The Potential of the Fibronectin Inhibitor Arg-Gly-Asp-Ser in the Development of Therapies for Glioblastoma.

Glioblastoma (GBM) is the most lethal and common malignant primary brain tumor in adults. An important feature that supports GBM aggressiveness is the unique composition of its extracellular matrix (ECM). Particularly, fibronectin plays an important role in cancer cell adhesion, differentiation, proliferation, and chemoresistance. Thus, herein, a hydrogel with mechanical properties compatible with the brain and the ability to disrupt the dynamic and reciprocal interaction between fibronectin and tumor cells was produced. High-molecular-weight hyaluronic acid (HMW-HA) functionalized with the inhibitory fibronectin peptide Arg-Gly-Asp-Ser (RGDS) was used to produce the polymeric matrix. Liposomes encapsulating doxorubicin (DOX) were also included in the hydrogel to kill GBM cells. The resulting hydrogel containing liposomes with therapeutic DOX concentrations presented rheological properties like a healthy brain. In vitro assays demonstrated that unmodified HMW-HA hydrogels only caused GBM cell killing after DOX incorporation. Conversely, RGDS-functionalized hydrogels displayed per se cytotoxicity. As GBM cells produce several proteolytic enzymes capable of disrupting the peptide-HA bond, we selected MMP-2 to illustrate this phenomenon. Therefore, RGDS internalization can induce GBM cell apoptosis. Importantly, RGDS-functionalized hydrogel incorporating DOX efficiently damaged GBM cells without affecting astrocyte viability, proving its safety. Overall, the results demonstrate the potential of the RGDS-functionalized hydrogel to develop safe and effective GBM treatments.

Decellularized kidney extracellular matrix-based hydrogels for renal tissue engineering.

Kidney regeneration is hindered by the limited pool of intrinsic reparative cells. Advanced therapies targeting renal regeneration have the potential to alleviate the clinical and financial burdens associated with kidney disease. Delivery systems for cells, extracellular vesicles, or growth factors aimed at enhancing regeneration can benefit from vehicles enabling targeted delivery and controlled release. Hydrogels, optimized to carry biological cargo while promoting regeneration, have emerged as promising candidates for this purpose. This study aims to develop a hydrogel from decellularized kidney extracellular matrix (DKECM) and explore its biocompatibility as a biomaterial for renal regeneration. The resulting hydrogel crosslinks with temperature and exhibits a high concentration of extracellular matrix. The decellularization process efficiently removes detergent residues, yielding a pathogen-free biomaterial that is non-hemolytic and devoid of α-gal epitope. Upon interaction with macrophages, the hydrogel induces differentiation into both pro-inflammatory and anti-inflammatory phenotypes, suggesting an adequate balance to promote biomaterial functionality in vivo. Renal progenitor cells encapsulated in the DKECM hydrogel demonstrate higher viability and proliferation than in commercial collagen-I hydrogels, while also expressing tubular cells and podocyte markers in long-term culture. Overall, the injectable biomaterial derived from porcine DKECM is anticipated to elicit minimal host reaction while fostering progenitor cell bioactivity, offering a potential avenue for enhancing renal regeneration in clinical settings. STATEMENT OF SIGNIFICANCE: The quest to improve treatments for kidney disease is crucial, given the challenges faced by patients on dialysis or waiting for transplants. Exciting new therapies combining biomaterials with cells can revolutionize kidney repair. In this study, researchers created a hydrogel from pig kidney. This gel could be used to deliver cells and other substances that help in kidney regeneration. Despite coming from pigs, it's safe for use in humans, with no harmful substances and reduced risk of immune reactions. Importantly, it promotes a balanced healing response in the body. This research not only advances our knowledge of kidney repair but also offers hope for more effective treatments for kidney diseases.

COX-2 inhibitor delivery system aiming intestinal inflammatory disorders.

Selective COX-2 inhibitors such as etoricoxib (ETX) are potentially indicated for the treatment of intestinal inflammatory disorders. However, their systemic administration provokes some off-site secondary effects, decreasing the desirable local effectiveness. To circumvent such limitations, herein an ETX delivery system based on electrospun fibrous meshes (eFMs) was proposed. ETX at different concentrations (1, 2, and 3 mg mL-1) was loaded into eFMs, which not affect the morphology and the mechanical properties of this drug delivery system (DDS). The ETX showed a burst release within the first 12 h, followed by a faster release until 36 h, gradually decreasing over time. Importantly, the ETX studied concentrations were not toxic to human colonic cells (i.e. epithelial and fibroblast). Moreover, the DDS loading the highest concentration of ETX, when tested with stimulated human macrophages, promoted a reduction of PGE2, IL-8 and TNF-α secretion. Therefore, the proposed DDS may constitute a safe and efficient treatment of colorectal diseases promoted by inflammatory disorders associated with COX-2.

Diaphragmatic hernia repair porcine model to compare the performance of biodegradable membranes against Gore-Tex®.

BACKGROUND: Patch repair of congenital diaphragmatic hernia (CDH) using Gore-Tex® is associated with infection, adhesions, hernia recurrence, long-term musculoskeletal sequels and poor tissue regeneration. To overcome these limitations, the performance of two novel biodegradable membranes was tested to repair CDH in a growing pig model. METHODS: Twelve male pigs were randomly assigned to 3 different groups of 4 animals each, determined by the type of patch used during thoracoscopic diaphragmatic hernia repair (Gore-Tex®, polycaprolactone electrospun membrane-PCLem, and decellularized human chorion membrane-dHCM). After 7 weeks, all animals were euthanized, followed by necropsy for diaphragmatic evaluation and histological analysis. RESULTS: Thoracoscopic defect creation and diaphragmatic repair were performed without any technical difficulty in all groups. However, hernia recurrence rate was 0% in Gore-Tex®, 50% in PCLem and 100% in dHCM groups. At euthanasia, Gore-Tex® patches appeared virtually unchanged and covered with a fibrotic capsule, while PCLem and dHCM patches were replaced by either floppy connective tissue or vascularized and floppy regenerated membranous tissue, respectively. CONCLUSION: Gore-Tex® was associated with a higher survival rate and lower recurrence. Nevertheless, the proposed biodegradable membranes were associated with better tissue integration when compared with Gore-Tex®.

Severe Hypertriglyceridemia: A 10-Year Review in a Portuguese Hospital.

INTRODUCTION: Severe hypertriglyceridemia (SHTG) is a rare condition associated with serious complications, such as acute pancreatitis (AP), and the best treatment is still a matter of discussion. The aim of this study is to outline the demographics, management, and outcomes (recurrence and mortality) of complications in patients with SHTG. MATERIAL AND METHODS: A retrospective, observational, and analytical study was carried out by obtaining clinical data from the electronic health records of patients with SHTG admitted to the Internal and Intensive Medicine units from the 1st of January 2009 to the 31st of December 2020 in a university hospital. RESULTS: The cohort included 17 patients. The most common complication was AP (13/17 = 76.5%). Admission to the intensive care unit (ICU) was observed in 84.2%. Among patients with AP, the most commonly administered therapies were insulin (82.4%) and fibrates (76.5%). Plasmapheresis was used in 58.8%, and the criteria for using this technique were mainly based on clinical and laboratory abnormalities. There were no deaths. The readmission rate at 30 days was 36.3%. CONCLUSION: This study shows the morbidity profile associated with SHTG, with a high level of ICU admissions and also a high level of the use of plasmapheresis. In our population, this approach had good results, and this should be highlighted as there are no clear international guidelines for this intervention. Distinguishing between patients with familial chylomicronemia syndrome or with multifactorial chylomicronemia is important as recent specific therapy for lipoprotein lipase (LPL) genetic deficit is available. In the near future, the performance of a genetic study should be considered in patients with SHTG as an attempt to avoid the high recurrence rate of complications of this disease.

Antioxidant and Anti-Inflammatory Activities of Stellera chamaejasme L. Roots and Aerial Parts Extracts.

Natural products, mainly plants, have a crucial role in folk medicine. Particularly, Stellera chamaejasme L. has been traditionally used in Mongolian medicine to treat various diseases, including chronic tracheitis, tuberculosis, and psoriasis. In this study, ethanol (EtOH) and dichloromethane (DCM) extracts of its roots (R) and aerial parts (AP) were evaluated for their antioxidant and anti-inflammatory activities. Thin-layer chromatography demonstrated the presence of flavonoids, namely kaempferol and quercetin-3-O-glucopyranoside, only in the EtOH-AP. Conversely, it showed that kaempferol, quercetin-3-O-glucopyranoside, coumarin, luteolin, rutin, morin, and riboflavin were not present in the other three extracts. The S. chamaejasme extracts exhibited strong antioxidant activity. In addition, the roots extracts presented the highest antioxidant activity against peroxyl radicals, with the EtOH-R being the most potent (IC50 = 0.90 ± 0.07 µg/mL). S. chamaejasme extracts also efficiently inhibited the production of one of the main pro-inflammatory cytokines, interleukin (IL)-6, in a dose-dependent manner by lipopolysaccharide-stimulated macrophages. Particularly, DCM-R was the strongest extract, reducing ≈ 91.5% of the IL-6 production. Since this extract was the most effective, gas chromatography-mass spectrometry (GC-MS) analyses were performed and demonstrated the presence of two fatty acids (palmitic acid and 9-octadecenoic acid), one fatty alcohol (1-hexadecanol), and one triterpenoid (squalene) that can contribute to the observed bioactivity. Herewith, S. chamaejasme extracts, mainly DCM-R, exhibit antioxidant and anti-inflammatory activities that could be applied as new and innovative natural formulations for the treatment of chronic inflammatory diseases.

Functional recovery of injured cavernous nerves achieved through endogenous nerve growth factor-containing bioactive fibrous membrane.

Radical prostatectomy is a highly successful treatment for prostate cancer, among the most prevalent manifestations of the illness. Damage of the cavernous nerve (CN) during prostatectomy is the main cause of postoperative erectile dysfunction (ED). In this study, the capability of a personalized bioactive fibrous membrane to regenerate injured CN was investigated. The fibrous membrane bioactivity is conferred by the selectively bound nerve growth factor (NGF) present in the rat urine. In a rat model of bilateral CN crush, the implanted bioactive fibrous membrane induces CN regeneration and restoration of erectile function, showing a significantly increased number of smooth muscle cells and content of endothelial and neuronal nitric oxide synthases (eNOS; nNOS). In addition, the bioactive fibrous membrane promotes nerve regeneration by increasing the number of myelinated axons and nNOS-positive cells, therefore reversing the CN fibrosis found in untreated rats or rats treated with a bare fibrous membrane. Therefore, this personalized regenerative strategy could overcome the recognized drawbacks of currently available treatments for CN injuries. It may constitute an effective treatment for prostate cancer patients suffering from ED after being subject to radical prostatectomy. STATEMENT OF SIGNIFICANCE: The present work introduces a unique strategy to address post-surgical ED resulting from CN injury during pelvic surgery (e.g., radical prostatectomy, radical cystoprostatectomy, abdominoperineal resection). It comprises a bioactive and cell-free fibrous implant, customized to enhance CN recovery. Pre-clinical results in a rat model of bilateral CN crush demonstrated that the bioactive fibrous implant can effectively heal injured CN, and restore penile structure and function. This implant selectively binds NGF from patient fluids (i.e. urine) due to its functionalized surface and high surface area. Moreover, its local implantation reduces adverse side effects. This tailored regenerative approach has the potential to revolutionize the treatment of ED in prostate cancer patients following radical prostatectomy, overcoming current treatment limitations.

Microfluidic Devices: A Tool for Nanoparticle Synthesis and Performance Evaluation.

The use of nanoparticles (NPs) in nanomedicine holds great promise for the treatment of diseases for which conventional therapies present serious limitations. Additionally, NPs can drastically improve early diagnosis and follow-up of many disorders. However, to harness their full capabilities, they must be precisely designed, produced, and tested in relevant models. Microfluidic systems can simulate dynamic fluid flows, gradients, specific microenvironments, and multiorgan complexes, providing an efficient and cost-effective approach for both NPs synthesis and screening. Microfluidic technologies allow for the synthesis of NPs under controlled conditions, enhancing batch-to-batch reproducibility. Moreover, due to the versatility of microfluidic devices, it is possible to generate and customize endless platforms for rapid and efficient in vitro and in vivo screening of NPs' performance. Indeed, microfluidic devices show great potential as advanced systems for small organism manipulation and immobilization. In this review, first we summarize the major microfluidic platforms that allow for controlled NPs synthesis. Next, we will discuss the most innovative microfluidic platforms that enable mimicking in vitro environments as well as give insights into organism-on-a-chip and their promising application for NPs screening. We conclude this review with a critical assessment of the current challenges and possible future directions of microfluidic systems in NPs synthesis and screening to impact the field of nanomedicine.

Combination Therapy With CD147-Targeted Nanoparticles Carrying Phenformin Decreases Lung Cancer Growth.

Lung cancer is one of the most fatal cancers worldwide. Resistance to conventional therapies remains a hindrance to patient treatment. Therefore, the development of more effective anti-cancer therapeutic strategies is imperative. Solid tumors exhibit a hyperglycolytic phenotype, leading to enhanced lactate production; and, consequently, its extrusion to the tumor microenvironment. Previous data reveals that inhibition of CD147, the chaperone of lactate transporters (MCTs), decreases lactate export in lung cancer cells and sensitizes them to phenformin, leading to a drastic decrease in cell growth. In this study, the development of anti-CD147 targeted liposomes (LUVs) carrying phenformin is envisioned, and their efficacy is evaluated to eliminate lung cancer cells. Herein, the therapeutic effect of free phenformin and anti-CD147 antibody, as well as the efficacy of anti-CD147 LUVs carrying phenformin on A549, H292, and PC-9 cell growth, metabolism, and invasion, are evaluated. Data reveals that phenformin decreases 2D and 3D-cancer cell growth and that the anti-CD147 antibody reduces cell invasion. Importantly, anti-CD147 LUVs carrying phenformin are internalized by cancer cells and impaired lung cancer cell growth in vitro and in vivo. Overall, these results provide evidence for the effectiveness of anti-CD147 LUVs carrying phenformin in compromising lung cancer cell aggressiveness.

Synthesis and Anti-Inflammatory Evaluation of a Library of Chiral Derivatives of Xanthones Conjugated with Proteinogenic Amino Acids.

In recent decades, the relationship between drug chirality and biological activity has been assuming enormous importance in medicinal chemistry. Particularly, chiral derivatives of xanthones (CDXs) have interesting biological activities, including enantioselective anti-inflammatory activity. Herein, the synthesis of a library of CDXs is described, by coupling a carboxyxanthone (1) with both enantiomers of proteinogenic amino esters as chiral building blocks (2-31), following the chiral pool strategy. The coupling reactions were performed at room temperature with good yields (from 44 to 99.9%) and very high enantiomeric purity, with most of them presenting an enantiomeric ratio close to 100%. To afford the respective amino acid derivatives (32-61), the ester group of the CDXs was hydrolyzed in mild alkaline conditions. Consequently, in this work, sixty new derivatives of CDXs were synthetized. The cytocompatibility and anti-inflammatory activity in the presence of M1 macrophages were studied for forty-four of the new synthesized CDXs. A significant decrease in the levels of a proinflammatory cytokine targeted in the treatment of several inflammatory diseases, namely interleukin 6 (IL-6), was achieved in the presence of many CDXs. The amino ester of L-tyrosine (X1AELT) was the most effective in reducing IL-6 production (52.2 ± 13.2%) by LPS-stimulated macrophages. Moreover, it was ≈1.2 times better than the D-enantiomer. Indeed, enantioselectivity was observed for the majority of the tested compounds. Thus, their evaluation as promising anti-inflammatory drugs should be considered.

Size-Dependent Polymeric Nanoparticle Distribution in a Static versus Dynamic Microfluidic Blood Vessel Model: Implications for Nanoparticle-Based Drug Delivery.

Nanoparticles (NPs) have been widely investigated in the nanomedicine field. One of the main challenges is to accurately predict the NP distribution and fate after administration. Microfluidic platforms acquired huge importance as tools to model the in vivo environment. In this study, we leveraged a microfluidic platform to produce FITC-labeled poly(lactide-co-glycolide)-block-poly(ethylene glycol) (PLGA-PEG) NPs with defined sizes of 30, 50, and 70 nm. The study aimed to compare the ability of NPs with differences of 20 nm in size to cross an endothelial barrier using static (Transwell inserts) and dynamic (microfluidic perfusion device) in vitro models. Our results evidence a size-dependent NP crossing in both models (30 > 50 > 70 nm) and highlight the bias deriving from the static model, which does not involve shear stresses. The permeation of each NP size was significantly higher in the static system than in the dynamic model at the earliest stages. However, it gradually decreased to levels comparable with those of the dynamic model. Overall, this work highlights clear differences in NP distribution over time in static versus dynamic conditions and distinct size-dependent patterns. These findings reinforce the need for accurate in vitro screening models that allow for more accurate predictions of in vivo performance.

The Influence of Feeder Cell-Derived Extracellular Matrix Density on Thymic Epithelial Cell Culture.

The thymus is responsible for the selection and development of T cells, having an essential role in the establishment of adaptive immunity. Thymic epithelial cells (TECs) are key players in T cell development interacting with thymocytes in the thymic 3D environment. Feeder-layer cells have been frequently used as platforms for the successful establishment of TEC cultures. Nevertheless, the role of the feeder cell-derived extracellular matrix (ECM) on TEC cultures was not previously reported. Therefore, this work aimed at assessing the effect of the ECM produced by feeder cells cultured at two different densities on the establishment of TEC culture. Due to the high surface area and porosity, electrospun fibrous meshes were used to support ECM deposition. The feeder cell-derived ECM was efficiently recovered after decellularization, maintaining the composition of major proteins. All the decellularized matrices were permeable and showed an increase in surface mechanical properties after decellularization. TEC cultures confirmed that the ECM density impacts cellular performance, with higher densities showing a decreased cellular activity. Our findings provide evidence that feeder cell-derived ECM is a suitable substrate for TEC culture and can potentially be applied in thymus bioengineering.

On the size-dependent internalization of sub-hundred polymeric nanoparticles.

The understanding of the interaction between nanoparticles (NPs) and cells is crucial to design nanocarriers with high therapeutic relevance. In this study, we exploited a microfluidics device to synthesize homogeneous suspensions of NPs with ≈ 30, 50, and 70 nm of size. Afterward, we investigated their level and mechanism of internalization when exposed to different types of cells (endothelial cells, macrophages, and fibroblasts). Our results show that all NPs were cytocompatible and internalized by the different cell types. However, NPs uptake was size-dependent, being the maximum uptake efficiency observed for the 30 nm NPs. Moreover, we demonstrate that size can lead to distinct interactions with different cells. For instance, 30 nm NPs were internalized with an increasing trend over time by endothelial cells, while a steady and a decreasing trend were observed when incubated with LPS-stimulated macrophages and fibroblasts, respectively. Finally, the use of different chemical inhibitors (chlorpromazine, cytochalasin-D, and nystatin), and low temperature (4 °C) indicated that phagocytosis/micropinocytosis are the main internalization mechanism for all NPs sizes. However, different endocytic pathways were initiated in the presence of particular NP sizes. In endothelial cells, for example, caveolin-mediated endocytosis occurs primarily in the presence of 50 nm NPs, whereas clathrin-mediated endocytosis substantially promotes the internalization of 70 nm NPs. This evidence demonstrates the importance of size in the NPs design for mediating interaction with specific cell types.

Development of bilayered porous silk scaffolds for thymus bioengineering.

The thymus coordinates the development and selection of T cells. It is structured into two main compartments: the cortex and the medulla. The replication of such complex 3D environment has been challenged by bioengineering approaches. Nevertheless, the effect of the scaffold microstructure on thymic epithelial cell (TEC) cultures has not been deeply investigated. Here, we developed bilayered porous silk fibroin scaffolds and tested their effect on TEC co-cultures. The small and large pore scaffolds presented a mean pore size of 84.33 ± 21.51 µm and 194.90 ± 61.38 µm, respectively. The highly porous bilayered scaffolds presented a high water absorption and water content (> 94 %), together with mechanical properties in the range of the native tissue. TEC (i.e., medullary (mTEC) and cortical (cTEC) cell lines) proliferation is increased in scaffolds with larger pores. The co-culture of both TEC lines in the bilayered porous silk scaffolds presents enhanced cell proliferation and metabolic activity when compared with mTEC in single culture. Also, when the co-culture occurred with cTEC in the small pores layer and mTEC in the large pores layer, a 9.2- and 18.9-fold increase in Foxn1 and Icam1 gene expression in cTEC is evident. These results suggest that scaffold microstructure and the co-culture influence TEC's behaviour. Bilayered silk scaffolds with adjusted microstructure are a valid alternative for TEC culture, having possible applications in advanced thymus bioengineering strategies.

Echinacea purpurea Fractions Represent Promising Plant-Based Anti-Inflammatory Formulations.

Echinacea purpurea is traditionally used in the treatment of inflammatory diseases. Therefore, we investigated the anti-inflammatory capacity of E. purpurea dichloromethanolic (DE) and ethanolic extracts obtained from flowers and roots (R). To identify the class of compounds responsible for the strongest bioactivity, the extracts were fractionated into phenol/carboxylic acid (F1) and alkylamide fraction (F2). The chemical fingerprint of bioactive compounds in the fractions was evaluated by LC-HRMS. E. purpurea extracts and fractions significantly reduced pro-inflammatory cytokines (interleukin 6 and/or tumor necrosis factor) and reactive oxygen and nitrogen species (ROS/RNS) production by lipopolysaccharide-stimulated primary human monocyte-derived macrophages. Dichloromethanolic extract obtained from roots (DE-R) demonstrated the strongest anti-inflammatory activity. Moreover, fractions exhibited greater anti-inflammatory activity than whole extract. Indeed, alkylamides must be the main compounds responsible for the anti-inflammatory activity of extracts; thus, the fractions presenting high content of these compounds presented greater bioactivity. It was demonstrated that alkylamides exert their anti-inflammatory activity through the downregulation of the phosphorylation of p38, ERK 1/2, STAT 3, and/or NF-κB signaling pathways, and/or downregulation of cyclooxygenase 2 expression. E. purpurea extracts and fractions, mainly DE-R-F2, are promising and powerful plant-based anti-inflammatory formulations that can be further used as a basis for the treatment of inflammatory diseases.

Intracellular Trafficking of Size-Tuned Nanoparticles for Drug Delivery.

Polymeric nanoparticles (NPs) are widely used as drug delivery systems in nanomedicine. Despite their widespread application, a comprehensive understanding of their intracellular trafficking remains elusive. In the present study, we focused on exploring the impact of a 20 nm difference in size on NP performance, including drug delivery capabilities and intracellular trafficking. For that, poly(ethylene glycol) methyl ether-block-poly(lactide-co-glycolide) (PLGA-PEG) NPs with sizes of 50 and 70 nm were precisely tailored. To assess their prowess in encapsulating and releasing therapeutic agents, we have employed doxorubicin (Dox), a well-established anticancer drug widely utilized in clinical settings, as a model drug. Then, the beneficial effect of the developed nanoformulations was evaluated in breast cancer cells. Finally, we performed a semiquantitative analysis of both NPs' uptake and intracellular localization by immunostaining lysosomes, early endosomes, and recycling endosomes. The results show that the smaller NPs (50 nm) were able to reduce the metabolic activity of cancer cells more efficiently than NPs of 70 nm, in a time and concentration-dependent manner. These findings are corroborated by intracellular trafficking studies that reveal an earlier and higher uptake of NPs, with 50 nm compared to the 70 nm ones, by the breast cancer cells. Consequently, this study demonstrates that NP size, even in small increments, has an important impact on their therapeutic effect.

Potential roles of hyaluronic acid in in vivo CAR T cell reprogramming for cancer immunotherapy.

Chimeric antigen receptor (CAR) T cell therapy has recently shown unprecedented clinical efficacy for cancer treatment, particularly of hematological malignancies. However, the complex manufacturing processes that involve ex vivo genetic modification of autologous T cells limits its therapeutic application. CAR T cells generated in vivo provide a valid alternative immunotherapy, "off-the-shelf", for cancer treatment. This approach requires carriers for the delivery of CAR-encoding constructs, which are plasmid DNA or messenger RNA, to T cells for CAR expression to help eradicate the tumor. As such, there are a growing number of studies reporting gene delivery systems for in vivo CAR T cell therapy based on viral vectors and polymeric nanoparticles. Hyaluronic acid (HA) is a natural biopolymer that can serve for gene delivery, because of its inherent properties of cell recognition and internalization, as well as its biodegradability, biocompatibility, and presence of functional groups for the chemical conjugation of targeting ligands. In this review, the potential of HA in the delivery of CAR constructs is discussed on the basis of previous experience of HA-based nanoparticles for gene therapy. Furthermore, current studies on CAR carriers for in vivo-generated CAR T cells are included, giving an idea of a rational design of HA-based systems for the more efficient delivery of CAR to circulating T cells.

On the Bioactivity of Echinacea purpurea Extracts to Modulate the Production of Inflammatory Mediators.

Inflammatory diseases are the focus of several clinical studies, due to limitations and serious side effects of available therapies. Plant-based drugs (e.g., salicylic acid, morphine) have become landmarks in the pharmaceutical field. Therefore, we investigated the immunomodulatory effects of flowers, leaves, and roots from Echinacea purpurea. Ethanolic (EE) and dichloromethanolic extracts (DE) were obtained using the Accelerated Solvent Extractor and aqueous extracts (AE) were prepared under stirring. Their chemical fingerprint was evaluated by liquid chromatography-high resolution mass spectrometry (LC-HRMS). The pro- and anti-inflammatory effects, as well as the reduction in intracellular reactive oxygen and nitrogen species (ROS/RNS), of the different extracts were evaluated using non-stimulated and lipopolysaccharide-stimulated macrophages. Interestingly, AE were able to stimulate macrophages to produce pro-inflammatory cytokines (tumor necrosis factor -TNF-α, interleukin -IL-1ß, and IL-6), and to generate ROS/RNS. Conversely, under an inflammatory scenario, all extracts reduced the amount of pro-inflammatory mediators. DE, alkylamides-enriched extracts, showed the strongest anti-inflammatory activity. Moreover, E. purpurea extracts demonstrated generally a more robust anti-inflammatory activity than clinically used anti-inflammatory drugs (dexamethasone, diclofenac, salicylic acid, and celecoxib). Therefore, E. purpurea extracts may be used to develop new effective therapeutic formulations for disorders in which the immune system is either overactive or impaired.

Laminin-2 immobilized on a 3D fibrous structure impacts cortical thymic epithelial cells behaviour and their interaction with thymocytes.

The tri-dimensionality of the thymic extracellular matrix (ECM) supports the crosstalk between thymocytes and thymic epithelial cells (TECs). The thymic ECM component laminin-2 is involved in the regulation of thymocytes and their interaction with cortical TECs (cTECs). Most in vitro studies use planar surfaces to study the interaction between ECM components and thymic cells. Herein, we developed a novel biofunctionalized culture system by immobilizing laminin-2 at the surface of porous and fibrous electrospun meshes. We aimed to study the interaction of cTECs with thymocytes in the presence of laminin-2 presented through this system. The results indicated that the presence of laminin-2, not its density, has a positive effect on the cell viability and proliferation of cTECs. qPCR results demonstrated that laminin-2 density influenced the expression of cTECs genes. An increased percentage of adherent CD4-CD8- thymocytes and a decreased percentage of CD4+CD8+ thymocytes were evident in higher laminin-2 concentrations. Higher concentrations decreased the expression of Il7 and Ccl25 in cTECs after thymocyte adhesion. Altogether, these results indicate that the interaction of thymocytes with the thymic cortical compartment is affected by laminin-2 density and supports the need for immobilized ECM proteins in porous and fibrous substrates for the study of thymus biology.