Circulating tumor cells (CTCs) serves a primary function in metastasis and recurrence of hepatocellular carcinoma (HCC). In the present study, in order to evaluate the analytical performance and clinical value of the liquid biopsy-based platform, a novel integrated subtraction enrichment and immunostaining-fluorescence in situ hybridization (iFISH®) platform was applied to analyze CTCs in patients with HCC undergoing liver transplantation (LT). In total, 30 patients with HCC undergoing LT and 10 healthy volunteers were enrolled. CTCs in peripheral blood that were obtained from each patient prior to LT and 3 months thereafter were detected using the iFISH® platform, and CellSearch® system was performed for each subject for comparison. Using iFISH® and CellSearch®, the percentage of CTCs in patients with pre-operative HCC were 70.00% and 26.67%, respectively. CTCs counted using iFISH® (iFISH-CTCs) were increased compared with CellSearch® (Cellsearch-CTCs) (P<0.01). A significant decrease in iFISH-CTCs was observed 3 months following LT (3.04±0.93/7.5 to 1.0±0.53/7.5 ml, P<0.05). Furthermore, patients with lower preoperative iFISH-CTCs level (<5/7.5 ml) had markedly increased recurrence-free survival compared with iFISH-CTCs (>5/7.5 ml, 15 vs. 5.5 months; P<0.01. iFISH® platform exhibits an increased analytical sensitivity, and may be used as a dynamic monitoring tool for CTCs, and CTCs may be a good prognostic indicator for patients with HCC undergoing LT.
Background & Aims
Sustained c-Jun N-terminal kinase (JNK) activation plays a major role in drug-induced liver injury (DILI). Stress-responsive microRNA-31 (miR-31) has been implicated in regulating different cellular damage, and JNK activation could induce miR-31 expression. However, the regulatory role of miR-31 in DILI has not been studied previously. We aimed to investigate whether miR-31 could ameliorate DILI and ascertain potential molecular mechanism.
miR-31 gene knockout (31-KO) and wild-type
mice were used to construct an acetaminophen (APAP)-induced DILI model. Primary mouse hepatocytes, as well as alpha mouse liver 12 (AML-12) cell lines, were used for in vitro experiments. Argonaute 2–associated RNA immunoprecipitation combined with high-throughput sequencing were performed to identify specific targets of miR-31.
31-KO mice showed a higher mortality rate, liver transaminase levels, and hepatic necrosis compared with those in wild-type mice after APAP-induced hepatotoxicity. The protective role of miR-31 on hepatocytes has been analyzed via constructing bone marrow chimeric mice. Mechanistically, we found that hepatic JNK phosphorylation increased significantly in 31-KO mice. This caused mitochondrial phosphorylated Src (p-Src) inactivation and more reactive oxygen species production, which directly amplifies hepatocyte necrotic cell death, while administration of JNK-specific inhibitor SP600125 could abrogate the differences. Moreover, bioinformatics analysis of RNA immunoprecipitation combined with high-throughput sequencing identified that guanosine triphosphatase, cell division cycle protein 42 (Cdc42), the upstream molecule of JNK signaling, was the specific target of miR-31 and could form a miR-31/Cdc42/phosphorylated mixed-lineage kinase 3 (p-MLK3) negative feedback loop to restrict JNK overactivation. Clinically, both miR-31 and phosphorylated JNK (p-JNK) were highly increased in liver tissues of DILI patients with different etiologies.
miR-31 can down-regulate Cdc42 to restrict overactivation of reactive oxygen species/JNK/mitochondria necrotic death loop in hepatocytes of APAP-induced DILI, which might provide a new therapeutic target for alleviating JNK overactivation–based liver injury.
The gas velocity and particle velocity in and out of the nozzle have great influence on the coating effect in the process of cold spray. An axis-symmetric two-dimensional mathematical model was presented to study the flow field in and out of cold spray nozzle, and the effect of different pressure, temperature, spraying distance and particle diameter on gas axial velocity and particle velocity were researched. The simulation results showed that the gas axial velocity and particle velocity increased when the gas pressure and temperature were increased, spray distance had little effect on the flow field in the nozzle, and small diameter particle was more easily accelerated.
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