Progenitor Cells Research Articles

PDGF-BB Deficiency in the Blood Serum from Aplastic Anemia Patients Affects Bone Marrow-Derived Multipotent Mesenchymal Stromal Cells

Aplastic anemia (AA) is characterized by bone marrow (BM) aplasia and pancytopenia. BM stromal microenvironment is closely intertwined with hematopoietic cells by reciprocal regulation. It is still unclear how hematopoietic deficiency affects the bone marrow stroma of the AA patients. Multipotent mesenchymal stromal cells (MMSCs) are the progenitors of stromal cells. In vitro, proliferation rate of MMSCs of AA patients is decreased compared to those of healthy donors. This may be explained by the influence of pathological environmental condition in the patients’ BM. The aim of the study was to compare the effect of AA patients’ sera on healthy donor MMSCs to healthy donors’ sera and to elucidate the nature of their difference. Proliferation test showed 3-fold decrease in number of MMSCs after incubation in medium supplemented with AA patients’ sera compared to donors’ serum samples. The degree of this effect correlated with the severity of thrombocytopenia in patients. The decrease in cell number was not associated with cell death, as the number of apoptotic cells defined by flow cytometry did not differ between the groups. ELISA revealed a decreased level of PDGF-BB in the patients’ sera compared to donors’ serum samples (69 ± 5 pg/mL vs 112 ± 21 pg/mL, respectively). The addition of recombinant PDGF-BB or healthy donor’s platelet lysate to the culture medium supplemented with AA patients’ serum restored its ability to support MMSCs growth. Thus, PDGF-BB deficiency is one of the environmental factors causing MMSCs damage in AA patients.

A small molecule K-3 promotes PDX1 expression and potentiates the differentiation of pluripotent stem cells into insulin-producing pancreatic β cells

Abstract Insulin-producing pancreatic β-like cells derived from human pluripotent stem cells (PSCs) are anticipated as a novel cell source for cell replacement therapy for diabetes patients. Here, we describe the identification of small molecule compounds that promote the differentiation of the PSCs into insulin-producing cells by high throughput screening with a chemical library composed of 55,000 compounds. The initial hit compound K-1 and one derivative K-3 increased the proportion of PSC-derived insulin-positive endocrine cells and their glucose-stimulated insulin secretory (GSIS) functions. K-3 preferentially acts on stage 3 pancreatic progenitor cells and increases the population expressing high levels of PDX1. As a result, the ratios of the PSC-derived PDX1 / NKX6.1 double-positive endocrine progenitor and INS / NKX6.1 double-positive mono-hormonal endocrine cells were increased. K-3 enhances the expression of functional pancreatic β cell markers and affects biological processes concerning organ development. K-3 also increased the yield of endocrine cells at the end of stage 5. The novel compound is a beneficial new tool for efficiently generating PSC-derived insulin-producing cells with high functionality and differentiation efficiency.

Abstract A007: Emergence of metastasis-initiating cells during lung adenocarcinoma progression

Abstract Metastatic lung adenocarcinoma (LUAD) is responsible for the most cancer-related deaths worldwide, and only a subset of cells is capable of metastasizing. Our lab has previously shown that these metastasis-initiating cells express the transmembrane glycoprotein called L1CAM and exhibit characteristics of a fetal lung developmental continuum, ranging from early progenitor states to more differentiated states. The exact signal by which progenitor cells with high metastatic potentials are generated remains unknown. Here, we demonstrate that L1CAM facilitates the signaling cascades necessary to initiate and maintain metastasis-initiating cells in LUAD. Mechanistically, L1CAM interacts with the planar cell polarity (PCP) complex and activates the transcription factor c-Jun. Activated c-Jun, as part of the AP-1 complex, binds to enhancer sites of progenitor genes and promotes the emergence of a metastasis-capable progenitor state. Pharmacological ablation of these metastatic progenitor cells by a cytotoxic L1CAM antibody effectively eliminated LUAD metastasis in vivo. Our findings identify a novel L1CAM-mediated mechanism that maintains a progenitor state essential for metastasis and can be targeted therapeutically. Citation Format: Jin Suk Park, Yasemin Kaygusuz, Carson Kenum, Mohamed I. Gatie, Lan He, Roshan Sharma, Ronan Chaligné, Abdul G. Khan, Ivo C. Lorenz, Paul J. Balderes, Tuomas Tammela, Anna-Katerina Hadjantonakis, Karuna Ganesh, Charles M. Rudin, Joan Massagué. Emergence of metastasis-initiating cells during lung adenocarcinoma progression [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr A007.

The Mechanistic Link Between Tau-Driven Proteotoxic Stress and Cellular Senescence in Alzheimer’s Disease

In Alzheimer’s disease (AD), tau dissociates from microtubules (MTs) due to hyperphosphorylation and misfolding. It is degraded by various mechanisms, including the 20S proteasome, chaperone-mediated autophagy (CMA), 26S proteasome, macroautophagy, and aggrephagy. Neurofibrillary tangles (NFTs) form upon the impairment of aggrephagy, and eventually, the ubiquitin chaperone valosin-containing protein (VCP) and heat shock 70 kDa protein (HSP70) are recruited to the sites of NFTs for the extraction of tau for the ubiquitin–proteasome system (UPS)-mediated degradation. However, the impairment of tau degradation in neurons allows tau to be secreted into the extracellular space. Secreted tau can be monomers, oligomers, and paired helical filaments (PHFs), which are seeding competent pathological tau that can be endocytosed/phagocytosed by healthy neurons, microglia, astrocytes, oligodendrocyte progenitor cells (OPCs), and oligodendrocytes, often causing proteotoxic stress and eventually triggers senescence. Senescent cells secrete various senescence-associated secretory phenotype (SASP) factors, which trigger cellular atrophy, causing decreased brain volume in human AD. However, the molecular mechanisms of proteotoxic stress and cellular senescence are not entirely understood and are an emerging area of research. Therefore, this comprehensive review summarizes pertinent studies that provided evidence for the sequential tau degradation, failure, and the mechanistic link between tau-driven proteotoxic stress and cellular senescence in AD.

Utility of acid citrate dextrose‐acidification for platelet volume reduction protocols in the transfusion service

AbstractBackgroundVolume‐reduced platelets can minimize circulatory overload, allergic transfusion reactions, or out‐of‐group plasma infusion. Our center adopted a volume reduction protocol that includes acidification with acid citrate dextrose solution A (ACD‐A) before centrifugation and without any rest period prior to resuspension allowing a better turnaround time for platelet issue.Study Design and MethodsThis report compares corrected count increments (CCIs) from full‐volume and ACD‐A acidified volume‐reduced human platelets in a retrospective study at a single hospital and in a mouse model.ResultsAt a pediatric tertiary care hospital, 530 patients received conventional apheresis platelets during the 20‐month study period. Among all patients, the expected 4‐h mean CCI was 9.8 (95% CI: 8.7, 10.9) for full‐volume platelets, and 8.8 (95% CI: 7.3, 10.6) for ACD‐acidified volume‐reduced platelets (p = .29). A statistically significant difference (p = .01) was identified in the expected 24‐h mean CCI: 6.3 (95% CI: 5.5–7.0) with full‐volume platelet, 4.7 (95% CI: 3.6–6.0) with ACD‐acidified volume‐reduced platelet. Limiting CCI calculations to patients with Hematology/Oncology/Hematopoietic Progenitor Cell Transplant diagnosis (n = 296, 56%) indicated a statistically significant difference in both 4‐ and 24‐h predicted CCIs, showing lower CCIs in ACD‐acidified volume‐reduced platelet, although these were still similar to the CCIs observed in all patients and considered to be clinically acceptable responses similar to other volume reduction protocols. The recovery of count‐adjusted, volume‐reduced platelets was significantly lower in mice, suggesting a procedure‐related defect.DiscussionACD‐A acidification of platelets before volume reduction decreases turnaround time for platelet issue and provides clinically allowable 4‐h and 24‐h platelet increments.

Abstract A032: Uncovering the impact of current therapies on LUAD dormant metastasis

Abstract Despite recent therapeutic advances, Lung Adenocarcinoma (LUAD) remains a leading cause of mortality worldwide, with most deaths attributed to metastasis. Metastasis is mediated by disseminated metastasis initiating cells (MICs) that can persist in a dormant state in distant sites for months to years and ultimately drive metastatic relapse, which is clinically incurable. Adjuvant therapy seeks to prevent cancer relapse, however, efforts to improve adjuvant treatments are hindered by an insufficient understanding of the molecular mechanisms supporting the long-term viability of dormant MICs. We have established that primitive LUAD cells characterized by the spontaneous expression of the pluripotency transcription factor SOX2 can enter an immune-evasive dormant state with downregulation of immune response mediators (STING, NK ligands, MHC-I) to evade immune surveillance during dormancy. Now, we hypothesize that dormant and immune evasive progenitor MICs may also escape therapies targeting growth pathways. We have generated latency competent cells (LCCs) that can remain in a latent state for months by isolating progenitor cells from the Kras LSL-G12D/+ ;Trp53 flox/flox mouse model and screening their metastatic potential and progenitor state in the B6 background. LCCs entering a slow-cycling state in response to dormancy signals remain sensitive to targeted KRAS inhibitors but contain a resistant subpopulation that survives. To specifically characterize the progenitor state in these LCC populations, we have developed and validated two strategies to fluorescently mark SOX2 expression and activity, respectively. As speculated, the resistant population is mostly composed of SOX2+ progenitor LCCs after treatment with KRAS inhibitor under dormancy signaling. Furthermore, induced-dormancy also drives an accumulation of cells in the SOX2+ progenitor state in these LCC-rich populations. Ongoing studies are focused on delineating the emergence of the SOX2+ progenitor state in LCCs during dormancy and the biological mechanisms driving their persistence and resistance to targeted therapies. We aim to define (1) the dependence of dormant LCCs on oncogenic signaling, the (2) effect of targeted therapy on MICs, and (3) strategies to overcome potential resistance mechanisms in dormant MICs. Our ultimate goal is to delineate strategies to clear latent MICs and prevent LUAD recurrence. Citation Format: Inês Godet, Zhenghan Wang, Joan Massagué. Uncovering the impact of current therapies on LUAD dormant metastasis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr A032.

Single cell and spatial transcriptomics highlight the interaction of club-like cells with immunosuppressive myeloid cells in prostate cancer.

Prostate cancer treatment resistance is a significant challenge facing the field. Genomic and transcriptomic profiling have partially elucidated the mechanisms through which cancer cells escape treatment, but their relation toward the tumor microenvironment (TME) remains elusive. Here we present a comprehensive transcriptomic landscape of the prostate TME at multiple points in the standard treatment timeline employing single-cell RNA-sequencing and spatial transcriptomics data from 120 patients. We identify club-like cells as a key epithelial cell subtype that acts as an interface between the prostate and the immune system. Tissue areas enriched with club-like cells have depleted androgen signaling and upregulated expression of luminal progenitor cell markers. Club-like cells display a senescence-associated secretory phenotype and their presence is linked to increased polymorphonuclear myeloid-derived suppressor cell (PMN-MDSC) activity. Our results indicate that club-like cells are associated with myeloid inflammation previously linked to androgen deprivation therapy resistance, providing a rationale for their therapeutic targeting.

SMPD4-mediated sphingolipid metabolism regulates brain and primary cilia development.

Genetic variants in multiple sphingolipid biosynthesis genes cause human brain disorders. A recent study looked at people from 12 unrelated families with variants in the gene SMPD4, a neutral sphingomyelinase that metabolizes sphingomyelin into ceramide at an early stage of the biosynthesis pathway. These individuals have severe developmental brain malformations, including microcephaly and cerebellar hypoplasia. The disease mechanism of SMPD4 was not known and so we pursued a new mouse model. We hypothesized that the role of SMPD4 in producing ceramide is important for making primary cilia, a crucial organelle mediating cellular signaling. We found that the mouse model has cerebellar hypoplasia due to failure of Purkinje cell development. Human induced pluripotent stem cells lacking SMPD4 exhibit neural progenitor cell death and have shortened primary cilia, which is rescued by adding exogenous ceramide. SMPD4 production of ceramide is crucial for human brain development.

Extracellular vesicles derived from endothelial progenitor cells modified by Houshiheisan promote angiogenesis and attenuate cerebral ischemic injury via miR-126/PIK3R2.

Angiogenesis following cerebral ischemia is crucial for restoring blood supply to the ischemic region. Extracellular vesicles (EVs) derived from endothelial progenitor cells (EPCs) offer potential therapeutic benefits in the treatment of cerebral ischemia. Houshiheisan (HSHS) has been shown to improve clinical outcomes in ischemic stroke patients, reduce cerebral ischemic damage in rats, and protect endothelial cells. However, the potential effects of HSHS-modified EPC-derived EVs (EVsHSHS) for cerebral ischemia remain unexplored. This study investigated the impact of EVsHSHS on angiogenesis using rats with permanent middle cerebral artery occlusion (pMCAO) and brain microvascular endothelial cells (BMECs) subjected to oxygen-glucose deprivation (OGD). Results demonstrated that EVsHSHS promoted the proliferation, migration, and tube formation of BMECs in vitro. In vivo, high doses of EVsHSHS exhibited better performance than equivalent doses of unmodified EPC-derived EVs in reducing cerebral infarction volume, improving cortical blood perfusion, decreasing neurological deficit scores, and increasing cortical microvessel density at day 7 post-modeling. The pro-angiogenic effects of EVsHSHS following cerebral ischemia were associated with the regulation of miR-126 and the PIK3R2/PI3K/AKT pathway.

Modeling extrahepatic hepatitis E virus infection in induced human primary neurons

Hepatitis E virus (HEV) infections are one of the most common causes of acute viral hepatitis, annually causing over 3 million symptomatic cases and 70,000 deaths worldwide. Historically, HEV was described as a hepatotropic virus, but has recently been associated with various extrahepatic manifestations including neurological disorders such as Guillain-Barré syndrome and neuralgic amyotrophy. However, the underlying pathogenesis of these neurological diseases remains largely unknown. The aim of this study was to investigate extrahepatic HEV manifestations in a neuronal model system using human-induced primary neurons (iPNs). Renal epithelial cells from human urine were reprogrammed to induced pluripotent stem cells to generate neuronal progenitor cells, which were subsequently differentiated into iPNs over 21 d. These iPNs supported HEV infection preferentially in neurite-bearing cells. Transcriptional profiling of the neuronal development process as well as viral infection dynamics in iPNs uncovered a lack of antiviral innate immune responses to HEV infection with only an intrinsic expression of distinct interferon-regulated genes and signaling molecules. Viral open reading frame 2 encoded capsid protein could be visualized by volumetric three-dimensional reconstitution within the neurites, which were reduced in length in an HEV inoculation-dependent manner. In conclusion, this neuron-derived human model system provides a powerful tool for studying extrahepatic manifestations of HEV infection. It further indicates a potential mechanism of pathogenesis driven by the interaction between host and viral factors.

TAD-dependent sub-TAD is required for enhancer-promoter interaction enabling the β-globin transcription.

Topologically associating domains (TADs) are chromatin domains in the eukaryotic genome. TADs often comprise several sub-TADs. The boundaries of TADs and sub-TADs are enriched in CTCF, an architectural protein. Deletion of CTCF-binding motifs at one boundary disrupts the domains, often resulting in a transcriptional decrease in genes inside the domains. However, it is not clear how TAD and sub-TAD affect each other in the domain formation. Unaffected gene transcription was observed in the β-globin locus when one boundary of TAD or sub-TAD was destroyed. Here, we disrupted β-globin TAD and sub-TAD by deleting CTCF motifs at both boundaries in MEL/ch11 cells. Disruption of TAD impaired sub-TAD, but sub-TAD disruption did not affect TAD. Both TAD and sub-TAD disruption compromised the β-globin transcription, accompanied by the loss of enhancer-promoter interactions. However, histone H3 occupancy and H3K27ac were largely maintained across the β-globin locus. Genome-wide analysis showed that putative enhancer-promoter interactions and gene transcription were decreased by the disruption of CTCF-mediated topological domains in neural progenitor cells. Collectively, our results indicate that there is unequal relationship between TAD and sub-TAD formation. TAD is likely not sufficient for gene transcription, and, therefore, sub-TAD appears to be required. TAD-dependently formed sub-TADs are considered to provide chromatin environments for enhancer-promoter interactions enabling gene transcription.

Rapid Disease Progression of Myelodysplastic Syndrome is Reflected in Transcriptomic and Functional Abnormalities of Bone Marrow MSCs.

Bone marrow (BM) mesenchymal stromal cells (MSCs) are important regulators of hematopoietic stem and progenitor cells (HSPCs). When transformed to a dysplastic phenotype, MSCs contribute to hematopoietic diseases such as myelodysplastic syndromes (MDS), but it remains unclear if there are specific properties in MDS-MSCs that contribute to the disease course. To understand this, we investigated MDS-MSCs from fast (MDSfast) vs slow (MDSslow) progressing disease groups and discovered differences between these groups. MDSfast-MSCs secrete more inflammatory factors, support myeloid-skewed differentiation of HSPCs, and importantly, show poorer response to hypomethylation as a key differentiator in GSEA analysis. When exposed to long-term in vivo stimulation with primary MDSfast-MSCs-based scaffolds, healthy donor (HD) HSPCs show elevated NF-κB expression, similar to leukemic HSPCs in MDS. Those "MDSfast-MSCs-primed" HD-HSPCs continue to show enhanced engraftment rates in secondary MDS-MSC-based scaffolds, providing evidence for the microenvironmental selection pressures in MDS towards leukemic HSPCs. Together, our data point towards a degree of co-development between MSCs and HSPCs during the progression of MDS, where changes in MDS-MSCs take place mainly at the transcriptomic and functional levels. These unique differences in MDS-MSCs can be utilized to improve disease prognostication and implement targeted therapy for unmet clinical needs.

Multi-Layered Implant Approach for Hemilaryngectomy Reconstruction in a Porcine Model.

Partial laryngectomies result in voice, swallowing, and airway impairment for thousands of patients in the United States each year. Treatment options for dynamic restoration of laryngeal function are limited. Thus, there is a need for new reconstructive approaches. Here, we evaluated early (4 week) outcomes of multi-layered mucosal-myochondral (MMC) implants when used to restore laryngeal form and function after hemilaryngectomy in a porcine model. Six Yucatan minipigs underwent transmural hemilaryngectomies followed by reconstruction with customized MMC implants aiming to provide site-appropriate localization of regenerated laryngeal tissues, while supporting laryngeal function. All implants were fabricated from polymeric collagen, with a subset of muscle and cartilage implants containing motor endplate-expressing muscle progenitor cells or cartilage-like cells differentiated from adipose stem cells, respectively. Vocalization and laryngeal electromyography (L-EMG) measurements with nerve conduction studies were performed post-operatively and compared with baseline along with gross and histological analyses of the healing response. All animals (n = 6) survived and maintained airway patency, safe swallowing, and phonation, without the use of tracheostomy and/or gastrostomy tubes. Histological evaluation indicated no adverse tissue reaction or implant degradation, showing progressive regenerative remodeling with mucosa reformation and ingrowth of new muscle and cartilage. Preliminary L-EMG suggested weak but detectable motor unit action potentials. Although vocalization duration, frequency, and intensity decreased post-operatively, all animals retained vocal capacity and parameter recovery was evident over the study duration. Engineered collagen polymeric implants in the presence or absence of autologous cell populations may serve as a feasible reconstructive option to restore dynamic function after hemilaryngectomy. Long-term follow-up is needed to further assess functional outcomes. NA Laryngoscope, 2024.

VEGFC Overexpression in Kidney Progenitor Cells Is a Model of Renal Lymphangiectasia.

Lymphangiogenesis is believed to be a protective response in the setting of multiple forms of kidney injury and mitigates the progression of interstitial fibrosis. To augment this protective response, promoting kidney lymphangiogenesis is being investigated as a potential treatment to slow the progression of kidney disease. As injury-related lymphangiogenesis is driven by signaling from the receptor VEGFR3 (vascular endothelial growth factor receptor 3) in response to the cognate growth factor VEGF (vascular endothelial growth factor)-C released by tubular epithelial cells, this signaling pathway is a candidate for future kidney therapeutics. However, the consequences to kidney development and function to targeting this signaling pathway remain poorly defined. We generated a new mouse model expressing Vegfc under regulation of the nephron progenitor Six2Cre driver strain (Six2Vegfc). Mice underwent a detailed phenotypic evaluation. Whole kidneys were processed for histology and 3-dimensional imaging. Six2Vegfc mice had reduced body weight and kidney function compared with littermate controls. Six2Vegfc kidneys demonstrated large peripelvic fluid-filled lesions with distortion of the pelvicalcyceal system which progressed in severity with age. Three-dimensional imaging showed a 3-fold increase in total cortical vascular density. Histology confirmed a substantial increase in LYVE1+ (lymphatic vessel endothelial hyaluronan receptor-1)/PDPN+ (podoplanin)/VEGFR3+ lymphatic capillaries extending alongside EMCN+ (endomucin) peritubular capillaries. There was no change in EMCN+ peritubular capillary density. Kidney lymphatic density was robustly increased in the Six2Vegfc mice. There were no changes in peritubular blood capillary density despite these endothelial cells also expressing VEGFR3. The model resulted in malformation of the lymphatic hilar plexus, resulting in severe hydronephrosis that resembled a human condition termed renal lymphangiectasia. This study defines the vascular consequences of augmenting VEGFC signaling during kidney development and provides new insight into human renal lymphatic malformations.

Endothelial progenitor cell-derived conditioned medium mitigates chronic cerebral ischemic injury through macrophage migration inhibitory factor-activated AKT pathway.

Chronic cerebral ischemia (CCI) is a significant health issue characterized by hypoperfusion due to damage or occlusion of the cerebral or carotid arteries. CCI may lead to progressive cognitive impairment that is considered as a prelude to neurodegenerative diseases, including dementia and Alzheimer's disease (AD). Endothelial progenitor cells (EPCs) have been implicated in vascular repair in ischemic cerebrovascular diseases, primarily by differentiating into endothelial cells (ECs) or through paracrine effects. However, the clinical transplantation of stem cell therapies remains limited. In this study, we investigated the effects of EPC-derived conditioned medium (EPC-CM) on the impaired vasculature and neurological function in a rodent model of CCI and the mechanism involved. EPC-CM was analyzed by cytokine array to identify key factors involved in angiogenesis and cellular senescence. The effects and mechanism of the candidate factors in the EPC-CM were validated in vitro using oxygen-glucose deprivation (OGD)-injured ECs and EPCs. The therapeutic effects of EPC-CM and the identified key factor were further examined in a rat model of CCI, which was induced by bilateral internal carotid artery ligation (BICAL). EPC-CM was administered via intracisternal injection one week post BICAL. The cerebral microvasculature and neurobehavior of the rats were examined three weeks after BICAL. Macrophage migration inhibitory factor (MIF) was identified as a key factor in the EPC-CM. Recombinant MIF protein promoted angiogenesis and prevented senescence in the injured EPCs and ECs. The effect was similar to that of the EPC-CM. These therapeutic effects were diminished when the EPC-CM was co-treated with MIF-specific antibody (Ab). Additionally, the vascular, motor, and cognitive improvements observed in the BICAL rats treated with EPC-CM were abolished by co-treated with MIF Ab. Furthermore, we found MIF promoted angiogenesis and anti-senescence via activating the AKT pathway. Inhibition of the AKT pathway diminished the protective effects of MIF in the in vitro study. We demonstrated that EPC-CM protected the brain from chronic ischemic injury and promoted functional recovery through MIF-mediated AKT pathway. These findings suggest EPC-CM holds potential as a novel cell-free therapeutic approach for treating CCI through the actions of MIF.

Mechanical stretch and chronotherapeutic techniques for progenitor cell transplantation and biomaterials

Mechanical stretch and chronotherapeutic techniques for progenitor cell transplantation and biomaterials

Computing hematopoiesis plasticity in response to genetic mutations and environmental stimulations.

Cell plasticity (CP), describing a dynamic cell state, plays a crucial role in maintaining homeostasis during organ morphogenesis, regeneration, and trauma-to-repair biological process. Single-cell-omics datasets provide an unprecedented resource to empower CP analysis. Hematopoiesis offers fertile opportunities to develop quantitative methods for understanding CP. In this study, we generated high-quality lineage-negative single-cell RNA-sequencing datasets under various conditions and introduced a working pipeline named scPlasticity to interrogate naïve and disturbed plasticity of hematopoietic stem and progenitor cells with mutational or environmental challenges. Using embedding methods UMAP or FA, a continuum of hematopoietic development is visually observed in wild type where the pipeline confirms a low proportion of hybrid cells ( P hc , with bias range: 0.4∼0.6) on a transition trajectory. Upon Tet2 mutation, a driver of leukemia, or treatment of DSS, an inducer of colitis, P hc is increased and plasticity of hematopoietic stem and progenitor cells was enhanced. We prioritized several transcription factors and signaling pathways, which are responsible for P hc alterations. In silico perturbation suggests knocking out EGR regulons or pathways of IL-1R1 and β-adrenoreceptor partially reverses P hc promoted by Tet2 mutation and inflammation.

The IL-1β inhibitor canakinumab in previously treated lower-risk myelodysplastic syndromes: a phase 2 clinical trial.

In myelodysplastic syndromes (MDS), the IL-1β pathway is upregulated, and previous studies using mouse models of founder MDS mutations demonstrated that it enhances hematopoietic stem and progenitor cells' (HSPCs') aberrant differentiation towards the myeloid lineage at the expense of erythropoiesis. To evaluate whether targeting the IL-1β signaling pathway can rescue ineffective erythropoiesis in patients with MDS, we designed a phase 2 non-randomized single-arm clinical trial (NCT04239157) to assess the safety profile and efficacy of the IL-1β inhibitor canakinumab in previously treated lower-risk MDS patients. We enrolled 25 patients with a median age of 74 years; 60% were male, 16% had lower-risk MDS, 84% had intermediate-1 risk MDS according to the International Prognostic Scoring System score, and 80% failed hypomethylating agent therapy. The study met the primary endpoint of defining the clinical activity of canakinumab, and the secondary objective of determining the safety profile, including the rate of transfusion independence, the duration of response, progression-free survival, leukemia-free survival, and overall survival. The overall response rate was 17.4%, with all responses including hematological improvement. Sequential post-hoc prospective single-cell RNA sequencing analyses of HSPCs and bone marrow mononuclear cells at different time points during therapy showed that canakinumab's on-target effects in hematopoietic populations expressing the IL-1β receptor decreased the TNF-mediated inflammatory signaling pathway but rescued ineffective erythropoiesis only in the context of lower genetic complexity. This study demonstrates that better stratification strategies could target lower-risk MDS patients more effectively.

Pharmacogenomic screening identifies and repurposes leucovorin and dyclonine as pro-oligodendrogenic compounds in brain repair.

Oligodendrocytes are critical for CNS myelin formation and are involved in preterm-birth brain injury (PBI) and multiple sclerosis (MS), both of which lack effective treatments. We present a pharmacogenomic approach that identifies compounds with potent pro-oligodendrogenic activity, selected through a scoring strategy (OligoScore) based on their modulation of oligodendrogenic and (re)myelination-related transcriptional programs. Through in vitro neural and oligodendrocyte progenitor cell (OPC) cultures, ex vivo cerebellar explants, and in vivo mouse models of PBI and MS, we identify FDA-approved leucovorin and dyclonine as promising candidates. In a neonatal chronic hypoxia mouse model mimicking PBI, both compounds promote neural progenitor cell proliferation and oligodendroglial fate acquisition, with leucovorin further enhancing differentiation. In an adult MS model of focal de/remyelination, they improve lesion repair by promoting OPC differentiation while preserving the OPC pool. Additionally, they shift microglia from a pro-inflammatory to a pro-regenerative profile and enhance myelin debris clearance. These findings support the repurposing of leucovorin and dyclonine for clinical trials targeting myelin disorders, offering potential therapeutic avenues for PBI and MS.

KAT6B is required for histone 3 lysine 9 acetylation and SOX gene expression in the developing brain.

Heterozygous mutations in the histone lysine acetyltransferase gene KAT6B (MYST4/MORF/QKF) underlie neurodevelopmental disorders, but the mechanistic roles of KAT6B remain poorly understood. Here, we show that loss of KAT6B in embryonic neural stem and progenitor cells (NSPCs) impaired cell proliferation, neuronal differentiation, and neurite outgrowth. Mechanistically, loss of KAT6B resulted in reduced acetylation at histone H3 lysine 9 and reduced expression of key nervous system development genes in NSPCs and the developing cortex, including the SOX gene family, in particular Sox2, which is a key driver of neural progenitor proliferation, multipotency and brain development. In the fetal cortex, KAT6B occupied the Sox2 locus. Loss of KAT6B caused a reduction in Sox2 promoter activity in NSPCs. Sox2 overexpression partially rescued the proliferative defect of Kat6b -/- NSPCs. Collectively, these results elucidate molecular requirements for KAT6B in brain development and identify key KAT6B targets in neural precursor cells and the developing brain.