Poly(ε-caprolactone) (PCL) is widely used as a scaffold material for tissue regeneration; however, its intrinsic hydrophobicity and slow biodegradation limit cell adhesion and tissue remodeling. This study aimed to improve the hydrophilicity and biodegradability of PCL-based scaffolds by incorporating the hydrophilic polymer poloxamer 407 (P407) while maintaining mechanical stability through hydroxyapatite (HA) reinforcement. The composite system was designed as a bioactive platform for dentin-pulp complex regeneration and other regenerative endodontic applications.

Parkinson's disease (PD) is an illness that causes both motor and non-motor symptoms in the patient which occurs as a result of a progressive loss of dopamine-producing neurons in the substantia nigra. Even though the success of symptomatic treatments is promising, at the same time there is currently no effective therapy that can halt or reverse disease progression. Key genes such as SNCA, LRRK2, and PINK1 are considered as the main hopefuls aspect for the treatment of Parkinson's because mutations of these genes are the reason for the appearance of the familial and sporadic kinds of the disease, respectively. The CRISPR-Cas system, a breakthrough genome-editing technology which enables precise and targeted genetic modifications, renders the possibilities of both PD research and therapy. Examining the mechanics of prime editing, base editing, and CRISPR-Cas9 highlights how effective and precise these methods are for modifying genes. An overview of recent developments in the use of CRISPR to create PD models is also included in the current review, with a focus on the roles these models play in clarifying disease pathways and locating new treatment targets. These models include isogenic cell lines, transgenic animals, and induced pluripotent stem cells (iPSCs). This review highlights the potential of CRISPR-based strategies to correct PD-associated mutations, modulate pathogenic gene expression, and develop neuroprotective interventions targeting key processes such as mitochondrial dysfunction. Furthermore, it critically evaluates the role of CRISPR-based technologies as transformative tools in PD research and therapy while highlighting key challenges for their clinical translation.

Spinal cord injury (SCI) leads to severe neurological dysfunction. This study investigated the role of exosome-derived netrin-1 (NTN1) in axonal repair after SCI.

Restoring innervation is essential for functional pulp regeneration. Apelin, an endogenous bioactive peptide widely expressed in the nervous system, has been reported to facilitate neuroprotection and neural differentiation. This study aims to investigate the role of Apelin-13 in promoting neural differentiation of human dental pulp stem cells (hDPSCs) and to elucidate the underlying molecular mechanisms.

Human chorionic villous mesenchymal stem cells (CV-MSCs) are a promising and effective treatment for tissue injury. Trophoblast dysfunction during pregnancy is significantly involved in the pathogenesis of preeclampsia (PE). This work aimed to understand how CV-MSCs regulate trophoblast function. In this study, we treated trophoblasts with CV-MSC-derived exosomes, and RNA-seq analysis was used to understand the differences in trophoblasts induced by exosomes. We examined the levels of TXNIP (thioredoxin-interactingprotein) and β-catenin in trophoblasts by immunohistochemistry, western blotting and qRT-PCR assays. Luciferase reporter assays and qRT-PCR assays were used to examine the role of miR135 b-5p in the effects of CV-MSC-derived exosomes. The growth and invasion of trophoblasts was evaluated using CCK-8 and Transwell assays. CV-MSC-derived exosomes markedly enhanced trophoblast proliferation and invasion. Furthermore, a significant decrease in TXNIP expression and activation of the β-catenin pathway in CV-MSC exosome-treated trophoblasts were observed. Consistent with these findings, TXNIP inhibition promoted trophoblast proliferation and invasion, similar to the effect of CV-MSC-derived exosomes, and this effect was accompanied by activation of the β-catenin pathway. In addition, overexpression of TXNIP inactivated the β-catenin pathway in trophoblasts and reduced the proliferation and invasion of trophoblasts. Importantly, miR135 b-5p was highly expressed in CV-MSC exosomes and interacted with TXNIP. MiR-135 b-5p overexpression significantly increased the proliferation and invasion of trophoblasts, which was attenuated by TXNIP overexpression. Our results suggest that TXNIP-dependent β-catenin pathway activation mediated by miR-135 b-5p, which was delivered by CV-MSC-derived exosomes, could promote the proliferation and invasion of trophoblasts.

Tissue engineering offers great promise for regenerating damaged organs including the heart. Although direct attachment of grafts at the target site is possible during surgery, minimally invasive delivery and suture-free approaches could reduce patient discomfort and allow repeat administration. However, for the therapy to be effective it is essential that the graft is successfully delivered to the epicardium and retained on target. Here, methacrylated alginate-based shape-memory patches labelled with 111InCl3 and loaded with luciferase expressing stem-cells were either injected towards the epicardium under ultrasound guidance or surgically grafted onto mouse hearts. Patch and cell location were serially tracked using SPECT-CT and bioluminescence imaging. Radiolabelling of shape-memory patches permitted serial tracking of graft location for seven days in-vivo, and revealed that injected patches rarely attached on-target whilst surgically implanted patches rapidly detached from the epicardium. In-vivo imaging was then used to evaluate modifications to biomaterial formulation and patch attachment strategies. This ultimately resulted in effective, suture-free surgical attachment of chitosan-coated patches loaded with luciferase-expressing human embryonic stem cell-derived epicardial cells onto the heart, illustrating a model therapeutic. This translational imaging approach facilitates iterative optimization of a novel biomaterial and could have wide-reaching applications for enhancing a range of regenerative therapies.

Stem cell-based therapies and liposome-integrated hydrogel matrix strategies are promising for articular cartilage regeneration. However, challenges persist due to the loss of stemness during in vitro expansion and the lack of control over stem cell behavior in vivo. Herein, we design a cartilage-inspired hydrogel scaffold similar to the native stem cell niche and expect to enhance chondrogenesis for cartilage repair. Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) are encapsulated within a composite hydrogel composed of methacrylate hyaluronic acid and collagen networks, mimicking the extracellular matrix of native cartilage. Additionally, embedded liposomes participate to form a lubrication mechanism, the boundary layer in the synovial joints, which lowers the coefficient of friction close to the native level. In vitro, our hydrogel niche system enhances cell-cell and cell-matrix interactions, induces hUC-MSCs mesenchymal condensation, establishes the hypoxic microenvironment formation, and therefore improves the chondrogenic differentiation. Moreover, SOX9, COL2, and HIF-1α expression are all upregulated, whereas MMP13 is downregulated. Meanwhile, in vivo implantation in an osteochondral defect model demonstrates our repair strategy is superior to the conventional hydrogel scaffolds. The advantages include the restoration of PRG4, the suppression of MMP13 and the increasement of ICRS II score to the native levels at 6 weeks post-implantation. These findings are important biophysical cues in regulating stem cell fate in a three-dimensional hydrogel niche and conducive to developing biomimetic scaffolds for cartilage repair.

Joint cartilage regeneration presents a significant challenge due to its limited self-repair capacity, hindered by local inflammation and metabolic dysregulation. Mitochondrial quality control (MQC) is crucial for maintaining chondrocyte differentiation and metabolism; however, strategies targeting MQC remain underexplored. Here, we generated an acellular delivery vehicle, cytoplastCXCR4, which was designed to facilitate mitochondria transport and restore MQC, from enucleated CXCR4 transfected mesenchymal stem cells (MSCs) through ultracentrifugation. CXCR4 was overexpressed to enhance migration toward inflamed cartilage via its interaction with SDF-1α, which is a chemokine elevated at the site of cartilage damage. The CXCR4 gene-transfected, nuclear-depleted cytoplastCXCR4 exhibited enhanced homing ability, via CXCR4-SDF-1α axis. In vitro results showed that cytoplastCXCR4 treatment improved mitochondrial quality-structure, oxygen consumption, and ATP synthesis-in osteoarthritic chondrocytes, thus re-establishing metabolic balance. Compared to exosomes, cytoplasts retain cellular bioactivity, including functional organelles and paracrine signaling. Notably, we observed mitochondrial transfer via cytoplast-derived vesicles, which contributed to the restoration of morphology and function in damaged mitochondria within injured chondrocytes. Furthermore, in vivo results demonstrated that cytoplastsCXCR4 had a longer retention time, and could precisely target the damaged cartilage and significantly facilitate cartilage repair compared with exosomes in a rat knee cartilage defect model. This study presents an MQC-targeted and mitochondria-preserving approach, offering an improved strategy for joint cartilage regeneration.

Embryonic stem cells (ESCs) constitute a unique lab resource, as they can differentiate to all other cell types, making them pluripotent. We established cynomolgus ESCs using ICSI-derived blastocysts. Cynomolgus ESCs expressed pluripotent markers in immunostaining and fluorescence-activated cell sorting analyses. To investigate its differentiation potential, we confirmed the expression of layer markers from the three germ layers after embryonic body differentiation. Moreover, our ESC shows a normal genotype and no mycoplasma contamination.

This protocol generates neural assembloids from human induced pluripotent stem cells (hiPSCs) to model human brain circuitry. It details the differentiation of excitatory-predominant hippocampal (Hc) and cortical (Cx) organoids and their fusion with inhibitory interneuron-predominant ganglionic eminence (GE) organoids. We then detail procedures for maintaining assembloids to promote interneuron migration and network integration, followed by functional assessment using two-photon calcium imaging to measure neuronal activity. For complete details on the use and execution of this protocol, please refer to McCrimmon et al.1.

To investigate the effect of irisin on the osteogenic differentiation of rat bone marrow mesenchymal stem cells (BMSCs) and bone regeneration in a femoral defect model. Rat BMSCs were treated with irisin in vitro, and osteogenic markers were analysed using reverse transcription quantitative polymerase chain reaction. In vivo, irisin-loaded composite scaffolds or blank scaffolds were implanted into rat femoral defects (n = 6 per group). Bone repair was assessed by micro-computed tomography, haematoxylin and eosin staining, and immunohistochemistry after 8 weeks. Irisin treatment significantly enhanced calcium nodule formation, as evidenced by increased staining intensity, size and number in both Alizarin Red and von Kossa assays. At the molecular level, Sost expression was markedly downregulated, whereas ALP, Lrp5, BMP2 and Smad1 were significantly upregulated at both mRNA levels (all p < 0.001). In vivo, rats implanted with irisin-composite scaffolds showed substantially improved bone repair, with higher bone volume fraction, bone mineral density and cortical bone volume compared with controls. Irisin promotes the osteogenic differentiation of rat BMSCs and accelerates bone regeneration in a critical-sized femoral defect model, potentially through suppression of Sost and subsequent activation of the Wnt and BMP signalling pathways. These findings highlight the therapeutic potential of irisin for bone tissue engineering applications.

Accurate and sensitive identification of cancer cells is of great importance for cancer diagnosis and prognosis. DNA circuits with high sensitivity have promise for detecting tumor-associated molecules, yet are constrained with limited tumor specificity due to the "on target, off cancer" effect. To address this issue, we propose a multi-pathway integrated DNA logic circuit (MDLC) system by integrating two AND-gate circuit modules for the highly specific and sensitive identification of cancer cells. Upon delivery into cancer cells, the MDLC system is synergistically activated by the three distinct tumor-specific biomarkers (O6-methylguanine-DNA methyltransferase, Apurinic/apyrimidinic endonuclease 1, and microRNA-21), thereby triggering a cascaded DNA circuit to generate amplified fluorescence signals. The MDLC system promises high specificity and broad applicability, which enables effective identification of three positive cancer cells (MCF-7, HepG2, and MDA-MB-231) from normal cells (MCF-10A). Further in vivo studies demonstrate the ability of this system to precisely recognize cancer cells and therefore excels in tumor imaging. Collectively, this work illustrates a simple and powerful strategy for developing DNA logic circuits, bringing new avenues for precise cancer diagnosis and biomedical applications.

Metanephric mesenchymal cells (MMCs) hold therapeutic potential for acute kidney injury (AKI), but their efficacy is limited, and the mechanisms underlying their action remain unclear. This study aimed to investigate whether catalpol-pretreated MMCs (MMCs-cata) could enhance the efficacy of AKI treatment by regulating key signaling pathways.

Psoriasis is a refractory immune-related disease. In recent years, it has been discovered that mesenchymal stem cells (MSCs) can be used as a new therapeutic approach for psoriasis, but their potential therapeutic mechanism remains unclear. This study aims to explore the role of MSCs in the treatment of psoriasis.

Invariant natural killer T (iNKT) cells, upon activation, exhibit antitumor roles by bridging innate and acquired immunity. To overcome the challenges in producing iNKT cells from patients with cancer, we previously developed allogeneic human induced pluripotent stem cell-derived iNKT (iPSC-iNKT) cells. However, the activation of iPSC-iNKT cells by glycolipid ligands remains a critical step for iNKT cell-mediated cancer therapy. To show the effect of iPSC-iNKT cell-mediated antitumor immunity, in this preclinical study, by taking advantage of a human immune cell-transplanted patient-derived xenograft model using human IL-7/15 knock-in NSG mice, we demonstrate that a combination of iPSC-iNKT cells and α-galactosylceramide-pulsed antigen-presenting cells (αGalCer/APC) induces robust antitumor effects. Single-cell analysis of tumor-infiltrating lymphocytes revealed that this combination therapy uniquely expanded tumor-reactive memory-phenotype CD4 and CD8 T cells. Taken together, upon activation by αGalCer/APC, iPSC-iNKT cells are capable of effectively inducing antitumor T cell immunity, making them a promising tool for generating personalized antitumor T cell immunity.

Cell competition is an evolutionarily conserved quality control mechanism that eliminates less-fit cells to ensure optimal tissue integrity during development, homeostasis, and regeneration. Beyond these physiological roles, recent evidence implicates a role for cell competition in disease, particularly in cancer, where it can function by either suppressing or promoting malignant progression. In this review, we provide an overview of the different molecular mechanisms that drive cell competition and their impact on cancer development and progression. We will evaluate the current state-of-the-art in vitro experimental systems that can be employed to study these processes. Ranging from classical 2D co-culture systems to advanced organoid and organ-on-chip platforms, these model systems collectively enhance our understanding of the complex cellular interactions that underlie the competitive differences between cells. By integrating insights from diverse model systems, we highlight how cell competition shapes tumor dynamics and discuss how this knowledge could inspire novel therapeutic strategies to prevent or control tumor growth.

Cornelia de Lange syndrome (CdLS) is a rare genetic disorder that affects almost any organ, including the central nervous system. It leads to a wide range of neurodevelopmental delays, and there are currently no available clinical treatments. CdLS is caused by pathogenic variants in one of the 7 genes coding for the cohesin complex, a multimeric structure responsible for sister chromatid cohesion, or for cohesin ring-interacting proteins. Additionally, altered regulation of molecular pathways during development, including the canonical WNT pathway, can cause CdLS malformations. In our study, we evaluated the positive effects of using lithium as an activator of the canonical WNT pathway to ameliorate neural CdLS phenotype. We have exploited accurate two-dimensional (2D) and three-dimensional (3D) human central nervous system in vitro models representing disease-related neurobiological phenotypes: induced pluripotent stem cells of human origin (hiPSCs) differentiated into neural precursors, neurons, and brain organoids (BOs). CdLS models demonstrate alterations in proliferation and differentiation capabilities when mimicking HDAC8 haploinsufficiency. Furthermore, RNA-seq analysis of BOs revealed that both neuronal differentiation and the WNT pathway are downregulated when treated with the HDAC8 inhibitor alone. Following lithium treatment, cells show an enhanced ability to differentiate into the neuronal lineage. Additionally, our working hypothesis is that a specific mechanism may exist that, by connecting lipid metabolism, canonical WNT pathway, and cell death, results in typical CdLS neurodevelopmental deficits.

Lung cancer, being the top cause of global cancer-related mortality, calls for effective treatments. RN7SK (7SK) is a long non-coding RNA (lncRNA) that plays a significant role in the regulation of gene transcription and thereby controls essential cellular activities. Limited evidence supports the anticancer potential of 7SK, and its suppressive effects have not been tested against lung cancer. This study explored the anticancer effects of RN7SK (7SK), a long non-coding RNA known to regulate gene transcription, through exosome-mediated delivery in lung cancer cells. Treatment with exosome-loaded 7SK (Exo-7SK) significantly elevated 7SK levels in non-small-cell lung cancer cells and suppressed key cancer traits. Exo-7SK reduced cell viability and proliferation, promoted apoptosis, and inhibited migration and invasion by shifting gene expression away from epithelial-mesenchymal transition. It also impaired spheroid formation and reduced spheroid dispersion and viability in 3D microfluidic cultures. In conclusion, our findings highlight the cancer-suppressive potential of exosome-mediated 7SK delivery against lung cancer, demonstrating significant efficacy in both 2D and 3D culture models. These observations warrant further confirmation in future studies employing advanced designs and clinically relevant models.