Search Results (316)

The Kindlin family of scaffold proteins plays key roles in integrin-mediated processes. Kindlin-1 and -2, encoded by the FERMT1 and FERMT2 genes, respectively, are expressed in the epidermis. Kindlin-1 plays protective roles against the development of cutaneous squamous cell carcinomas (cSCCs) in epidermal keratinocytes. However, the role of Kindlin-2 in transformed epidermal keratinocytes has remained virtually unexplored. In this study, we used siRNA approaches to generate Kindlin-2-depleted cells in three isogenic transformed keratinocyte lines. PM1, MET1, and MET4 cells model, respectively, a precancerous lesion, a primary cSCC, and a metastatic lesion of the latter. MET1 cells express both Kindlin-1 and -2. However, Kindlin-1 was not detectable in PM1 and MET4 cells. FERMT2 silencing in PM1 and MET4, but not in MET1 cells, reduced proliferation and the ability to adhere to culture surfaces and spreading. Furthermore, Kindlin-2-depleted PM1 and MET4, but not MET1 cells, exhibited decreased numbers of focal adhesions, as well as an altered F-actin and microtubule cytoskeletal organization. Significantly, FERMT2 silencing reduced the directional migration in all three cell types. These findings are consistent with the concept that, in the absence of other Kindlin orthologues, Kindlin-2 plays a prominent role in the modulation of the proliferation, spreading, focal adhesion assembly, and motility of transformed keratinocytes, as exemplified by PM1 and MET4 cells. Full article

This study involved the fabrication of poly (vinyl alcohol) (PVA) nanocomposite films using the solution-casting process, which incorporated strontium-coated silver oxide (Sr-Ag₂O) nanoparticles generated by a plant-extract assisted method. Various characterization techniques, such as XRD, SEM, TEM, UV, and FTIR, showed the formation and uniform distribution of Sr-Ag₂O nanoparticles in the PVA film, which are biocompatible nanocomposite films. The presence of hydroxyl groups leads to appreciable mixing and interaction between the Sr-Ag₂O nanoparticles and the PVA polymer. Mechanical and thermal results suggest enhanced tensile strength and increased thermal stability. In addition, the sample of PVA/Sr-Ag₂O (1.94/0.06 wt. ratio) nanocomposite film showed decreased hydrophilicity, lower hemolysis, non-toxicity, and appreciable cell migration activity, with nearly 19.95% cell migration compared to the standard drug, and the presence of Sr-Ag₂O nanoparticles favored the adhesion and spreading of cells, which triggered the reduction in the gaps. These research findings suggest that PVA/Sr-Ag₂O nanocomposite films with good mechanical, antimicrobial, non-toxic, and biocompatible properties could be applied in biological wound-healing applications. Full article

Due to their biocompatibility, biodegradability, injectability, and self-setting properties, calcium–magnesium phosphate cements (MCPCs) have proven to be effective biomaterials for bone defect filling. Two types of MCPC powders based on the magnesium whitlockite or stanfieldite phases with MgO with different magnesium contents (20 and 60%) were synthesised. The effects of magnesium ions (Mg²⁺) on functional properties such as setting time, temperature, mechanical strength, injectability, cohesion, and in vitro degradation kinetics, as well as cytocompatibility in the MG-63 cell line and the osteogenic differentiation of BM hMSCs in vitro, were analysed. The introduction of NaHA into the cement liquid results in an increase in injectability of up to 83%, provides a compressive strength of up to 22 MPa, and shows a reasonable setting time of about 20 min without an exothermic reaction. These cements had the ability to support MG-63 cell adhesion, proliferation, and spread and the osteogenic differentiation of BM hMSCs in vitro, stimulating ALPL, SP7, and RUNX2 gene expression and ALPL production. The combination of the studied physicochemical and biological properties of the developed cement compositions characterises them as bioactive, cytocompatible, and promising biomaterials for bone defect reconstruction. Full article

Electrospun fibers, possessing biomimetic characteristics similar to fibrous extracellular matrices, have attracted widespread attention as scaffold materials for skin tissue engineering. The topographical structure of electrospun fibers plays a critical role in determining cell behavior. However, the effects of fiber topography on human skin fibroblasts (HSFs) remain unclear. In this study, electrospinning technology was employed to investigate how parallel and crossed fiber architectures influence the spreading morphology, proliferation, and migration of HSFs. The results demonstrated that cells exhibited spindle-shaped elongation along single fibers; on closely spaced parallel fibers, cells formed cross-adhesions between adjacent fibers, with a fiber spacing of 30–60 μm serving as the threshold range for distinguishing individual cell behaviors. At fiber intersections, a characteristic spacing of 100 μm distinguished three distinct cellular responses: anchoring, turning, and bridging. The probability of a cell altering its preexisting migration path depended on its ability to extend laterally and reach adjacent fibers, which was constrained by the upper limit of the cell body’s minor axis. This study elucidated the unique role of the electrospun fiber topography in guiding cellular decision-making in complex microenvironments, provided important insights into topography-triggered cell migration, and highlighted the practical significance of material-guided strategies in tissue engineering. Full article

Biocompatibility remains a central issue for introducing biomaterials and nanomedicines into the clinic, requiring safety, functionality, toxicity prevention, and the control of foreign body reactions. Therefore, it is necessary to evaluate multiple biomaterial parameters and molecular interactions affecting cell functions, like apoptosis, adhesion, proliferation, or spreading, as well as intracellular signals and cellular microenvironment status. Although conventional well-established in vitro techniques are helpful at the first stages of bio and nanomaterials development, high-throughput techniques expand the screening and designing possibilities. This review presents high-throughput functional proteomics approaches, focused on protein microarrays and mass spectrometry techniques, for the evaluation of biocompatibility in the new era of biomedicine. Full article

Background:Enterococcus spp. is the third leading cause of healthcare-associated infections in the American continent, often because of the virulence factors that protect the bacterium against host defenses and facilitate tissue attachment and genetic material exchange. In addition, vancomycin, considered a last-resort treatment, has shown reduced efficacy in Enterococcus spp. strains. However, the relationship between bacterial resistance and virulence factors remains unclear. This study intends to evaluate the prevalence of glycopeptide-resistant genotypes and virulence factors in Enterococcus spp. strains. Methods: Over six months, 159 Enterococcus spp. strains causing nosocomial infections were analyzed. Multiplex PCR was performed to identify species, glycopeptide-resistant genotypes, and 12 virulence factors. Results: The most abundant species identified were Enterococcus faecalis and E. faecium. Vancomycin resistance was observed in 10.7% of the isolates, and the vanA genotype was present in 47% of resistant samples. The main virulence factors detected were acm (54%), which is related to cell adhesion; gel E (66%), a metalloproteinase linked to tissue damage; and the sex pheromones cpd (64%) and ccf (84%), which are involved in horizontal gene transfer. A significant association was found between the prevalence of acm, ccf, and cpd in VRE isolates, indicating the potential dissemination of genes to emerging strains via horizontal gene transfer. In addition, a new E. faecium, which displayed five virulence factors and harbored the vanA sequence type, was identified and registered as ST2700. Conclusions:Enterococcus faecalis and E. faecium are clinically critical due to multidrug resistance and virulence factors like acm, which aids host colonization. Genes ccf and cpd promote resistance spread via horizontal transfer, while the emerging ST2700 strain requires urgent monitoring to curb its virulent, drug-resistant spread. Full article

Chronic diabetic wounds affect 15–20% of patients and are characterized by impaired healing due to disrupted hemostasis, inflammation, proliferation, and extracellular matrix (ECM) remodeling. Low-level light therapy (LLLT) has emerged as a promising noninvasive strategy for enhancing tissue regeneration. Here, we developed a multispectral pulsed LED system combining red and near-infrared light to stimulate wound healing. In vitro photostimulation of human keratinocytes and fibroblasts on biomimetic hydrogels enhanced adhesion, spreading, migration, and proliferation via increased focal adhesion kinase (pFAK), paxillin, and F-actin expression. In vivo, daily LED treatment of streptozotocin-induced diabetic wounds accelerated closure and improved ECM remodeling. Histological and molecular analyses revealed elevated levels of MMPs, interleukins, collagen, fibronectin, FGF2, and TGF-β1, supporting regenerative healing without excessive fibrosis. These findings demonstrate that multispectral pulsed photobiomodulation enhances diabetic wound healing through focal adhesion-mediated cell migration and ECM remodeling, offering a cost-effective and clinically translatable approach for chronic wound therapy. Full article

Adhesion between Schwann cells (SCs, a type of glial cell in the peripheral nervous system) and their underlying substrates is a fundamental process that holds critical importance for the proper functioning of the peripheral nervous system. Conducting further in-depth research into the adhesion mechanisms of nerve cells is of paramount significance, as it can pave the way for the development of highly effective biomaterials and facilitate the repair of nerve injuries. Thyroid Receptor Interaction Protein 6 (TRIP6), a member of the ZYXIN family of LIM domain-containing proteins, serves as a key component of focal adhesions. It plays a pivotal role in regulating a diverse array of cellular responses, including the reorganization of the actin cytoskeleton and cell adhesion. Accumulated data indicate that RSC96 cells (rat Schwann cells), which are rat Schwann cells, exhibit integrin-based mechanosensitivity during the initial phase of adhesion, specifically within the first 24 h. This enables the cells to sense and respond to alterations in matrix stiffness. The results of immunofluorescence staining experiments revealed intriguing findings. An increase in matrix stiffness not only led to significant changes in the morphological parameters of RSC96 ells, such as circularity, aspect ratio, and cell spreading area, but also enhanced the expression levels of TRIP6, focal adhesion kinase (FAK), and vinculin within these cells. These changes collectively promoted the adhesion of RSC96 cells to the matrix. Furthermore, when TRIP6 expression was silenced in RSC96 cells cultured on hydrogels, a notable decrease in the expression of both FAK and vinculin was observed. This, in turn, had a detrimental impact on cell adhesion. In summary, the present study strongly suggests that TRIP6 may play a crucial role in promoting the adhesion of RSC96 cells to polyacrylamide hydrogels with varying stiffness. This research not only offers a fresh perspective on the study of the integrin-mediated force regulation of cell adhesion but also lays a solid foundation for potential applications in tissue engineering, regenerative medicine, and other related fields. Full article

Fibronectin (FN), a primary component of the extracellular matrix (ECM), features multiple structural domains closely linked to various cellular behaviors, including migration, spreading, adhesion, and proliferation. The FN3 domain, which contains the RGD sequence, is critical in tissue repair because it enables interaction with integrin receptors on the cell surface. However, the large molecular weight of wild-type FN presents challenges for its large-scale production through heterologous expression. Therefore, this study focused on cloning the FN3 functional domain of full-length FN for expression and validation. This study selected Bacillus subtilis as the expression host due to its prominent advantages, including efficient protein secretion, absence of endotoxins, and minimal codon bias. The recombinant vector pHT43-FN3 was successfully constructed through homologous recombination technology and transformed into Bacillus subtilis WB800N. The FN3 protein was successfully expressed after induction with IPTG. Following purification of the recombinant FN protein using a His-tag nickel column, SDS-PAGE analysis showed that the molecular weight of FN3 was approximately 27.3 kDa. Western blot analysis confirmed the correct expression of FN3, and the BCA protein assay kit determined a protein yield of 5.4 mg/L. CCK8 testing demonstrated the good biocompatibility of FN3. In vitro cell experiments showed that FN3 significantly promoted cell migration at a 20 μg/mL concentration and enhanced cell adhesion at 10 μg/mL. In summary, this study successfully utilized Bacillus subtilis to express the FN3 functional domain peptide from FN protein and has validated its ability to promote cell migration and adhesion. These findings not only provide a strategy for the expression of FN protein in B. subtilis, but also establish an experimental foundation for the potential application of FN3 protein in tissue repair fields such as cutaneous wound healing and cartilage regeneration. Full article

Electrospun collagen-based fibrous mats are of increasing interest for cell culture, regenerative medicine, and tissue engineering. The focus of this investigation is on the assessment of the electrospinning ability of bovine split hide collagen (BSHC), the effect of glycosaminoglycan (GAG) incorporation on the mats’ structural morphology, and the impact on the adhesion and proliferation of human adipose-derived mesenchymal stem cells (hAD-MSCs). Electrospun mats were prepared using benign and fluoroalcohol solutions of BSHC and BSHC/GAGs under varied operation conditions. SEM observations and analysis were employed to characterize the structural morphology of the mats. Several parameters were used to evaluate the hAD-MSC behavior: cytotoxicity, cell morphology, cell number and spreading area, cytoskeleton, focal adhesion contacts, and cell proliferation. Electrospinning using benign solvents was impossible. However, fiber mats were successfully prepared from hexafluoropropanol (HFP) solutions. Different structural morphologies and fiber diameters of the electrospun mats were observed depending on the composition and concentration of the electrospinning solutions. Both BSHC and BSHC/GAG mats supported the in vitro adhesion, growth, and differentiation of hAD-MSCs, with some variations based on their composition and structural morphology. The absence of cytotoxicity and the good hAD-MSC adhesiveness make them promising substrates for cell adhesion, proliferation, and further stem cell differentiation. Full article

This study investigated the impact of the nanocrystalline atomic layer-deposited hydroxyapatite (ALD-HA) coating of titanium (Ti) surface on the attachment and proliferation of human gingival fibroblasts (HGFs). Ti discs were divided into ALD-HA-coated and non-coated (NC) controls. HGFs were harvested from gingival biopsies of patients subjected to extraction of their third molar. The cells were cultivated on the Ti discs for 2 and 24 h to evaluate the initial cell attachment using confocal microscopy. Spreading of cells and the signals of focal adhesion proteins were measured. Moreover, the adhesion proteins vinculin and paxillin expression levels were evaluated using Western blot after 3 d of cultivation. In addition, the proliferation of HGF was assessed by cultivating the cells on Ti discs for 1, 3, and 7 d. Fibroblast spreading was significantly greater on ALD-HA surfaces than on NC surfaces after 2 h (p < 0.001). In addition, the signals of vinculin and paxillin were significantly higher on the ALD-HA than on the NC surfaces at 2 and 24 h. The confocal microscope analysis also revealed significantly higher expression of focal adhesion molecules on ALD-HA surfaces at both time points. Furthermore, the cell proliferation rate was significantly higher at d 3 (p = 0.022) and d 7 (p < 0.001) on the ALD-HA compared to the NC surfaces. These findings indicate that ALD-HA coating enhances focal adhesion formation, cell spreading, and proliferation on Ti surfaces, suggesting its potential to improve gingival tissue attachment to Ti implant surfaces. Full article

Bone graft substitutes combining the mechanical features of poly-ε-caprolactone (PCL) and the bioactivity of β-tricalcium phosphate (β-TCP) have been widely reported in the literature. Surprisingly, however, very little is known about the incorporation of carbonate at a biomimicking level. The authors studied β-TCP/PCL composites at 20 wt.% and 40 wt.%, either enriched or not with sodium bicarbonate (at 2 wt.% and 4 wt.%), through SEM and EDX analyses; surface free energy estimation; pH measurement after 1, 2, and 3 days of incubation in cell media; nanoindentation; and a protein adsorption test with bovine serum albumin. The early biological response was assessed using adipose mesenchymal stem cells, as an established in vitro model, via cellular adhesion (20 min), spreading (24 h), and viability assays (1, 3, 7 days). By increasing the β-TCP content, the composites’ hardnesses and Young’s moduli (EiT) were improved, as well as their protein adsorption compared to neat PCL. Sodium bicarbonate increased the polar component of the surface energy, alkalinized the composite with a higher β-TCP content, and attenuated its early negative cell response. Further investigation is needed to deepen the knowledge of the mechanisms underpinning the mechanical features and long-term biological behavior. Full article

Constructing the surface structures of hydroxyapatite (HA) materials is a promising strategy for orchestrating the cell behaviors of bone marrow mesenchymal stem cells (BMSCs), beneficial for advancing BMSC-based tissue repair and regenerative therapies. The majority of previous strategies have focused on fabricating artificial micro-/nano-scale geometric topographies or patterns on HA surfaces. Yet, constructing surface crystal defects has received insufficient attention and application, despite their importance as highlighted by theoretical calculations. This is largely due to the instability of crystal defects, which tend to be eliminated during crystallization. Here, given the fact that stepped structures are rich in stable crystal defects along their edges and kinks, we crafted HA dishes featuring stepped surfaces and utilized them to establish cell culture models of BMSCs. The outcomes revealed that the stepped structures markedly altered the physicochemical properties of HA surfaces and affected the cytoskeleton structures, spreading area, cell morphology, and focal adhesions of BMSCs in the cell culture model, resulting in inhibited cell adhesion. Given that YAP is a key mechanical sensitive factor, and its nuclear translocation is closely tied to cytoskeletal reorganization, the nuclear translocation efficiency of YAP has been investigated. The results showed that a changed cell adhesion could affect the nuclear translocation efficiency of YAP, which would be an important reason for the change in proliferation and differentiation ability of BMSCs. This work not only enhances the understanding of the responses of BMSCs to HA surface structures but also facilitates the design and optimization of HA materials. Moreover, our manufacturing method is facile and efficient, positioning it to potentially integrate with other processing techniques for the more effective and precise regulation of BMSCs. Full article

Multipotent vascular stem cells (MVSCs) are found in the vascular wall and surrounding tissues and possess the ability to differentiate into mesenchymal lineages. Previous studies have shown that MVSCs can be activated in response to vascular injury and differentiate into vascular smooth muscle cells (SMCs), contributing to vascular remodeling and microvessel formation. However, it remains unclear as to whether and how microenvironmental changes in the extracellular matrix, such as substrate stiffness, modulates MVSC differentiation under pathological conditions. This study demonstrated that MVSCs cultured on stiff substrates exhibited increased cell spreading, stronger cell adhesion, and a higher expression of SMC markers, including myosin heavy chain (MHC), myocardin (MYCD), calponin 1 (CNN1), and smooth muscle α-actin (SMA). In contrast, MVSCs on soft substrates showed an elevated expression of the chondrogenic markers aggrecan 1 (AGC1) and collagen-II (COL2A1). The presence of TGF-β1 further increased the expression of SMC markers on stiff substrates and chondrogenic markers on the soft substrates. Collectively, these results establish substrate stiffness as a key regulator of MVSC lineage commitment through cytoskeletal reorganization, with TGF-β1 acting as a biochemical amplifier. Our findings highlight the substrate-stiffness-dependent differentiation of MVSCs and provide mechanistic insights into the role of MVSCs in vascular remodeling during atherosclerosis development and blood vessel regeneration. Full article

Nowadays, the spread of foodborne diseases and the growing concerns about antibiotic resistance have shifted the focus of researchers towards the use of substances of natural origin. Phenyllactic acid (PLA), a naturally produced compound, has already demonstrated antimicrobial properties against pathogenic microorganisms and those responsible for food spoilage. This study examines the antibacterial and antibiofilm properties of PLA against foodborne pathogens such as Salmonella enterica Derby and Escherichia coli O26. The study showed that PLA effectively inhibited both biofilm formation and bacterial planktonic growth, with minimal inhibitory concentrations (MICs) ranging from 2 to 2.75 mg mL⁻¹. A dose-dependent inhibition of biofilm formation was observed, reaching approximately 90% for Salmonella strains and 50% for E. coli at 1.5 mg mL⁻¹. The cytotoxicity evaluation on Caco-2 cells showed that PLA was well-tolerated at concentrations up to 2 mg mL⁻¹. PLA’s effectiveness was also demonstrated in real food matrices, where its application in minced beef stored at 4 °C significantly reduced microbial populations, unlike in untreated samples where bacterial counts increased. PLA showed a good ability to inhibit biofilm formation and eradicate a mature biofilm, measuring the total bacterial biofilm biomass. Additionally, PLA was found to be biocompatible in Caco-2 cells, confirming that it poses no health risk at the tested concentrations. The study also observed that PLA reduced bacterial adhesion to intestinal cells, suggesting its potential in preventing intestinal bacterial colonization. These results highlight PLA as a promising natural antimicrobial agent for food preservation, with potential applications in sustainable packaging and controlling microbial contamination in food processing. Future studies should further explore PLA’s long-term stability and its interactions in complex food environments. Full article