Evaluation of chicken embryo extract and egg yolk extract as alternatives to basic cell culture medium supplement.

BACKGROUND: Fetal calf serum (FCS), an existing cell culture supplement, is effective but has several drawbacks, including being expensive, requiring a lengthy process of production, and requiring a hard currency. With this in mind, we planned to evaluate chick embryo extract and egg yolk extracts in cell culture as alternatives to fetal calf serum (FCS). METHODS: Specific pathogen-free eggs were purchased from the National Veterinary Institute, Bishoftu, Ethiopia, and incubated in a humidified incubator at 37 °C for 11 days. Egg yolk extract (EYE) and chick embryo extract (CEE) were collected after the egg was opened with caution not to destroy the yolk sack or the chick embryo itself. Chick fibroblasts and Vero cells were cultured in minimum essential medium (MEM) supplemented with egg yolk extract or chick embryo extract at ratios of 0:10, 1:9, 2.5:7.5, and 5:5% fetal calf serum. RESULTS: Fibroblast cell attachment was better in media supplemented with 5% CEE and 5% FCS. The confluency was also greater than 50% at this concentration. Vero cells cultured with 5% CEE and 5% FCS also exhibited very good cell attachment and a confluency of up to 70%. Viability and confluency were also observed at 5%:5% ratios of 50 and 70%, respectively. CONCLUSION: This investigation evaluated these two extracts as cell culture supplements and revealed promising results as alternatives to fetal calf serum. The limitation of this study is that it only used two cell types and additional cell lines, and different ratios should be tested. With the above findings, further research using different cell lines, ratios and conditions is warranted.

Sil1-deficient fibroblasts generate an aberrant extracellular matrix leading to tendon disorganisation in Marinesco-Sjögren syndrome.

BACKGROUND: Marinesco-Sjögren syndrome (MSS) is an autosomal recessive neuromuscular disorder that arises in early childhood and is characterized by congenital cataracts, myopathy associated with muscle weakness, and degeneration of Purkinje neurons leading to ataxia. About 60% of MSS patients have loss-of-function mutations in the SIL1 gene. Sil1 is an endoplasmic reticulum (ER) protein required for the release of ADP from the master chaperone Bip, which in turn will release the folded proteins. The expression of non-functional Sil1 leads to the accumulation of unfolded proteins in the ER and this triggers the unfolded protein response (UPR). A dysfunctional UPR could be a key element in the pathogenesis of MSS, although our knowledge of the molecular pathology of MSS is still incomplete. METHODS: RNA-Seq transcriptomics was analysed using the String database and the Ingenuity Pathway Analysis platform. Fluorescence confocal microscopy was used to study the remodelling of the extracellular matrix (ECM). Transmission electron microscopy (TEM) was used to reveal the morphology of the ECM in vitro and in mouse tendon. RESULTS: Our transcriptomic analysis, performed on patient-derived fibroblasts, revealed 664 differentially expressed (DE) transcripts. Enrichment analysis of DE genes confirmed that the patient fibroblasts have a membrane trafficking issue. Furthermore, this analysis indicated that the extracellular space/ECM and the cell adhesion machinery, which together account for around 300 transcripts, could be affected in MSS. Functional assays showed that patient fibroblasts have a reduced capacity of ECM remodelling, reduced motility, and slower spreading during adhesion to Petri dishes. TEM micrographs of negative-stained ECM samples from these fibroblasts show differences of filaments in terms of morphology and size. Finally, structural analysis of the myotendinous junction of the soleus muscle and surrounding regions of the Achilles tendon revealed a disorganization of collagen fibres in the mouse model of MSS (woozy). CONCLUSIONS: ECM alterations can affect the proper functioning of several organs, including those damaged in MSS such as the central nervous system, skeletal muscle, bone and lens. On this basis, we propose that aberrant ECM is a key pathological feature of MSS and may help explain most of its clinical manifestations.

Synthesis of Heterostructured TiO2 Nanopores/Nanotubes by Anodizing at High Voltages.

This paper reports on the coating of heterostructured TiO2 nanopores/nanotubes on Ti substrates by anodizing at high voltages to design surfaces for biomedical implants. As the anodized voltage from 50 V to 350 V was applied, the microstructure of the coating shifted from regular TiO2 nanotubes to heterostructured TiO2 nanopores/nanotubes. In addition, the dimension of the heterostructured TiO2 nanopores/nanotubes was a function of voltage. The electrochemical characteristics of TiO2 nanotubes and heterostructured TiO2 nanopores/nanotubes were evaluated in simulated body fluid (SBF) solution. The creation of heterostructured TiO2 nanopores/nanotubes on Ti substrates resulted in a significant increase in BHK cell attachment compared to that of the Ti substrates and the TiO2 nanotubes.

Fabricating Multiphasic Angiogenic Scaffolds Using Amyloid/Roxadustat-Assisted High-Temperature Protein Printing.

Repairing multiphasic defects is cumbersome. This study presents new soft and hard scaffold designs aimed at facilitating the regeneration of multiphasic defects by enhancing angiogenesis and improving cell attachment. Here, the nonimmunogenic, nontoxic, and cost-effective human serum albumin (HSA) fibril (HSA-F) was used to fabricate thermostable (up to 90 °C) and hard printable polymers. Additionally, using a 10.0 mg/mL HSA-F, an innovative hydrogel was synthesized in a mixture with 2.0% chitosan-conjugated arginine, which can gel in a cell-friendly and pH physiological environment (pH 7.4). The presence of HSA-F in both hard and soft scaffolds led to an increase in significant attachment of the scaffolds to the human periodontal ligament fibroblast (PDLF), human umbilical vein endothelial cell (HUVEC), and human osteoblast. Further studies showed that migration (up to 157%), proliferation (up to 400%), and metabolism (up to 210%) of these cells have also improved in the direction of tissue repair. By examining different in vitro and ex ovo experiments, we observed that the final multiphasic scaffold can increase blood vessel density in the process of per-vascularization as well as angiogenesis. By providing a coculture environment including PDLF and HUVEC, important cross-talk between these two cells prevails in the presence of roxadustat drug, a proangiogenic in this study. In vitro and ex ovo results demonstrated significant enhancements in the angiogenic response and cell attachment, indicating the effectiveness of the proposed design. This approach holds promise for the regeneration of complex tissue defects by providing a conducive environment for vascularization and cellular integration, thus promoting tissue healing.

Application and Development of Electrospun Nanofiber Scaffolds for Bone Tissue Engineering.

Nanofiber scaffolds have gained significant attention in the field of bone tissue engineering. Electrospinning, a straightforward and efficient technique for producing nanofibers, has been extensively researched. When used in bone tissue engineering scaffolds, electrospun nanofibers with suitable surface properties promote new bone tissue growth and enhance cell adhesion. Recent advancements in electrospinning technology have provided innovative approaches for scaffold fabrication in bone tissue engineering. This review comprehensively examines the utilization of electrospun nanofibers in bone tissue engineering scaffolds and evaluates the relevant literature. The review begins by presenting the fundamental principles and methodologies of electrospinning. It then discusses various materials used in the production of electrospun nanofiber scaffolds for bone tissue engineering, including natural and synthetic polymers, as well as certain inorganic materials. The challenges associated with these materials are also described. The review focuses on novel electrospinning techniques for scaffold construction in bone tissue engineering, such as multilayer nanofibers, multifluid electrospinning, and the integration of electrospinning with other methods. Recent advancements in electrospinning technology have enabled the fabrication of precisely aligned nanofiber scaffolds with nanoscale architectures. These innovative methods also facilitate the fabrication of biomimetic structures, wherein bioactive substances can be incorporated and released in a controlled manner for drug delivery purposes. Moreover, they address issues encountered with traditional electrospun nanofibers, such as mechanical characteristics and biocompatibility. Consequently, the development and implementation of novel electrospinning technologies have revolutionized scaffold fabrication for bone tissue engineering.

Cold Atmospheric Plasma Improves the Colonization of Titanium with Primary Human Osteoblasts: An In Vitro Study.

Several studies have shown that cold atmospheric plasma (CAP) treatment can favourably modify titanium surfaces to promote osteoblast colonization. The aim of this study was to investigate the initial attachment of primary human osteoblasts to plasma-treated titanium. Micro-structured titanium discs were treated with cold atmospheric plasma followed by the application of primary human osteoblasts. The microwave plasma source used in this study uses helium as a carrier gas and was developed at the Leibniz Institute for Surface Modification in Leipzig, Germany. Primary human osteoblasts were analyzed by fluorescence and cell biological tests (alkaline phosphatase activity and cell proliferation using WST-1 assay). The tests were performed after 4, 12, and 24 h and showed statistically significant increased levels of cell activity after plasma treatment. The results of this study indicate that plasma treatment improves the initial attachment of primary human osteoblasts to titanium. For the first time, the positive effect of cold atmospheric plasma treatment of micro-structured titanium on the initial colonization with primary human osteoblasts has been demonstrated. Overall, this study demonstrates the excellent biocompatibility of micro-structured titanium. The results of this study support efforts to use cold atmospheric plasmas in implantology, both for preimplantation conditioning and for regeneration of lost attachment due to peri-implantitis.

Mapping the microcarrier design pathway to modernise clinical mesenchymal stromal cell expansion.

Microcarrier expansion systems show exciting potential to revolutionise mesenchymal stromal cell (MSC)-based clinical therapies by providing an opportunity for economical large-scale expansion of donor- and patient-derived cells. The poor reproducibility and efficiency of cell expansion on commercial polystyrene microcarriers have driven the development of novel microcarriers with tuneable physical, mechanical, and cell-instructive properties. These new microcarriers show innovation toward improving cell expansion outcomes, although their limited biological characterisation and compatibility with dynamic culture systems suggest the need to realign the microcarrier design pathway. Clear headway has been made toward developing infrastructure necessary for scaling up these technologies; however, key challenges remain in characterising the wholistic effects of microcarrier properties on the biological fate and function of expanded MSCs.

Human cytokeratin 1 plays a role in the interaction of Pteropine orthoreovirus with Hek293 cells but not HeLa cells.

Pteropine orthoreovirus (PRV) causes respiratory tract infections in humans. Despite its emergence as a zoonotic and respiratory virus, little is known about its cell tropism, which hampers progress in fully understanding its pathogenesis in humans. Hek293 cells are most susceptible to PRV infection, while HeLa cells are the least. Human cytokeratin 1 (CK1) was identified as the protein that interacts with PRV. The immunofluorescence assay and qPCR results revealed prior treatment with anti-CK1 may provide Hek293 cells protection against PRV. The KRT1-knockout Hek293 cells were less susceptible to PRV infection. Further study into the pathogenesis of PRV in humans is needed.

Moving grating-based laser heterodyne digital holographic microscopy system for measuring dynamic phase of living cell attachment.

We propose a laser heterodyne digital holography microscopy system based on a moving grating, which uses the Doppler principle between a moving grating and beam to achieve a low-frequency bias between the diffracted beams, abandoning traditional heterodyne digital holography that requires multiple acousto-optic modulators. The dynamic phase distribution obtained using the laser heterodyne digital holography phase-reconstruction algorithm was more realistic and analyzable than the results of the angular spectrum algorithm. The structure and algorithm were used to capture the shape characteristics of mouse fibroblasts after ~2 h of incubation (37°C, 5% CO2), and the dynamic phase distribution of the cells was monitored in real-time during the attachment process. The system proposed in this study, with its high spatial resolution and high-precision phase measurement capability, is suitable for both static and live cells.

Gelatin-tyramine addition and low hydrogel density improves cell attachment, migration, and metabolic activity in vitro and tissue response in vivo in enzymatically crosslinkable dextran-hyaluronic acid hydrogels.

Hydrogels are receiving increasing attention for their use in 3D cell culture, tissue engineering, and bioprinting applications. Each application places specific mechanical and biological demands on these hydrogels. We developed a hydrogel toolbox based on enzymatically crosslinkable polysaccharides via tyramine (TA) moieties, allowing for rapid and tunable crosslinking with well-defined stiffness and high cell viability. Including gelatin modified with TA moieties (Gel-TA) improved the hydrogels' biological properties; 3 T3 fibroblasts and HUVECs attached to and proliferated on the enriched hydrogels at minute Gel-TA concentrations, in contrast to bare or unmodified gelatin-enriched hydrogels. Moreover, we were able to switch HUVECs from a quiescent to a migratory phenotype simply by altering the ligand concentration, demonstrating the potential to easily control cell fate. In encapsulation studies, Gel-TA significantly improved the metabolic activity of 3 T3 fibroblasts in soft hydrogels. Furthermore, we showed rapid migration and network formation in Gel-TA enriched hydrogels in contrast to a non-migratory behavior in non-enriched polysaccharide hydrogels. Finally, low hydrogel density significantly improves tissue response in vivo with large infiltration and low fibrotic reaction. Further development by adding ECM proteins, peptides, and growth factor adhesion sites will lead to a toolbox for hydrogels tailored toward their desired application.

Concentration- and Solvent-Induced Chiral Tuning by Manipulating Non-Proteinogenic Amino Acids in Glycoconjugate Supra-Scaffolds: Interaction with Protein, and Streptomycin Delivery.

We showed solvent- and concentration-triggered chiral tuning of the fibrous assemblies of two novel glycoconjugates Z-P(Gly)-Glu and Z-F(4-N)-Glu made by chemical attachment of Cbz-protected [short as Z)] non-proteinogenic amino acids L-phenylglycine [short as P(Gly)] and 4-Nitro-L-phenylalanine [short as F(4-N)] with D-glucosamine [short as Glu]. Both biomimetic gelators can form self-healing and shape-persistent gels with a very low critical gelator concentration in water as well as in various organic solvents, indicating they are ambidextrous supergelators. Detailed spectroscopic studies suggested ß-sheet secondary structure formation during anisotropic self-aggregation of the gelators which resulted in the formation of hierarchical left-handed helical fibers in acetone with an interlayer spacing of 2.4 nm. After the physical characterization of the gels, serum protein interaction with the gelators was assessed, indicating they may be ideal for biomedical applications. Further, both gelators are benign, non-immunogenic, non-allergenic, and non-toxic in nature, which was confirmed by performing the blood parameters and liver function tests on Wister rats. Streptomycin-loaded hydrogels showed efficacious antibacterial activity in vitro and in vivo as well. Finally, cell attachment and biocompatibility of the hydrogels were demonstrated which opens a newer avenue for promising biomedical and therapeutic applications.

Cryo-Electrospinning Generates Highly Porous Fiber Scaffolds Which Improves Trabecular Meshwork Cell Infiltration.

Human trabecular meshwork is a sieve-like tissue with large pores, which plays a vital role in aqueous humor outflow. Dysfunction of this tissue can occur, which leads to glaucoma and permanent vision loss. Replacement of trabecular meshwork with a tissue-engineered device is the ultimate objective. This study aimed to create a biomimetic structure of trabecular meshwork using electrospinning. Conventional electrospinning was compared to cryogenic electrospinning, the latter being an adaptation of conventional electrospinning whereby dry ice is incorporated in the fiber collector system. The dry ice causes ice crystals to form in-between the fibers, increasing the inter-fiber spacing, which is retained following sublimation. Structural characterization demonstrated cryo-scaffolds to have closer recapitulation of the trabecular meshwork, in terms of pore size, porosity, and thickness. The attachment of a healthy, human trabecular meshwork cell line (NTM5) to the scaffold was not influenced by the fabrication method. The main objective was to assess cell infiltration. Cryo-scaffolds supported cell penetration deep within their structure after seven days, whereas cells remained on the outer surface for conventional scaffolds. This study demonstrates the suitability of cryogenic electrospinning for the close recapitulation of trabecular meshwork and its potential as a 3D in vitro model and, in time, a tissue-engineered device.

Development of cultivable alginate fibers for an ideal cell-cultivated meat scaffold and production of hybrid cultured meat.

Slaughtering animals for meat pose several challenges, including environmental pollution and ethical concerns. Scaffold-based cell-cultivated meat has been proposed as a solution to these problems, however, the utilization of animal-derived materials for scaffolding or the high cost of production remains a significant challenge. Alginate is an ideal material for cell-cultivated meat scaffolds but has poor cell adhesion properties. To address this issue, we achieved 82 % cell adhesion coverage by controlling the specific structure generated during the ionic crosslinking process of alginate. Post 11 days of culture; we evaluated cell adhesion, differentiation, and aligned cell networks. The cell growth increased by 12.7 % compared to the initial seeding concentration. Finally, we created hybrid cell-cultivated meat by combining single-cell protein from mycelium and cell-cultivated meat. This is non-animal based, edible, cost-effective, and has a desirable texture by blending cell-cultivated meat with a meat analogue. In summary, the creation of improved alginate fibers can effectively tackle various obstacles encountered in the manufacturing of cell-cultivated meat. This includes enhancing cell adhesion, reducing costs, and streamlining the production procedure.

Fabrication andin vitroevaluation of photo cross-linkable silk fibroin-epsilon-poly-L-lysine hydrogel for wound repair.

An optimal wound-healing hydrogel requires effective antibacterial properties and a favorable cell adhesion and proliferation environment. AlthoughBombyx morisilk fibroin (SF) possesses inherent wound-healing properties, it lacks these essential qualities. This study aimed to fabricate a novel photo-polymerizable hydrogel by utilizing SF's wound-healing efficiency and the epsilon-poly-L-lysine (EPL) antimicrobial activity. The SF was modified with three different concentrations of glycidyl methacrylate (GMA) to obtain SF-GMA(L), SF-GMA(M), and SF-GMA(H). A methacrylated EPL (EPL-GMA) was also produced. Then, SF-GMA was mixed with EPL-GMA to produce photo-crosslinkable SF-GMA-EPL hydrogels. The SF-GMA(L)-EPL, SF-GMA(M)-EPL, and SF-GMA(H)-EPL hydrogels, fabricated with 20% EPL-GMA, demonstrated maximum antimicrobial activity and mammalian cell adhesion ability. The hydroxyl radical (•OH) scavenging efficiency of the hydrogels was tested and shown to be between 69% and 74%. These hydrogels also exhibited 60% efficiency in removing bacterial lipopolysaccharides. The water absorption ability of the hydrogels was consistent with the size of their internal pores. The hydrogels exhibited a slow degradation fashion, and their degradation products appeared cytocompatible. Finally, the elastomeric properties of the hydrogels were determined, and a storage modulus (G') of 300-600 Pa was demonstrated. In conclusion, the hydrogels created in this study possess excellent biological and physical properties to support wound healing.

Endogenous Galectin-1 Modulates Cell Biological Properties of Immortalized Retinal Pigment Epithelial Cells In Vitro.

In the eye, an increase in galectin-1 is associated with various chorioretinal diseases, in which retinal pigment epithelium (RPE) cells play a crucial role in disease development and progression. Since little is known about the function of endogenous galectin-1 in these cells, we developed a galectin-1-deficient immortalized RPE cell line (ARPE-19-LGALS1-/-) using a sgRNA/Cas9 all-in-one expression vector and investigated its cell biological properties. Galectin-1 deficiency was confirmed by Western blot analysis and immunocytochemistry. Cell viability and proliferation were significantly decreased in ARPE-19-LGALS1-/- cells when compared to wild-type controls. Further on, an increased attachment of galectin-1-deficient RPE cells was observed by cell adhesion assay when compared to control cells. The diminished viability and proliferation, as well as the enhanced adhesion of galectin-1-deficient ARPE-19 cells, could be blocked, at least in part, by the additional treatment with human recombinant galectin-1. In addition, a significantly reduced migration was detected in ARPE-19-LGALS1-/- cells. In comparison to control cells, galectin-1-deficient RPE cells had enhanced expression of sm-α-actin and N-cadherin, whereas expression of E-cadherin showed no significant alteration. Finally, a compensatory expression of galectin-8 mRNA was observed in ARPE-19-LGALS1-/- cells. In conclusion, in RPE cells, endogenous galectin-1 has crucial functions for various cell biological processes, including viability, proliferation, migration, adherence, and retaining the epithelial phenotype.

Enhanced bacterial adhesion force by rifampicin resistance promotes microbial colonization on PE plastic compared to non-resistant biofilm formation.

The microbial biofilm formed on plastics, is ubiquitous in the environment. However, the effects of antibiotic resistance on the development of the biofilm on plastics, especially with regard to initial cell attachment, remain unclear. In this study, we investigated the initial bacterial adhesion and subsequent biofilm growth of a rifampin (Rif) resistant E. coli (RRE) and a normal gram-positive B. subtilis on a typical plastic (polyethylene, PE). The experiments were conducted in different antibiotic solutions, including Rif, sulfamethoxazole (SMX), and kanamycin (KM), with concentrations ranging from 1 to 1000 µg/L to simulate different aquatic environments. The AFM-based single-cell adhesion force determination revealed that Rif resistance strengthened the adhesion force of RRE to PE in the environment rich in Rif rather than SMX and KM. The enhanced adhesion force may be due to the higher secretion of extracellular polymeric substances (EPS), particularly proteins, by RRE in the presence of Rif compared to the other two antibiotics. In addition, the higher ATP level of RRE would facilitate the initial adhesion and subsequent biofilm growth. Transcriptome analysis of RRE separately cultured in Rif and SMX environments demonstrated a clear correlation between the expression of Rif resistance and the augmented bacterial adhesion and cellular activity. Biofilm biomass analysis confirmed the promotion effect of Rif resistance on biofilm growth when compared to non-resistant biofilms, establishing a novel association with the augmentation of microbial adhesion force. Our study highlights concerns related to the dissemination of antibiotic resistance during microbial colonization on plastic that may arise from antibiotic resistance.

The evaluation of the osteopromoting capabilities of composites based on biopolymers and gold/silver nanoparticles doped bioactive glasses on an experimental rat bone defect.

The most important concept behind using bone scaffolds is the biocompatibility of the material to avoid a local inflammatory response and must have the following properties: osteoinduction, osteoconductivity, angiogenesis, and mechanical support for cell growth. Gold nanoparticles/gold and silver nanoparticles -containing bioactive glasses in biopolymer composites have been used to enhance bone regeneration. These composites were testedin vitroon fibroblast and osteoblast cell lines using MTT tests, immunofluorescence, scanning electron microscopy analysis, andin vivoin an experimental bone defect in Sprague-Dawley rats. Both composites promoted adequate biological effects on human fibroblastic BJ (CRL 2522TM) cell lines and human osteoblastic cells isolated from the human patella in terms of cell proliferation, morphology, migration, and attachment. Most importantly, they did not cause cellular apoptosis and necrosis. According to the histological and immunohistochemical results, both composites were osteoinductive and promoted new bone formation at 60 d. Evidence from this study suggests that the small amount of silver content does not influence negatively thein vitroorin vivoresults. In addition, we obtained accurate results proving that the existence of apatite layer and proteins on the surface of the recovered composite, supports the validity ofin vitrobioactivity research.

De Novo Design of a Versatile Peptide-Based Coating to Impart Targeted Functionality at the Surface of Native Polystyrene.

Peptide sequence periodicity is a simple design tool that can be used to generate functional peptide-based surface coatings. De novo-designed peptide N3-PEG-VK16 is characterized by a hydrophobic periodicity of two that avidly binds to native polystyrene priming its surface for subsequent targeted functionalization via chemical ligation. The peptidic portion of N3-PEG-VK16 is responsible for surface binding, converting polystyrene's hydrophobic surface into a wettable and electrostatically charged environment that facilitates cell attachment. Native polystyrene surfaces are coated by simple peptide adsorption from an aqueous buffered solution, and the resulting primed surface is easily functionalized by cycloaddition chemistry. Herein, we show that ligating a vitronectin-derived peptide to primed polystyrene surfaces enables adhesion, expansion, long-term culture, and phenotype maintenance of human induced pluripotent stem cells. To demonstrate scope, we also show that additional functional ligands can be used, for example, nerve growth factor protein, to control neurite outgrowth.

Mechanical force drives the initial mesenchymal-epithelial interaction during skin organoid development.

Rationale: Stem cells self-organize to form organoids that generate mini-organs that resemble the physiologically-developed ones. The mechanism by which the stem cells acquire the initial potential for generating mini-organs remains elusive. Here we used skin organoids as an example to study how mechanical force drives initial epidermal-dermal interaction which potentiates skin organoids to regenerate hair follicles. Methods: Live imaging analysis, single-cell RNA-sequencing analysis, and immunofluorescence were used to analyze the contractile force of dermal cells in skin organoids. Bulk RNA-sequencing analysis, calcium probe detection, and functional perturbations were used to verify that calcium signaling pathways respond to the contractile force of dermal cells. In vitro mechanical loading experiment was used to prove that the stretching force triggers the epidermal Piezo1 expression which negatively regulates dermal cell attachment. Transplantation assay was used to test the regenerative ability of skin organoids. Results: We found that dermal cell-derived contraction force drives the movement of dermal cells surrounding the epidermal aggregates to trigger initial mesenchymal-epithelial interaction (MEI). In response to dermal cell contraction force, the arrangement of the dermal cytoskeleton was negatively regulated by the calcium signaling pathway which further influences dermal-epidermal attachment. The native contraction force generated from the dermal cell movement exerts a stretching force on the adjacent epidermal cells, activating the stretching force sensor Piezo1 in the epidermal basal cells during organoid culture. Epidermal Piezo1 in turn drives strong MEI to negatively regulate dermal cell attachment. Proper initial MEI by mechanical-chemical coupling during organoid culture is required for hair regeneration upon transplantation of the skin organoids into the back of the nude mice. Conclusion: Our study demonstrated that mechanical-chemical cascade drives the initial event of MEI during skin organoid development, which is fundamental to the organoid, developmental, and regenerative biology fields.

Effects of Cold Atmospheric Plasma Pre-Treatment of Titanium on the Biological Activity of Primary Human Gingival Fibroblasts.

Cold atmospheric plasma treatment (CAP) enables the contactless modification of titanium. This study aimed to investigate the attachment of primary human gingival fibroblasts on titanium. Machined and microstructured titanium discs were exposed to cold atmospheric plasma, followed by the application of primary human gingival fibroblasts onto the disc. The fibroblast cultures were analyzed by fluorescence, scanning electron microscopy and cell-biological tests. The treated titanium displayed a more homogeneous and denser fibroblast coverage, while its biological behavior was not altered. This study demonstrated for the first time the beneficial effect of CAP treatment on the initial attachment of primary human gingival fibroblasts on titanium. The results support the application of CAP in the context of pre-implantation conditioning, as well as of peri-implant disease treatment.