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At least 991 records · Page 55

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DOE OSTI2017

The inhibition of lactate dehydrogenase A hinders the transcription of histone 2B gene independently from the block of aerobic glycolysis

Most cancer cells use aerobic glycolysis to fuel their growth and many efforts are made to selectively block this metabolic pathway in cancer cells by inhibiting lactate dehydrogenase A (LDHA). However, LDHA is a moonlighting protein which exerts functions also in the nucleus as a factor associated to transcriptional complexes. Here we found that two small molecules which inhibit the enzymatic activity of LDHA hinder the transcription of histone 2B gene independently from the block of aerobic glycolysis. Moreover, we observed that silencing this gene reduces cell replication, hence suggesting that the inhibition of LDHA can also affect the proliferation of normal non-glycolysing dividing cells. - Highlights: • Blocking aerobic glycolysis is an approach to impair proliferation of cancer cells. • Small inhibitors of LDHA block aerobic glycolysis. • LDHA is also involved in the transcription of histone 2B gene. • LDHA inhibitors block histone 2B transcription. • LDHA inhibitors can hinder the proliferation also of non-glycolysing normal cells.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2017

Age-related increase in Wnt inhibitor causes a senescence-like phenotype in human cardiac stem cells

Aging of cardiac stem/progenitor cells (CSCs) impairs heart regeneration and leads to unsatisfactory outcomes of cell-based therapies. As the precise mechanisms underlying CSC aging remain unclear, the use of therapeutic strategies for elderly patients with heart failure is severely delayed. In this study, we used human cardiosphere-derived cells (CDCs), a subtype of CSC found in the postnatal heart, to identify secreted factor(s) associated with CSC aging. Human CDCs were isolated from heart failure patients of various ages (2–83 years old). Gene expression of key soluble factors was compared between CDCs derived from young and elderly patients. Among these factors, SFRP1, a gene encoding a Wnt antagonist, was significantly up-regulated in CDCs from elderly patients (≥65 years old). sFRP1 levels was increased significantly also in CDCs, whose senescent phenotype was induced by anti-cancer drug treatment. These results suggest the participation of sFRP1 in CSC aging. We show that the administration of recombinant sFRP1 induced cellular senescence in CDCs derived from young patients, as indicated by increased levels of markers such as p16, and a senescence-associated secretory phenotype. In addition, co-administration of recombinant sFRP1 could abrogate the accelerated CDC proliferation induced by Wnt3A. Taken together, our results suggest that canonical Wnt signaling and its antagonist, sFRP1, regulate proliferation of human CSCs. Furthermore, excess sFRP1 in elderly patients causes CSC aging. - Highlights: • Wnt signaling regulates proliferation of human cardiac stem cells. • Expression of sFRP1, which is a Wnt antagonist, is up-regulated in elderly patients with heart failure. • Expression of sFRP1 is increased in anti-cancer drug-induced senescent human cardiac stem cells. • sFRP1 causes cellular senescence of young patients-derived cardiac stem cells.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2017

MicroRNA-193-5p modulates angiogenesis through IGF2 in type 2 diabetic cardiomyopathy

Background: Patients with diabetic cardiomyopathy are often associated with increasing risk of heart failure. In this work, we used animal model to characterize the angiogenic effect of microRNA-193-5p, miR-193-5p in type 2 diabetic Goto-Kakizaki (GK) rats' myocardial microvascular endothelial cells, MMEC(GK). Methods: MiR-193-5p in MMEC(GK) was compared to its expression in Wistar rat MMEC. In MMEC(GK), miR-193-5p was downregulated through viral infection. Its angiogenic effects on MMEC(GK) migration and proliferation were assessed by transwell and MTT assays, respectively. Downstream target of miR-193-5p, insulin growth factor 2 (IGF2), was assessed by dual-luciferase activity, qRT-PCR and western blot assays, respectively. In miR-193-5p-downregulated MMEC(GK), IGF2 was further de-regulated to assess its mechanism in miR-193-5p-downreuglation induced angiogenic regulation. Results: MiR-193-5p is overexpressed in MMEC(GK). Its downregulation has significantly angiogenic effect by inducing migration and proliferation in MMEC(GK). IGF2 was demonstrated to be directly regulated by miR-193-5p in MMEC(GK). In addition, IGF2 inhibition in miR-193-5p-downregulated MMEC(GK)’s severely hindered cell migration and proliferation. Conclusion: MiR-193-5p is an active angiogenic factor in diabetic cardiomyopathy, possibly through inverse regulation on its downstream IGF2 gene.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Umbilical cord-derived mesenchymal stem cells alleviated inflammation and inhibited apoptosis in interstitial cystitis via AKT/mTOR signaling pathway

Highlights: • UC-MSCs alleviated inflammation in rat model and cell model of IC. • UC-MSCs promoted proliferation of urothelium. • UC-MSCs inhibited apoptosis of urothelium. • UC-MSCs activated AKT/mTOR signaling pathway to cure IC. Interstitial cystitis (IC) is a bladder syndrome characterized by pelvic pain and urinary frequency without infection or other identifiable pathology. There are no effective treatments to cure IC. This study investigated the effects of human umbilical cord-derived mesenchymal stem cells (UC-MSCs) injection on IC rat model. Furthermore, we used a coculture system to find the possible molecular mechanism on the human uroepithelial cells (SV-HUC-1), which was the cell model of IC. A rat model of IC was established via systemic injection with cyclophosphamide (CYP) and a cell model of IC was induced by being exposed to tumor necrosis factor (TNF)-α (10 ng/ml). After one week, UC-MSCs injection significantly ameliorated the bladder voiding function in IC rat model. And the Histo- and immunohistochemical analyses showed that UC-MSCs can repair impaired bladder, reduce mast cell infiltration and inhibit apoptosis of urothelium. ELISA results showed that UC-MSCs can decrease IL-1β, IL-6 and TNF-α in bladder. In the coculture system, UC-MSCs can promote proliferation of impaired SV-HUC-1 cells, and inhibit apoptosis. However, while knocked down EGF secreted by UC-MSCs with siRNA, the effects would be weaken. Western blot showed that UC-MSCs increase protein expression levels of p-AKT and p-mTOR in SV-HUC-1 cells, and decrease the levels of cleaved caspase-3. Taken together, we provide evidence that UC-MSCs therapy can successfully alleviate IC in a preclinical animal Model and cell model by alleviating inflammation, promoting proliferation and inhibiting apoptosis. In addition, we demonstrate that the AKT/mTOR signaling pathway was activated.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Long non-coding RNA LUCAT1 modulates methotrexate resistance in osteosarcoma via miR-200c/ABCB1 axis

Highlights: • LncRNA LUCAT1 and ABCB1 protein are both up-regulated in osteosarcoma methotrexate-resistant cells. • LncRNA LUCAT1 knockdown suppress the expression levels drug resistance related genes, proliferation, invasion and tumor growth of osteosarcoma cells in vitro and vivo. • Bioinformatics predictive tools and luciferase assay reveal the LUCAT1/miR-200c/ABCB1 pathway. • Rescue experiments confirm the combined role of LUCAT1, miR-200c and ABCB1 on osteosarcoma methotrexate resistance. Long non-coding RNAs (lncRNAs) have been verified to participate in the tumorigenesis of multiple cancers. Nevertheless, the deepgoing role molecular mechanisms of lncRNAs on osteosarcoma chemoresistance remain unclear. In present study, we investigate the function of lncRNA LUCAT1 on osteosarcoma methotrexate (MTX) resistant phenotype and discover the potential regulatory mechanism. Results showed that LUCAT1 was up-regulated in MTX-resistant cells (MG63/MTX, HOS/MTX) compared to that in parental cells. LncRNA LUCAT1 and ABCB1 protein expression levels were both up-regulated when induced by different concentration of methotrexate. In vitro and vivo, LUCAT1 knockdown decreased the expression levels drug resistance related genes (MDR1, MRP5, LRP1), proliferation, invasion and tumor growth of osteosarcoma cells. Bioinformatics tools and luciferase assay reveled that miR-200c both targeted the 3′-UTR of LUCAT1 and ABCB1 mRNA, suggesting the modulation of LUCAT1 on ABCB1 through sponging miR-200c. Rescue experiments confirmed the combined role of LUCAT1, miR-200c and ABCB1 on osteosarcoma proliferation, invasion and methotrexate resistance. Overall, results indicate the vital role of LUCAT1 in the methotrexate resistance regulation through miR-200c/ABCB1 pathway, providing a novel insight and treatment strategy for osteosarcoma drug resistance.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Human adipose-derived stem cells and simvastatin-functionalized biomimetic calcium phosphate to construct a novel tissue-engineered bone

Highlights: • Performance of a strategy of constructing a novel tissue-engineered bone (TEB). • Simvastatin could release in a sustained manner after incorporated in the TEB. • This novel TEB could promote the bone formation capability of hASCs more effectively. To repair bone defects, we evaluate the in-vitro and in-vivo osteogenic activities of a novel tissue-engineered bone (TEB) by elaborately combining biomimetic calcium phosphate (BioCaP) granules with internally-incorporated simvastatin (SIM) and human adipose-derived stem cells (hASCs). First, we constructed BioCaP with SIM internally incorporated (SIM-BioCaP). Then we characterized the morphology and chemical composition of SIM-BioCaP. The release kinetics of SIM was monitored in vitro spectroscopically. Thereafter, we explored the in-vitro cellular responses of hASCs to SIM-BioCaP by performing scanning electron microscopy observation, proliferation assay, alkaline phosphatase (ALP) activity assay, alizarin red staining and real-time PCR. Finally, we investigated the in-vivo osteogenic activities of the novel TEB in a subcutaneous bone induction model in nude mice. We found that SIM was successfully incorporated internally in BioCaP and showed a slow release manner without significantly affecting the attachment and proliferation of hASCs. The released SIM from BioCaP could significantly enhance the proliferation, ALP activities, mineralized nodules formation and osteogenic genes of hASCs. The in-vivo tests showed this TEB could induce new bone formation while the other groups could not. Taken together, the present data show that this novel TEB represented a very promising construct to treat critical-volume bone defects.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Effects of the RNA-binding protein, KSRP, on innate immune response against Helicobacter pylori infection in mice

Highlights: • H. pylori infection decreased expression of KSRP. • KSRP overexpression triggered HMOX1 and CXCL-2 ectopic expression. • KSRP overexpression facilitated H. pylori proliferation and colonization. • KSRP overexpression triggered more severe gastric mucosal damage. Helicobacter pylori (H. pylori) contributes to various gastric diseases such as chronic gastritis, gastric ulcer, and gastric carcinoma. Host innate immune response against the pathogen plays a significant role in elimination of pathogen infection. Importantly, pathogen elimination is closely related to numerous inflammatory-related genes that participate in complex biological response of cells to harmful stimuli. Here we studied effects of the KH-type splicing regulatory protein (KSRP), a RNA-binding protein, on innate immune response against H. pylori infection. We found that H. pylori infection downregulated KSRP expression directly, and that KSRP overexpression repressed upregulation of CXCL-2 expression induced by H. pylori and facilitated H. pylori proliferation in vitro. Similarly, KSRP overexpression in H. pylori mice also facilitated H. pylori proliferation and colonization, and induced more severe gastric mucosal damage. Intriguingly, CXCL-2 and HMOX-1 were upregulated in H. pylori infected mice after KSRP overexpression. This difference in expression of these genes implicated that KSRP was closely associated with and directly participated in the innate immune response against H. pylori. These results were beneficial for understanding the in vivo function of KSRP on innate immune response against pathogen infection.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Angiotensin II type 1 receptor-associated protein regulates carotid intimal hyperplasia through controlling apoptosis of vascular smooth muscle cells

Highlights: • ATRAP as a negative regulator of AT1 receptor plays an inhibitory role in VSMCs proliferation both in vitro and in vivo. • ATRAP triggers the marked apoptosis of VSMCs in balloon-injuried carotid arteries and depresses neointimal formation. • ATRAP induces VSMCs apoptosis through the influence of the PI3K/Akt signaling pathway. Intimal hyperplasia is the main cause of restenosis after carotid artery injury, and the underlying mechanism involves the proliferation and migration of vascular smooth muscle cells (VSMCs). Angiotensin II Type 1 Receptor-Associated Protein (ATRAP) has been reported to withstand intimal hyperplasia by inhibiting VSMCs proliferation and migration; however, whether the beneficial effect of ATRAP associates with VSMCs apoptosis remains unclarified. We demonstrated that the adenoviral-mediated overexpression of ATRAP induced VSMC apoptosis, alleviating the balloon injury-induced neointima formation in rats. Under the condition of Angiotensin-II stimulation, ATRAP overexpression induced the apoptosis of rat VSMCs by depressing the PI3K-Akt signaling; whereas up-regulation of Akt by PTEN inhibitor abolished the apoptotic death. Thus, ATRAP regulates carotid intimal hyperplasia through controlling the PI3K-Akt signal-mediated VSMCs apoptosis.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Tsc1 controls the development and function of alveolar macrophages

Highlights: • Deletion of Tsc1 leads to the impaired development and a great reduction of AMs. • Tsc1 suppresses the proliferation of AMs. • Tsc1-deficient AMs exhibits increased ROS production and phagocytosis. Alveolar macrophages (AMs) are pivotal for maintaining the lung homeostasis, but how the development and function of AMs regulated remains largely unknown. In the present study, we demonstrated that the number of AMs was controlled by the Tsc1 protein. Cd11c-specific deletion of Tsc1 caused inefficient transition from pre-AMs to AMs in lung, which led to a great reduction of AM population. Ablation of Tsc1 downregulated the expression of surface marker CD64 and SiglecF on AMs. We further showed that conditional knockout of Tsc1 led to enhanced proliferation and increased reactive oxygen species (ROS) production and phagocytosis in AMs. These results indicated that Tsc1 was a critical regulator of development, proliferation and function in AMs.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Inhibition of MALAT1 sensitizes liver cancer cells to 5-flurouracil by regulating apoptosis through IKKα/NF-κB pathway

Highlights: • MALAT1 knockdown inhibits liver cancer cell proliferation. • MALAT1 silence enhances apoptosis and liver cancer cells sensibility to 5-FU incubation. • MALAT1 deficiency related increase in sensitivity of liver cancer cells was associated with regulation of NF-κB. • MALAT1 knockdown reduces tumor growth in vivo. Metastasis associated lung adenocarcinoma transcript 1 (MALAT1) is involved in tumor cell growth process. However, its role and molecular mechanism in liver cancer is still not fully understood. In this study, we found that MALAT1 was significantly expressed in liver cancer cell lines. And knockdown of MALAT1 suppressed proliferation, migration and invasion of HepG2 cells, accompanied with decrease of Rho-associated coiled-coil-forming protein kinase 1 (ROCK1), α-smooth muscle actin (α-SMA), N-cadherin, Vimentin and TWIST. Significantly, MALAT1 deletion sensitized HepG2 cells to 5-FU-induced cell cycle arrest in G1 phase, as evidenced by the significant reduction in Cyclin D1 and CDK4 and increase in p53, p21 and p27 protein levels. In addition, MALAT1 knockdown triggered 5-FU induced apoptosis in HepG2 cells by inducing intrinsic apoptosis-related signals, including Cyto-c, Apaf-1, cleaved Caspase-9/-7/-3 and poly (ADP-ribose) polymerase (PARP). Furthermore, phosphorylated nuclear factor-κB (p-NF-κB) was also down-regulated by MALAT1 silence. Importantly, suppression of IKKα/NF-κB significantly elevated apoptosis and reduced liver cancer cell viability in MALAT1-knockdown cells with 5-FU incubation. The nude mice transplantation model also confirmed the promoted sensitivity of MALAT1-silenced HepG2 cells to 5-FU by blocking tumor cell proliferation and inducing apoptosis. Therefore, our data supplied a potential mechanism by which knockdown of MALAT1 might play an important role in augmenting sensitivity of HepG2 cells to 5-FU in therapeutic approaches, demonstrating suppressing of MALAT1 may serve as a combination with chemotherapeutic agents in liver cancer treatment.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Long non-protein coding RNA DANCR functions as a competing endogenous RNA to regulate osteoarthritis progression via miR-577/SphK2 axis

Highlights: • DANCR was increased in OA. • DANCR inhibition suppressed OA chondrocytes proliferation and induced cell apoptosis. • DANCR-miR-577/SphK2 axis play important roles in OA. Long noncoding RNAs (lncRNAs) have been known to be involved in multiple diverse diseases, including osteoarthritis (OA). This study aimed to explore the role of differentiation antagonizing non-protein coding RNA (DANCR) in OA and identify the potential molecular mechanisms. The expression of DANCR in cartilage samples from patients with OA was detected using quantitative reverse transcription–polymerase chain reaction. The effects of DANCR on the viability of OA chondrocytes and apoptosis were explored using cell counting kit 8 assay and flow cytometry assay, respectively. Additionally, the interaction among DANCR, miR-577, and SphK2 was explored using dual-luciferase reporter and RIP assays. The present study found that DANCR was significantly upregulated in patients with OA. Functional assays demonstrated that DANCR inhibition suppressed the proliferation of OA chondrocytes and induced cell apoptosis. The study also showed that DANCR acted as a competitive endogenous RNA to sponge miR-577, which targeted the mRNA of SphK2 to regulate the survival of OA chondrocytes. In conclusion, the study revealed that lncRNA DANCR might promote the proliferation of OA chondrocytes and reduce apoptosis through the miR-577/SphK2 axis. Thus, lncRNA DANCR might be considered as a potential therapeutic target for OA treatment.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Influence and mechanism of Angiotensin 1-7 on biological properties of normal prostate epithelial cells

Highlights: • Ang1-7 inhibit proliferation and EMT of prostate epithelial cells by inactivation of PI3K axis. • Ang1-7 can counteract oxidant stress in prostate by inhibition of VEGF and upregulation of NOS3. • Ang1-7 increase the mRNA level of antiapoptotic genes: BIRC5 and BCL-2 in PNT1A cells. • Ang1-7 can upregulate not only AT1-7/MAS expression but also classic angiotensin receptors. • Ang1-7 can upregulate the AR and ESR2 receptor but dowregulate ESR1 in PNT1A. The ACE2/Ang1-7/MAS axis was involved in the cell proliferation, migration and apoptosis of many types of reproductive tissues. The research was conducted on prostate epithelial cells, immortalized by Simian Virus 40. We examined the influence of Ang 1–7 on biological properties of PNT1A cells after 24- or 48-h treatment. The employed selective antagonists of angiotensin receptors allowed evaluation of the receptor mediating Ang1-7 action. Our data clearly indicate that Ang1-7 can decrease cell proliferation and epithelial-to-mesenchymal transition of PNT1A cells via inactivation of PI3K axis and modulation of expression of the NF-kB gene family. Furthermore, it counteracts oxidant stress and inflammation in prostate cells by inhibition of VEGF expression and MMPs activation as well as by modulating the level of ERα and ERβ. On the other hand, this heptapeptide can promote cell survival by alteration of expression of anti- and pro-apoptotic members as well as compensatory up-regulation of AR expression. Summary, the results confirm the existence of a complicated dependence networks between the various elements of the local RAS and steroid hormone receptor pathways in prostate gland. Furthermore, shows the chances of using ACE2/Ang1-7/MAS pathway as a novel therapeutic target in prevention and treatment of prostate diseases.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

FOXP4-AS1 participates in the development and progression of osteosarcoma by downregulating LATS1 via binding to LSD1 and EZH2

Highlights: • We first confirmed that FOXP4-AS1 is overexpressed in OS, and is the independent risk factor in OS prognosis. • We further clarified that Upregulated FOXP4-AS1 promotes the proliferation, migration and cell cycle, but inhibits apoptosis of OS cells. • Finally, we demonstrated that FOXP4-AS1 participates in the development and progression of OS by downregulating LATS1. Osteosarcoma (OS) is the most common malignant bone tumor in children and adolescents. LncRNA has been confirmed to participate in a variety of cancers. The purpose of this study was to explore the effect of FOXP4-AS1 on the development of osteosarcoma (OS) and its underlying mechanism. FOXP4-AS1 expressions in 60 OS tissues and paracancerous tissues were detected by qRT-PCR (quantitative real-time polymerase chain reaction). We confirmed that FOXP4-AS1 was overexpressed in OS tissues than that of paracancerous tissues. The disease-free survival and overall survival of OS patients were not correlated with age, gender and tumor location, but remarkably correlated with FOXP4-AS1 expression, tumor size and lung metastasis. For in vitro experiments, MG63 cells expressed a higher expression of FOXP4-AS1, whereas U2OS cells expressed a lower expression, which were selected for the following studies. Overexpressed FOXP4-AS1 led to enhanced proliferation, migration and invasion, shortened G0/G1 phase, as well as inhibited cell cycle. Knockdown of FOXP4-AS1 in MG63 cells obtained the opposite results. Furthermore, RIP assay indicated that FOXP4-AS1 could inhibit LATS1 expression by binding to LSD1 and EZH2, so as to participate in OS development. In conclusion, these results revealed that FOXP4-AS1 is overexpressed in OS, and is the independent risk factor in OS prognosis. Upregulated FOXP4-AS1 promotes the proliferation, migration and cell cycle, but inhibits apoptosis of OS cells. Furthermore, FOXP4-AS1 participates in the development and progression of OS by downregulating LATS1 via binding to LSD1 and EZH2.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

CAR-mediated repression of Cdkn1a(p21) is accompanied by the Akt activation

Highlights: • CAR-mediated liver growth is accompanied by the decrease in PTEN. • CAR-mediated liver growth is accompanied by the Akt activation. • CAR-mediated Akt activation is accompanied by the decrease in Foxo1. • CAR activation decreased the level of p21. It was shown that CAR participates in the regulation of many cell processes. Thus, the activation of CAR causes a proliferating effect in the liver, which provides grounds to consider CAR as a therapeutic target when having a partial resection of this organ. Even though a lot of work has been done on the function of CAR in regulating hepatocyte proliferation, very little has been done on its complex mediating mechanism. This study, therefore, showed that the liver growth resulting from CAR activation leads to the decline in the level of PTEN protein and subsequent Akt activation in mouse liver. The increase of Akt activation produced by CAR agonist was accompanied by a decrease in the level of Foxo1, which was correlated with decreased expression of Foxo1 target genes, including Cdkn1a(p21). Moreover, the study also demonstrated that there exists a negative regulatory impact of CAR on the relationship between Foxo1 and targeted Cdkn1a(p21) promoter. Therefore, the study results revealed an essential function of CAR-Akt-Foxo1 signalling pathway in controlling hepatocyte proliferation by repressing the cell cycle regulator Cdkn1a (p21).

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

MiR-455-3p suppresses renal fibrosis through repression of ROCK2 expression in diabetic nephropathy

Highlights: • Downregulation of miR-455-3p expression in DN. • ROCK2 as a target of miR-455-3p. • Re-introduction of ROCK2 attenuated the effect of miR-455-3p on cell proliferation, ECM accumulation and EMT in vitro. • Reductions in proinflammatory levels and renal fibrosis by miR-455-3p usages, in vivo. Emerging evidence has shown that microRNAs (miRNAs) play a mediatory role in the pathogenesis of diabetic nephropathy (DN), but the function of the involved miRNAs is still incomplete. Here, we found that miR-455-3p was down-regulated in the human mesangial cells (HMC) and human proximal tubule epithelial cells (HK-2) stimulated with high glucose (HG) or transforming growth factor beta 1 (TGF-β1). Rho-associated coiled coil-containing protein kinase 2 (ROCK2) was identified as a directed target of miR-455-3p. Overexpression of ROCK2 significantly attenuated the inhibitory effects of miR-455-3p on cell proliferation, extracellular matrix (ECM) synthesis and epithelial-mesenchymal transition (EMT) in HG-treated cells. Furthermore, the DN model was prepared by using high-fat feeding combined with Streptozotocin (STZ) induced rats, and the DN group was treated by injecting miR-455-3p agomir. The results of periodic acid-Schiff (PAS) and Masson staining showed that miR-455-3p overexpression improved the pathological changes of glomerular hypertrophy, mesangial amplification, and renal fibrosis. Additionally, miR-455-3p overexpression decreased ROCK2, proliferating cell nuclear antigen (PCNA) and Collagen I levels, and also reduced inflammatory cytokines TNF-α, MCP-1 and IL-1β levels in vivo. Altogether, these results suggest that miR-455-3p plays an essential role in the treatment of renal fibrosis through repressing ROCK2 expression; and miR-455-3p might be an effective therapy for DN.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Effects of amiloride on physiological activity of stem cells of human lung cancer and possible mechanism

Lung cancer is a common malignant tumor, the cancer stem cells (CSCs) were regarded responsible for the development of cancer tissue. The effects of amiloride on lung cancer stem cells and the possible mechanism were not much investigated. In this study, human NCI-H1975 lung CSCs were selected by flow cytometry, and the effects of amiloride at different concentrations (0, 12.5, 25, 50, and 100 μmol/L) were evaluated on proliferation, migration, invasion and apoptosis of CSCs using cell counting kit-8 and Transwell migration assays as well as flow cytometry. Wstern blot analysis was performed to investigate the effect of amiloride on the level of proteins in uPA system, NF-kB pathway, and PI3K-AKT-mTOR pathway in CSCs. As a result, we found that amiloride inhibited proliferation, migration and invasion of lung CSCs, and promoted apoptosis. Further, we found that amiloride decreased levels of target proteins in the uPA system, as well as the NF-kB and PI3K-AKT-mTOR pathways. These results indicated that amiloride could inhibit proliferation, migration and invasion of lung CSCs, and promotes apoptosis, these effects may be related to decreased levels of proteins in the uPA system, the NF-kB pathway, and the PI3K-AKT-mTOR pathway.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

ANGPTL2 deletion inhibits osteoclast generation by modulating NF-κB/MAPKs/Cyclin pathways

Highlights: • ANGPTL2 is associated with the osteoclast development. • Inhibition of ANGPTL2 reduces the proliferation of osteoclast precursors. • ANGPTL2 deficiency alleviates ovariectomy (OVX)-induced bone loss. Osteoclasts are multinucleated cells essential for bone-resorption. Successful repair of bone defciencies still remains a great challenge worldwide. The signaling factor angiopoietin-like protein 2 (ANGPTL2), one of eight ANGPTL proteins, functions in maintenance of tissue homeostasis partly through regulating inflammation. In the study, ANGPTL2 expression was promoted during osteoclast development and that suppressing ANGPTL2 alleviated osteoclast production regulated by macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-κB ligand (RANKL). The results suggested that ANGPTL2 knockdown inhibited M-CSF-caused proliferation of osteoclast precursor cells. Further, ANGPTL2 silence reduced nuclear factor of activated T cell c 1 (NFATC1) and NFATC4 expressions in M-CSF-treated cells, along with decreased Runx2, OPN and Colla1. Moreover, silencing ANGPTL2 down-regulated M-CSF-promoted expressions of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, and chemoattractant protein-1 (CCL-2). Consistently, ANGPTL2 knockdown reduced M-CSF-enhanced activation of IKKα, IκBα and nuclear factor κB (NF-κB) and mitogen-activated protein kinases (MAPKs) (p38 MAPK, ERK1/2 MAPK and JNK MAPK). Additionally, knockdown of ANGPTL2 inhibited the induction of Cyclin D1, Cyclin D2 and Cyclin E1 due to M-CSF exposure. In vivo, we confirmed that ANGPTL2 knockout (KO) mice were protected against osteoporosis induced by ovariectomy (OVX), as proved by the improved bone loss and bone mineral density (BMD). Decreased expression of NFATCs was also observed in OVX-induced mice in the absence of ANGPTL2. Elevated release of pro-inflammatory cytokines was abrogated by ANGPTL2 knockout in femoral heads of mice with OVX operation, accompanied with a significant reduction of phosphorylated NF-κB and MAPKs signaling pathways. And down-regulated expression of Cyclin D1, Cyclin D2 and Cyclin E1 was observed in OVX-operated mice with ANGPTL2 knockout. Therefore, our study indicated that ANGPTL2 played an essential role in osteoclast generation through regulating the proliferation and inflammation of osteoclast lineage cells, providing new insights into the therapeutic strategy to alleviate bone loss.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

ZIC5 facilitates the growth of hepatocellular carcinoma through activating Wnt/β-catenin pathway

Highlights: • High ZIC5 expression in specimens indicates poor prognosis of HCC patients. • ZIC5 was required in proliferation and tumorigenicity of HCC. • ZIC5 promotes proliferation and tumorigenicity of HCC cells. • ZIC5 promoted growth of HCC in a β-catenin dependant manner. The incidence and mortality of hepatocellular carcinoma (HCC) is high, but the mechanisms underlying the growth and progression of HCC have not been elucidated. Recently, the ZIC family member 5 (ZIC5) is emerging as an oncogene in various types of tumors. However, its expression and biological role in HCC have not been reported. This study first demonstrated that ZIC5 was up-regulated in HCC specimens, and high ZIC5 expression indicated poor prognosis of HCC patients. In addition, over-expressed ZIC5 promoted the proliferation, migration and invasion of HCC cell lines Huh7 and HepG2 in vitro and in vivo, while ZIC5 knockdown achieved the opposite effects. Actually, ZIC5 increased the expression of genes participating in Wnt/β-catenin pathway such as β-catenin and CyclinD1. ZIC5 also promoted β-catenin to enter the nucleus of HCC cells. Furthermore, silencing β-catenin abated the promoting role of ZIC5 in HCC. Overall, this study reveals a novel mechanism of ZIC5/β-catenin that mediates the invasion and metastasis of HCC and ZIC5 serves as a novel indicator for prognosis of HCC patients.

60 APPLIED LIFE SCIENCES↗