Microstructure and electric properties of lead lanthanum titanate thin film under transverse electric fields

Author name used in this publication: N. Chong2001-2002 > Academic research: refereed > Publication in refereed journalVersion of RecordPublishe

Wnt-C59 arrests stemness and suppresses growth of nasopharyngeal carcinoma in mice by inhibiting the Wnt pathway in the tumor microenvironment

Wnt/β-catenin signaling is responsible for the generation of cancer stem cells (CSCs) in many human tumors, including nasopharyngeal carcinoma (NPC). Recent studies demonstrate that Wnt or PORCN inhibitor, Wnt-C59, inhibits tumor growth in MMTV-WNT1 transgenic mice. The effect of Wnt-C59 in human tumors is not clear. In this study, the NPC cell lines investigated manifest heterogeneous responses to Wnt-C59 treatment. Wnt-C59 decreased tumor growth of SUNE1 cells in mice immediately following the administration of Wnt-C59. Mice injected with HNE1 cells did not develop visible tumors after the treatment of Wnt-C59, while control mice developed 100% tumors. Wnt-C59 inhibited stemness properties of NPC cells in a dosage-dependent manner by arresting sphere formation in both HNE1 and SUNE1 cells. Thus, Wnt-C59 has the potential to eradicate CSCs in human tumors. Active β-catenin and Axin2 proteins were strongly expressed in stromal cells surrounding growing tumors, confirming the importance of Wnt signaling activities in the microenvironment being driving forces for cell growth. These novel findings confirm the ability of Wnt-C59 to suppress Wnt-driven undifferentiated cell growth in NPC. Both anti-Wnt signaling and anti-CSC approaches are feasible strategies in cancer therapy.published_or_final_versio

Super-resolution microscopy reveals coupling between mammalian centriole subdistal appendages and distal appendages

Copyright © 2020 Chong et al. Subdistal appendages (sDAPs) are centriolar elements that are observed proximal to the distal appendages (DAPs) in vertebrates. Despite the obvious presence of sDAPs, structural and functional understanding of them remains elusive. Here, by combining super-resolved localization analysis and CRISPR-Cas9 genetic perturbation, we find that although DAPs and sDAPs are primarily responsible for distinct functions in ciliogenesis and microtubule anchoring, respectively, the presence of one element actually affects the positioning of the other. Specifically, we find dual layers of both ODF2 and CEP89, where their localizations are differentially regulated by DAP and sDAP integrity. DAP depletion relaxes longitudinal occupancy of sDAP protein ninein to cover the DAP region, implying a role of DAPs in sDAP positioning. Removing sDAPs alter the distal border of centrosomal g-tubulins, illustrating a new role of sDAPs. Together, our results provide an architectural framework for sDAPs that sheds light on functional understanding, surprisingly revealing coupling between DAPs and sDAPs.Ministry of Science and Technology, Taiwan (107-2112-M-001-037, 107-2313-B-001-009, 108-2313-B-010-001, 108-2628-B-010-007, 108-2638-B-010-001 -MY2); Academia Sinica (2317-1040300); National Institutes of Health (GM088253)

A novel Family Dignity Intervention (FDI) for enhancing and informing holistic palliative care in Asia: study protocol for a randomized controlled trial

Background The lack of a holistic approach to palliative care can lead to a fractured sense of dignity at the end of life, resulting in depression, hopelessness, feelings of being a burden to others, and the loss of the will to live among terminally ill patients. Building on the clinical foundation of Dignity Therapy, together with the empirical understanding of dignity-related concerns of Asian families facing terminal illness, a novel Family Dignity Intervention (FDI) has been developed for Asian palliative care. FDI comprises a recorded interview with a patient and their primary family caregiver, which is transcribed, edited into a legacy document, and returned to the dyads for sharing with the rest of the patient’s family. The aims of this study are to assess the feasibility, acceptability and potential effectiveness of FDI in reducing psychosocial, emotional, spiritual, and psychophysiological distress in community-dwelling and in-patient, Asian, older terminally ill patients and their families living in Singapore. Methods/design An open-label randomized controlled trial. One hundred and twenty-six patient-family dyads are randomly allocated to one of two groups: (1) an intervention group (FDI offered in addition to standard psychological care) and (2) a control group (standard psychological care). Both quantitative and qualitative outcomes are assessed in face-to-face interviews at baseline, 3 days and 2 weeks after intervention, as well as during an exit interview with family caregivers at 2 months post bereavement. Primary outcome measures include sense of dignity for patients and psychological distress for caregivers. Secondary outcomes include meaning in life, quality of life, spirituality, hopefulness, perceived support, and psychophysiological wellbeing, as well as bereavement outcomes for caregivers. Qualitative data are analyzed using the Framework method. Discussion To date, there is no available palliative care intervention for dignity enhancement in Asia. This first-of-its-kind study develops and tests an evidence-based, family driven, psycho-socio-spiritual intervention for enhancing dignity and wellbeing among Asian patients and families facing mortality. It addresses a critical gap in the provision of holistic palliative care. The expected outcomes will contribute to advancements in both theories and practices of palliative care for Singapore and its neighboring regions while serving to inform similar developments in other Asian communities. Trial registration ClinicalTrials.gov, ID: NCT03200730. Registered on 26 June 2017

An Unexpected Role for the Clock Protein Timeless in Developmental Apoptosis

Background: Programmed cell death is critical not only in adult tissue homeostasis but for embryogenesis as well. One of the earliest steps in development, formation of the proamniotic cavity, involves coordinated apoptosis of embryonic cells. Recent work from our group demonstrated that c-Src protein-tyrosine kinase activity triggers differentiation of mouse embryonic stem (mES) cells to primitive ectoderm-like cells. In this report, we identified Timeless (Tim), the mammalian ortholog of a Drosophila circadian rhythm protein, as a binding partner and substrate for c-Src and probed its role in the differentiation of mES cells. Methodology/Principal Findings: To determine whether Tim is involved in ES cell differentiation, Tim protein levels were stably suppressed using shRNA. Tim-defective ES cell lines were then tested for embryoid body (EB) formation, which models early mammalian development. Remarkably, confocal microscopy revealed that EBs formed from the Tim-knockdown ES cells failed to cavitate. Cells retained within the centers of the failed cavities strongly expressed the pluripotency marker Oct4, suggesting that further development is arrested without Tim. Immunoblots revealed reduced basal Caspase activity in the Tim-defective EBs compared to wild-type controls. Furthermore, EBs formed from Tim-knockdown cells demonstrated resistance to staurosporine-induced apoptosis, consistent with a link between Tim and programmed cell death during cavitation. Conclusions/Significance: Our data demonstrate a novel function for the clock protein Tim during a key stage of early development. Specifically, EBs formed from ES cells lacking Tim showed reduced caspase activity and failed to cavitate. As a consequence, further development was halted, and the cells present in the failed cavity remained pluripotent. These findings reveal a new function for Tim in the coordination of ES cell differentiation, and raise the intriguing possibility that circadian rhythms and early development may be intimately linked. © 2011 O'Reilly et al

Differential sensitivity of Src-family kinases to activation by SH3 domain displacement

Src-family kinases (SFKs) are non-receptor protein-tyrosine kinases involved in a variety of signaling pathways in virtually every cell type. The SFKs share a common negative regulatory mechanism that involves intramolecular interactions of the SH3 domain with the PPII helix formed by the SH2-kinase linker as well as the SH2 domain with a conserved phosphotyrosine residue in the C-terminal tail. Growing evidence suggests that individual SFKs may exhibit distinct activation mechanisms dictated by the relative strengths of these intramolecular interactions. To elucidate the role of the SH3:linker interaction in the regulation of individual SFKs, we used a synthetic SH3 domain-binding peptide (VSL12) to probe the sensitivity of downregulated c-Src, Hck, Lyn and Fyn to SH3-based activation in a kinetic kinase assay. All four SFKs responded to VSL12 binding with enhanced kinase activity, demonstrating a conserved role for SH3:linker interaction in the control of catalytic function. However, the sensitivity and extent of SH3-based activation varied over a wide range. In addition, autophosphorylation of the activation loops of c-Src and Hck did not override regulatory control by SH3:linker displacement, demonstrating that these modes of activation are independent. Our results show that despite the similarity of their downregulated conformations, individual Src-family members show diverse responses to activation by domain displacement which may reflect their adaptation to specific signaling environments in vivo. © 2014 Moroco et al

HS1, a Lyn Kinase Substrate, Is Abnormally Expressed in B-Chronic Lymphocytic Leukemia and Correlates with Response to Fludarabine-Based Regimen

In B-Chronic Lymphocytic Leukemia (B-CLL) kinase Lyn is overexpressed, active, abnormally distributed, and part of a cytosolic complex involving hematopoietic lineage cell-specific protein 1 (HS1). These aberrant properties of Lyn could partially explain leukemic cells’ defective apoptosis, directly or through its substrates, for example, HS1 that has been associated to apoptosis in different cell types. To verify the hypothesis of HS1 involvement in Lyn-mediated leukemic cell survival, we investigated HS1 protein in 71 untreated B-CLL patients and 26 healthy controls. We found HS1 overexpressed in leukemic as compared to normal B lymphocytes (1.38±0.54 vs 0.86±0.29, p<0.01), and when HS1 levels were correlated to clinical parameters we found a higher expression of HS1 in poor-prognosis patients. Moreover, HS1 levels significantly decreased in ex vivo leukemic cells of patients responding to a fludarabine-containing regimen. We also observed that HS1 is partially localized in the nucleus of neoplastic B cells. All these data add new information on HS1 study, hypothesizing a pivotal role of HS1 in Lyn-mediated modulation of leukemic cells’ survival and focusing, one more time, the attention on the BCR-Lyn axis as a putative target for new therapeutic strategies in this disorder

Multiparameter Phospho-Flow Analysis of Lymphocytes in Early Rheumatoid Arthritis: Implications for Diagnosis and Monitoring Drug Therapy

The precise mechanisms involved in the initiation and progression of rheumatoid arthritis (RA) are not known. Early stages of RA often have non-specific symptoms, delaying diagnosis and therapy. Additionally, there are currently no established means to predict clinical responsiveness to therapy. Immune cell activation is a critical component therefore we examined the cellular activation of peripheral blood mononuclear cells (PBMCs) in the early stages of RA, in order to develop a novel diagnostic modality.PBMCs were isolated from individuals diagnosed with early RA (ERA) (n = 38), longstanding RA (n = 10), osteoarthritis (OA) (n = 19) and from healthy individuals (n = 10). PBMCs were examined for activation of 15 signaling effectors, using phosphorylation status as a measure of activation in immunophenotyped cells, by flow cytometry (phospho-flow). CD3+CD4+, CD3+CD8+ and CD20+ cells isolated from patients with ERA, RA and OA exhibited activation of multiple phospho-epitopes. ERA patient PBMCs showed a bias towards phosphorylation-activation in the CD4+ and CD20+ compartments compared to OA PBMCs, where phospho-activation was primarily observed in CD8+ cells. The ratio of phospho (p)-AKT/p-p38 was significantly elevated in patients with ERA and may have diagnostic potential. The mean fluorescent intensity (MFI) levels for p-AKT and p-H3 in CD4+, CD8+ and CD20+ T cells correlated directly with physician global assessment scores (MDGA) and DAS (disease activity score). Stratification by medications revealed that patients receiving leflunomide, systemic steroids or anti-TNF therapy had significant reductions in phospho-specific activation compared with patients not receiving these therapies. Correlative trends between medication-associated reductions in the levels of phosphorylation of specific signaling effectors and lower disease activity were observed.Phospho-flow analysis identified phosphorylation-activation of specific signaling effectors in the PB from patients with ERA. Notably, phosphorylation of these signaling effectors did not distinguish ERA from late RA, suggesting that the activation status of discrete cell populations is already established early in disease. However, when the ratio of MFI values for p-AKT and p-p38 is >1.5, there is a high likelihood of having a diagnosis of RA. Our results suggest that longitudinal sampling of patients undergoing therapy may result in phospho-signatures that are predictive of drug responsiveness

Phosphoprotein Associated with Glycosphingolipid-Enriched Microdomains Differentially Modulates Src Kinase Activity in Brain Maturation

Src family kinases (SFK) control multiple processes during brain development and function. We show here that the phosphoprotein associated with glycosphigolipid-enriched microdomains (PAG)/Csk binding protein (Cbp) modulates SFK activity in the brain. The timing and localization of PAG expression overlap with Fyn and Src, both of which we find associated to PAG. We demonstrate in newborn (P1) mice that PAG negatively regulates Src family kinases (SFK). P1 Pag1-/- mouse brains show decreased recruitment of Csk into lipid rafts, reduced phosphorylation of the inhibitory tyrosines within SFKs, and an increase in SFK activity of >/ = 50%. While in brain of P1 mice, PAG and Csk are highly and ubiquitously expressed, little Csk is found in adult brain suggesting altered modes of SFK regulation. In adult brain Pag1-deficiency has no effect upon Csk-distribution or inhibitory tyrosine phosphorylation, but kinase activity is now reduced (−20–30%), pointing to the development of a compensatory mechanism that may involve PSD93. The distribution of the Csk-homologous kinase CHK is not altered. Importantly, since the activities of Fyn and Src are decreased in adult Pag1-/- mice, thus presenting the reversed phenotype of P1, this provides the first in vivo evidence for a Csk-independent positive regulatory function for PAG in the brain