Phosphodiesterases in the Liver as Potential Therapeutic Targets of Cirrhotic Portal Hypertension.

Liver cirrhosis is a frequent condition with high impact on patients' life expectancy and health care systems. Cirrhotic portal hypertension (PH) gradually develops with deteriorating liver function and can lead to life-threatening complications. Other than an increase in intrahepatic flow resistance due to morphological remodeling of the organ, a functional dysregulation of the sinusoids, the smallest functional units of liver vasculature, plays a pivotal role. Vascular tone is primarily regulated by the nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathway, wherein soluble guanylate cyclase (sGC) and phosphodiesterase-5 (PDE-5) are key enzymes. Recent data showed characteristic alterations in the expression of these regulatory enzymes or metabolite levels in liver cirrhosis. Additionally, a disturbed zonation of the components of this pathway along the sinusoids was detected. This review describes current knowledge of the pathophysiology of PH with focus on the enzymes regulating cGMP availability, i.e., sGC and PDE-5. The results have primarily been obtained in animal models of liver cirrhosis. However, clinical and histochemical data suggest that the new biochemical model we propose can be applied to human liver cirrhosis. The role of PDE-5 as potential target for medical therapy of PH is discussed.

The soluble guanylate cyclase stimulator riociguat reduces fibrogenesis and portal pressure in cirrhotic rats.

In cirrhotic patients, portal hypertension (PHT) deteriorates survival, yet treatment options are limited. A major contributor to increased intrahepatic vasoconstriction in PHT is dysfunctional nitric-oxide signaling. Soluble guanylate cyclase (sGC) is the receptor of nitric-oxide and can be stimulated by riociguat. Riociguat is approved for pulmonary hypertension but has not been studied in liver cirrhosis. In this study we assessed the effects of riociguat on PHT and liver fibrosis in cholestatic (bile duct ligation, BDL) and toxic (carbon-tetrachloride, CCl4) rat models. In cirrhotic livers sGC expression was upregulated. In BDL rats, riociguat reduced liver fibrosis and decreased portal pressure without affecting systemic hemodynamics. In an early BDL disease stage, riociguat decreased bile duct proliferation, improved sinusoidal vascular dysfunction and inhibited angiogenesis. In advanced BDL riociguat exhibited anti-inflammatory effects. In CCl4 rats the beneficial effects of riociguat treatment were less pronounced and confined to an early disease stage. Similarly, in patients with cholestatic cirrhosis and PHT nitrates (that induce sGC activity) decreased portal pressure more effectively than in patients with non-cholestatic etiology. We also found an improvement of transaminases in patients with pulmonary hypertension receiving riociguat. Our findings support the clinical development of sGC stimulators in patients with cirrhotic PHT.

Case report of a modified Meso-Rex bypass as a treatment technique for late-onset portal vein cavernous transformation with portal hypertension after adult deceased-donor liver transplantation.

RATIONALE: Portal vein thrombosis is a complication after liver transplantation and cavernous transformation of the portal vein (CTPV) is a result of portal vein thrombosis, with symptoms of portal hypertension revealed by an enhanced CT scan. Meso-Rex bypass is an artificial shunt connecting the left portal vein to the superior mesenteric vein and is mainly used for idiopathic cavernomas. This technique is also used for post-transplant portal vein thrombosis in pediatric patients thereby bypassing obstructed sites of the extrahepatic portal vein. Here we report about an adult patient who was treated by connecting the cystic part of the portal vein to the splenic vein instead of the superior mesenteric vein. PATIENTS CONCERN: An adult male patient with post-liver transplantation portal vein cavernous transformation suffered from hypersplenism and elevated hepatic enzymes. DIAGNOSIS: The last follow up revealed irregular and obvious hypersplenism, and splenomegaly had occurred, while an enhanced CT scan revealed serious esophagogastric varices and CTPV in addition to occluded right and common PV trunks. INTERVENTION: The patient was treated by connecting the cystic part of the portal vein to the splenic vein instead of the superior mesenteric vein. OUTCOME: After the operation, a satisfactory velocity was confirmed 1 month postoperatively and the shunt still remained patent at the 6-month postoperation follow-up. LESSONS: A Meso-Rex bypass intervention connecting the left portal vein to the splenic vein instead of the superior mesenteric vein after liver transplantation in an adult patient with right and common portal vein occlusions has been successfully performed as an alternative approach.

Effects of Nuclear Factor-E2-related factor 2/Heme Oxygenase 1 on splanchnic hemodynamics in experimental cirrhosis with portal hypertension.

OBJECTIVE: We explored the effects of Nuclear Factor-E2-related factor 2 (Nrf2) and Heme Oxygenase 1 (HO-1) on splanchnic hemodynamics in portal hypertensive rats. METHODS: Experimental cirrhosis with portal hypertension was induced by intraperitoneal injection of carbon tetrachloride. The expression of proteins was examined by immunoblotting. Hemodynamic studies were performed by radioactive microspheres. The vascular perfusion system was used to measure the contractile response of mesentery arterioles in rats. RESULTS: Nrf2 expression in the nucleus and HO-1 expression in cytoplasm was significantly enhanced in portal hypertensive rats. Portal pressure, as well as regional blood flow, increased significantly in portal hypertension and can be blocked by tin protoporphyrin IX. The expression of endogenous nitric oxide synthase and vascular endothelial growth factors increased significantly compared to normal rats, while HO-1 inhibition decreased the expression of these proteins significantly. The contractile response of mesenteric arteries decreased in portal hypertension, but can be partially recovered through tin protoporphyrin IX treatment. CONCLUSIONS: The expression of Nrf2/HO-1 increased in mesenteric arteries of portal hypertensive rats, which was related to oxidative stress. HO-1was involved in increased portal pressure and anomaly splanchnic hemodynamics in portal hypertensive rats.

Thalidomide ameliorates portal hypertension via nitric oxide synthase independent reduced systolic blood pressure.

AIM: Portal hypertension is a common complication of liver cirrhosis and significantly increases mortality and morbidity. Previous reports have suggested that the compound thalidomide attenuates portal hypertension (PHT). However, the mechanism for this action is not fully elucidated. One hypothesis is that thalidomide destabilizes tumor necrosis factor α (TNFα) mRNA and therefore diminishes TNFα induction of nitric oxide synthase (NOS) and the production of nitric oxide (NO). To examine this hypothesis, we utilized the murine partial portal vein ligation (PVL) PHT model in combination with endothelial or inducible NOS isoform gene knockout mice. METHODS: Wild type, inducible nitric oxide synthase (iNOS)(-/-) and endothelial nitric oxide synthase (eNOS)(-/-) mice received either PVL or sham surgery and were given either thalidomide or vehicle. Serum nitrate (total nitrate, NOx) was measured daily for 7 d as a surrogate of NO synthesis. Serum TNFα level was quantified by enzyme-linked immunosorbent assay. TNFα mRNA was quantified in liver and aorta tissue by reverse transcription-polymerase chain reaction. PHT was determined by recording splenic pulp pressure (SPP) and abdominal aortic flow after 0-7 d. Response to thalidomide was determined by measurement of SPP and mean arterial pressure (MAP). RESULTS: SPP, abdominal aortic flow (Qao) and plasma NOx were increased in wild type and iNOS(-/-) PVL mice when compared to sham operated control mice. In contrast, SPP, Qao and plasma NOx were not increased in eNOS(-/-) PVL mice when compared to sham controls. Serum TNFα level in both sham and PVL mice was below the detection limit of the commercial ELISA used. Therefore, the effect of thalidomide on serum TNFα levels was undetermined in wild type, eNOS(-/-) or iNOS(-/-) mice. Thalidomide acutely increased plasma NOx in wild type and eNOS(-/-) mice but not iNOS(-/-) mice. Moreover, thalidomide temporarily (0-90 min) decreased mean arterial pressure, SPP and Qao in wild type, eNOS(-/-) and iNOS(-/-) PVL mice, after which time levels returned to the respective baseline. CONCLUSION: Thalidomide does not reduce portal pressure in the murine PVL model by modulation of NO biosynthesis. Rather, thalidomide reduces PHT by decreasing MAP by an undetermined mechanism.

Inhibition of Janus kinase-2 signalling pathway ameliorates portal hypertensive syndrome in partial portal hypertensive and liver cirrhosis rats.

BACKGROUND AND AIMS: JAK2/STAT3 signalling promotes fibrosis, angiogenesis and inflammation in many diseases; however, the role of this pathway in portal hypertension remains obscure. This study aimed to explore the function of JAK2/STAT3 signalling in portal hypertension and estimate the potential therapeutic effect of treatment with the specific inhibitor AG490. METHODS: Rats induced by partial portal vein ligation and common bile duct ligation were treated with AG490 for two weeks. Haemodynamic parameters were assessed. The levels of phospho-STAT3 protein and related cytokines were detected by western blotting of splanchnic organs. Liver, spleen and intestine characterization was performed using histological analyses. Peripheral blood cell counts were also detected. RESULTS: High levels of phospho-STAT3 protein were detected in portal hypertensive rats. AG490 effectively inhibited JAK2/STAT3 signalling and its downstream cytokines and provided protective effects by decreasing splanchnic neovascularization and inflammation and by attenuating portal pressure and hyperdynamic splanchnic circulation. In cirrhosis rats, AG490 inhibited intrahepatic fibrosis, angiogenesis and inflammation. AG490 improved the peripheral blood cell counts and the splenomegaly observed in these rats. CONCLUSIONS: JAK2/STAT3 signalling is essential in portal hypertension, and targeting JAK2/STAT3 may be a promising therapy to treat this condition.

Novel LIPA mutations in Mexican siblings with lysosomal acid lipase deficiency.

Lysosomal acid lipase (LAL) deficiency is an under-recognized lysosomal disease caused by deficient enzymatic activity of LAL. In this report we describe two affected female Mexican siblings with early hepatic complications. At two months of age, the first sibling presented with alternating episodes of diarrhea and constipation, and later with hepatomegaly, elevated transaminases, high levels of total and low-density lipoprotein cholesterol, and low levels of high-density lipoprotein. Portal hypertension and grade 2 esophageal varices were detected at four years of age. The second sibling presented with hepatomegaly, elevated transaminases and mildly elevated low-density lipoprotein and low high-density lipoprotein at six months of age. LAL activity was deficient in both patients. Sequencing of LIPA revealed two previously unreported heterozygous mutations in exon 4: c.253C>A and c.294C>G. These cases highlight the clinical continuum between the so-called Wolman disease and cholesteryl ester storage disease, and underscore that LAL deficiency represents a single disease with a degree of clinical heterogeneity.

Hepatic dimethylarginine-dimethylaminohydrolase1 is reduced in cirrhosis and is a target for therapy in portal hypertension.

BACKGROUND & AIMS: Portal hypertension is characterized by reduced hepatic eNOS activity. Asymmetric-dimethylarginine (ADMA), an eNOS inhibitor, is elevated in cirrhosis and correlates with the severity of portal hypertension. Dimethylarginine dimethylaminohydrolase-1 (DDAH-1) is the key enzyme metabolizing hepatic ADMA. This study characterized DDAH-1 in cirrhosis, and explored hepatic DDAH-1 reconstitution through farnesoid X receptor (FXR) agonism and DDAH-1 gene therapy. METHODS: DDAH-1 immunohistochemistry was conducted on human cirrhosis and healthy liver tissue. Subsequently, sham-operated or bile-duct-ligated (BDL) cirrhosis rats were treated with the FXR agonist obeticholic acid (OA, 5 mg/kg) or vehicle for 5 days. Further, animals underwent hydrodynamic injection with DDAH-1-expressing plasmid or saline control, which resulted in the following groups: sham+saline, BDL+saline, BDL+DDAH-1-plasmid. Portal pressure (PP) measurements were performed. Plasma ALT was measured by COBAS INTEGRA, DDAH-1 expression by qPCR and Western blot, eNOS activity by radiometric assay. RESULTS: Immunohistochemistry and Western-blotting confirmed hepatic DDAH-1 was restricted to hepatocytes, and expression decreased significantly in cirrhosis. In BDL rats, reduced DDAH-1 expression was associated with elevated hepatic ADMA, reduced eNOS activity and high PP. OA treatment significantly increased DDAH-1 expression, reduced hepatic tissue ADMA, and increased liver NO generation. PP was significantly reduced in BDL+OA vs. BDL+vehicle (8±1 vs. 13.5±0.6 mmHg; p<0.01) with no change in the mean arterial pressure (MAP). Similarly, DDAH-1 hydrodynamic injection significantly increased hepatic DDAH-1 gene and protein expression, and significantly reduced PP in BDL+DDAH-1 vs. BDL+saline (p<0.01). CONCLUSIONS: This study demonstrates DDAH-1 is a specific molecular target for portal pressure reduction, through actions on ADMA-mediated regulation of eNOS activity. Our data support translational studies, targeting DDAH-1 in cirrhosis and portal hypertension.

Glycogen phosphorylase BB levels are associated with haemodynamic parameters in cirrhotic patients undergoing transjugular intrahepatic portosystemic shunt insertion.

BACKGROUND: Transjugular intrahepatic portosystemic shunts (TIPS) have become a widely accepted tool in the treatment of patients with symptomatic portal hypertension. The aim of our study was to assess glycogen phosphorylase BB (GPBB) concentration in relation to echocardiographic and haemodynamic parameters in patients before and after TIPS insertion. METHODS: The study population consisted of 55 patients (38 men and 17 women, age 55.6±8.9 years, range 37-74 years) with liver cirrhosis treated with transjugular portosystemic shunting. GPBB, echocardiographic, and haemodynamic parameters were measured before TIPS insertion and 24 h after the procedure. GPBB concentrations were assessed using the Cardiac Array for Evidence Investigator protein biochip. Correlation between parameters was assessed using the Spearman's coefficient. RESULTS: Serum post-procedural GPBB concentrations were increased in comparison with baseline (5.58 vs. 2.67 µg/L, P<0.001). GPBB concentration after TIPS significantly correlated with baseline systemic vascular resistence (r=0.330; P=0.017) and cardiac index (r=0.313; P=0.025). CONCLUSION: GPBB concentration measurement may be a useful tool for monitoring myocardial ischemia during a TIPS procedure.

Glytan decreases portal pressure via mesentery vasoconstriction in portal hypertensive rats.

AIM: To investigate the effects of Glytan on splanchnic hemodynamics and its reduction of portal pressure in portal hypertensive rats. METHODS: Glytan (Ganluotong in Chinese), is composed of salvianolic acid B and diammonium glycyrrhizinate. Portal hypertension (PHT) was induced in the rats by common bile duct ligation (BDL). Hemodynamic studies were performed using the colored microsphere method. Radioimmunoassay (RIA) was used to determine endothelin (ET)-1 levels in the mesenteric circulation. Western blotting methods were used to investigate the effect of Glytan on ET A receptor (ETAR), ET B receptor (ETBR), endothelial NO synthase (eNOS), G-protein-coupled receptor kinase (GRK)2, and ß-arrestin 2 expression in the mesentery. The mRNA of ETAR and ETBR was determined using real-time polymerase chain reaction. RESULTS: Treatment with Glytan reduced portal pressure (PP) and portal territory blood flow (PTBF) and increased both mean arterial pressure (MAP) and splanchnic vascular resistance (SVR). Especially at 4 wk, PP decreased by about 40%, while MAP increased by 13%, SVR increased by 12%, and PTBF decreased by about 21%. The effect of blood flow reduction was greatest in the mesentery (about 33%) at 4 wk. The mesenteric circulation ET-1 levels of BDL rats were lower and negatively correlated with PP at 4 wk. Glytan can increase mesenteric ET-1 content and inhibit ETBR, eNOS, GRK2, and ß-arrestin 2 expression in the mesentery. Moreover, Glytan showed no effect on the expression of ETAR protein and mRNA. CONCLUSION: The decreased PP and PTBF observed after Glytan treatment were related to increased mesenteric vasoconstriction and increased receptor sensitivity to vasoconstrictor.

Antioxidant properties of glutamine and its role in VEGF-Akt pathways in portal hypertension gastropathy.

AIM: To investigate the effects of glutamine on oxidative/nitrosative stress and the vascular endothelial growth factor (VEGF)-Akt-endothelial nitric oxide synthase (eNOS) signaling pathway in an experimental model of portal hypertension induced by partial portal vein ligation (PPVL). METHODS: Portal hypertension was induced by PPVL. The PPVL model consists of a partial obstruction of the portal vein, performed using a 20 G blunt needle as a guide, which is gently removed after the procedure. PPVL model was performed for 14 d beginning treatment with glutamine on the seventh day. On the fifteenth day, the mesenteric vein pressure was checked and the stomach was removed to test immunoreactivity and oxidative stress markers. We evaluated the expression and the immunoreactivity of proteins involved in the VEGF-Akt-eNOS pathway by Western blotting and immunohistochemical analysis. Oxidative stress was measured by quantification of the cytosolic concentration of thiobarbituric acid reactive substances (TBARS) as well as the levels of total glutathione (GSH), superoxide dismutase (SOD) activity, nitric oxide (NO) production and nitrotyrosine immunoreactivity. RESULTS: All data are presented as the mean ± SE. The production of TBARS and NO was significantly increased in PPVL animals. A reduction of SOD activity was detected in PPVL + G group. In the immunohistochemical analyses of nitrotyrosine, Akt and eNOS, the PPVL group exhibited significant increases, whereas decreases were observed in the PPVL + G group, but no difference in VEGF was detected between these groups. Western blotting analysis detected increased expression of phosphatidylinositol-3-kinase (PI3K), P-Akt and eNOS in the PPVL group compared with the PPVL + G group, which was not observed for the expression of VEGF when comparing these groups. Glutamine administration markedly alleviated oxidative/nitrosative stress, normalized SOD activity, increased levels of total GSH and blocked NO overproduction as well as the formation of peroxynitrite. CONCLUSION: Glutamine treatment demonstrated to reduce oxidative damage but does not reduce angiogenesis induced by PH in gastric tissue, demonstrating a beneficial role for the PI3K-Akt-eNOS pathway.

Amelioration of carbon tetrachloride-induced cirrhosis and portal hypertension in rat using adenoviral gene transfer of Akt.

AIM: To investigate whether a virus constitutively expressing active Akt is useful to prevent cirrhosis induced by carbon tetrachloride (CCl4). METHODS: Using cre-loxp technique, we created an Ad-myr-HA-Akt virus, in which Akt is labeled by a HA tag and its expression is driven by myr promoter. Further, through measuring enzyme levels and histological structure, we determined the efficacy of this Ad-myr-HA-Akt virus in inhibiting the development of cirrhosis induced by CCl4 in rats. Lastly, using western blotting, we examined the expression levels and/or phosphorylation status of Akt, apoptotic mediators, endothelial nitric oxide synthase (eNOS), and markers for hepatic stellate cells activation to understand the underlying mechanisms of protective role of this virus. RESULTS: The Ad-myr-HA-Akt virus was confirmed using polymerase chain reaction amplification of inserted Akt gene and sequencing for full length of inserted fragment, which was consistent with the sequence reported in the GenBank. The concentrations of Ad-myr-HA-Akt and adenoviral enhanced green fluorescent protein (Ad-EGFP) virus used in the current study were 5.5 × 10(11) vp/mL. The portal vein diameter, peak velocity of blood flow, portal blood flow and congestion index were significantly increased in untreated, saline and Ad-EGFP cirrhosis groups when compared to normal control after the virus was introduced to animal through tail veil injection. In contrast, these parameters in the Akt cirrhosis group were comparable to normal control group. Compared to the normal control, the liver function (Alanine aminotransferase, Aspartate aminotransferase and Albumin) was significantly impaired in the untreated, saline and Ad-EGFP cirrhosis groups. The Akt cirrhosis group showed significant improvement of liver function when compared to the untreated, saline and Ad-EGFP cirrhosis groups. The Hyp level and portal vein pressure in Akt cirrhosis groups were also significantly lower than other cirrhosis groups. The results of HE and Van Gieson staining indicated that Akt group has better preservation of histological structure and less fibrosis than other cirrhosis groups. The percentage of apoptotic cell was greatly less in Akt cirrhosis group than in other cirrhosis groups. Akt group showed positive HA tag and an increased level of phosphorylated Akt as well as decreased levels of Fas. In contrast, Caspase-3 and Caspase-9 levels in Akt group were significantly lower than other cirrhosis groups. Noticeable decrease of DR5 and α-SMA and increase of phosphorylated eNOS were observed in the Akt group when compared to other cirrhosis groups. The NO level in liver was significantly higher in Akt group than other cirrhosis groups, which was consistent with the level of phosphorylated eNOS in these groups. CONCLUSION: This study suggest that Ad-myr-HA-Akt virus is a useful tool to prevent CCl4-induced cirrhosis in rat model and Akt pathway may be a therapeutic target for human cirrhosis.

Small liver remnants are more vulnerable to ischemia/reperfusion injury after extended hepatectomies: a case-control study.

BACKGROUND: There is evidence that small-for-size liver grafts are more vulnerable to ischemia/reperfusion injury after liver transplantation. We hypothesized that ischemic injury is more pronounced in small liver remnants after major hepatectomies. METHODS: Fifteen patients underwent extended hepatectomy with remnant liver mass less than 30% of standard liver weight (study group). These patients were matched with patients who underwent minor liver resection, with liver remnants equal to or more than 70% of standard liver weight (control group). Ischemia/reperfusion injury was assessed by tissue caspase-3 activity postoperatively as well as peak aspartate aminotransferase (AST) values and a-glutathione S-transferase (α-GST) levels adjusted for remnant liver weight. In addition, caspase-3 activity and adjusted serum markers of hepatocyte injury were correlated with the degree of postoperative portal hypertension. RESULTS: Caspase-3 activity was higher in patients with small liver remnants (22.66±6.57 vs. 12.60±4.06 count per high-power field, p<0.001). Serum markers of hepatocyte injury, when adjusted per gram of liver remnant, were found to be higher in the study group than in the control group (AST: 1.26±0.25 vs. 0.54±0.11 IU g(-1), p<0.001; α-GST: 0.14±0.02 vs. 0.08±0.01 IU g(-1), p<0.001). Tissue caspase-3 expression in the small liver remnant group correlated with both AST and α-GST levels adjusted per gram of liver remnant (r2=0.51, p=0.005 and r2=0.71, p<0.001, respectively). Significant correlations between postoperative portal hypertension and the same markers as well as caspase-3 activity were also demonstrated. CONCLUSION: Liver remnants less than 30% of standard liver weight are much more susceptible to ischemia/reperfusion injury than controls twice the size. Adjustment of serum markers of hepatocyte injury to the liver remnant weight depicts injury more accurately.

Clinical analysis of 15 cases of liver nodular regenerative hyperplasia.

Nodular regenerative hyperplasia (NRH) of liver may be one of the leading causes of non-cirrhotic intrahepatic portal hypertension (NCIPH), although the exact relationship is currently unknown. Diagnosis of NRH is relatively difficult and involves surgical pathology, and thus it is necessary to improve the preoperative recognition of NRH. Here, we analyze 15 cases of NRH to better understand this disease. All the liver specimens were microscopically examined by hematoxylin-eosin staining and reticulin and Masson trichrome staining. Diagnoses of NRH were confirmed by pathological examination. Clinically, NRH presents as diffused liver lesions with mildly increased liver enzymes. Portal hypertension is the most common clinical manifestation presenting prominently as splenomegaly, hypersplenism, and esophageal varices bleeding. NRH is often associated with autoimmune or collagen vascular diseases, and such patients often present with a variety of positive autoantibodies and increased erythrocyte sedimentation rate (ESR), Ig and γ %. Pathological examination of the liver showed diffuse small regenerative nodules without fibrous septa and obstructive portal venopathy. For those patients with portal hypertension of unknown cause and preserved liver function, especially, those combined with autoimmune diseases, NRH should be considered.

Therapeutic effect of captopril, pentoxifylline, and cordyceps sinensis in pre-hepatic portal hypertensive rats.

BACKGROUND/AIM: Portal hypertension is an important and potentially fatal complication of liver disease whereby cellular and fibrotic alterations manifest to increase portal venous pressure. The aim of this study is to investigate the effect of captopril, pentoxifylline (PTX), and cordyceps sinensis in pre-hepatic portal hypertensive rats. SETTINGS AND DESIGN: Wistar male rats were divided at random into 3 main groups: the first group: control rats. The second group: sham-operated rats and the third group: prehepatic portal hypertensive rats (PHPHT) induced by regulated pre-hepatic portal vein ligation. After 14 days, Group 3 was subdivided into 5 subgroups. Subgroup (1): portal vein-ligated (PVL) was killed at once; Subgroup (2): received distilled water for 30 days (untreated PVL group); subgroups 3-5 were treated with captopril (60 mg/kg, orally); PTX (100 mg/kg, orally); and C. sinensis (200 mg/kg, orally), respectively, as a single daily dose for 30 days. PATIENTS AND METHODS: Portal pressure, nitric oxide (NO), antioxidant enzymes, Liver enzymes, and creatinine levels were measured to evaluate the status of the liver state. RESULTS: Portal vein ligation produced significant increments in liver enzymes, NO, creatinine and portal pressure concomitant with significant decrements in glutathione content and superoxide dismutase activity. Treatment with captopril, PTX, and C. sinensis resulted in a significant reduction in liver enzymes, NO, creatinine and portal pressure and observable increase in antioxidant enzymes. CONCLUSIONS: captopril, PTX, and C. sinensis have promising effect in controlling PHPHT and reducing hyperdynamic circulatory state through reduction of portal pressure and NO level.

Droxidopa, an oral norepinephrine precursor, improves hemodynamic and renal alterations of portal hypertensive rats.

UNLABELLED: We aimed to evaluate the effects of droxidopa (an oral synthetic precursor of norepinephrine) on the hemodynamic and renal alterations of portal hypertensive rats. Sham, portal vein-ligated (PVL), and 4-week biliary duct-ligated (BDL) rats received a single oral dose of droxidopa (25-50 mg/kg) or vehicle and hemodynamic parameters were monitored for 2 hours. Two groups of BDL and cirrhotic rats induced by carbon tetrachloride (CCl(4) ) were treated for 5 days with droxidopa (15 mg/kg, twice daily, orally); hemodynamic parameters and blood and urinary parameters were assessed. The droxidopa effect on the Rho kinase (RhoK) / protein kinase B (AKT) / endothelial nitric oxide synthase (eNOS) pathways was analyzed by western blot in superior mesenteric artery (SMA). The acute administration of droxidopa in PVL and BDL rats caused a significant and maintained increase in arterial pressure and mesenteric arterial resistance, with a significant decrease of mesenteric arterial and portal blood flow, without changing portal pressure and renal blood flow. Two-hour diuresis greatly increased. Carbidopa (DOPA decarboxylase inhibitor) blunted all effects of droxidopa. Chronic droxidopa therapy in BDL rats produced the same beneficial hemodynamic effects observed in the acute study, did not alter liver function parameters, and caused a 50% increase in 24-hour diuresis volume (7.4 ± 0.9 mL/100g in BDL vehicle versus 11.8 ± 2.5 mL/100g in BDL droxidopa; P = 0.01). Droxidopa-treated rats also showed a decreased ratio of p-eNOS/eNOS and p-AKT/AKT and increased activity of RhoK in SMA. The same chronic treatment in CCl(4) rats caused similar hemodynamic effects and produced significant increases in diuresis volume and 24-hour natriuresis (0.08 ± 0.02 mmol/100g in CCl(4) vehicle versus 0.23 ± 0.03 mmol/100g in CCl(4) droxidopa; P = 0.014). CONCLUSION: Droxidopa might be an effective therapeutic agent for hemodynamic and renal alterations of liver cirrhosis and should be tested in cirrhosis patients.

G-protein-coupled receptor kinase interactor-1 (GIT1) is a new endothelial nitric-oxide synthase (eNOS) interactor with functional effects on vascular homeostasis.

Endothelial cell nitric-oxide (NO) synthase (eNOS), the enzyme responsible for synthesis of NO in the vasculature, undergoes extensive post-translational modifications that modulate its activity. Here we have identified a novel eNOS interactor, G-protein-coupled receptor (GPCR) kinase interactor-1 (GIT1), which plays an unexpected role in GPCR stimulated NO signaling. GIT1 interacted with eNOS in the endothelial cell cytoplasm, and this robust association was associated with stimulatory eNOS phosphorylation (Ser(1177)), enzyme activation, and NO synthesis. GIT1 knockdown had the opposite effect. Additionally, GIT1 expression was reduced in sinusoidal endothelial cells after liver injury, consistent with previously described endothelial dysfunction in this disease. Re-expression of GIT1 after liver injury rescued the endothelial phenotype. These data emphasize the role of GPCR signaling partners in eNOS function and have fundamental implications for vascular disorders involving dysregulated eNOS.