In the present study, we assessed the clinical value of circulating tumor cells (CTC) with stem-like phenotypes for diagnosis, prognosis, and surveillance in hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) by an optimized qPCR-based detection platform. Differing subsets of CTCs were investigated, and a multimarker diagnostic CTC panel was constructed in a multicenter patient study with independent validation (total = 1,006), including healthy individuals and patients with chronic hepatitis B infection (CHB), liver cirrhosis (LC), benign hepatic lesion (BHL), and HBV-related HCC, with area under the receiver operating characteristic curve (AUC-ROC) reflecting diagnostic accuracy. The role of the CTC panel in treatment response surveillance and its prognostic significance were further investigated. The AUC of the CTC panel was 0.88 in the training set [sensitivity = 72.5%, specificity = 95.0%, positive predictive value (PPV) = 92.4, negative predictive value (NPV) = 77.8] and 0.93 in the validation set (sensitivity = 82.1%, specificity = 94.2%, PPV = 89.9, NPV = 89.3). This panel performed equally well in detecting early-stage and α-fetoprotein-negative HCC, as well as differentiating HCC from CHB, LC, and BHL. The CTC load was decreased significantly after tumor resection, and patients with persistently high CTC load showed a propensity of tumor recurrence after surgery. The prognostic significance of the CTC panel in predicting tumor recurrence was further confirmed [training: HR = 2.692; 95% confidence interval (CI), 1.617-4.483; < 0.001; and validation: HR = 3.127; 95% CI, 1.360-7.190; = 0.007]. Our CTC panel showed high sensitivity and specificity in HCC diagnosis and could be a real-time parameter for risk prediction and treatment monitoring, enabling early decision-making to tailor effective antitumor strategies. .

We recently showed that lovastatin attenuates cyclosporin A (CsA)-induced damage of cortical collecting duct (CCD) principal cells by reducing intracellular cholesterol. Previous studies showed that, in cell expression models or artificial membranes, exogenous cholesterol directly inhibits inward rectifier potassium channels, including Kir1.1 (Kcnj1; the gene locus for renal outer medullary K + [ROMK1] channels). Therefore, we hypothesized that lovastatin might stimulate ROMK1 by reducing cholesterol in CCD cells. Western blots showed that mpkCCD c14 cells express ROMK1 channels with molecular masses that approximate the molecular masses of ROMK1 in renal tubules detected before and after treatment with DTT. Confocal microscopy showed that ROMK1 channels were not in the microvilli, where cholesterolrich lipid rafts are located, but rather, the planar regions of the apical membrane of mpkCCD c14 cells. Furthermore, phosphatidylinositol-4,5-bisphosphate [PI(4,5)P 2 ], an activator of ROMK channels, was detected mainly in the microvilli under resting conditions along with the kinase responsible for PI(4,5) P 2 synthesis, phosphatidylinositol-4-phosphate 5-kinase, type I g [PI(4)P5K I g], which may explain the low basal open probability and increased sensitivity to tetraethylammonium observed here for this channel. Notably, lovastatin induced PI(4)P5K I g diffusion into planar regions and elevated PI(4,5)P 2 and ROMK1 open probability in these regions through a cholesterol-associated mechanism. However, exogenous cholesterol alone did not induce these effects. These results suggest that lovastatin stimulates ROMK1 channels, at least in part, by inducing PI(4,5)P 2 synthesis in planar regions of the renal CCD cell apical membrane, suggesting that lovastatin could reduce cyclosporin-induced nephropathy and associated hyperkalemia.

Heat shock protein 90 (HSP90) is a ubiquitous molecular chaperone involved in the proper conformation of many proteins. HSP90 inhibitors (17-dimethyl aminoethylamino-17-demethoxygeldanamycin hydrochloride [17-DMAG]) bind to and inactivate HSP90, suppressing some key signaling pathways involved in the inflammatory process. Since considerable evidence suggests that inflammation accounts for the progression of cerebral ischemic injury, we investigated whether 17-DMAG can modulate inflammatory responses in middle cerebral artery occluded (MCAO) mice. Male C57/BL6 mice were pretreated with 17-DMAG or vehicle for 7 d before being subjected to transient occlusion of middle cerebral artery and reperfusion. Mice were evaluated at 24 h after MCAO for neurological deficit scoring. Moreover, the mechanism of the anti-inflammatory effect of 17-DMAG was investigated with a focus on nuclear factor kappa B (NF-κB) pathway. 17-DMAG significantly reduced cerebral infarction and improved neurological outcome. 17-DMAG suppressed activation of microglia and decreased phosphorylation of inhibitory (I)κB and subsequent nuclear translocation of p65, which eventually downregulated expression of NF-κB-regulated genes. These results suggest that 17-DMAG has a promising therapeutic effect in ischemic stroke treatment through an anti-inflammatory mechanism.Key words heat shock protein 90 (HSP90); stroke; inflammation Acute ischemic stroke is one of the most frequent causes of death and permanent disability in adults worldwide.1) Although various mechanisms are involved in the pathogenesis of stroke, recent work has demonstrated that inflammatory responses accompanying necrotic brain injury contribute to ischemic pathology, and anti-inflammatory strategies have become popular.2) Nuclear factor-kappa B (NF-κB) is a central regulator of inflammatory response in ischemic stroke, whose activation is required for the transcriptional induction of many proinflammatory mediators involved in innate immunity, such as cellular adhesion molecules, cytokines, and growth factors.3) Indeed, it has been evidenced that factors that modulate the activity of NF-κB could potentially regulate inflammatory processes in ischemic stroke. 4)Heat shock protein 90 (HSP90) is a ubiquitous molecular chaperone which plays essential roles in the folding, activation, and assembly of client proteins. Blocking the ATP-binding site of HSP90 by inhibitors, results in degradation of some client proteins (e.g. inhibitory (I)κB kinase (IKK)) via an ubiquitin-proteasome-dependent pathway.6) Previous studies showed that inhibition of HSP90 by injections of geldanamycin (GA, an HSP90 inhibitor) into the cerebral ventricles protects ischemic stroke through stimulation of heat shock gene transcription (such as HSP70 and HSF1).7) 17-Dimethylaminoethylamino-17-demethoxygeldanamycin hydrochloride (17-DMAG) is a more water soluble HSP90 inhibitor and can be administered orally. However little is known about anti-inflammatory effect of HSP90 inhibitor during ischemic stroke and whether 17...