1 receptor through a NO-dependentpathway (Zipprich 2007; Zipprich et al. 2010). Both adenosine and NO are well known as mainmediators being involved in the vasodilation of the splanchnic circulation in cirrhosis (Wiest andGroszmann 2002; Iwakiri and Groszmann 2006, 2007). The site of resistance of the hepatic arteriesis located in the presinusoidal arterioles (Takasaki and Hano 2001) where NO production ispreserved, while in the sinusoidal and postsinusoidal areas, deficiency of NO facilitates the effectsof vasoconstrictors and thus contributes to the increased intrahepatic resistance in portal hyperten-sion (Loureiro-Silva et al. 2003). In addition, H2S has been shown to be involved in the maintenanceof perfusion in liver cirrhosis via the regulation of hepatic cystathionine-gamma-lyase (CSE)expression by the farnesoid X receptor (Renga et al. 2009). Accordingly, the administration ofa synthetic farnesoid X receptor ligand to carbon tetrachloride-treated rats protected against thedownregulation of CSE expression, increased H2S generation, reduced portal pressure, and atten-uated the endothelial dysfunction of isolated and perfused cirrhotic rat livers (Renga et al. 2009).

In contrast to the situation found in cirrhotic livers, the maintenance of the HABR in livertransplants (Henderson et al. 1992; Smyrniotis et al. 2002; Demetris et al. 2006) can have detri-mental effects (Eipel et al. 2010b). As increased flow in reduced-size livers is predominantly carriedthrough the portal vein, the intact HABR causes a reduction of the hepatic arterial flow, potentiallyharming organ integrity and function (Henderson et al. 1992) (Fig. 1). Small-for-size livers, as seenin split-liver transplantation or after extended hepatectomy, are associated with portal hypertensionand high intravascular shear stress (Glanemann et al. 2005) and – as a consequence of an intactbuffer response – with poor hepatic arterial flow and vasospasm. In severe cases, this leads tofunctional de-arterialization, ischemic cholangitis, and parenchymal infarcts, also called “small-for-size syndrome” (Demetris et al. 2006). Of note, the HABR-induced decrease of the hepatic arterialblood flow can successfully be counteracted by surgical (Eipel et al. 2010b) (Fig. 3) and pharma-cological interventions, such as application of adenosine (Kelly et al. 2009) or NO (Cantréet al. 2008). Reduced portal venous flow by means of splenic artery ligation, splenic arteryembolization, or splenectomy has been shown to ameliorate the overactive HABR (Sato


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et al. 2002; Troisi and de Hemptinne 2003; Quintini et al. 2008) (Fig. 3). Besides the modulation ofthe portal venous inflow from the spleen, established techniques such as portacaval or mesocavalshunts lead to a reduction of portal hyperperfusion and an increase of hepatic arterial inflow byreversion of the HABR (Boillot et al. 2003; Taniguchi et al. 2007; Sánchez-Cabús et al. 2013; Tuet al. 2013) (Fig. 3). Extending this information, Eipel and coworkers could show that 90 %hepatectomy caused a fourfold rise in portal venous blood flow, a slight decrease of hepatic arterialblood flow with a 50 % reduction in hepatic tissue pO2, and a high mortality (Eipel et al. 2010a).These detrimental effects could be avoided by splenectomy before extended hepatectomy, whichreduced portal venous inflow and caused a rise in hepatic arterial blood flow with doubling in tissuepO2. Ligation of the hepatic artery abolished the beneficial effect of splenectomy, indicating that thesalutary effect of splenectomy in small-for-size livers has to be attributed to a rise in hepatic arterialblood flow with sufficient oxygen supply rather than to a reduced portal venous hyperperfusion tothe remnant liver (Eipel et al. 2010a).

Anatomy of the Hepatic Microvasculature

Hepatic microvascularization comprises two afferent vessels (arterial and portal terminal branches),the sinusoids and the terminal hepatic or postsinusoidal venule. The knowledge on the anatomy ofthe hepatic microvascular bed, in particular the terminal distribution of the afferent vessels and thesinusoids, originates from detailed studies using light microscopy, intravital fluorescence micros-copy, and transmission and scanning electron microscopy (Burkel 1970; Kardon and Kessel 1980;Yamamoto et al. 1985; Menger et al. 1991; Pannarale et al. 2007).

The hepatic artery and the hepatic portal vein enter the liver at its hilus at which they undergorepeated branching into the terminal vessels, i.e., terminal hepatic arterioles and terminal portalvenules with a diameter of 15–35 mm and a length of 50–70 mm (Oda et al. 2003), which thenprovide the blood supply to the hepatic tissue nourishing sinusoids. Sinusoids are the hepaticcapillaries and, however, present with two specific peculiarities, namely, the presence ofa fenestrated endothelium and the absence of a basement membrane (Wisse 1970). The branchesof the hepatic artery and the hepatic portal vein travel through the liver parenchyma parallel to each

IVC

Liver

HASpA

SpleenBDPV

Fig. 3 Surgical interventions for modulation of the hepatic inflow, showing the portocaval shunt (white arrow), ligationof the splenic artery (black arrow) and splenectomy (grey arrow). BD: Bile duct; HA: Hepatic artery; IVC: Inferior venacava; PV: Portal vein; SpA: Splenic artery (modified from Ref. (Eipel et al. 2010b))


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other within the portal tracts, being accompanied by branches of the main bile duct and the mainlymphatic vessels. Except for the cirrhotic liver with deterioration of trans-sinusoidal macromolec-ular exchange, where lymph vessels massively expand due to their drainage function and gainvisibility within the portal tracts (Vollmar et al. 1997), lymphatic vessels are often collapsed andinconspicuous. Thus, only three structures are regularly visible within the portal tracts with theconsequence that they are often referred to as portal triads (Figs. 2 and 4a).

After passage through the sinusoids, blood drains into postsinusoidal venules and is collected insmall branches of hepatic veins, several of which may combine, increase in diameter, and reach thesublobular vein and hepatic veins, which then leave the liver on the dorsal surface and extend to theextrahepatic inferior caval vein (McCuskey 2000). The proper architecture of the liver parenchymawith concepts of the liver lobule (Figs. 2 and 4b, c), the portal unit, the liver acinus, and otherstructures has been the object of controversial discussions for centuries (Sasse et al. 1992). Thoughthe liver microcirculation can be organized both into morphological and functional units, thehexagonal hepatic lobule seems to be the concept most consistent with existing evidence(Ekataksin and Kaneda 1999; Malarkey et al. 2005) (Figs. 2 and 4b, c) and has displaced the unittermed “simple liver acinus,” being first proposed by Rappaport (1958, 1973). The hepatic micro-circulatory unit in principle consists of the two terminal afferent vessels within the portal tractsdistributed along the lobular periphery, the network of sinusoids running between the liver cords andthe efferent postsinusoidal venule in the center of the lobule (Figs. 2 and 4b, c). This functionalsubstructure conforms to the basic idea of a microcirculatory unit and is also in line withthe proposed gradient of hepatocellular metabolic heterogeneity along the sinusoidal path(Teutsch et al. 1999).

Hepatic arterioles communicate with portal venules (McKee Olds and Stafford 1930; Bloch 1955;Knisely et al. 1957; Burkel 1970; McCuskey 2000; Oda et al. 2003, 2006) in that hepatic arterioleswind themselves around the portal venules sending short branches (i) to the portal venules, theso-called arterio-portal anastomoses, and (ii) to the capillaries of the peribiliary plexus, whichsupport the bile duct and drain into the sinusoids via arterio-sinus twigs. In the very periportalregion of liver parenchyma, these little branches consist of a complete basement membrane andunfenestrated endothelium, while further downstream they resemble true sinusoids, as they lose theirbasement membrane and become fenestrated. The terminal hepatic arteriole-derived capillariesfurther supply (iii) the portal venular wall as vasa vasorum and (iv) the connective tissue including

Fig. 4 Histology of the liver. (a) Haematoxylin- and eosin-stained liver section from a normal mouse showing the portaltract which consists of the portal vein (PV), branches of the hepatic artery (HA), and tributaries to the bile duct (BD). (b)Histological representation of the hexagonal hepatic lobule (black lines) with the central vein in the middle and the portaltriads at the periphery, as well as the diamond-shaped hepatic acinus (red line) which is centered on the line connectingtwo portal triads and extends outwards to the two adjacent central veins. (c) Immunohistochemical staining for collagen1expression in carbon tetrachloride-treated mouse liver. This matrix protein is highly expressed by hepatic stellate cells inthe perisinusoidal space upon liver injury. It can clearly be seen that collagen1 expression is restricted primarily to thearea of the limiting plate, displaying the structure of the hepatic lobule


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the nerves of the portal tract (Oda et al. 2003, 2006). Using the enzyme histochemistry of alkalinephosphatase activity, the identification of endothelial cells of the arteriolar capillaries within themicrovascular system allowed the distinct evaluation of the terminal distribution of the hepaticartery and its involvement in the control of hepatic sinusoidal blood flow (Oda et al. 2006). Ourgroup could demonstrate that shunting of blood via hepatic arterio-portal anastomoses is essentialfor the maintenance of nutritional microvascular supply and oxygen delivery in HABR of rat livers(Richter et al. 2001b). However, there are also reports about the existence of liver areas beingexclusively supplied by arterial or portal venous blood, causing a considerable heterogeneity withinthe hepatic microcirculation (Timm and Vollmar 2013) (Fig. 5).

The capillary bed, namely, the hepatic sinusoids (Fig. 6a), serves to supply tissue with oxygen andnutrients and removes metabolic products. Between the liver cords, sinusoids run straight from theperiportal to the pericentral area over a length of approximately 250 mmwith an increase in diameterfrom ~7 to ~15 mm (Fig. 6a). Sinusoids communicate with each other through shorterinterconnecting sinusoids running across the liver cell cords. The endothelial lining of sinusoids ischaracterized by cells with flattened processes and open fenestrations, being arranged in clusters of10–50 pores, forming the so-called sieve plates with a diameter of 150–175 nm, and occupyingabout 6–8 % of the endothelial surface (Fig. 7). In response to alterations of sinusoidal blood flowand perfusion pressure, these fenestrae can contract and dilate and thus act as a selective sievingbarrier to control the extensive exchange of material between the blood and the liver cells, and viceversa, contributing to the homeostatic control of the hepatic microcirculation. Their distinctnonuniform distribution in size and number, i.e., a decrease in diameter but an increase of frequency

Fig. 5 Analysis of portal venous blood flow by injection of a low-molecular fluorescent compound (sodium fluorescein)that almost freely diffuses from the intravascular sinusoidal towards the extravascular space and is therefore generallyused for tissue contrast enhancement during in vivo microscopy of the liver. After injection via the V. lienalis, onlineobservations of the dye appearing in the liver showed that this low-molecular substance disperses inhomogeneouslywithin the liver, resulting in a differential hepatocellular uptake. Images show wide areas of acini marked by intense,moderate and no staining corresponding to high, moderate and negligible portal venous blood supply (with permissionof Microvasc Res from Ref. (Timm and Vollmar 2013))


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from periportal to pericentral zones, allows the higher centrilobular porosity (Wisse et al. 1985;Braet and Wisse 2002). These membrane-bound pores lack a diaphragm and a basal lamina, thuscontrasting the fenestrae described in the kidney, pancreas, and brain (Braet and Soon 2005).Besides their contribution to liver microcirculatory homeostasis, they exert scavenger function bytheir high receptor-mediated endocytotic capacity and clear the blood of many macromolecularwaste products (Smedsrød et al. 1994). In addition, they have been shown to play a yet overlookedfunction in liver immunology by allowing naive T cells to interact with hepatocytes and thusimmune tolerance, as lymphocytes pass through the liver (Warren et al. 2006). Meanwhile, innova-tions in correlative imaging techniques by using combined light, probe, laser, and various electronmicroscope techniques and their application in the hepatic endothelial cell model allow novelinsights into the subcellular components and supramolecular structures of the liver sieve (Braetet al. 2007, 2009). They will help to better understand their biological relevance in various diseases,such as fibrosis, cirrhosis, atherosclerosis, and cancer (Braet and Wisse 2002). The most recently

Fig. 7 Lumen of a hepatic sinusoid with the endothelial cell coating, displaying the typical fenestrations arranged asclusters and forming the so-called sieve plates. Freeze-fracture technique and scanning electron microscopy of the ratliver are shown. Magnification x12,000 (with permission of Physiol Rev from Ref. (Vollmar and Menger 2009))

Fig. 6 Representative intravital fluorescence microscopic images of the mouse liver in epi-illumination techniquewhich show the hepatic microcirculation with the sinusoidal perfusion under physiologic (a) and pathologic conditions(b). (a) In detail, communicating networks are developed between the portal venous and the hepatic arterial circulation.After repeated branching, the terminal hepatic arterioles and terminal portal venules supply the blood to the hepaticsinusoids in which the blood flow direction (arrows) occurs from periportal to pericentral. Main sinusoids run straightbetween the liver cell cords over a length of�250 mm and communicate with each other through shorter interconnectingsinusoids running across the liver cell cords. (b) Deterioration of the hepatic microcirculation due to an acute liverfailure, e.g., in galactosamine/lipopolysaccharide-treated mice, is characterized by a pronounced perfusion failure of upto 80% non-perfused sinusoids. Note the hemorrhagic tissue with conglomerates of erythrocytes representing the severeliver injury


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described synchrotron radiation micro-computed tomography will further provide an improvedunderstanding of the pathogenesis of hepatic disease by detecting hepatic sinusoids and theiralterations in three-dimensional structures of the damaged liver (Yoon et al. 2013).

The space of Disse, i.e., the space between the sinusoidal endothelial cells (SECs) and themicrovilli-rich hepatocellular surface, is the location of the fat- and vitamin A-storing perisinusoidalcells, known as Ito or hepatic stellate cells (HSCs) (Friedman 2008) (Fig. 8). With their cell bodiesoften found in the recesses between hepatocytes, HSCs encircle the sinusoidal tube with theirperisinusoidal processes and numerous fingerlike secondary branches (Blomhoff and Wake 1991).Their perisinusoidal orientation within the space of Disse but in particular their endowment withdesmin, alpha smooth muscle actin, and microfilaments and, thus, remarkable capacity for cellularcontraction constitute HSCs as the dominant regulator of blood flow through hepatic sinusoids(Rockey 2001; Soon and Yee 2008) (Fig. 9). Besides that, HSCs are well known for their importancein retinol metabolism and as key actors in the hepatic fibrogenic response to injury (Vollmaret al. 1998; Tacke and Weiskirchen 2012). Most recently, HSC-specific gene targeting has beendeveloped to combat HSC activation-associated transformation into extracellular matrix-synthesizing myofibroblasts and thus fibrogenesis (Bansal et al. 2011; Poelstra and Schuppan2011; Reetz et al. 2013) (Fig. 9).

In contrast to the perisinusoidal location of HSCs, Kupffer cells (KCs) are anchored to the luminalsite of the endothelium and, thus, exposed to the bloodstream (Fig. 8). By their large bodiesprotruding into the sinusoidal lumen, KCs represent a flow hindrance and are – next toHSCs – also considered to contribute to blood flow regulation through sinusoids (Arii and Imamura2000; McCuskey 2000; Abshagen et al. 2008). KCs are attached to the endothelium by cytoplasmicprocesses, which sometimes also anchor across the lumen to the opposite sinusoidal wall. In contrastto the homogeneous distribution of HSCs throughout the liver lobule (Suematsu et al. 1993; Vollmaret al. 1998), there is a gradient of KCs with higher numbers, larger size, and greater phagocyticactivity in periportal than pericentral regions (Bouwens et al. 1986, 1992). This results in a zonaldistribution with a specific kinetics of phagocytosis (Vollmar et al. 1994b).

Fig. 8 Schematic illustration of the structure of the liver cell cord. The hepatic parenchyma consisting of hepatocytes(HC) is separated from the sinusoids by the space of Disse. The sinusoid itself is composed of nonparenchymal cells,such as fenestrated sinusoidal endothelial cells (SEC), hepatic stellate cells (HSC), which are located in the Space ofDisse and resident liver macrophages, so called Kupffer cells (KC) which are located inside the sinusoid


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Regulation of the Hepatic Microcirculation

Physiology of the Hepatic MicrocirculationThe hepatic microcirculation is mainly controlled by the contractile cells of the sinusoids, such asHSCs, SECs, and KCs. Due to the interposition of the liver in an arterial high-pressure and a portalvenous and venous low-pressure system, there are steep blood pressure gradients from the portalvenous system with ~50 mmH2O but in particular from the hepatic arteriolar system with ~300–400mmH2O into the sinusoids with ~10–20 mmH2O (Nakata et al. 1960). They are suggested to bemaintained by the existence of sphincter-like specialized structures at the entrance to and at the exitfrom the sinusoids (Knisely et al. 1957), though the existence of sphincter-like structures per se andtheir constitution of cellular components are not definitely clarified (for details, see Vollmar andMenger 2009).

In contrast to that, there is clear evidence that the hepatic microvascular blood flow is regulatedand redistributed at the level of the microcirculation, namely, by HSCs, KCs, and SECs, and theirbroad responsiveness to vasoactive mediators. Dominant players in the delicate control of thevascular tone under both physiologic and pathologic conditions are the endothelins (ETs) and thegaseous mediators NO, CO, and H2S.

Hepatic cellular sources of ETs, with ET-1 being the most predominant player among the twoother isopeptides ET-2 and ET-3, are SECs, HSCs, and KCs (Rockey et al. 1998; Pannen 2002).ET-1 acts via the ETA receptor and results in a constriction of the effector cell. On the contrary,binding of ET-1 to the ETB1 receptor on endothelial cells may result in a NO-dependent relaxation of

Fig. 9 Representative images of primary hepatic stellate cells (HSCs) in vitro (upper panel) and in vivo (lower panel).(a) Immunohistochemical staining of alpha smooth muscle actin (αSMA) of culture-activated HSCs showingmyofibroblast-like phenotype with long processes. (b) Vitamin A-autofluorescence of primary HSCs upon UV lightepi-illumination. (c) Fluorescence microscopic images of αSMA expression (blue) of adenoviral-transduced (greenfluorescent protein) HSCs. (d) Immunohistochemical staining for cytoskeletal αSMA expression (brown) in carbontetrachloride-injured mouse liver. This protein is highly expressed upon liver injury in activated HSCs. (e) Represen-tative intravital fluorescence microscopic images of the autofluorescence of vitamin A in the lipid droplets of HSCs uponUV light epi-illumination. (f) Representative images of GFP-positive activated HSCs upon adenoviral gene transfer,displayed in higher magnification showing a myofibroblast-like phenotype of HSCs as well as sinusoidal lining byprocesses of HSCs


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adjacent vascular smooth muscle cells and pericytes, while ETB2 receptor stimulation induces cellcontraction. The function and significance of ET-1 strongly depend not only on the local generationby the abovementioned cellular sources but also on the magnitude of receptor expression and itstemporal and spatial variations. ET receptor expression is found highest on HSCs, followed by SECsand KCs (Housset et al. 1993). ET-1 also causes reversible and graded contraction of isolated stellatecells in culture (Pinzani et al. 1992; Rockey 1995) and narrows the sinusoidal lumen in isolatedperfused livers (Zhang et al. 1995; Rockey andWeisiger 1996) as well as in livers with intact afferentand efferent nerves upon systemic or intraportal infusion (Bauer et al. 1994). Intravital fluorescencehigh-resolution microscopy could demonstrate that the ET-1-induced reduction of sinusoidal diam-eter colocalized with the body of stellate cells, underlining that these liver-specific pericytes serve ascontractile targets for ET (Zhang et al. 1995). Besides this intrasinusoidal site of action of ET,intraportal infusion of ETcauses also an intense contraction of perivascular smooth muscle cells andprotrusion of endothelial cells into the lumen of preterminal portal venules. This indicates that thesemicrovascular segments can function as presinusoidal quasi-sphincters (Kaneda et al. 1998).

Within the hepatic microcirculation, counterplayers of the ETs are the mediators NO and CO,guaranteeing the critical balance between vasoconstriction and vasodilation of hepatic sinusoids(Pannen et al. 1996a). In hepatic arteriolar resistance vessels, the constitutive NO synthase ofendothelial cells (eNOS) produces and releases NO (Shah et al. 1997), which acts in a paracrinefashion by diffusing into smooth muscle cells. There, NO reacts with the ferrous iron in the hemeprosthetic group of the soluble guanylate cyclase that increases the concentration of cyclic guano-sine monophosphate and activates a cell signaling pathway, which results in smooth musclerelaxation (Rodeberg et al. 1995; Tran et al. 2003). In line with that, either L-arginine or NO donorsincrease liver blood flow which can be attenuated by inhibitors of NOS or cyclic guanosinemonophosphate (Zhang et al. 1997; Mittal et al. 1994; Shah et al. 1997). The higher shear stressand shear stress-dependent release of NO in the arterial compared to the venous bed may beresponsible for the fact that NO serves as a potent vasodilator in the hepatic arterial circulationbut exerts only a minor vasodilatory effect in the portal venous bed (Pannen and Bauer 1998).

Using cultured SEC exposed to flow in a parallel-plate flow chamber, it has been shown that thesecells respond to an increase in flowwith an enhanced production of NO (Shah et al. 1997). NO is alsoreleased from isolated perfused livers in a time- and flow-dependent manner (Shah et al. 1997). Asthe hepatic sinusoids contain no smooth muscle layer, it can be assumed that basal and flow-dependent NO release regulates vessel resistance bymodulating stellate cells. This view is supportedby the fact that adenoviral gene vector-based expression of neuronal NOS in SECs, HSCs, andhepatocytes increases NO production and inhibits ET-1-induced contractility of perisinusoidalstellate cells in normal livers (Yu et al. 2000).

There is convincing evidence by the sophisticated work of Suematsu and coworkers using dual-color digital microfluorography that CO – comparably to NO – functions as an endogenousmodulator of hepatic sinusoidal perfusion through a relaxing mechanism involving HSCs, as themajor sites of action of CO corresponded to local sinusoidal segments colocalized with HSCs(Suematsu et al. 1995; Goda et al. 1998). Next to biliverdin and iron, CO is released in equimolarconcentrations through the physiological microsomal catabolism of the heme molecule by theenzyme heme oxygenase (HO), which consists of two distinct isoenzymes termed HO-1 andHO-2. Under physiologic conditions, HO-1, the inducible form, is only found in KCs, while theconstitutive form HO-2 is distributed to parenchymal cells. It has been shown that CO derived fromHO-2 in hepatocytes contributes to active relaxation of the sinusoids (Goda et al. 1998). In addition,functional blockade of the CO-releasing HO-1 by zinc protoporphyrin IX inhibited baseline COgeneration and caused an increase of resistance, a discrete pattern of constriction in sinusoids, and


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a reduction of the sinusoidal perfusion velocity (Suematsu et al. 1995). Further, CO maintains portalvenous vascular tone in a relaxed state, while there is no intrinsic CO-mediated vasodilation in thehepatic artery (Pannen and Bauer 1998).

Though the third gaseous mediator H2S is much less studied with respect to the regulation of thehepatic microcirculation, H2S is known for its potent vasodilatory effects in both the systemic andthe splanchnic circulation (Fiorucci et al. 2005, 2006). In normal livers, H2S released from eitherexogenous (sodium hydrosulfide supplementation) or endogenous (L-cysteine supplementation)sources reverses the norepinephrine-induced increase of portal pressure (Fiorucci et al. 2005). UsingRT-PCR, Western blot analysis, and immunohistochemistry, our group could demonstrate theexpression of both H2S-synthesizing enzymes, i.e., the CSE and the cystathionine-beta-synthetase(CBS), in the aorta, vena cava, hepatic artery, and portal vein, as well as in hepatic parenchymaltissue (Siebert et al. 2008) (Fig. 10). SECs express none of the H2S-generating enzymes CSE andCBS and do not release H2S (Fiorucci et al. 2005). This is in line with the observation that H2Sreleased by normal rat livers is not modified by an increased shear stress and that modulation ofintrahepatic resistance by H2S is not dependent on NO, underscoring sites different from that ofSECs being involved in the synthesis of H2S (Fiorucci et al. 2005). Indeed, HSCs express CSE andcan release H2S (Fiorucci et al. 2005). L-Cysteine, a substrate of CSE, decreased vasoconstriction innormal rat livers, and the H2S-induced relaxation of hepatic microcirculation was attenuated byglibenclamide, an adenosine triphosphate-sensitive K+ channel inhibitor (Fiorucci et al. 2005),underlining the vasorelaxant action of H2S via the activation of adenosine triphosphate-dependent K+channels (Tang et al. 2005).

In summary, blood flow regulation through sinusoids resides in the sinusoids themselves. Withinthe extraordinary complex interplay of cellular components and vasoactive mediator systems, NOand CO may exert synergistic vasodilatory effects due to their common stimulation of solubleguanylate cyclase. Further, NO may either attenuate or enhance HO-1 gene expression (Motterliniet al. 1996; Hoetzel et al. 2001). NO and CO can functionally antagonize the vasoconstrictive effectsof ETs (Suematsu et al. 1996). NO limits ET-1 release from endothelial cells (Brunner et al. 1995)and is capable of terminating ET signaling (Goligorsky et al. 1994). Finally, NO is a physiologicalmodulator of the endogenous production of H2S by increasing the expression and activity of CSE(Zhao et al. 2001). However, it has been shown that – though endothelial dysfunction caused byhyperhomocysteinemia comprises defective NO bioavailability – homocysteine-induced impair-ment of NO release could be reversed by H2S in a NO-independent manner (Distrutti et al. 2008),implying an additive rather than a synergistic effect of NO and H2S in the liver microcirculation.

Dysfunction of the Hepatic Microcirculation in InjuryBesides inflammation, which will not be addressed in this book chapter, hepatic injury is caused bymicrocirculatory dysfunction, involving the action of potent vasoactive mediators, such as ETs,thromboxane A2, angiotensin II, and catecholamines as vasoconstrictive and NO, CO, H2S, andprostaglandins as vasodilative substances (Vollmar and Menger 2009). While under physiologicconditions the influence of these vasoactive mediators on intrahepatic blood flow is rather small andtheir effects are balanced, any adverse stimulus, however, may lead to an upregulation of constrictoror dilator influences at all microvascular segments of blood flow regulation. These are mostlymatched neither for time nor for space and may therefore cause a imbalance of vasomotor activityand vasotonus. Thus, the maintenance of a critical balance seems to be an attractive concept for anadequate regulation of hepatic blood flow, aiming at limiting microcirculatory dysfunction-associated liver injury (Pannen 2002; Palmes et al. 2005a).


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Reactive oxygen species, endotoxins, cytokines, hypoxia, and vascular shear stress can induce ETgene expression (Rubanyi and Polokoff 1994). Because all of these mediators play an essential rolein numerous pathological states of the liver, they have been described to be associated with increasedgeneration and plasma concentrations of ET-1 and substantial changes of the ET receptor expressionpattern (for a review, see Vollmar and Menger 2009). Intriguingly, the downregulation of the ETAreceptor expression under pathologic conditions, such as hepatic ischemia/reperfusion (Soninet al. 1999; Yokoyama et al. 2000; Kim and Lee 2004), endotoxemia (Sonin et al. 1999; Bavejaet al. 2002; Kim and Lee 2004), and portal vein ligation (Yokoyama et al. 2001), may inducevasodilation by binding of ET-1 to the ETB1 receptor on SECs, thereby attenuating its own

Fig. 10 Representative RT-PCR analysis of cystathionine-gamma-lyase (CSE) (a) and cystathionine-beta-synthetase(CBS) mRNA expression (e) in vascular and liver tissue. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) servedas internal control. Representative immunohistochemical images of CSE (left) and CBS expression (right) of the hepaticartery (HA; b) and portal vein (PV; F), aorta and vena cava (V. cava) (c, g), as well as liver tissue, displaying a portal triad(d, h); arteria hepatica (A. hepatica) (with permission of Am J Physiol Gastrointest Liver Physiol from Ref. (Siebert et al.2008))


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vasoconstrictive effects mediated via ETA and ETB2 receptors on HSCs (Pannen 2002). This impliesa self-restricting process, because in the absence of this ET-mediated vasodilatory effect, the extentof ischemia/reperfusion-induced liver injury has been found to be even greater (Pannen et al. 1998).In general, under pathologic conditions, the hepatic microcirculation is sensitized to ET-1. EnhancedET-1 responses occur at both sinusoidal and presinusoidal levels, as – for example – shock- andresuscitation-primed livers respond to ET-1 with a greater increase in portal driving pressure, totalportal resistance, and zero-flow pressures as well as with a more pronounced decrease in portal flowand sinusoidal diameters (Pannen et al. 2001). Comparably, microvascular hyperresponsiveness toET-1 has been described in endotoxin-pretreated livers (Pannen et al. 1996b) or in livers duringpolymicrobial sepsis (Baveja et al. 2002).

ET-induced vasoconstriction can be functionally antagonized by the vasodilative NO and CO. Invitro studies showing that CO-exposed endothelial cells react with an increase of endothelial cyclicguanosine monophosphate content and a decrease of ET-1 expression (Morita and Kourembanas1995) or that NO donors reduce ET-1 secretion from endothelial cells (Brunner et al. 1995;Mitsutomi et al. 1999) imply that the close interaction of these vasoactive mediators could neutralizethe effects of each other. However, there is large body of evidence that an imbalance of theexpression of stress-induced vasoactive mediators is responsible for the alterations of the livermicrocirculation and the subsequent tissue injury.

Marked vasoregulatory imbalances due to the loss of the delicate equilibrium of vasoactivemediators were shown by the ischemia- and endotoxemia-induced upregulation of mRNA encodingET-1 (2.5- and 6-fold), HO-1 (2- and 2.5-fold), and inducible nitric oxide synthase (iNOS) (6.4- and>24-fold) (Sonin et al. 1999). In addition, HO-1 (16-fold) and ET-1 (9-fold) mRNA, but not iNOS,are found upregulated in hemorrhagic shock-exposed rat livers (Rensing et al. 1999; Rensinget al. 2002). Also microcirculatory injury in small-for-size liver grafts is related to an upregulationof ET-1 and iNOS, leading to a deterioration of intracellular homeostasis, as reflected by thedownregulation of HO-1 (Liang et al. 2003). The remarkable potential of these dysregulated enzymesystems in deterioration of the hepatic microcirculation is underlined by the fact that NO donors andET receptor blockers protect the postischemic liver microcirculation by maintaining the balancebetween NO and ET (Mitsuoka et al. 1999; Scommotau et al. 1999; Uhlmann et al. 2000). In linewith this, postischemic reperfusion injury of the liver benefits from blockade of excessive iNOSexpression, decreasing NO and hydroxyl radical production (Lin et al. 2004). There is evidence thatthe constitutive release of NO by the eNOS is mandatory as a physiological counterpart for the actionof ET, while the excessive release of NO by the iNOS can be harmful and might contribute tomicrovascular dysfunction and hepatic injury, not least because of the subsequent nitrogen speciesproduction and nitrosative stress (van Golen et al. 2013). In line with this, it has been shown thatrepeated cycles of hepatic ischemia and reperfusion caused a gradual attenuation of activatedphospho-eNOS but a gradual increase of iNOS expression (Miyake et al. 2013). The protectivefunction of eNOS and the rather injurious effect of an upregulation of iNOS (Matejovic et al. 2004;Eum et al. 2007) are further underscored by the fact that the administration of arginine together witha selective iNOS inhibitor during the early phase of sepsis restores plasma arginine, reducesoxidative stress by maintaining NO derived from constitutive NOS, and attenuates neuroendocrinestress responses (Xie et al. 2008). In contrast, unselective inhibition of NO activities aggravates liverinjury (Takemura et al. 2000).

Comparably to eNOS, HO-1 upregulation confers cytoprotection in many models of organ andtissue injury (Wunder et al. 2004; Ryter and Choi 2007; Fang et al. 2011) and CO ameliorateshepatobiliary dysfunction caused by heme overloading in endotoxemic livers (Kyokane et al. 2001).Accordingly, HO-1 deficiency exhibits a heightened and dysregulated inflammatory response to


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endotoxin (Tracz et al. 2007). Of interest, the cytoprotection of the HO-1 metabolite bilirubin ismediated at least in part through the inhibition of iNOS expression, underlining the close interactionof these mediators controlling sinusoidal vasotonus (Wang et al. 2004). Cirrhotic rat livers showeddecreased expression of both HO isoenzymes and reduced HO activity when compared to normal ratlivers (Van Landeghem et al. 2009). Consequently, the increased intrahepatic vascular resistance ofcirrhotic rats could be attenuated by perfusion with the CO-releasing molecule (CORM)-2 and theHO-1 inducer hemin, while the inhibition of HO by the HO inhibitor zinc protoporphyrin IX causeda dose-related increase in intrahepatic vascular resistance in normal and cirrhotic liver. Thus, innormal liver, the hemodynamically relevant CO production seems to occur extrasinusoidally, whileintrasinusoidally HO-1 may predominantly regulate the microcirculation in cirrhotic livers (vanLandeghem et al. 2009).

Currently, there is very little information in the literature regarding the effect of H2S on the hepaticmicrocirculation, particularly during pathologic conditions, such as sepsis. Despite being protectivein many disease states, portal infusion of H2S increased portal pressure, decreased sinusoidaldiameter, and increased sinusoidal heterogeneity and net constriction in an endotoxin model ofsepsis in rats (Norris et al. 2013a). Increased H2S levels during sepsis were further shown tocontribute to the hypersensitization of the sinusoid to the vasoconstrictor effect of ET-1 (Norriset al. 2013b). The CSE inhibitor DL-propargylglycine significantly abrogated the increase insinusoidal constriction and attenuated the increase in NADH fluorescence following ET-1 exposureduring endotoxemia, suggesting an improvement in hepatic oxygen availability (Norriset al. 2013a). Thus, the vasoconstrictor action of H2S on the hepatic sinusoid may contribute tothe progression of sepsis by enhancing microvascular perfusion failure (Norris et al. 2013a, b).

Besides HSC-mediated vasoconstriction (Bauer et al. 1994; Pannen et al. 1998; Clemens andZhang 1999) and impaired sinusoidal vasomotor control, several other mechanisms can lead tosinusoidal perfusion failure. Structural peculiarities of the sinusoid, including diameter, tortuosity ofpath, branching pattern and number, size, and activity of KCs with the regional differences betweenperiportal and pericentral areas, predispose to flow heterogeneities within the liver. These causea gradient of perfusion failure, which is most pronounced in the periportal segment of the sinusoids(Vollmar et al. 1994a; Brock et al. 1999). Edema of SECs and entrapment of activated leukocytesmay also lead to an increase of flow hindrance, which further induces perfusion heterogeneity andperfusion deficits (Chaudry et al. 1981; Vollmar et al. 1994c, d, 1996b). In addition, inflammation-and injury-associated adherence of leukocytes in postsinusoidal venules (Vollmar et al. 1994d,1995) may alter sinusoidal perfusion due to an increase of blood viscosity (Chien 1985) and, hence,vascular resistance (Braide et al. 1984). Further, perfusion failure in sinusoids is thought to be causedby sluggish blood flow, intravascular hemoconcentration, and procoagulant conditions (Mengeret al. 1999).

Failure of sinusoidal perfusion is discussed as key factor in the pathogenesis of tissue injury inwarm ischemia/reperfusion (Vollmar et al. 1994a; Khandoga et al. 2006; Burkhardt et al. 2008), coldpreservation and transplantation (Koeppel et al. 1996; Mehrabi et al. 2003; Rentsch et al. 2005),shock and resuscitation (Vollmar et al. 1994c; Roesner et al. 2006; Kubulus et al. 2008),endotoxemia (Li et al. 2004; Singer et al. 2006; Slotta et al. 2006), acute liver failure (Palmeset al. 2005b; Eipel et al. 2007; Le Minh et al. 2007; Kuhla et al. 2013) (Fig. 6b), bile duct ligation(Abrahám et al. 2008; Laschke et al. 2008), and drug-induced hepatotoxicity (Bauer et al. 2000;Randle et al. 2008). The severity of sinusoidal perfusion failure is particularly dependent on thenature of the injurious stimuli, on the animal and strain of species used, and on the time point ofassessment but can amount to 40–60 % in models of severe liver injury, such as reperfusion upon24 h of cold storage (Koeppel et al. 1996) or galactosamine/endotoxin exposure for induction of


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fulminant liver failure (Le Minh et al. 2007; Kuhla et al. 2013). Microcirculatory failure in the liverafter stress is characterized by perfusion heterogeneity, resulting in a mismatch between oxygen andnutrient supply and demand. The impaired nutritive blood flow accompanied by reduced oxygenavailability decreases cellular levels of high-energy phosphates and contributes to early and latehepatocellular injury and dysfunction, being underlined by significant correlations between micro-circulatory disorders and hepatocellular disintegration or liver dysfunction (Vollmar et al. 1994a,1996a) as well as between reduced blood oxygenation index and increased transaminases (Gotoet al. 1992). This underlines the determinant role of an intact hepatic microcirculation for theadequate organ integrity and function (Menger et al. 1999).

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