Pseudocontinuous ASL was collected using 30 pairs of tag and control acquisition using a 3-dimensional gradient-echo spin-echo (GRASE) acquisition. All images were registered to a high-resolution anatomical atlas. Average CBF measurements

within regions of contrast-enhancement and T2 hyperintensity were evaluated between the two modalities. Additionally, voxel-wise correlation between CBF measurements obtained with DSC and ASL were assessed. Results demonstrated a positive linear correlation between DSC and ASL measurements of CBF when regional average values were compared; however, PS-341 ic50 a statistically significant voxel-wise correlation was only observed in around 30-40% of patients. These results suggest DSC and ASL may provide regionally similar, but spatially different measurements of

CBF. Magnetic resonance imaging (MRI) is the mainstay of brain tumor imaging, both in diagnosis and treatment. Traditionally, clinicians rely on contrast enhancement to characterize the relative degree of malignancy in suptratentorial Tanespimycin tumors. However, with increasing evidence for the critical role of angiogenesis in determination of tumor malignancy and growth potential, imaging modalities capable of quantifying cerebral blood flow (CBF) have become attractive alternatives. Several studies have shown that higher grade brain tumors have significantly higher perfusion measurements than low-grade tumors,[1-3] suggesting that CBF measurements may be a better method

for characterizing brain tumor angiogenesis and monitor treatment response. As antiangiogenic therapy is now the standard of care for recurrent malignant gliomas, there is a significant need for monitoring changes in cerebral blood flow within Oxalosuccinic acid areas of suspected tumor independent of contrast enhancement. The gold standard for perfusion MR imaging is dynamic susceptibility contrast (DSC) MRI, which uses a bolus injection of paramagnetic contrast agent, usually gadolinium, as a nondiffusible tracer for CBF. Calculations of CBF, CBV (cerebral blood volume; the fraction of tissue volume occupied by blood), and mean transit time (MTT = CBF/CBV, the time it takes for blood to pass through the vasculature within the tissue of interest) can be made simultaneously. However, this requires deconvolving the arterial input function (AIF) from the time series data. As a result, few studies have been done on the reproducibility of DSC measurements of CBF. Arterial spin labeling (ASL) is a continually evolving noninvasive technique for quantifying CBF. ASL uses magnetically tagged blood water as an endogenous, diffusible tracer for blood flow. Specifically, blood in a feeding artery is subjected to an inversion pulse, and the magnetization can be followed as it is transferred to brain tissue by capillary exchange at a rate dependent on perfusion of the tissue.

Hence, our data support that IL-6 is strongly associated with the severity of liver diseases. CXCL9, CXCL10 and CXCL11 appear to be particularly important in chronic HCV infection by promoting the development of intrahepatic

inflammation that leads to fibrogenesis.[22, 23] These chemokines are also significantly elevated in patients with necroinflammatory activity of acute GSK1120212 nmr and chronic hepatitis C.[24, 25] In our study, serum CXCL9 and CXCL10 were higher in patients with chronic HBV infection than in healthy individuals, which was in agreement with a previous report.[12] Moreover, the serum CXCR3-associated chemokines CXCL9, CXCL10 and CXCL11 were all well correlated with serum values of AST, ALT and bilirubin. Because we observed a

significant correlation between these chemokines and IL-6, our findings suggest that CXCR3-associated chemokines may too contribute to necroinflammatory activity in chronic HBV infection. Rapamycin solubility dmso However, there were insufficient histological data in our study to assess whether IL-6 and CXCR3-associated chemokines were correlated with degree of fibrosis, in addition to a lack of biochemical evidence of inflammation. We furthermore showed a striking negative association between HBsAg concentration and levels of IL-6 and CXCR3-associated chemokines. As HBsAg was also negatively correlated with transaminases and bilirubin, this HBsAg decline may be linked to increased immunological activity. Interestingly, this study demonstrated a beneficial role of IL-22 in achieving a VR during ETV therapy. IL-22

is an IL-10 family cytokine PFKL that is important for the modulation of tissue responses during inflammation and is expressed by many types of lymphocytes of both the innate and adaptive immune systems, most notably T-helper 17 cells, γδ T cells, natural killer cells and lymphoid tissue inducer-like cells. The IL-22 receptor is highly expressed on hepatocytes.[26, 27] At present, several studies support a protective role of IL-22 in the prevention of hepatocellular damage, although there is evidence indicating dual protective and pathogenic roles for this cytokine in the liver.[17, 28-30] Some groups have examined the association between IL-22 and liver fibrosis in humans and mice.[31, 32] In one report, tumor-infiltrating lymphocytes in HCC exhibited elevated IL-22 expression, and these IL-22+ lymphocytes promoted tumor growth and metastasis in mice.[33] Although human patients with chronic hepatitis B show increased percentages of T-helper 17 cells in the peripheral blood and liver and an increased concentration of IL-22 in the serum,[14, 34] there have been no reports on treatment outcome in patients with chronic HBV infection during ETV therapy. In our study, IL-22 levels decreased over time in both the VR and non-VR groups, but they were consistently higher in the VR group.

Methods: In 2 Phase 3 IBS-C trials, patients meeting Rome II IBS-C criteria received once-daily 290-μg linaclotide or placebo. Patients reported daily abdominal pain, discomfort, bloating, fullness, cramping, spontaneous bowel movement [SBM] and complete SBM [CSBM] frequency, stool consistency, and straining. AR of IBS-C symptoms was reported weekly (yes/no). CMC thresholds for these symptoms were estimated using receiver-operating-characteristic methods with AR as an anchor (pooled 12-week trial data). Linaclotide/placebo results were compared using these thresholds to define 12-week responders.

Distribution of agreement between weekly AR and FDA responder criteria (≥30% reduction in worst abdominal pain and increase in CSBMs of ≥1 from baseline) was assessed. Results: AR-based CMC thresholds for abdominal symptom percent improvement from baseline Ganetespib mw ranged from 21.9–33.6%, while change-from-baseline thresholds for improvement in SBMs and CSBMs/week were 2.1 and 0.7, respectively. Responder rates for AR and FDA criteria were

greater in linaclotide vs placebo groups. Analysis of weekly responder rates revealed considerable agreement (≥70%) between AR and FDA responder EPs. These binary responses were reflected by greater rates of clinically meaningful improvement for all abdominal/bowel symptoms for linaclotide NVP-BKM120 solubility dmso vs placebo (all endpoints P < .001). Numbers needed to treat ranged from 5.1–6.4 for abdominal symptoms and 2.4–3.7 for bowel symptoms. Conclusion: Responder analyses based on CMC

thresholds estimated using AR indicated that linaclotide resulted in a higher percentage of patients experiencing clinically meaningful improvement in IBS-C symptoms vs placebo. There was considerable agreement between AR and FDA responder endpoints. Key Word(s): 1. IBS-C; 2. linaclotide; 3. abdominal symptoms; 4. relief; Presenting Author: JOSEISIDRO MINERO Additional Authors: EIRA Edoxaban CERDA Corresponding Author: JOSEISIDRO MINERO Affiliations: HCM Objective: Background. The small intestinal bacterial overgrowth (SIBO) has been considered part of the pathophysiology of irritable bowel syndrome and treatment with rifaximin 400 mg orally every 8 hours for 10 days have been proposed, since it decreased SIBO by 80%. The need of retreatment and the time lapse of it have not been established. Purpose: To determine the time lapse of retreatment with rifaximin in patients with SIBO, evaluated with Global Symptom Scale (GSS), Bristol Scale (BS) and glucose breath test. Methods: Material and Methods: SIBO patients (H breath test positive) that have been treated with rifaximin 400 mg every 8 hours for 10 days were followed for one year, GSS and BS were fulfilled at 6 and 12 months and glucose hydrogen breath tests were performed at 12 months. Patients with relapsing symptoms were treated again with rifaximin 400 mg every 8 hours for 10 days. Results: Results. We evaluated 10 patients.

Indeed, daily administration of Cxcl9

concomitantly to CCl4 strongly inhibited the formation of new blood vessels compared with vehicle-treated mice. JQ1 research buy The difference between Cxcl9 und vehicle-treated mice was evident by quantification of CD31-positive cells (Fig. 6A) as well as vWF-positive cells (Supporting Fig. 7). Furthermore, as determined by contrast-enhanced ultrasound, the microvascular perfusion of the liver was significantly reduced in Cxcl9-treated mice compared with vehicle-treated mice, supporting a reduced density of vessels in the livers of these mice (Fig. 6B). In line with the direct interaction between Cxcl9 and VEGF pathways in vitro, the antiangiogenic properties of Cxcl9 were also linked to a strong decrease in VEGF protein levels within the liver in vivo (Supporting Fig. 8A). Importantly, alongside reduced neoangiogenesis, mice treated with Cxcl9 also had a strongly reduced severity of liver fibrosis compared with vehicle-treated mice (Fig. 7A). This difference was evident after quantification of Sirius red-stained liver tissues (Fig. 7B), biochemical measurement of hepatic hydroxyproline

contents (Fig. 7C), and by assessment of intrahepatic Col1α1 mRNA expression (Supporting Fig. 8B). As Cxcl9 might have direct chemotactic effects on liver infiltrating Afatinib cells, we also determined the number of Th1-polarized, IFN-γ-positive cells in the livers of Cxcl9 and vehicle-treated mice. However, the number of IFN-γ-positive cells was not different between the groups (Supporting Fig. 8C), arguing against a major influence of the immune system on the phenotype observed after Cxcl9 treatment. Instead,

the content of α-SMA in the liver was strongly reduced by Cxcl9 treatment (Fig. 7D), suggesting that a main effect of Cxcl9 in vivo is the modulation of stellate cell activation alongside with reduced neoangiogenesis and endothelial cell inhibition. In the current study we provide evidence that the Cxcr3 chemokine system is an important modulator of neoangiogenesis in the murine liver and that the Cxcr3 ligand Cxcl9 has the potential to ameliorate neoangiogenesis and liver fibrosis aminophylline in vivo. In recent studies we demonstrated that mice deficient in the chemokine receptor Cxcr3 are more prone to liver fibrosis in different experimental models. 7 These results were in line with earlier findings of the importance of Cxcr3 in models of pulmonary and renal fibrosis. 12, 13 The effects of Cxcr3 ligands in liver disease models were mainly explained by reduced recruitment of Th1-polarized or regulatory T cells. 7, 10, 11 Furthermore, a direct inhibitory effect of CXCL9 on collagen secretion of stellate cells was identified. 7 However, another important feature of Cxcr3 ligands is their strong angiostatic function 16, 23 and their close correlations to VEGF concentrations in vivo.

30, 31, 33, 55, 57 In support of our hypothesis, we found MitoQ inhibited the formation 4-HNE protein adduct formation

and 3-NT levels, indicators of the antioxidant action of MitoQ (Figs. 1, 2). The pattern of 4-HNE and 3-NT staining demonstrate a strong gradient extending from the pericentral region deep into the periportal region of the liver and is consistent with similar studies of ethanol-induced liver injury.58 The enhancement of the oxidative/nitrosative mTOR inhibitor stress gradient is linked to several factors including exacerbation of the hypoxic gradient developed in the liver acini and LPS-induced cytokine production in chronic ethanol consumption. MitoQ treatment in LPS-induced inflammation has been shown to involve decreases in proinflammatory cytokines such as interleukin (IL)-1β, IL-6, and IL-8 and an increase in the antiinflammatory cytokine IL-10 levels.59 However, in the present study we found that MitoQ did not significantly change the expression of iNOS, suggesting that its mode of action is downstream of cytokine signaling PDGFR inhibitor (Fig. 2B). Because it has been shown that mitochondria and specifically MitoQ can modify the cellular response to hypoxia, we next examined this pathway.30,

33, 40 Induction of tissue hypoxia and HIF1α in the liver is a hallmark of alcohol-induced liver disease.29, 60 Furthermore, iNOS-derived NO has been shown to inhibit prolyl hydroxylase enzyme activity by competing for the iron(II) in the catalytic site of the enzymes during normoxia and changes mitochondrial function with increased ROS formation.7, 50, 61 Our data are consistent with this literature because we found chronic ethanol-induced HIF1α expression/stabilization Akt inhibitor (Fig. 3A), increased ROS, and increased iNOS (Fig. 2B). MitoQ treatment inhibited ethanol-induced HIF1α expression in the liver (Fig. 3A), whereas iNOS expression remained

unaltered (Fig. 2B). Recently, it has been shown that HIF1α in hepatocytes is a major determinant in the pathogenesis of alcoholic steatosis.29 Taken together, we propose that MitoQ inhibits the mitochondria-dependent induction of HIF1α through suppression of increased mitochondrial ROS in response to NO exposure or damage to the mitochondrion by peroxynitrite. To form peroxynitrite, superoxide must also be formed in response to ethanol consumption and could come from a number of sources, including the mitochondrion or NADPH oxidase.10, 14 Because it has been shown that MitoQ can directly scavenge peroxynitrite, this is a likely mechanism through which activation of HIF1α is prevented.36 Ethanol feeding leads to inhibition of mitochondrial protein synthesis, which is largely responsible for the changes in the activities of the mitochondrial respiratory chain complexes.

Conclusion: Insulin, promotes HepaRG-hepatocyte differentiation and proliferation, however maintained high levels of signaling through this pathway impair maturation and lead to aberrant lipid accumulation. These results may facilitate refinement of current strategies designed to produce mature cells from various progenitor sources in vitro. Disclosures: The following people have nothing to disclose: Luke A. Noon, Alicia MartinezRomero, Jose E. O’Connor, https://www.selleckchem.com/products/forskolin.html Anne Comlu, Pascale Bouillé, Christiane Guillouzco, Deborah J. Burks Aim: To assess the utility of autologous mesenchymal stem cells (MSCs) peripheral vein infusion as a possible therapeutic modality and to confirm the supportive role of the stem

cell (SC) treatment for patients with end-stage liver diseases Methods: Forty patients with post-HCV end-stage liver diseases were randomized into 2 groups. Group 1, comprising 20 patients

and they have received granulocyte colony stimulating factor (GCSF) for 5 days followed by autologous MSC peripheral veins infusion. Group 2, comprising 20 patients and they have received regular liver supportive treatment and have served as Palbociclib ic50 a control group. Results: In the infused group (Gr. I), There was near normalization of liver enzymes and improvement in liver synthetic function (S. Albumin, Prothrombin time and concentration) in 54%. There was significant changes in albumin (p=0.000), bilirubin (p=0.002), INR (p=0.017), prothrombin conc. (p=0.029), AST (p=0.156) and ALT levels (p=0.029). Also, in Gr. I, there Sodium butyrate was stabilization of the clinical and biochemical status in 13% of cases. None of the patients in the control group (Gr. II) showed any significant improvement. Hepatic fibrosis was assessed in the treated group by detection of procollagen Ill C peptide level (PIIICP) (9.4 ± 4.2) and procollagen Ill N peptide level (PIIINP) (440 ± 189) (Pre treatment Value) in the patients’ serum. It was reported a decrease in the level of PIIICP (8.1 ± 2.6) and PIIINP (388 ± 102) after stem cell therapy (three months treatment Value) but they have not reached the significant value (6

months treatment Value) (p=0.7), however there was a significant correlation coefficient after three months between the serum level of the PIIINP and prothrombin concentration (p=-0.5) and between the serum level of the PIIICP and ascites (p=0.550) Conclusion: Our data showed that autologous mesenchymal stem cells infusion into the peripheral veins was effective and showed the same result as intrahepatic infusion from our previous studies (Salama et al, 2011) and confirmed the supportive role of mesenchymal stem cell treatment for end-stage liver disease with satisfactory tolerability and beneficial effects on liver synthetic functions and hepatic fibrosis. We have also observed in this study that the serum albumin has been improved within the first two weeks and prothrombin concentration improvement was delayed for almost one month.

Again, the contextual switch governing DC behavior in diverse states of liver injury remains uncertain. The role of Kupffer cells in APAP-induced liver toxicity is controversial13,

16, 18 but may perhaps be similar to the role of DC. Ju et al.13 showed an increased susceptibility to APAP-induced liver injury when effectively depleting mice of Kupffer cells with gadolinium chloride. The mechanism of the Kupffer hepatoprotective effect is thought learn more to be related to decreased expression of IL-10, an antiinflammatory cytokine, in Kupffer cell-depleted mice. In our current study, DC did not produce detectible IL-10 after APAP treatment and their depletion did not affect serum or liver IL-10 levels (Fig. 3D). Interestingly, Bamboat et al34 demonstrated that in liver ischemia/reperfusion injury, liver DCs were responsible for IL-10 production by way of TLR9 activation. DC-mediated IL-10 production was shown to

suppress monocyte inflammatory function and reduce hepatic injury.34 However, because DC do not produce detectible IL-10 after APAP challenge, it appears that the mechanism responsible for the DC protective role in APAP-induced hepatotoxicity is distinct from that of ischemia/reperfusion liver injury. Our mechanistic investigations further show that the exacerbated liver toxicity associated with DC depletion is independent DNA Damage inhibitor of associated elevations in neutrophils or inflammatory monocytes, as well as independent of systemic elevations in TNF-α, MCP-1, and IL-6 and unrelated to IFN-α (Supporting Fig. 10). Our study implicates DC in APAP-induced liver injury as an innate protector that can be compared to the role of DC in sepsis. Sepsis is characterized by an intense hyperinflammatory response designed to eliminate an underlying infectious source.35 However, there is growing evidence that an initial proinflammatory cascade is thought to be followed by an activation of a compensatory antiinflammatory response syndrome that leads to immune suppression and subsequent poorer clinical outcomes.35, 36 DC loss appears to play a significant role in the pathogenesis of sepsis.37 By using

cecal ligation and puncture (CLP) surgery as a model for sepsis in rodents, recent CYTH4 investigations have shown that a decline in DC counts occur in conjunction with immune dysfunction, suggesting that DC may even have a protective role against the development of immunosuppression in sepsis.37 The mechanism of DC loss appears to be related to extensive apoptosis of DC during sepsis. This was demonstrated in a study in which mouse spleens showed a significant increase in caspase 3-mediated apoptosis in follicular dendritic cells 36-48 hours after CLP insult compared to controls.37, 38 Furthermore, Scumpia et al.39 showed that DC-deficient mice had increased mortality after CLP. Similarly, in our study, DC depletion exacerbated APAP-mediated hepatic injury and led to a significant increase in mortality.

Bacterial translocation and subsequent monocyte accumulation may also stimulate pulmonary angiogenesis in HPS, which may be partly controlled by genetic factors. However, there remains a need for more human experimental data to support the development of new therapies targeting these proposed mechanisms. The presence of HPS should be considered in all patients with liver disease

who complain of dyspnea, which is common in cirrhosis, but which is present in 50% of patients with HPS.[13] A more specific symptom is platypnea (dyspnea that increases from the supine to the erect position), which may be associated check details with orthodoxia (hypoxia that is worse when erect). Finger clubbing is very common in HPS. In one study, it was found in almost 50% of HPS patients compared with 2% in those without the disorder.[54] This striking difference implies that one should always suspect HPS in patients with chronic liver disease and clubbing. Patients with severe HPS may be sufficiently hypoxic to appear cyanosed at rest, and the rare finding of cyanosis and clubbing in a cirrhotic patient is highly suggestive of the presence of severe HPS.[54] Although some studies show that spider nevi are often seen in HPS, there is no major difference in their prevalence in cirrhotics with HPS compared with control cirrhotic patients with

similar liver disease.[13] BAY 73-4506 cell line The diagnosis of HPS depends on establishing that impaired gas exchange in a patient

with liver disease is due to pulmonary vascular dilatation. In most cases, the results of arterial blood gases and a study to detect intrapulmonary shunting (see later) are sufficiently specific to do this once other intrinsic cardiorespiratory diseases are excluded. Pulse oximetry can be a useful monitoring tool in the outpatient setting, and has been proposed as a screening mafosfamide tool for HPS in the cirrhotic population, with a cut-off value of ≤ 97% providing a high sensitivity and moderate specificity for an arterial oxygen tension (PaO2) ≤ 70 mmHg, but is less sensitive in mild HPS.[55, 56] However, in order to confirm the diagnosis, arterial blood gas estimation should be undertaken with the patient in a sitting position, breathing room air. The degree of gas exchange abnormality that is required for the diagnosis of HPS remains controversial. The most sensitive marker is an increase in the alveolar–arterial oxygen gradient (PA-aO2). Recommended cut-off values for the diagnosis of HPS are PaO2 ≤ 80 mmHg or PA-aO2 ≥ 15 mmHg. To avoid a complex calculation to correct for the increase in PA-aO2 that occurs with age, cut-off values of PaO2 ≤ 70 mmHg or PA-aO2 ≥ 20 mmHg are suggested in patients older than 64 years[2] (Table 1). Two methods of defining intrapulmonary dilatation are available: contrast echocardiography, most often using microbubbles as the contrast, and radioactive lung perfusion scan using macroaggregated albumin (MAA).

Goldman et al.[75] showed NO mediated inhibition of the activity of the sodium/hydrogen exchanger that can pump protons out of cells to maintain intracellular pH. Subsequently, intracellular acidification induced NO exposure that led to DNA damage,

toxicity, and neoplastic development. In addition, Sirolimus molecular weight there have been several animal model studies that investigated the involvement of NO in the pathogenesis of esophageal carcinogenesis. Accumulating previous animal model studies demonstrated that administering nitrite with ascorbic acid or other anti-oxidants to rats induced tumors in their forestomach, which is contiguous with the esophagus and lined by squamous epithelium.[76-78] The tumors did not occur if only nitrite was administered. Hence, the combination of nitrite and anti-oxidants generated a high concentration of NO, which had a mutagenic effect on the epithelium.[79] Kuroiwa et al.[80] extended these studies by showing that administration of exogenous NO (sodium nitrite plus ascorbic acid) to an acid-type reflux model of rat induced esophageal squamous cell carcinoma, although the treatment failed to induce the development of Barrett’s esophagus or esophageal adenocarcinoma. This study may indicate that NO alone in the absence of refluxed duodenal

contents is incapable of causing columnar transformation of the esophagus, although click here the treatment might elicit squamous carcinogenesis. In another study, Kumagai et al.[81] demonstrated that thioproline (a nitrite scavenger)

inhibited the development of esophageal adenocarcinoma by gastroduodenal reflux in rats, suggesting the involvement of reactive nitrogen species such as NO, peroxynitrite, and nitroso compounds in the pathogenesis of esophageal adenocarcinoma. Since the 1980s, crotamiton numerous studies have shown that the incidence of esophageal adenocarcinoma has been increasing rapidly in many western countries; however, the precise cause for the cancer endemic remains unclear.[82] In terms of a mass survey, a 20-fold increase in use of chemical nitrogenous fertilizers and associated increased dietary nitrate exposure post-war may be potentially responsible for the marked increase in the incidence of the esophageal adenocarcinoma.[10, 11, 20] Thus far, epidemiological studies have not shown an association of nitrate intake with esophageal adenocarcinoma.[83, 84] Additionally, a recent epidemiological study has demonstrated a paradoxical association between nitrate intake and Barrett’s esophagus depending on gender, that is, total nitrate intake was inversely associated with Barrett’s esophagus in men, while it was positively associated with the pre-malignant condition in women.[85] These studies may suggest that individual susceptibility to esophageal adenocarcinoma does not depend on nitrate intake. However, other co-factors are also closely involved in the chemical reaction of luminal NO generation in the lower esophagus.

00024, Wilcoxon signed-rank test; Fig. 4A). Next, we asked whether DNMT1 mRNA expression was inversely correlated with the levels of miR-152 in HBV-related HCC tissues. Twenty HCCs were analyzed for the expression levels of DNMT1 mRNAs and for miR-152 expression by real-time PCR. A statistically significant inverse correlation was observed between DNMT1 mRNA and miR-152 (n = 20, r = −0.462, P = 0.02, Pearson’s correlation; Fig. 4B). These data showed the reciprocal regulation of Idelalisib manufacturer the tumor suppressor miR-152 and its target DNMT1 in human HCCs and suggested that miR-152 may play a causal role in DNA methylation leading to liver cancer in chronic hepatitis B patients. Because

miR-152 was frequently down-regulated in HBV-positive HCC tissues in comparison with the adjacent noncancerous liver specimens, whereas DNMT1 expression appeared to be inversely correlated, we wondered if the reduction of miR-152 expression selleck chemicals could be driven by the increase in DNMT1 expression. We enhanced or inhibited DNMT1 expression

by transfecting the DNMT1 expression vector (pcDNA3.1-DNMT1) or DNMT1 siRNA into HepG2 and Huh-7 cells with the pcDNA3.1 vector and control siRNA, respectively, as the negative controls (Fig. 5A,B). After 48 hours of transfection, we measured the miR-152 expression levels of these cells and found that none of them were significantly changed in comparison with the negative controls (Fig. 5C,D). We investigated whether the enforced expression of miR-152 could also functionally result in DNA hypermethylation. GDM was measured with an LC-MS/MS method in HepG2 and HepG2.2.15 cell lines after 72 hours of transfection with miR-152 inhibitor or miR-152 mimics, respectively. The miRNA negative control and inhibitor negative control served as negative controls for the mimics and inhibitor, respectively. We observed a reduction of 6.31% to 4.08% in GDM for the HepG2.2.15 cell

line treated with miR-152 mimics in comparison with the negative controls Aprepitant and an increase of 4.55% to 5.88% in GDM for the HepG2 cell line treated with miR-152 inhibitors (Fig. 6A). Several genes have been found to be silenced by DNA methylation induced by HBx via interactions with de novo DNMTs in HBV-HCC, including GSTP1 and CDH1. To assess whether inhibiting the expression of miR-152 could also lead to hypermethylation and silencing of these genes in HCC, we measured the DNA methylation levels of CDH1 and GSTP1 by bisulfate sequencing in the HepG2 cell line after transfection with the miR-152 inhibitor or miRNA inhibitor negative control. The GSTP1 DNA methylation level was increased in HepG2 cells transfected with the miR-152 inhibitor in comparison with the miRNA inhibitor negative control (mean of 85.8% versus 57.4%, P < 0.005, Wilcoxon rank test; Fig. 6C) Likewise, the CDH1 DNA methylation level was increased in HepG2 cells transfected with the miR-152 inhibitor in comparison with the control (mean of 23.8% versus 0%, P < 0.