Cirrhosis-downregulated LSECtin can be retrieved by cytokines, shifts the TLR-induced LSECs secretome and correlates with the hepatic Th response.

BACKGROUND AND AIMS: We evaluated tolerogenic C-type lectin LSECtin loss in cirrhosis and its potential regulation by cytokines. METHODS: Liver tissue from patients with cirrhosis and healthy controls, immortalised and generated LSECtin-CRISPR immortalised LSECs, and murine primary LSECs from the CCl4 model were handled. RESULTS: LSECtin expression was reduced in liver tissue from cirrhotic patients, and it decreased from compensated to decompensated disease. Increased phosphorylation of MAPK, Akt and NFkB was observed upon LSECtin stimulation in LSEC murine cell line, showing a pattern of inflammatory and chemotactic cytokines either restrained (IL-10, CCL4) or unrestrained (TNF-α, IL-1ß, IL-6, CCL2). CD44 attenuated whereas LAG-3 increased all substrates phosphorylation in combination with TLR4 and TLR2 ligands except for NFkB. TNF-α, IL-1 ß, IL-6 and CCL2 were restrained by LSECtin crosslinking on TLRs studied. Conversely, IL-10 and CCL4 were upregulated, suggesting a LSECtin-TLRs synergistic effect. Also, LSECtin was significantly induced after IL-13 stimulation or combined with anti-inflammatory cytokines in cirrhotic and immortalised LSECs. Th17 and regulatory T cells were progressively increased in the hepatic tissue from compensated to decompensated patients. A significant inverse correlation was present between gene expression levels of CLEC4G/LSECtin and RORγT and FOXP3 in liver tissues. CONCLUSION: LSECtin restrains TLR proinflammatory secretome induced on LSECs by interfering immune response control, survival and MAPKs signalling pathways. The cytokine-dependent induction of LSECtin and the association between LSECtin loss and Th17 cell subset expansion in the liver, provides a solid background for exploring LSECtin retrieval as a mechanism to reprogram LSEC homeostatic function hampered during cirrhosis.

Neuroblastoma RAS viral oncogene homolog (N-RAS) deficiency aggravates liver injury and fibrosis.

Progressive hepatic damage and fibrosis are major features of chronic liver diseases of different etiology, yet the underlying molecular mechanisms remain to be fully defined. N-RAS, a member of the RAS family of small guanine nucleotide-binding proteins also encompassing the highly homologous H-RAS and K-RAS isoforms, was previously reported to modulate cell death and renal fibrosis; however, its role in liver damage and fibrogenesis remains unknown. Here, we approached this question by using N-RAS deficient (N-RAS-/-) mice and two experimental models of liver injury and fibrosis, namely carbon tetrachloride (CCl4) intoxication and bile duct ligation (BDL). In wild-type (N-RAS+/+) mice both hepatotoxic procedures augmented N-RAS expression in the liver. Compared to N-RAS+/+ counterparts, N-RAS-/- mice subjected to either CCl4 or BDL showed exacerbated liver injury and fibrosis, which was associated with enhanced hepatic stellate cell (HSC) activation and leukocyte infiltration in the damaged liver. At the molecular level, after CCl4 or BDL, N-RAS-/- livers exhibited augmented expression of necroptotic death markers along with JNK1/2 hyperactivation. In line with this, N-RAS ablation in a human hepatocytic cell line resulted in enhanced activation of JNK and necroptosis mediators in response to cell death stimuli. Of note, loss of hepatic N-RAS expression was characteristic of chronic liver disease patients with fibrosis. Collectively, our study unveils a novel role for N-RAS as a negative controller of the progression of liver injury and fibrogenesis, by critically downregulating signaling pathways leading to hepatocyte necroptosis. Furthermore, it suggests that N-RAS may be of potential clinical value as prognostic biomarker of progressive fibrotic liver damage, or as a novel therapeutic target for the treatment of chronic liver disease.

The Hepatic Sinusoid in Chronic Liver Disease: The Optimal Milieu for Cancer.

The liver sinusoids are a unique type of microvascular beds. The specialized phenotype of sinusoidal cells is essential for their communication, and for the function of all hepatic cell types, including hepatocytes. Liver sinusoidal endothelial cells (LSECs) conform the inner layer of the sinusoids, which is permeable due to the fenestrae across the cytoplasm; hepatic stellate cells (HSCs) surround LSECs, regulate the vascular tone, and synthetize the extracellular matrix, and Kupffer cells (KCs) are the liver-resident macrophages. Upon injury, the harmonic equilibrium in sinusoidal communication is disrupted, leading to phenotypic alterations that may affect the function of the whole liver if the damage persists. Understanding how the specialized sinusoidal cells work in coordination with each other in healthy livers and chronic liver disease is of the utmost importance for the discovery of new therapeutic targets and the design of novel pharmacological strategies. In this manuscript, we summarize the current knowledge on the role of sinusoidal cells and their communication both in health and chronic liver diseases, and their potential pharmacologic modulation. Finally, we discuss how alterations occurring during chronic injury may contribute to the development of hepatocellular carcinoma, which is usually developed in the background of chronic liver disease.

Transcriptomic Profiling of the Liver Sinusoidal Endothelium during Cirrhosis Reveals Stage-Specific Secretory Signature.

The poor prognosis of chronic liver disease (CLD) generates the need to investigate the evolving mechanisms of disease progression, thus disclosing therapeutic targets before development of clinical complications. Considering the central role of liver sinusoidal endothelial cells (LSECs) in pre-neoplastic advanced CLD, the present study aimed at investigating the progression of CLD from an endothelial holistic perspective. RNAseq defined the transcriptome of primary LSECs isolated from three pre-clinical models of advanced CLD, during the progression of the disease, and from fresh human cirrhotic tissue. At each stage of the disease, the effects of LSECs secretome on neighboring cells and proteomic analysis of LSECs-derived extracellular vesicles (EVs) were also determined. CLD was associated with deep common modifications in the transcriptome of LSECs in the pre-clinical models. Pathway enrichment analysis showed predominance of genes related with pro-oncogenic, cellular communication processes, and EVs biogenesis during CLD progression. Crosstalk experiments revealed endothelial EVs as potent angiocrine effectors. The proteome of LSECs EVs showed stage-specific signatures, including over-expression of tropomyosin-1. Proof-of-principle experiments treating cirrhotic HSCs with recombinant tropomyosin-1 suggested de-activating effects. Our data provide the basis for discovering novel biomarkers and therapeutic targets for new disease-modifying treatments for patients with advanced CLD.

Role of liver sinusoidal endothelial cells in liver diseases.

Liver sinusoidal endothelial cells (LSECs) form the wall of the hepatic sinusoids. Unlike other capillaries, they lack an organized basement membrane and have cytoplasm that is penetrated by open fenestrae, making the hepatic microvascular endothelium discontinuous. LSECs have essential roles in the maintenance of hepatic homeostasis, including regulation of the vascular tone, inflammation and thrombosis, and they are essential for control of the hepatic immune response. On a background of acute or chronic liver injury, LSECs modify their phenotype and negatively affect neighbouring cells and liver disease pathophysiology. This Review describes the main functions and phenotypic dysregulations of LSECs in liver diseases, specifically in the context of acute injury (ischaemia-reperfusion injury, drug-induced liver injury and bacterial and viral infection), chronic liver disease (metabolism-associated liver disease, alcoholic steatohepatitis and chronic hepatotoxic injury) and hepatocellular carcinoma, and provides a comprehensive update of the role of LSECs as therapeutic targets for liver disease. Finally, we discuss the open questions in the field of LSEC pathobiology and future avenues of research.

Cell Death in Hepatocellular Carcinoma: Pathogenesis and Therapeutic Opportunities.

Hepatocellular carcinoma (HCC) is the most prevalent primary liver cancer and the third leading cause of cancer death worldwide. Closely associated with liver inflammation and fibrosis, hepatocyte cell death is a common trigger for acute and chronic liver disease arising from different etiologies, including viral hepatitis, alcohol abuse, and fatty liver. In this review, we discuss the contribution of different types of cell death, including apoptosis, necroptosis, pyroptosis, or autophagy, to the progression of liver disease and the development of HCC. Interestingly, inflammasomes have recently emerged as pivotal innate sensors with a highly pathogenic role in various liver diseases. In this regard, an increased inflammatory response would act as a key element promoting a pro-oncogenic microenvironment that may result not only in tumor growth, but also in the formation of a premetastatic niche. Importantly, nonparenchymal hepatic cells, such as liver sinusoidal endothelial cells, hepatic stellate cells, and hepatic macrophages, play an important role in establishing the tumor microenvironment, stimulating tumorigenesis by paracrine communication through cytokines and/or angiocrine factors. Finally, we update the potential therapeutic options to inhibit tumorigenesis, and we propose different mechanisms to consider in the tumor microenvironment field for HCC resolution.

Pan-PPAR agonist lanifibranor improves portal hypertension and hepatic fibrosis in experimental advanced chronic liver disease.

BACKGROUND & AIMS: In advanced chronic liver disease (ACLD), deregulated hepatic necroinflammatory processes play a key role in the development of liver microvascular dysfunction, fibrogenesis, and increased hepatic vascular tone, resulting in progression of ACLD and portal hypertension. Given the current lack of an effective treatment, we aimed to characterise the effects of the pan-peroxisome proliferator-activated receptor (pan-PPAR) agonist lanifibranor in 2 preclinical models of ACLD, as well as in liver cells from patients with ACLD. METHODS: Cirrhotic rats (thioacetamide or common bile duct ligation; TAA or cBDL) randomly received lanifibranor (100 mg/kg/day, po) or vehicle for 14 days (n = 12/group). PPAR expression, systemic and hepatic haemodynamics, presence of ascites, liver sinusoidal endothelial cell (LSEC) phenotype, hepatic stellate cell (HSC) activation, serum transaminases and albumin, hepatic macrophage infiltration, cytokine expression, and liver fibrosis were determined. Hepatic cells were isolated from the livers of patients with cirrhosis and their phenotype was evaluated after treatment with either lanifibranor or vehicle. RESULTS: TAA-cirrhotic rats receiving lanifibranor showed significantly lower portal pressure compared with vehicle-treated animals (-15%; p = 0.003) without decreasing portal blood flow, indicating improved hepatic vascular resistance. Moreover, lanifibranor-treated TAA-rats showed decreased ascites, improved LSEC and HSC phenotypes, ameliorated hepatic microvascular function, reduced hepatic inflammation, and significant fibrosis regression (-32%; p = 0.020). These findings were confirmed in the cBDL rat model as well as in human liver cells from patients with cirrhosis, which exhibited phenotypic improvement upon treatment with lanifibranor. CONCLUSIONS: Lanifibranor ameliorates fibrosis and portal hypertension in preclinical models of decompensated cirrhosis. Promising results in human hepatic cells further support its clinical evaluation for the treatment of ACLD. LAY SUMMARY: Advanced chronic liver disease (ACLD) constitutes a serious public health issue for which safe and effective treatments are lacking. This study shows that lanifibranor improves portal hypertension and liver fibrosis, 2 key elements of the pathophysiology of ACLD, in preclinical models of the disease. Evaluation of lanifibranor in liver cells from patients with ACLD further supports its beneficial effects.

Human amniotic stem cells improve hepatic microvascular dysfunction and portal hypertension in cirrhotic rats.

BACKGROUND AND AIMS: Portal hypertension is the main consequence of cirrhosis, responsible for the complications defining clinical decompensation. The only cure for decompensated cirrhosis is liver transplantation, but it is a limited resource and opens the possibility of regenerative therapy. We investigated the potential of primary human amniotic membrane-derived mesenchymal stromal (hAMSCs) and epithelial (hAECs) stem cells for the treatment of portal hypertension and decompensated cirrhosis. METHODS: In vitro: Primary liver sinusoidal endothelial cells (LSECs) and hepatic stellate cells (HSCs) from cirrhotic rats (chronic CCl4 inhalation) were co-cultured with hAMSCs, hAECs or vehicle for 24 hours, and their RNA profile was analysed. In vivo: CCl4-cirrhotic rats received 4x106 hAMSCs, 4x106 hAECs, or vehicle (NaCl 0.9%) (intraperitoneal). At 2-weeks we analysed: a) portal pressure (PP) and hepatic microvascular function; b) LSECs and HSCs phenotype; c) hepatic fibrosis and inflammation. RESULTS: In vitro experiments revealed sinusoidal cell phenotype amelioration when co-cultured with stem cells. Cirrhotic rats receiving stem cells, particularly hAMSCs, had significantly lower PP than vehicle-treated animals, together with improved liver microcirculatory function. This hemodynamic amelioration was associated with improvement in LSECs capillarization and HSCs de-activation, though hepatic collagen was not reduced. Rats that received amnion derived stem cells had markedly reduced hepatic inflammation and oxidative stress. Finally, liver function tests significantly improved in rats receiving hAMSCs. CONCLUSIONS: This preclinical study shows that infusion of human amniotic stem cells effectively decreases PP by ameliorating liver microcirculation, suggesting that it may represent a new treatment option for advanced cirrhosis with portal hypertension.

Rilpivirine attenuates liver fibrosis through selective STAT1-mediated apoptosis in hepatic stellate cells.

OBJECTIVE: Liver fibrosis constitutes a major health problem worldwide due to its rapidly increasing prevalence and the lack of specific and effective treatments. Growing evidence suggests that signalling through cytokine-activated Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathways regulates liver fibrosis and regeneration. Rilpivirine (RPV) is a widely used anti-HIV drug not reported to produce hepatotoxicity. We aimed to describe the potential hepatoprotective effects of RPV in different models of chronic liver injury, focusing on JAK-STAT signalling regulation. DESIGN: The effects of RPV on hepatic steatosis, inflammation and fibrogenesis were studied in a nutritional mouse model of non-alcoholic fatty liver disease, carbon tetrachloride-induced fibrosis and bile duct ligation-induced fibrosis. Primary human hepatic stellate cells (hHSC) and human cell lines LX-2 and Hep3B were used to investigate the underlying molecular mechanisms. RESULTS: RPV exerted a clear anti-inflammatory and antifibrotic effect in all the in vivo models of liver injury employed, and enhanced STAT3-dependent proliferation in hepatocytes and apoptosis in HSC through selective STAT1 activation. These results were reproduced in vitro; RPV undermined STAT3 activation and triggered STAT1-mediated pathways and apoptosis in HSC. Interestingly, this selective pro-apoptotic effect completely disappeared when STAT1 was silenced. Conditioned medium experiments showed that HSC apoptosis activated STAT3 in hepatocytes in an interleukin-6-dependent mechanism. CONCLUSION: RPV ameliorates liver fibrosis through selective STAT1-dependent induction of apoptosis in HSC, which exert paracrinal effects in hepatocytes, thus promoting liver regeneration. RPV's actions may represent an effective strategy to treat chronic liver diseases of different aetiologies and help identify novel therapeutic targets.

Ischemia/Reperfusion Injury in the Aged Liver: The Importance of the Sinusoidal Endothelium in Developing Therapeutic Strategies for the Elderly.

The liver endothelium plays a key role in the progression and resolution of liver diseases in young and adult individuals. However, its role in older people remains unknown. We have herein evaluated the importance of the sinusoidal endothelium in the pathophysiology of acute liver injury, and investigated the applicability of simvastatin, in aged animals. Eighteen-months-old male Wistar rats underwent 60 minutes of partial warm ischemia followed by 2 hours of reperfusion (WIR). A group of aged rats received simvastatin for 3 days before WIR. Endothelial phenotype, parenchymal injury, oxidative and nitrosative stress, and fenestrae dynamics were analyzed. The effects of WIR and simvastatin were investigated in primary LSEC from aged animals. The results of this study demonstrated that WIR significantly damages the liver endothelium and its effects are markedly worse in old animals. WIR-aged livers exhibited reduced vasodilation and sinusoidal capillarization, associated with liver damage and cellular stress. Simvastatin prevented the detrimental effects of WIR in aged livers. In conclusion, the liver sinusoidal endothelium of old animals is highly vulnerable to acute insult, thus targeted protection is especially relevant in preventing liver damage. Simvastatin represents a useful therapeutic strategy in aging.

Restoration of liver sinusoidal cell phenotypes by statins improves portal hypertension and histology in rats with NASH.

Non-alcoholic steatohepatitis (NASH) is a common chronic liver disorder in developed countries, with the associated clinical complications driven by portal hypertension (PH). PH may precede fibrosis development, probably due to endothelial dysfunction at early stages of the disease. Our aim was to characterize liver sinusoidal endothelial cell (LSEC) dedifferentiation/capillarization and its contribution to PH in NASH, together with assessing statins capability to revert endothelial function improving early NASH stages. Sprague-Dawley rats were fed with high fat glucose-fructose diet (HFGFD), or control diet (CD) for 8 weeks and then treated with simvastatin (sim) (10 mg·kg-1·day-1), atorvastatin (ato) (10 mg·kg-1·day-1) or vehicle during 2 weeks. Biochemical, histological and hemodynamic determinations were carried out. Sinusoidal endothelial dysfunction was assessed in individualized sorted LSEC and hepatic stellate cells (HSC) from animal groups and in whole liver samples. HFGFD rats showed full NASH features without fibrosis but with significantly increased portal pressure compared with CD rats (10.47 ± 0.37 mmHg vs 8.30 ± 0.22 mmHg; p < 0.001). Moreover, HFGFD rats showed a higher percentage of capillarized (CD32b-/CD11b-) LSEC (8% vs 1%, p = 0.005) showing a contractile phenotype associated to HSC activation. Statin treatments caused a significant portal pressure reduction (sim: 9.29 ± 0.25 mmHg, p < 0.01; ato: 8.85 ± 0.30 mmHg, p < 0.001), NASH histology reversion, along with significant recovery of LSEC differentiation and a regression of HSC activation to a more quiescent phenotype. In an early NASH model without fibrosis with PH, LSEC transition to capillarization and HSC activation are reverted by statin treatment inducing portal pressure decrease and NASH features improvement.

A Nutraceutical Rich in Docosahexaenoic Acid Improves Portal Hypertension in a Preclinical Model of Advanced Chronic Liver Disease.

Inflammation and oxidative stress play a key role in the pathophysiology of advanced chronic liver disease (ACLD) and portal hypertension (PH). Considering the current lack of effective treatments, we evaluated an anti-inflammatory and antioxidant nutraceutical rich in docosahexaenoic acid (DHA) as a possible therapy for ACLD. We investigated the effects of two-week DHA supplementation (500 mg/kg) on hepatic fatty acids, PH, oxidative stress, inflammation, and hepatic stellate cell (HSC) phenotype in rats with ACLD. Additionally, the effects of DHA were evaluated in murine macrophages and human HSC. In contrast to vehicle-treated animals, cirrhotic rats receiving DHA reestablished a healthy hepatic fatty acid profile, which was associated with an improvement in PH. The mechanisms underlying this hemodynamic improvement included a reduction in oxidative stress and inflammation, as well as a marked HSC deactivation, confirmed in human HSC. Experiments with cultured macrophages showed that treatment with DHA protects against pro-inflammatory insults. The present preclinical study demonstrates that a nutraceutical rich in DHA significantly improves PH in chronic liver disease mainly by suppressing inflammation and oxidative stress-driven HSC activation, encouraging its evaluation as a new treatment for PH and cirrhosis.

Emricasan Ameliorates Portal Hypertension and Liver Fibrosis in Cirrhotic Rats Through a Hepatocyte-Mediated Paracrine Mechanism.

In cirrhosis, liver microvascular dysfunction is a key factor increasing hepatic vascular resistance to portal blood flow, which leads to portal hypertension. De-regulated inflammatory and pro-apoptotic processes due to chronic injury play important roles in the dysfunction of liver sinusoidal cells. The present study aimed at characterizing the effects of the pan-caspase inhibitor emricasan on systemic and hepatic hemodynamics, hepatic cells phenotype, and underlying mechanisms in preclinical models of advanced chronic liver disease. We investigated the effects of 7-day emricasan on hepatic and systemic hemodynamics, liver function, hepatic microcirculatory function, inflammation, fibrosis, hepatic cells phenotype, and paracrine interactions in rats with advanced cirrhosis due to chronic CCl4 administration. The hepato-protective effects of emricasan were additionally investigated in cells isolated from human cirrhotic livers. Cirrhotic rats receiving emricasan showed significantly lower portal pressure than vehicle-treated animals with no changes in portal blood flow, indicating improved vascular resistance. Hemodynamic improvement was associated with significantly better liver function, reduced hepatic inflammation, improved phenotype of hepatocytes, liver sinusoidal endothelial cells, hepatic stellate cells and macrophages, and reduced fibrosis. In vitro experiments demonstrated that emricasan exerted its benefits directly improving hepatocytes' expression of specific markers and synthetic capacity, and ameliorated nonparenchymal cells through a paracrine mechanism mediated by small extracellular vesicles released by hepatocytes. Conclusion: This study demonstrates that emricasan improves liver sinusoidal microvascular dysfunction in cirrhosis, which leads to marked amelioration in fibrosis, portal hypertension and liver function, and therefore encourages its clinical evaluation in the treatment of advanced chronic liver disease.

Improved hemodynamic and liver function in portal hypertensive cirrhotic rats after administration of B. pseudocatenulatum CECT 7765.

PURPOSE: Evaluating whether changes in gut microbiota induced by a bifidobacterial strain may have an effect on the hepatic vascular function in portal hypertensive cirrhotic rats. METHODS: Bile duct ligation (BDL) was performed in rats. A subgroup of animals received B. pseudocatenulatum CECT7765 (109 cfu/daily ig.) for 1 week prior to laparotomy. Hemodynamic, biochemical and inflammatory markers were evaluated. Ileal microbiota composition was identified. Statistical analysis was performed. RESULTS: Sham-operated (n = 6), BDL (n = 6) and BDL treated with bifidobacteria (n = 8) rats were included. B. pseudocatenulatum CECT7765 significantly decreased proteobacteria (p = 0.001) and increased Bacteroidetes (p = 0.001) relative abundance. The bifidobacteria decreased the Firmicutes/Bacteroidetes ratio in the BDL model (p = 0.03). BDL with bifidobacteria vs BDL rats showed: significantly reduced portal vein area, portal flow, congestion index, alkaline phosphatase and total bilirubin, significantly increased serum cytokines and nitric oxide levels, gene expression levels of bile acids receptor FXR and endothelial nitric oxide synthase. Quantitative changes in the Clostridiales and Bacteroidales orders were independently associated with variations in portal vein area and portal flow, while changes in the Proteobacteria phylum were independently associated with congestion. Variations in all liver function markers significantly correlated with total OTUs mainly in the Firmicutes, but only changes in the Clostridiales were independently associated with alkaline phosphatase in the ANCOVA analysis. CONCLUSION: Hemodynamic alterations and liver dysfunction induced by BDL in rats are partially restored after oral administration of B. pseudocatenulatum CECT7765. Results provide a proof-of-concept for the beneficial effect of this bifidobacterial strain in reducing complications derived from portal hypertension in cirrhosis.

4 in 1: Antibody-free protocol for isolating the main hepatic cells from healthy and cirrhotic single rat livers.

Liver cells isolated from pre-clinical models are essential tools for studying liver (patho)physiology, and also for screening new therapeutic options. We aimed at developing a new antibody-free isolation method able to obtain the four main hepatic cell types (hepatocytes, liver sinusoidal endothelial cells [LSEC], hepatic macrophages [HMΦ] and hepatic stellate cells [HSC]) from a single rat liver. Control and cirrhotic (CCl4 and TAA) rat livers (n = 6) were perfused, digested with collagenase and mechanically disaggregated obtaining a multicellular suspension. Hepatocytes were purified by low revolution centrifugations while non-parenchymal cells were subjected to differential centrifugation. Two different fractions were obtained: HSC and mixed LSEC + HMΦ. Further LSEC and HMΦ enrichment was achieved by selective adherence time to collagen-coated substrates. Isolated cells showed high viability (80%-95%) and purity (>95%) and were characterized as functional: hepatocytes synthetized albumin and urea, LSEC maintained endocytic capacity and in vivo fenestrae distribution, HMΦ increased expression of inflammatory markers in response to LPS and HSC were activated upon in vitro culture. The 4 in 1 protocol allows the simultaneous isolation of highly pure and functional hepatic cell sub-populations from control or cirrhotic single livers without antibody selection.

Hepatic microcirculation and mechanisms of portal hypertension.

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.

Advances in therapeutic options for portal hypertension.

Portal hypertension represents one of the major clinical consequences of chronic liver disease, having a deep impact on patients' prognosis and survival. Its pathophysiology defines a pathological increase in the intrahepatic vascular resistance as the primary factor in its development, being subsequently aggravated by a paradoxical increase in portal blood inflow. Although extensive preclinical and clinical research in the field has been developed in recent decades, no effective treatment targeting its primary mechanism has been defined. The present review critically summarizes the current knowledge in portal hypertension therapeutics, focusing on those strategies driven by the disease pathophysiology and underlying cellular mechanisms.

Effects of aging on liver microcirculatory function and sinusoidal phenotype.

The socioeconomic and medical improvements of the last decades have led to a relevant increase in the median age of worldwide population. Although numerous studies described the impact of aging in different organs and the systemic vasculature, relatively little is known about liver function and hepatic microcirculatory status in the elderly. In this study, we aimed at characterizing the phenotype of the aged liver in a rat model of healthy aging, particularly focusing on the microcirculatory function and the molecular status of each hepatic cell type in the sinusoid. Moreover, major findings of the study were validated in young and aged human livers. Our results demonstrate that healthy aging is associated with hepatic and sinusoidal dysfunction, with elevated hepatic vascular resistance and increased portal pressure. Underlying mechanisms of such hemodynamic disturbances included typical molecular changes in the cells of the hepatic sinusoid and deterioration in hepatocyte function. In a specific manner, liver sinusoidal endothelial cells presented a dysfunctional phenotype with diminished vasodilators synthesis, hepatic macrophages exhibited a proinflammatory state, while hepatic stellate cells spontaneously displayed an activated profile. In an important way, major changes in sinusoidal markers were confirmed in livers from aged humans. In conclusion, our study demonstrates for the first time that aging is accompanied by significant liver sinusoidal deregulation suggesting enhanced sinusoidal vulnerability to chronic or acute injuries.

Simvastatin Prevents Progression of Acute on Chronic Liver Failure in Rats With Cirrhosis and Portal Hypertension.

BACKGROUND & AIMS: Cirrhosis and its clinical consequences can be aggravated by bacterial infections, ultimately leading to the development of acute on chronic liver failure (ACLF), characterized by acute decompensation, organ failure, and high mortality within 28 days. Little is known about cellular and molecular mechanisms of ACLF in patients with cirrhosis, so no therapeutic options are available. We developed a sepsis-associated preclinical model of ACLF to facilitate studies of pathogenesis and evaluate the protective effects of simvastatin. METHODS: Male Wistar rats inhaled CCl4 until they developed cirrhosis (at 10 weeks) or cirrhosis with ascites (at 15-16 weeks). Male Sprague-Dawley rats received bile-duct ligation for 28 days or intraperitoneal thioacetamide for 10 weeks to induce cirrhosis. After induction of cirrhosis, some rats received a single injection of lipopolysaccharide (LPS) to induce ACLF; some were given simvastatin or vehicle (control) 4 hours or 24 hours before induction of ACLF. We collected data on changes in hepatic and systemic hemodynamics, hepatic microvascular phenotype and function, and survival times. Liver tissues and plasma were collected and analyzed by immunoblots, quantitative polymerase chain reaction, immuno(fluoro)histochemistry and immunoassays. RESULTS: Administration of LPS aggravated portal hypertension in rats with cirrhosis by increasing the severity of intrahepatic microvascular dysfunction, exacerbating hepatic inflammation, increasing oxidative stress, and recruiting hepatic stellate cells and neutrophils. Rats with cirrhosis given LPS had significantly shorter survival times than rats with cirrhosis given the control. Simvastatin prevented most of ACLF-derived complications and increased survival times. Simvastatin appeared to increase hepatic sinusoidal function and reduce portal hypertension and markers of inflammation and oxidation. The drug significantly reduced levels of transaminases, total bilirubin, and ammonia, as well as LPS-mediated activation of hepatic stellate cells in liver tissues of rats with cirrhosis. CONCLUSIONS: In studies of rats with cirrhosis, we found administration of LPS to promote development of ACLF, aggravating the complications of chronic liver disease and decreasing survival times. Simvastatin reduced LPS-induced inflammation and liver damage in rats with ACLF, supporting its use in treatment of patients with advanced chronic liver disease.