Clinical Pathways and Outcomes of Andexanet Alfa Administration for the Reversal of Critical Bleeding in Patients on Oral Direct Factor Xa Inhibitors.

Background Andexanet is U.S. Food and Drug Administration (FDA) approved for the reversal of critical bleeding from factor Xa inhibitors and off-label for surgical reversal. Data are lacking on andexanet administration processes. Methods We retrospectively studied patients at a 23-hospital system who received andexanet from November 2019 to March 2023. Abstractors coded demographics, comorbidities, anticoagulant use, andexanet indication, and process times. The primary outcome was presentation-to-andexanet time; diagnosis, ordering, and administration times were calculated. Secondary outcomes included in-hospital postandexanet major thromboembolism/bleeding and mortality. Results In total, 141 patients were analyzed. Andexanet indications were predominantly neurologic bleeding (85.8%). Twenty-four patients (17.0%) were transferred from nontertiary/academic centers to tertiary/academic centers. The median presentation-to-administration time was 192.5 minutes (interquartile range [IQR]: 108.0-337.0 minutes). Components were as follows: 72.5 minutes (IQR: 39.0-137.5 minutes) for bleeding diagnosis; 35.5 minutes (IQR: 0-96.5 minutes) for andexanet ordering; and 53.0 minutes (IQR: 38.5-78.5 minutes) for administration, which was longer at tertiary/academic hospitals (ratio 1.5, 95% confidence interval [CI]: 1.2-2.0, p = 0.002). Gastrointestinal or other critical bleeding (ratio 2.59, 95% CI: 1.67-4.02, p < 0.001), and tertiary/academic center treatment (ratio 1.58, 95% CI: 1.15-2.18, p = 0.005), were associated with increased time. Major thromboembolism, bleeding, and mortality occurred in 10.6, 12.0, and 22.9% of patients, respectively. Conclusions In our cohort, the median presentation-to-administration time was over 3 hours. Cumulative times were longer at tertiary/academic hospitals and for gastrointestinal/other bleeding. Postandexanet major thromboembolism/bleeding occurred more at tertiary/academic hospitals, possibly related to transfers. Prospective studies may elucidate clinical decision-making bottlenecks.

Depression and anxiety among patients treated for bladder cancer: examining clinical, demographic, and psychosocial predictors.

INTRODUCTION AND OBJECTIVE: The significant impact of bladder cancer and treatment on patient health related quality of life (HRQoL) and emotional wellbeing has been documented. Increasing evidence from cancer research emphasizes the importance of examining patients supportive care needs and received social support as factors that could influence their emotional adjustment. The purpose of the study was to assess the demographic and clinical predictors of depression and anxiety among bladder cancer patients and its associations with patient reported supportive care needs and perceived availability of social support. METHODS: A cross-sectional design was used to investigate the study questions. Bladder cancer patients were recruited from the Bladder Cancer Advocacy Network (BCAN) to complete a questionnaire that included the Hospital Anxiety and Depression Scale (HADS), bladder cancer patient need survey (BCNAS-32), and the social provisions scale (SPS). The inclusion criteria restricted our sample to include bladder cancer patients who were English speakers, aged 18-85 years, and were able and willing to provide informed consent. Patients who had metastatic disease, cancer recurrence, or other primary cancers at the time of assessment were excluded from the study. RESULTS: Participants included 159 bladder cancer patients. The mean age was 62±9.4 years and 51% were male. Almost two-thirds (62%) of patients reported a diagnosis of muscle invasive bladder cancer (MIBC), 25% patient reported clinically significant levels of anxiety, 17% reported clinically significant levels of depression, and 13% and 17% reported abnormal borderline abnormal levels for anxiety and depression, respectively. Univariate regression analyses revealed significant associations between HADS total score, HADS depression and anxiety subscales, patient age, physical functioning/daily living needs, sexuality needs, and perceived social support with higher total scores, anxiety, and depression scores associated with younger age, higher unmet needs, and lower levels of social support. Multivariate regression analyses, showed similar findings confirming the associations depicted by the univariate regression analyses. CONCLUSIONS: Bladder cancer patients experience significant levels of depression and anxiety and these levels are associated with patient age, supportive care unmet needs and lack of social support. Patient focused interventions could be tailored to address these issues with the goal to improve patient HRQoL and emotional adjustment.

Vaccine-breakthrough SARS-CoV-2 infections in people with multiple sclerosis and related conditions: An observational study by the New York COVID-19 Neuro-Immunology Consortium (NYCNIC-2).

BACKGROUND: People with MS (PwMS) and related conditions treated with anti-CD20 and S1P modulating therapies exhibit attenuated immune responses to SARS-CoV-2 vaccines. It remains unclear whether humoral/T-cell responses are valid surrogates for postvaccine immunity. OBJECTIVE: To characterize COVID-19 vaccine-breakthrough infections in this population. METHODS: We conducted a prospective multicenter cohort study of PwMS and related CNS autoimmune conditions with confirmed breakthrough infections. Postvaccination antibody response, disease-modifying therapies (DMTs) at the time of vaccination, and DMT at the time of infection were assessed. RESULTS: Two hundred nine patients had 211 breakthrough infections. Use of anti-CD20 agents at time of infection was associated with increased infection severity (p = 0.0474, odds ratio (OR) = 5.923) for infections during the Omicron surge and demonstrated a trend among the total cohort (p = 0.0533). However, neither use of anti-CD20 agents at the time of vaccination nor postvaccination antibody response was associated with hospitalization risk. Anti-CD20 therapies were relatively overrepresented compared to a similar prevaccination-era COVID-19 cohort. CONCLUSION: Use of anti-CD20 therapies during vaccine breakthrough COVID-19 infection is associated with higher severity. However, the attenuated postvaccination humoral response associated with anti-CD20 therapy use during vaccination may not drive increased infection severity. Further studies are necessary to determine if this attenuated vaccine response may be associated with an increased likelihood of breakthrough infection.

Effect of access site choice on inferior vena cava filter angulation and outcomes.

BACKGROUND: In the present study, we compared the outcomes of inferior vena cava (IVC) filter placement between the femoral vein (FV) and internal jugular (IJ) vein access sites. METHODS: We performed a retrospective study using the Vascular Quality Initiative database to assess patients who had undergone IVC filter placement from 2013 to 2019. The patients were placed into two groups according to the access site location: FV and IJ vein. The FV group included patients with access via the right and left FVs and other leg veins, and the IJ group included patients with access via the right or left IJ vein. The primary outcome was the rate of filter angulation. The secondary outcomes included access site complications such as deep vein thrombosis, hematoma, and bleeding requiring transfusion. RESULTS: Of 13,221 patients, 8214 (63%) had undergone IVC filter placement via FV access and 4789 (37%) via IJ access. The remaining 218 patients had had an unknown access site or were excluded. Within the IJ group, 4696 (98.0%) had undergone access via the right IJ and 93 (2%) via the left IJ. Within the FV (common femoral, femoral, or other infrainguinal veins) group, 7007 (85.3%) had undergone access via the right FV and 1207 (14.6%) via the left FV. The mean patient age was 63 ± 15.9 years, the mean body mass index was 30.9 ± 9.60 kg/m2, and 6788 of the patients were men (52.0%). The most common indication for filter placement was a contraindication to anticoagulation because of a recent or active bleeding episode (30%), followed by planned surgery (22%), new deep vein thrombosis/pulmonary embolism (7%), fall risk (5%), and trauma (4%). Infrarenal filters had been placed in 97.9% of the patients. Univariate analysis identified body mass index and suprarenal placement as independent risk factors for angulation. The final multivariate analysis showed a significant increase in angulation (0.9% vs 0.34%; odds ratio, 1.46; 95% confidence interval, 1.02-2.11; P = .04) and increased access site complications (0.25% vs 0.07%; odds ratio, 2.068; 95% confidence interval, 1.01-4.23; P = .048) in the FV access group. No significant correlation between the access site and retrieval rate was found (P = .9270). CONCLUSIONS: Placement of IVC filters via IJ access showed a lower rate of filter angulation in the IVC and fewer access site complications compared with FV access.

Outcomes of Endovascular Management of Isolated Profunda Femoris Artery Occlusive Disease.

BACKGROUND: Endovascular management of isolated profunda femoris artery occlusive disease has not been well studied. Our aim is to analyze the outcomes of endovascular management of profunda artery occlusive disease. METHODS: This is a retrospective analysis using data from the Vascular Quality Initiative. All patients from 2013 to 2018 treated percutaneously for isolated profunda artery occlusive disease were included. Endovascular treatment included plain balloon alone, stent, stent graft, atherectomy, and drug-coated balloon without any concomitant endovascular or surgical treatment. Demographic, procedural, and follow-up data were obtained. Primary end points were primary patency, improvement of symptoms, and need for reintervention. Univariate and multivariable analysis was used to assess for significant variables. RESULTS: Of the 105,568 lower extremity endovascular interventions performed during this time period, there were 361 procedures (0.3%) performed on 341 patients for isolated profunda artery occlusive disease. The average age of these patients was 67.8 years (+/- 11.8), with 59.8% being men. The most common indication for treatment of the profunda artery was claudication (44.8%), followed by tissue loss (28.5%) and rest pain (26.0%). The most common treatment modality was plain balloon (58.5%), followed by stent (18.6%), drug-coated balloon (10.0%), atherectomy (9.4%), and stent graft (3.6%). At a mean follow-up of 13 months (+/-4.6), data were available for 238 patients (69.7%). Overall primary patency at 13 months was 92.9%. There was no significant difference in terms of patency for each treatment modality (Table I). Preoperative ambulatory status, aspirin, and statin were significantly associated with patency. At most recent follow-up, 67% of patients had improvement of their symptoms, whereas 29% were unchanged. Reintervention data were available for 247 patients, with a reintervention rate of 15.8% (n = 39) and a mean reintervention time of 226 days (+/- 173), with the majority of reinterventions (62%) occurring in the plain balloon group. Reinterventions were primarily endovascular (64%) with 9 patients (23%) undergoing surgical reintervention. CONCLUSIONS: Endovascular management of profunda femoris artery occlusive disease has acceptable one-year patency rates with low reintervention rates. Endovascular treatment may be an acceptable alternative to selected patients who are high-risk for surgery.

The Effects of a Comprehensive Multidisciplinary Outpatient Diabetes Program on Hospital Readmission Rates in Patients with Diabetes: A Randomized Controlled Prospective Study.

OBJECTIVE: The diagnosis of diabetes mellitus is associated with an increased risk of hospital readmissions. The goal of this study was to determine whether there was a difference in the rates of 30-day and 365-day hospital readmissions between diabetic patients who, upon their discharge, received diabetes care in a standard primary care setting and those who received their care in a specialized multidisciplinary diabetes program. METHODS: This was a randomized controlled prospective study. RESULTS: One hundred and ninety two consecutive patients were recruited into the study, 95 (49%) into standard care (control group) and 97 (51%) into a multidisciplinary diabetes program (intervention group). The 30-day overall hospital readmission rates (including both emergency department and hospital readmissions) were 19% in the control group and 7% in the intervention group (P = .02). The 365-day overall hospital readmission rates were 38% in the control group and 14% in the intervention group (P = .0002). CONCLUSION: Patients with diabetes who are assigned to a specialized multidisciplinary diabetes program upon their discharge exhibit significantly reduced hospital readmission rates at 30 days and 365 days after discharge.

Belimumab promotes negative selection of activated autoreactive B cells in systemic lupus erythematosus patients.

Belimumab has therapeutic benefit in active systemic lupus erythematosus (SLE), especially in patients with high-titer anti-dsDNA antibodies. We asked whether the profound B cell loss in belimumab-treated SLE patients is accompanied by shifts in the immunoglobulin repertoire. We enrolled 15 patients who had been continuously treated with belimumab for more than 7 years, 17 matched controls, and 5 patients who were studied before and after drug initiation. VH genes of sort-purified mature B cells and plasmablasts were subjected to next-generation sequencing. We found that B cell-activating factor (BAFF) regulates the transitional B cell checkpoint, with conservation of transitional 1 (T1) cells and approximately 90% loss of T3 and naive B cells after chronic belimumab treatment. Class-switched memory B cells, B1 B cells, and plasmablasts were also substantially depleted. Next-generation sequencing revealed no redistribution of VH, DH, or JH family usage and no effect of belimumab on representation of the autoreactive VH4-34 gene or CDR3 composition in unmutated IgM sequences, suggesting a minimal effect on selection of the naive B cell repertoire. Interestingly, a significantly greater loss of VH4-34 was observed among mutated IgM and plasmablast sequences in chronic belimumab-treated subjects than in controls, suggesting that belimumab promotes negative selection of activated autoreactive B cells.

Signalling via the osteopontin and high mobility group box-1 axis drives the fibrogenic response to liver injury.

OBJECTIVE: Liver fibrosis is associated with significant collagen-I deposition largely produced by activated hepatic stellate cells (HSCs); yet, the link between hepatocyte damage and the HSC profibrogenic response remains unclear. Here we show significant induction of osteopontin (OPN) and high-mobility group box-1 (HMGB1) in liver fibrosis. Since OPN was identified as upstream of HMGB1, we hypothesised that OPN could participate in the pathogenesis of liver fibrosis by increasing HMGB1 to upregulate collagen-I expression. DESIGN AND RESULTS: Patients with long-term hepatitis C virus (HCV) progressing in disease stage displayed enhanced hepatic OPN and HMGB1 immunostaining, which correlated with fibrosis stage, whereas it remained similar in non-progressors. Hepatocyte cytoplasmic OPN and HMGB1 expression was significant while loss of nuclear HMGB1 occurred in patients with HCV-induced fibrosis compared with healthy explants. Well-established liver fibrosis along with marked induction of HMGB1 occurred in CCl4-injected OpnHep transgenic yet it was less in wild type and almost absent in Opn-/- mice. Hmgb1 ablation in hepatocytes (Hmgb1ΔHep) protected mice from CCl4-induced liver fibrosis. Coculture with hepatocytes that secrete OPN plus HMGB1 and challenge with recombinant OPN (rOPN) or HMGB1 (rHMGB1) enhanced collagen-I expression in HSCs, which was blunted by neutralising antibodies (Abs) and by Opn or Hmgb1 ablation. rOPN induced acetylation of HMGB1 in HSCs due to increased NADPH oxidase activity and the associated decrease in histone deacetylases 1/2 leading to upregulation of collagen-I. Last, rHMGB1 signalled via receptor for advanced glycation end-products and activated the PI3K-pAkt1/2/3 pathway to upregulate collagen-I. CONCLUSIONS: During liver fibrosis, the increase in OPN induces HMGB1, which acts as a downstream alarmin driving collagen-I synthesis in HSCs.

High mobility group box-1 (HMGB1) participates in the pathogenesis of alcoholic liver disease (ALD).

Growing clinical and experimental evidence suggests that sterile inflammation contributes to alcoholic liver disease (ALD). High mobility group box-1 (HMGB1) is highly induced during liver injury; however, a link between this alarmin and ALD has not been established. Thus, the aim of this work was to determine whether HMGB1 contributes to the pathogenesis of ALD. Liver biopsies from patients with ALD showed a robust increase in HMGB1 expression and translocation, which correlated with disease stage, compared with healthy explants. Similar findings were observed in chronic ethanol-fed wild-type (WT) mice. Using primary cell culture, we validated the ability of hepatocytes from ethanol-fed mice to secrete a large amount of HMGB1. Secretion was time- and dose-dependent and responsive to prooxidants and antioxidants. Selective ablation of Hmgb1 in hepatocytes protected mice from alcohol-induced liver injury due to increased carnitine palmitoyltransferase-1, phosphorylated 5'AMP-activated protein kinase-α, and phosphorylated peroxisome proliferator-activated receptor-α expression along with elevated LDL plus VLDL export. Native and post-translationally modified HMGB1 were detected in humans and mice with ALD. In liver and serum from control mice and in serum from healthy volunteers, the lysine residues within the peptides containing nuclear localization signals (NLSs) 1 and 2 were non-acetylated, and all cysteine residues were reduced. However, in livers from ethanol-fed mice, in addition to all thiol/non-acetylated isoforms of HMGB1, we observed acetylated NLS1 and NLS2, a unique phosphorylation site in serine 35, and an increase in oxidation of HMGB1 to the disulfide isoform. In serum from ethanol-fed mice and from patients with ALD, there was disulfide-bonded hyperacetylated HMGB1, disulfide-bonded non-acetylated HMGB1, and HMGB1 phosphorylated in serine 35. Hepatocytes appeared to be a major source of these HMGB1 isoforms. Thus, hepatocyte HMGB1 participates in the pathogenesis of ALD and undergoes post-translational modifications (PTMs) that could condition its toxic effects.

Osteopontin induces ductular reaction contributing to liver fibrosis.

OBJECTIVE: In human chronic liver disease, there is association between ductular reaction (DR) and fibrosis; yet, the mechanism triggering its onset and its role in scar formation remains unknown. Since we previously showed that osteopontin (OPN) is highly induced during drug-induced liver fibrosis, we hypothesised that OPN could drive oval cells (OC) expansion and DR and signal to hepatic stellate cells (HSC) to promote scarring. RESULTS: In vivo studies demonstrated increased OPN expression in biliary epithelial cells (BEC) and in OC in thioacetamide (TAA)-treated mice. OPN ablation protected mice from TAA and bile duct ligation-induced liver injury, DR and scarring. This was associated with greater hepatocyte proliferation, lower OC expansion and DR along with less fibrosis, suggesting that OPN could activate the OC compartment to differentiate into BEC, which could then signal to HSC to enhance scarring. Since TAA-treated wild-type mice and cirrhotic patients showed TGF-ß(+) BEC, which were lacking in TAA-treated Opn(-/-) mice and in healthy human explants, this suggested that OPN could regulate TGF-ß, a profibrogenic factor. In vitro experiments confirmed that recombinant OPN (rOPN) decreases hepatocyte proliferation and increases OC and BEC proliferation. To evaluate how BEC regulate collagen-I production in HSC, co-cultures were established. Co-cultured BEC upregulated OPN and TGF-ß expression and enhanced collagen-I synthesis by HSC. Lastly, recombinant TGF-ß (rTGFß) and rOPN promoted BEC proliferation and neutralisation of OPN and TGF-ß reduced collagen-I expression in co-cultured HSC. CONCLUSIONS: OPN emerges as a key matricellular protein driving DR and contributing to scarring and liver fibrosis via TGF-ß.

Osteopontin binding to lipopolysaccharide lowers tumor necrosis factor-α and prevents early alcohol-induced liver injury in mice.

UNLABELLED: Although osteopontin (OPN) is induced in alcoholic patients, its role in the pathophysiology of alcoholic liver disease (ALD) remains unclear. Increased translocation of lipopolysaccharide (LPS) from the gut is key for the onset of ALD because it promotes macrophage infiltration and activation, tumor necrosis factor-α (TNFα) production, and liver injury. Since OPN is protective for the intestinal mucosa, we postulated that enhancing OPN expression in the liver and consequently in the blood and/or in the gut could protect from early alcohol-induced liver injury. Wild-type (WT), OPN knockout (Opn(-/-)), and transgenic mice overexpressing OPN in hepatocytes (Opn(HEP) Tg) were fed either the control or the ethanol Lieber-DeCarli diet. Ethanol increased hepatic, plasma, biliary, and fecal OPN more in Opn(HEP) Tg than in WT mice. Steatosis was less in ethanol-treated Opn(HEP) Tg mice as shown by decreased liver-to-body weight ratio, hepatic triglycerides, the steatosis score, oil red-O staining, and lipid peroxidation. There was also less inflammation and liver injury as demonstrated by lower alanine aminotransferase (ALT) activity, hepatocyte ballooning degeneration, LPS levels, the inflammation score, and the number of macrophages and TNFα(+) cells. To establish if OPN could limit LPS availability and its noxious effects in the liver, binding studies were performed. OPN showed binding affinity for LPS which prevented macrophage activation, reactive oxygen, and nitrogen species generation and TNFα production. Treatment with milk OPN (m-OPN) blocked LPS translocation in vivo and protected from early alcohol-induced liver injury. CONCLUSION: Natural induction plus forced overexpression of OPN in the liver or treatment with m-OPN protect from early alcohol-induced liver injury by blocking the gut-derived LPS and TNFα effects in the liver.

Osteopontin delays resolution of liver fibrosis.

To date, considerable progress has been made both in the mechanisms driving liver fibrosis and in the prevention of disease progression. Resolution of liver fibrosis is an emerging field in hepatology; yet, the mediators involved remain elusive. Earlier work from our laboratory demonstrated that the matricellular cytokine osteopontin (OPN) is pro-fibrogenic by promoting hepatic stellate cell (HSC) activation and extracellular matrix (ECM) deposition in vitro and in vivo and specifically by governing fibrillar collagen-I expression, the key pro-fibrogenic protein. Here we hypothesized that OPN could also delay the resolution of liver fibrosis by sustaining collagen-I synthesis or by preventing its degradation. To demonstrate this, wild-type (WT) and OPN-knockout (Opn(-/-)) mice were administered thioacetamide (TAA) in the drinking water for 4 months. Half of the mice were killed at 4 months to assess the extent of fibrosis at the peak of injury, and the rest of the mice were killed 2 months after TAA withdrawal to determine the rate of fibrosis resolution. Following TAA cessation, livers from Opn(-/-) mice showed no centrilobular and parenchymal necrosis along with faster ECM remodeling than WT mice. The latter was quantified by less fibrillar collagen-I immunostaining. Western blot analysis demonstrated a significant decrease in fibrillar collagen-I and in tissue inhibitor of metalloproteinase-1 (TIMP-1) in Opn(-/-) mice undergoing fibrosis resolution compared with WT mice. In conclusion, these results suggest that OPN delays liver fibrosis resolution due to sustained fibrillar collagen-I deposition; hence, inhibiting OPN could be an effective therapeutic strategy for resolving liver fibrosis.

Milk osteopontin, a nutritional approach to prevent alcohol-induced liver injury.

Alcohol consumption is a leading cause of liver disease worldwide; thus, there is an urgent need to develop novel therapeutic interventions. Key events for the onset and progression of alcoholic liver disease result in part from the gut-to-liver interaction. Osteopontin is a cytokine present at high concentration in human milk, umbilical cord, and infants' plasma with beneficial potential. We hypothesized that dietary administration of milk osteopontin could prevent alcohol-induced liver injury perhaps by maintaining gut integrity and averting hepatic inflammation and steatosis. Wild-type mice were fed either the control or the ethanol Lieber-DeCarli diets alone or in combination with milk osteopontin for 3 wk, and parameters of gut and liver damage were measured. Milk osteopontin protected the stomach and the gut by increasing gland height, crypt cell plus enterocyte proliferation, and mucin content in addition to lowering macrophages, plasmacytes, lymphocytes, and neutrophils in the mucosa and submucosa in alcohol-fed mice. Milk osteopontin targeted the gut-liver axis, preserving the expression of tight-junction proteins in alcohol-fed mice thus maintaining intestinal integrity and permeability. There was protection from liver injury since transaminases, the activity scores, triglyceride levels, neutrophil infiltration, 3-nitrotyrosine residues, lipid peroxidation end products, translocation of gram-negative bacteria, lipopolysaccharide levels, and tumor necrosis factor-α were lower in cotreated than in ethanol-fed mice. Furthermore, milk osteopontin diminished ethanol-mediated liver injury in OPN knockout mice. Milk osteopontin could be a simple effective nutritional therapeutic strategy to prevent alcohol hepatotoxicity due, among others, to gut protective, anti-inflammatory, and anti-steatotic actions.

CYP2E1 and oxidant stress in alcoholic and non-alcoholic fatty liver disease.

Alcoholic (ALD) and non-alcoholic fatty liver diseases (NAFLD) are clinical conditions leading to hepatocellular injury and inflammation resulting from alcohol consumption, high fat diet, obesity and diabetes, among others. Oxidant stress is a major contributing factor to the pathogenesis of ALD and NAFLD. Multiple studies have shown that generation of reactive oxygen species (ROS) is key for the progression of fatty liver to steatohepatitis. Cytochrome P450 2E1 (CYP2E1) plays a critical role in ROS generation and CYP2E1 is also induced by alcohol itself. This review summarizes the role of CYP2E1 in ALD and NAFLD.

Osteopontin, an oxidant stress sensitive cytokine, up-regulates collagen-I via integrin α(V)ß(3) engagement and PI3K/pAkt/NFκB signaling.

UNLABELLED: A key feature in the pathogenesis of liver fibrosis is fibrillar Collagen-I deposition; yet, mediators that could be key therapeutic targets remain elusive. We hypothesized that osteopontin (OPN), an extracellular matrix (ECM) cytokine expressed in hepatic stellate cells (HSCs), could drive fibrogenesis by modulating the HSC pro-fibrogenic phenotype and Collagen-I expression. Recombinant OPN (rOPN) up-regulated Collagen-I protein in primary HSCs in a transforming growth factor beta (TGFß)-independent fashion, whereas it down-regulated matrix metalloprotease-13 (MMP13), thus favoring scarring. rOPN activated primary HSCs, confirmed by increased α-smooth muscle actin (αSMA) expression and enhanced their invasive and wound-healing potential. HSCs isolated from wild-type (WT) mice were more profibrogenic than those from OPN knockout (Opn(-/-)) mice and infection of primary HSCs with an Ad-OPN increased Collagen-I, indicating correlation between both proteins. OPN induction of Collagen-I occurred via integrin α(v)ß(3) engagement and activation of the phosphoinositide 3-kinase/phosphorylated Akt/nuclear factor kappa B (PI3K/pAkt/NFκB)-signaling pathway, whereas cluster of differentiation 44 (CD44) binding and mammalian target of rapamycin/70-kDa ribosomal protein S6 kinase (mTOR/p70S6K) were not involved. Neutralization of integrin α(v) ß(3) prevented the OPN-mediated activation of the PI3K/pAkt/NFκB-signaling cascade and Collagen-I up-regulation. Likewise, inhibition of PI3K and NFκB blocked the OPN-mediated Collagen-I increase. Hepatitis C Virus (HCV) cirrhotic patients showed coinduction of Collagen-I and cleaved OPN compared to healthy individuals. Acute and chronic liver injury by CCl(4) injection or thioacetamide (TAA) treatment elevated OPN expression. Reactive oxygen species up-regulated OPN in vitro and in vivo and antioxidants prevented this effect. Transgenic mice overexpressing OPN in hepatocytes (Opn(HEP) Tg) mice developed spontaneous liver fibrosis compared to WT mice. Last, chronic CCl(4) injection and TAA treatment caused more liver fibrosis to WT than to Opn(-/-) mice and the reverse occurred in Opn(HEP) Tg mice. CONCLUSION: OPN emerges as a key cytokine within the ECM protein network driving the increase in Collagen-I protein contributing to scarring and liver fibrosis.

Role of nitric oxide in the regulation of fibrogenic factors in experimental liver fibrosis in mice.

Previously, we have shown that an increased expression level of iNOS but a reduction in the expression of eNOS is associated with increased oxidative stress markers in CCl4-induced experimental liver fibrosis. The present study aimed to investigate the effect of L-arginine and 5-methylisothiourea hemisulfate (SMT) in the expression of profibrogenic factors in chronic liver injury. ICR mice were treated with CCl4 with or without treatment of L-arginine, an NO donor, or SMT, an iNOS inhibitor. The expression of matrix metalloptroteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs), α-smooth muscle actin (α-SMA), tumor necrosis factor-α (TNF-α) and cyclooxygenase-2 (COX-2) were investigated by RT-PCR. The activity of the MMP-2 and MMP-9 were measured by zymography. Our results showed that CCl4-treated mice showed significant up-regulation of expression of pro-fibrogenic factors, TNF-α and COX-2. Treatment with L-arginine or SMT showed a significant reduction in CCl4-induced expression of these pro-fibrogenic factors, TNF-α and COX-2. In conclusion, both SMT and L-arginine effectively attenuated the progression of CCl4-induced liver fibrosis. SMT suppresses iNOS mediated NO production. However, L-arginine augments NO production. The similar effect of the two drugs on liver fibrosis indicates that there may be two distinct pathways of NOS mediated fibrogenesis in chronic liver injury by iNOS and eNOS. Our results suggest that eNOS-mediated liver fibrogenesis may play a more important role than that of iNOS in chronic liver injury. Taken together, these results support the contention that NO plays an active role in the progression of liver fibrosis and hepatocellular damage.

Endothelial nitric oxide synthase is a critical factor in experimental liver fibrosis.

Reduced expression of endothelial nitric oxide synthase (eNOS) in chronic liver disease can reduce hepatic perfusion and accelerate fibrosis. The relationship between eNOS expression and liver fibrogenesis remains unclear. We investigated whether L-arginine attenuated chronic liver fibrosis through eNOS expression. Chronic liver injury was induced by administration of carbon tetrachloride (CCl(4)) to mice for 8 weeks. 5-Methylisothiourea hemisulphate (SMT), an iNOS inhibitor, or L-arginine, a NOS substrate were injected subcutaneously. CCl(4)-induced hepatotoxicity, oxidative stress and accumulation of collagen were detected in the liver. The expression levels of inducible NOS (iNOS) and nuclear factor kappa-B (NF-kappaB) activity in the liver after CCl(4) treatment were increased but eNOS expression and activator protein-1 (AP-1) activity were decreased. Both SMT and L-arginine effectively reduced CCl(4) induced oxidative stress and collagen formation, but L-arginine showed a significantly greater suppression of collagen formation, iNOS expression and NF-kappaB activity. L-arginine also restored the level of eNOS and AP-1 activity. L-arginine was more effective than SMT in suppressing liver fibrosis. L-arginine might improve NO production which facilitates hepatic blood flow and thus retards liver fibrogenesis. Our results showed that the reduced eNOS expression in CCl(4)-treated mice was reversed by L-arginine. Furthermore, L-arginine also reversed the reduced AP-1 activity, an eNOS promoter.

A voluntary oral-feeding rat model for pathological alcoholic liver injury.

The variety of animal models used in the study of alcoholic liver disease reflects the formidable task of developing a model that replicates the human disease. We show that oral feeding of fatty acids derived from fish oil and ethanol induces fatty liver, necrosis, inflammation, and fibrosis. Together with the study of oxidative and nitrosative stress markers, cytokines, proteasome function, and protein studies, this model has provided an inexpensive and technically simple method of establishing pathological alcoholic liver injury.

Green tea polyphenols as an anti-oxidant and anti-inflammatory agent for cardiovascular protection.

Our review aims to examine the cellular and molecular mechanisms of cardiovascular protection of green tea polyphenols, particularly epigallocatechin gallate (EGCG), which focuses on the anti-oxidative and anti-inflammatory effects. EGCG is the major and the most active component in green tea. Studies have shown that EGCG protects cellular damage by inhibiting DNA damage and oxidation of LDL. One of the protective properties of EGCG is its ability to scavenge free radicals. EGCG can also reduce the inflammatory response associated with local tissue injuries such as the hepatocellular necrosis in acute liver injury induced by carbon tetrachloride. The protective effect of EGCG is due to its ability to decrease lipid peroxidation, oxidative stress and the production of nitric oxide (NO) radicals by inhibiting the expression of iNOS. EGCG also ameliorates the overproduction of pro-inflammatory cytokines and mediators, reduces the activity of NF-kappaB and AP-1 and the subsequent formation of peroxynitrite with NO and reactive oxygen species. Thus, EGCG effectively mitigates cellular damage by lowering the inflammatory reaction and reducing the lipid peroxidation and NO generated radicals leading to the oxidative stress. Green tea is proposed to be a dietary supplement in the prevention of cardiovascular diseases in which oxidative stress and proinflammation are the principal causes.