bluuebunny/crossref_metadata_2025_split huggingface dataset:
FOXA2 PROMOTES PROSTATE CANCER BONE COLONIZATION
Authors: connelly zachary m. , Yang Shu , Orr A. Wayne , Yu Xiuping 
Published: June 2021

BackgroundIt is estimated that 26,000 men will die from prostate cancer (PCa) in 2017. Androgen deprivation therapy (ADT) is the gold standard treatment for advanced PCa. Initially, patients respond to treatment, however, these tumors almost invariably progress to castrate‐resistant PCa, for which there is no cure. Most PCa patients who fail ADT develop metastasis and preferentially relocate to the bones. Bone metastasis results in significant morbidity and mortality as the average time to death is approximately 3–5 years with no treatment available. Understanding the mechanisms by which PCa grow in the bone is critical for the development of novel therapeutics to treat and decrease tumor‐mediated bone destruction.MethodsGene expression profiling studies have found that FOXA2, a forkhead transcription factor that is expressed in embryonic prostate and neuroendocrine PCa, is present in a subset of metastatic PCa specimens. Our preliminary study found FOXA2 expression in a sample set of human PCa bone metastases, suggesting an involvement of FOXA2 in human PCa bone metastases. Also, we found high levels of FOXA2 in aggressive PCa PC3 cells, but not in PCa LNCaP cells. PC3 cells generate osteolytic lesions in bone, whereas LNCaP cells minimally grow following bone inoculation. To establish FOXA2's role in promoting PCa metastasis, FOXA2 was stably knocked down in PC3 cells and overexpressed in LNCaP cells.ResultsWe found that FOXA2 knockdown in PC3 cells resulted in a significant decrease in PC3 mediated in vivo bone destruction following intra‐tibial injection. To understand how FOXA2 is facilitating these changes, we examined the expression of integrins and observed that FOXA2 knockdown decreased the expression of collagen‐binding integrins α1 and αv in PC3 cells. Furthermore, we found FOXA2 knockdown decreased PC3 cells' adhesion and spreading on collagen I (a major component of bone ECM) coated surfaces. The Foxa2‐controlled expression of integrins α1 and αv and the resulting changes in the adherence and spreading would provide a mechanism for PCa cells colonize bone and initiate the bone‐destruction cycle. Additionally, FOXA2 knockdown in PC3 cells resulted in a significant decrease in expression of parathyroid hormone‐related protein (PTHrP), a bone remodeling‐associated protein. We observed that PTHrP mRNA was decreased in FOXA2 knockdown cells and protein in FOXA2 knockdown subcutaneous grafts. When PC3 FOXA2 knockdown cells were co‐cultured with osteoblast and osteoclast, osteoclast markers were decreased.ConclusionsTaken together, FOXA2 plays a major role in facilitating PCa's ability to colonize bone, and further, promote osteoclast activation.Support or Funding InformationFeist‐Weiller Cancer Center/Foundation of Legacy Funds The Office of Research SEED awardsThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Receptor Tyrosine Kinase Signalling in the Absence of Kinase Activity and Cancer of Non‐Genetic Origin
Authors: Ladbury John 
Published: June 2021

FGFR2‐expressing cancer cells with low concentrations of the adaptor protein Grb2 show high prevalence for metastatic outcome. In non‐stimulated cells the SH3 domain (and not the SH2 domain(s)) of Plcγ1 directly competes for a proline‐rich binding site at the very C‐terminus of FGFR2 with the C‐terminal SH3 domain of Grb2. Reduction of Grb2 concentration permits access of Plcγ1 to the receptor. Recruitment of Plcγ1 in this way is sufficient to up‐regulate phospholipase activity. This results in increased cell proliferation, motility and promotion of cell invasive behaviour in the absence of extracellular receptor stimulation. Therefore tumour growth and metastatic outcome can be dictated by the constitutive competition between Grb2 and Plcγ1 for the phosphorylation‐independent binding site on FGFR2. Since the majority of receptor tyrosine kinases have proline‐rich sequences in their C‐termini, the possibility of a second tier of signal transduction in the absence of growth factor stimulation, or kinase‐activating mutations emerges – leading to cancer of non‐genetic origin. Further examples of this second tier of signalling have been investigated and mechanistic detail of how protein concentration fluctuations in response to cellular stress can lead to pathogenesis has been established.Support or Funding InformationThis work is funded by CRUKThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The role of sphingosine kinase 2 in promoting multiple myeloma cell invasive growth
Authors: Zhang Hannah , Zhao Derrick , Li Xiaojiaoyang , Liu Runping , Wang Xuan , Zhou Huiping 
Published: June 2021

Multiple myeloma (MM) is a heterogeneous disease with a prognosis that varies with patient genetics, disease burden, tumor biology, and treatments. Effective therapy for high risk MM is currently unavailable due to the limited understanding of its pathophysiology. Sphingosine 1‐phosphate (S1P) has been identified as an important signaling molecule involved in regulating various cancer cell growths. Two protein kinases responsible for S1P synthesis, sphingosine kinase 1 and 2 (SphK1 and SphK2), have been identified. During the last decade, numerous studies have shown that SphK1 plays a critical role in promoting various cancer cell growths. However, only limited attention has been paid to SphK2. Our recent studies reported that activation of SphK2 is linked to invasive growth of cholangiocarcinoma. However, the role of SphK2 in MM cell growth is not clear and is the focus of this study.MethodsStudies were conducted in the human MM cell lines, KMS‐11, KMS‐12BM, KMS‐20, KMS‐21BM, KMS‐26, KMS‐28BM, KMS‐34, MOLP‐8, LP‐1, MM.1S. The expression of SphK1 and SphK2 was determined by real‐time RT‐PCR and Western blot analysis and confirmed by DNA sequencing. The SphK2 inhibitor, K145, and gene‐specific shRNA were used to inhibit SphK2 activation and expression. Cell proliferation was determined using a CCK‐8 kit. Cell apoptosis was analyzed by Annexin V‐FITC/PI staining followed by flow cytometry.ResultsThe expression level of SphK2 in all tested MM cell lines was significantly higher than SphK1. S1P significantly induced proliferation and growth of MM cells, which was inhibited by K145 as well as gene specific shRNA targeting SphK2. The SphK2 expression level is correlated to invasiveness of MM cell growth and its sensitivity to K145.Discussion/ConclusionSphK1 and SphK2 are all expressed in human MM cancer cells. However, the SphK2 expression level is much higher than SphK1. In this study, we were able to show that SphK2 plays a critical role in S1P‐mediated invasive growth of MM cells. The results in this study suggest that targeting SphK2 may represent a novel therapy for MM.Support or Funding InformationThis work was partially supported by National Institutes of Health Grant R01 DK104893 and R01DK‐057543; VA Merit Award I01BX001390; National Natural Science Foundation of China Grants 81070245 and 81270489.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigating the Potential for Na+‐H+ Exchanger Isoform 1 (NHE1) Inhibitors as Adjuvant Therapies in the Treatment of Ovarian Cancer
Authors: Corradi Anna R. , Provost Joseph J. , Wallert Mark 
Published: June 2021

Each year in the United States over 200,000 women are diagnosed with ovarian cancer leading to more than 130,000 deaths. A major challenge in ovarian cancer treatment is the development of drug resistance to the therapeutic agents. Increases in the expression and activity Na+‐H+ Exchanger Isoform 1 (NHE1) was recently demonstrated to regulate cell proliferation in ovarian cancer patients. NHE1 is an 815 amino acid transmembrane transport protein that regulates intracellular pH. We hypothesize that combining common ovarian cancer therapeutic agents along with compounds that directly or indirectly inhibit NHE1 will enhance the ability of the therapeutic agents to slow ovarian cancer cell proliferation and migration. Three ovarian adenocarcinoma cell lines; CAOV‐3, SKOV‐3, and OVCAR‐3 are used in this research. Using a proliferation assay, we assessed the impact of different ovarian cancer therapeutics (Rucaparib, Olaparib, Paclitaxel and Cisplatin) in these three cell lines in the presence and absence of inhibitors that decrease NHE1 activity directly or indirectly. Our previous work demonstrated that the NHE1 inhibitor cariporide and a number of protein kinase inhibitors decreased cell proliferation rates in all three cell types. Treatment with Cisplatin reduced cell proliferation most significantly in the CAOV‐3 line. Paclitaxel and Cisplatin each had significant and comparable impacts on the SKOV‐3 and OVCAR‐3 cell lines. Cell migration was evaluated using electric cell‐substrate impedance sensing (ECIS). In the SKOV‐3 and CAOV‐3 cells, inhibition with cariporide and the Rsk inhibitor BID1870 each dramatically reduced the rate of cell migration. The research presented here supports the role of NHE1 in cell proliferation, and for the first time implicates NHE1 in the regulation of cell migration in ovarian cancer cells. Data will be presented that evaluated the potential for the inhibition of NHE1 as an adjuvant therapy in ovarian cancer.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Proof‐of‐Concept in a 3D Culture Model of Glioma Invasion: Towards Personalized Therapeutics in Brain Cancer
Authors: van Pel Derek M. , Naus Christian C. , Sin Wun Chey 
Published: June 2021

Glioma is a highly aggressive form of brain cancer, with some subtypes having 5‐year survival rates of less than 5%. Tumor cell invasion into the surrounding parenchyma seems to be the primary driver of these poor outcomes, as most gliomas recur within 2 cm of the original surgically‐resected tumor. Many current approaches to the development of anticancer therapy attempt to target genetic weaknesses in a particular cancer, but may not take into account the microenvironment experienced by a tumor and patient‐specific genetic differences in susceptibility to treatment.The primary aim of our research program is to overcome these limitations by ultimately developing 3D cerebral organoids generated from patient‐derived induced pluripotent stem cells (iPSCs), in which we will screen to identify an optimal treatment regimen by combining the organoids with patient‐derived tumor cells. As a proof‐of‐concept of this approach, we developed a system in which we could model the invasion of human glioma cells into mouse neural progenitor cell‐derived spheroids. Here we demonstrate the utility of such an approach, and find that we can follow invasion of human tumor cells using cell tracking dyes and 3D laser scanning confocal microscopy, both in real time and in fixed samples. These results were validated using conventional cryosectioning. We go on to show broad applicability of this system by using two different tumor cell lines, and two different mouse genetic backgrounds, attempting to mimic differences that might be observed in tumor and patient brain tissue, respectively. Having obtained this proof‐of‐concept, we will next recapitulate these results in a fully humanized system using spheroids and then cerebral organoids from human iPSCs, before moving on to patient‐derived cultures. This stepwise approach could ultimately lead to truly personalized treatments for brain cancer.Support or Funding InformationDvP is supported by the UBC Faculty of Medicine, the UBC Summer Student Research Program, and the Mach‐Gaensslen Foundation. CCN is supported by the Canada Research Chair program and a grant from the Canadian Cancer Society. WCS is supported by the Canadian Cancer Society.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Role of N‐type Calcium Channels in inflammatory pain and associated sensory nerve growth
Authors: Pitake Saumitra , Mishra Santosh 
Published: June 2021

Voltage‐gated calcium channels (VGCC) play an important role in pain neurotransmission by releasing neuropeptides substance P and calcitonin gene‐related peptide from the sensory neurons. Here, we identified a new role of VGCC in pain signaling by sensory nerve growth at the periphery in the skin. In the present study, we first assessed the role of N‐type calcium channels (Cav2.2) in inflammatory pain. Complete Freund's Adjuvant (CFA), a known inflammogen was injected into the hind‐paws of C57Bl6 mice to induce inflammation and dorsal root ganglia from the lumbar region (L4–L6) were isolated and cultured 24‐hour post injection. Voltage clamp recordings revealed a significant increase in Cav2.2 currents between contralateral and ipsilateral DRGs. Current clamp analysis of these DRGs showed a significant increase in action potential frequency upon CFA injection. A significant increase in Cav2.2 currents was also observed when CFA (1:1000) was directly added to DRG culture. The increase in Cav2.2 currents was significantly decreased upon addition of ω‐conotoxin (1 μM) into DRG culture. Immunocytochemical analysis of cultured DRGs with CFA (1:1000) revealed a significant increase in TRPV1, Cav2.2 and neuronal filament markers such as Synaptophysin and neuronal marker β‐III tubulin (TUJ1), indicate increased neuronal synthesis and neurite formation at the periphery. This was further corroborated by Western Blot analysis. Altogether, our study suggests a possible role of VGCC‐dependent sensory nerve growth that might be responsible for increased sensitivity to minimal stimuli in chronic pain.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Role of Circadian Deadenylase Nocturnin in the Mitochondria
Authors: Ojo Anne , Laothamatas Isara , Green Carla 
Published: June 2021

The circadian rhythm is an evolutionarily conserved process that drives rhythmic behavior and physiology with a period of approximately 24 hours. Circadian rhythms are generated by the heterodimeric transcription factors CLOCK/BMAL1. CLOCK/BMAL1 binds to E‐boxes across the genome and drives the transcription of many genes, including their own repressors Per and Cry. Transcription activation by CLOCK/BMAL1 and its inhibitory feedback loop generated by PER/CRY result in the rhythmic expression of thousands of clock‐controlled genes. Nocturnin (mNOC) is among those genes regulated by CLOCK/BMAL1. mNOC is highly expressed in the liver, where its expression is at maximum in the evening and minimum in the morning. Compared to WT mice, mNOC KO mice exhibit a resistance to high‐fat diet (HFD) induced obesity. Protein sequence analysis reveals that mNOC contains an EEP domain that is conserved in the CCR4 family, whose members function as deadenylases. Interestingly, mNOC also contains a putative mitochondrial targeting signal (MTS) in between two translation initiation sites. To investigate if mNOC does localize to the mitochondria, we performed immunocytochemistry in HEK293 cells overexpressing mNOC that does or does not code for the MTS; and performed a mitochondrial fractionation followed by proteinase k protection assay. To examine mNOC's function in the mitochondria, we overexpressed mNOC in HEK293 cells and performed RT‐qPCR to examine mitochondrial RNA stability. We found that mNOC is able to localize to the mitochondria, but we did not see changes in mitochondrial RNA levels. Further experiments will help conclude if mNOC is targeting mitochondrial translation.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Immunoregulatory role of melatonin in Helicobacter pylori‐induced gastric diseases
Authors: Jianhua Luo , Hui Zhang , Mi Zhang , Jun Song , Li Li , Dancen Lin , Meifang Lin , Feng Zhang , Hui Liu , Ruixiang Zhou 
Published: June 2021

Melatonin has shown important immunoregulatory activities in inflammatory disorders; however, the immunoregulatory role of melatonin in Helicobacter pylori‐induced gastric diseases remains unclear until now. This study aimed to examine the in‐vivo activity of melatonin against H. pylori‐induced gastric diseases and explore the immunoregulatory mechanisms. HE staining showed obvious inflammatory cell infiltration in the gastric mucosa and submucosa, and the culture of the mouse stomach homogenate was positive for the rapid urease test. Immunohistochemistry revealed extensive MT1 and MT2 expression in the gastric mucosa of H. pylori‐infected mice. ELISA measured lower TGF‐β1 levels in the liver of H. pylori‐infected mice than in uninfected mice 2 weeks post‐treatment with 25, 50 or 100 mg/kg melatonin, while higher TGF‐β1 levels were detected 4 and 6 weeks post‐treatment. Western blotting determined lower Foxp3 expression in the spleen of infected mice than in uninfected mice 2 weeks post‐treatment with 25, 50 or 100 mg/kg melatonin, and higher Foxp3 levels were detected 4 weeks post‐treatment with 50 and 100 mg/kg melatonin; however, the Foxp3 level was comparative in infected and uninfected mice 6 weeks post‐treatment. In addition, higher RORβ expression was found in the gastric specimens of infected mice than in uninfected mice 2 weeks post‐treatment with 25, 50 or 100 mg/kg melatonin, and higher RORβ expression was detected 6 weeks post‐treatment with 25 mg/kg melatonin. At 2, 4 and 6 weeks post‐treatment with 25, 50 and 100 mg/kg melatonin, higher TGF‐β1 expression was detected in the liver of infected mice than in uninfected mice. The results demonstrate that melatonin may alleviate H. pylori‐induced gastric diseases through mediating TGF‐β1 and Foxp3 expression via membrane and nuclear receptors.Support or Funding InformationNational Natural Science Foundation Youth Fund:81302601 NSFC(National Natural Science Foundation of China):30971541This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Determining the Effects of Chronic Interferon Stimulation on Heme Oxygenase‐1 in Human Monocyte Derived Macrophages
Authors: Cobo Bryan , Garza Rolando , Kolson Dennis L. 
Published: June 2021

HIV‐associated neurocognitive disorders (HAND) affect ~40% of the HIV infected population even with the use of combined antiretroviral therapy (cART). We previously reported that the critical cytoprotective enzyme, Heme oxygenase‐1 (HO‐1) is reduced in the brains of HIV infected individuals and that this reduction correlates with increased expression of type 1 interferon response genes and markers of monocyte activation. Furthermore, reduction of HO‐1 protein expression is accompanied by increased HO‐1 RNA expression. We also showed that chronic type 2 interferon (IFN) stimulation in astrocytes leads to HO‐1 protein degradation via an immunoproteasome mediated mechanism. Here, we aim to determine the effects of chronic type 1 and type 2 IFN stimulation on human monocyte derived macrophage (MDM) HO‐1 protein expression and elucidate the mechanisms by which IFNs cause dysregulation of HO‐1 protein. To address this, we treated primary human MDMs with type 1 or type 2 IFNs for 24 hours or 10 days. We collected protein lysates for western blot analysis and total RNA for quantitative PCR. Supernatants were collected to measure glutamate production and supernatant neurotoxicity. We observed that chronic type 1 and type 2 IFN stimulation (0.1 ng/ml) did not alter HO‐1 protein expression, however, we observed increased HO‐1 RNA expression with chronic type 1 IFN stimulation. Furthermore, we observed increased immunoproteosome subunit expression and loss of constitutive proteasome expression. No differences were observed in glutamate production or supernant neurotoxicity. These results suggest that chronic type 1 and type 2 IFN stimulation induce immunoproteasome expression and can subsequently lead to accelerated HO‐1 protein degradation, which may contribute to neurodegeneration.Support or Funding InformationI would like to thank Mr. Rolando Garza and Dr. Dennis L. Kolson for their insightful comments and for allowing me to aid with their research project this summer. I would also like to express my gratitude towards my program advisors, Dr. Arnaldo Diaz Vazquez and Dr. Raquel Castellanos, the Office of Research and Diversity Training at Penn, and the following organizations that made this opportunity possible: MARC program @ UPR‐RP (NIH Grant 5T34GM007821‐37) and the Perelman School of Medicine @ UPenn.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Mechanotransduction via HA LFA‐1 promotes Kindlin‐3/RACK1/Orai1 engagement to mediate Calcium in PMN
Authors: Morikis Vasilios Aris , Simon Scott 
Published: June 2021

Absence of Kindlin‐3 severely impairs β2‐integrin function on neutrophils, results in severe bleeding and resistance to arterial thrombosis and immunodeficiency. Kindlin‐3 binds LFA‐1 β‐integrin tail independent of Talin‐1 and is associated with integrin conversion to a high‐affinity state and the facilitation of micro‐cluster formation that allow for long‐lived shear resistance bonds with ICAM‐1. However, the molecular and mechanical events that link LFA‐1 to Kindlin‐3 at spatially distinct sites to allow for subsequent engagement with scaffolding protein RACK1 and calcium release activated channel Orai1 has not been rigorously shown. We show that an allosteric shift of LFA‐1 due to force generation on the β2‐integrin cytodomain promotes interaction with Kindlin‐3 F3 subdomain and when sufficient force is transduced through this mechanical linkage induces the onset of calcium flux. We utilize real time two‐line total internal reflection fluorescent and epifluorescence microscopy on neutrophil‐like PLB‐985 cells arrested to ICAM‐1 within our microfluidic vascular mimetic. We have been able to identify the stoichiometric binding rate of Kindlin‐3 and subsequent RACK‐1 recruitment to high‐affinity LFA‐1 sites in response to increasing shear profiles prior to calcium flux. In addition to outside‐in signaling we have shown that force acting on LFA‐1 amplifies calcium influx during CXCR1/2 ligation in a Kindlin‐3 dependent manner. To assess the spatial proximity and association between Kindlin‐3/RACK1 with focal adhesion clusters we are developing a FRET pair between the Kindlin‐3 PH domain and RACK1 propellers 5 to 7 to further elucidate the spatiotemporal events and requirements for outside‐in signaling and neutrophil progression to a migratory phenotype. This has allowed for us to identify key forces and proteins required for outside‐in signaling during neutrophil transition from rolling to arrest which will help further glyocomimetic development to treat various disease states.Support or Funding InformationThe work was supported by grants AI047294 (SIS).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.