TRIM44 Promotes Rabies Virus Replication by Autophagy-Dependent Mechanism.

Tripartite motif (TRIM) proteins are a multifunctional E3 ubiquitin ligase family that participates in various cellular processes. Recent studies have shown that TRIM proteins play important roles in regulating host-virus interactions through specific pathways, but their involvement in response to rabies virus (RABV) infection remains poorly understood. Here, we identified that several TRIM proteins are upregulated in mouse neuroblastoma cells (NA) after infection with the rabies virus using RNA-seq sequencing. Among them, TRIM44 was found to regulate RABV replication. This is supported by the observations that downregulation of TRIM44 inhibits RABV replication, while overexpression of TRIM44 promotes RABV replication. Mechanistically, TRIM44-induced RABV replication is brought about by activating autophagy, as inhibition of autophagy with 3-MA attenuates TRIM44-induced RABV replication. Additionally, we found that inhibition of autophagy with rapamycin reverses the TRIM44-knockdown-induced decrease in LC3B expression and autophagosome formation as well as RABV replication. The results suggest that TRIM44 promotes RABV replication by an autophagy-dependent mechanism. Our work identifies TRIM44 as a key host factor for RABV replication, and targeting TRIM44 expression may represent an effective therapeutic strategy.

Pyruvate Kinase Differentially Alters Metabolic Signatures during Head and Neck Carcinogenesis.

During glycolysis, the muscle isoform of pyruvate kinase PKM2 produces ATP in exchange for dephosphorylation of phosphoenolpyruvate (PEP) into pyruvate. PKM2 has been considered as a tumor-promoting factor in most cancers, whereas the regulatory role of PKM2 during head and neck carcinogenesis remained to be delineated. PKM2 mRNA and protein expression was examined in head and neck tumorous specimens. The role of PKM2 in controlling cellular malignancy was determined in shRNA-mediated PKM2-deficient head and neck squamous cell carcinoma (HNSC) cells. In agreement with the results in other cancers, PKM2 expression is enriched in both mouse and human HNSC tissues. Nevertheless, PKM2 mRNA expression reversely correlated with tumor stage, and greater recurrence-free survival rates are evident in the PKM2high HNSC population, arguing that PKM2 may be tumor-suppressive. Multifaceted analyses showed a greater in vivo xenografic tumor growth and an enhanced cisplatin resistance in response to PKM2 loss, whereas PKM2 silencing led to reduced cell motility. At the molecular level, metabolic shifts towards mitochondrial metabolism and activation of oncogenic Protein kinase B (PKB/Akt) and extracellular signal-regulated kinase (ERK) signals were detected in PKM2-silencing HNSC cells. In sum, our findings demonstrated that PKM2 differentially modulated head and neck tumorigenicity via metabolic reprogramming.

Nanoparticle-Mediated Therapy with miR-198 Sensitizes Pancreatic Cancer to Gemcitabine Treatment through Downregulation of VCP-Mediated Autophagy.

Pancreatic ductal adenocarcinoma (PDAC) remains an extremely aggressive disease characterized by rapidly acquired multi-drug resistance, including to first-line chemotherapeutic agent gemcitabine. Autophagy is a process that is often exploited by cancer and is one of several intrinsic factors associated with resistance to gemcitabine. We have previously found that miR-198 acts as a tumor suppressor in PDAC through the targeting of factors including Valosin-containing protein (VCP). VCP has been reported to play an important role in autophagic flux. In this study, we investigated whether the repression of VCP through miR-198 administration disrupts the autophagy process and sensitizes PDAC cells to gemcitabine treatment in vitro. Moreover, we used LGA-PEI (LPNP) nanoparticles to effectively administer miR-198 to tumors in vivo, inducing tumor sensitization to gemcitabine and leading to a significant reduction in tumor burden and metastases and a concomitant downregulation of VCP expression and autophagy maturation. Our results indicate a potential therapeutic strategy for targeting gemcitabine resistant PDAC and establishes the use of LPNPs for effective therapeutic delivery of nucleic acids in vitro and in vivo.

Comparison of Longitudinal Skeletal Thigh Muscle Findings With Magnetic Resonance Imaging in Patients With Peripheral Artery Disease With-Versus-Without Diabetes Mellitus.

The aim of this secondary analysis of ELIMIT (The Effect of Lipid Modification on Peripheral Artery Disease after Endovascular Intervention Trial) was to determine longitudinal changes over 24 months in skeletal thigh muscle volumes and individual muscle compartments in patients with peripheral artery disease (PAD) with and without diabetes. A total of 48 patients with available magnetic resonance imaging of the distal superficial femoral artery at baseline and 2 years were included in this analysis. Muscle volumes and superficial femoral artery wall, lumen, and total vessel volumes were quantified. Intrareader reproducibility of muscle tracings was assessed with the intraclass correlation coefficient using a 2-way model. Baseline characteristics were similar between patients with PAD with and without diabetes, except for smoking history (p = 0.049), cholesterol levels (p <0.050), and calf walking pain (p = 0.049). Interobserver reproducibility of the muscle volume tracings was excellent for all muscle groups (all intraclass correlation coefficients >0.86, confidence interval 0.69 to 0.94). Total muscle and total leg volumes increased significantly between baseline and 24 months among patients with PAD without diabetes (31 ± 6.4 cm3 vs 32 ± 7.0 cm3, p <0.001; 18 ± 4.4 cm3 vs 19 ± 4.8 cm3, p = 0.045), whereas there was no change in patients with PAD and diabetes. Total muscle volume was inversely associated with age and body mass index in patients with PAD both with and without diabetes (p <0.05). In conclusion, magnetic resonance imaging-quantified thigh muscle volumes are highly reproducible and may be of interest in assessing PAD patients with and without diabetes.

Two Antibody-Guided Lactic-co-Glycolic Acid-Polyethylenimine (LGA-PEI) Nanoparticle Delivery Systems for Therapeutic Nucleic Acids.

We previously reported a new polymer, lactic-co-glycolic acid-polyethylenimine (LGA-PEI), as an improved nanoparticle (NP) delivery for therapeutic nucleic acids (TNAs). Here, we further developed two antibody (Ab)-conjugated LGA-PEI NP technologies for active-targeting delivery of TNAs. LGA-PEI was covalently conjugated with a single-chain variable fragment antibody (scFv) against mesothelin (MSLN), a biomarker for pancreatic cancer (PC), or a special Ab fragment crystallizable region-binding peptide (FcBP), which binds to any full Ab (IgG). TNAs used in the current study included tumor suppressor microRNA mimics (miR-198 and miR-520h) and non-coding RNA X-inactive specific transcript (XIST) fragments; green fluorescence protein gene (GFP plasmid DNA) was also used as an example of plasmid DNA. MSLN scFv-LGA-PEI NPs with TNAs significantly improved their binding and internalization in PC cells with high expression of MSLN in vitro and in vivo. Anti-epidermal growth factor receptor (EGFR) monoclonal Ab (Cetuximab) binding to FcBP-LGA-PEI showed active-targeting delivery of TNAs to EGFR-expressing PC cells.

A subset of cytotoxic effector memory T cells enhances CAR T cell efficacy in a model of pancreatic ductal adenocarcinoma.

In humans, the natural killer (NK) cell marker CD161 identifies several subsets of T cells, including a polyclonal CD8 αß T cell receptor-expressing subset with characteristic specificity for tissue-localized viruses. This subset also displays enhanced cytotoxic and memory phenotypes. Here, we characterized this unique T cell subset and determined its potential suitability for use in chimeric antigen receptor (CAR) T cell therapy. In mice, gene expression profiling among the CD161-equivalent CD8+ T cell populations (CD8+NK1.1+) revealed substantial up-regulation of granzymes, perforin, killer lectin-like receptors, and innate signaling molecules in comparison to CD8+NK1.1- T cells. Adoptive transfer of CD8+NK1.1+ cells from previously exposed animals offered substantially enhanced protection and improved survival against melanoma tumors and influenza infection compared to CD8+NK1.1- cells. Freshly isolated human CD8+CD61+ T cells exhibited heightened allogeneic killing activity in comparison to CD8+CD61- T cells or total peripheral blood mononuclear cells (PBMCs). To determine whether this subset might improve the antitumor efficacy of CAR T cell therapy against solid tumors, we compared bulk PBMCs, CD8+CD161-, and CD8+CD161+ T cells transduced with a human epidermal growth factor receptor-2 (HER2)-specific CAR construct. In vitro, CD8+CD161+ CAR-transduced T cells killed HER2+ targets faster and with greater efficiency. Similarly, these cells mediated enhanced in vivo antitumor efficacy in xenograft models of HER2+ pancreatic ductal adenocarcinoma, exhibiting elevated expression of granzymes and reduced expression of exhaustion markers. These data suggest that this T cell subset presents an opportunity to improve CAR T cell therapy for the treatment of solid tumors.

Long-Term Results of Concurrent Chemoradiotherapy Combined with Anti-EGFR Monoclonal Antibody Prior to Surgery in Locally Advanced Cervical Cancer: A Single-Institute Prospective Study.

PURPOSE: We aimed to evaluate the long-term survival outcomes of concurrent chemoradiotherapy (CCRT) combined with nimotuzumab followed by surgery in patients with locally advanced cervical cancer (LACC). PATIENTS AND METHODS: Patients received whole pelvic intensity-modulated radiation therapy (IMRT) and concomitantly with weekly cisplatin (40 mg/m2) or nedaplatin (30 mg/m2) and weekly nimotuzumab (200 mg). After assessment of the treatment response, patients then underwent radical surgery. RESULTS: Between June 2013 and July 2016, 33 patients with FIGO IB2-IIIB cervical cancer were recruited. Clinical complete response and partial response were observed in 8 (24.3%) and 23 patients (69.7%), respectively. Twenty-seven patients (81.8%) were successfully treated with radical hysterectomy and pelvic lymphadenectomy: 9 (33.3%) showed pathological complete response; 10 (37.1%) showed partial response and 8 (29.6%) presented with persistent macroscopic/microscopic residual carcinoma. For the intention-to-treat population, the median follow-up time was 53.7 months. Locoregional recurrence and distant metastases were observed in three and seven patients, respectively. The 5-year overall survival, progression-free survival, locoregional recurrence-free survival, and distant metastasis-free survival were 81.5%, 72.7%, 90.9%, and 78.3%, respectively. Both acute and late toxicities were manageable and mainly limited to grade 1 or 2. CONCLUSION: Concurrent chemoradiotherapy combined with nimotuzumab followed by surgery for patients with LACC is safe and results in excellent long-term treatment outcomes. Further randomized controlled studies are warranted to confirm the findings.

PDGFRA in vascular adventitial MSCs promotes neointima formation in arteriovenous fistula in chronic kidney disease.

Chronic kidney disease (CKD) induces the failure of arteriovenous fistulas (AVFs) and promotes the differentiation of vascular adventitial GLI1-positive mesenchymal stem cells (GMCs). However, the roles of GMCs in forming neointima in AVFs remain unknown. GMCs isolated from CKD mice showed increased potential capacity of differentiation into myofibroblast-like cells. Increased activation of expression of PDGFRA and hedgehog (HH) signaling were detected in adventitial cells of AVFs from patients with end-stage kidney disease and CKD mice. PDGFRA was translocated and accumulated in early endosome when sonic hedgehog was overexpressed. In endosome, PDGFRA-mediated activation of TGFB1/SMAD signaling promoted the differentiation of GMCs into myofibroblasts, extracellular matrix deposition, and vascular fibrosis. These responses resulted in neointima formation and AVF failure. KO of Pdgfra or inhibition of HH signaling in GMCs suppressed the differentiation of GMCs into myofibroblasts. In vivo, specific KO of Pdgfra inhibited GMC activation and vascular fibrosis, resulting in suppression of neointima formation and improvement of AVF patency despite CKD. Our findings could yield strategies for maintaining AVF functions.

Corrigendum: Regulatory Role of Hexokinase 2 in Modulating Head and Neck Tumorigenesis.

[This corrects the article DOI: 10.3389/fonc.2020.00176.].

Regulatory Role of Hexokinase 2 in Modulating Head and Neck Tumorigenesis.

To support great demand of cell growth, cancer cells preferentially obtain energy and biomacromolecules by glycolysis over mitochondrial oxidative phosphorylation (OxPhos). Among all glycolytic enzymes, hexokinase (HK), a rate-limiting enzyme at the first step of glycolysis to catalyze cellular glucose into glucose-6-phosphate, is herein emphasized. Four HK isoforms, HK1-HK4, were discovered in nature. It was shown that HK2 expression is enriched in many tumor cells and correlated with poorer survival rates in most neoplastic cells. HK2-mediated regulations for cell malignancy and mechanistic cues in regulating head and neck tumorigenesis, however, are not fully elucidated. Cellular malignancy index, such as cell growth, cellular motility, and treatment sensitivity, and molecular alterations were determined in HK2-deficient head and neck squamous cell carcinoma (HNSCC) cells. By using various cancer databases, HK2, but not HK1, positively correlates with HNSCC progression in a stage-dependent manner. A high HK2 expression was detected in head and neck cancerous tissues compared with their normal counterparts, both in mouse and human subjects. Loss of HK2 in HNSCC cells resulted in reduced cell (in vitro) and tumor (in vivo) growth, as well as decreased epithelial-mesenchymal transition-mediated cell movement; in contrast, HK2-deficient HNSCC cells exhibited greater sensitivity to chemotherapeutic drugs cisplatin and 5-fluorouracil but are more resistant to photodynamic therapy, indicating that HK2 expression could selectively define treatment sensitivity in HNSCC cells. At the molecular level, it was found that HK2 alteration drove metabolic reprogramming toward OxPhos and modulated oncogenic Akt and mutant TP53-mediated signals in HNSCC cells. In summary, the present study showed that HK2 suppression could lessen HNSCC oncogenicity and modulate therapeutic sensitivity, thereby being an ideal therapeutic target for HNSCCs.

Adsorption of miR-218 by lncRNA HOTAIR regulates PDE7A and affects glioma cell proliferation, invasion, and apoptosis.

OBJECTIVE: To evaluate the role of targeted adsorption of miR-218 by long-chain non-coding RNAHOTAIR to regulate PDE7A on glioma cell proliferation, invasion, and apoptosis. METHODS: The expressions of lncRNA HOTAIR, miR-218, and PDE7A in glioma tissues and normal parcancer tissues, NHA and glioma cell lines were determined, and correlations among the three genes were analyzed. The subcellular localization of lncRNA HOTAIR was determined by fluorescent in situ hybridization. Dual-luciferase reporter assay was used to validate the targeted relationship between lncRNA HOTAIR/miR-218/PDE7A. Glioma cells were grouped to receive intervention of lncRNA HOTAIR or miR-218. MTT, transwell, and flow cytometry were performed to determine the proliferation, invasion, and apoptosis of cells. RESULTS: Compared with the normal tissues and cells, the expression of lncRNA HOTAIR was increased while miR-218 was suppressed in glioma tissues samples and cells (all P<0.05). Inhibition of lncRNA HOTAIR expression, was able to induce apoptosis and suppress the proliferation and invasion of cells (all P<0.05). LncRNA HOTAIR is mainly localized in the cytoplasm, and is able to adsorb miR-218 as ceRNA. The effect of knockdown of HOTAIR on glioma cells could be partially rescued by miR-218 inhibitor. The expression of PDE7A was enhanced in glioma tissues and cells compared to normal tissues and cells (all P<0.05), which positively correlated with the expression of HOTAIR (r=0.546, P<0.05) and negatively correlated with the expression of miR-218 (r=0.363, P<0.05). The targeted relationship between miR-218 and PDE7A was validated: Overexpression of miR-218 was able to suppress the proliferation and invasion of glioma cells and restrain apoptosis compared to the miR-NC group (all P<0.05). The effect of miR-218 on glioma cells could be partially rescued by PDE7A. CONCLUSION: lncRNA HOTAIR can adsorb miR-218 to regulate expression of PDE7A and promote the malignant biologic behavior of glioma cells.

Mesothelin and TGF-α predict pancreatic cancer cell sensitivity to EGFR inhibitors and effective combination treatment with trametinib.

Clinical trials of EGFR inhibitors in combination with gemcitabine for the treatment of pancreatic ductal adenocarcinoma (PDAC) have generated mixed results partially due to the poorly defined effectiveness of EGFR inhibitors in PDAC. Here, we studied a panel of PDAC cell lines to compare the IC50s of the EGFR inhibitors gefitinib and cetuximab. We found that gefitinib induced biphasic inhibition in over 50% of PDAC cells, with the initial growth inhibition occurring at nanomolar concentrations and a second growth inhibition occurring outside the clinical range. In contrast to gefitinib, cetuximab produced a single phase growth inhibition in a subset of PDAC cells. Using this sensitivity data, we screened for correlations between cell morphology proteins and EGFR ligands to EGFR inhibitor sensitivity, and found that mesothelin and the EGFR ligand TGF-α have a strong correlation to gefitinib and cetuximab sensitivity. Analysis of downstream signaling pathways indicated that plc-γ1 and c-myc were consistently inhibited by EGFR inhibitor treatment in sensitive cell lines. While an inconsistent additive effect was observed with either cetuximab or gefitinib in combination with gemcitabine, the cell pathway data indicated consistent ERK activation, leading us to pursue EGFR inhibitors in combination with trametinib, a MEK1/2 inhibitor. Both cetuximab and gefitinib in combination with trametinib produced an additive effect in all EGFR sensitive cell lines. Our results indicate that mesothelin and TGF-α can predict PDAC sensitivity to EGFR inhibitors and a combination of EGFR inhibitors with trametinib could be a novel effective treatment for PDAC.

Notch signaling in bone marrow-derived FSP-1 cells initiates neointima formation in arteriovenous fistulas.

Neointima formation is a major contributor to arteriovenous fistula (AVF) failure. We have previously shown that activation of the Notch signaling pathway contributes to neointima formation by promoting the migration of vascular smooth muscle cells (VSMCs) into the venous anastomosis. In the current study we investigated the mechanisms underlying the dedifferentiation and migration of VSMCs, and in particular the role of bone marrow-derived fibroblast specific protein 1 (FSP-1)+ cells, another cell type found in models of vascular injury. Using VSMC-specific reporter mice, we found that most of the VSMCs participating in AVF neointima formation originated from dedifferentiated VSMCs. We also observed infiltration of bone marrow-derived FSP-1+ cells into the arterial anastomosis where they could interact with VSMCs. In vitro, conditioned media from FSP-1+ cells stimulated VSMC proliferation and phenotype switching. Activated Notch signaling transformed FSP-1+ cells into type I macrophages and stimulated secretion of cytokines and growth factors. Pretreatment with a Notch inhibitor or knockout of the canonical downstream factor RBP-Jκ in bone marrow-derived FSP1+ cells decreased FSP1+ cell infiltration into murine AVFs, attenuating VSMC dedifferentiation and neointima formation. Our results suggest that targeting Notch signaling could provide a new therapeutic strategy to improve AVF patency.

Ginsenoside Rb1 Blocks Ritonavir-Induced Oxidative Stress and eNOS Downregulation through Activation of Estrogen Receptor-Beta and Upregulation of SOD in Human Endothelial Cells.

We have previously shown that ritonavir (RTV), a highly active anti-retroviral therapy (HAART) drug, can cause endothelial dysfunction through oxidative stress. Several antioxidants including ginsenoside Rb1, a compound with antioxidant effect, can effectively block this side effect of RTV in endothelial cells. In the current study, we explored a mechanism by which ginsenoside Rb1 could protect these cells via binding of estrogen receptors (ERs). We found that several human endothelial cell lines differentially expressed ER-ß and had very low levels of ER-α. RTV treatment significantly increased the production of reactive oxygen species (ROS) and decreased the expression of endothelial nitric oxidase synthase (eNOS) and superoxide dismutase (SOD) in HUVECs, while Rb1 effectively blocked these effects of RTV. These effects of Rb1 were effectively inhibited by silencing ER-ß, indicating that ginsenoside Rb1 requires ER-ß for its antioxidant activity in inhibiting the deleterious effect of RTV in human endothelial cells. Furthermore, Rb1 specifically activated ER-ß transactivation activity by ER-ß luciferase reporter assay. Rb1 competitively bound to ER-ß, which was determined by the high sensitive fluorescent polarization assay.

A prospective study on neoadjuvant chemoradiotherapy plus anti-EGFR monoclonal antibody followed by surgery for locally advanced cervical cancer.

BACKGROUND: To investigate the efficacy and safety of neoadjuvant chemoradiotherapy plus anti-epidermal growth factor receptor monoclonal antibody followed by surgery for locally advanced cervical cancer (LACC). PATIENTS AND METHODS: Patients with histologically proven LACC were enrolled into this prospective study. All patients received intensity-modulated radiation therapy with conventional fractionation. Weekly cisplatin or nedaplatin was administered concurrently with intensity-modulated radiation therapy. Nimotuzumab, a humanized anti-epidermal growth factor receptor monoclonal antibody, was given at a dose of 200 mg per week for 6 cycles. Approximately 1 month after the completion of neoadjuvant treatment, the patients were assessed for clinical tumor response and operability based on MRI and gynecological examination. For those who were considered to be candidates for surgery, radical hysterectomy, and pelvic lymph node dissection were performed 5-6 weeks after the completion of neoadjuvant therapy. RESULTS: Twenty-eight patients were enrolled. Clinical complete response and partial response were found in 8 (28.5%) and 20 (71.5%) patients, respectively. Four patients were not eligible for surgery and 2 patients refused surgery although they were assessed as surgical candidates. They were not included in this analysis. Radical hysterectomy and pelvic lymph node dissection were performed for the remaining 22 patients. Among them, 8 (36.4%) had complete pathology response, 9 (40.9%) presented with persistent atypical cells or cervical intraepithelial neoplasia, and 5 (22.7%) presented with macroscopic and/or microscopic residual disease, according to the pathological evaluation. Median follow-up time was 22 months (range, 5-39 months). The 2-year locoregional control rate, progression-free survival rate, distant metastasis-free survival rate, and overall survival rate were 95.0%, 85.2%, 84.0%, and 90.0%, respectively. Acute toxicities were mild in general and easily manageable. Chronic toxicities were mainly limited to grade 1. No severe late toxicities were observed. CONCLUSION: Concurrent chemoradiotherapy plus nimotuzumab followed by surgery is highly effective and safe in LACC. Further studies are warranted to confirm the findings.

Sustained Thromboresistant Bioactivity with Reduced Intimal Hyperplasia of Heparin-Bonded Polytetrafluoroethylene Propaten Graft in a Chronic Canine Femoral Artery Bypass Model.

BACKGROUND: Bypass graft thrombosis remains a significant mode of failure in prosthetic graft revascularization. The purpose of this investigation was to evaluate the long-term thromboresistant effect of heparin-bonded expanded polytetrafluoroethylene (ePTFE) graft using Carmeda BioActive Surface technology in a canine model. METHODS: Bilateral femorofemoral artery bypass grafts with ePTFE grafts were performed in 25 adult grayhound dogs. In each animal, a heparin-bonded ePTFE graft (Propaten, WL Gore) was placed on one side, whereas a control nonheparin graft was placed on the contralateral side. The graft patency was assessed at 1, 6, 12, 18, and 24 months (n = 5 per group) following the bypass. Heparin bioactivity of the graft material was analyzed. The effect of intimal hyperplasia was also assessed. RESULTS: All bypass grafts were patent at 1 month. Significantly greater patency rates were noted in the Propaten group compared to the control group at 12, 18, and 24 months, which were 84%, 80%, and 80% vs. 55%, 35%, and 20%, respectively (P < 0.02). There was a significant reduction in the anastomotic neointimal area and neointimal cell proliferation in Propaten grafts compared with control grafts at all groups between 6 and 24 months (P < 0.05). Heparin bioactivity as measured by antithrombin binding assay was demonstrated in the Propaten graft between 1 and 24 months. Mean heparin activities on Propaten grafts ranged from 26.3 ± 6.4 pmol/cm2 to 18.4 ± 8.7 pmol/cm2 between 1 and 24 months, which were significantly greater than the control group (P < 0.001). Differences between mean heparin activities of explanted Propaten graft samples at the various time points were nonsignificant (P > 0.05). CONCLUSIONS: Heparin-bonded ePTFE graft provides a thromboresistant surface and reduced anastomotic intimal hyperplasia at 2 years. The stable heparin bioactivity of the Propaten graft confers an advantage in long-term graft patency.

Comparison of ASPIRE Mechanical Thrombectomy Versus AngioJet Thrombectomy System in a Porcine Iliac Vein Thrombosis Model.

BACKGROUND: Percutaneous mechanical thrombectomy device has become an important therapeutic armamentarium in the management of venous thromboembolism. In this study, we compare the efficacy and safety profile of the AngioJet thrombectomy device and ASPIRE thrombectomy system in a porcine venous thrombosis model. METHODS: Twelve adult pigs underwent bilateral iliac venous thrombosis created by using a stent graft thrombosis model and subsequently underwent either AngioJet (n = 6) or ASPIRE mechanical thrombectomy (n = 6) 1 week later. Intravascular ultrasound (IVUS) was used to assess thrombectomy efficacy, and computed tomography was used to evaluate pulmonary embolism (PE). Hemolytic effect was measured by plasma-free hemoglobin (PfHgb). Iliac vein thrombogenicity was evaluated with radiolabeled platelet and fibrin deposition. Veins were harvested and evaluated with light microscopy and scanning electron microscopy (SEM). RESULTS: Similar thrombectomy efficacy by IVUS evaluation was noted in both groups. Significant greater PE and hemolysis were identified in the AngioJet group compared to the ASPIRE group. The AngioJet group had greater reduction in WBC and platelet compared to the ASPIRE group. No difference was found in thrombogenicity, light microscopic evaluation, or SEM. CONCLUSIONS: Both devices had similar thrombectomy efficacy and thrombogenicity response. The ASPIRE catheter incurred less PE and hemolysis compared to the AngioJet device. Vessel wall response by histological analysis and SEM was similar in both groups.

Toll-like receptor 3 adjuvant in combination with virus-like particles elicit a humoral response against HIV.

Human Immunodeficiency Virus (HIV) Virus-Like Particles (VLPs) composed of HIVIIIB Gag and HIVBaL gp120/gp41 envelope are a pseudovirion vaccine capable of presenting antigens in their native conformations. To enhance the immunogenicity of the HIV Env antigen, VLPs were coupled to VesiVax Conjugatable Adjuvant Lipid Vesicles (CALV) containing one of four toll-like-receptor (TLR) ligands, each activating a receptor with distinct cellular localization and downstream pathways. C57BL/6 mice were vaccinated by intranasal prime followed by two sub-cheek boosts and their sera immunoglobulin and neutralizing potency were measured over a duration of 3months after vaccination. PBS control, VLPs alone, CALV+VLPs, and VLPs complexed with CALV and ligands for TLR2 (PAM3CAG), TLR3 (dsRNA), TLR4 (MPLA), or TLR7/8 (resiquimod) were evaluated based on antibody titer, IgG1 and IgG2c class switching, germinal center formation, T follicular cells and potency of neutralizing antibodies. Consistently, the TLR3 ligand dsRNA complexed to CALV and in combination with VLPs (CALV(dsRNA)+VLPs) induced the strongest response. CALV(dsRNA)+VLPs induced the highest titers against the recombinant vaccine antigens clade B Bal gp120 and pr55 Gag. Additionally, CALV(dsRNA)+VLPs induced cross-clade antibodies, represented by high titers of antibody to clade c 96ZM651 gp120. CALV(dsRNA)+VLPs induced predominantly IgG2c over IgG1, a response associated with T helper type 1 (Th1)-like cytokines. In turn, CALV(dsRNA)+VLP immunized mice generated the most potent neutralizing antibodies against HIV strain MN.3. Finally, at time of sacrifice, a significant increase in germinal center B cells and T follicular cells was detected in mice which received CALV(dsRNA)+VLPs compared to PBS. Our results indicate that CALV(dsRNA) is a superior adjuvant for HIV VLPs in generating a Th1-like immunoglobulin profile, while prolonging lymph node germinal centers, T follicular cells and generating neutralizing antibodies to a highly sensitive tier 1A variant of HIV.

Overexpression of Semaphorin-3E enhances pancreatic cancer cell growth and associates with poor patient survival.

Semaphorin-3E (Sema3E) is a member of an axon guidance gene family, and has recently been reported to contribute to tumor progression and metastasis. However, its role in pancreatic cancer is yet unknown and uncharacterized. In this study, we showed that Sema3E is overexpressed in human pancreatic cancer, and that high Sema3E levels are associated with tumor progression and poor survival. Interestingly, we also observed Sema3E expression in the nucleus, even though Sema3E is reported to be a secreted protein. Overexpression of Sema3E in pancreatic cancer cells promoted cell proliferation and migration in vitro, and increased tumor incidence and growth in vivo. Conversely, knockout of Sema3E suppressed cancer cell proliferation and migration in vitro, and reduced tumor incidence and size in vivo. Moreover, Sema3E induced cell proliferation via acting through the MAPK/ERK pathway. Collectively, these results reveal an undiscovered role of Sema3E in promoting pancreatic cancer pathogenesis, suggesting that Sema3E may be a suitable prognostic marker and therapeutic target for pancreatic cancer.

New polymer of lactic-co-glycolic acid-modified polyethylenimine for nucleic acid delivery.

AIM: To develop an improved delivery system for nucleic acids. MATERIALS & METHODS: We designed, synthesized and characterized a new polymer of lactic-co-glycolic acid-modified polyethylenimine (LGA-PEI). Functions of LGA-PEI polymer were determined. RESULTS: The new LGA-PEI polymer spontaneously formed nanoparticles (NPs) with DNA or RNA, and showed higher DNA or RNA loading efficiency, higher or comparable transfection efficacy, and lower cytotoxicity in several cell types including PANC-1, Jurkat and HEK293 cells, when compared with lipofectamine 2000, branched or linear PEI (25 kDa). In nude mouse models, LGA-PEI showed higher delivery efficiency of plasmid DNA or miRNA mimic into pancreatic and ovarian xenograft tumors. LGA-PEI/DNA NPs showed much lower toxicity than control PEI NPs in mouse models. CONCLUSION: The new LGA-PEI polymer is a safer and more effective system to deliver DNA or RNA than PEI.