Physical-Mechanical Modifications of Eggs for Food-Processing During Storage

Abstract Physical-mechanical properties of egg constituents and their modifications during storage and poststorage greatly influence the efficiency of food processing, such as the separation of white and yolk by mechanical shelling. Thick albumen height, Haugh unit, yolk index and vitelline membrane-yolk system strength of eggs from Hy-Line White and Lohmann Brown hens were analyzed during 7 mo of storage at 0°C performing 3 poststorage treatments: i) immediately after refrigeration, T1; ii) after a further 6 h at 18°C after refrigeration, T2; and iii) after a week at 18°C after refrigeration, T3. For all qualitative parameters considered, this last poststorage treatment appeared to be the factor that produced the highest decrements; with respect to the first poststorage treatment, a further week at 18°C after refrigeration can involve mean decreases of about 19, 14, 14, and 16% in thick albumen height, Haugh unit, yolk index, and vitelline membrane-yolk system strength (in terms of maximum force), respectively. During about 7 mo of storage at 0°C, the latter parameter decreases, on average, by 10%. Increasing the storage time, physical-mechanical behavior was sometimes divergent from the observed trends

Tendon-like Electrospun PLGA Scaffolds with Optimized Physical Cues Induced Tenogenic Differentiation and Boosted Immunomodulatory Properties on Amniotic Epithelial Stem Cells.

Introduction: The advanced strategies in the field of Tissue Engineering might render possible overcoming the unsatisfactory results of conventional treatments to deal with tendinopathies. In this context, the design of tendon biomimetic electrospun scaffolds engineered with Amniotic Epithelial Stem Cells (AECs), which have shown a high teno-regenerative and immunomodulatory potential in tendon-defect models, can represent a promising solution for tendon regeneration. Methods: Poly(lactide-co-glycolic) acid (PLGA) scaffolds were fabricated using the electrospinning technique to mimic the native tendon biomechanics and extracellular matrix by optimizing: fiber alignment and diameter size (1.27 and 2.5 µm), and surface chemistry using the Cold Atmospheric Plasma (CAP) Technique. Moreover, the teno-inductive and immunomodulatory effects of these parameters on AECs have been also assessed. Results: The fabricated PLGA scaffolds with highly aligned fibers and small diameter size (1.27 µm) induced a stepwise tenogenic differentiation on AECs with an early epithelial-mesenchymal transition (EMT), followed by their tenogenic differentiation. Indeed, SCX, an early tendon marker, was significantly more efficiently translated into the downstream effector TNMD, a mature tendon marker. Moreover, 1.27 µm fiber diameter induced on AECs a higher expression of anti-inflammatory interleukin mRNAs (IL-4 and IL-10). The CAP treated PLGA scaffolds showed an improved cell adhesion and infiltration without altering their topological structure and teno-inductive properties. In fact, AECs engineered with CAP treated fibers, expressed in their cytoplasm TNMD. Moreover, CAP treatment did not alter the mechanical properties of PLGA scaffolds. Conclusions: The developed electrospun PLGA scaffolds with the optimized features represent an ideal tendon-like construct that could be applied in in-vivo models to evaluate their biosafety and teno-regenerative potential

Amniotic Epithelial Stem Cells Counteract Acidic Degradation By-Products of Electrospun PLGA Scaffold by Improving their Immunomodulatory Profile In Vitro

Electrospun poly(lactic-co-glycolic acid) (PLGA) scaffolds with highly aligned fibers (ha-PLGA) represent promising materials in the field of tendon tissue engineering (TE) due to their characteristics in mimicking fibrous extracellular matrix (ECM) of tendon native tissue. Among these properties, scaffold biodegradability must be controlled allowing its replacement by a neo-formed native tendon tissue in a controlled manner. In this study, ha-PLGA were subjected to hydrolytic degradation up to 20 weeks, under di-H2 O and PBS conditions according to ISO 10993-13:2010. These were then characterized for their physical, morphological, and mechanical features. In vitro cytotoxicity tests were conducted on ovine amniotic epithelial stem cells (oAECs), up to 7 days, to assess the effect of non-buffered and buffered PLGA by-products at different concentrations on cell viability and their stimuli on oAECs’ immunomodulatory properties. The ha-PLGA scaffolds degraded slowly as evidenced by a slight decrease in mass loss (14%) and average molecular weight (35%), with estimated degradation half-time of about 40 weeks under di-H2 O. The ultrastructure morphology of the scaffolds showed no significant fiber degradation even after 20 weeks, but alteration of fiber alignment was already evident at week 1. Moreover, mechanical properties decreased throughout the degradation times under wet as well as dry PBS conditions. The influence of acid degradation media on oAECs was dose-dependent, with a considerable effect at 7 days’ culture point. This effect was notably reduced by using buffered media. To a certain level, cells were able to compensate the generated inflammation-like microenvironment by upregulating IL-10 gene expression and favoring an anti-inflammatory rather than pro-inflammatory response. These in vitro results are essential to better understand the degradation behavior of ha-PLGA in vivo and the effect of their degradation by-products on affecting cell performance. Indeed, buffering the degradation milieu could represent a promising strategy to balance scaffold degradation. These findings give good hope with reference to the in vivo condition characterized by physiological buffering systems

Hypoxia-Mimetic CoCl2 Agent Enhances Pro-Angiogenic Activities in Ovine Amniotic Epithelial Cells-Derived Conditioned Medium

Amniotic epithelial stem cells (AECs) are largely studied for their pro-regenerative properties. However, it remains undetermined if low oxygen (O2) levels that AECs experience in vivo can be of value in maintaining their biological properties after isolation. To this aim, the present study has been designed to evaluate the effects of a hypoxia-mimetic agent, cobalt chloride (CoCl2), on AECs’ stemness and angiogenic activities. First, a CoCl2 dose-effect was performed to select the concentration able to induce hypoxia, through HIF-1α stabilization, without promoting any cytotoxicity effect assessed through the analysis of cell vitality, proliferation, and apoptotic-related events. Then, the identified CoCl2 dose was evaluated on the expression and angiogenic properties of AECs’ stemness markers (OCT-4, NANOG, SOX-2) by analysing VEGF expression, angiogenic chemokines’ profiles, and AEC-derived conditioned media activity through an in vitro angiogenic xeno-assay. Results demonstrated that AECs are sensitive to the cytotoxicity effects of CoCl2. The unique concentration leading to HIF-1α stabilization and nuclear translocation was 10 µM, preserving cell viability and proliferation up to 48 h. CoCl2 exposure did not modulate stemness markers in AECs while progressively decreasing VEGF expression. On the contrary, CoCl2 treatment promoted a significant short-term release of angiogenic chemokines in culture media (CM). The enrichment in bio-active factors was confirmed by the ability of CoCl2-derived CM to induce HUVEC growth and the cells’ organization in tubule-like structures. These findings demonstrate that an ap-propriate dose of CoCl2 can be adopted as a hypoxia-mimetic agent in AECs. The short-term, chemical-induced hypoxic condition can be targeted to enhance AECs’ pro-angiogenic properties by providing a novel approach for stem cell-free therapy protocols

Tendon biomimetic 3D scaffold enhance amniotic epithelial stem cells biological potential

Tendon tissue engineering represents an emerging field whose aim focuses on the design of 3D tendon biomimetic scaffolds that should ideally combine adequate physical, mechanical, biological and functional properties of the native tissue. In this research, it was designed a bundle tendon-like PLGA 3D scaffold with highly aligned fibers on which the structure and mechanical properties were evaluated. Moreover, it was assessed scaffold’s teno-differentiative and immuno-inductive ability on amniotic epithelial stem cells (AECs). The fabricated PLGA 3D scaffolds mimic macroscopically and microscopically the structure of native tendon tissue and its biomechanical properties. Biologically, AECs seeded on the fabricated 3D scaffolds acquired a spindle tenocyte-like morphology after just 24h compared to the AECs cultured on petri dishes (CTR) which maintained their cobblestone morphology. The phenotypic change of the engineered AECs was also confirmed by visualizing TNMD protein expression, a mature tendon marker, within their cytoplasm and supported by the analysis of tendon-related genes (SCX, COL1, and TNMD) that were significantly upregulated at 7-day culture, while no TNMD protein expression or significant increase in tendon-related genes was found in CTR cells. Moreover, the 3D construct induced on AECs an upregulation of IL-10, an anti-inflammatory cytokine, maintaining basal levels of IL-12, a pro-inflammatory cytokine, showing a favorable IL10/IL12 ratio. In conclusion, the fabricated PLGA 3D scaffolds are tendon biomimetic in terms of ultrastructure and biomechanics, making them also suitable for surgical purposes. Moreover, these constructs revealed a high teno- and immuno-inductive potential on AECs and thus represent potential candidates for tendon regeneration

Timed rise from floor as a predictor of disease progression in Duchenne muscular dystrophy: An observational study

The role of timed items, and more specifically, of the time to rise from the floor, has been reported as an early prognostic factor for disease progression and loss of ambulation. The aim of our study was to investigate the possible effect of the time to rise from the floor test on the changes observed on the 6MWT over 12 months in a cohort of ambulant Duchenne boys.A total of 487 12-month data points were collected from 215 ambulant Duchenne boys. The age ranged between 5.0 and 20.0 years (mean 8.48 ±2.48 DS).The results of the time to rise from the floor at baseline ranged from 1.2 to 29.4 seconds in the boys who could perform the test. 49 patients were unable to perform the test at baseline and 87 at 12 month The 6MWT values ranged from 82 to 567 meters at baseline. 3 patients lost the ability to perform the 6mwt at 12 months. The correlation between time to rise from the floor and 6MWT at baseline was high (r = 0.6, p<0.01).Both time to rise from the floor and baseline 6MWT were relevant for predicting 6MWT changes in the group above the age of 7 years, with no interaction between the two measures, as the impact of time to rise from the floor on 6MWT change was similar in the patients below and above 350 m. Our results suggest that, time to rise from the floor can be considered an additional important prognostic factor of 12 month changes on the 6MWT and, more generally, of disease progression

Tendon Immune Regeneration: Insights on the Synergetic Role of Stem and Immune Cells during Tendon Regeneration

Tendon disorders represent a very common pathology in today’s population, and tendinopathies that account 30% of tendon-related injuries, affect yearly millions of people which in turn cause huge socioeconomic and health repercussions worldwide. Inflammation plays a prominent role in the development of tendon pathologies, and advances in understanding the underlying mechanisms during the inflammatory state have provided additional insights into its potential role in tendon dis-orders. Different cell compartments, in combination with secreted immune modulators, have shown to control and modulate the inflammatory response during tendinopathies. Stromal compartment represented by tenocytes has shown to display an important role in orchestrating the inflammatory response during tendon injuries due to the interplay they exhibit with the immune-sensing and infiltrating compartments, which belong to resident and recruited immune cells. The use of stem cells or their derived secretomes within the regenerative medicine field might represent synergic new therapeutical approaches that can be used to tune the reaction of immune cells within the damaged tissues. To this end, promising opportunities are headed to the stimulation of macrophages polarization towards anti-inflammatory phenotype together with the recruitment of stem cells, that possess immunomodulatory properties, able to infiltrate within the damaged tissues and improve tendinopathies resolution. Indeed, the comprehension of the interactions between tenocytes or stem cells with the immune cells might considerably modulate the immune reaction solving hence the inflammatory response and preventing fibrotic tissue formation. The purpose of this review is to compare the roles of distinct cell compartments during tendon homeostasis and injury. Furthermore, the role of immune cells in this field, as well as their interactions with stem cells and tenocytes during tendon regeneration, will be discussed to gain insights into new ways for dealing with tendinopathies

Scaffold-Mediated Immunoengineering as Innovative Strategy for Tendon Regeneration

Tendon injuries are at the frontier of innovative approaches to public health concerns and sectoral policy objectives. Indeed, these injuries remain difficult to manage due to tendon’s poor healing ability ascribable to a hypo-cellularity and low vascularity, leading to the formation of a fibrotic tissue affecting its functionality. Tissue engineering represents a promising solution for the regeneration of damaged tendons with the aim to stimulate tissue regeneration or to produce functional implantable biomaterials. However, any technological advancement must take into consideration the role of the immune system in tissue regeneration and the potential of biomaterial scaffolds to control the immune signaling, creating a pro-regenerative environment. In this context, immunoengineering has emerged as a new discipline, developing innovative strategies for tendon injuries. It aims at designing scaffolds, in combination with engineered bioactive molecules and/or stem cells, able to modulate the interaction between the transplanted biomaterial-scaffold and the host tissue allowing a pro-regenerative immune response, therefore hindering fibrosis occurrence at the injury site and guiding tendon regeneration. Thus, this review is aimed at giving an overview on the role exerted from different tissue engineering actors in leading immunoregeneration by crosstalking with stem and immune cells to generate new paradigms in designing regenerative medicine approaches for tendon injuries