Utilization of GelMA with phosphate glass fibers for glial cell alignment

Glial cell alignment in tissue engineered constructs is essential for achieving functional outcomes in neural recovery. While gelatin methacrylate (GelMA) hydrogel offers superior biocompatibility along with permissive structure and tailorable mechanical properties, phosphate glass fibers (PGFs) can provide physical cues for directionality of neural growth. Aligned PGFs were fabricated by a melt quenching and fiber drawing method and utilized with synthesized GelMA hydrogel. The mechanical properties of GelMA and biocompatibility of the GelMA-PGFs composite were investigated in vitro using rat glial cells. GelMA with 86% methacrylation degree were photo-crosslinked using 0.1%wt photo-initiator (PI). Photocrosslinking under UV exposure for 60 s was used to produce hydrogels (GelMA-60). PGFs were introduced into the GelMA before crosslinking. Storage modulus and loss modulus of GelMA-60 was 24.73 ± 2.52 and 1.08 ± 0.23 kN/m2 , respectively. Increased cell alignment was observed in GelMA-PGFs compared with GelMA hydrogel alone. These findings suggest GelMA-PGFs can provide glial cells with physical cues necessary to achieve cell alignment. This approach could further be used to achieve glial cell alignment in bioengineered constructs designed to bridge damaged nerve tissue

The functional relationship between transglutaminase 2 and transforming growth factor β1 in the regulation of angiogenesis and endothelial-mesenchymal transition

The importance of transglutaminase 2 (TG2) in angiogenesis has been highlighted in recent studies, but other roles of this multi-functional enzyme in endothelial cell (EC) function still remains to be fully elucidated. We previously showed that the extracellular TG2 is involved in maintaining tubule formation in ECs by a mechanism involving matrix-bound vascular endothelial growth factor (VEGF) signalling. Here, by using the ECs and fibroblast co-culture and ECs 3D culture models, we demonstrate a further role for TG2 in both endothelial tubule formation and in tubule loss, which involves its role in the regulation of transforming growth factor β1 (TGFβ1) and Smad signalling. We demonstrate that inhibition of tubule formation by TG2 inhibitors can be restored by add-back of exogenous TGFβ1 at pg/ml levels and show that TG2 -/- mouse ECs are unable to form tubules in 3D culture and display negligible Smad signalling compared to wild-type cells. Loss of tubule formation in the TG2 -/- ECs can be reconstituted by transduction with TG2. We demonstrate that extracellular TG2 also has an important role in TGFβ1-induced transition of ECs into myofibroblast-like cells (endothelial-mesenchymal transition), resulting in loss of EC tubules and tubule formation. Our data also indicate that TG2 may have a role in regulating TGFβ signalling through entrapment of active TGFβ1 into the extracellular matrix. In conclusion, our work demonstrates that TG2 has multi-functional roles in ECs where its ability to fine-tune of TGFβ1 signalling means it can be involved in both endothelial tubule formation and tubule rarefaction

The role of tissue transglutaminase (TG2) in regulating the tumour progression of the mouse colon carcinoma CT26

The multifunctional enzyme tissue transglutaminase (TG2) is reported to both mediate and inhibit tumour progression. To elucidate these different roles of TG2, we established a series of stable-transfected mouse colon carcinoma CT26 cells expressing a catalytically active (wild type) and a transamidating-inactive TG2 (Cys277Ser) mutant. Comparison of the TG2-transfected cells with the empty vector control indicated no differences in cell proliferation, apoptosis and susceptibility to doxorubicin, which correlated with no detectable changes in the activation of the transcription factor NF-?B. TG2-transfected cells showed increased expression of integrin ß3, and were more adherent and less migratory on fibronectin than control cells. Direct interaction of TG2 with ß3 integrins was demonstrated by immunoprecipitation, suggesting that TG2 acts as a coreceptor for fibronectin with ß3 integrins. All cells expressed the same level of TGFß receptors I and II, but only cells transfected with active TG2 had increased levels of TGFß1 and matrix-deposited fibronectin, which could be inhibited by TG2 site-directed inhibitors. Moreover, only cells transfected with active TG2 were capable of inhibiting tumour growth when compared to the empty vector controls. We conclude that in this colon carcinoma model increased levels of active TG2 are unfavourable to tumour growth due to their role in activation of TGFß1 and increased matrix deposition, which in turn favours increased cell adhesion and a lowered migratory and invasive behaviour

Identifying Bioglass and Liquid Exfoliation of Graphite/MWCNT Mixtures Through UV–Vis Spectroscopy

Carbon allotropes such as graphene and multiwalled carbon nanotube (MWCNT) are studied for an extensive range of applications. Various exfoliation techniques were employed to yield the best form of generated allotropes. Liquid phase exfoliation utilizes the technique of sonication of these allotropes in the solvent, results in the best desired form of the high quality, safe, simple and economically viable final product. This study discusses liquid-phase exfoliation of graphene and MWCNT in chloroform. Their absorbance intensity has shown a contrast solubility profile with respect to different weight percentages of each allotrope. The comparative study was further analyzed with modification of bioglass (BG) within the suspensions, of which hazards in agglomerations of allotropes’ particles as concentration increases could potentially give a prevention insight for better preparation and processing materials formulation. Hence the study aims in reporting UV absorbance intensity of various weight percentages of liquid exfoliated graphene and MWCNT particles, with the addition of BG in chloroform due to their unique structures and remarkable properties and their exploitation in diverse potential applications, including the biomedical engineering field especially in the field of bone tissue engineering

Determination of water content in Dehydrated Mammalian cells using terahertz pulsed imaging: A feasibility study

Mammalian cells are involved in a range of biotechnological applications and more recently have been increasingly exploited in regenerative medicine. Critical to successful applications involving mammalian cells are their long-term storage and transport, for which cryopreservation in liquid nitrogen is the most frequently used strategy. However, cryopreservation suffers from high costs, difficulties in transport logistics and the use of undesirable additives (e.g. animal sera or DMSO). An alternative approach, proposed as low cost, low maintenance and process-compatible, is viable desiccation of mammalian cells. Several groups claim to have achieved this, but the extent of desiccation in the cell samples concerned is not always clear, in part because of difficulties in determining very low water content. Although several techniques exist that are frequently used to quantify the amount of water in samples (e.g. FTIR spectroscopy, thermogravimetric analysis (TGA), NMR spectroscopy), the complexity of sample preparation, as well as the costs and time constraints involved are disadvantageous. Here, we assess a novel, rapid and low cost technique, i.e. terahertz (THz) spectroscopy, for the quantification of water content within dehydrated mammalian cell samples

Human Three-Dimensional Models for Studying Skin Pathogens

Skin is the most exposed surface of the human body, separating the microbe-rich external environment, from the sterile inner part. When skin is breached or its homeostasis is perturbed, bacterial, fungal and viral pathogens can cause local infections or use the skin as an entry site to spread to other organs. In the last decades, it has become clear that skin provides niches for permanent microbial colonization, and it actively interacts with microorganisms. This crosstalk promotes skin homeostasis and immune maturation, preventing expansion of harmful organisms. Skin commensals, however, are often found to be skin most prevalent and dangerous pathogens. Despite the medical interest, mechanisms of colonization and invasion for most skin pathogens are poorly understood. This limitation is due to the lack of reliable skin models. Indeed, animal models do not adequately mimic neither the anatomy nor the immune response of human skin. Human 3D skin models overcome these limitations and can provide new insights into the molecular mechanisms of microbial pathogenesis. Herein, we address the strengths and weaknesses of different types of human skin models and we review the main findings obtained using these models to study skin pathogens