Protocol to generate an in vitro model to study vascular calcification using human endothelial and smooth muscle cells

Vascular calcification is a systemic disease characterized by calcium salt deposition within vascular walls. Here, we present a protocol for establishing an advanced dynamic in vitro co-culture system using endothelial and smooth muscle cells to replicate vascular tissue complexity. We describe steps for cell culture and seeding in a double-flow bioreactor that recreates the action of blood in humans. We then detail the induction of calcification, setting up of the bioreactor, followed by cell viability assessment and calcium quantification

A Different Trend of Heat Shock Proteins 90 mRNA and Protein Inhepatocellular Carcinoma Cell Line-Secreted Extracellular Vesicles

Primary Hepatocellular Carcinoma (HCC) does not usually show any symptoms at the early stage and the use of biomarkers is necessary to aid in diagnosis. Recently Extracellular Vesicles (EVs), submicron membrane- bound structures secreted from different cell types containing a wide variety of bioactive molecules, have increased the attention in many cancers, including HCC, becoming an auspicious candidate as biomarkers and therapy in the scenario of limited diagnostic and treatment option. Many indications have shown that heat shock proteins (Hsps) are important modulators in treatment resistance and invasion of HCC becoming attractive therapeutic targets. In particular, Hsp90α/β isoforms have been found to play critical roles in regulating the proliferation, apoptosis, and metastasis of tumor cells, suggesting for these proteins a role as targets for modern anticancer therapies. The study aimedto verify the presence of Hsp90α/β in EVs secreted by an HCC tumor cell line (HepG2) and by a non-tumorigenic hepatocyte cell line (WRL68), both at protein and mRNA levels, and to analyze their expression variations. The result showed that Hsp90s are transported by the EVs as protein but not at the mRNA level. To build new therapeutic targets using EVs or other organelles as performed on exosomes in recent studies, it is essential to evaluate the action at the pre or post-transcriptional level given their different behavior in transporting proteins or mRNA

Circulatory inflammation molecules and extracellular matrix proteoglycans: local and systemic modulated markers in an atherogenesis model

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Proteomic Modulation in TGF-β-Treated Cholangiocytes Induced by Curcumin Nanoparticles

Curcumin is a natural polyphenol that exhibits a variety of beneficial effects on health, including anti-inflammatory, antioxidant, and hepato-protective properties. Due to its poor water solubility and membrane permeability, in the present study, we prepared and characterized a water-stable, freely dispersible nanoformulation of curcumin. Although the potential of curcumin nanoformulations in the hepatic field has been studied, there are no investigations on their effect in fibrotic pathological conditions involving cholangiocytes. Exploiting an in vitro model of transforming growth factor-  (TGF- )-stimulated cholangiocytes, we applied the Sequential Window Acquisition of All Theoretical Mass Spectra (SWATH-MS)-based quantitative proteomic approaches to study the proteome modulation induced by curcumin nanoformulation. Our results confirmed the well-documented anti-inflammatory properties of this nutraceutic, highlighting the induction of programmed cell death as a mechanism to counteract the cellular damages induced by TGF- . Moreover, curcumin nanoformulation positively influenced the expression of several proteins involved in TGF- -mediated fibrosis. Given the crucial importance of deregulated cholangiocyte functions during cholangiopathies, our results provide the basis for a better understanding of the mechanisms associated with this pathology and could represent a rationale for the development of more targeted therapies

Type-specific inflammatory responses of vascular cells activated by interaction with virgin and aged microplastics

Microplastics (MPs) are recognized as a major environmental problem due to their ubiquitous presence in ecosystems and bioaccumulation in food chains. Not only humans are continuously exposed to these pollutants through ingestion and inhalation, but recent findings suggest they may trigger vascular inflammation and potentially worsen the clinical conditions of cardiovascular patients. Here we combine headspace analysis by needle trap microextraction-gas chromatography-mass spectrometry (HS-NTME-GC-MS) and biological assays to evaluate the effects of polystyrene, high- and low-density polyethylene MPs on phenotype, metabolic activity, and pro-inflammatory status of Vascular Smooth Muscle Cells (VSMCs) the most prominent cells in vascular walls. Virgin and artificially aged MPs (4 weeks at 40 °C and 750 W/m2 simulated solar irradiation) were comparatively tested at 1 mg/mL to simulate a realistic exposure scenario. Our results clearly show the activation of oxidative stress and inflammatory processes when VSMCs were cultured with aged polymers, with significant overexpression of IL-6 and TNF-α. In addition, volatile organic compounds (VOCs), including pentane, acrolein, propanal, and hexanal as the main components, were released by VSMCs into the headspace. Type-specific VOC response profiles were induced on vascular cells from different MPs

A proteomics approach to the study of bleomycin- induced lung fibrosis

Idiopathic pulmonary fibrosis (IPF) is the most severe lung fibrotic form and very few pharmacological therapies are available at present. Key events in the onset of the disease are the activation of fibroblasts to myofibroblasts and the production and release of extracellular matrix (ECM) and molecular factors. Primary murine lung fibroblasts were isolated and their activation induced by Bleomycin (BLM) treatment. Extracellular Vesicles (EV) were isolated and protein extracted. Released soluble proteins (Secretome) and EV-derived proteins were reduced, alkylated and trypsin digested. A nano-LC-MS/MS SWATHTM approach was used for the proteomics analyses. Specific proteins with a putative role in the transition from physiological to fibrotic conditions, such as several matrix metalloproteinases (MMPs), osteopontin (OPN), chitinase-3-like protein1 (CHI3L1) and CD44 resulted differentially released from BLM-treated fibroblasts as compared with untreated lung fibroblasts. Our results provide further understanding of the pathophysiological features of lung fibrosis, and suggest specific target for pharmacological treatments

Inflammatory and antioxidant pattern unbalance in "clopidogrel-resistant" patients during acute coronary syndrome.

Background. In acute coronary syndrome (ACS), inflammation and redox response are associated with increased residual platelet reactivity (RPR) on clopidogrel therapy. We investigated whether clopidogrel interaction affects platelet function and modulates factors related to inflammation and oxidation in ACS patients differently responding to clopidogrel. Material andMethods. Platelet aggregation was measured in 29 ACS patients on dual (aspirin/clopidogrel) antiplatelet therapy. Nonresponders (NR) were defined as RPR ≥70% by ADP. Several inflammatory and redox parameters were assayed and platelet proteome was determined. Results. Eight (28%) out of 29 ACS patients resulted NR to clopidogrel. At 24 hours, the levels of Th2-type cytokines IL-4, IFN, andMCP-1 were higher in NR, while blood GSH (r-GSHbl) levels were lower in NR than responders (R). Proteomic analysis evidenced an upregulated level of platelet adhesion molecule, CD226, and a downregulation of the antioxidant peroxiredoxin-4. In R patients the proinflammatory cytokine IL-6 decreased, while the anti-inflammatory cytokine IL-1Ra increased. Conclusions. In patients with high RPR on clopidogrel therapy, an unbalance of inflammatory factors, platelet adhesion molecules, and circulatory and platelet antioxidantmolecules was observed during the acute phase. Proinflammatory milieu persists in nonresponders for a long time after the acute event while antioxidant blood factors tend to conform to normal responsiveness

The Grizzly, March 25, 1986

The Day U.C. Shut Down • R.A. Selections Underway • Suggestions Being Taken for Forum Topics • Letter: Proud Parents Commend U.C.\u27s Talent • Learn to Swim • New Course for Lifeguards • BreMiller\u27s Diversified Interests • What are You Doing After Graduation? • Matthews an NEH Grant Recipient • Briefs: USGA Represented at Leadership Conference; Ray Bunt Honored at Luncheon; Science Fair in Helfferich; Senior Symposium Topics; Women\u27s Track • Men\u27s Lacrosse Club Starts New Season • Bears Even Record • Lady Bears Look to Win it all • Women\u27s Lacrosse to Begin • Gymnasts Deserted by Coach Morrison • A Reminiscent Backflip • Woody\u27s Hannah a Hit • Equipment Donatedhttps://digitalcommons.ursinus.edu/grizzlynews/1161/thumbnail.jp