Current approaches for bone repair predominantly target localized delivery of growth factors that are aimed at the coupling of angiogenesis and osteogenesis. However, delayed revascularization and regeneration of critical-sized bone defects are still challenging. In this study, we engineer an ossification center-like organoid (OCO) that consist of an inner-core bone morphogenetic and neurotrophic spheroid generated via MSCs-loaded 3D printing, alongside the interstitially distributed outer-shell proangiogenic neurotrophic phase. Our results demonstrate that collective implantation of OCOs achieves rapid bone bridging with successive OC-like bone ossicles formation across the bone defect in a "divide-and-conquer" way. Single-cell RNA sequencing analysis unveils a developmentally mimicking stem cell community that dominated with Krt8+ skeletal stem cells (SSCs) is uniquely recruited by the pro-regenerative in-situ organoid fusion and maturation. Particularly noteworthy is the specific expansion of Krt8+ SSCs concomitant with the simultaneous reduction of Has1+ migratory fibroblasts (MFs) post-OCO implantation. Furthermore, cross-species comparisons employing machine learning reveal high resemblance of the relative Krt8+ SSCs/Has1+ MFs composition in bone regeneration with that in public data from developmental bone tissues. Our findings advocate an approach akin to "divide-and-conquer" utilizing engineered OC-like organoids for prompt regeneration of large-sized bone defects.

Peripheral neuropathy is a common complication in diabetes, affecting around 50% of the diabetic population. Co-occurrence of diabetic peripheral neuropathy (DPN) and diabetic bone disease has led to the hypothesis that DPN influences bone metabolism, although little experimental evidence has yet supported this premise. To investigate, mice were fed a high-fat diet (HFD) followed by phenotyping of skeletal-innervating neurons and bone architectural parameters. Results showed that HFD feeding resulted in a marked decrease in skeletal innervation (69%-41% reduction in Beta-III-Tubulin-stained nerves, 38% reduction in CGRP-stained nerves in long bone periosteum). These changes in skeletal innervation were associated with significant alterations in bone mass and in cortical and trabecular bone microarchitecture of long bones. Single-cell RNA sequencing (scRNA-Seq) of sensory neurons and bone tissue was next utilized to reconstruct potential nerve-to-bone signaling interactions, including implication of sensory nerve-derived neurotrophins (Bdnf), neuropeptides (Gal, Calca and Calcb), and other morphogens (Vegfa, Pdgfa, and Angpt2). Moreover, scRNA-Seq identified marked shifts in periosteal cell transcriptional changes within HFD-fed conditions, including a reduction in cell proliferation, an increase in adipogenic differentiation markers, and reductions in WNT, TGFβ, and MAPK signaling activity. When isolated, periosteal cells from HFD-fed mice showed deficits in proliferative and osteogenic differentiation potential. Moreover, these cellular changes in proliferation and differentiation capacity were restored by treatment of HFD-exposed periosteal cells to sensory neuron-conditioned medium. In summary, HFD modeling of type 2 diabetes results in skeletal polyneuropathy. Moreover, the combination of multi-tissue scRNA-Seq and isolated in vitro studies strengthen the case for altered nerve-to-bone signaling in diabetic bone disease.

Hematopoietic stem cells (HSC) with multilineage potential are critical for T cell reconstitution after allogeneic hematopoietic cell transplantation (allo-HCT). The Kitlo HSC subset is enriched for multipotential precursors, but their T cell potential remains poorly characterized. Using a preclinical allo-HCT mouse model, we demonstrate that Kitlo HSCs provide superior thymic recovery and T cell reconstitution, resulting in improved immune responses to post-transplant infection. Kitlo HSCs with augmented bone marrow (BM) lymphopoiesis mitigate age-associated thymic alterations and enhance T cell recovery in middle-aged mice. Mechanistically, chromatin profiling reveals Kitlo HSCs exhibiting higher activity of lymphoid-specifying transcription factors, such as, ZBTB1. Zbtb1 deletion diminishes HSC engraftment and T cell potential; by contrast, reinstating Zbtb1 in megakaryocytic-biased Kithi HSCs rescues hematopoietic engraftment and T cell potential in vitro and in vivo. Furthermore, age-associated decline in Kitlo HSCs is associated with diminished T lymphopoietic potential in aged BM precursors; meanwhile, Kitlo HSCs in aged mice maintain enhanced lymphoid potential, but their per-cell capacity is diminished. Lastly, we observe an analogous human BM KITlo HSC subset with enhanced lymphoid potential. Our results thus uncover an age-related epigenetic regulation of lymphoid-competent Kitlo HSCs for T cell reconstitution.

Duplication 15q (dup15q) syndrome is a leading genetic cause of autism spectrum disorder, offering a key model for studying autism-related mechanisms. Using single-cell and single-nucleus RNA sequencing of cortical organoids from dup15q patient-derived iPSCs and post-mortem brain samples, we identify increased glycolysis, disrupted layer-specific marker expression, and aberrant morphology in deep-layer neurons during fetal-stage organoid development. In adolescent-adult postmortem brains, upper-layer neurons exhibit heightened transcriptional burden related to synaptic signaling, a pattern shared with idiopathic autism. Using spatial transcriptomics, we confirm these cell-type-specific disruptions in brain tissue. By gene co-expression network analysis, we reveal disease-associated modules that are well preserved between postmortem and organoid samples, suggesting metabolic dysregulation that may lead to altered neuron projection, synaptic dysfunction, and neuron hyperexcitability in dup15q syndrome.

Cell-based therapies are promising for treating solid tumors, but challenges like tumor heterogeneity, antigen escape, and immunosuppressive microenvironments hinder their efficacy. Inducible suicide gene systems, often viewed solely as safety mechanisms, offer an underappreciated opportunity to enhance cellular therapies. These systems, triggered by various mechanisms (prodrugs, ligands, antibodies, or small molecules), enable controlled elimination of therapeutic cells. Recent developments demonstrate that this controlled cell death, especially when inducing immunogenic cell death (ICD), can kill even resistant tumor cells and reshape the tumor microenvironment (TME) from suppressive to stimulatory. This review highlights the transformative potential of integrating these suicide systems into cell therapies, overcoming key limitations, and amplifying antitumor responses while ensuring safety.

Liver diseases pose a major global health burden, and progress in understanding liver pathophysiology and therapeutic development is hampered by the lack of predictive human models. Hepatic organoids (i.e 3D in vitro structures derived from primary, progenitor, or pluripotent stem cells) offer a physiologically relevant alternative to traditional 2D cultures and animal models. This review provides a comprehensive overview of hepatic organoid systems and their translational potential.

Curcumae Rhizoma, a traditional Chinese medicine derived from the dried rhizome of Curcuma species, has long been used to treat liver-related disorders. In the classical medical text Taiping Shenghui Fang, it was recorded for the treatment of hepatocellular carcinoma (HCC) characterized by Qi stagnation and blood stasis. Its major chemical constituents include curcumin and Zedoary Turmeric Oil (ZTO). While curcumin has been extensively studied for its pharmacological activities, the therapeutic potential and mechanism of ZTO in HCC remain insufficiently understood.

Switching from a contractile to a synthetic phenotype of pulmonary arterial smooth muscle cells (PASMCs) is known to play a crucial role in pulmonary arterial hypertension (PAH). We investigated how hypoxia and mechanical stress mediate the phenotypic switching of PASMCs.

Glioblastoma multiforme (GBM) is a highly aggressive brain cancer marked by aggressive growth and therapeutic resistance. A growing body of research highlights substantial spatial heterogeneity within GBM tumors as a critical factor contributing to treatment failure. Advanced molecular techniques, including third-generation genomics, high-resolution metabolomics, and single-cell and spatial transcriptomics, have illuminated the detailed genetic and epigenetic landscape, revealing a complex interplay of molecular modifications. A key determinant of spatial heterogeneity is differential perfusion, which leads to the formation of distinct microenvironmental niches characterized by varying oxygen, nutrient, and growth factor availability. This ITH impacts not only GBM cancer cells but also the entire tumor microenvironment, including immune and other stromal cells. Interactions between cancer cells and the surrounding stroma significantly modulate immune surveillance, frequently promoting tumor malignancy. Perfusion further dictates the plasticity in GBM, enabling their transformation into the aggressive mesenchymal subtype. This review examines how such perfusion-driven differences affect GBM intra-tumoral heterogeneity, focussing on the aspects of immune regulation, and treatment resistance. We discuss emerging therapeutic strategies that target perfusion-induced heterogeneity, including anti-angiogenic and immunotherapeutic approaches. Further, the review emphasizes the importance of the tumor microenvironment and highlights the complex interplay of factors driving GBM progression, paving the way for more effective and personalized treatment strategies aimed at enhancing patient survival.

Programmed death-ligand 1 (PD-L1), a critical immune checkpoint ligand, is overexpressed in several malignancies. The newly identified protein posttranslational modification lactylation, occurring on lysine residues, is extensively involved in various biological processes. However, PD-L1 lactylation and its role in tumorigenesis remain unclear. In this study, we discover that lactylation of PD-L1 suppresses liver cancer growth by inhibiting cholesterol synthesis. Acetyltransferase E1A-binding protein p300 (p300) catalyzes the lactylation of PD-L1 at the lysine 189 residue (K189). Histone deacetylase 2-dependent delactylation of PD-L1 K189 promotes vimentin-mediated nuclear translocation of PD-L1. Functionally, PD-L1 K189 delactylation accelerates liver cancer growth both in vitro and in vivo by facilitating cholesterol production. Clinically, an antibody against PD-L1 K189 lactylation reveals that PD-L1 delactylation is positively associated with the progression of liver cancer histological grade. Mechanistically, PD-L1 K189 delactylation upregulates SQLE, a rate-limiting enzyme in cholesterol biosynthesis, by increasing SQLE transcription activity via the transcription factor YY1. Therefore, our findings demonstrate that lactylation-dependent regulation of PD-L1 promotes liver cancer growth.

Gene discovery studies in individuals with diabetes diagnosed within 6 months of life (neonatal diabetes, NDM) can provide unique insights into the development and function of human pancreatic beta-cells. We describe the identification of homozygous PAX4 loss-of-function variants in 2 unrelated individuals with NDM: a p.(Arg126*) stop-gain variant and a c.-352_104del deletion affecting the first 4 PAX4 exons. We confirmed the p.(Arg126*) variant causes nonsense mediated decay in CRISPR-edited human induced pluripotent stem cell (iPSC)-derived pancreatic endoderm cells. Integrated analysis of CUT&RUN and RNA-sequencing in PAX4-depleted islet cell models identified genes directly regulated by PAX4 involved in both pancreatic islet development and glucose-stimulated insulin secretion. Both probands had transient NDM which remitted in early infancy but relapsed at the ages of 2.4 and 6.7 years, demonstrating that in contrast to mouse models, PAX4 is not essential for the development of human pancreatic beta-cells.

A new paradigm for drug development and patient therapeutic strategies is required, especially for complex, heterogeneous diseases including metabolic dysfunction-associated steatotic liver disease (MASLD). Heterogeneity in MASLD patients is driven by genetics, various co-morbidities, gut microbiota composition, lifestyle, environment and demographics that produce multiple patient disease presentations and outcomes. Existing drug development methods have had limited success for complex, heterogeneous diseases like MASLD where only a fraction of patients respond to specific treatments, prediction of a therapeutic response is not presently possible and the cost of the new classes of drugs are high. However, it is now possible to generate patient digital twins (PDTs) that are computational models of patients using clinomics and other "omics" data collected from patients to make various predictions including responses to therapeutics. PDTs are then integrated with patient biomimetic twins (PBTs) that are patient-derived organoids or induced pluripotent stem cells that are then differentiated into the optimal number of organ-specific cells to produce organ experimental models. The PBTs mimic key aspects of the patient's pathophysiology, enabling predictions to be tested. In conclusion: integration of patient digital twins and patient biomimetic twins has the potential to create a powerful precision medicine platform, yet there are challenges.

Aging is characterized by the progressive deterioration of cell and tissue functions. The liver, which regulates metabolic homeostasis, detoxification, and immune responses, undergoes structural and functional changes with age. These include increasing genomic instability, telomere attrition, epigenetic alterations, loss of proteostasis, deregulated nutrient-sensing and intracellular communication, mitochondrial dysfunction, cell senescence, stem cell exhaustion, chronic inflammation, disabled macroautophagy, and dysbiosis. These alterations contribute to hepatocyte dysfunction, impaired regenerative responses, and fibrosis risk, which all exacerbate existing liver diseases. Senescence involves irreversible cell cycle arrest resulting in an inflammatory, senescence-associated secretory cell phenotype. Senescent hepatocytes, liver sinusoidal endothelial cells, hepatic stellate cells, and Kupffer cells accumulate in the aged liver, creating an inflammatory and fibrotic microenvironment that promotes tumorigenesis. As the burden of aging-related liver disease increases, therapeutic strategies targeting hepatic senescence have gained attention. We review these, along with the mechanisms and pathogenic effects of liver aging.

Follicular lymphoma (FL) is characterized by the expansion of neoplastic follicle structures and is suggested to have a distinctive form of T cell immunity. However, the heterogeneity and role of follicular T cells beyond T follicular helper (TFH) cells remain largely unexplored in FL. Here, we performed multi-omics analyses of follicular T cells in FL leveraging pan-cancer single-cell mapping, spatially resolved single-cell transcriptomics and multiplex protein profiling, and functional characterization. We identified transcriptionally and spatially distinct non-TFH follicular T cell subsets that expand in FL. These subsets exhibit enhanced anti-tumorigenic properties and form unique spatial niches. Their phenotypes were replicated under interleukin-21-predominant conditions, revealing discrete self-regulatory cellular ecosystems that generate these subsets and may underlie FL clinical behaviors. Furthermore, these subsets robustly stratify FL prognoses, independently of existing prognostic markers. Our findings highlight previously unrecognized immunity that could advance our understanding of lymphoma and improve patient management.

The ineffective hematopoiesis of myelodysplastic syndrome (MDS) suggests that hematopoietic stem and progenitor cells (HSPCs) are defective. Here, we demonstrate that NUP98::HOXD13 (NHD13) MDS mice have significantly decreased functional HSPCs. Moreover, in contrast to wild-type (WT) bone marrow (BM), lineage-positive (Lin+) BM cells from NHD13 mice have self-renewal potential. Specific subsets of NHD13 Lin+ cells that express B220 and Kit antigens were able to self-renew and generate MDS in WT recipients. Although this unique B220+Kit+ phenotype could be found in WT as well as NHD13 BM, the population was markedly increased in NHD13 BM. Further characterization using Mac1 and Gr1 markers revealed that both Mac1+Gr1+B220+Kit+ and Mac1-Gr1- B220+Kit+ populations showed self-renewal and led to an MDS phenotype in WT recipients. Taken together, these findings demonstrate that as normal hematopoiesis derived from typical HSPCs decreases in NHD13 mice, committed hematopoietic progenitor cells proliferate, self-renew, and initiate MDS.

Olfactory neurogenesis occurs throughout the lives of vertebrates, including in humans, and relies on the continuous differentiation and integration of neurons into a complex network. How progenitor cells convert fluctuations in cell-cell signaling into streamlined fate decisions over both space and time is poorly understood. Here, we track multicellular dynamics in the zebrafish olfactory epithelium, undertake targeted perturbations, and find that neurogenesis is driven by mutual antagonism between Notch signaling and insulinoma-associated 1a (Insm1a) that is responsive to inter-organ retinoic acid signaling. Single-cell analysis reveals that olfactory neurons emerge from transient groups of cells termed cellular neighborhoods. Stochastic modeling shows that neighborhood self-assembly is maintained by a tightly regulated bistable toggle switch. Differentiating cells migrate apically in response to brain-derived neurotrophic factor (BDNF) to take up residence as mature sensory neurons. Cumulatively, these findings reveal how stochastic signaling networks spatiotemporally regulate a balance between progenitors and derivatives, driving sustained neurogenesis in an intricate organ system.

Accumulating evidence indicates that maturation limits cardiomyocyte proliferation. We expand on that theory by co-culturing human induced pluripotent stem cell (hiPSC)-cardiomyocytes (CM) with epicardial cells (EPCs) and epicardial-derived cells in both 2D co-cultures and 3D engineered heart tissues (EHTs). In 2D co-cultures, the percentage of proliferating CM increased in parallel with stark electrophysiologic improvements. Single-cell transcriptomics revealed a significant shift in the bulk CM population of the epicardial-CM co-cultures as characterized by more fetal-like myofilament isoforms but with enhanced pathways associated with electrochemical maturation. The 3D-EHTs containing EPCs showed more limited proliferation but a similar improvement in CM electrophysiologic function. Next, epicardial-derived fibroblasts (EPD-FBs) were added to the EHTs containing EPCs, and we observed significant myofilament maturation and increased force generation. Our results suggest that some aspects of CM maturation (i.e., electrochemical) can occur when proliferation rates are relatively high, and that sarcomere-associated mechanical maturation occurs at later developmental stages when proliferation has largely ceased.

Multiciliated ependymal and neural stem cells are key cell populations of the subventricular zone. Recent findings revealed that at least a subpopulation of radial glial cells during embryogenesis can be bipotent and produce both neural stem cells and ependymal cells. The balance between these cell populations is orchestrated by Geminin superfamily, ensuring optimal niche function. However, whether cell fate decisions are definitive or dynamic and whether potential regional differences exist remain elusive. In this review, we delve into the shared origins of different subventricular zone cell populations, and we explore the potential interplay among them. Moreover, we compile evidence on the de-differentiation capacity of ependymal cells and their controversial neural stem cell function under specific conditions, with emphasis on the possible implication of a rare population of biciliated (E2) ependymal cells. Understanding the mechanisms regulating cell fate decisions may unravel ependymal cells' therapeutic potential in therapies targeting various human diseases.

Understanding epithelial stem cell differentiation and morphogenesis during breast tissue development is essential, as disruption in these processes underlie breast cancer formation. We used a next-generation single-cell-derived organoid model to investigate how individual stem cells give rise to complex tissue. We show that discoidin domain receptor 1 (DDR1) inhibition traps cells in a bipotent state, blocking alveolar morphogenesis and luminal cell expansion, which is necessary for complex epithelium formation. Disrupting RUNX1 function produced nearly identical phenotypes, underscoring its critical role downstream of DDR1. Mechanistically, DDR1 affects the interaction and expression of RUNX1 and its cofactor core binding factor beta (CBFβ), thereby regulating its activity. Mutational analyses in breast cancer patients reveal frequent alterations in the DDR1-RUNX1 signaling axis, particularly co-occurring mutations. Together, these findings uncover DDR1-RUNX1 as a central signaling pathway driving breast epithelial differentiation, whose dysregulation may contribute fundamentally to breast cancer pathogenesis.

Common forms of Alzheimer's disease (AD) are complex and polygenic. We have created a research resource that seeks to capture the extremes of polygenic risk in a collection of human induced pluripotent stem cell (iPSC) lines from over 100 donors: the IPMAR Resource (iPSC Platform to Model Alzheimer's Disease Risk). Donors were selected from a large UK cohort of 6,000+ research-diagnosed early or late-onset AD cases and elderly cognitively healthy controls, many of whom have lived through the age of risk for disease development (>85 years). We include iPSC with extremes of global AD polygenic risk (high-risk late-onset AD: 34; high-risk early-onset AD: 29; low-risk control: 27) as well as those reflecting complement pathway-specific genetic risk (high-risk AD: 9; low-risk controls: 10). All iPSC have associated clinical, longitudinal, and genetic datasets and will be available through collaboration or from cell (EBiSC) and data (DPUK) repositories.