YAP Inhibits the Apoptosis and Migration of Human Rectal Cancer Cells via Suppression of JNK-Drp1-Mitochondrial Fission-HtrA2/Omi Pathways.

BACKGROUND/AIMS: The Hippo-Yap pathway is associated with tumor development and progression. However, little evidence is available concerning its role in cancer cell apoptosis and migration via mitochondrial homeostasis. Here, we identify mitochondrial fission as a regulator of the Hippo-Yap pathway in human rectal cancer tumorigenesis and metastasis. METHODS: In this study, we performed loss-of function assays concerning Yap in RCC via shRNA. Cellular viability and apoptosis were measured via MTT, the TUNEL assay and trypan blue staining. Mitochondrial function was assessed via JC1 staining, the mPTP opening assay, mitochondrial respiratory function analysis, electron microscopy and immunofluorescence analysis of HtrA2/Omi. Mitophagy and mitochondrial fission were assessed via western blots and immunofluorescence. Cell migration was evaluated via the Transwell assay, wound-healing assay and immunofluorescence analysis of F-actin. The interaction between JNK and Yap was detected via co-immunoprecipitation and Yap recombinant mutagenic plasmid transfection. Western blots were used to analyze signaling pathways in conjunction with JNK inhibitors or HtrA2/Omi siRNA. RESULTS: Yap is upregulated in human rectal cancer cells, where its expression correlates positively with cell survival and migration. Functional studies established that silencing of Yap drove JNK phosphorylation, which induced Drp1 activation and translocation to the surface of mitochondria, initiating mitochondrial fission. Excessive mitochondrial fission mediated HtrA2/Omi leakage from the mitochondria into the cytoplasm, where HtrA2/Omi triggered cellular apoptosis via the mitochondrial apoptosis pathway. Moreover, released HtrA2/Omi also phosphorylated cofilin and inhibited cofilin-mediated F-actin polymerization. F-actin collapse perturbed lamellipodia formation and therefore impaired cellular migration and invasion. CONCLUSION: Collectively, our results demonstrate that Hippo-Yap can serve as a tumor promoter in human rectal cancer and acts by restricting JNK/Drp1/mitochondrial fission/ HtrA2/Omi, with potential implications for new approaches to human rectal cancer therapy.

EHD3 promotes gastric cancer progression via Wnt/ß-catenin/EMT pathway and associates with clinical prognosis and immune infiltration.

Gastric cancer (GC) shows high levels of heterogeneity and predicts a poor prognosis. The expressions of EHD3 are found to be misregulated in a number of tumors. However, the clinical significance and potential function of EHD3 expression in GC patients remain unknown. In this study, we found that EHD3 expression was distinctly increased in GC specimens and cell lines in both TCGA datasets and our cohort. High levels of EHD3 expression were linked to worse outcomes for patients with GC in clinical tests. Nomogram based on multivariate assays displayed good predictive accuracy for GC patients, as evidenced by C-indices and calibration graphs. Low levels of EHD3 mRNA were discovered in GC tissues due to EHD3 methylation's negative regulation of EHD3. In addition, EHD3 was observed to be related to several immune cells and might play a role in successful immunotherapy. Functionally, it was verified that knockdown of EHD3 remarkably suppressed the proliferation, migration and invasion of GC cells in vitro and in vivo. Results of Western blot confirmed that knockdown of EHD3 suppressed the expressions of ß-catenin, MMP-9, and N-cadherin, while promoting the expression of E-cadherin. Overall, this research identified a novel GC-related gene EHD3 which might be a novel prognostic biomarker involved in tumor microenvironment. EHD3 promoted the proliferation and metastasis of GC cells through influencing the Wnt/ß-catenin/EMT signaling pathway, suggesting it as a novel treatment target for GC patients.

Nanosilver-Decorated Biodegradable Mesoporous Organosilica Nanoparticles for GSH-Responsive Gentamicin Release and Synergistic Treatment of Antibiotic-Resistant Bacteria.

PURPOSE: Antibiotic-resistant bacteria are pathogens that have emerged as a serious public health risk. Thus, there is an urgent need to develop a new generation of anti-bacterial materials to kill antibiotic-resistant bacteria. METHODS: Nanosilver-decorated mesoporous organosilica nanoparticles (Ag-MONs) were fabricated for co-delivery of gentamicin (GEN) and nanosilver. After investigating the glutathione (GSH)-responsive matrix degradation and controlled release of both GEN and silver ions, the anti-bacterial activities of Ag-MONs@GEN were systematically determined against several antibiotic-susceptible and antibiotic-resistant bacteria including Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Enterococcus faecalis. Furthermore, the cytotoxic profiles of Ag-MONs@GEN were evaluated. RESULTS: The GEN-loaded nanoplatform (Ag-MONs@GEN) showed glutathione-responsive matrix degradation, resulting in the simultaneous controlled release of GEN and silver ions. Ag-MONs@GEN exhibited excellent anti-bacterial activities than Ag-MONs and GEN alone via inducing ROS generation, especially enhancing synergetic effects against four antibiotic-resistant bacteria including Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Enterococcus faecalis. Moreover, the IC50 values of Ag-MONs@GEN in L929 and HUVECs cells were 313.6 ± 15.9 and 295.7 ± 12.3 µg/mL, respectively, which were much higher than their corresponding minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values. CONCLUSION: Our study advanced the development of Ag-MONs@GEN for the synergistic and safe treatment of antibiotic-resistant bacteria.

Upregulation of lncRNA SNHG1 is associated with metastasis and poor prognosis in cancers: A meta-analysis.

BACKGROUND: Accumulating evidence suggested that the expression level of long noncoding RNA small nucleolar RNA host gene 1 (lncRNA SNHG1) was upregulated in various cancers, and high expression of SNHG1 was associated with metastasis and prognosis in patients with cancer.The relationship between SNHG1 expression and metastasis or prognosis in malignant tumors was investigated in this meta-analysis. METHODS: A systematic search was performed in PubMed, Web of Science, and Cochrane Library from inception until May 31, 2018. Hazard ratio (HR) or odds ratio (OR) with 95% confidence intervals (95% CIs) were calculated to demonstrate prognostic value of SNHG1 using Stata 12.0 software. RESULTS: A total of 10 studies including 1129 patients were finally enrolled in the meta-analysis based on the inclusion and exclusion criteria. Increased SNHG1 expression was significantly associated with lymph node metastasis (OR = 3.28, 95% CI = 2.02-5.33) and advanced TNM stage (OR = 0.26, 95% CI = 0.16-0.43). Moreover, high expression of SNHG1 could predict poor overall survival (HR = 2.32, 95% CI = 1.90-2.83), event-free survival (HR = 1.58, 95% CI = 1.06-2.35), recurrence-free survival (HR = 2.15, 95% CI = 1.23-3.77), progression-free survival (HR = 2.75, 95% CI = 1.70-4.46), and disease-free survival (HR = 1.93, 95% CI = 1.10-3.40) in patients with cancer. CONCLUSION: The present meta-analysis demonstrated that upregulation of lncRNA SNHG1 might serve as a useful prognostic biomarker in various cancers.

Gene expression profiling by mRNA sequencing reveals dysregulation of core genes in Rictor deficient T-ALL mouse model.

T-cell acute lymphoblastic leukemia (T-ALL) is a neoplastic disorder with peak incidence in children and young adults. The mTOR complex is an important component of the PI3K/Akt/mTOR signaling cascade and holds great promise for the treatment of hematopoietic malignancies. Previous studies have shown that the depression of Rictor, one of the components of the mTOR complex, prevents myeloproliferative disorders and leukemia However, knowledge of the progression of mTOR has not greatly improved the prognosis of T-ALL. To identify potential prognostic biomarkers for T-ALL, a whole-genome expression profile of Rictior deficient T-ALL mice was performed. As a result, 1475 differentially expressed genes (DEGs) were identified. Network analysis revealed 46 genes with a high network degree and fold-change value. Kaplan-Meier analysis identified ten crucial genes which significantly associated with survival in Rictor deficient T-ALL mice. These findings provide potential therapeutic targets in leukemia and bear immediate relevance to patients with leukemia.

Long Non-coding RNA ITIH4-AS1 Accelerates the Proliferation and Metastasis of Colorectal Cancer by Activating JAK/STAT3 Signaling.

Accumulating evidence has uncovered long non-coding RNAs (lncRNAs) as central regulators in the pathogenesis of diverse human cancers including colorectal cancer (CRC). The present study discovered that a novel lncRNA ITIH4 antisense RNA 1 (ITHI4-AS1) was frequently under-expressed in most normal human tissues, including colon tissues. Therefore, we aimed to investigate the role of ITHI4-AS1 in CRC. Interestingly, a significant overexpression of ITIH4-AS1 was observed in CRC cell lines relative to normal NCM460 cells. Also, we investigated the facilitating role of ITIH4-AS1 in CRC cell growth and metastasis both in vitro and in vivo. Additionally, we explained that ITIH4-AS1 upregulation in CRC was attributed to downregulation or even depletion of RE1 silencing transcription factor (REST), a presently identified transcriptional repressor for ITIH4-AS1. Meanwhile, the contribution of ITIH4-AS1 to CRC development was validated to rely on the activation of the JAK/STAT3 pathway. More importantly, we verified that FUS interacted with both ITIH4-AS1 and STAT3, and that ITIH4-AS1 evoked nuclear translocation of phosphorylated (p)-STAT3 in CRC through recruiting FUS. In summary, our findings unveiled for the first time that REST downregulation-enhanced ITIH4-AS1 exerts pro-tumor functions in CRC through FUS-dependent activation of the JAK/STAT3 pathway, implying that targeting ITIH4-AS1 may be a novel effective strategy for CRC therapy.

Rictor is required for early B cell development in bone marrow.

The development of early B cells, which are generated from hematopoietic stem cells (HSCs) in a series of well-characterized stages in bone marrow (BM), represents a paradigm for terminal differentiation processes. Akt is primarily regulated by phosphorylation at Thr308 by PDK1 and at Ser473 by mTORC2, and Akt signaling plays a key role in hematopoiesis. However, the role of mTORC2 in the development of early B cells remains poorly understood. In this study, we investigated the functional role of mTORC2 by specifically deleting an integral component, Rictor, in a hematopoietic system. We demonstrated that the deletion of Rictor induced an aberrant increase in the FoxO1 and Rag-1 proteins in BM B cells and that this increase was accompanied by a significant decrease in the abundance of B cells in the peripheral blood (PB) and the spleen, suggesting impaired development of early B cells in adult mouse BM. A BM transplantation assay revealed that the B cell differentiation defect induced by Rictor deletion was not affected by the BM microenvironment, thus indicating a cell-intrinsic mechanism. Furthermore, the knockdown of FoxO1 in Rictor-deleted HSCs and hematopoietic progenitor cells (HPCs) promoted the maturation of B cells in the BM of recipient mice. In addition, we revealed that treatment with rapamycin (an mTORC1 inhibitor) aggravated the deficiency in B cell development in the PB and BM. Taken together, our results provide further evidence that Rictor regulates the development of early B cells in a cell-intrinsic manner by modifying the expression of FoxO1 and Rag-1.

Rictor/mammalian target of rapamycin 2 regulates the development of Notch1 induced murine T-cell acute lymphoblastic leukemia via forkhead box O3.

Mammalian target of rapamycin (mTOR) is composed of two distinct biochemical complexes, mTORC1 and mTORC2. In response to nutrients and growth factors, mTORC1 is known to control cellular growth by regulating the translational regulators S6 kinase 1 and 4E binding protein 1, whereas mTORC2 mediates cell proliferation and survival by activating Akt through phosphorylation at Ser473. Studies have shown that the deregulation of mTORC2 leads to the development of myeloproliferative disorder and leukemia in the phosphatase and tensin homolog deleted on chromosome ten (PTEN)-deleted mouse model. However, the mechanism by which mTORC2 specifically affects leukemogenesis is still not fully understood. Here, we investigated the role of mTORC2 in NOTCH1-driven T-cell acute lymphoblastic leukemia (T-ALL) in a Rictor-deficient mouse model. We found that, by deleting Rictor, an essential component of mTORC2, leukemia progression was significantly suppressed by arresting a greater proportion of Rictor(△/△) leukemic cells at the G0 phase of the cell cycle. Furthermore, the absence of Rictor led to the overexpression of chemotaxis-related genes, such as CCR2, CCR4 and CXCR4, which contributed to the homing and migration of Rictor-deficient T-ALL cells to the spleen but not the bone marrow. In addition, we demonstrated that inactivation of mTORC2 caused the overexpression of forkhead box O3 and its downstream effectors and eased the progression of leukemia in T-ALL mice. Our study thus indicates that forkhead box O3 could be a potential drug target for the treatment of T-ALL leukemia.

Identification of functional cooperative mutations of SETD2 in human acute leukemia.

Acute leukemia characterized by chromosomal rearrangements requires additional molecular disruptions to develop into full-blown malignancy, yet the cooperative mechanisms remain elusive. Using whole-genome sequencing of a pair of monozygotic twins discordant for MLL (also called KMT2A) gene-rearranged leukemia, we identified a transforming MLL-NRIP3 fusion gene and biallelic mutations in SETD2 (encoding a histone H3K36 methyltransferase). Moreover, loss-of-function point mutations in SETD2 were recurrent (6.2%) in 241 patients with acute leukemia and were associated with multiple major chromosomal aberrations. We observed a global loss of H3K36 trimethylation (H3K36me3) in leukemic blasts with mutations in SETD2. In the presence of a genetic lesion, downregulation of SETD2 contributed to both initiation and progression during leukemia development by promoting the self-renewal potential of leukemia stem cells. Therefore, our study provides compelling evidence for SETD2 as a new tumor suppressor. Disruption of the SETD2-H3K36me3 pathway is a distinct epigenetic mechanism for leukemia development.

[Recent advances of studies on role of mTOR signaling in aging of hematopoietic and other organ systems-review].

Mammalian target of rapamycin (mTOR) is a serine/threonine protein kinase, which plays an essential role in cell growth, proliferation and survival. mTOR regulates the transcription of mRNA, synthesis of ribosome and gene expression for metabolism. By forming mTOR complex, it regulates cellular activities by phosphorylating its downstream proteins, such as S6 protein kinase and 4E-BP1. In recent years, the role of mTORC1 in regulating aging is gradually recognized. Studies of physiological function and the regulatory mechanisms of mTOR signaling can not only help to better understand the aging mechanism for cells or organs, but also provide insights as to finding potential new drug targets for aging related diseases. This review focuses on recent advances of mTOR and aging related diseases in hematopoietic and other organ systems.

Distinct sensitivity of CD8+ CD4- and CD8+ CD4+ leukemic cell subpopulations to cyclophosphamide and rapamycin in Notch1-induced T-ALL mouse model.

The Notch1 signaling pathway plays an essential role in cell growth and differentiation. Over-expression of the intracellular Notch1 domain (ICN1) in murine hematopoietic cells is able to induce robust T-cell acute lymphoblastic leukemia (T-ALL) in mice. Here we explored the drug sensitivity of T-ALL cells in two subpopulations of CD8(+)CD4(+) and CD8(+)CD4(-) cells in Notch1-induced T-ALL mice. We found that Notch1 induced T-ALL cells could be decreased by chemotherapeutic drug cyclophosphamide (CTX). CD8(+)CD4(-) T-ALL cells were more sensitive to CTX treatment than CD8(+)CD4(+) T-ALL cells. The percentage of apoptotic cells induced by CTX treatment was higher in CD8(+)CD4(-) T-ALL cells. T-ALL cells were also inhibited by inhibitor of mTORC1 rapamycin. CD8(+)CD4(+) T-ALL cells were more susceptible to rapamycin treatment than CD8(+)CD4(-) T-ALL cells. Rapamycin treatment selectively arrested more CD8(+)CD4(+) T-ALL cells at G0 phase of cell cycle. A combination of the two drugs significantly improved overall survival of T-ALL bearing mice when compared with CTX or rapamycin alone. These results indicated that CD8(+)CD4(+) and CD8(+)CD4(-) leukemia cell populations had distinct drug sensitivity.

[Regulatory mechanisms of PI3K/AKT signaling pathway in acute leukemia].

This study was aimed to analyze the expression profiles of PI3K/AKT signaling pathway genes from bone marrow samples of AML and ALL patients and normal samples. AML, ALL and normal bone marrow samples were collected from 6 AML, 6 ALL patients and 4 normal persons. The expression of PI3K/AKT signaling pathway genes including PTEN, CCND1, mTOR, RICTOR, FOXO1 were detected by real-time fluorescent quantification RT-PCR while GAPDH gene expression was used as an internal reference. The relative gene expression level was calculated by the method of the 2(-ΔΔCt). The results showed that the gene expression profiles were different between normal and leukemic groups. PTEN, mTOR and RICTOR expression levels were down-regulated, while FOXO1 and CCND1 levels were up-regulated in AML and ALL. PTEN was down-regulated in 10 out of the 12 samples; mTOR was down-regulated in 9 out of the 12 samples; RICTOR was down-regulated in 7 out of the 12 samples; FOXO1 was up-regulated in 9 out of the 12 samples and CCND1 was up-regulated in 7 out of the 12 samples. It is concluded that PI3K/AKT signal pathway is activated in both AML and ALL leukemic cells.