Technology-Enabled Health Care Collaboration in Pediatric Chronic Illness: Pre-post Interventional Study for Feasibility, Acceptability, and Clinical Impact of an Electronic Health Record–Linked Platform for Patient-Clinician Partnership

Background: Mobile health (mHealth) technology has the potential to support the Chronic Care Model\u27s vision of closed feedback loops and patient-clinician partnerships. Objective: This study aims to evaluate the feasibility, acceptability, and short-term impact of an electronic health record-linked mHealth platform (Orchestra) supporting patient and clinician collaboration through real-time, bidirectional data sharing. Methods: We conducted a 6-month prospective, pre-post, proof-of-concept study of Orchestra among patients and parents in the Cincinnati Children\u27s Hospital inflammatory bowel disease (IBD) and cystic fibrosis (CF) clinics. Participants and clinicians used Orchestra during and between visits to complete and view patient-reported outcome (PRO) measures and previsit plans. Surveys completed at baseline and at 3- and 6-month follow-up visits plus data from the platform were used to assess outcomes including PRO completion rates, weekly platform use, disease self-efficacy, and impact on care. Analyses included descriptive statistics; pre-post comparisons; Pearson correlations; and, if applicable, effect sizes. Results: We enrolled 92 participants (CF: n = 52 and IBD: n = 40), and 73% (67/92) completed the study. Average PRO completion was 61%, and average weekly platform use was 80%. Participants reported improvement in self-efficacy from baseline to 6 months (7.90 to 8.44; P = .006). At 6 months, most participants reported that the platform was useful (36/40, 90%) and had a positive impact on their care, including improved visit quality (33/40, 83%), visit collaboration (35/40, 88%), and visit preparation (31/40, 78%). PRO completion was positively associated with multiple indicators of care impact at 3 and 6 months. Conclusions: Use of an mHealth tool to support closed feedback loops through real-time data sharing and patient-clinician collaboration is feasible and shows indications of acceptability and promise as a strategy for improving pediatric chronic illness management

The novel estrogen-induced gene EIG121 regulates autophagy and promotes cell survival under stress

We previously identified a novel estrogen-induced gene, EIG121, as being differentially regulated in endometrioid and nonendometrioid endometrial carcinoma. The function of EIG121 was unknown. Using a tetracycline-inducible system, we found that overexpression of EIG121, but not of LacZ, caused a profound suppression of cell growth. Subcellular fractionation and immunofluroscent labeling indicated that EIG121 was a transmembrane protein localized in the plasma membrane-late endosome–lysosome compartments. Deletion of the putative transmembrane domain abolished the membrane association. In cells overexpressing EIG121, cytoplasmic vacuoles accumulated after EIG121 induction, and the autophagosome marker LC3 translocated into punctuate, dot-like structures. Electron microscopy revealed that in cells overexpressing EIG121, autophagosomes were markedly increased. Overexpression of EIG121 also increased the cells containing acidic vesicles and induced lysosomal degradation of long-lived proteins. In MCF-7 cells, both EIG121 and LC3 were rapidly degraded by a lysosomal mechanism after starvation. Knockdown of EIG121 blocked starvation-induced LC3 degradation. By itself, knockdown of EIG121 did not affect cell survival. When combined with starvation or cytotoxic agents, EIG121 knockdown greatly increased apoptosis. Our results suggest that EIG121 is associated with the endosome–lysosome compartments and may have an important role in autophagy. Under unfavorable conditions such as starvation and exposure to cytotoxic agents, EIG121 may protect cells from cell death by upregulating the autophagy pathway

CLU blocks HDACI-mediated killing of neuroblastoma

Clusterin is a ubiquitously expressed glycoprotein with multiple binding partners including IL-6, Ku70, and Bax. Clusterin blocks apoptosis by binding to activated Bax and sequestering it in the cytoplasm, thereby preventing Bax from entering mitochondria, releasing cytochrome c, and triggering apoptosis. Because increased clusterin expression correlates with aggressive behavior in tumors, clusterin inhibition might be beneficial in cancer treatment. Our recent findings indicated that, in neuroblastoma cells, cytoplasmic Bax also binds to Ku70; when Ku70 is acetylated, Bax is released and can initiate cell death. Therefore, increasing Ku70 acetylation, such as by using histone deacetylase inhibitors, may be therapeutically useful in promoting cell death in neuroblastoma tumors. Since clusterin, Bax, and Ku70 form a complex, it seemed likely that clusterin would mediate its anti-apoptotic effects by inhibiting Ku70 acetylation and blocking Bax release. Our results, however, demonstrate that while clusterin level does indeed determine the sensitivity of neuroblastoma cells to histone deacetylase inhibitor-induced cell death, it does so without affecting histone deacetylase-inhibitor-induced Ku70 acetylation. Our results suggest that in neuroblastoma, clusterin exerts its anti-apoptotic effects downstream of Ku70 acetylation, likely by directly blocking Bax activation

Acetonic Extract of Buxus sempervirens Induces Cell Cycle Arrest, Apoptosis and Autophagy in Breast Cancer Cells

Plants are an invaluable source of potential new anti-cancer drugs. Here, we investigated the cytotoxic activity of the acetonic extract of Buxus sempervirens on five breast cancer cell lines, MCF7, MCF10CA1a and T47D, three aggressive triple positive breast cancer cell lines, and BT-20 and MDA-MB-435, which are triple negative breast cancer cell lines. As a control, MCF10A, a spontaneously immortalized but non-tumoral cell line has been used. The acetonic extract of Buxus sempervirens showed cytotoxic activity towards all the five studied breast cancer cell lines with an IC50 ranging from 7.74 µg/ml to 12.5 µg/ml. Most importantly, the plant extract was less toxic towards MCF10A with an IC50 of 19.24 µg/ml. Fluorescence-activated cell sorting (FACS) analysis showed that the plant extract induced cell death and cell cycle arrest in G0/G1 phase in MCF7, T47D, MCF10CA1a and BT-20 cell lines, concomitant to cyclin D1 downregulation. Application of MCF7 and MCF10CA1a respective IC50 did not show such effects on the control cell line MCF10A. Propidium iodide/Annexin V double staining revealed a pre-apoptotic cell population with extract-treated MCF10CA1a, T47D and BT-20 cells. Transmission electron microscopy analyses indicated the occurrence of autophagy in MCF7 and MCF10CA1a cell lines. Immunofluorescence and Western blot assays confirmed the processing of microtubule-associated protein LC3 in the treated cancer cells. Moreover, we have demonstrated the upregulation of Beclin-1 in these cell lines and downregulation of Survivin and p21. Also, Caspase-3 detection in treated BT-20 and T47D confirmed the occurrence of apoptosis in these cells. Our findings indicate that Buxus sempervirens extract exhibit promising anti-cancer activity by triggering both autophagic cell death and apoptosis, suggesting that this plant may contain potential anti-cancer agents for single or combinatory cancer therapy against breast cancer

O-Glycosylation Regulates Ubiquitination and Degradation of the Anti-Inflammatory Protein A20 to Accelerate Atherosclerosis in Diabetic ApoE-Null Mice

Background: Accelerated atherosclerosis is the leading cause of morbidity and mortality in diabetic patients. Hyperglycemia is a recognized independent risk factor for heightened atherogenesis in diabetes mellitus (DM). However, our understanding of the mechanisms underlying glucose damage to the vasculature remains incomplete. Methodology/Principal Findings: High glucose and hyperglycemia reduced upregulation of the NF-κB inhibitory and atheroprotective protein A20 in human coronary endothelial (EC) and smooth muscle cell (SMC) cultures challenged with Tumor Necrosis Factor alpha (TNF), aortae of diabetic mice following Lipopolysaccharide (LPS) injection used as an inflammatory insult and in failed vein-grafts of diabetic patients. Decreased vascular expression of A20 did not relate to defective transcription, as A20 mRNA levels were similar or even higher in EC/SMC cultured in high glucose, in vessels of diabetic C57BL/6 and FBV/N mice, and in failed vein grafts of diabetic patients, when compared to controls. Rather, decreased A20 expression correlated with post-translational O-Glucosamine-N-Acetylation (O-GlcNAcylation) and ubiquitination of A20, targeting it for proteasomal degradation. Restoring A20 levels by inhibiting O-GlcNAcylation, blocking proteasome activity, or overexpressing A20, blocked upregulation of the receptor for advanced glycation end-products (RAGE) and phosphorylation of PKCβII, two prime atherogenic signals triggered by high glucose in EC/SMC. A20 gene transfer to the aortic arch of diabetic ApoE null mice that develop accelerated atherosclerosis, attenuated vascular expression of RAGE and phospho-PKCβII, significantly reducing atherosclerosis. Conclusions: High glucose/hyperglycemia regulate vascular A20 expression via O-GlcNAcylation-dependent ubiquitination and proteasomal degradation. This could be key to the pathogenesis of accelerated atherosclerosis in diabetes

A20 Modulates Lipid Metabolism and Energy Production to Promote Liver Regeneration

Background: Liver Regeneration is clinically of major importance in the setting of liver injury, resection or transplantation. We have demonstrated that the NF-$\kappa$B inhibitory protein A20 significantly improves recovery of liver function and mass following extended liver resection (LR) in mice. In this study, we explored the Systems Biology modulated by A20 following extended LR in mice. Methodology and Principal Findings: We performed transcriptional profiling using Affymetrix-Mouse 430.2 arrays on liver mRNA retrieved from recombinant adenovirus A20 (rAd.A20) and rAd.$\beta$galactosidase treated livers, before and 24 hours after 78% LR. A20 overexpression impacted 1595 genes that were enriched for biological processes related to inflammatory and immune responses, cellular proliferation, energy production, oxidoreductase activity, and lipid and fatty acid metabolism. These pathways were modulated by A20 in a manner that favored decreased inflammation, heightened proliferation, and optimized metabolic control and energy production. Promoter analysis identified several transcriptional factors that implemented the effects of A20, including NF-$\kappa$B, CEBPA, OCT-1, OCT-4 and EGR1. Interactive scale-free network analysis captured the key genes that delivered the specific functions of A20. Most of these genes were affected at basal level and after resection. We validated a number of A20's target genes by real-time PCR, including p21, the mitochondrial solute carriers SLC25a10 and SLC25a13, and the fatty acid metabolism regulator, peroxisome proliferator activated receptor alpha. This resulted in greater energy production in A20-expressing livers following LR, as demonstrated by increased enzymatic activity of cytochrome c oxidase, or mitochondrial complex IV. Conclusion: This Systems Biology-based analysis unravels novel mechanisms supporting the pro-regenerative function of A20 in the liver, by optimizing energy production through improved lipid/fatty acid metabolism, and down-regulated inflammation. These findings support pursuit of A20-based therapies to improve patients' outcomes in the context of extreme liver injury and extensive LR for tumor treatment or donation

Autophagy Interplay with Apoptosis and Cell Cycle Regulation in the Growth Inhibiting Effect of Resveratrol in Glioma Cells

Prognosis of patients with glioblastoma (GBM) remains very poor, thus making the development of new drugs urgent. Resveratrol (Rsv) is a natural compound that has several beneficial effects such as neuroprotection and cytotoxicity for several GBM cell lines. Here we evaluated the mechanism of action of Rsv on human GBM cell lines, focusing on the role of autophagy and its crosstalk with apoptosis and cell cycle control. We further evaluated the role of autophagy and the effect of Rsv on GBM Cancer Stem Cells (gCSCs), involved in GBM resistance and recurrence. Glioma cells treated with Rsv was tested for autophagy, apoptosis, necrosis, cell cycle and phosphorylation or expression levels of key players of these processes. Rsv induced the formation of autophagosomes in three human GBM cell lines, accompanied by an upregulation of autophagy proteins Atg5, beclin-1 and LC3-II. Inhibition of Rsv-induced autophagy triggered apoptosis, with an increase in Bax and cleavage of caspase-3. While inhibition of apoptosis or autophagy alone did not revert Rsv-induced toxicity, inhibition of both processes blocked this toxicity. Rsv also induced a S-G2/M phase arrest, accompanied by an increase on levels of pCdc2(Y15), cyclin A, E and B, and pRb (S807/811) and a decrease of cyclin D1. Interestingly, this arrest was dependent on the induction of autophagy, since inhibition of Rsv-induced autophagy abolishes cell cycle arrest and returns the phosphorylation of Cdc2(Y15) and Rb(S807/811), and levels of cyclin A, and B to control levels. Finally, inhibition of autophagy or treatment with Rsv decreased the sphere formation and the percentage of CD133 and OCT4-positive cells, markers of gCSCs. In conclusion, the crosstalk among autophagy, cell cycle and apoptosis, together with the biology of gCSCs, has to be considered in tailoring pharmacological interventions aimed to reduce glioma growth using compounds with multiple targets such as Rsv