A non-invasive, sensor-based approach to exploit the autofluorescence of saffron (Crocus sativus L.) for on-site evaluation of aging.

So far, compliance with ISO 3632 standard specifications for top-quality saffron guarantees good agricultural and post-harvest production practices. Tracking early-stage oxidation remains challenging. Our study aims to address this issue by exploring the visible, fluorescence, and near-infrared spectra of category I saffron. Using a multi-spectral sensor, we tested fresh and artificially aged saffron in powder form. High autofluorescence intensities at 600-700 nm allowed calibration for the 'content of aged saffron'. Samples with minimum coloring strength (200-220 units) were classified as 70% aged, while those exceeding maximum aroma strength (50 units) as 100% aged. Consistent patterns across origin, age, and processing history indicated potential for objectively assessing early-oxidation markers. Further analyses uncovered multiple contributing fluorophores, including cis-apocarotenoids, correlated with FTIR-based aging markers. Our findings underscore that sensing autofluorescence of traded saffron presents an innovative quality diagnostic approach, paving new research pathways for assessing the remaining shelf-life along its supply chain.

In vitro strategies for mimicking dynamic cell-ECM reciprocity in 3D culture models.

The extracellular microenvironment regulates cell decisions through the accurate presentation at the cell surface of a complex array of biochemical and biophysical signals that are mediated by the structure and composition of the extracellular matrix (ECM). On the one hand, the cells actively remodel the ECM, which on the other hand affects cell functions. This cell-ECM dynamic reciprocity is central in regulating and controlling morphogenetic and histogenetic processes. Misregulation within the extracellular space can cause aberrant bidirectional interactions between cells and ECM, resulting in dysfunctional tissues and pathological states. Therefore, tissue engineering approaches, aiming at reproducing organs and tissues in vitro, should realistically recapitulate the native cell-microenvironment crosstalk that is central for the correct functionality of tissue-engineered constructs. In this review, we will describe the most updated bioengineering approaches to recapitulate the native cell microenvironment and reproduce functional tissues and organs in vitro. We have highlighted the limitations of the use of exogenous scaffolds in recapitulating the regulatory/instructive and signal repository role of the native cell microenvironment. By contrast, strategies to reproduce human tissues and organs by inducing cells to synthetize their own ECM acting as a provisional scaffold to control and guide further tissue development and maturation hold the potential to allow the engineering of fully functional histologically competent three-dimensional (3D) tissues.

Effect of peristaltic-like movement on bioengineered intestinal tube.

The intestine is a highly heterogeneous hollow organ with biological, mechanical and chemical differences between lumen and wall. A functional human intestine model able to recreate the in vivo dynamic nature as well as the native tissue morphology is demanded for disease research and â€‹drug discovery. Here, we present a system, which combines an engineered three-dimensional (3D) tubular-shaped intestine model (3D In-tube) with a custom-made microbioreactor to impart the key aspects of the in vivo microenvironment of the human intestine, mimicking the rhythmic peristaltic movement. We adapted a previously established bottom-up tissue engineering approach, to produce the 3D tubular-shaped lamina propria and designed a glass microbioreactor to induce the air-liquid interface â€‹condition and peristaltic-like motion. Our results demonstrate the production of a villi-like protrusion and a correct spatial differentiation of the intestinal epithelial cells in enterocyte-like as well as mucus-producing-like cells on the lumen side of the 3D In-tube. This dynamic platform offers a proof-of-concept model of the human intestine.

A novel engineered dermis for in vitro photodamage research.

The realization of biologically relevant human tissue equivalents as an in vitro model to investigate human diseases, as well as to test the efficacy or toxicity of novel compounds, is emerging as a new challenge in tissue engineering. Currently, the in vitro three-dimensional (3D) dermis model mainly involves the use of cells embedded in exogenous non-human matrices. However, such models feature biological and functional disparities with native dermis, therefore limiting their relevance to the in vivo situation. The purpose of this study was to provide a reliable endogenous human dermal equivalent (HDE) able to recapitulate the extracellular matrix (ECM) remodelling of the native dermis occurring after external damage. To this end, UVA irradiation was used to induce photodamage to both the HDE and to a fibroblast-populated collagen matrix. The photodamage was investigated at the cellular and ECM level and the results showed that, although a cellular response was detected in both systems, no ECM reorganization characteristic of the in vivo photo-aged dermis could be detected in the fibroblast-populated collagen matrix. In contrast in the HDE, the neosynthesized ECM recapitulated the characteristic ageing behaviour of the dermis found in vivo, in terms of collagen and hyaluronic acid synthesis as well as collagen organization remodelling. This study therefore demonstrates the role of the endogenous ECM in recapitulating in vitro the functionality of the human dermis and the proposed HDE as a novel tool for photoprotection trials. Copyright © 2016 John Wiley & Sons, Ltd.

Engineered cardiac micromodules for the in vitro fabrication of 3D endogenous macro-tissues.

The in vitro fabrication of an endogenous cardiac muscle would have a high impact for both in vitro studies concerning cardiac tissue physiology and pathology, as well as in vivo application to potentially repair infarcted myocardium. To reach this aim, we engineered a new class of cardiac tissue precursor (CTP), specifically conceived in order to promote the synthesis and the assembly of a cardiac extracellular matrix (ECM). The CTPs were obtained by culturing a mixed cardiac cell population, composed of myocyte and non-myocyte cells, into porous gelatin microspheres in a dynamic bioreactor. By engineering the culture conditions, the CTP developed both beating properties and an endogenous immature cardiac ECM. By following a bottom-up approach, a macrotissue was fabricated by molding and packing the engineered tissue precursor in a maturation chamber. During the macrotissue formation, the tissue precursors acted as cardiac tissue depots by promoting the formation of an endogenous and interconnected cardiac network embedding the cells and the microbeads. The myocytes cell fraction pulled on ECM network and induced its compaction against the internal posts represented by the initial porous microbeads. This reciprocal interplay induced ECM consolidation without the use of external biophysical stimuli by leading to the formation of a beating and endogenous macrotissue. We have thus engineered a new class of cardiac micromodules and show its potential for the fabrication of endogenous cardiac tissue models useful for in vitro studies that involve the cardiac tissue remodeling.

A micro-perfusion bioreactor for on line investigation of ECM remodeling under hydrodynamic and biochemical stimulation.

Tissue-on-chip (TOC) systems aim at replicating complex biological dynamics in vitro with the potential either to improve the understanding of human biology or to develop more accurate therapeutic strategies. To replicate faithfully the intricate interrelationships between cells and their surrounding microenvironment, the three-dimensional (3D) tissue model must possess a responsive extracellular matrix (ECM). ECM remodeling plays a pivotal role in guiding cells and tissues functions and such aspect is somewhat denied during in vitro studies. For this purpose, we fabricated a micro-perfusion bioreactor capable to sustain the viability of 3D engineered tissue models recapitulating the process of the native ECM deposition and assembly. Engineered human dermis micro-tissue precursors (HD-µTP) were used as building blocks to generate a final tissue. HD-µTP were loaded in the perfusion space of the micro-perfusion bioreactor and, under the superimposition of different fluid dynamic regimes and biochemical stimulation, they synthesized new collagen proteins that were, then, assembled in the perfusion space forming a continuum of cells embedded in their own ECM. The micro-perfusion bioreactor was fabricated to allow the on-line monitoring of the oxygen consumption and the assembly of the newly formed collagen network via real time acquisition of the second harmonic generation (SHG) signal. The possibility to detect the collagen reorganization due to both fluid dynamic and biochemical stimulation, let us to define the optimal perfusion configuration in order to obtain a TOC system based on an endogenous and responsive ECM.

Biophysical properties of dermal building-blocks affects extra cellular matrix assembly in 3D endogenous macrotissue.

The fabrication of functional tissue units is one of the major challenges in tissue engineering due to their in vitro use in tissue-on-chip systems, as well as in modular tissue engineering for the construction of macrotissue analogs. In this work, we aim to engineer dermal tissue micromodules obtained by culturing human dermal fibroblasts into porous gelatine microscaffold. We proved that such stromal cells coupled with gelatine microscaffolds are able to synthesize and to assemble an endogenous extracellular matrix (ECM) resulting in tissue micromodules, which evolve their biophysical features over the time. In particular, we found a time-dependent variation of oxygen consumption kinetic parameters, of newly formed ECM stiffness and of micromodules self-aggregation properties. As consequence when used as building blocks to fabricate larger tissues, the initial tissue micromodules state strongly affects the ECM organization and maturation in the final macrotissue. Such results highlight the role of the micromodules properties in controlling the formation of three-dimensional macrotissue in vitro, defining an innovative design criterion for selecting tissue-building blocks for modular tissue engineering.

Cord blood unit bankability can be predicted by prenatal sonographic parameters.

OBJECTIVE: To identify possible sonographic prenatal parameters and postnatal parameters in order to obtain more bankable cord blood units (CBUs) containing a high number of primitive progenitor cells, allowing CBUs to be used as a source of haematopoietic progenitors for clinical transplantation. STUDY DESIGN: Prospective study undertaken in the Department of Gynaecology, Obstetrics and Reproductive Science, Second University of Naples, Italy. In total, 219 unrelated CBU donors were enrolled. Ultrasound parameters (biparietal diameter, head circumference, abdominal circumference, femur length, estimation of fetal weight, umbilical artery pulsatility index), collected at hospital admission, together with birth weight and placental weight, were correlated with bankable CBU parameters (CBU volume, total nucleated cell count, CD34+ cell count). RESULTS: Femur length and abdominal circumference correlated positively with bankable CBUs. Receiver operating curve analysis showed that these parameters can identify bankable CBUs. CONCLUSIONS: This is the first prospective study to show the relationship between ultrasonographic fetal parameters at term and the possibility of obtaining high-quality CBUs. As such, cord blood banking could be improved worldwide by performing low-cost ultrasonographic scans.

Time and space evolution of transport properties in agarose-chondrocyte constructs.

During the development of de novo synthesized cartilage tissue engineered constructs, transport and biophysical properties are expected to change in time and space. Monitoring and control of the evolution of these parameters are of crucial importance to process biohybrid constructs in vitro. The aim of this work was to measure fluid and macromolecular transport and evolution of mechanical properties of tissue-engineered cartilage constructs as a function of culture time and extracellular matrix (ECM) production. It was found, in agreement with other literature reports, that mechanical and fluid transport properties of the constructs correlated well with time of culture and glycosaminoglycan (GAG) content. Further, diffusion coefficients of 2 probes, dextran (500 kDa) and bovine serum albumin (BSA), correlated well with GAG production. Diffusion coefficients (D) were measured with high spatial and temporal resolution by fluorescent recovery after photobleaching (FRAP). Diffusivity steadily decreases with time while it does not vary through the thickness of the specimen. On the basis of these results, an empirical relationship between diffusion coefficient and GAG content was proposed for the 2 probes analyzed. The results of this study provide useful information to optimize and control the tissue culture process in vitro.