Downregulation of FEM1C enhances metastasis and proliferation in colorectal cancer.

BACKGROUND: Feminization-1 (FEM-1) is considered a substrate recognition subunit of CUL2-RING E3 ubiquitin ligase complexes, which refers to sex determination by modulating TRA-1 stability in C. elegans. The function of mammalian orthologous gene of FEM-1 remains to be elucidated. METHODS: The expression of FEM1C in colorectal cancer (CRC) cells was interfered by small interference RNA (siRNA) transfection, and Cell counting kit-8 (CCK-8) assay, colony formation assay and transwell assay were performed. In order to estimate the function on metastasis, stable knockdown FEM1C cells were used to established liver and lung metastasis models. In addition, the expression of FEM1C in normal tissues, adenomas and tumor tissues were analyzed, and the relationship between FEM1C expression level and prognosis was analyzed by Kaplan-Meier method. RESULTS: Here, we report that the elimination of FEM1C, one of the members of FEM-1, significantly promoted the migration and invasion of colorectal cancer (CRC) cells in vitro and promoted liver and lung metastases in vivo. It also showed that the removal of FEM1C improved the proliferation ability of CRC cells. In particular, the cell shape changed from epithelial to fibroblast-like morphology. The tight cell monolayer was transformed into a dispersed distribution. Furthermore, it was demonstrated that FEM1C is down-regulated in tissues of CRC compared to normal tissues, and the high expression of FEM1C positively correlates with a good prognosis in patients with CRC. GSEA analysis showed that EMT signatures was enriched in FEM1C knockdown groups. CONCLUSIONS: Down-regulation of FEM1C promotes proliferation and metastasis, and FEM1C may be a tumor suppressor in the development of CRC.

LncRNA ENSG00000254615 Modulates Proliferation and 5-FU Resistance by Regulating p21 and Cyclin D1 in Colorectal Cancer.

5-Fluorouracil (5-FU) resistance is an urgent problem of colorectal cancer (CRC) chemotherapy that needs to be resolved. To investigate 5-FU-associated lncRNAs for CRC might be of great significant. LncRNA ENSG00000254615 was detected by RNA-sequencing. ENSG00000254615 were detected highly expressed in 5-FU-sensitive CRC cells and tissue specimens, and inhibited cell proliferation and attenuated 5-FU resistance in vitro and in vivo. Furthermore, ENSG00000254615 participated in the regulation of p21 and Cyclin D1. Taken together, we proposed that ENSG00000254615 inhibits proliferation and attenuates 5-FU resistance of CRC by regulating p21 and Cyclin D1 expression.

m6A methyltransferase METTL3 suppresses colorectal cancer proliferation and migration through p38/ERK pathways.

Purpose: Although many biological processes are involved in the modification of N6-methyladenosine (m6A), the exact role of m6A in the development of malignant tumors remains unclear. Methyltransferase 3 (METTL3) is a major RNA N6-methyladenosine methyltransferase. We aimed to explore the role of METTL3 in colorectal cancer (CRC) carcinogenesis and disease progression. Methods: In this study, immunohistochemistry was performed with a tissue microarray. qRT-PCR and Western blots were used to evaluate the expression of METTL3 in CRC cells. The effect of METTL3 on cell proliferation, migration and invasion of CRC cells was examined by IncuCyte Live Cell Analysis System and transwell assay, respectively. Results: The results suggested that positive expression of METTL3 was significantly associated with longer survival time (P=0.011). We next demonstrated that overexpression of METTL3 could inhibit proliferation, migration and invasion in CRC cells, while downregulation of METTL3 shows the opposite result. Furthermore, downregulation of METTL3 resulted in activation of p-p38 and p-ERK. Moreover, the inhibitors of p38 or ERK kinase could significantly reverse the effect of migration and invasion, which was induced by knockdown of METTL3. Conclusion: We concluded that METTL3 played a tumor-suppressive role in CRC cell proliferation, migration and invasion through p38/ERK pathways, which indicated that METTL3 might be a novel marker for CRC carcinogenesis, progression and survival.

Echocardiographic Assessment for the Detection of Cardiotoxicity Due to Vascular Endothelial Growth Factor Inhibitor Therapy in Metastatic Renal Cell and Colorectal Cancers.

BACKGROUND: Cardio-oncology is a recently established discipline that focuses on the management of patients with cancer who are at risk for developing cardiovascular complications as a result of their underlying oncologic treatment. In metastatic colorectal cancer (mCRC) and metastatic renal cell carcinoma (mRCC), vascular endothelial growth factor inhibitor (VEGF-i) therapy is commonly used to improve overall survival. Although these novel anticancer drugs may lead to the development of cardiotoxicity, whether early detection of cardiac dysfunction using serial echocardiography could potentially prevent the development of heart failure in this patient population requires further study. The aim of this study was to investigate the role of two-dimensional speckle-tracking echocardiography in the detection of cardiotoxicity due to VEGF-i therapy in patients with mCRC or mRCC. METHODS: Patients with mRCC or mCRC were evaluated using serial echocardiography at baseline and 1, 3, and 6 months following VEGF-i treatment. RESULTS: A total of 40 patients (34 men; mean age, 63 ± 9 years) receiving VEGF-i therapy were prospectively recruited at two academic centers: 26 (65%) were receiving sunitinib, eight (20%) pazopanib, and six (15%) bevacizumab. The following observations were made: (1) 8% of patients developed clinically asymptomatic cancer therapeutics-related cardiac dysfunction; (2) 30% of patients developed clinically significant decreases in global longitudinal strain, a marker for early subclinical cardiac dysfunction; (3) baseline abnormalities in global longitudinal strain may identify a subset of patients at higher risk for developing cancer therapeutics-related cardiac dysfunction; and (4) new or worsening hypertension was the most common adverse cardiovascular event, afflicting nearly one third of the study population. CONCLUSIONS: Cardiac dysfunction defined by serial changes in myocardial strain assessed using two-dimensional speckle-tracking echocardiography occurs in patients undergoing treatment with VEGF-i for mCRC or mRCC, which may provide an opportunity for preventive interventions.

An affinity peptide exerts antiviral activity by strongly binding nervous necrosis virus to block viral entry.

Nervous necrosis virus (NNV) causes viral nervous necrosis (VNN), a disease that leads to almost 100% mortality among larvae and juvenile fish, severely affecting the aquaculture industry. VNN vaccines based on inactivated viruses or virus-like particles (VLPs) are unsuitable for fish fry with immature adaptive immune systems. Here, we applied an anti-NNV strategy based on affinity peptides (AFPs). Three phage display peptide libraries were screened against RBS, the VLP of orange-spotted grouper nervous necrosis virus (OGNNV). From the positive clones, a dodecapeptide with the highest binding capacity (BC) to RBS was selected. This AFP agglutinated or disrupted virion particles, inhibiting RBS entry into sea bass (SB) cells. To enhance BC and solubility, we amended the AFP sequence as "LHWDFQSWVPLL" and named as 12C. One to three copies of 12C in tandem were prokaryotically expressed with a maltose binding protein (MBP) linked by a flexible peptide. Of the recombinant proteins expressed, MBP-triple-12C (MBP-T12C) exhibited the highest BC, efficiently blocked RBS entry, and strongly inhibited OGNNV infection at viral entry. Moreover, MBP-T12C bound the VLPs of all NNV serotypes, displaying broad-spectrum anti-NNV ability, and recognized only OGNNV and mud crab virus, demonstrating binding specificity. Therefore, these anti-NNV AFPs specifically bound NNV, aggregating or disrupting the viral particles, to reduce the contact probability between the virus and cell surface, subsequently inhibiting viral infection. Our results not only provided a candidate of anti-NNV AFP, but a framework for the development of antiviral AFP.

A Potent and Specific CD38 Inhibitor Ameliorates Age-Related Metabolic Dysfunction by Reversing Tissue NAD+ Decline.

Aging is characterized by the development of metabolic dysfunction and frailty. Recent studies show that a reduction in nicotinamide adenine dinucleotide (NAD+) is a key factor for the development of age-associated metabolic decline. We recently demonstrated that the NADase CD38 has a central role in age-related NAD+ decline. Here we show that a highly potent and specific thiazoloquin(az)olin(on)e CD38 inhibitor, 78c, reverses age-related NAD+ decline and improves several physiological and metabolic parameters of aging, including glucose tolerance, muscle function, exercise capacity, and cardiac function in mouse models of natural and accelerated aging. The physiological effects of 78c depend on tissue NAD+ levels and were reversed by inhibition of NAD+ synthesis. 78c increased NAD+ levels, resulting in activation of pro-longevity and health span-related factors, including sirtuins, AMPK, and PARPs. Furthermore, in animals treated with 78c we observed inhibition of pathways that negatively affect health span, such as mTOR-S6K and ERK, and attenuation of telomere-associated DNA damage, a marker of cellular aging. Together, our results detail a novel pharmacological strategy for prevention and/or reversal of age-related NAD+ decline and subsequent metabolic dysfunction.

Hemocyanin from Shrimp Litopenaeus vannamei Has Antiproliferative Effect against HeLa Cell In Vitro.

Hemocyanin (HMC) has been shown to participate in multiple roles of immune defence. In this study, we investigated the antiproliferative effect and underpinning mechanism of HMC from Litopenaeus vannamei in vitro. Sulforhodamine B (SRB) assay indicated that HMC could dramatically inhibit the growth of HeLa cells, but not 293T cells under the same conditions. Moreover, typical morphological features of apoptosis in HeLa cells including the formation of apoptotic body-like vesicles, chromatin condensation and margination were observed by using 4, 6-diamidino-2- phenylindole dihydrochloride (DAPI) staining and fluorescence analysis. An apoptotic DNA ladder from 180 to 300 bp was also detected. Furthermore, 10 variation proteins associated with apoptosis pathway, viz. G3PDH isoforms 1/2 (G3PDH1/2), aldosereductase, ectodemal dysplasia receptor associated death receptor domain isoform CRA_a (EDARADD), heat shock 60kD protein 1 variant 1 (HSP60), heat shock 70kDa protein 5 precursor (HSP70), heat shock protein 90kDa beta member 1 precursor (HSP90), 14-3-3 protein ζ/δ, Ran and ubiquitin activating enzyme E1(UBE1), were identified from HMC-treated HeLa cells by the proteomic and quantitative real-time RT-PCR strategies. Importantly, the reactive oxygen species (ROS), mitochondrial membrane potential (Δψm) and caspase-9/3 activities were changed significantly in HMC-treated HeLa cells. Together, the data suggests that L. vannamei HMC mediates antiproliferative properties through the apoptosis mechanism involving the mitochondria triggered pathway.

Structural analysis and insertion study reveal the ideal sites for surface displaying foreign peptides on a betanodavirus-like particle.

Betanodavirus infection causes fatal disease of viral nervous necrosis in many cultured marine and freshwater fish worldwide and the virus-like particles (VLP) are effective vaccines against betanodavirus. But vaccine and viral vector designs of betanodavirus VLP based on their structures remain lacking. Here, the three-dimensional structure of orange-spotted grouper nervous necrosis virus (OGNNV) VLP (RBS) at 3.9 Å reveals the organization of capsid proteins (CP). Based on the structural results, seven putative important sites were selected to genetically insert a 6× histidine (His)-tag for VLP formation screen, resulting in four His-tagged VLP (HV) at positions N-terminus, Ala220, Pro292 and C-terminus. The His-tags of N-terminal HV (NHV) were concealed inside virions while those of 220HV and C-terminal HV (CHV) were displayed at the outer surface. NHV, 220HV and CHV maintained the same cell entry ability as RBS in the Asian sea bass (SB) cell line, indicating that their similar surface structures can be recognized by the cellular entry receptor(s). For application of vaccine design, chromatography-purified CHV could provoke NNV-specific antibody responses as strong as those of RBS in a sea bass immunization assay. Furthermore, in carrying capacity assays, N-terminus and Ala220 can only carry short peptides and C-terminus can even accommodate large protein such as GFP to generate fluorescent VLP (CGV). For application of a viral vector, CGV could be real-time visualized to enter SB cells in invasion study. All the results confirmed that the C-terminus of CP is a suitable site to accommodate foreign peptides for vaccine design and viral vector development.

Regadenoson Stress Real-Time Myocardial Perfusion Echocardiography for Detection of Coronary Artery Disease: Feasibility and Accuracy of Two Different Ultrasound Contrast Agents.

BACKGROUND: The aim of this study was to compare the efficacy of myocardial perfusion (MP) and wall motion (WM) analysis obtained with real-time myocardial contrast echocardiography (RTMCE) and two widely used contrast agents in detecting coronary artery disease after injection of the vasodilator regadenoson. METHODS: One hundred fifty patients were studied at two academic centers using regadenoson (400-µg intravenous bolus) vasodilator stress RTMCE (7.5% Optison infusion [n = 50] or 1.5% Definity infusion [n = 100]). Both MP and WM with RTMCE were analyzed at rest and after regadenoson bolus. Comparisons of WM and MP sensitivity, specificity, and accuracy were made. Quantitative angiography was performed in all patients within 1 month of the regadenoson stress study (>50% and >70% diameter stenosis was considered significant). Reviewers were blinded to all clinical and quantitative angiographic data. RESULTS: Rate-pressure product after regadenoson was higher in Optison than Definity patients (P = .004). Using a 50% diameter stenosis on quantitative angiography as a reference standard, overall sensitivity, specificity, and accuracy for combined WM and MP analysis were not different for both agents (Optison, 77%, 64%, and 73%; Definity, 80%, 74%, and 78%; P = NS). The sensitivity, specificity, and accuracy of WM analysis alone for Optison were 68%, 71%, and 69% compared with 60%, 72%, and 66% for Definity (P = NS). Adding MP analysis improved the sensitivity and accuracy of Definity for detecting both >50% and >70% stenoses (P < .001 vs WM), while MP analysis did not improve the sensitivity of Optison for detecting either >50 or >70% stenoses. CONCLUSIONS: RTMCE during regadenoson stress using either Optison or Definity is a rapid and effective method for the detection of coronary artery disease. The ability of MP imaging to improve WM accuracy may depend on the rate-pressure product achieved.

Relationship between HgbA1c and myocardial blood flow reserve in patients with type 2 diabetes mellitus: noninvasive assessment using real-time myocardial perfusion echocardiography.

To study the relationship between glycosylated hemoglobin (HgbA1c) and myocardial perfusion in type 2 diabetes mellitus (T2DM) patients, we prospectively enrolled 24 patients with known or suspected coronary artery disease (CAD) who underwent adenosine stress by real-time myocardial perfusion echocardiography (RTMPE). HgbA1c was measured at time of RTMPE. Microbubble velocity (ß min(-1)), myocardial blood flow (MBF, mL/min/g), and myocardial blood flow reserve (MBFR) were quantified. Quantitative MCE analysis was feasible in all patients (272/384 segments, 71%). Those with HgbA1c > 7.1% had significantly lower ßreserve and MBFR than those with HgbA1c ≤ 7.1% (P < 0.05). In patients with suspected CAD, there was a significant inverse correlation between MBFR and HgbA1c (r = -0.279, P = 0.01); however, in those with known CAD, this relationship was not significant (r = -0.117, P = 0.129). Using a MBFR cutoff value > 2 as normal, HgbA1c > 7.1% significantly increased the risk for abnormal MBFR, (adjusted odds ratio: 1.92, 95% CI: 1.12-3.35, P = 0.02). Optimal glycemic control is associated with preservation of MBFR as determined by RTMPE, in T2DM patients at risk for CAD.

Evaluation of myocardial viability after myocardial infarction with intravenous real-time myocardial contrast echocardiography.

The myocardial viability after myocardial infarction was evaluated by intravenous myocardial contrast echocardiography. Intravenous real-time myocardial contrast echocardiography was performed on 18 patients with myocardial infarction before coronary revascularization. Follow-up echocardiography was performed 3 months after coronary revascularization. Segmental wall motion was assessed using 18-segment LV model and classified as normal, hypokinesis, akinesis and dyskinesis. Viable myocardium was defined by evident improvement of segmental wall motion 3 months after coronary revascularization. Myocardial perfusion was assessed by visual interpretation and divided into 3 conditions: homogeneous opacification; partial or reduced opacification or subendocardial contrast defect; contrast defect. The former two conditions were used as the standard to define the viable myocardium. The results showed that 109 abnormal wall motion segments were detected among 18 patients with myocardial infarction, including 47 segments of hypokinesis, 56 segments of akinesis and 6 segments of dyskinesis. The wall motion of 2 segments with hypokinesis before coronary revascularization which showed homogeneous opacification, 14 of 24 segments with hypokinese and 20 of 24 segments with akinese before coronary revascularization which showed partial or reduced opacification or subendocardial contrast defect was improved 3 months after coronary revascularization. In our study, the sensitivity and specificity of evaluation of myocardial viability after myocardial infarction by intravenous real-time myocardial contrast echocardiography were 94.7% and 78.9%, respectively. It was concluded that intravenous real-time myocardial contrast echocardiography could accurately evaluate myocardial viability after myocardial infarction.