Effects of a Dielectric Barrier Discharge (DBD) treatment on chitosan/polyethylene oxide nanofibers and their cellular interactions

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An efficient, economical slow-freezing method for large-scale human embryonic stem cell banking

Human embryonic stem cells (hESCs) are one of the most interesting cell types for tissue engineering, cell therapy, basic scientific research, and drug screening. Fast advancement in these areas requires the availability of large amounts of safe and well-characterized hESCs from hESC banks. Therefore, optimized freezing protocols, allowing the cryopreservation of large amounts of hESC without direct contact with liquid nitrogen, need to be established. In this study, 6 different cryoprotector combinations [dimethylsulfoxide (DMSO), ethylene glycol, and hydroxyethylstarch (HES)] combined with 2 different application methods were screened with the VUB01 cell line, to establish a new slow-freezing protocol with high recovery rates and a good expansion capacity. Our best conditions were confirmed in 4 other hESC lines: H1, H9, 181, and UGent2. To our knowledge, this is the first time that HES is evaluated as a cryoprotector for hESCs. The use of 5% DMSO + 5% HES combined with a new detachment protocol leads to efficient hESC cryopreservation. This protocol involves treating the hESC colonies with cell dissociation solution, a mild dissociation solution uncommonly used for hESC culture. A recovery ratio ranging from 45.5% to 168.2% was obtained, and these were significantly different from the other tested conditions (Student's t-test, P < 0.05). The cryopreserved hESCs were morphologically comparable to control cells, exhibited a good expansion profile, were positive for pluripotent expression markers, and could still differentiate into the 3 germ layers. This new protocol allows efficient and economical hESC cryopreservation, ideal for hESC banking

A 3D cell death assay to quantitatively determine ferroptosis in spheroids

The failure of drug efficacy in clinical trials remains a big issue in cancer research. This is largely due to the limitations of two-dimensional (2D) cell cultures, the most used tool in drug screening. Nowadays, three-dimensional (3D) cultures, including spheroids, are acknowledged to be a better model of the in vivo environment, but detailed cell death assays for 3D cultures (including those for ferroptosis) are scarce. In this work, we show that a new cell death analysis method, named 3D Cell Death Assay (3DELTA), can efficiently determine different cell death types including ferroptosis and quantitatively assess cell death in tumour spheroids. Our method uses Sytox dyes as a cell death marker and Triton X-100, which efficiently permeabilizes all cells in spheroids, was used to establish 100% cell death. After optimization of Sytox concentration, Triton X-100 concentration and timing, we showed that the 3DELTA method was able to detect signals from all cells without the need to disaggregate spheroids. Moreover, in this work we demonstrated that 2D experiments cannot be extrapolated to 3D cultures as 3D cultures are less sensitive to cell death induction. In conclusion, 3DELTA is a more cost-effective way to identify and measure cell death type in 3D cultures, including spheroids.</jats:p

Effects of pre- and post-electrospinning plasma treatments on electrospun PCL nanofibers to improve cell interactions

In this study, liquid plasma treatment was used to improve the morphology of Poly-CaproLactone (PCL) NanoFibers (NFs), followed by performing a Dielectric Barrier Discharge (DBD) plasma surface modification to enhance the hydrophilicity of electrospun mats generated from plasma-modified PCL solutions. Cell interaction studies performed after 1 day and 7 days clearly revealed the highly increased cellular interactions on the double plasma-treated nanofibers compared to the pristine ones due to the combination of (1) a better NF morphology and (2) an increased surface hydrophilicity

The role of scaffold architecture and composition on the bone formation by adipose-derived stem cells

Scaffold architecture and composition are crucial parameters determining the initial cell spatial distribution and consequently bone tissue formation. Three-dimensional poly-ε-caprolactone (PCL) scaffolds with a 0/90° lay-down pattern were plotted and subjected to 1) an oxygen plasma (PCL O) or 2) a post-argon plasma modification with gelatine and fibronectin (PCL Fn). These scaffolds with an open pore structure were compared with more compact scaffolds fabricated by conventional processing techniques: oxidized polylactic acid (LA O) and collagen (COL) scaffolds. Human adipose tissue derived stem cell/scaffold interaction was studied. The study revealed that the biomimetic surface modification of plotted scaffolds did not increase the seeding efficiency. The proliferation and colonization was superior for PCL Fn in comparison with PCL O. The plotted PCL Fn was completely colonized throughout the scaffold whereas conventional scaffolds only at the edge. Protein-based scaffolds (PCL Fn and COL) enhanced the differentiation, although plotted scaffolds showed a delay in their differentiation compared with compact scaffolds. In conclusion, protein modification of plotted PCL scaffolds enhances uniform tissue formation but shows a delayed differentiation in comparison with compact scaffolds. The present study demonstrates that biomimetic PCL scaffolds could serve as a guiding template to obtain a uniform bone tissue formation in vivo

Texture profile analysis reveals a stiffer ovarian cortex after testosterone therapy : a pilot study

Purpose: The importance of the surrounding ovarian stromal cells and extracellular matrix in the development and maturation of follicles has recently gained attention. An aberrant extracellular matrix has been described in ovaries of patients with polycystic ovary syndrome where a more rigid structural environment, possibly induced by endogenous testosterone, impairs normal folliculogenesis. In this context, we describe the textural parameters of the ovarian cortex of transgender men after prolonged testosterone administration compared to the textural parameters of the non-exposed ovarian cortex originating from female oncological patients. Methods: Texture profile analysis (TPA) was performed on ovarian cortex (5 x 5 mm) of oncological and transgender patients in order to measure stiffness, hardness, cohesiveness, and springiness of the ovarian cortex (LRXplus universal testing system). Statistical analysis was performed using repeated measurements mixed models and the Spearman rank order correlation test (IBM SPSS Statistics 23). Results: A total of 36 frozen-thawed cortical strips (5 x 5 mm) were subjected to TPA. The superficial part of cortex fragments originating from transgender persons (fragments < 1.4 mm; N = 10) appeared to be significantly stiffer compared to cortex derived from oncology patients (fragments < 1.4 mm; N = 7) (6.78 +/- 1.38 N/mm versus 5.41 +/- 0.9 N/mm respectively, p = 0.036). Conclusions: This is the first application of TPA in ovarian cortex to study the physical properties. Comparing the physical properties, we objectively describe an increased cortical stiffness in the most outer part of the ovarian cortex following prolonged testosterone administration in transgender men compared to the ovarian cortex of oncological patients. This preliminary and novel approach could be the start of future research to understand the physical properties of ovarian tissue

Investigating the stability of plasma polymerized acrylic acid coatings

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