CD34 + hematopoietic stem/progenitor cells (HSPCs) are vasculogenic and hypoxia is a strong stimulus for the vasoreparative functions of these cells. Angiotensinconverting enzyme 2 (ACE2)/angiotensin-(1-7)/Mas receptor (MasR) pathway stimulates vasoprotective functions of CD34 + cells. This study tested if ACE2 and MasR are involved in the hypoxic stimulation of CD34 + cells. Cells were isolated from circulating mononuclear cells derived from healthy subjects (n = 46) and were exposed to normoxia (20% O 2 ) or hypoxia (1% O 2 ). Luciferase reporter assays were carried out in cells transduced with lentivirus carrying ACE2-or MasR-or a scramble-3′-untranslated region gene with a firefly luciferase reporter. Expressions or activities of ACE, angiotensin receptor Type 1 (AT1R), ACE2, and MasR were determined. In vitro observations were verified in HSPCs derived from mice undergoing hindlimb ischemia (HLI). In vitro exposure to hypoxia-increased proliferation and migration of CD34 + cells in basal conditions or in response to vascular endothelial growth factor (VEGF) or stromal-derived factor 1α (SDF) compared with normoxia. Expression of ACE2 or MasR was increased relative to normoxia while ACE or AT1R expressions were unaltered. Luciferase activity was increased by hypoxia in cells transfected with the luciferase reporter plasmids coding for the ACE2-or MasR promoters relatively to the control. The effects of hypoxia were mimicked by VEGF or SDF under normoxia. Hypoxia-induced ADAM17-dependent shedding of functional ACE2 fragments. In mice undergoing HLI, increased expression/activity of ACE2 and MasR were observed in the circulating HSPCs. This study provides compelling evidence for the hypoxic upregulation of ACE2 and MasR in CD34 + cells, which likely contributes to vascular repair. K E Y W O R D S ACE2, CD34 + cells, hypoxia, Mas receptor 1 | BACKGROUND Accumulated evidence based on numerous laboratory and clinical studies strongly support the vasoregenerative functions of bone marrow-derived CD34 + hematopoietic stem/progenitor cells (HSPCs) and their therapeutic potential for the treatment of ischemic vascular disorders (Mackie & Losordo, 2011; Schachinger

Aberrant activation of Wnt signaling has been implicated in human osteosarcoma, which may provide a genetic vulnerability that can be targeted in osteosarcoma treatment. To test whether Wnt activation is necessary for osteosarcoma growth, colony formation, invasion, and metastasis, we treated human osteosarcoma cells with a small molecule inhibitor of Wnt/β-catenin, PRI-724, which suppresses Wnt/β-catenin-mediated transcription. We found increased protein levels of endogenous active-β-catenin in five human osteosarcoma cell lines. Treatment with PRI-724 was sufficient to inhibit human osteosarcoma 143B and SJSA-1 cell proliferation. Suppressed Wnt signaling was confirmed by decreased protein levels of the Wnt target Cyclin D1. Furthermore, we revealed significant inhibitory effects on cell migration, invasion, and colony formation in the human osteosarcoma cells. Using deposited data from next generation sequencing studies, we analyzed somatic mutations and gene expression of components in the Wnt/β-catenin pathway. We found somatic mutations and upregulated gene expression of many components in the Wnt/ β-catenin pathway, indicating activated Wnt signaling. Taken together, our results illustrate the critical role of Wnt/β-catenin signaling in human osteosarcoma pathogenesis and growth, as well as the therapeutic potential of Wnt inhibitors in the treatment of human osteosarcoma.

Silica nanoparticles (SiNPs) have been used as vehicles for drug delivery, molecular detection, and cellular manipulations in nanoneuromedicine. SiNPs may cause adverse effects in brain including neurotoxicity, neuroinflammation, neurodegeneration and enhancing levels of amyloid beta protein (Aβ); all pathological hallmarks of Alzheimer’s disease. Therefore, the extent to which SiNPs influence Aβ generation and the underlying mechanisms by which this occurs deserves investigation. Our studies were focused on the effects of SiNPs on endolysosomes which uptake, traffic, and mediate the actions of SiNPs. These organelles are also where amyloidogenesis largely originates. We found that SiNPs, in primary cultured hippocampal neurons, accumulated in endolysosomes and caused a rapid and persistent deacidification of endolysosomes. SiNPs significantly reduced endolysosome calcium stores as indicated by a significant reduction in the ability of the lysosomotropic agent GPN to release calcium from endolysosomes. SiNPs increased Aβ1–40 secretion whereas two agents that acidified endolysosomes, ML-SA1 and CGS21680, blocked SiNPs-induced deacidification and increased generation of Aβ1–40. Our findings suggest that SiNP-induced deacidification of and calcium release from endolysosomes might be mechanistically-linked to increased amyloidogenesis. The use of SiNPs might not be the best nanomaterial for therapeutic strategies against AD and other neurological disorders linked to endolysosome dysfunction.

Although many cancer prognoses have improved in the past 50 years due to advancements in treatments, there has been little improvement in therapies for small-cell lung cancer (SCLC). One promising avenue to improve treatment for SCLC is to understand its underlying genetic alterations that drive its formation, growth, and cellular heterogeneity. RB1 loss is one key driver of SCLC, and RB1 loss has been associated with an increase in pluripotency factors such as SOX2. SOX2 is highly expressed and amplified in SCLC and has been associated with SCLC growth. Using a genetically engineered mouse model, we have shown that Sox2 is required for efficient SCLC formation. Furthermore, genome-scale binding assays have indicated that SOX2 can regulate key SCLC pathways such as NEUROD1 and MYC. These data suggest that SOX2 can be associated with the switch of SCLC from an ASCL1 subtype to a NEUROD1 subtype. Understanding this genetic switch is key to understanding such processes as SCLC progression, cellular heterogeneity, and treatment resistance. Implications: Understanding the molecular mechanisms of SCLC initiation and development are key to opening new potential therapeutic options for this devastating disease.

Although many cancer prognoses have improved in the past fifty years due to advancements in treatments, there has been little to no improvement in therapies for small cell lung cancer (SCLC) which currently has a five-year survival rate of less than 7%. One promising avenue to improve treatment for SCLC is to understand its underlying genetic alterations that drive its formation and growth. One such mutation in SCLC, which appears in many cancers, is of the Rb gene. When mutated, Rb causes hyperproliferation and loss of cellular identity. Normally Rb promotes differentiation by regulating lineage specific transcription factors including regulation of pluripotency factors such as Sox2. However, there is evidence that when certain tissues lose Rb, Sox2 becomes upregulated and promotes oncogenesis. To better understand the relationship between Rb and Sox2 and to uncover new treatments for SCLC we have studied the role of Sox2in Rb loss initiated tumors by investigating both the tumor initiation in SCLC genetically engineered mouse models, as well as tumor maintenance in SCLC cell lines.

Small Cell Lung Cancer (SCLC) is often a heterogeneous tumor, where dynamic regulation of key transcription factors can drive multiple populations of phenotypically different cells which contribute differentially to tumor dynamics. This tumor is characterized by a very low 2-year survival rate, high rates of metastasis, and rapid acquisition of chemoresistance. The heterogeneous nature of this tumor makes it difficult to study and to treat, as it is not clear how or when this heterogeneity arises. Here we describe temporal, single-cell analysis of SCLC to investigate tumor initiation and chemoresistance in both SCLC xenografts and an autochthonous SCLC model. We identify an early population of tumor cells with high expression of AP-1 network genes that are critical for tumor growth. Furthermore, we have identified and validated the cancer testis antigens (CTAs) PAGE5 and GAGE2A as mediators of chemoresistance in human SCLC. CTAs have successfully been targeted in other tumor types and may be a promising avenue for targeted therapy in SCLC. Implications: Understanding the evolutionary dynamics of SCLC can shed light on key mechanisms such as cellular plasticity, heterogeneity, and chemoresistance.

<p>Determining effective concentrations for treatment of SCLC cell lines with cisplatin and etoposide.</p>

<p>Similarity analysis to determine subclonal relationships.</p>