Label-Free Optical Sensing and Medical Grade Resins: An Advanced Approach to Investigate Cell–Material Interaction and Biocompatibility

: The Corning Epic® label-free (ELF) system is an innovative technology widely used in drug discovery, immunotherapy, G-protein-associated studies, and biocompatibility tests. Here, we challenge the use of ELF to further investigate the biocompatibility of resins used in manufacturing of blood filters, a category of medical devices representing life-saving therapies for the increasing number of patients with kidney failure. The biocompatibility assays were carried out by developing a cell model aimed at mimicking the clinical use of the blood filters and complementing the existing cytotoxicity assay requested by ISO10993-5. Experiments were performed by putting fibroblasts in both direct contact with two types of selected resins, and indirect contact by means of homemade customized well inserts that were precisely designed and developed for this technology. For both types of contact, fibroblasts were cultured in medium and human plasma. ELF tests confirmed the biocompatibility of both resins, highlighting a statistically significant different biological behavior of a polyaromatic resin compared to control and ion-exchanged resin, when materials were in indirect contact and soaking with plasma. Overall, the ELF test is able to mimic clinical scenarios and represents a promising approach to investigate biocompatibility, showing peculiar biological behaviors and suggesting the activation of specific intracellular pathways

Novel bioprinted 3D model to human fibrosis investigation

Fibrosis is shared in multiple diseases with progressive tissue stiffening, organ failure and limited therapeutic options. This unmet need is also due to the lack of adequate pre-clinical models to mimic fibrosis and to be challenged novel by anti-fibrotic therapeutic venues. Here using bioprinting, we designed a novel 3D model where normal human healthy fibroblasts have been encapsulated in type I collagen. After stimulation by Transforming Growth factor beta (TGFβ), embedded cells differentiated into myofibroblasts and enhanced the contractile activity, as confirmed by the high level of α − smooth muscle actin (αSMA) and F-actin expression. As functional assays, SEM analysis revealed that after TGFβ stimulus the 3D microarchitecture of the scaffold was dramatically remolded with an increased fibronectin deposition with an abnormal collagen fibrillar pattern. Picrius Sirius Red staining additionally revealed that TGFβ stimulation enhanced of two logarithm the collagen fibrils neoformation in comparison with control. These data indicate that by bioprinting technology, it is possible to generate a reproducible and functional 3D platform to mimic fibrosis as key tool for drug discovery and impacting on animal experimentation and reducing costs and time in addressing fibrosis

Microfragmented adipose tissue is associated with improved ex vivo performance linked to HOXB7 and b-FGF expression

Introduction: Adipose tissue (AT) has become a source of mesenchymal stromal/stem cells (MSC) for regenerative medicine applications, in particular skeletal disorders. Several enzymatic or mechanical procedures have been proposed to process AT with the aim to isolate cells that can be locally implanted. How AT is processed may impact its properties. Thus, we compared AT processed by centrifugation (C-AT) to microfragmentation (MF-AT). Focusing on MF-AT, we subsequently assessed the impact of synovial fluid (SF) alone on both MF-AT and isolated AT-MSC to better understand their cartilage repair mechanisms. Materials and methods: MF-AT and C-AT from the same donors were compared by histology and qRT-PCR immediately after isolation or as ex vivo cultures using a micro-tissue pellet system. The in vitro impact of SF on MF-AT and AT-MSC was assessed by histological staining and molecular analysis. Results: The main AT histological features (i.e., increased extracellular matrix and cellularity) of the freshly isolated or ex vivo-cultured MF-AT persisted compared to C-AT, which rapidly deteriorated during culture. Based on our previous studies of HOX genes in MSC, we investigated the involvement of Homeobox Protein HOX-B7 (HOXB7) and its target basic Fibroblast Growth Factor (bFGF) in the molecular mechanism underlying the improved performance of MF-AT. Indeed, both these biomarkers were more prominent in freshly isolated MF-AT compared to C-AT. SF alone preserved the AT histological features of MF-AT, together with HOXB7 and bFGF expression. Increased cell performance was also observed in isolated AT-MSC after SF treatment concomitant with enhanced HOXB7 expression, although there was no apparent association with bFGF. Conclusions: Our findings show that MF has a positive effect on the maintenance of AT histology and may trigger the expression of trophic factors that improve tissue repair by processed AT

Autologous Marrow Mesenchymal Stem Cell Driving Bone Regeneration in a Rabbit Model of Femoral Head Osteonecrosis

Osteonecrosis of the femoral head (ONFH) is a progressive degenerative disease that ultimately requires a total hip replacement. Mesenchymal stromal/stem cells (MSCs), particularly the ones isolated from bone marrow (BM), could be promising tools to restore bone tissue in ONFH. Here, we established a rabbit model to mimic the pathogenic features of human ONFH and to challenge an autologous MSC-based treatment. ON has been originally induced by the synergic combination of surgery and steroid administration. Autologous BM-MSCs were then implanted in the FH, aiming to restore the damaged tissue. Histological analyses confirmed bone formation in the BM-MSC treated rabbit femurs but not in the controls. In addition, the model also allowed investigations on BM-MSCs isolated before (ON-BM-MSCs) and after (ON+BM-MSCs) ON induction to dissect the impact of ON damage on MSC behavior in an affected microenvironment, accounting for those clinical approaches foreseeing MSCs generally isolated from affected patients. BM-MSCs, isolated before and after ON induction, revealed similar growth rates, immunophenotypic profiles, and differentiation abilities regardless of the ON. Our data support the use of ON+BM-MSCs as a promising autologous therapeutic tool to treat ON, paving the way for a more consolidated use into the clinical settings

Human Adipose Mesenchymal Stromal/Stem Cells Improve Fat Transplantation Performance

The resorption rate of autologous fat transfer (AFT) is 40–60% of the implanted tissue, requiring new surgical strategies for tissue reconstruction. We previously demonstrated in a rabbit model that AFT may be empowered by adipose-derived mesenchymal stromal/stem cells (AD-MSCs), which improve graft persistence by exerting proangiogenic/anti-inflammatory effects. However, their fate after implantation requires more investigation. We report a xenograft model of adipose tissue engineering in which NOD/SCID mice underwent AFT with/without human autologous AD-MSCs and were monitored for 180 days (d). The effect of AD-MSCs on AFT grafting was also monitored by evaluating the expression of CD31 and F4/80 markers. Green fluorescent protein-positive AD-MSCs (AD-MSC-GFP) were detected in fibroblastoid cells 7 days after transplantation and in mature adipocytes at 60 days, indicating both persistence and differentiation of the implanted cells. This evidence also correlated with the persistence of a higher graft weight in AFT-AD-MSC compared to AFT alone treated mice. An observation up to 180 d revealed a lower resorption rate and reduced lipidic cyst formation in the AFT-AD-MSC group, suggesting a long-term action of AD-MSCs in support of AFT performance and an anti-inflammatory/proangiogenic activity. Together, these data indicate the protective role of adipose progenitors in autologous AFT tissue resorption

Parco archeologico del Piano della Civita e allestimento del Museo Archeologico Roger Lambrechts in Artena

Dieci anni di ricerche storiche, scavi archeologici, restauri paesaggistici ed edilizi, ed infine di progetti e realizzazioni nel territorio del comune di Artena sono qui raccontati dai loro protagonisti. La pubblicazione si suddivide in due parti: nella prima parte viene messa in luce la figura dell’archeologo belga prof. Roger Lambrechts, attraverso i ricordi dei suoi collaboratori. La città storica preromana sul Piano della Civita e la città seicentesca di Scipione Borghese vengono delineati in due distinti saggi. Nella seconda parte vengono illustrate la realizzazione del Parco Archeologico del Piano della Civita, il restauro del Granaio Borghese ed il nuovo Museo Archeologico intitolato a Roger Lambrechts. Un saggio di Andrea Bruschi affronta il tema cruciale degli interventi di trasformazione delle preesistenze storiche anche alla luce delle molteplici esperienze realizzate su questo stesso tema dagli architetti Di Noto e Montuori

GRUPPO MONTUORI

La scheda illustra il progetto redatto come gruppo di studio dell'Università degli Studi Roma Tre, Facoltà di architettura, nell'ambito della ricerca Nazionale PRIN sugli archivi di architettur

Controlled release of vancomycin from freeze-dried chitosan salts coated with different fatty acids by spray-drying

The aim of this study was to describe a controlled drug release system based on chitosan salts for vancomycin hydrochloride delivery. Chitosan aspartate (CH-Asp), chitosan glutamate (CH-Glu) and chitosan hydrochloride (CH-HCl) were prepared by freeze-drying and coated with stearic, palmitic, myristic and lauric acids by spray-drying technique. Vancomycin hydrochloride was used as a peptidic model drug whose sustained release should minimize its inactivation in the upper part of the gastrointestinal tract. This study evaluated, in vitro, the influence of chitosan salts on the release behaviour of vancomycin hydrochloride from the freeze-dried and spray-dried systems at pH 2.0 and 7.4

SOLUBLE TRAIL-ARMED HUMAN AD-MSC AS NOVEL CELL THERAPY APPROACH FOR PANCREATIC DUCTAL ADENOCARCINOMA

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive adult tumors and its prognosis is still poor since the number of deaths almost equal the number of new cases. New therapeutic approaches are therefore urgently needed. In our model human adipose mesenchy.mal stromallstem cells (ADMSC) have been armed with a novel soluble TRAIL variant that is constantly released by modified progenitors (sTRAIL AD-MSC). The wild type TRAIL form is known to act as a trimer stabilized by a zinc-binding site. In this srudy, by gene engineering, we allow AD-MSC to secrete a trimeric zincindependent soluble TRAIL variant. The molecule has been then challenged in vitro and in vivo, either using sTRAIL AD-MSC supernatant or injecting sTRAIL AD-MSC cells in a PDAC xenotransplant rnodel, We demonstrated that sTRAIL was stable at 37°C far at least 24 hours and was able to induce apoptosis in the PDAC lines BxPC-3 and MIA PaCa-2 and, more interestingly, against primary PDAC cells. Moreover, sTRAIL released by AD-MSC was able to significantly counteract tumor growth with a reduction of the cytokeratin-7 positive cells and by an anti-angiogenic effect. In parallel, a retrospective study on PDAC specimens form patients (n = 19) has been conducted in order to investigate TRAIL DR4, DR5 and OPG receptor expression in "real" PDAC tissue and generate insights on the possible clinical translation of our approach. Our results suggest that MSC can be vehicles for novel TRAIL variants opening novel opportunities for PDAC treatrnent by multiple mechanisms