ArticlePDF AvailableLiterature Review

Role of liver sinusoidal endothelial cells in nonalcoholic fatty liver disease

February 2019
Journal of Hepatology 70(6)

February 2019
70(6)

DOI:10.1016/j.jhep.2019.02.012

Authors:

Université de Paris

Non-alcoholic fatty liver disease (NAFLD) and its complications are an expanding health problem associated with the metabolic syndrome. Liver sinusoidal endothelial cells (LSECs) are highly specialized endothelial cells localized at the interface between the blood derived from the gut and the adipose tissue on the one side, and other liver cells on the other side. In physiological conditions, LSECs are gatekeepers of liver homeostasis. LSECs display anti-inflammatory and anti-fibrogenic properties by preventing Kupffer cell and hepatic stellate cell activation and regulating intrahepatic vascular resistance and portal pressure. This review focusses on changes occurring in LSECs in NAFLD and on their consequences on NAFLD progression and complications. Capillarization, namely the loss of LSEC fenestrae, and LSEC dysfunction, namely the loss of the ability of LSECs to generate vasodilator agents in response to increased shear stress both occur early in NAFLD. These LSEC changes favour steatosis development and set the stage for NAFLD progression. At the stage of non-alcoholic steatohepatitis, altered LSECs release inflammatory mediators and contribute to the recruitment of inflammatory cells, thus promoting liver injury and inflammation. Altered LSECs also fail to maintain hepatic stellate cell quiescence and release fibrogenic mediators, including Hedgehog signalling molecules, promoting liver fibrosis. Liver angiogenesis is increased in NAFLD and contributes to liver inflammation and fibrosis, but also to hepatocellular carcinoma development. Thus, improving LSEC health appears to be a promising approach to prevent NAFLD progression and complications.

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Role of liver sinusoidal endothelial cells in non-alcoholic fatty

liver disease

Adel Hammoutene

1,2

, Pierre-Emmanuel Rautou

1,3,4,5,

⇑

Summary

Non-alcoholic fatty liver disease (NAFLD) and its complications are an expanding health problem asso-

ciated with the metabolic syndrome. Liver sinusoidal endothelial cells (LSECs) are highly specialized

endothelial cells localized at the interface between the blood derived from the gut and the adipose tissue

on the one side, and other liver cells on the other side. In physiological conditions, LSECs are gatekeepers

of liver homeostasis. LSECs display anti-inﬂammatory and anti-ﬁbrogenic properties by preventing

Kupffer cell and hepatic stellate cell activation and regulating intrahepatic vascular resistance and portal

pressure. This review focusses on changes occurring in LSECs in NAFLD and on their consequences on

NAFLD progression and complications. Capillarization, namely the loss of LSEC fenestrae, and LSEC dys-

function, namely the loss of the ability of LSECs to generate vasodilator agents in response to increased

shear stress both occur early in NAFLD. These LSEC changes favour steatosis development and set the

stage for NAFLD progression. At the stage of non-alcoholic steatohepatitis, altered LSECs release inﬂam-

matory mediators and contribute to the recruitment of inﬂammatory cells, thus promoting liver injury

and inﬂammation. Altered LSECs also fail to maintain hepatic stellate cell quiescence and release ﬁbro-

genic mediators, including Hedgehog signalling molecules, promoting liver ﬁbrosis. Liver angiogenesis is

increased in NAFLD and contributes to liver inﬂammation and ﬁbrosis, but also to hepatocellular carci-

noma development. Thus, improving LSEC health appears to be a promising approach to prevent NAFLD

progression and complications.

Introduction

Non-alcoholic fatty liver disease (NAFLD) encom-

passes a spectrum of conditions including simple

steatosis and non-alcoholic steatohepatitis

(NASH), deﬁned as the association of steatosis,

hepatocellular damage, inﬂammation and varying

degrees of ﬁbrosis.

NAFLD is associated with obe-

sity, insulin resistance and/or type 2 diabetes and

other metabolic abnormalities, collectively termed

the metabolic syndrome.

2,3

NAFLD is an expanding

health problem with an estimated global preva-

lence of 25%.

2,4

A recent modelling approach esti-

mated that NAFLD cases in the United States will

expand from 83 million in 2015, corresponding

to about 25% of the population, to 100 million in

2030, corresponding to more than 33% of the pop-

ulation.

While simple steatosis is generally

benign, NASH can progress to both cirrhosis and

end-stage liver disease. NASH is currently a lead-

ing cause of liver disease among adults awaiting

liver transplantation in Europe and in the United

States and is projected to become the most com-

mon indication for liver transplantation in the

next decade.

2,4

Importantly, patients with meta-

bolic syndrome and NASH also develop hepatocel-

lular carcinoma (HCC) in the absence of

cirrhosis.

5,6

Despite its prevalence and severity,

there is no approved therapy for NASH and avail-

able treatments only aim to control associated

conditions.

Understanding the mechanisms of

NAFLD, and in particular how simple steatosis pro-

gresses to NASH and then to cirrhosis and/or liver

cancer, is of the utmost importance.

The current view of the pathogenesis of NASH

centres on the response of hepatocytes to insulin

resistance and lipotoxicity. The immune system

and hepatic stellate cell activation are regarded

as secondary events.

The vascular endothelium,

representing the interface between blood and

other tissues of the body, is not only a physical

barrier but is implicated in different physiological

roles, such as haemostasis, metabolite transporta-

tion, inﬂammation, thrombosis, angiogenesis and

vascular tone. The liver endothelium is mainly

formed of liver sinusoidal endothelial cells (LSECs)

which are highly specialized endothelial cells at

the interface between blood derived from the vis-

ceral adipose tissue and the gut, on the one side,

and hepatic stellate cells and hepatocytes, on the

other side. LSECs have a unique phenotype in the

human body as they lack a basement membrane

and have a multitude of fenestrae organized into

sieves, that regulate the transport of macro-

molecules, including lipids and lipoproteins,

across the sinusoid.

7,8

This review will speciﬁcally

focus on the role of LSECs in the pathophysiology

of NAFLD and its complications.

Review JOURNAL

OF HEPATOLOGY

Keywords: Endothelium;

Steatosis; Capillarization;

Endothelial dysfunction;

Angiogenesis.

Received 5 January 2019;

received in revised form 10

2019; accepted 13 February

2019

Inserm, UMR-970, Paris Cardio-

vascular Research Center, PARCC,

Paris, France;

University Paris Descartes, Paris,

France;

INSERM, UMR1149, Centre de

Recherche sur l’Inﬂammation,

Paris, France;

University Paris Diderot, Paris,

France;

Service d’Hépatologie, Centre de

Référence des Maladies Vascu-

laires du Foie, DHU Unity, Pôle des

Maladies de l’Appareil Digestif,

Hôpital Beaujon, AP-HP, Clichy,

France

⇑

Corresponding author.

Address: Service d’hépatologie,

Hôpital Beaujon, 100 boulevard

du Général Leclerc, 92100 Cli-

chy, France. Tel.: +33 1 40 87 52

83; fax: +33 1 40 87 55 30.

E-mail address: pierre-emma-

nuel.rautou@inserm.fr

(P.-E. Rautou).

Journal of Hepatology 2019 vol. 70 j1278–1291

LSECs and simple steatosis

Role of LSECs in lipid transfer in the normal

liver

Dietary lipids present in the circulation have to be

transported through the vascular endothelium to

be metabolized by tissues. In physiological condi-

tions, LSECs are major regulators of the bidirec-

tional lipid exchange between the blood and the

liver parenchyma. First, LSEC fenestrae allow for

efﬁcient transfer of lipoproteins, chylomicron

remnants (small lipoproteins derived from chy-

lomicrons generated by enterocytes from dietary

lipids), and other macromolecules, from the sinu-

soidal blood to the space of Disse, where they

are taken up by hepatocytes.

9–11

LSEC fenestrae

form a selective barrier for lipids. Indeed, older

studies using radiolabelled lipoproteins showed

that larger lipoproteins do not cross LSEC fenestrae

and remain in the lumen of the sinusoid.

More-

over, when LSECs lose their fenestrae following

vascular endothelial growth factor (VEGF) path-

way disruption,

11,13,14

uptake of ﬂuorescently

labelled lipids is impaired.

11,15

Second, LSECs also

regulate lipid transfer through their high capacity

for endocytosis, as shown by their ability to

rapidly take-up oxidized or acetylated-low density

lipoproteins.

8,16,17

LSEC capillarization occurs early in NAFLD and

promotes steatosis

LSECs undergo morphological and functional

changes during liver steatosis.

18–21

One of the

most remarkable phenotypic changes is the loss

of fenestrae, also called defenestration or sinu-

soidal capillarization, associated with the forma-

tion of a basement membrane on the abluminal

surface of LSECs. Several independent groups

reported that sinusoidal capillarization appears

very early in NAFLD.

18–20

Miyao and colleagues

observed that defenestration begins after 1 week

of choline-deﬁcient, L-amino acid-deﬁned diet in

mice.

Similar observations were made in rats

fed a high-fat diet for 3 weeks.

Triggers for sinusoidal capillarization are not

fully identiﬁed, but we can speculate that exces-

sive dietary macronutrients, including lipids, car-

bohydrates, and gut microbiota-derived products

play a role.

Cogger and coworkers demon-

strated in mice challenged with several diets

varying in content of macronutrients and energy

that LSEC porosity and fenestrae frequency are

inversely correlated with dietary fat intake, while

fenestrae diameter is inversely correlated with

protein or carbohydrate intake.

In this study,

authors also found a negative correlation

between LSEC fenestrae (frequency, porosity and

diameter) and circulating free fatty acid (FFA)

levels.

In vitro studies suggested that defenes-

tration occurs following excessive lipid exposure.

For instance, exposure of human primary LSECs

to oxidized low-density lipoprotein (ox-LDL)

reduces the diameter and the porosity of the

fenestrae.

The effect of FFA on fenestrae has also

been tested in primary rat LSECs, but ﬁrm conclu-

sions cannot be drawn as the authors did not test

several concentrations of FFA but rather the pres-

ence vs. the absence of FFA, which does not ade-

quately mimic in vivo conditions.

Recent

evidence point to gut microbiota in the pathogen-

esis of NAFLD.

3,23

Cogger and colleagues showed

that LSEC fenestrae inversely correlated with the

abundance of Bacteroidetes in the gut and posi-

tively correlated with the abundance of Firmi-

cutes.

Moreover, it has been shown that a

single injection of endotoxin in rats induces a

decrease in both diameter and number of fenes-

trae suggesting that gut microbiota-derived prod-

ucts may contribute to LSEC capillarization,

although caution is needed since the concentra-

tion of endotoxin used in that study was high.

In turn, capillarization favours liver steatosis

(Fig. 1), as observed in mice deﬁcient in plas-

malemma vesicle-associated protein (PLVAP), an

endothelial-speciﬁc integral membrane glycopro-

tein required for the formation of endothelial fen-

estrae.

These mice exhibit a pronounced

reduction in the number of LSEC fenestrae, associ-

ated with a decrease in sinusoidal permeability,

and spontaneously develop extensive steatosis.

These mice also have hyperlipoproteinemia and

increased triglyceridemia due to the retention of

chylomicron remnants in the blood. As mentioned

above, Vegfb

-/-

mice also exhibit a reduction in the

number of LSEC fenestrae and less uptake of

labelled oleic acid due to capillarization, but

steatosis was not evaluated.

A ﬁrst hypothesis explaining this consequence

of capillarization on steatosis could be that

reduced LSEC permeability impairs the passage

of hepatocyte-derived very low-density lipopro-

tein toward the sinusoidal lumen, thus inducing

cholesterol and triglyceride retention in the liver.

However, these lipoproteins may escape the liver

through the lymphatic system.

As an alternative

explanation, Herrnberger et al. proposed that chy-

lomicron remnants originating from the blood,

and required for synthesis of very low-density

lipoprotein by hepatocytes, cannot reach hepato-

cytes due to LSEC capillarization; their absence

in hepatocytes might then stimulate de novo hep-

atic lipid synthesis and induce steatosis as a com-

pensatory mechanism.

25,27

However, there is no

available data to ascertain this hypothesis. Simi-

larly, Fraser and collaborators postulated that, fol-

lowing LSEC capillarization, chylomicron

remnants and dietary cholesterol no longer cross

the fenestrae to inhibit HMGCoA reductase, the

rate limiting enzyme for hepatocyte cholesterol

biosynthesis, consequently activating endogenous

cholesterol synthesis in hepatocytes.

Taken together, these data suggest that fea-

tures of metabolic syndrome are associated with

LSEC capillarization, which promotes steatosis

(Fig. 1).

Key point

LSEC capillarization and

dysfunction occur very

early in NAFLD progression

and contribute to dietary

induced steatosis.

JOURNAL

OF HEPATOLOGY

Journal of Hepatology 2019 vol. 70 j1278–1291 1279

LSECs dysfunction occurs early in NAFLD and

promotes steatosis

Liver steatosis is associated with an increased por-

tal pressure and increased intrahepatic vascular

resistance.

29–32

In patients, hepatic venous pres-

sure gradient correlates with the degree of steato-

sis.

Using Doppler ﬂowmetry, Seifalian and

colleagues observed impaired sinusoidal perfusion

in human fatty liver grafts compared with normal

liver grafts.

Similar results were obtained in rab-

bits and rats with diet-induced steatosis, with an

inverse correlation between hepatic parenchymal

microcirculation and the severity of steatosis.

34,35

This increased intrahepatic vascular resistance

has a mechanical and a dynamic component. The

mechanical part is due to the compression of the

sinusoidal lumen by enlarged fat-laden hepato-

cytes.

29,36–38

The dynamic part is due to liver

endothelial dysfunction. Endothelial dysfunction

is a pathological condition, common to all vascular

beds, deﬁned as the inability of blood vessels to

dilate in response to increased blood ﬂow.

Endothelial dysfunction is generally indicated by

the loss of nitric oxide bioavailability due to eNOS

(also called NOS3) inhibition.

Several lines of

evidence show that LSEC dysfunction occurs in

fatty livers and is involved in the increased intra-

hepatic vascular resistance associated with steato-

sis.

40–43

eNOS activation and liver nitric oxide

content are reduced in mice and rats fed a high-

fat diet or a diet rich in saturated fatty acids for

4 weeks.

40,41

Isolated-perfused liver experiments

performed in these animals showed augmented

portal perfusion pressure and reduced vasodila-

tory response to acetylcholine, indicating liver

endothelial dysfunction. These changes were

observed in the absence of inﬂammation and

ﬁbrosis, suggesting that endothelial dysfunction

is an early feature associated with steatosis in

NAFLD.

41,42

Several mechanisms could account for this liver

endothelial dysfunction associated with steatosis

(Fig. 2). First, LSECs dysfunction can be induced

by overabundance of lipids during steatosis. In

vitro experiments showed that stimulation of

human primary LSECs with ox-LDL downregulates

eNOS expression through the ox-LDL receptor,

LOX1.

In addition, exposure of primary LSECs

to palmitic acid also attenuates nitric oxide

bioavailability through peroxynitrite production

by NOX1, a nitric oxide consuming enzyme highly

expressed in LSECs of mice fed a high-fat diet.

Second, steatosis induces insulin resistance in

LSECs, leading to impairment of insulin-

dependent vasodilation.

41,45,46

This effect is due,

on the one hand, to the downregulation of eNOS

activity,

and on the other hand to the upregula-

tion of iNOS (also called NOS2), the inducible form

of NOS which can cause endothelial dysfunction

through increased nitro-oxidative stress.

46–48

Interestingly, V-PYRRO/NO – a diazeniumdiolate

ion metabolized in the liver that spontaneously

Chylomicron remnants

VLDL synthesis

Endogenous

cholesterol and

triglycerides

VLDL LDL

Peripheral

tissues

Fenestrae

Lumen

Space of Disse

Physiological conditions

LSEC

Chylomicron

remnants

VLDL synthesis

↑ Endogenous

cholesterol and

triglycerides

synthesis

VLDL Lymphatic vessels

Metabolic syndrome

ox-LDL/FFA

Carbohydrates/proteins

Gut-microbiota products

Steatosis

Capillarization

Circulating

chylomicron

remnants

Hyperlipidemia

Lumen

Space of Disse Peripheral

circulation

Capillarization

LSEC

Hepatocyte

Exogenous

cholesterol and

triglycerides

VLDL

export

↓ Exogenous

cholesterol and

triglycerides

Fig. 1. LSEC capillarization promotes steatosis. In physiological conditions, chylomicron remnants cross LSEC fenestrae and

provide cholesterol and triglycerides for VLDL synthesis. VLDL are then released by hepatocytes and reach blood ﬂow through

fenestrae.In metabolic syndrome conditions, LSEC capillarization arises early in the course of NAFLD, possibly because of

exposure of LSECs to dietary macronutrients. In turn, LSEC capillarization promotes steatosis, possibly because capillarization

blocks the transfer of chylomicron remnants to hepatocytes, thus stimulating endogenous cholesterol and triglyceride

synthesis, as a compensatory mechanism for the synthesis of VLDL, which reach blood ﬂow through the lymphatic system. FFA,

free fatty acids; LDL, low-density lipoprotein; LSECs, liver sinusoidal endothelial cells; NAFLD, non-alcoholic fatty liver disease;

ox-LDL, oxidized low-density lipoprotein; VLDL; very low-density lipoprotein.

Key point

Intrahepatic vascular

resistance is increased

even when steatosis is the

only histological feature of

NAFLD. This is due to the

combination of a compres-

sion of sinusoids by fat-

laden enlarged hepatocytes

and of a dysfunction of

LSECs due to reduced nitric

oxide bioavailability.

Review

1280 Journal of Hepatology 2019 vol. 70 j1278–1291

decomposes to nitric oxide with a very short half-

life at physiological pH and that triggers cyclic

guanosine 30,50-monophosphate (cGMP) synthesis

– improves hepatic microcirculation in mice with

steatosis induced by a high-fat diet.

Third, the

gut microbiota also seems to contribute to liver

endothelial dysfunction. Indeed, Garcia-Lezana

and colleagues demonstrated that restoration of

a healthy microbiota via faecal transplantation

normalizes portal hypertension by improving

intrahepatic vascular resistance and endothelial

dysfunction in rats.

In turn, LSEC dysfunction favours steatosis

(Fig. 2). Indeed, deﬁciency in nitric oxide in

eNos

/

mice results in massive fat droplet deposi-

tion and increases liver triglyceride content.

40,50

Nitric oxide contributes to the regulation of hep-

atic lipid content by limiting citrate synthesis in

mitochondria, which is involved in fatty acid pro-

duction.

Nitric oxide also attenuates synthesis of

fatty acids in isolated cultured rat hepatocytes by

nitrosylating acetyl-CoA

and activating AMP-

activated protein kinase,

52–54

an inhibitor of

glycerol-3-phosphate acyltransferase and thus of

triacylglycerol synthesis.

55,56

In addition, nitric

oxide also allows efﬁcient fatty acid beta-

oxidation through s-nitrosylation of very long-

chain acyl-CoA dehydrogenase in hepatocytes.

Interestingly, therapies augmenting nitric oxide

availability in the liver ameliorate steatosis. The

V-PYRRO/NO or the improvement of nitric oxide/

cGMP signalling with the phosphodiesterase-5

inhibitor sildenaﬁl protect against liver steatosis

in mice fed a high-fat diet.

40,58,59

Moreover, treat-

ment with simvastatin, a drug able to increase

expression and activity of eNOS expression in the

liver, decreases steatosis induced by a high-fat diet

in rats.

To summarize, steatosis is associated with LSEC

dysfunction which in turn worsens steatosis

(Fig. 2).

Angiogenesis and steatosis

Angiogenesis, deﬁned as the formation of new

vessels from pre-existing vessels, is a key event

in the progression of NAFLD.

61–65

VEGF is the mas-

ter pro-angiogenic regulator of this process sup-

ported by activation of hypoxia inducible factors

(HIFs) in hypoxic areas.

Serum VEGF levels are higher in patients with

biopsy-proven steatosis than in healthy individu-

als.

62,63,65

In animal models, liver expression of

VEGF and CD105, an endothelial cell marker,

increase after 3 days of methionine- and choline-

deﬁcient diet in obese and diabetic db/db trans-

genic mice and after 1 week of this diet in

C57BL6/J mice, before NASH appears.

However,

3 studies reported that new vessels develop in

the livers of patients with NASH, but not in

individuals with simple steatosis or normal

livers.

66–68

This suggests that molecular events

associated with upregulation of angiogenic factors

start early in the course of NAFLD, while angiogen-

esis appears later, as detailed below.

LSECs in NASH

LSECs contribute to oxidative stress in NASH

In response to lipotoxicity, hepatocytes generate

reactive oxygen species (ROS) and initiate a robust

inﬂammatory response that accentuates liver

injury.

1,3

In addition to hepatocytes,

1,3

the lipo-

toxic response also occurs in LSECs contributing

to ROS generation.

44,69

Indeed, ROS have been

detected not only in hepatocytes but also in sinu-

soidal cells in patients with NASH.

Circulating

lipids seem to account for the oxidative stress in

LSECs. Indeed, exposure of murine LSECs to palmi-

tic acid upregulates NOX1 expression, an enzyme

implicated in ROS production.

In addition, stim-

ulation of human primary LSECs with ox-LDL

increases ROS generation after binding to

LOX1.

This oxidative stress in LSECs contributes

to NASH. Indeed, mice with a global deﬁciency in

NOX1, which is highly expressed in LSECs in

NAFLD, had attenuated liver lesions when fed a

high-fat diet, as shown by lower serum ALT level

and lower hepatic cleaved caspase-3 expression.

Therefore, in NASH, ROS production takes place

not only in hepatocytes, but also to some extent

in LSECs, and seems to contribute to hepatocyte

injury.

Anti-inﬂammatory role of LSECs at initial stages

of NASH

Progression of simple steatosis to steatohepatitis is

accompanied by adhesion of leukocytes to the

sinusoidal endothelium followed by inﬁltration of

leukocytes within liver parenchyma to form

inﬂammatory foci.

Moderate and resolved

inﬂammatory responses are beneﬁcial to the liver

as they promote the re-establishment of home-

ostasis, contribute to tissue repair and exert hep-

atoprotective effects.

However, chronic

inﬂammation, as seen in NASH, leads to death of

hepatocytes and causes damage to the liver

parenchyma.

In physiological conditions, LSECs

constitute a barrier regulating the entry of circulat-

ing leukocytes within liver parenchyma and play-

ing an anti-inﬂammatory role.

73–76

At early

stages of NAFLD progression, some evidence indi-

cates that LSECs also exhibit anti-inﬂammatory

functions.

40,77

Indeed, Tateya and colleagues ele-

gantly demonstrated that nitric oxide derived from

LSECs inhibits Kupffer cell activation in mice fed a

high-fat diet for a short period of time (8 weeks).

In vitro, both human and murine LSECs

exposed to FFA for a short period (16 hours) exhibit

a downregulation of pro-inﬂammatory chemoki-

nes involved in monocyte and macrophage

recruitment, through a MAPK dependent

pathway.

Key point

Molecular events associ-

ated with angiogenesis are

initiated during simple

steatosis, but angiogenesis

itself is only detected in

NASH.

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OF HEPATOLOGY

Journal of Hepatology 2019 vol. 70 j1278–1291 1281

LSECs promotes liver inﬂammation at more

advanced stages of NASH

As mentioned, LSEC alterations arise early in

NAFLD progression, prior to liver inﬂamma-

tion.

18,41

Indeed, LSEC capillarization precedes

Kupffer cell activation

18,41

and liver nitric oxide

content falls before liver NF-kB activation and

TNF

, IL-6 and ICAM-1 upregulation.

30,40,41

LSEC

capillarization and dysfunction are permissive

for establishment of liver inﬂammation. Indeed,

mice deﬁcient in eNOS exhibit an accelerated

hepatic inﬂammatory response, while improving

nitric oxide/cGMP signalling with the

phosphodiesterase-5 inhibitor sildenaﬁl or with

simvastatin prevents liver inﬂammation in

rodents fed a high-fat diet.

40,60

During NALFD progression, LSECs then acquire

a pro-inﬂammatory phenotype and functions

(Fig. 3). LSECs pro-inﬂammatory phenotype during

NASH is characterized by progressive overexpres-

sion of adhesion molecules including ICAM-1,

VCAM-1 and VAP-1 (AOC3) at the surface of LSECs,

as observed in mouse models of NASH.

68,78–82

LSECs also produce a number of pro-

inﬂammatory mediators in NASH, including TNF

IL-6, IL-1 and MCP1 (CCL2).

68,82–84

This pro-inﬂammatory phenotype of LSECs in

NASH is associated with pro-inﬂammatory func-

tions (Fig. 3). First, dysfunctional LSECs fail to

maintain Kupffer cell quiescence.

Second, the

release of inﬂammatory mediators by LSECs con-

tributes to the inﬂammatory response by activat-

ing neighbouring Kupffer cells, and by favouring

recruitment, adhesion and transmigration of blood

leukocytes.

82,85,86

The mechanisms of interaction

between leukocytes and LSECs in NASH have been

reviewed elsewhere in detail and are summarized

in Fig. 3.

80,87–89

LSECs’ expression of ICAM-1,

VCAM-1 and VAP-1 is crucial for these interactions

since in vivo and in vitro studies showed reduced

leukocyte adhesion to hepatic sinusoids when

these receptors are blocked or not functional.

80,90

Moreover, inhibition of the VCAM-1 ligand, VLA-

4 (or ITGA4), on monocytes using an anti-VLA-4

antibody inhibits adhesion and transendothelial

migration of monocytes across LSECs – from

wild-type mice fed a high-fat diet and from ob/ob

obese mice – and improves liver inﬂammation.

Steatosis

↓NO

Acetyl CoA

FenestraeLumen

Space of Disse

Physiological conditions

eNOS

Citrate

Fatty acids

Triglycerides

GPAT

Citrate

AMPK

Acetyl CoA

Metabolic syndrome

↓NO

Fatty acids

↑Triglycerides

Citrate

↓eNOS LSEC

dysfunction

↑iNOS

↑NOX1

ox-LDL, FFA

Insulin resistance ↑ IHVR

Peroxynit rites

Lumen

Space of Disse

Intrahepatic

vascular tone

LSEC LSEC

Hepatocyte

Fatty acids

β-oxidation

ACC

AMPK

ACC

LSEC

dysfunction

Hepatocyte

enlargement

Nitrosylation

Citrate

Fatty acids

β-oxi dation

GPAT

↓NO

Nitrosylation

Fig. 2. LSEC dysfunction promotes steatosis. In physiological conditions, LSECs release NO which regulates intrahepatic

vascular tone on the one hand, and hepatic lipid metabolism on the other hand. NO limits hepatic lipid content by inhibiting

hepatic de novo lipogenesis, through a limitation of citrate synthesis in mitochondria, an inhibition of ACC and of GPAT, and by

promoting fatty acids beta-oxidation. In metabolic syndrome conditions, overabundance of lipids and insulin resistance lead to

downregulation of eNOS activity and to upregulation of iNOS and of NOX1 (a nitric oxide consuming enzyme), causing nitro-

oxidative stress through peroxynitrite production and eventually endothelial dysfunction. Reduced NO availability promotes

steatosis. Liver steatosis is associated with an increased intrahepatic vascular resistance which has a mechanical component,

due to the compression of the sinusoidal lumen by enlarged fat-laden hepatocytes, and a dynamic component, due to a liver

endothelial dysfunction. ACC, acetyl CoA carboxylase; AMPK, AMP-activated protein kinase; eNOS, endothelial nitric oxide

synthase; GPAT, glycerol-3-phosphate acyltransferase; LSECs, liver sinusoidal endothelial cells; IHRV; intrahepatic vascular

resistance; iNOS, inducible nitric oxide synthase; NO, nitric oxide; NOX1, NADPH oxidase 1.

Key point

Lipotoxicity and inﬂam-

mation induce endothelial

inﬂammation. Activated

LSECs release cytokines

and chemokines and over-

express adhesion mole-

cules, thus sustaining liver

inﬂammation.

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1282 Journal of Hepatology 2019 vol. 70 j1278–1291

Although the stimuli responsible for LSECs’

inﬂammatory phenotype and functions in NASH

are not ﬁrmly identiﬁed, several mediators are

potential candidates. This includes products

derived from the visceral adipose tissue, such as

ox-LDL, FFA and adipokines. Indeed, in vitro

studies showed that stimulation of LSECs with

ox-LDL and FFA (palmitate) activate NF-kB and

TLR-4, respectively.

21,44,91

Moreover, circulating

concentrations of several adipokines, including

TNF

and IL-6, are increased in the portal vein in

the context of metabolic syndrome, and may

contribute to LSECs inﬂammatory phenotype.

The gut microbiota also has an emerging role in

NASH as a source of inﬂammatory stimuli.

1,3

Increased intestinal permeability and elevated

plasma concentrations of lipopolysaccharide

(LPS)

93,94

observed in NASH may also contribute

to LSECs’ pro-inﬂammatory function.

Besides

mediators derived from the portal vein, hepato-

cytes and liver inﬂammatory cells also release

inﬂammatory mediators in NASH that can activate

LSECs.

1,95

To summarize, while LSECs play an anti-

inﬂammatory role in the initial stages of NAFLD,

a switch towards pro-inﬂammatory functions

occurs during the course of NAFLD development,

paving the way for NASH progression.

Angiogenesis in liver inﬂammation in NASH

Pathologic angiogenesis increases with

NASH

61,68,96,97

(Fig. 4). Indeed, several studies

reported the formation of new vessels in the liver

of patients with NASH. Moreover, serum VEGF and

sVEGFR1 levels are higher in patients with steato-

sis and biopsy-proven NASH than in healthy indi-

viduals.

62,66–68,98

Similarly, in animal models of

NASH, liver vasculature is disrupted and hepatic

expression of VEGF and CD105 is increased.

Several mechanisms trigger angiogenesis dur-

ing NASH. First, chronic inﬂammation promotes

angiogenesis. Indeed, chronic inﬂammation sus-

tains tissue hypoxia and induces transcription of

NF-κB

Lipotoxicity

Inflammation

Gut microbiota-

derived products

Activated Kupffer cell

Quiescent

Kupffer cell

Transendothelial

migration

Lymphocytes,

Monocytes, Neutrophils,

NK/NKT cells

Leukocytes

Chronic

inflammation

Space of Disse

Sinusoidal lumen

MCP1, TNFα,

IL-6, IL-1

↓ NO

Endothelial dysfunction

VCAM-1, ICAM- 1, VAP-1

LSEC

Fig. 3. LSECs acquire pro-inﬂammatory functions in NASH. Lipotoxicity, inﬂammation and

gut microbiota-derived products induce LSECs’ inﬂammatory phenotype and function

mediated by NF-kB activation, which orchestrates the release of pro-inﬂammatory mediators

and the overexpression of adhesion molecules. Inﬂammatory mediators and LSEC dysfunction

promote Kupffer cell activation and leukocyte chemoattraction. Adhesion molecule overex-

pression allows adhesion and transendothelial migration of the recruited leukocytes in the

hepatic parenchyma. ICAM-1, intercellular adhesion molecule-1; IL-1, interleukin 1; IL-6,

interleukin 6; LSECs, liver sinusoidal endothelial cells; MCP1, monocyte chemoattractant

protein-1; NASH, non-alcoholic steatohepatitis; NF-kB, nuclear factor kappa B; NK, natural

killer; NO, nitric oxide; TNF

, tumor necrosis factor alpha; VAP-1, vascular adhesion protein-

1; VCAM-1, vascular cell adhesion molecule-1.

Steatosis Inflammation Fibrosis

↑Angiogenesis

Anti-angiogenic

therapies

Angiopoeitin-2

VEGF

Leptin

Hepatocytes injury

Inflammation

Fibrosis

Hypoxia

Anti-VEGFR2

Blocking

Angiopoietin2-Tie

interaction

(L1-10 peptibody)

Angiotensin-II

receptor

blockers

Hepatocytes- derived

microvesicles

Fig. 4. Angiogenesis in NASH. Hypoxia, liver injury, lipids, oxidative stress, inﬂammation and ﬁbrosis induce the release of

pro-angiogenic factors, such as hepatocyte-derived microvesicles, VEGF and Angiopoietin-2, from parenchymal and non-

parenchymal cells including LSECs, promoting pathologic angiogenesis. Adipokines, such as leptin, also exhibit pro-angiogenic

activity contributing to pathologic angiogenesis in NASH. In turn, angiogenesis promotes liver inﬂammation and ﬁbrosis as

shown by anti-angiogenic therapies which prevent liver inﬂammation and ﬁbrosis in experimental models of NASH. LSECs,

liver sinusoidal endothelial cells; NASH, non-alcoholic steatohepatitis; VEGF, vascular endothelial growth factor; VEGFR2,

vascular endothelial growth factor receptor 2.

Key point

Inﬂammation promotes

angiogenesis that in turn

worsens liver inﬂamma-

tion as demonstrated by

the anti-inﬂammatory

effect of anti-angiogenic

therapies.

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Journal of Hepatology 2019 vol. 70 j1278–1291 1283

angiogenic genes modulated by HIF-1

61,99

Pro-

inﬂammatory mediators also elicit a direct angio-

genic response through the induction of HIF-1

transcriptional activity and VEGF production.

Moreover, cytokines and ROS released during

NASH can activate the MAPK/ERK pathway, a sig-

nalling pathway involved in cell migration and

angiogenesis.

Second, hepatocyte-derived

microvesicles link lipotoxicity with angiogenesis.

Indeed, hepatocytes exposed in vitro to excessive

amounts of saturated FFA, that mimics steatosis,

release microvesicles with a pro-angiogenic activ-

ity.

100

Likewise, mice fed a methionine- and

choline-deﬁcient diet have high circulating levels

of hepatocyte-derived microvesicles able to

induce angiogenesis. Third, angiopoietin-2 is

another mechanism of liver angiogenesis in NASH.

Angiopoietins are key regulators of angiogenesis.

Although angiopoietins-1 and 2 contribute to vas-

cular stability and quiescence in physiological

conditions, angiopoietin-2 promotes pathological

angiogenesis in inﬂammatory conditions.

101

Lefere

and coworkers recently showed that serum

angiopoietin-2 levels are increased in patients

with NASH and correlate with liver steatosis,

inﬂammation and hepatocyte ballooning, but not

with liver ﬁbrosis.

Similar ﬁndings were

observed with 2 murine models of NASH, namely

mice fed a methionine- and choline-deﬁcient diet

and mice with neonatal injection of streptozotocin

followed by 16 weeks of western diet.

The main

source of hepatic angiopoietin-2 was LSECs.

Inhibiting angiopoietin-2 levels using the

angiopoietin-2/Tie2 receptor inhibiting peptibody

L1-10 reduced hepatic angiogenesis and normal-

ized vascular microarchitecture.

In turn, angiogenesis promotes inﬂammation

since various strategies of inhibition of angiogenesis

all improve liver inﬂammation (Fig. 4). Coulon and

colleagues showed in a mouse model of NASH that

treatment with anti-VEGFR2 antibody improves

liver vasculature and decreases liver inﬂammatory

gene expression, both using preventive and thera-

peutic approaches.

Lefere and colleagues showed

that blocking angiopoietin-2/Tie2 interaction with

the L1-10 peptibody also alleviates liver injury and

inﬂammation in mice fed a methionine- and

choline-deﬁcient diet.

Importantly, this effect of

L1-10 therapy is at least partly mediatedby an effect

on LSECs since L1-10 treatment downregulates

VCAM-1, ICAM-1 and MCP1 expression in liver

endothelial cells isolated from mice fed a

methionine- and choline-deﬁcient diet.

This

anti-inﬂammatory effect of anti-angiogenic treat-

ment is not speciﬁc for NASH, as it is observed in

most models of chronic liver disease, namely carbon

tetrachloride, and bile duct ligation.

102–106

To summarize, inﬂammation stimulates angio-

genesis that in turn worsens inﬂammation, as

shown by the anti-inﬂammatory effect of anti-

angiogenic therapies (Fig. 4).

LSECs in NASH-related liver ﬁbrosis

Liver ﬁbrosis is deﬁned as the excessive deposi-

tion of extracellular matrix in liver parenchyma.

The main mechanism leading to liver ﬁbrosis is a

long-standing wound healing process caused by

hepatocellular injury and inﬂammation and

mediated by hepatic stellate cell activation.

1,107

Hepatic stellate cells are nonparenchymal cells

close to LSECs, in the space of Disse, which store

retinoids in physiological conditions and shift

their phenotype to an activated myoﬁbroblastic

state during liver injury and inﬂammation,

wherein they secrete large amounts of extracel-

lular matrix compounds.

107

As detailed above,

LSECs are major effectors of liver inﬂammation

in NASH, and consequently also promote hepatic

ﬁbrosis. For example, LSECs overexpress VAP-1

during inﬂammation which, in addition to its

pro-inﬂammatory functions in NASH, is directly

involved in hepatic stellate cell activation.

Inhi-

bition or deﬁciency in VAP-1 in mice fed a

methionine- and choline-deﬁcient diet or a

high-fat diet attenuates liver ﬁbrosis.

LSECs

also contribute to liver ﬁbrosis through capillar-

ization and endothelial dysfunction, as detailed

in the following sections.

LSEC capillarization promotes liver ﬁbrosis

Capillarization is observed in patients and animal

models of NASH, preceding ﬁbrosis,

18,41,108–112

but also promoting its development (Box 1).

Indeed, PLVAP deﬁcient mice, displaying a pro-

nounced reduction in the number of LSEC fenes-

trae, spontaneously develop perisinusoidal liver

ﬁbrosis.

Experiments performed using cultured LSECs

and hepatic stellate cells highlighted the impor-

tance of cross-talk between these cells types in

regulating each other’s phenotype. While healthy

LSECs maintain hepatic stellate cell quiescence,

capillarized LSECs lose this ability

112,113

(Fig. 5).

A vicious cycle between LSEC capillarization and

hepatic stellate cell activation then occurs during

the ﬁbrotic process.

In NASH, ballooned hepatocytes produce

Hedgehog molecules.

114

The Hedgehog pathway

regulates capillarization, as inhibition of Hedge-

hog signalling prevents capillarization and par-

tially reverts the phenotype of LSECs from a

dedifferentiated state to their differentiated

LSECs alterations are observed in various dietary or genetic models of NAFLD, without

apparent specificity. Indeed, LSECs capillarization is observed in animals fed a choline-

deficient, L-amino acid-defined diet or a high-fat diet.18−20 LSECs dysfunction can be induced

by a high-fat diet, or a rich saturated fatty acids diet, or a high-fat high-glucose and

high-fructose diet.40,41,43 Markers of LSECs inflammatory phenotype are observed in animals

fed a high-fat diet or a methionine and choline deficient diet.68,82 Markers of angiogenesis

appear after methionine and choline deficient diet in db/db obese mice and in C57BL/6 mice

and in rats fed a choline-deficient, L-amino acid-defined diet.63,129

Box 1. LSEC changes are not speciﬁc for certain animal models of NASH. LSEC, liver

sinusoidal endothelial cells; NASH, non-alcoholic steatohepatitis.

Key point

LSEC capillarization and

dysfunction precede liver

ﬁbrosis and are permissive

for it, through the loss of

the ability of LSECs to

maintain quiescence of

hepatic stellate cells.

Review

1284 Journal of Hepatology 2019 vol. 70 j1278–1291

state.

115

LSECs are thus Hedgehog-sensitive cells,

but they are also Hedgehog producing cells. Simi-

larly, quiescent hepatic stellate cells are

Hedgehog-sensitive cells, while activated hepatic

stellate cells become Hedgehog-producing cells,

which are also able to release macrovesicles

loaded with Hedgehog signalling molecules that

interact with LSECs.

116,117

It is thus tempting to

speculate that during NASH, Hedgehog ligands

are released by hepatocytes and LSECs, thus

activating LSECs themselves, as well as quiescent

hepatic stellate cells, by autocrine and paracrine

effects. Activated hepatic stellate cells can then

secrete Hedgehog molecules, promoting LSEC

capillarization which in turn favours hepatic

stellate cell activation, promoting the ﬁbrogenic

process.

LSECs dysfunction promotes liver ﬁbrosis

Endothelial dysfunction appears very early in the

course of NAFLD and precedes ﬁbrosis in animal

models of NASH.

30,40,41

Several lines of evidence

suggest that liver endothelial dysfunction

contributes to liver ﬁbrosis. First, in rats fed a

high-fat diet, simvastatin increases liver eNOS

expression and ameliorates liver ﬁbrosis.

Second, eNOS inhibition using L-NAME blocks

the ability of healthy LSECs to keep hepatic stellate

cells quiescent.

118,119

Third, an activator of soluble

guanylate cyclase, a receptor for nitric oxide, can

recapitulate the effect of healthy LSECs and

reverse hepatic stellate cell activation.

However,

a limitation of these studies is that most

approaches might not only act on endothelial

function but also on LSEC capillarization.

112,118

Indeed, although L-NAME blocks LSEC-induced

hepatic stellate cell quiescence, addition of a nitric

oxide donor does not directly reverse hepatic stel-

late cell activation in vitro,

suggesting that addi-

tional LSEC-derived factors could be responsible

for the reversion of activated hepatic stellate cells

to quiescence.

To summarize, these data demonstrate that

capillarization and LSEC dysfunction not only pre-

cede liver ﬁbrosis, but also promote it (Fig. 5). In

their differentiated state, LSECs are able to main-

tain hepatic stellate cell quiescence, making differ-

entiated LSECs gatekeepers of ﬁbrosis in NASH, as

in other chronic liver diseases.

Liver endothelial-to-mesenchymal transition: a

process promoting liver ﬁbrosis?

Another important process that links endothelial

cells to organ ﬁbrosis is endothelial-to-

mesenchymal transition, i.e. the mechanism by

which endothelial cells convert into myoﬁbroblasts

and contribute to extracellular matrix deposi-

tion.

120,121

Endothelial-to-mesenchymal transition

occurs in various ﬁbrotic cardiovascular and pul-

monary diseases.

120–122

In vitro studies showed

that healthy LSECs produce a modest amount of col-

lagen and ﬁbronectin.

123

Capillarized LSECs secrete

ﬁbrogenic factors, such as TGF-b1, and extracellular

matrix proteins, such as ﬁbronectin and laminin,

that may be considered as an endothelial-to-

mesenchymal transition, as well as stimulating

activation of neighbouring hepatic stellate

cells.

8,124,125

In the liver disease ﬁeld, only 1 study

has described endothelial-to-mesenchymal transi-

tion in vivo, in patients with alcohol- or hepatitis C

virus-related cirrhosis and in mice treated with car-

bon tetrachloride.

126

This process might also occur

during liver ﬁbrosis in NASH, but further studies are

required.

Altered

LSEC

Fibronectine

Laminine

TGF-β

Hh molecules

Endothelial dysfunction

Capillarization

↑ Hh signaling

Hh molecules

Activated

HSC

Quiescent

HSC

ECM secretion

Space of Disse

Lumen

Hh molecules

↓NO

↑ Fibrosis

Fig. 5. LSECs in NASH-related ﬁbrosis. Healthy LSECs are gatekeepers of liver ﬁbrosis by

maintaining HSCs quiescence through NO, while altered LSECs (following capillarization and

LSEC dysfunction) lose this ability. In addition, altered LSECs release proﬁbrogenic molecules

such as TGF-b, Hedgehog molecules, laminin and ﬁbronectin, which activate HSCs. Activated

HSCs produces Hedgehog molecules reinforcing their own activation and LSEC capillarization.

Activated HSCs then produce large amounts of extracellular matrix thus inducing liver ﬁbrosis.

ECM, extracellular matrix; Hh, Hedgehog; HSCs, hepatic stellate cells; LSECs, liver sinusoidal

endothelial cell; NO, nitric oxide; TGF-b, transforming growth factor-b.

Stabilin

1/2

LYVE-1 CD32b

(SE-1)

ICAM-1Integrins

αvβ3

αvβ5

Higher

expression of

Integrins

αvβ3, αvβ5

Lower

expression of

ICAM-1

Lower adherence

capacity with

leukocytes

Higher adherence capacity with

cancerous cells

T-cells tolerance towards

cancer-associated

antigens

Immunosuppressive environment

Liver endothelial cell

Tumour progression

Fig. 6. Role of liver endothelial cells in HCC development in chronic liver diseases (not

speciﬁc for NAFLD). During HCC progression, endothelial cells sequentially lose their speciﬁc

markers including stabilin-1, stabilin-2, LYVE-1 and CD32b (SE-1). Conversely, endothelial

expression of integrins increases, facilitating adhesion of liver cancer cells. In parallel,

endothelial expression of ICAM-1 decreases, leading to a lower ability of leukocyte to adhere

and inﬁltrate HCC. Endothelial cells within HCC can also alter tumour-associated immune

responses via their ability to confer T cell tolerance towards cancer-associated antigens and

to create an immunosuppressive environment. ICAM-1, intercellular adhesion molecule-1;

LYVE-1, lymphatic vessel endothelial hyaluronic acid receptor 1; HCC, hepatocellular

carcinoma; NAFLD, non-alcoholic fatty liver disease.

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Angiogenesis in NASH-related liver ﬁbrosis

Liver angiogenesis correlates with liver ﬁbrosis in

patients with NASH.

66,67

The mechanisms leading

to liver angiogenesis in NASH-related ﬁbrosis

include those mentioned above, namely tissue

hypoxia,

hepatocyte-derived microvesi-

cles

100,127

and angiopoietin-2,

but also leptin.

Leptin concentrations are increased in the serum

of patients with NAFLD.

128

This adipocytokine

has both pro-angiogenic effects,

129

and direct

pro-ﬁbrogenic effects, through the upregulation

of TGF-bin LSECs and Kupffer cells.

130

In turn, angiogenesis promotes liver ﬁbrosis

since several approaches inhibiting liver angiogen-

esis prevent NASH-related ﬁbrosis (Fig. 4). First, in

the study by Kitade and colleagues, neither angio-

genesis nor ﬁbrosis were observed in the absence

of leptin signalling in a rat model of NASH.

129

Sec-

ond, blocking the release of pro-angiogenic

microvesicles from fat laden-hepatocytes or

inhibiting their binding to target cells protects

mice from steatohepatitis-induced pathologic

angiogenesis and results in a reduction in liver

ﬁbrosis.

100

Third, Zhou and coworkers recently

showed that a speciﬁc deletion of the physiologi-

cal inhibitor of angiogenesis, prolyl-hydroxylase-

2, in endothelial cells results in an overexpression

of angiopoietin-2 and TGF-b1 in the liver and pro-

motes dietary-induced liver ﬁbrosis in mice.

131,132

Whether this pro-ﬁbrotic effect of endothelial

prolyl-hydroxylase-2 deﬁciency is directly

induced by promoting angiogenesis remains to

be demonstrated. Fourth, 2 studies reported that

inhibiting angiotensin-II receptor using telmisar-

tan or candesartan inhibits liver angiogenesis

and ﬁbrosis in rats fed a choline-deﬁcient, L-

amino acid-deﬁned diet.

96,97

Finally, Lefere and

colleagues demonstrated that blocking angiogene-

sis by inhibiting the interaction between

angiopoietin-2/Tie2 using the L1-10 peptibody

improves liver ﬁbrosis in preventive and therapeu-

tic strategies in mice fed a methionine- and

choline-deﬁcient diet. Therapeutic application of

L1-10 peptibody also prevents liver ﬁbrosis in dia-

betic mice with NASH (streptozotocin and western

diet model).

It should be noted that an anti-

ﬁbrotic effect of anti-angiogenic treatments has

also been observed in models of chronic liver dis-

ease without NASH.

102–106,133–139

LSECs in NASH-induced HCC

In most cases, HCC develops on a background of

chronic liver disease (70–90% of all patients). The

role of liver endothelial cells in HCC development,

outside the NAFLD setting, has been reviewed

elsewhere and is summarized in Fig. 6.

8,76,140–143

Patients with metabolic syndrome and NAFLD

also develop HCC in the absence of underlying cir-

rhosis, suggesting oncogenic pathways speciﬁc for

NAFLD.

5,6

Adipokines and angiogenesis associated

with NAFLD seem to account – at least partly – for

this speciﬁc link between NAFLD and HCC.

Circulating concentrations of the adipokine

FABP4 are increased in patients with NAFLD with-

out HCC and correlate with liver inﬂammation

and ﬁbrosis.

144

Interestingly, Laouirem and col-

leagues recently demonstrated that LSECs exposed

to conditions mimicking NAFLD – namely high con-

centrations of glucose, insulin, or VEGFA – release

FABP4. They also observed that FABP4 released by

LSECs exerts pro-oncogenic effects, since it induces

hepatocyte proliferation. In mice fed a high-fat diet,

speciﬁc inhibition of FABP4 reduces HCC growth.

145

We can speculate that FABP4 from LSECs not only

contributes to HCC progression but also to HCC

development in a NAFLD setting (Fig. 7).

In NAFLD, angiogenesis is highly stimulated

and promotes NAFLD-associated HCC, since vari-

ous inhibitors of angiogenesis all prevent HCC

development. First, leptin-mediated angiogenesis

has been demonstrated to be involved in HCC

development as neither angiogenesis nor HCC

develop in the absence of leptin signalling in

Zucker rats fed a choline-deﬁcient, L-amino acid-

deﬁned diet.

129

Second, Yoshiji and colleagues

showed that a conventional anti-angiogenic treat-

ment with sorafenib inhibits the appearance of

preneoplastic lesions in rats fed a choline-

deﬁcient, L-amino acid-deﬁned diet.

146

In this

study, authors also demonstrated that a treatment

combining low doses of sorafenib with the

angiotensin-II receptor inhibitor losartan also suc-

cessfully inhibited preneoplastic lesions.

146

Third,

Tamaki and colleagues demonstrated that inhibi-

tion of angiotensin-II receptor with telmisartan

inhibits HIF-1

activity and VEGF expression and

prevents HCC development in the liver of rats

fed a choline-deﬁcient, L-amino acid-deﬁned diet

for 48 weeks.

Finally, Lefere and coworkers

recently showed that therapeutic inhibition of

angiopoietin-2 alleviates steatohepatitis and pre-

vents NASH-associated HCC progression in mice.

Adipokines

↑ FABP4

↑ Leptin

FABP4

↑ VEGF

↑ Angiopoeitin-2 ↑Angiogenesis

Anti-angiogenic

therapies

Angiotensin-II receptor blockers

(Losartan,Telmisartan)

Blocking Angiopoietin2-Tie

interaction (L1-10 peptibody)

Sorafenib

HCC development

and progression

Metabolic syndrome

NAFLD

Fig. 7. Role of liver endothelial cells in hepatocellular carcinoma development in the

NAFLD setting. Circulating concentrations of angiocrine factors, such as VEGF and

angiopoeitin-2, and adipokines, such as leptin are increased in NAFLD. These mediators

induce angiogenesis which promotes HCC growth. The adipokine FABP4 is released by adipose

tissue and endothelial cells and contributes to HCC development and progression. FABP4, fatty

acid binding protein 4; HCC, hepatocellular carcinoma; NAFLD, non-alcoholic fatty liver

disease.

Key point

Adipokines contribute to

HCC development in

NAFLD.

Key point

Liver angiogenesis corre-

lates with the severity of

liver ﬁbrosis and promotes

its development. Once

established, ﬁbrosis stimu-

lates angiogenesis by

increasing tissue hypoxia.

Blocking pathologic angio-

genesis prevents liver

ﬁbrosis.

Key point

NALFD associated angio-

genesis promotes HCC.

Blocking pathologic angio-

genesis prevents HCC

development and

progression.

Review

1286 Journal of Hepatology 2019 vol. 70 j1278–1291

Gaps in knowledge and future directions

Even if our understanding of the role of LSECs in

NAFLD has progressed over the last years, several

aspects remain elusive. First, triggers responsible

for LSEC alterations in NAFLD are mostly unknown.

It has been suggested that mediators derived from

the visceral adipose tissue and the gut are respon-

sible, but this has not been convincingly estab-

lished. Indeed, available in vitro studies

considered each mediator individually and not in

combination, as in vivo in the portal venous

blood.

21,44,77

Second, mechanisms underlying

endothelial changes in NAFLD, including capillar-

ization, need to be deﬁned which might provide

new therapeutic targets for NAFLD. Third, the role

of LSECs in NASH-related cirrhosis has not been

speciﬁcally investigated. Whether LSEC function

and phenotype differ in cirrhosis related to NASH

from cirrhosis related to other causes remains to

be assessed.

147

Fourth, although NAFLD is well rec-

ognized as favouring HCC development, we are still

at a very early stage of understanding how LSEC

changes might favour HCC development.

Conclusion

LSECs are gatekeepers of liver homeostasis in phys-

iological conditions. In NAFLD, sinusoidal endothe-

lial alterations, including capillarization and LSEC

dysfunction, occur early in disease progression, at

the stage of simple steatosis. These initial changes,

triggered by lipotoxicity, adipokines, inﬂammation

and gut microbiota-derived products are associ-

ated with a loss of the ability of LSECs to prevent

liver inﬂammation and ﬁbrosis associated with

NASH. Indeed, altered LSECs fail to maintain Kupf-

fer cells and hepatic stellate cells in a quiescent

state. At the stage of NASH, altered LSECs con-

tribute to liver angiogenesis, inﬂammation, ﬁbrosis

and HCC. Improving LSEC health has great thera-

peutic potential for NAFLD. The current challenge

is the identiﬁcation of strategies to speciﬁcally tar-

get LSECs in order to modulate their activity.

Financial support

This work was supported by the ‘‘Institut National

de la Santé et de la Recherche Médicale” (ATIP

AVENIR), Paris Descartes University, the ‘‘Agence

Nationale pour la Recherche” (ANR-14-CE12-

0011, ANR-14-CE35-0022, ANR-18-CE14-0006-

01) and by the ‘‘Association Française pour l’Etude

du foie” (AFEF 2014). A.H. was supported by the

‘‘Ministère de l’Enseignement Supérieur et de la

Recherche”.

Conﬂict of interest

The authors declare no conﬂicts of interest that

pertain to this work.

Please refer to the accompanying ICMJE disclo-

sure forms for further details.

Authors’ contributions

A.H. and P-E.R wrote the manuscript.

Supplementary data

Supplementary data to this article can be found

online at https://doi.org/10.1016/j.jhep.2019.02.

012.

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Semaphorin-3A regulates liver sinusoidal endothelial cell porosity and promotes hepatic steatosis

Article

Full-text available

Jun 2024

Prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD), formerly known as non-alcoholic fatty liver disease, increases worldwide and associates with type 2 diabetes and other cardiometabolic diseases. Here we demonstrate that Sema3a is elevated in liver sinusoidal endothelial cells of animal models for obesity, type 2 diabetes and MASLD. In primary human liver sinusoidal endothelial cells, saturated fatty acids induce expression of SEMA3A , and loss of a single allele is sufficient to reduce hepatic fat content in diet-induced obese mice. We show that semaphorin-3A regulates the number of fenestrae through a signaling cascade that involves neuropilin-1 and phosphorylation of cofilin-1 by LIM domain kinase 1. Finally, inducible vascular deletion of Sema3a in adult diet-induced obese mice reduces hepatic fat content and elevates very low-density lipoprotein secretion. Thus, we identified a molecular pathway linking hyperlipidemia to microvascular defenestration and early development of MASLD.

Gut Microbiota and Sinusoidal Vasoregulation in MASLD: A Portal Perspective

Article

Full-text available

Jun 2024

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a common condition with heterogeneous outcomes difficult to predict at the individual level. Feared complications of advanced MASLD are linked to clinically significant portal hypertension and are initiated by functional and mechanical changes in the unique sinusoidal capillary network of the liver. Early sinusoidal vasoregulatory changes in MASLD lead to increased intrahepatic vascular resistance and represent the beginning of portal hypertension. In addition, the composition and function of gut microbiota in MASLD are distinctly different from the healthy state, and multiple lines of evidence demonstrate the association of dysbiosis with these vasoregulatory changes. The gut microbiota is involved in the biotransformation of nutrients, production of de novo metabolites, release of microbial structural components, and impairment of the intestinal barrier with impact on innate immune responses, metabolism, inflammation, fibrosis, and vasoregulation in the liver and beyond. The gut–liver axis is a conceptual framework in which portal circulation is the primary connection between gut microbiota and the liver. Accordingly, biochemical and hemodynamic attributes of portal circulation may hold the key to better understanding and predicting disease progression in MASLD. However, many specific details remain hidden due to limited access to the portal circulation, indicating a major unmet need for the development of innovative diagnostic tools to analyze portal metabolites and explore their effect on health and disease. We also need to safely and reliably monitor portal hemodynamics with the goal of providing preventive and curative interventions in all stages of MASLD. Here, we review recent advances that link portal metabolomics to altered sinusoidal vasoregulation and may allow for new insights into the development of portal hypertension in MASLD.

Role of post‐hepatectomy portal hypertension in predicting the outcome of hepatocellular carcinoma after curative resection

Article

Full-text available

Jun 2024

Aims Portal venous pressure frequently increases after a major hepatectomy. We aimed to assess the outcomes of patients with hepatocellular carcinoma (HCC) with different post‐hepatectomy portal pressure statuses and the predictive factors correlated with prognosis and post‐hepatectomy portal hypertension (PHPH). Methods Data from consecutive patients who underwent curative‐intent hepatectomy for HCC at the Eastern Hepatobiliary Surgery Hospital and Mengchao Hepatobiliary Hospital of Fujian Medical University between January 2008 and December 2016 were retrospectively analyzed. Patients were followed up until March 1, 2022. PHPH was defined as new‐onset portal hypertension (PH) in patients without preoperative PH within 1 month of hepatectomy. Patients were classified into two groups according to the presence or absence of PHPH: non‐PH and PHPH. The clinicopathological characteristics were compared between the two groups. Univariate and multivariate analyses were used to identify independent risk factors for early and late recurrence of HCC, overall survival (OS), and PHPH. Results Disease‐free survival rates were significantly higher in the non‐PH group ( n = 1068) than those in the PHPH group ( n = 423) (62.0%, 39.0%, and 31.0% vs. 46.2%, 24.5%, and 19.3% at 1, 3, and 5 years, respectively; all p < 0.001). The 1‐, 3‐, and 5‐year OS rates were 91.0%, 66.4%, and 51.4% in the non‐PH group and 80.0%, 48.9%, and 32.6% in the PHPH group, respectively (all p < 0.001). Multivariate analysis revealed that PHPH was independently associated with early recurrence of HCC and poor OS (hazard ratio [HR] 1.476, 95% confidence interval [CI] 1.279–1.704, p < 0.001 and HR 1.601, 95% CI 1.383–1.853, p < 0.001, respectively). Furthermore, hepatitis B virus infection, cirrhosis, preoperative alanine aminotransferase > 40 U/L, and major hepatectomy were identified as independent risk factors for PHPH. Conclusions PHPH was associated with early recurrence of HCC and poor OS but not with late recurrence. Therefore, PHPH may be an attractive target for therapeutic interventions and follow‐up surveillance to improve prognosis.

A Systematic Review of Statins for the Treatment of Nonalcoholic Steatohepatitis: Safety, Efficacy, and Mechanism of Action

Article

Full-text available

Apr 2024
MOLECULES

Nonalcoholic fatty liver disease (NAFLD) is the liver component of a cluster of conditions, while its subtype, nonalcoholic steatohepatitis (NASH), emerges as a potentially progressive liver disorder that harbors the risk of evolving into cirrhosis and culminating in hepatocellular carcinoma (HCC). NASH and cardiovascular disease (CVD) have common risk factors, but compared to liver-related causes, the most common cause of death in NASH patients is CVD. Within the pharmacological armamentarium, statins, celebrated for their lipid-modulating prowess, have now garnered attention for their expansive therapeutic potential in NASH. Evidence from a plethora of studies suggests that statins not only manifest anti-inflammatory and antifibrotic properties but also impart a multifaceted beneficial impact on hepatic health. In this review, we used “statin”, “NAFLD”, “NASH”, and “CVD” as the major keywords and conducted a literature search using the PubMed and Web of Science databases to determine the safety and efficacy of statins in patients and animals with NASH and NAFLD, and the mechanism of statin therapy for NASH. Simultaneously, we reviewed the important role of the intestinal microbiota in statin therapy for NASH, as it is hoped that statins will provide new insights into modulating the harmful inflammatory microbiota in the gut and reducing systemic inflammation in NASH patients.

A novel efficient strategy to generate liver sinusoidal endothelial cells from human pluripotent stem cells

Article

Full-text available

Jun 2024

Liver sinusoidal endothelial cells (LSECs) are highly specialized endothelial cells (ECs) that play an important role in liver development and regeneration. Additionally, it is involved in various pathological processes, including steatosis, inflammation, fibrosis and hepatocellular carcinoma. However, the rapid dedifferentiation of LSECs after culture greatly limits their use in vitro modeling for biomedical applications. In this study, we developed a highly efficient protocol to induce LSEC-like cells from human induced pluripotent stem cells (hiPSCs) in only 8 days. Using single-cell transcriptomic analysis, we identified several novel LSEC-specific markers, such as EPAS1, LIFR, and NID1, as well as several previously revealed markers, such as CLEC4M, CLEC1B, CRHBP and FCN3. These LSEC markers are specifically expressed in our LSEC-like cells. Furthermore, hiPSC-derived cells expressed LSEC-specific proteins and exhibited LSEC-related functions, such as the uptake of acetylated low density lipoprotein (ac-LDL) and immune complex endocytosis. Overall, this study confirmed that our novel protocol allowed hiPSCs to rapidly acquire an LSEC-like phenotype and function in vitro. The ability to generate LSECs efficiently and rapidly may help to more precisely mimic liver development and disease progression in a liver-specific multicellular microenvironment, offering new insights into the development of novel therapeutic strategies.

Loss of fenestrae in liver sinusoidal endothelial cells contributes to MASLD

Article

Jun 2024

The Role of Liver Sinusoidal Endothelial Cells in NAFLD Disease

Article

Jan 2024

玥勤黄

Angiocrine signaling in sinusoidal homeostasis and liver diseases

Article

May 2024
J HEPATOL

The liver sinusoid in chronic liver disease: NAFLD and NASH

Chapter

Jan 2024

Comprehensive study of the interplay between immunological and metabolic factors in hepatic steatosis

Article

Apr 2024

Endothelial fatty liver binding protein 4: a new targetable mediator in hepatocellular carcinoma related to metabolic syndrome

Article

Full-text available

Apr 2019

Metabolic syndrome (MS) is becoming the leading risk factor for hepatocellular carcinoma (HCC). HCC development related to MS may occur in advanced or non-advanced liver fibrosis, suggesting specific molecular pathways. Among these pathways, basal inflammatory state and adipokines production are involved. The aim of this study was to evaluate the role of fatty acid-binding protein 4 (FABP4). In this study, we demonstrate the specific overexpression of FABP4 in human HCC samples from patients with MS compared to other risk factors for chronic liver disease with FABP4 expression restricted to peritumoral endothelial cells. In vitro, glucose, insulin, VEGFA and hypoxia upregulated endothelial FABP4, which was reversed by metformin through mTOR pathway inhibition. FABP4 exerts oncogenic effects on hepatoma cell lines by upregulating the angiogenesis gene signature and pathways involved in the cell cycle, leading to increased cell proliferation and migration, and downregulating HIF1 pathway; effects were reversed in the presence of a specific FABP4 inhibitor (BMS309403). We showed the role of microvesicles as FABP4 vectors between endothelial and tumor cells. In vivo, BMS309403 significantly reduces tumor growth in heterotopic and orthotopic xenografted mice model. In conclusion, this study demonstrates the emerging oncogenic role of liver endothelial cells through FABP4 in HCC related to MS, and highlights new anti-neoplastic mechanism of metformin.

The gut microbiota in infants of obese mothers increases inflammation and susceptibility to NAFLD

Article

Full-text available

Oct 2018

Maternal obesity is associated with increased risk for offspring obesity and non-alcoholic fatty liver disease (NAFLD), but the causal drivers of this association are unclear. Early colonization of the infant gut by microbes plays a critical role in establishing immunity and metabolic function. Here, we compare germ-free mice colonized with stool microbes (MB) from 2-week-old infants born to obese (Inf-ObMB) or normal-weight (Inf-NWMB) mothers. Inf-ObMB-colonized mice demonstrate increased hepatic gene expression for endoplasmic reticulum stress and innate immunity together with histological signs of periportal inflammation, a histological pattern more commonly reported in pediatric cases of NAFLD. Inf-ObMB mice show increased intestinal permeability, reduced macrophage phagocytosis, and dampened cytokine production suggestive of impaired macrophage function. Furthermore, exposure to a Western-style diet in Inf-ObMB mice promotes excess weight gain and accelerates NAFLD. Overall, these results provide functional evidence supporting a causative role of maternal obesity-associated infant dysbiosis in childhood obesity and NAFLD.

Capillarization of Hepatic Sinusoids in Man

Article

Mar 1963
GASTROENTEROLOGY

Progressive changes in the hepatic perisinusoidal tissue space of Disse in chronic liver diseases ultimately lead to development of a basement membrane and capillarization of sinusoids. The transformation of an open circulation to a closed one decreases the effectiveness of hepatic circulation and aggravates hepatocellular insufficiency.

Hepatic microcirculation and mechanisms of portal hypertension

Article

Dec 2018
NAT REV GASTRO HEPAT

The liver microcirculatory milieu, mainly composed of liver sinusoidal endothelial cells (LSECs), hepatic stellate cells (HSCs) and hepatic macrophages, has an essential role in liver homeostasis, including in preserving hepatocyte function, regulating the vascular tone and controlling inflammation. Liver microcirculatory dysfunction is one of the key mechanisms that promotes the progression of chronic liver disease (also termed cirrhosis) and the development of its major clinical complication, portal hypertension. In the present Review, we describe the current knowledge of liver microcirculatory dysfunction in cirrhotic portal hypertension and appraise the preclinical models used to study the liver circulation. We also provide a comprehensive summary of the promising therapeutic options to target the liver microvasculature in cirrhosis.

The role of macrophages in nonalcoholic fatty liver disease and nonalcoholic steatohepatitis

Article

Nov 2018
NAT REV GASTRO HEPAT

Nonalcoholic fatty liver disease (NAFLD) and its inflammatory and often progressive subtype nonalcoholic steatohepatitis (NASH) are becoming the leading cause of liver-related morbidity and mortality worldwide, and a primary indication for liver transplantation. The pathophysiology of NASH is multifactorial and not yet completely understood; however, innate immunity is a major contributing factor in which liver-resident macrophages (Kupffer cells) and recruited macrophages play a central part in disease progression. In this Review, we assess the evidence for macrophage involvement in the development of steatosis, inflammation and fibrosis in NASH. In this process, not only the polarization of liver macrophages towards a pro-inflammatory phenotype is important, but adipose tissue macrophages, especially in the visceral compartment, also contribute to disease severity and insulin resistance. Macrophage activation is mediated by factors such as endotoxins and translocated bacteria owing to increased intestinal permeability, factors released from damaged or lipoapoptotic hepatocytes, as well as alterations in gut microbiota and defined nutritional components, including certain free fatty acids, cholesterol and their metabolites. Reflecting the important role of macrophages in NASH, we also review studies investigating drugs that target macrophage recruitment to the liver, macrophage polarization and their inflammatory effects as potential treatment options for patients with NASH.

Non-Alcoholic Fatty Liver Disease—A Global Public Health Perspective

Article

Nov 2018
J HEPATOL

Zobair M. Younossi

As the epidemics of obesity and type 2 diabetes mellitus increase worldwide, the prevalence of non-alcoholic fatty liver disease (NAFLD) is increasing proportionately. The subtype of NAFLD which can be characterised as non-alcoholic steatohepatitis (NASH) is a potentially progressive liver disease that can lead to cirrhosis, hepatocellular carcinoma, liver transplantation, and death. NAFLD is also associated with extrahepatic manifestations such as chronic kidney disease, cardiovascular disease and sleep apnoea. NAFLD and NASH carry a large economic burden and create poor health-related quality of life. Despite this important burden, we are only beginning to understand its mechanisms of pathogenesis and the contribution of environmental and genetic factors to the risk of developing a progressive course of disease. Research is underway to identify appropriate non-invasive diagnostic methods and effective treatments. Although the risk of liver-related mortality is increased in patients with NAFLD and liver fibrosis stages F3 or F4, the leading cause of death is cardiovascular disease. Given the rapidly growing global burden of NAFLD and NASH, efforts must continue to find accurate non-invasive diagnostic and prognostic biomarkers, to develop effective treatments for individuals with advanced NASH and prevention methods for individuals at high risk of NAFLD and progressive liver disease.

Angiopoietin‐2 promotes pathological angiogenesis and is a novel therapeutic target in murine non‐alcoholic fatty liver disease

Article

Sep 2018

Angiogenesis contributes to the development of non-alcoholic steatohepatitis (NASH) and promotes inflammation, fibrosis and progression to hepatocellular carcinoma (HCC). Angiopoietin-2 (Ang-2) is a key regulator of angiogenesis. We aimed to investigate the role of Ang-2 and its potential as therapeutic target in NASH using human samples, in vivo mouse models and in vitro assays. Serum Ang-2 levels were determined in 104 obese patients undergoing bariatric surgery and concomitant liver biopsy. The effect of the Ang-2/Tie2 receptor inhibiting peptibody L1-10 was evaluated in the methionine-choline deficient (MCD) and streptozotocin-western diet NAFLD mouse models, and in vitro on endothelial cells and bone marrow-derived macrophages. The hepatic vasculature was visualized with μCT scans and scanning electron microscopy of vascular casts. Serum Ang-2 levels were increased in patients with histological NASH compared to patients with simple steatosis and correlated with hepatic CD34 immunoreactivity as a marker of hepatic angiogenesis. Serum and hepatic Ang-2 levels were similarly increased in mice with steatohepatitis. Both preventive and therapeutic L1-10 treatment reduced hepatocyte ballooning and fibrosis in MCD diet-fed mice, and was associated with reduced hepatic angiogenesis and normalization of the vascular micro-architecture. Liver-isolated endothelial cells and monocytes from MCD-fed L1-10-treated mice showed reduced expression of leukocyte adhesion and inflammatory markers, respectively, compared to cells from untreated MCD diet-fed mice. In the streptozotocin-western diet model, therapeutic Ang-2 inhibition was able to reverse NASH and attenuate HCC progression. In vitro, L1-10 treatment mitigated the increased cytokine production in LPS-stimulated endothelial cells, but not macrophages. Conclusion: Our findings provide evidence for Ang-2 inhibition as a new therapeutic strategy to target pathological angiogenesis in NASH. This article is protected by copyright. All rights reserved.

Global Perspectives on Non-alcoholic Fatty Liver Disease and Non-alcoholic Steatohepatitis

Article

Sep 2018

Over the past 2 decades, nonalcoholic fatty liver disease (NAFLD) has grown from a relatively unknown disease to the most common cause of CLD in the world. In fact, 25% of the world's population is currently thought to have NAFLD. Non‐alcoholic steatohepatitis (NASH) is the subtype of NAFLD that can progress to cirrhosis, hepatocellular carcinoma, and death. NAFLD and NASH are found in not only adults—there is a high prevalence in children and adolescents. Due to NAFLD's close association with type 2 diabetes (T2DM) and obesity, the latest models predict the prevalence of NAFLD and NASH will increase, causing a tremendous clinical and economic burden and poor patient‐reported outcomes. Nonetheless, there is no accurate non‐invasive method to detect NASH and treatment is limited to life style modifications. To examine the state of NAFLD among different regions and understand the global trajectory of this disease, an international group of experts came together during 2017 AASLD Global NAFLD Forum. We provide a summary of this forum and an assessment of the current state of NAFLD and NASH worldwide. This article is protected by copyright. All rights reserved.

Tracking Fenestrae Dynamics in Live Murine Liver Sinusoidal Endothelial Cells

Article

Jan 2019

The fenestrae of liver sinusoidal endothelial cells (LSEC) allow passive transport of solutes, macromolecules and particulate material between the sinusoidal lumen and the liver parenchymal cells. Until recently, fenestrae and fenestrae-associated structures were mainly investigated using electron microscopy on chemically fixed LSECs. Hence, the knowledge about their dynamic properties remained until to-date largely elusive. Recent progress in atomic force microscopy (AFM) has allowed the study of live cells in three-dimensions (X, Y & Z) over a prolonged time (t) and this at unprecedented speeds and resolving power. Hence, we employed those latest advances in AFM imaging on living LSECs. As a result, we were able to monitor as first the position, size and number of fenestrae and sieve plates using 4-D AFM (X, Y, Z & t) on intact LSEC in vitro. During those time-lapse experimentations, dynamic data were collected on the origin and morpho-functional properties of the filtration apparatus of LSECs. We present herein structural evidence on single laying and grouped fenestrae thereby elucidating their dynamic nature from formation to disappearance. We also collected data on the lifespan of fenestrae. More especially, the formation and closing of entire sieve plates were observed and recorded how the continuous rearrangement of sieve plates affect the structure of fenestrae within them. We observed also the dawn and rise of fenestrae-forming centres and defenestration centres in LSECs under different experimental conditions. Conclusion: No other multimodal biomedical high-resolution imaging technique to-date has allowed collecting new fine structural information on the lifespan, formation, and disappearance of LSEC fenestrae. By doing so, we gathered also evidence of three different pathways implemented in the loss of fenestrae that result in defenestrated LSECs. This article is protected by copyright. All rights reserved.

Risk of Hepatocellular Cancer in Patients With Non-Alcoholic Fatty Liver Disease

Article

Aug 2018
GASTROENTEROLOGY

Background: There are limited data on the risk of hepatocellular cancer (HCC) in patients with non-alcoholic fatty liver disease (NAFLD). We aimed to estimate the risk of incident HCC among patients with NAFLD. Methods: We conducted a retrospective cohort study from a total of 130 facilities in the Veterans Health Administration. Patients with NAFLD diagnosed between 1/1/2004 and 12/31/2008 were included and followed until HCC diagnosis, death or 12/31/2015. We also identified a gender and age-matched control cohort without NAFLD. We ascertained all new HCC cases from the Central Cancer Registry and manual chart reviews. We calculated incidence rates for HCC by NAFLD status as well as in subgroups of NAFLD patients. We used competing risk models to compare the risk of HCC in patients with vs. those without NAFLD. We reviewed electronic medical records of all HCC cases that developed in NAFLD patients without cirrhosis. Results: We compared 296,707 NAFLD patients with 296,707 matched controls. During 2,382,289 person-years [PY] of follow-up, 490 NAFLD patients developed HCC (0.21/1000 PY). HCC incidence was significantly higher among NAFLD patients vs. controls (0.02/1000 PY; hazard ratio, 7.62, 95% confidence interval=5.76-10.09). Among patients with NAFLD, those with cirrhosis had the highest annual incidence of HCC (10.6 /1000 PY). Among patients with NAFLD cirrhosis, HCC risk ranged from 1.6 to 23.7 per 1000 PY based on other demographic characteristics; the risk of HCC was the highest in older Hispanics with cirrhosis. In medical record reviews, 20% of NAFLD patients with HCC had no evidence of cirrhosis. Conclusions: Risk of HCC was higher in NAFLD patients than that observed in general clinical population. Most HCC cases in NAFLD developed in patients with cirrhosis. The absolute risk of HCC was higher than the accepted thresholds for HCC surveillance for most patients with NAFLD cirrhosis.

Role of liver sinusoidal endothelial cells in nonalcoholic fatty liver disease

Abstract

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