VCAM-1 Targeted Lipopolyplexes as Vehicles for Efficient Delivery of shRNA-Runx2 to Osteoblast-Differentiated Valvular Interstitial Cells; Implications in Calcific Valve Disease Treatment.

Calcific aortic valve disease (CAVD) is a progressive inflammatory disorder characterized by extracellular matrix remodeling and valvular interstitial cells (VIC) osteodifferentiation leading to valve leaflets calcification and impairment movement. Runx2, the master transcription factor involved in VIC osteodifferentiation, modulates the expression of other osteogenic molecules. Previously, we have demonstrated that the osteoblastic phenotypic shift of cultured VIC is impeded by Runx2 silencing using fullerene (C60)-polyethyleneimine (PEI)/short hairpin (sh)RNA-Runx2 (shRunx2) polyplexes. Since the use of polyplexes for in vivo delivery is limited by their instability in the plasma and the non-specific tissue interactions, we designed and obtained targeted, lipid-enveloped polyplexes (lipopolyplexes) suitable for (1) systemic administration and (2) targeted delivery of shRunx2 to osteoblast-differentiated VIC (oVIC). Vascular cell adhesion molecule (VCAM)-1 expressed on the surface of oVIC was used as a target, and a peptide with high affinity for VCAM-1 was coupled to the surface of lipopolyplexes encapsulating C60-PEI/shRunx2 (V-LPP/shRunx2). We report here that V-LPP/shRunx2 lipopolyplexes are cyto- and hemo-compatible and specifically taken up by oVIC. These lipopolyplexes are functional as they downregulate the Runx2 gene and protein expression, and their uptake leads to a significant decrease in the expression of osteogenic molecules (OSP, BSP, BMP-2). These results identify V-LPP/shRunx2 as a new, appropriately directed vehicle that could be instrumental in developing novel strategies for blocking the progression of CAVD using a targeted nanomedicine approach.

Achalasia at the Crossroads Between Specialties.

Background: Achalasia is a condition that can be treated either by surgery, such as Heller myotomy associated with an antireflux procedure, or by pneumatic dilation, the choice of type of treatment being a widely debated topic nowadays. Methods: We selected patients with the diagnosis of achalasia, calculating the Eckardt score on admission and they were treated by pneumatic dilation, respectively by myotomy associated with fundoplication. Therapeutic success at the end of treatment was analyzed, as well as that in subsequent follow-ups. At the same time, other important aspects of the study were quality of life and complications. Results: Forty-eight patients were included, 20 in the group of those treated by pneumatic dilation, and 28 treated by surgery. The results of the therapeutic success were to the advantage of the surgery, both after the completion of the treatment, and in the follow-ups from one year to 2 years (96.4% vs 90%, respectively 92.9% vs. 85%). The quality of life was better in patients with pneumatic dilation throughout the period. Conclusions: Surgical treatment of achalasia has a higher success rate than pneumatic dilation, but the latter is associated with a better quality of life.

Nanocarriers of shRNA-Runx2 directed to collagen IV as a nanotherapeutic system to target calcific aortic valve disease.

Runx2 is a key transcription factor involved in valvular interstitial cells (VIC) osteodifferentiation, a process actively entwined with the calcific aortic valve disease (CAVD). We hypothesize that a strategy intended to silence Runx2 could be a valuable novel therapeutic option for CAVD. To this intent, we aimed at (i) developing targeted nanoparticles for efficient delivery of short hairpin (sh)RNA sequences specific for Runx2 to the aortic valve employing a relevant mouse model for CAVD and (ii) investigate their therapeutic potential in osteoblast-differentiated VIC (oVIC) cultivated into a 3D scaffold. Since collagen IV was used as a target, a peptide that binds specifically to collagen IV (Cp) was conjugated to the surface of lipopolyplexes encapsulating shRNA-Runx2 (Cp-LPP/shRunx2). The results showed that Cp-LPP/shRunx2 were (i) cytocompatible; (ii) efficiently taken up by 3D-cultured oVIC; (iii) diminished the osteodifferentiation of human VIC (cultured in a 3D hydrogel-derived from native aortic root) by reducing osteogenic molecules expression, alkaline phosphatase activity, and calcium concentration; and (iv) were recruited in aortic valve leaflets in a murine model of atherosclerosis. Taken together, these data recommend Cp-LPP/shRunx2 as a novel targeted nanotherapy to block the progression of CAVD, with a good perspective to be introduced in practical use.

Predictors of Economic Outcomes among Romanian Youth: The Influence of Education-An Empirical Approach Based on Elastic Net Regression.

Young people have to be provided with opportunities to access prosperous, resilient and fulfilling lives. Investing in education and skills is considered one of the most important ways to support young people's well-being and to enable them to enjoy good career prospects. Using the framework of human capital theory, we explored the role of education among the factors explaining wage variation among Romanian youth. We built our analysis on micro-data for Romania from the EU Statistics on Income and Living Conditions 2020. In order to identify the most important factors influencing the wage distribution, we employed the elastic net regression approach. Moreover, we considered the phenomenon of expansion of education and ran the analysis by alternately using a traditional measure for education and a relative measure reflecting the theory of education as positional good. We ran the analysis for different cohorts of the population, focusing the discussion on the results for young people. Our findings confirm the importance of education for wage distribution together with other factors of influence, such as gender, degree of urbanization, region, sector of employment and working experience. Our conclusions are relevant for designing more effective educational and social policies to deal with various disadvantages faced by youth in Romania.

VLA4-Enhanced Allogeneic Endothelial Progenitor Cell-Based Therapy Preserves the Aortic Valve Function in a Mouse Model of Dyslipidemia and Diabetes.

The number and function of endothelial progenitor cells (EPCs) are reduced in diabetes, contributing to deteriorated vascular repair and the occurrence of cardiovascular complications. Here, we present the results of treating early diabetic dyslipidemic mice or dyslipidemic with disease-matched EPCs modified to overexpress VLA4 (VLA4-EPCs) as compared with the treatment of EPCs transfected with GFP (GFP-EPCs) as well as EPCs from healthy animals. Organ imaging of injected PKH26-stained cells showed little pulmonary first-pass effects and distribution in highly vascularized organs, with splenic removal from circulation, mostly in non-diabetic animals. Plasma measurements showed pronounced dyslipidemia in all animals and glycaemia indicative of diabetes in streptozotocin-injected animals. Echocardiographic measurements performed 3 days after the treatment showed significantly improved aortic valve function in animals treated with VLA4-overexpressing EPCs compared with GFP-EPCs, and similar results in the groups treated with healthy EPCs and VLA4-EPCs. Immunohistochemical analyses revealed active inflammation and remodelling in all groups but different profiles, with higher MMP9 and lower P-selectin levels in GFP-EPCs, treated animals. In conclusion, our experiments show that genetically modified allogeneic EPCs might be a safe treatment option, with bioavailability in the desired target compartments and the ability to preserve aortic valve function in dyslipidemia and diabetes.

What Encourages Longer Educational Careers in Tertiary Education? A Three-Level Approach for the Case of Romanian Universities.

Students' commitment and engagement in the educational process are shaped by a dense combination of factors, with effects on educational attainment and on the length of their educational careers. Decisions of prolonging education by enrolling in master's degrees are beneficial for both individuals and societies, as such programs provide higher levels of specialized skills Longer educational careers are favored by a mix of factors acting at the level of individual, university, or wider environment. We focus our study on exploring factors conducive for students' intentions to pursue master's degrees considering longer educational careers as desirable outcomes. Thus, this article investigates how the individual and environmental factors interplay and shape the predisposition of students to prolong their educational career by enrolling in master's degrees. For this, we applied three-level logistic regression models for a sample of 502 students enrolled in their final year of bachelor studies grouped by universities and universities grouped by counties. The empirical results revealed that the final grade, the father level of education, the type of working contract, and job seniority are individual-level determinants influencing the decision of enrolment in a master's program. At the university level, the type of university and the university performance score positively impact the students' decision to enroll in a master's program. At the county level, the empirical evidence pointed out the significance of determinants such as the proportion of students enrolled in bachelor studies; participation rate in education and training; employment level in high-technology sectors (HTC), total-knowledge intensive sectors (KIS), and knowledge-intensive high-technology sectors (KIS_HTC); proportion of persons with tertiary education employed in science and technology; proportion of scientists and engineers; local development; R&D expenditure, personnel, and researchers in the business sector; average gross earnings; density of active firms; birth rate of companies; proportion of innovative enterprises or those introducing product innovations on the decision to enroll in a master's program.

P-selectin targeted RAGE-shRNA lipoplexes alleviate atherosclerosis-associated inflammation.

The receptor for advanced glycation end products (RAGE) plays a central role in the chronic inflammatory process associated with atherosclerosis development. We aimed to develop lipoplexes carrying RAGE-short hairpin (sh) RNA, targeted to the adhesion molecule P-selectin, selectively expressed on the surface of activated endothelium (Psel-lipo/shRAGE) to down-regulate RAGE expression as a therapeutic strategy for atherosclerosis. In vitro, Psel-lipo/shRAGE lipoplexes were efficiently taken up by activated endothelial cells (EC), decreased the expression of RAGE protein, and proved to be functional by reducing the monocyte adhesion to activated EC. In ApoE-deficient mice, the targeted lipoplexes accumulated specifically and efficiently transfected the aorta. The repeated administration of Psel-lipo/shRAGE lipoplexes, twice per week for one month: i) reduced the expression of RAGE protein in the aorta by decreasing the expression of NF-kB and TNF-α; ii) diminished the plasma levels of TNF-α, IL6, IL-1ß, and MCP-1; iii) inhibited the atherosclerotic plaque development and iv) had no significant adverse effects. In conclusion, the newly developed Psel-lipo/shRAGE lipoplexes reduce the inflammatory processes associated with RAGE signaling and the progression of atherosclerosis in ApoE-deficient mice. Downregulation of RAGE employing these lipoplexes may represent a promising new targeted therapy to block atherosclerosis progression.

Perceived Academic Self-Efficacy among Romanian Upper Secondary Education Students.

Perceived academic self-efficacy represents an important component of students' mental health and well-being. The link between efficacy beliefs and level of academic performances relies on the fact that they regulate the anxiety and foster motivation, school engagement, effort, and perseverance of students. This paper aims to identify factors that are conducive for more efficacious beliefs in different socio-economic and educational contexts. We build our analysis on data collected from a sample of Romanian upper secondary education students on their beliefs in relation to their ability to perform at the baccalaureate exam. We employ decision tree models in order to unveil the way factors interact and predict perceived academic self-efficacy, with focus on the positive support received from parents and teachers, as well as on features of the school environment. Our results can be useful for building more resilient educational environments that support mental health and academic well-being of students.

Evaluation of VCAM-1 Targeted Naringenin/Indocyanine Green-Loaded Lipid Nanoemulsions as Theranostic Nanoplatforms in Inflammation.

Naringenin, an anti-inflammatory citrus flavonoid, is restrained from large-scale use by its reduced water solubility and bioavailability. To overcome these limitations, naringenin was loaded into lipid nanoemulsions directed towards vascular cell adhesion molecule (VCAM)-1, exposed by activated endothelium, and delivered intravenously in a murine model of lipopolysaccharide (LPS)-induced inflammation. To follow the in vivo bio-distribution, naringenin-loaded nanoemulsions were labeled with near-infrared probe Indocyanine Green (ICG). Based on ICG fluorescence, a VCAM-1-dependent retention of nanoemulsions was detected in the heart and aorta, while ultra-high-performance liquid chromatography (UHPLC) measurements showed a target-selective accumulation of naringenin in the heart and lungs. Correlated, fluorescence and UHPLC data indicated a mixed behavior of the VCAM-1 directed nanoparticles, which were driven not only by the targeting moiety but also by passive retention. The treatment with naringenin-loaded nanoemulsions reduced the mRNA levels of some inflammatory mediators in organs harvested from mice with acute inflammation, indicative of their anti-inflammatory potential. The data support a novel theranostic nanoplatform for inflammation, the naringenin/ICG-loaded nanoparticles that either by passive accumulation or effective targeting of the activated endothelium can be employed for imaging inflamed vascular areas and efficient delivery of the encapsulated therapeutic agent.

Preparation regimen is more important than patient-related factors: a randomized trial comparing a standard bowel preparation before colonoscopy with an individualized approach.

BACKGROUND: Optimal bowel preparation is one of the most important factors affecting the quality of colonoscopy. Several patient-related factors are known to influence the quality of bowel cleansing but randomized trials in this area are lacking. We aimed to compare an individualized bowel prep strategy based on patient characteristics to a standard preparation regimen. MATERIAL AND METHODS: We conducted an endoscopist-blinded multicenter randomized control-trial. The Boston Bowel Prep Score (BBPS) was used to assess quality of bowel preparation and a 10 point visual analogue scale to assess patient comfort during bowel prep. Patients were randomised to either the standard regimens of split-dose 4L polyethylene-glycol (group A), split-dose sodium picosulphate/magnesium citrate (group B) or to either of the two depending on their responses to a 3-item questionnaire (individualized preparation, group C). RESULTS: 185 patients were randomized during the study period and 143 patients were included in the final analysis. Patients in the individualized group had a median BBPS of 7 compared to a median of 6 in the standard group (p = 0.7). Also, there was no significant difference in patients' comfort scores, irrespective of study group or laxative regimen. However, on multivariable analysis, a split-dose 4L polyethylene-glycol was an independent predictor for achieving a BBPS>6 (OR 3.7, 95% CI 1.4-9.8), regardless of patient-related factors. CONCLUSION: The choice of laxative seems to be more important than patient-related factors in predicting bowel cleansing. Comfort during bowel prep is not influenced by the type of strategy used.