Biogenesis of Rab14-positive endosome buds at Golgi-endosome contacts by the RhoBTB3-SHIP164-Vps26B complex.

Early endosomes (EEs) are crucial in cargo sorting within vesicular trafficking. While cargoes destined for degradation are retained in EEs and eventually transported to lysosomes, recycled cargoes for the plasma membrane (PM) or the Golgi undergo segregation into specialized membrane structures known as EE buds during cargo sorting. Despite this significance, the molecular basis of the membrane expansion during EE bud formation has been poorly understood. In this study, we identify a protein complex comprising SHIP164, an ATPase RhoBTB3, and a retromer subunit Vps26B, which promotes the formation of EE buds at Golgi-EE contacts. Our findings reveal that Vps26B acts as a novel Rab14 effector, and Rab14 activity regulates the association of SHIP164 with EEs. Depletion of SHIP164 leads to enlarged Rab14+ EEs without buds, a phenotype rescued by wild-type SHIP164 but not the lipid transfer-defective mutants. Suppression of RhoBTB3 or Vps26B mirrors the effects of SHIP164 depletion. Together, we propose a lipid transport-dependent pathway mediated by the RhoBTB3-SHIP164-Vps26B complex at Golgi-EE contacts, which is essential for EE budding.

Erratum: MicroRNA-376c-3p Facilitates Human Hepatocellular Carcinoma Progression via Repressing AT-Rich Interaction Domain 2: Erratum.

[This corrects the article DOI: 10.7150/jca.27939.].

A Possible Role of VPS13B in the Formation of Golgi-Lipid Droplet Contacts Associating with the ER.

While the physical interactions between the Golgi apparatus (Golgi) and lipid droplets (LDs) have been suggested through system-level imaging, the Golgi-LD membrane contact sites (MCSs) remain largely uncharacterized. Here, we show evidence to support the existence of Golgi-LD MCSs in HEK293 cells. We further suggest that vacuolar protein sorting-associated protein 13B (VPS13B) localizes to and promotes the formation of Golgi-LD contacts upon oleic acid (OA) stimulation using 3D high-resolution microscopy. Depletion of VPS13B moderately affects the formation of Golgi-LD contacts upon OA treatment in addition to the fragmentation of the Golgi. Although cellular functions of VPS13B-mediated contacts are still elusive, these findings may provide a new insight into related diseases caused by loss-of-function mutations of VPS13B.

Tex2 is required for lysosomal functions at TMEM55-dependent ER membrane contact sites.

ER tubules form and maintain membrane contact sites (MCSs) with late endosomes/lysosomes (LE/lys). The molecular composition and cellular functions of these MCSs are poorly understood. Here, we find that Tex2, an SMP domain-containing lipid transfer protein conserved in metazoan and yeast, is a tubular ER protein and is recruited to ER-LE/lys MCSs by TMEM55, phosphatases that convert PI(4,5)P2 to PI5P on LE/lys. We show that the Tex2-TMEM55 interaction occurs between an N-terminal region of Tex2 and a catalytic motif in the PTase domain of TMEM55. The Tex2-TMEM55 interaction can be regulated by endosome-resident type 2 PI4K activities. Functionally, Tex2 knockout results in defects in lysosomal trafficking, digestive capacity, and lipid composition of LE/lys membranes. Together, our data identify Tex2 as a tubular ER protein that resides at TMEM55-dependent ER-LE/lys MCSs required for lysosomal functions.

Erratum: MicroRNA-376c-3p Facilitates Human Hepatocellular Carcinoma Progression via Repressing AT-Rich Interaction Domain 2: Erratum.

[This corrects the article DOI: 10.7150/jca.27939.].

Long non-coding RNA myocardial infarction associated transcript promotes the proliferation of cholangiocarcinoma cells by targeting miR-551b-3p/CCND1 axis.

Accumulating reports have demonstrated that long non-coding RNAs (lncRNAs) play critical roles in the occurrence and metastasis of cholangiocarcinoma (CCA). LncRNA myocardial infarction associated transcript (MIAT) has been widely reported in hepatocellular carcinoma, pancreatic cancer and colorectal cancer, but the relationship between MIAT and CCA progression has not yet been investigated. In the present study, we found that the expression of MIAT in CCA tissues was prominently higher than that in normal bile duct tissues. Moreover, TCGA-CHOL data in the GEPIA platform further revealed the upregulated expression of MIAT in CCA tissues. Additionally, quantitative real-time PCR results showed that MIAT expression was increased in CCA cell lines compared to the human intrahepatic biliary epithelial cell line. Functionally, MIAT knockdown significantly inhibited cell proliferation and induced G0/G1 phase arrest as well as apoptosis in HuCCT-1 and QBC939 cells. Conversely, ectopic expression of MIAT obviously facilitated the proliferation, cell cycle progression and apoptosis resistance of RBE cells. Mechanistically, MIAT directly interacted with miR-551b-3p and inversely modulated miR-551-3p level in CCA cells. Furthermore, MIAT knockdown reduced the expression of cyclin D1 (CCND1), which was rescued by miR-551b-3p silencing in HuCCT-1 cells. Importantly, CCND1 restoration partially reversed MIAT knockdown-induced proliferation inhibition, G0/G1 phase arrest and apoptosis in HuCCT-1 cells. In conclusion, MIAT was frequently overexpressed in CCA. MIAT contributed to the growth of CCA cells by targeting miR-551b-3p/CCND1 axis.

MicroRNA-551b-3p inhibits tumour growth of human cholangiocarcinoma by targeting Cyclin D1.

MicroRNAs (miRNAs) are powerful regulators in the tumorigenesis of cholangiocarcinoma (CCA). Previous studies report that miR-551b-3p acts as an oncogenic factor in ovarian cancer, but plays a tumour suppressive role in gastric cancer. However, the expression pattern and potential function of miR-551b-3p were still unclear in CCA. Therefore, this study aimed to explore the expression of miR-551b-3p and its role as well as molecular mechanism in CCA. Analysis of TCGA dataset suggested that miR-551b-3p was under-expressed in CCA tissues compared to normal bile duct tissues. Furthermore, our data confirmed the decreased levels of miR-551b-3p in CCA samples and cell lines. Interestingly, TCGA data suggested that low miR-551b-3p level indicated reduced overall survival of CCA patients. Gain- and loss-of-function experiments found that miR-551b-3p inhibited the proliferation, G1-S phase transition and induced apoptosis of CCA cells. In vivo experiments revealed that ectopic expression of miR-551b-3p inhibited tumour growth of CCA in mice. Further investigation demonstrated that miR-551b-3p directly bond to the 3'-UTR of Cyclin D1 (CCND1) mRNA and negatively regulated the abundance of CCND1 in CCA cells. An inverse correlation between miR-551b-3p expression and the level of CCND1 mRNA was detected in CCA tissues from TCGA dataset. Notably, CCND1 knockdown showed similar effects to miR-551b-3p overexpression in HuCCT-1 cells. CCND1 restoration rescued miR-551b-3p-induced inhibition of proliferation, G1 phase arrest and apoptosis in HuCCT-1 cells. In summary, miR-551b-3p inhibits the expression of CCND1 to suppress CCA cell proliferation and induce apoptosis, which may provide a theoretical basis for improving CCA treatment.

MicroRNA-376c-3p Facilitates Human Hepatocellular Carcinoma Progression via Repressing AT-Rich Interaction Domain 2.

Hepatocellular carcinoma (HCC), accounting for approximately 90% of liver cancer, is the most lethal malignant tumors in the world. Large amount of evidence indicate that microRNAs (miRNAs) contribute to the tumorigenesis and progression of HCC. Among them, miR-376c-3p was recently identified as a tumor-related miRNA and is up-regulated in HBV-related HCC. But, the clinical significance of miR-376c-3p and its biological function in HCC progression are still unclear. Here, we confirmed that miR-376c-3p expression level in HCC was markedly higher than that in noncancerous tissues. Up-regulation of miR-376c-3p was detected in four different HCC cell lines. High miR-376c-3p expression correlated with poor prognostic features, such as large tumor size and venous infiltration. Follow-up data indicated that high miR-376c-3p level evidently correlated with poor clinical outcomes of HCC patients. Moreover, knockdown of miR-376c-3p repressed HCC cell growth, migration and invasion in vitro. miR-376c-3p overexpression facilitated these malignant behaviors of Bel-7402 cells. Mechanistically, miR-376c-3p posttranscriptionally repressed ARID2 expression by directly interacting with its 3'-UTR. Furthermore, an obvious negative correlation between miR-376c-3p and ARID2 mRNA expression in HCC tissues was confirmed. Notably, miR-376c-3p knockdown suppressed HCC growth and metastasis in nude mice. Gain-of-function experiments showed that ARID2 inhibited cell growth and mobility of Hep3B cells. Subsequently, ARID2 knockdown rescued miR-376c-3p silencing attenuated Hep3B cell proliferation and mobility. Our results suggest that miR-376c-3p exerts an oncogenic role in HCC progression.

Manipulation of autophagy: a novelly potential therapeutic strategy for retinal neovascularization.

BACKGROUND: The relationship between the role of VEGF and autophagy in the process of retinal angiogenesis is still unclear. In this study, we explored this issue by using the mouse retinal vascular endothelial cell (RVEC) as a model. METHODS: RVECs were divided into the following groups: control, hypoxia (H), 3-methyladenine (3-MA) + H, VEGF + H, 3-MA + VEGF+H, anti-VEGF antibody + H, 3-MA+ anti-VEGF antibody + H. We then examined activation of autophagy by detecting formation of autophagosomes with transmission electron microscopy (TEM) and by counting the number of green fluorescent protein-positive (GFP+) puncta in RVECs. The turnover of microtubule associated protein 1 light chain 3 B (LC3B) and VEGF were examined by western blot. Cell migratory capacity was measured with wound healing assay and transwell assay. The capillary formation assay was performed to investigate the angiogenic capacity. RESULTS: Hypoxia led to an increased number of autophagosome and of the GFP+ puncta, an increased ratio of LC3B-II/I and enhanced migratory and capillary-formation capacities of RVECs. Pre-treatment with 3-MA attenuated activation of autophagy and abrogated the enhanced cell migration and capillary formation under hypoxia. Exposure to VEGF significantly increased migratory and capillary formation capacities of RVECs under hypoxia and 3-MA decreased VEGF-induced angiogenesis without its expression. Formation of autophagosome, the number of GFP+ puncta of RVECs and expression of LC3B-II/I were both elevated in cells treated with anti-VEGF antibody and these effects were partially inhibited by 3-MA pretreatment. CONCLUSION: Our present data may identify autophagic response as a novel target for enhancing the therapeutic efficacy of angiogenesis inhibitors.

microRNA­196b promotes cell migration and invasion by targeting FOXP2 in hepatocellular carcinoma.

Accumulating evidence indicates that microRNAs (miRNAs) play important roles in tumorigenesis and metastasis. Recent research has shown that miR­196b is implicated in metastasis by regulating the migration and invasion of cancer cells. However, the clinical significance of miR­196b and its role as well as the underlying mechanisms in hepatocellular carcinoma (HCC) remain largely unknown. Here, we detected miR­196b expression in HCC and matched non-tumor tissues with qRT­PCR. We found that miR­196b displayed higher expression in HCC patient tissues and cells. Clinical analysis revealed that high miR­196 expression was correlated with venous infiltration, advanced TNM stage and poor prognosis. Functionally, we demonstrated that miR­196b promoted the migration and invasion of HCC cells in vitro. Moreover, miR­196b knockdown restrained pulmonary metastasis in vivo. Mechanistically, we confirmed that miR­196b could directly bind to 3'UTR of forkhead box P2 (FOXP2) mRNA and repress its expression. miR­196b and FOXP2 showed a negative correlation in HCC tissues. More importantly, upregulation of FOXP2 antagonized miR­196b­mediated migration and invasion in Hep3B cells. Furthermore, FOXP2 knockdown partially reversed the anti­metastatic function of the miR­196b inhibitor on HCCLM3 cells. Taken together, we demonstrated that miR­196b may function as a prognostic biomarker and suppressed FOXP2 expression, subsequently leading to the metastasis of HCC. Our findings highlight a novel mechanism of miR­196b in the progression of HCC and identify miR­196b/FOXP2 axis as a promising target for HCC.

Immature colon carcinoma transcript-1 promotes cell growth of hepatocellular carcinoma via facilitating cell cycle progression and apoptosis resistance.

Immature colon carcinoma transcript-1 (ICT1) is a newly identified oncogene, which regulates proliferation, cell cycle progression and apoptosis of cancer cells. However, the clinical significance, biological function and underlying mechanisms of ICT1 in hepatocellular carcinoma (HCC) remain poorly known. In the present study, we showed that the expression of ICT1 in HCC tissues were notably overexpressed compared to corresponding non-tumor tissues. Accordingly, the relative levels of ICT1 were upregulated in HCC cell lines compared with LO2 cells. The positive expression of ICT1 was correlated with large tumor size and advanced TNM tumor stage. Kaplan-Meier plots indicated that ICT1-positive expression in HCC patients showed a prominent shorter survival. In addition, ICT1 knockdown inhibited proliferation and cell cycle progression, and induced apoptosis in HepG2 cells. While, ICT1 overexpression showed opposite effects on these cellular processes of Hep3B cells. In vivo experiments demonstrated that ICT1 deficiency reduced the growth of subcutaneous HCC in nude mice. Notably, ICT1 knockdown reduced the levels of CDK1, cyclin B1 and Bcl-2 and increased the expression of Bax in HepG2 cells. ICT1 overexpression resulted in upregulation of CDK1, cyclin B1 and Bcl-2, and downregulation of Bax in Hep3B cells. Furthermore, microRNA-134 (miR-134) was recognized as a direct upstream regulator and inversely modulated ICT1 abundance in HCC cells. Altogether, our data support that miR-134 regulation of ICT1 facilitates malignant phenotype of HCC cells probably via cell cycle and apoptosis-associated proteins including CDK1, cyclin B1, Bcl-2 and Bax.

SRPX2, an independent prognostic marker, promotes cell migration and invasion in hepatocellular carcinoma.

Sushi repeat-containing protein X-linked 2 (SRPX2), a novel chondroitin sulfate proteoglycan, is overexpressed in human cancer. Recent studies have reported that SRPX2 overexpression is observed in gastrointestinal cancer, and promotes migration and invasion of cancer cells. While, the clinical significance and biological function of SRPX2 remain rarely known in hepatocellular carcinoma (HCC). Here, we found that the levels of SRPX2 in HCC tissues were notably overexpressed compared to non-cancerous specimens. Accordingly, the levels of SRPX2 were obviously up-regulated in HCC cells compared with LO2 cells. The positive expression of SRPX2 was prominently correlated with venous infiltration and advanced TNM tumor stage. Furthermore, SRPX2 expression acted as an independent prognostic marker for HCC patients. SRPX2 knockdown prominently inhibited the invasion and migration of HCCLM3 cells, while SRPX2 restoration enhanced these cellular biological behaviors of Hep3B cells in vitro. Moreover, SRPX2 knockdown suppressed pulmonary metastasis of HCCLM3 cells in nude mice. Mechanically, SRPX2 knockdown reduced the levels of phosphorylated focal adhesion kinase (p-FAK), p-AKT, matrix metallopeptidase 2 (MMP2) and MMP9 in HCCLM3 cells. In turn, SRPX2 overexpression promoted the activation of FAK/AKT pathway and increased MMP2/9 expression in Hep3B cells. Thus, SRPX2 contributes to migration and invasion of HCC cells probably by targeting FAK/AKT pathway-mediated MMP2/9 expression. SRPX2 potentially acts as an independent prognostic predictor and a drug-target for HCC patients.

MicroRNA-33a promotes cell proliferation and inhibits apoptosis by targeting PPARα in human hepatocellular carcinoma.

MicroRNA-33a (miR-33a) is dysregulated in a number of human cancers, where it functions as an oncogenic miRNA. However, the clinical significance of miR-33a and its underlying molecular pathways regarding the progression of hepatocellular carcinoma (HCC) are currently unknown. In the present study, it was observed that the level of miR-33a expression was significantly increased in HCC tissues, relative to adjacent non-tumor tissues. Increased miR-33a expression was significantly correlated with poor prognostic features of HCC, including larger tumor size, higher Edmondson-Steiner grading and higher tumor-node-metastasis tumor stage. Furthermore, high levels of miR-33a expression were associated with decreases in the 5-year overall survival rate and recurrence-free survival of patients with HCC. In addition, functional experiments indicated that overexpression of miR-33a led to increased proliferation and reduced apoptosis of the HCC cell line Huh7, while knockdown of miR-33a decreased proliferation and induced apoptosis in the HCC cell line HepG2. Furthermore, peroxisome proliferator activated receptor alpha (PPARα) was identified as a direct target of miR-33a in HCC. Upregulation of miR-33a was found to reduce the levels of PPARα expression in Huh7 cells, while inhibition of miR-33a lead to a downregulation in PPARα expression in HepG2 cells. Collectively, these results suggest that miR-33a regulates the proliferation and apoptosis of HCC cells, and is a potential prognostic marker of HCC.

An information retrieval system for medical records & documents.

The forms of the medical records are different from one institute to another. Moreover, medical records are always stored in free text. Consequently, medical records almost can not be logically analyzed and understood by machines. In this paper, we have applied the information retrieval (IR) technique in the using of medical records. We have implemented an IR system for the users, such as doctors and patients, to query similar or related medical records to support diagnosis and treatment. Knowledge retrieval for reuse is the key idea of this system.