Tissue engineering. Part C, Methods最新文献

Cell membrane isolation is essential for diverse biological investigations, ranging from fundamental research to advanced therapeutic applications. This study compared two methods-differential centrifugation and discontinuous sucrose density gradient ultracentrifugation-for isolating cell membranes from the human natural killer (NK) cell line (KHYG-1). The aim was to identify the method that minimizes contamination from nuclear, mitochondrial, and cytosolic components. Differential centrifugation yielded approximately 8 mg of cell membrane, whereas sucrose density gradient ultracentrifugation produced about 5 mg. Despite the lower yield, the latter method exhibited superior performance due to significantly reduced contamination. This protocol is adaptable to various cell types, offering a reliable approach for producing cell membrane-coated mimics for therapeutic use. The increasing demand for isolated cell membranes in biomedical applications highlights the importance of optimizing isolation techniques.

Experimental analyses of knee joint contractures have traditionally utilized a 6-month rabbit model as the gold standard. However, this model is time-intensive and costly. The purpose of this study was to develop an abbreviated rabbit model of knee contractures and compare it to the well-established longer model. Twenty female New Zealand White rabbits were divided into two equal groups and prospectively studied to assess knee passive extension angles (PEA), contracture angles (CA), and terminal posterior capsular stiffness. Experimental knees were immobilized for either 4 weeks (n = 10) with an 8-week remobilization period in the abbreviated model (i.e., 3 months) or for 8 weeks (n = 10) with a 16-week remobilization period in the standard model (i.e., 6 months). PEAs were assessed at remobilization and several time points using differing vertical forces. At sacrifice, terminal biomechanical data were collected to assess posterior capsular stiffness. Analysis of PEAs in live animals at each torque value and time point demonstrated increased PEAs and decreased CAs in the 3-month abbreviated model as compared to the 6-month standard model. At sacrifice, biomechanical analysis demonstrated that the posterior capsules of the 3-month experimental limbs were significantly more stiff than the contralateral limb (2.4 vs. 0.05 Ncm/°, p < 0.0001), but significantly less stiff compared to the 6-month experimental limbs (2.4 vs. 4.7 Ncm/°, p < 0.0001). Our study suggests that the 6-month standard rabbit knee model of arthrofibrosis should continue to be used in the laboratory assessment of arthrofibrosis. However, the abbreviated model may be beneficial under selected experimental conditions.

To investigate the histomorphometric performance of two-stage maxillary sinus floor elevation (TMSFE) with various bone substitutes in the treatment of atrophic posterior maxilla. Four databases (PubMed, Embase, Web of Science, and The Cochrane Library) were searched from the beginning of database establishment to August 8, 2023. The included articles were limited to the English language. A systematic search was performed to identify randomized controlled trials assessing the histological performance of various biomaterials in TMSFE with a follow-up of 5-8 months. The main outcome was an area of new bone, and an additional outcome was residual graft material. Extracted data were analyzed by using a Bayesian approach (the Markov chain Monte Carlo) to establish ranks of various biomaterials in R language. Finally, the search identified 22 studies that reported 22 trials on bone area (17 kinds of biomaterials) and 12 studies on residual graft materials (12 kinds of biomaterials) after the exclusion of one study disconnected from the network plot. No local inconsistency could be found in studies regarding bone formation, while no closed loop was detected in residual graft material. The top 3 probabilities of biomaterials in terms of bone formation were Allograft + Xenograft (AG + X) (87.14%), X + Polymer (75.69%), and Autogenous Bone + Bioactive Glass (AB + BG) (71.44%). AG + X had the highest probability (87.14%) of being the most optimal treatment for bone formation. Biphasic calcium phosphate + Fibrin sealant (BCP + FS) was ranked as the slowest absorbing biomaterial (78.27%) in TMSFE. Within the limitations of the current network meta-analysis, AG + X may represent an optimal biomaterial for bone formation in TMSFE. The use of X in combination with other biomaterials demonstrates superior osteogenic effects in TMSFE. BCP + FS exhibited strong mechanical properties during a short-term observational period. The present findings suggest that AB is not the only feasible standard for bone grafts.

Stem cells play a critical role in the regeneration process by proliferating and differentiating to form new bone tissue. However, stem cells tend to lose their stemness and pluripotency during in vitro expansion, resulting in reduced bone regeneration capacity after osteogenic induction. Our aim is to enhance the osteogenic impact of human umbilical cord-derived mesenchymal stromal cells (hUC-MSCs) through spontaneous spheroid in vitro. The pluripotency and osteogenesis-related genes up-regulated in hUC-MSCs can be enhanced in spontaneous spheroids in vitro. For in vivo testing, spontaneous spheroids were transplanted into mice using beta-tricalcium phosphate as a scaffold. Transplant samples were stained using hematoxylin and eosin (HE), immunohistochemistry, and TRAP staining. The samples showed new bone formation, upregulated SP7 and OCN expression, and more vigorous bone metabolism in the Sph-OI group than the other groups. However, new bone formation was mainly immature bone. Overall, our findings demonstrate that hUC-MSC spheroids possess remarkable pluripotency, with the spontaneous spheroids formed following osteogenic induction exhibiting enhanced osteogenic differentiation potential and bone regeneration capacity. However, optimizing the osteogenic differentiation process and elucidating the underlying mechanisms of bone regeneration are critical scientific issues that urgently need to be addressed to enable its application in bone regeneration.

Hepatocellular carcinoma (HCC) is an aggressive liver tumor with a unique metabolic profile and a shift to glycolytic metabolism. This review discusses the contribution of pyruvate kinase M2 (PKM2) to HCC development and its potential as a target for therapy. We carried out a broad literature review on PKM2, focusing on its role in the glycolytic pathway and special interactions with key signaling pathways like Phosphoinositide 3-kinase/Protein kinase B/Mammalian target of rapamycin (PI3K/AKT/mTOR) and Mitogen-activated protein kinase (MAPK). PKM2 also performs a dual role in energy metabolism and signal transduction in HCC. PKM2 is paramount in the induction of HCC by regulating cellular metabolism and oncogenic signaling pathways. It promotes tumor growth, survival, and metastasis through interaction with the PI3K/AKT/mTOR and MAPK pathways. PKM2 is a key factor in HCC pathogenesis, with a dual impact on metabolism and signaling. Its properties may open the way for developing novel therapeutic interventions against HCC. Thus, PKM2 inhibition may offer further opportunities for tumor growth blockade, which could meaningfully improve patients' clinical outcomes.

Sympathetic innervation plays a critical role in regulating vascular function, yet its influence on vascular regeneration and reinnervation following ischemic injury remains poorly understood. This study develops and validates murine models of localized sympathetic denervation using 6-hydroxydopamine (6-OHDA) to enable study of the sympathetic nervous system's impact on vascular systems during tissue repair. Two methods of 6-OHDA administration were employed: a single topical application during open surgery and minimally invasive weekly subcutaneous injections. The topical application model achieved temporary denervation lasting 1 week without causing vascular damage, while the subcutaneous injection model provided sustained denervation for up to 4 weeks with minimal inflammation and no significant changes to vascular architecture. To investigate the effects of denervation in an ischemic context, these models were combined with a hindlimb ischemia model. Ischemia induced persistent denervation in both 6-OHDA-treated and control limbs, with limited sympathetic nerve regeneration observed over 4 weeks. Despite persistent denervation, microvascular density and perfusion recovery in ischemic muscles were comparable between denervated and control groups. This suggests that ischemia governs vascular regeneration independently of sympathetic input. These results demonstrate that localized 6-OHDA administration provides a versatile tool for achieving controlled sympathetic denervation in peripheral arteries. These models provide a novel platform for studying vascular regeneration and reinnervation under both normal and ischemic conditions, offering novel insights into the interactions between neural regulation and vascular repair processes. This work lays the foundation for future research into neural-vascular crosstalk and new possibilities for developing regenerative therapies targeting the autonomic regulation of vascular health.

In vitro experiments, a crucial component of preclinical research, are widely used due to their accessibility and controlled conditions. However, traditional two-dimensional (2D) cell models are limited in their ability to simulate the complex interactions in organ systems. To address it, emerging technologies have shifted cell cultures from 2D to three-dimensional (3D), offering improved in vitro-in vivo correlation for traditional in vitro screening. Reconstructed human epidermis (RHE) is a 3D skin tissue model that closely mimics human skin in both structure and function. We established a sodium dodecyl sulfate (SDS)-induced epidermal injury model on RHE, and the result demonstrated that treating RHE with a 2.5 mg/mL SDS solution for 24 h could cause a significant epidermal damage. We also treated it with common clinical repair biomaterials, to screen the key indicator of SDS-induced 3D epidermal injury model, which includes several chemokines such as regulated upon activation normal T-cell expressed and secreted and interferon-γ-induced protein 10 that triggered inflammatory responses, and the important component protein of barrier structure-filaggrin and loricrin. In this study, we provide a platform for biomaterials evaluation that offers support and complementarities for in vitro experiments of skin repair.

Nanozymes, as innovative enzyme mimics, hold significant promise for wound care, including antibacterial properties and tissue regeneration. Given their potential to transform wound management, this study utilizes advanced bibliometric tools to provide a comprehensive analysis of the nanozyme research landscape. The analysis covers various aspects, including publication trends, institutional contributions, journal coverage, and author involvement, offering a holistic view of research dynamics. It reveals the evolution of nanozyme research across different phases of wound healing by examining keyword co-occurrence frequencies and timeline developments. In addition, the study identifies emerging research clusters within these phases, focusing on three key areas: enhancing nanozyme performance, integrating them with hydrogel matrices, and developing responsiveness to external stimuli. These clusters highlight the increasing sophistication and diversity of nanozyme-based solutions for wound care. Furthermore, the study explores the intersection of nanozyme research with artificial intelligence (AI) and wearable sensors. This integration presents unprecedented opportunities for real-time monitoring, personalized treatment plans, and predictive analytics in wound care. The findings indicate a growing interest in this interdisciplinary field, pinpointing research frontiers centered around AI-driven wound assessment, continuous monitoring through wearable technologies, and the application of AI algorithms in nanozyme-based wound dressings. In summary, this bibliometric study provides a comprehensive global overview of research trends, key literature, hotspots, and emerging frontiers in nanozyme-based wound care. By investigating the synergy between AI, wearable sensors, and nanozymes, it elucidates the potential for novel and personalized treatment strategies in this rapidly advancing field.

Mouse embryonic fibroblasts (MEFs) have been widely used as feeder cells in embryonic stem cell cultures because they can mimic the embryonic microenvironment. Milk fat globule-epidermal growth factor 8 (MFGE8) is expressed during mouse gonadal development, 10.5-13.5 embryonic, and is also found in MEF-conditioned medium (MEF-CM). Feeder-less culture of human-induced pluripotent stem cells (iPSCs) with MEF-CM significantly decreased the number of adherent cells when an inhibitory antibody against MFGE8 was used. The concentration of mouse MFGE8 in MEF-CM, as measured by an ELISA (Enzyme-Linked Immunosorbent Assay), was 0.16-1.24 μg/mL. Mouse MFGE8 and human MFGE8 have partially different molecular structures. Both the recombinant mouse MFGE8 and human MFGE8 significantly promoted cell adhesion of human iPSCs at medium-added concentrations of 2 μg/mL. This cell adhesion was also strongly inhibited by Arginylglycylaspartic acid (RGD) inhibitors, suggesting that it is dependent on the RGD sequence. The integrin αVβ5 expressed in iPSCs was thought to be involved in binding to the RGD sequence. MEF-CMs have long been an essential bio-derived material for the feeder culture method of iPSC culture. This study demonstrates that MFGE8 in MEF-CM is a functional factor in the promoting of cell adhesion of human iPSCs. Furthermore, the use of MFGE8-containing media demonstrates that iPSCs can be established and cultured while maintaining pluripotency and inducing three germ layer differentiation. The results of this study suggest the possibility of using MFGE8 as a scaffold material suitable for inducing differentiation when reproducing in vivo maturation in vitro.