establishing-situ-closed-circuit-perfusion-lower-abdominal-organs

Establishing In Situ Closed Circuit Perfusion of Lower Abdominal Organs and Hind Limbs in Mice

Ex vivo perfusion is an important physiological tool to study the function of isolated organs (e.g. liver, kidneys). At the same time, due to the small ...

The First Hospital of Jilin University

University of Colorado Anschutz Medical Campus

Jilin University

X-linked inhibitor of apoptosis protein (XIAP) has been shown to be highly expressed in lung cancer, but not in normal lung tissue, which makes it an attractive target for lung cancer treatment. Celecoxib (CXB), a cyclooxygenase-2 inhibitor, is in wide clinical use for the treatment and prevention of non-small cell lung cancer (NSCLC). Therefore, in our study, we combined short hairpin RNA (shRNA) targeted to XIAP (XIAP-shRNA) with CXB and tested the effects of this combination on lung cancer cells to identify more effective therapeutics against lung cancer. An XIAP-shRNA plasmid was constructed and transfected into the A549 NSCLC cell line. The cells were then treated with CXB and XIAP-shRNA alone or in combination for indicated time periods, and the treatments were assessed for their effects on cell proliferation, apoptosis, migration, invasion and receptor signaling using the MTT, TUNEL, wound healing and Matrigel invasion assays and western blotting, respectively. In addition, an NSCLC xenograft model was prepared to observe tumor growth. It was found that both CXB and XIAP‑shRNA significantly inhibited cell proliferation, migration and invasion, and induced cell apoptosis in vitro, as well as suppressed tumor growth in vivo. Moreover, the combination of the agents significantly enhanced these effects compared to the single agent treatments. We also found that the combination treatment significantly suppressed constitutive phosphorylation of PI3K and AKT, which may contribute to the inhibition of tumor growth. These findings suggest that the combination of XIAP‑shRNA and CXB is a promising drug candidate for the treatment of NSCLC.

Combined treatment of XIAP-targeting shRNA and celecoxib synergistically inhibits the tumor growth of non‑small cell lung cancer cells in vitro and in vivo.

In this study, 5-fluorouracil (5-FU) and LY294002 (LY)-loaded PEGylated nanoliposome was prepared to target esophageal squamous cell carcinoma (ESCC). The particles were characterized in terms of physicochemical and biological parameters. The co-delivery of autophagy inhibitor and chemotherapeutic drug in a single carrier was successfully accomplished. The two components from 5-FU and LY-loaded PEGylated nanoliposome (FLNP) released in a controlled manner with LY relatively released faster compared to that of 5-FU. FLNP showed a receptor-mediated cellular uptake that will allow the gradual release of drug in the acidic environment. The cellular uptake of nanoparticles (NP) was further confirmed by FACS analysis. The combination of 5-FU and LY resulted in higher cytotoxic effect compared to that of individual drugs. Most importantly, FLNP exhibited a significantly higher anticancer effect in cancer cells compared to that of free cocktail combinations. The faster release of LY from FLNP leads to autophagy inhibition that improves the sensitivity of cancer cells towards 5-FU, resulting in more cell death. Consistently, FLNP induced a greater apoptosis (~ 48%) of cancer cells compared to that of any other groups. Western blot analysis clearly showed that 5-FU and LY individually increased the expression of caspase-3 and PARP, while as expected FLNP induced a remarkable expression of these protein markers indicating the superior anticancer effect. We believe that the programmed release of autophagy inhibitor and chemotherapeutic drug from a single nanocarrier will increase the prospect of anticancer therapy in ESCC.

Autophagy Inhibitor (LY294002) and 5-fluorouracil (5-FU) Combination-Based Nanoliposome for Enhanced Efficacy Against Esophageal Squamous Cell Carcinoma.

Radioresistance hinders the therapeutic outcomes of radiotherapy in non‑small cell lung cancer (NSCLC). Although long non‑coding RNAs (lncRNAs) have been demonstrated to participate in the regulation of multiple cell behaviors, whether they can modulate the radiosensitivity of NSCLC and the underlying molecular mechanisms have not been well investigated. In the present study, it was revealed that NSCLC NCI‑H460 cells were more sensitive to ionizing radiation (IR) than A549 cells. Using the RNA‑Seq method, four highly differentially expressed lncRNAs were identified, including the growth arrest‑specific transcript 5 (GAS5), syntaxin binding protein 5 antisense RNA 1 (STXBP5‑AS1), metastasis associated lung adenocarcinoma transcript 1 (MALAT1) and X‑inactive specific transcript (XIST), which were predicted to play roles in the acquisition of radiosensitivity. Using real‑time quantitative PCR (qPCR), it was demonstrated that lncRNA GAS5 was significantly upregulated in NCI‑H460 cells but not in A549 cells during IR. Mechanistically, it was demonstrated that overexpression of lncRNA GAS5 decreased the level of microRNA‑21 (miR‑21). Overexpression of lncRNA GAS5 or suppression of miR‑21 markedly increased the IR‑induced cell apoptosis of A549 cells. It was also demonstrated that overexpression of lncRNA GAS5 increased PTEN expression and suppressed Akt phosphorylation through the modulation of miR‑21. Notably, it was revealed that IR enhanced the interaction between lncRNA GAS5 and the miR‑21/PTEN/Akt axis. In summary, the present findings revealed that lncRNA GAS5 has a radiosensitization effect on NSCLC, indicating the potential application of lncRNA GAS5 in NSCLC radiotherapy.

Long non‑coding RNA GAS5 increases the radiosensitivity of A549 cells through interaction with the miR‑21/PTEN/Akt axis.

Lung adenocarcinoma (LUAD) is one of the most common diagnosed pathological categories of lung cancer. Long noncoding RNAs (lncRNAs) have been manifested to be key regulators in modulating multiple cancers. Nevertheless, the pathologic role of lncRNA WDFY3-AS2 in LUAD remains elusive. The relative messenger RNA and protein levels were assessed by quantitative reverse transcription-polymerase chain reaction and Western blot analyses, respectively. Colony formation, carboxyfluorescein succinimidyl ester, terminal deoxynucleotidyl transferase dUTP nick-end labeling, wound-healing, and transwell invasion assays were performed to study the underlying role of WDFY3-AS2 in LUAD. Luciferase reporter assay, chromatin immunoprecipitation, RNA pull down, and RNA immunoprecipitation assays were conducted to probe into the interactions between relevant genes. WDFY3-AS2 expression was elevated in LUAD and WDFY3-AS2 transcription was activated by transcription factor USF1. Silencing WDFY3-AS2 could suppress cell proliferation, migration, and invasion, whereas accelerate cell apoptosis in LUAD. Molecular mechanism assays revealed that WDFY3-AS2 could bind to miR-491-5p and miR-491-5p inhibition could reverse the inhibitory effect of WDFY3-AS2 silence on LUAD progression. Besides, zinc finger protein 703 (ZNF703) was identified as a downstream target of miR-491-5p and its expression could be upregulated by WDFY3-AS2. Further, rescue assays uncovered that ZNF703 overexpression could restore the suppressive influence of silenced WDFY3-AS2 on LUAD development. USF1-acitvated WDFY3-AS2 promotes LUAD progression via targeting miR-491-5p/ZNF703 axis, suggesting the potential value of WDFY3-AS2 as a novel target for LUAD treatment.

USF1-induced overexpression of long noncoding RNA WDFY3-AS2 promotes lung adenocarcinoma progression via targeting miR-491-5p/ZNF703 axis.

Developing wound dressing that inhibits bacterial infection for treating complex wound healing processes has been a research hotspot. Here, we report the fabrication of Cu-MOFs (HKUST-1) incorporated electrospun chitosan/polyvinyl alcohol (HKUST-1/chitosan/PVA) fibers through the blending electrospinning for wound therapy. HKUST-1/chitosan/PVA fibers displayed satisfying physical properties, such as mechanical property, water uptake, water vapor transmission rate, etc. Cytotoxicity test indicated that HKUST-1/chitosan/PVA fibers were biocompatible and could support cell adhesion. Due to the HKUST-1 incorporation, HKUST-1/chitosan/PVA fibers exhibited the good antibacterial activity against Escherichia coli and Staphylococcus aureus with 99% antibacterial efficiency. Furthermore, in animal studies, compared with commercial chitosan dressings and chitosan/PVA fibers, HKUST-1/chitosan/PVA fibers were more efficient to heal the wound with less inflammation. In summary, the HKUST-1/chitosan/PVA fibers with good physicochemical property, biocompatibility and antibacterial property is an excellent wound dressing for full-thickness skin repair.

Incorporation of metal-organic frameworks into electrospun chitosan/poly (vinyl alcohol) nanofibrous membrane with enhanced antibacterial activity for wound dressing application.

Epidermal growth factor receptor- tyrosine kinase inhibitors (EGFR-TKIs) have shown promise against non-small cell lung cancers (NSCLCs) in clinics but the utility is often short-lived because of T790M mutations in EGFR that help evade TKIs' action. Osimertinib is the third and latest generation TKI that targets EGFRs with T790M mutations. However, there are already reports on acquired resistance against Osimertinib. Recent work has revealed the role that miRNAs, particularly tumor suppressor let-7c, play in the invasiveness and acquired resistance of NSCLCs, but the mechanistic details, particularly in Osimertinib resistance, remain elusive. Using two cells lines, H1975 (endogenous T790M mutation) and HCC827-T790M (with acquired T790M mutation), we found that let-7c is a regulator of EMT, as well as it affects CSC phenotype. In both the cell lines, transfection with pre-let-7c led to reversal of EMT as studied through EMT markers e-cadherin and ZEB1. This resulted in reduced proliferation and invasion. Conversely, reduced expression of let-7c through anti-let-7c transfections significantly increased proliferation and invasion of lung cancer cells. Expression of let-7c was functionally relevant as EMT correlated with resistance to Osimertinib. High let-7c expression reversed EMT and made cells sensitive to Osimertinib, and vice versa. WNT1 and TCF-4 were found to be two targets of let-7c which were epigenetic suppressed by let-7c through increased methylation. In vivo, pre-let-7c inhibited while anti-let-7c potentiated tumor growth and WNT1 and TCF-4 were downregulated in xenografts with pre-let-7c. Silencing of both WNT1 and TCF-4 resulted in potentiation of Osimertinib action. Our results suggest an important role of let-7c in regulating EMT and the resulting Osimertinib resistance in T790M NSCLCs. More clinical studies need to be performed to fully understand the translational relevance of this novel mechanism.

Let-7c regulated epithelial-mesenchymal transition leads to osimertinib resistance in NSCLC cells with EGFR T790M mutations.

Non-small cell lung cancer (NSCLC) is a type of cancer that is associated with high prevalence and high mortality rates in China. Therefore, it is of importance to identify the mechanisms underlying NSCLC progression. In the present study, reverse transcription-quantitative PCR was performed to measure the expression of microRNA (miR)-365b in NSCLC cell lines. In addition, the biological roles of miR-365b and N-acetylgalactosaminyltransferase 4 (GALNT4) were investigated by manipulating the expression levels of miR-365b and GALNT4 in NSCLC cells. It was found that miR-365b expression was reduced in NSCLC tissues and cells. Overexpression of miR-365b inhibited NSCLC cell proliferation whilst promoting apoptosis, but miR-365b knockdown promoted NSCLC cell proliferation. In addition, it was demonstrated that miR-365b regulated the proliferation and apoptosis of NSCLC cells by targeting GALNT4 expression. Collectively, the present study identified a miR-365b/GALNT4 regulatory axis in NSCLC, suggesting that miR-365b may serve as a therapeutic target for NSCLC.

Article	Year
Combined treatment of XIAP-targeting shRNA and celecoxib synergistically inhibits the tumor growth of non‑small cell lung cancer cells in vitro and in vivo. Oncology reports\| PubMed ID: 25524805	2015
Autophagy Inhibitor (LY294002) and 5-fluorouracil (5-FU) Combination-Based Nanoliposome for Enhanced Efficacy Against Esophageal Squamous Cell Carcinoma. Nanoscale research letters\| PubMed ID: 30328537	2018
Long non‑coding RNA GAS5 increases the radiosensitivity of A549 cells through interaction with the miR‑21/PTEN/Akt axis. Oncology reports\| PubMed ID: 32020207	2020
USF1-induced overexpression of long noncoding RNA WDFY3-AS2 promotes lung adenocarcinoma progression via targeting miR-491-5p/ZNF703 axis. Molecular carcinogenesis\| PubMed ID: 32275336	2020
Incorporation of metal-organic frameworks into electrospun chitosan/poly (vinyl alcohol) nanofibrous membrane with enhanced antibacterial activity for wound dressing application. International journal of biological macromolecules\| PubMed ID: 32353508	2020
Let-7c regulated epithelial-mesenchymal transition leads to osimertinib resistance in NSCLC cells with EGFR T790M mutations. Scientific reports\| PubMed ID: 32641854	2020
miR-365b regulates the development of non-small cell lung cancer via GALNT4. Experimental and therapeutic medicine\| PubMed ID: 32765677	2020

Ping Ren

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