Nitrogen Deficiency Enhances Eggplant Defense against Western Flower Thrips via the Induction of the Jasmonate Pathway.

Plant nutrition is connected to defense against insect herbivores, but the exact mechanism underlying the effect of the nitrogen (N) supply on the anti-herbivore capacity of eggplants (Solanum melongena) has not been studied in detail. Therefore, we examined the impact of low (LN, 0.5 mM) and high (HN, 5 mM) nitrate levels on eggplant resistance against the western flower thrips Frankliniella occidentalis (WFT), a major destructive eggplant pest. Our results showed that LN plants displayed enhanced defense responses to WFT compared to HN plants. This included increased transcript levels of key genes in the jasmonic acid (JA) pathway, the accumulation of JA-amido conjugates (jasmonoyl-isoleucine, jasmonoyl-phenylalanine, and jasmonoyl-valine), JA precursor (12-oxophytodienoic acid), and methyl jasmonate, higher transcript levels of defense marker genes (MPK3, MPK7, and WRKY53), and increased activities of polyphenol oxidase and peroxidase upon a WFT attack. Our findings suggest that N deficiency can prime JA-mediated defense responses in eggplants, resulting in increased anti-herbivore resistance.

Fast, multiplexable and efficient somatic gene deletions in adult mouse skeletal muscle fibers using AAV-CRISPR/Cas9.

Molecular screens comparing different disease states to identify candidate genes rely on the availability of fast, reliable and multiplexable systems to interrogate genes of interest. CRISPR/Cas9-based reverse genetics is a promising method to eventually achieve this. However, such methods are sorely lacking for multi-nucleated muscle fibers, since highly efficient nuclei editing is a requisite to robustly inactive candidate genes. Here, we couple Cre-mediated skeletal muscle fiber-specific Cas9 expression with myotropic adeno-associated virus-mediated sgRNA delivery to establish a system for highly effective somatic gene deletions in mice. Using well-characterized genes, we show that local or systemic inactivation of these genes copy the phenotype of traditional gene-knockout mouse models. Thus, this proof-of-principle study establishes a method to unravel the function of individual genes or entire signaling pathways in adult skeletal muscle fibers without the cumbersome requirement of generating knockout mice.

Hypoxia-induced reprogramming of glucose-dependent metabolic pathways maintains the stemness of human bone marrow-derived endothelial progenitor cells.

The benefits of hypoxia for maintaining the stemness of cultured human bone marrow-derived endothelial progenitor cells (BM EPCs) have previously been demonstrated but the mechanisms responsible remain unclear. Growing evidences suggest that cellular metabolism plays an important role in regulating stem cell fate and self-renewal. Here we aimed to detect the changes of glucose metabolism and to explore its role on maintaining the stemness of BM EPCs under hypoxia. We identified the metabolic status of BM EPCs by using extracellular flux analysis, LC-MS/MS, and 13C tracing HPLC-QE-MS, and found that hypoxia induced glucose metabolic reprogramming, which manifested as increased glycolysis and pentose phosphate pathway (PPP), decreased tricarboxylic acid (TCA) and mitochondrial respiration. We further pharmacologically altered the metabolic status of cells by employing various of inhibitors of key enzymes of glycolysis, PPP, TCA cycle and mitochondria electron transport chain (ETC). We found that inhibiting glycolysis or PPP impaired cell proliferation either under normoxia or hypoxia. On the contrary, inhibiting pyruvate oxidation, TCA or ETC promoted cell proliferation under normoxia mimicking hypoxic conditions. Moreover, promoting pyruvate oxidation reverses the maintenance effect of hypoxia on cell stemness. Taken together, our data suggest that hypoxia induced glucose metabolic reprogramming maintains the stemness of BM EPCs, and artificial manipulation of cell metabolism can be an effective way for regulating the stemness of BM EPCs, thereby improving the efficiency of cell expansion in vitro.

Social Media Data Mining of Antitobacco Campaign Messages: Machine Learning Analysis of Facebook Posts.

BACKGROUND: Social media platforms provide a valuable source of public health information, as one-third of US adults seek specific health information online. Many antitobacco campaigns have recognized such trends among youth and have shifted their advertising time and effort toward digital platforms. Timely evidence is needed to inform the adaptation of antitobacco campaigns to changing social media platforms. OBJECTIVE: In this study, we conducted a content analysis of major antitobacco campaigns on Facebook using machine learning and natural language processing (NLP) methods, as well as a traditional approach, to investigate the factors that may influence effective antismoking information dissemination and user engagement. METHODS: We collected 3515 posts and 28,125 associated comments from 7 large national and local antitobacco campaigns on Facebook between 2018 and 2021, including the Real Cost, Truth, CDC Tobacco Free (formally known as Tips from Former Smokers, where "CDC" refers to the Centers for Disease Control and Prevention), the Tobacco Prevention Toolkit, Behind the Haze VA, the Campaign for Tobacco-Free Kids, and Smoke Free US campaigns. NLP methods were used for content analysis, including parsimonious rule-based models for sentiment analysis and topic modeling. Logistic regression models were fitted to examine the relationship of antismoking message-framing strategies and viewer responses and engagement. RESULTS: We found that large campaigns from government and nonprofit organizations had more user engagements compared to local and smaller campaigns. Facebook users were more likely to engage in negatively framed campaign posts. Negative posts tended to receive more negative comments (odds ratio [OR] 1.40, 95% CI 1.20-1.65). Positively framed posts generated more negative comments (OR 1.41, 95% CI 1.19-1.66) as well as positive comments (OR 1.29, 95% CI 1.13-1.48). Our content analysis and topic modeling uncovered that the most popular campaign posts tended to be informational (ie, providing new information), where the key phrases included talking about harmful chemicals (n=43, 43%) as well as the risk to pets (n=17, 17%). CONCLUSIONS: Facebook users tend to engage more in antitobacco educational campaigns that are framed negatively. The most popular campaign posts are those providing new information, with key phrases and topics discussing harmful chemicals and risks of secondhand smoke for pets. Educational campaign designers can use such insights to increase the reach of antismoking campaigns and promote behavioral changes.

Associations between use of specific social media sites and electronic cigarette use among college students.

OBJECTIVE: To examine dose-response associations between use of specific social media sites and the use of electronic cigarettes (e-cigarettes) and traditional cigarettes. METHODS: This was a cross-sectional study of 298 first-year college students enrolled in the fall 2019 semester at a large state university. Heckman selection and Probit model were used to estimate associations between use of specific social media sites and e-cigarette/traditional cigarette use. RESULTS: Each additional hour per day spent on Snapchat was associated with a 4.61% increase in the probability of lifetime e-cigarette use. In addition, among current e-cigarette users, more time spent on Snapchat was associated with more frequent e-cigarette use (marginal effects: 0.13, p = 0.001). Facebook, Twitter, Snapchat and Instagram were not associated with traditional cigarette smoking. CONCLUSION: Snapchat was the only major social media platform associated with both lifetime and current e-cigarette use.

The Association Between Tobacco Retailer Outlet Density and Prevalence of Cigarette Smoking in Virginia.

OBJECTIVE: We examine the association between tobacco retail outlet density and adult smoking prevalence at the county level in Virginia, controlling for spatial autocorrelations. AIMS AND METHODS: Pooling data from 2020 County Health Rankings (compiled data from various sources including, but not limited to, the National Center for Health Statistics-Mortality Files, the Behavioral Risk Factor Surveillance System (BRFSS), and the American Community Survey) and Counter Tools, we conducted regression analyses that accounted for spatial autocorrelation (spatial lag models, LMlag) and adjusted for county-level access to healthcare, demographics, SES, environmental factors, risk conditions or behaviors, and population health measures. RESULTS: Our estimates provide evidence that every increase of one tobacco retail outlet per 1000 persons was associated with 1.16 percentage points (95% CI: 0.80-1.52) higher smoking prevalence at the county level in Virginia after controlling for spatial autocorrelation. The effect of outlet density was largely explained by social determinants of health such as SES, risky conditions or behaviors, and environmental factors. We further noticed that the impact of social determinants of health were closely related and can be explained by indicators of population health (rates of mental distress (ß = 1.49, 95% CI: 1.31-1.67) and physical inactivity (ß = 0.07, 95% CI: 0.04-0.10). CONCLUSIONS: Although higher tobacco outlet density was associated with an increase in county-level smoking prevalence, the impact of outlet density was largely explained by social determinants of health and mental illness. Improving well-being at the community level could be a promising strategy in future tobacco control policies. IMPLICATION: The influence of tobacco outlet density seems to be explained by other social determinants of health and population level of mental or physical health. Thus, efforts to reduce tobacco use and consequent negative health effects should explore the impact of improving regional living standards. However, a sole focus on economic growth may not be sufficient, whereas a focus on such things as promoting work-life balance and improving overall well-being at the community level may be more.

E-cigarette use during pregnancy and its association with adverse birth outcomes in the US.

The popularity of e-cigarette use among young adults is a growing concern. However, little is known about factors associated with e-cigarette use in pregnant women and birth outcomes. In this retrospective cohort study, we evaluated the influence of several factors on behavioral changes in e-cigarette use before and during pregnancy, and assessed the association between e-cigarette use and subsequent birth outcomes among pregnant women. The Population Assessment of Tobacco and Health (PATH) study, a government-sponsored national longitudinal study based in the US, Waves 1 through 4 (2013-2018) were used. Multivariate logistic regressions were conducted to estimate behavioral changes in e-cigarette use during pregnancy and subsequent influence on high-risk birth (e.g., preterm birth, low birth weight, birth defects, etc.) and fetal death. Although pregnant women who quit vaping before pregnancy (OR = 1.14, 95% CI 0.54-2.40) or had any use during pregnancy (OR = 1.19, 95% CI 0.38-3.73) showed non-differential risk of having a high-risk birth in comparison to women who did not initiate vaping, we observed that the usage of mint/menthol flavor was correlated with higher risk of fetus death (OR = 3.27, 95% CI 1.17-9.19). Healthcare providers should encourage e-cigarette users to quit prior to and during early pregnancy.

Dual roles of mTORC1-dependent activation of the ubiquitin-proteasome system in muscle proteostasis.

Muscle size is controlled by the PI3K-PKB/Akt-mTORC1-FoxO pathway, which integrates signals from growth factors, energy and amino acids to activate protein synthesis and inhibit protein breakdown. While mTORC1 activity is necessary for PKB/Akt-induced muscle hypertrophy, its constant activation alone induces muscle atrophy. Here we show that this paradox is based on mTORC1 activity promoting protein breakdown through the ubiquitin-proteasome system (UPS) by simultaneously inducing ubiquitin E3 ligase expression via feedback inhibition of PKB/Akt and proteasome biogenesis via Nuclear Factor Erythroid 2-Like 1 (Nrf1). Muscle growth was restored by reactivation of PKB/Akt, but not by Nrf1 knockdown, implicating ubiquitination as the limiting step. However, both PKB/Akt activation and proteasome depletion by Nrf1 knockdown led to an immediate disruption of proteome integrity with rapid accumulation of damaged material. These data highlight the physiological importance of mTORC1-mediated PKB/Akt inhibition and point to juxtaposed roles of the UPS in atrophy and proteome integrity.

Does Physical Activity Predict Obesity-A Machine Learning and Statistical Method-Based Analysis.

BACKGROUND: Obesity prevalence has become one of the most prominent issues in global public health. Physical activity has been recognized as a key player in the obesity epidemic. OBJECTIVES: The objectives of this study are to (1) examine the relationship between physical activity and weight status and (2) assess the performance and predictive power of a set of popular machine learning and traditional statistical methods. METHODS: National Health and Nutrition Examination Survey (NHANES, 2003 to 2006) data were used. A total of 7162 participants met our inclusion criteria (3682 males and 3480 females), with average age ranging from 48.6 (normal weight) to 52.1 years old (overweight). Eleven classifying algorithms-including logistic regression, naïve Bayes, Radial Basis Function (RBF), local k-nearest neighbors (k-NN), classification via regression (CVR), random subspace, decision table, multiobjective evolutionary fuzzy classifier, random tree, J48, and multilayer perceptron-were implemented and evaluated, and they were compared with traditional logistic regression model estimates. RESULTS: With physical activity and basic demographic status, of all methods analyzed, the random subspace classifier algorithm achieved the highest overall accuracy and area under the receiver operating characteristic (ROC) curve (AUC). The duration of vigorous-intensity activity in one week and the duration of moderate-intensity activity in one week were important attributes. In general, most algorithms showed similar performance. Logistic regression was middle-ranking in terms of overall accuracy, sensitivity, specificity, and AUC among all methods. CONCLUSIONS: Physical activity was an important factor in predicting weight status, with gender, age, and race/ethnicity being less but still essential factors associated with weight outcomes. Tailored intervention policies and programs should target the differences rooted in these demographic factors to curb the increase in the prevalence of obesity and reduce disparities among sub-demographic populations.

Comparison of three lymph node staging systems in evaluating the prognosis of patients with pT3 esophageal squamous cell carcinoma.

To explore the prognostic value of three lymph node staging systems, including number of positive lymph nodes (pN), lymph node ratio (LNR), and log odds of positive lymph nodes (LODDS), in patients with pT3 stage esophageal squamous cell carcinoma (ESCC). Data from 1667 patients with pT3 stage ESCC who underwent surgical resection were reviewed. The log-rank test was used to assess the differences in overall survival (OS) between groups. Multivariate analysis was performed to identify independent prognostic factors. The receiver operating characteristic curve was used to assess the prognostic accuracy of the three staging methods. The median survival time for the entire group was 48.0 months, and the 1-, 3- and 5-year OS rates were 83.9%, 55.1% and 66.6%, respectively. All three lymph node staging systems were significantly correlated with OS in univariate and multivariate analyses. However, LNR and LODDS staging systems could more accurately predict survival than the pN staging system in patients with < 15 lymph nodes dissected, while LODDS have the best prognostic homogeneity. All three staging systems could be used for prognostic assessment in pT3 stage ESCC. But LODDS staging system might be superior to the others due to its prognostic homogeneity.

The nuclear gene rpl18 regulates erythroid maturation via JAK2-STAT3 signaling in zebrafish model of Diamond-Blackfan anemia.

Diamond-Blackfan anemia (DBA) is a rare, inherited bone marrow failure syndrome, characterized by red blood cell aplasia, developmental abnormalities, and enhanced risk of malignancy. However, the underlying pathogenesis of DBA is yet to be understood. Recently, mutations in the gene encoding ribosomal protein (RP) L18 were identified in DBA patients. RPL18 is a crucial component of the ribosomal large subunit but its role in hematopoiesis remains unknown. To genetically model the ribosomal defect identified in DBA, we generated a rpl18 mutant line in zebrafish, using CRISPR/Cas9 system. Molecular characterization of this mutant line demonstrated that Rpl18 deficiency mirrored the erythroid defects of DBA, namely a lack of mature red blood cells. Rpl18 deficiency caused an increase in p53 activation and JAK2-STAT3 activity. Furthermore, we found inhibitors of JAK2 or STAT3 phosphorylation could rescue anemia in rpl18 mutants. Our research provides a new in vivo model of Rpl18 deficiency and suggests involvement of signal pathway of JAK2-STAT3 in the DBA pathogenesis.

mTORC1 signalling is not essential for the maintenance of muscle mass and function in adult sedentary mice.

BACKGROUND: The balance between protein synthesis and degradation (proteostasis) is a determining factor for muscle size and function. Signalling via the mammalian target of rapamycin complex 1 (mTORC1) regulates proteostasis in skeletal muscle by affecting protein synthesis and autophagosomal protein degradation. Indeed, genetic inactivation of mTORC1 in developing and growing muscle causes atrophy resulting in a lethal myopathy. However, systemic dampening of mTORC1 signalling by its allosteric inhibitor rapamycin is beneficial at the organismal level and increases lifespan. Whether the beneficial effect of rapamycin comes at the expense of muscle mass and function is yet to be established. METHODS: We conditionally ablated the gene coding for the mTORC1-essential component raptor in muscle fibres of adult mice [inducible raptor muscle-specific knockout (iRAmKO)]. We performed detailed phenotypic and biochemical analyses of iRAmKO mice and compared them with muscle-specific raptor knockout (RAmKO) mice, which lack raptor in developing muscle fibres. We also used polysome profiling and proteomics to assess protein translation and associated signalling in skeletal muscle of iRAmKO mice. RESULTS: Analysis at different time points reveal that, as in RAmKO mice, the proportion of oxidative fibres decreases, but slow-type fibres increase in iRAmKO mice. Nevertheless, no significant decrease in body and muscle mass or muscle fibre area was detected up to 5 months post-raptor depletion. Similarly, ex vivo muscle force was not significantly reduced in iRAmKO mice. Despite stable muscle size and function, inducible raptor depletion significantly reduced the expression of key components of the translation machinery and overall translation rates. CONCLUSIONS: Raptor depletion and hence complete inhibition of mTORC1 signalling in fully grown muscle leads to metabolic and morphological changes without inducing muscle atrophy even after 5 months. Together, our data indicate that maintenance of muscle size does not require mTORC1 signalling, suggesting that rapamycin treatment is unlikely to negatively affect muscle mass and function.

Loss of rps9 in Zebrafish Leads to p53-Dependent Anemia.

Ribosome is a vital molecular machine for protein translation in the cell. Defects in several ribosomal proteins including RPS19, RPL11 and RPS14 have been observed in two types of anemia: Diamond Blackfan Anemia and 5q- syndrome. In zebrafish, deficiency of these ribosomal proteins shows similar anemic phenotype. It remains to be determined if any other ribosome proteins are similarly involved in regulating erythropoiesis. Here we generated mutations in zebrafish rps9, a rarely studied ribosomal protein gene, and investigated its function. Analysis of this mutant demonstrates that rps9 disruption leads to impairment of erythrocyte maturation, resulting in anemia. In addition, the overall phenotype including the anemic state is p53-dependent in rps9 mutants. Furthermore, this anemic state can be partially relieved by the treatment of L-leucine, and dexamethasone, which have been previously used in rescuing the phenotype of other ribosomal protein mutants. Finally, by comparing the phenotype, we show that there are considerable differences in morphology, cytomorphology, and hemoglobin levels for four ribosomal protein mutants in zebrafish. Based on the observed difference, we suggest that the level of anemic severity correlates with the delayed status of erythrocyte maturation in zebrafish models.

MMP9 inhibition increases erythropoiesis in RPS14-deficient del(5q) MDS models through suppression of TGF-ß pathways.

The del(5q) myelodysplastic syndrome (MDS) is a distinct subtype of MDS, associated with deletion of the ribosomal protein S14 (RPS14) gene that results in macrocytic anemia. This study sought to identify novel targets for the treatment of patients with del(5q) MDS by performing an in vivo drug screen using an rps14-deficient zebrafish model. From this, we identified the secreted gelatinase matrix metalloproteinase 9 (MMP9). MMP9 inhibitors significantly improved the erythroid defect in rps14-deficient zebrafish. Similarly, treatment with MMP9 inhibitors increased the number of colony forming unit-erythroid colonies and the CD71+ erythroid population from RPS14 knockdown human BMCD34+ cells. Importantly, we found that MMP9 expression is upregulated in RPS14-deficient cells by monocyte chemoattractant protein 1. Double knockdown of MMP9 and RPS14 increased the CD71+ population compared with RPS14 single knockdown, suggesting that increased expression of MMP9 contributes to the erythroid defect observed in RPS14-deficient cells. In addition, transforming growth factor ß (TGF-ß) signaling is activated in RPS14 knockdown cells, and treatment with SB431542, a TGF-ß inhibitor, improved the defective erythroid development of RPS14-deficient models. We found that recombinant MMP9 treatment decreases the CD71+ population through increased SMAD2/3 phosphorylation, suggesting that MMP9 directly activates TGF-ß signaling in RPS14-deficient cells. Finally, we confirmed that MMP9 inhibitors reduce SMAD2/3 phosphorylation in RPS14-deficient cells to rescue the erythroid defect. In summary, these study results support a novel role for MMP9 in the pathogenesis of del(5q) MDS and the potential for the clinical use of MMP9 inhibitors in the treatment of patients with del(5q) MDS.

Bone marrow mesenchymal stem cell-derived exosomes protect against myocardial infarction by promoting autophagy.

Exosomes have been demonstrated to be effective in the treatment of a variety of cardiac disorders. However, the effects of mesenchymal stem cell (MSC) exosomes on myocardial infarction is yet to be determined. The current study aimed to investigate the potential therapeutic effects of MSC exosomes on myocardial injuries that are caused by myocardial infarction. MSCs were isolated from rat bone marrow and were used for exosome enrichment using culture medium. Confirmation that MSCs and exosomes had been successfully extracted was performed using flow cytometry, electron microscopy and western blot analysis. A rat myocardial ischemia reperfusion (I/R) model was established by ligation of the left anterior descending coronary artery. Rat myocardial injuries were determined using 2,3,5-triphenyltetrazolium chloride, Masson and TUNEL staining. H9c2 cell proliferation, apoptosis and migration were analyzed using 5-ethynyl-2'-deoxyuridine, Hoechst staining, flow cytometry and Transwell assays. Marker gene expression was evaluated using reverse transcription-quantitative PCR, western blot analysis and immunofluorescence. Rat MSC exosomes were revealed to suppress myocardial injury and the myocardiocyte functions that were induced by I/R. The results also demonstrated decreased apoptotic protease activating factor-1 and increased autophagy-related protein 13 expression. The H9c2 cell proliferation and migration inhibition, as well as cell apoptosis during hypoxia-reoxygenation (H/R), were suppressed by rat MSC exosomes, with an alteration of the expression of apoptotic and autophagic genes also being demonstrated. The application of autophagy inhibitor 3-methyladenine significantly mitigated the effect of exosomes on H9c2 cell proliferation and apoptosis, which were induced by H/R. Rat MSC exosomes inhibited myocardial infarction pathogenesis, possibly by regulating autophagy.

Oct4 upregulates osteopontin via Egr1 and is associated with poor outcome in human lung cancer.

BACKGROUND: Roles of cancer stem cells and early growth response gene 1 (Egr1) in carcinogenesis have been extensively studied in lung cancer. However, the role of Egr1 in the metastasis of lung cancer remains undetermined, especially in regard to stem cell-related pathways. METHODS: Egr1, osteopontin (OPN) and Oct4 expression in human lung cancer was determined by performing immunohistochemistry. Immunoblotting, ELISA, luciferase reporter assay, chromatin immunoprecipitation assay and RT-PCR were performed to validate the regulation of Oct4-Egr1-OPN axis. Moreover, the effect of Oct4-Egr1-OPN axis on lung cancer progression was evaluated by cell migration assay and mice study. RESULTS: We detected Oct4, Egr1, and OPN expression in clinical specimens from 79 lung cancer patients, including 72 adenocarcinomas and 7 squamous cell carcinomas. High expression of Oct4, Egr1, and OPN accounted for 53, 51, and 57% of the patients, respectively. All of the three biomarkers were positively correlated in clinical human lung cancer. Patients with high expression of OPN were significantly associated with shorter disease-free survivals than those with low expression of OPN (p < 0.05). In lung cancer cells, Oct4 transactivated the Egr1 promoter and upregulated Egr1 expression. In a human lung cancer xenograft model, Oct4-overexpressing tumors expressed elevated levels of Egr1. Furthermore, overexpression of Oct4 in lung cancer cells increased the metastatic potential. CONCLUSIONS: Egr1 exerts a promoting effect on cancer metastasis in Oct4-overexpressing lung cancer. Thus, therapeutic strategies targeting the Oct4/Egr1/OPN axis may be further explored for the treatment of lung cancer, especially when lung cancer is refractory to conventional treatment due to cancer stem cells.

mTORC1 and PKB/Akt control the muscle response to denervation by regulating autophagy and HDAC4.

Loss of innervation of skeletal muscle is a determinant event in several muscle diseases. Although several effectors have been identified, the pathways controlling the integrated muscle response to denervation remain largely unknown. Here, we demonstrate that PKB/Akt and mTORC1 play important roles in regulating muscle homeostasis and maintaining neuromuscular endplates after nerve injury. To allow dynamic changes in autophagy, mTORC1 activation must be tightly balanced following denervation. Acutely activating or inhibiting mTORC1 impairs autophagy regulation and alters homeostasis in denervated muscle. Importantly, PKB/Akt inhibition, conferred by sustained mTORC1 activation, abrogates denervation-induced synaptic remodeling and causes neuromuscular endplate degeneration. We establish that PKB/Akt activation promotes the nuclear import of HDAC4 and is thereby required for epigenetic changes and synaptic gene up-regulation upon denervation. Hence, our study unveils yet-unknown functions of PKB/Akt-mTORC1 signaling in the muscle response to nerve injury, with important implications for neuromuscular integrity in various pathological conditions.

Rescue of spinal muscular atrophy mouse models with AAV9-Exon-specific U1 snRNA.

Spinal Muscular Atrophy results from loss-of-function mutations in SMN1 but correcting aberrant splicing of SMN2 offers hope of a cure. However, current splice therapy requires repeated infusions and is expensive. We previously rescued SMA mice by promoting the inclusion of a defective exon in SMN2 with germline expression of Exon-Specific U1 snRNAs (ExspeU1). Here we tested viral delivery of SMN2 ExspeU1s encoded by adeno-associated virus AAV9. Strikingly the virus increased SMN2 exon 7 inclusion and SMN protein levels and rescued the phenotype of mild and severe SMA mice. In the severe mouse, the treatment improved the neuromuscular function and increased the life span from 10 to 219 days. ExspeU1 expression persisted for 1 month and was effective at around one five-hundredth of the concentration of the endogenous U1snRNA. RNA-seq analysis revealed our potential drug rescues aberrant SMA expression and splicing profiles, which are mostly related to DNA damage, cell-cycle control and acute phase response. Vastly overexpressing ExspeU1 more than 100-fold above the therapeutic level in human cells did not significantly alter global gene expression or splicing. These results indicate that AAV-mediated delivery of a modified U1snRNP particle may be a novel therapeutic option against SMA.

A novel 3D breast-cancer-on-chip platform for therapeutic evaluation of drug delivery systems.

The ability to rapidly screen drugs and drug delivery systems with a more accurate tumor model to better predict their in vivo performance is of great importance in drug development, because there have been some limitations in currently used tumor models. To address this problem, we developed an in vitro breast tumor model on a chip, composed of a microvessel wall, the extracellular matrix (ECM) and uniformly sized multicellular tumor spheroids (MCTS), for the evaluation of nanoparticle-based drug delivery systems. A carbon dots (CDs)-based drug delivery system was synthesized as a model to evaluate the real-time monitoring ability of the system transport through the endothelium and the penetrability into MCTS with a high spatio-temporal resolution on the established platform. Moreover, a modified 96-well plate was used to hold the microfluidic devices for in situ cytotoxicity assays of the MCTS by a microplate reader. Our findings revealed that the synthesized drug delivery system could be transported across an endothelial monolayer within 3 h and was nontoxic to the cells throughout the experiment. In addition, we demonstrated the capabilities of this model by assessing the delivery and efficacy of the drug delivery system in BT549 and T47D spheroids, two cell lines representative of triple negative breast cancer (TNBC) and non-TNBC, respectively. This microfluidic platform enables evaluation of dynamic transport behavior and in situ cytotoxicity evaluation in one system. The established platform provides a more accurate and low-cost in vitro model for rapid drug screening in pre-clinical studies.

Adsorptive interaction of cationic pharmaceuticals on activated charcoal: Experimental determination and QSAR modelling.

Due to high mobility and specific toxic actions of the ionizable pharmaceuticals in surface water with a normal range of pH, the pharmaceuticals should be removed before being discharged. Therefore, this study investigated the adsorptive interactions between cationic pharmaceuticals and a popular adsorbent (i.e., activated charcoal) frequently used in water treatment processes. For that, we performed isotherm experiments and then the results were plotted by Langmuir model to determine the adsorption affinity (b) and capacity (qm). Afterwards, to interpret the adsorption behaviors, two simple prediction models were developed based on quantitative structure-activity relationships (QSAR). In the modelling, molecular weight (MW), polar surface area (PSA), and octanol-water partitioning coefficient (log P) were used as model parameters. In the results, the combinations of these three parameters could predict the adsorption affinity and capacity in R2 of 0.85 and 0.80, respectively. The robustness of models was validated by leave-one-out cross-validation (Q2LOO) and the estimated Q2LOO values were 0.60 and 0.55 for the adsorption affinity and capacity, respectively, which are higher than the acceptability of standard i.e., 0.5.