Vascular tissue - boon or bane? How pathogens usurp long-distance transport in plants and the defence mechanisms deployed to counteract them.

Evolutionary emergence of specialised vascular tissues has enabled plants to coordinate their growth and adjust to unfavourable external conditions. Whilst holding a pivotal role in long-distance transport, both xylem and phloem can be encroached on by various biotic factors for systemic invasion and hijacking of nutrients. Therefore, a complete understanding of the strategies deployed by plants against such pathogens to restrict their entry and establishment within plant tissues, is of key importance for the future development of disease-tolerant crops. In this review, we aim to describe how microorganisms exploit the plant vascular system as a route for gaining access and control of different host tissues and metabolic pathways. Highlighting several biological examples, we detail the wide range of host responses triggered to prevent or hinder vascular colonisation and effectively minimise damage upon biotic invasions.

Mitochondrial Transplantation Promotes Protective Effector and Memory CD4+ T Cell Response During Mycobacterium Tuberculosis Infection and Diminishes Exhaustion and Senescence in Elderly CD4+ T cells.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (M.tb), is a major global health concern, particularly affecting those with weakened immune systems, including the elderly. CD4+ T cell response is crucial for immunity against M.tb, but chronic infections and aging can lead to T cell exhaustion and senescence, worsening TB disease. Mitochondrial dysfunction, prevalent in aging and chronic diseases, disrupts cellular metabolism, increases oxidative stress, and impairs T-cell functions. This study investigates the effect of mitochondrial transplantation (mito-transfer) on CD4+ T cell differentiation and function in aged mouse models and human CD4+ T cells from elderly individuals. Mito-transfer in naïve CD4+ T cells is found to promote protective effector and memory T cell generation during M.tb infection in mice. Additionally, it improves elderly human T cell function by increasing mitochondrial mass and altering cytokine production, thereby reducing markers of exhaustion and senescence. These findings suggest mito-transfer as a novel approach to enhance aged CD4+ T cell functionality, potentially benefiting immune responses in the elderly and chronic TB patients. This has broader implications for diseases where mitochondrial dysfunction contributes to T-cell exhaustion and senescence.

The progress of induced pluripotent stem cells derived from pigs: a mini review of recent advances.

Pigs (Sus scrofa) are widely acknowledged as an important large mammalian animal model due to their similarity to human physiology, genetics, and immunology. Leveraging the full potential of this model presents significant opportunities for major advancements in the fields of comparative biology, disease modeling, and regenerative medicine. Thus, the derivation of pluripotent stem cells from this species can offer new tools for disease modeling and serve as a stepping stone to test future autologous or allogeneic cell-based therapies. Over the past few decades, great progress has been made in establishing porcine pluripotent stem cells (pPSCs), including embryonic stem cells (pESCs) derived from pre- and peri-implantation embryos, and porcine induced pluripotent stem cells (piPSCs) using a variety of cellular reprogramming strategies. However, the stabilization of pPSCs was not as straightforward as directly applying the culture conditions developed and optimized for murine or primate PSCs. Therefore, it has historically been challenging to establish stable pPSC lines that could pass stringent pluripotency tests. Here, we review recent advances in the establishment of stable porcine PSCs. We focus on the evolving derivation methods that eventually led to the establishment of pESCs and transgene-free piPSCs, as well as current challenges and opportunities in this rapidly advancing field.

Deciphering two decades of cellular reprogramming in cancer: A bibliometric analysis of evolving trends and research frontiers.

Recent research has reevaluated the traditional view of cancer's linear progression and recurrence by introducing cellular reprogramming a process in which cancer cells can their state under certain conditions. This change is driven by a combination of genetic and epigenetic factors, with pivotal roles played by key genes, and pathways, notably Wnt and Notch. The complexity of cancer's behavior is further influenced by factors such as the epithelial-mesenchymal transition (EMT) and therapy-induced stress, both of which are significant contributors to cancer recurrence. In this context bibliometric analysis emerges as a crucial tool for evaluating the impacts and trends within scientific literature. Our study utilized bibliometrics to analysis the role of cellular reprogramming oncology over the past two decades, highlighting its potential to improve cancer treatment outcomes. In conducting this analysis, we searched for literature search on cellular reprogramming (CR) in the Web of Science database, covering the years 2002-2022. We employed visualization tools like Citespace, VOSviewer, and Bibliometrix to analyze the collected data resulting in a dataset of 3102 articles. The United States and China emerged as leading contributors to this field, with the University of Texas MD Anderson Cancer Center being the most prolific institution. Menendez was the most influential scholar in this research domain. Cancers was the journal with the most publications on this subject. The most local-cited document was the article titled "Hallmarks of Cancer: The Next Generation". A comprehensive analysis has been conducted based on keywords and cited references. In recent years, the research emphasis has shifted to "extracellular vesicles," "cancer therapy," and "cellular plasticity". Therefore, this analysis uses bibliometrics to chart cutting-edge progress in cancer's cellular reprogramming, aiding experts to quickly understand and innovate in this crucial area.

Dissecting gene activation and chromatin remodeling dynamics in single human cells undergoing reprogramming.

During cell fate transitions, cells remodel their transcriptome, chromatin, and epigenome; however, it has been difficult to determine the temporal dynamics and cause-effect relationship between these changes at the single-cell level. Here, we employ the heterokaryon-mediated reprogramming system as a single-cell model to dissect key temporal events during early stages of pluripotency conversion using super-resolution imaging. We reveal that, following heterokaryon formation, the somatic nucleus undergoes global chromatin decompaction and removal of repressive histone modifications H3K9me3 and H3K27me3 without acquisition of active modifications H3K4me3 and H3K9ac. The pluripotency gene OCT4 (POU5F1) shows nascent and mature RNA transcription within the first 24 h after cell fusion without requiring an initial open chromatin configuration at its locus. NANOG, conversely, has significant nascent RNA transcription only at 48 h after cell fusion but, strikingly, exhibits genomic reopening early on. These findings suggest that the temporal relationship between chromatin compaction and gene activation during cellular reprogramming is gene context dependent.

Role of small molecules as drug candidates for reprogramming somatic cells into induced pluripotent stem cells: A comprehensive review.

With the use of specific genetic factors and recent developments in cellular reprogramming, it is now possible to generate lineage-committed cells or induced pluripotent stem cells (iPSCs) from readily available and common somatic cell types. However, there are still significant doubts regarding the safety and effectiveness of the current genetic methods for reprogramming cells, as well as the conventional culture methods for maintaining stem cells. Small molecules that target specific epigenetic processes, signaling pathways, and other cellular processes can be used as a complementary approach to manipulate cell fate to achieve a desired objective. It has been discovered that a growing number of small molecules can support lineage differentiation, maintain stem cell self-renewal potential, and facilitate reprogramming by either increasing the efficiency of reprogramming or acting as a genetic reprogramming factor substitute. However, ongoing challenges include improving reprogramming efficiency, ensuring the safety of small molecules, and addressing issues with incomplete epigenetic resetting. Small molecule iPSCs have significant clinical applications in regenerative medicine and personalized therapies. This review emphasizes the versatility and potential safety benefits of small molecules in overcoming challenges associated with the iPSCs reprogramming process.

Dexamethasone-Integrated Mesenchymal Stem Cells for Systemic Lupus Erythematosus Treatment via Multiple Immunomodulatory Mechanisms.

The therapeutic application of mesenchymal stem cells (MSCs) has good potential as a treatment strategy for systemic lupus erythematosus (SLE), but traditional MSC therapy still has limitations in effectively modulating immune cells. Herein, we present a promising strategy based on dexamethasone liposome-integrated MSCs (Dexlip-MSCs) for treating SLE via multiple immunomodulatory pathways. This therapeutic strategy prolonged the circulation time of dexamethasone liposomes in vivo, restrained CD4+T-cell proliferation, and inhibited the release of proinflammatory mediators (IFN-γ and TNF-α) by CD4+T cells. In addition, Dexlip-MSCs initiated cellular reprogramming by activating the glucocorticoid receptor (GR) signaling pathway to upregulate the expression of anti-inflammatory factors such as cysteine-rich secretory protein LCCL-containing domain 2 (CRISPLD2) and downregulate the expression of proinflammatory factors. In addition, Dexlip-MSCs synergistically increased the anti-inflammatory inhibitory effect of CD4+T cells through the release of dexamethasone liposomes or Dex-integrated MSC-derived exosomes (Dex-MSC-EXOs). Based on these synergistic biological effects, we demonstrated that Dexlip-MSCs alleviated disease progression in MRL/lpr mice more effectively than Dexlip or MSCs alone. These features indicate that our stem cell delivery strategy is a promising therapeutic approach for clinical SLE treatment.

A Youthful Touch: Reversal of Aging Hallmarks by Cell Reprogramming.

BACKGROUND: With the elderly population projected to double by 2050, there is an urgent need to address the increasing prevalence of age-related debilitating diseases and ultimately minimize discrepancies between the rising lifespan and stagnant health span. Cellular reprogramming by overexpression of Oct3/4, Klf4, Sox2, and cMyc (OKSM) transcription factors is gaining attention in this context thanks to demonstrated rejuvenating effects in human cell cultures and live mice, many of which can be uncoupled from dedifferentiation and loss of cell identity. SUMMARY: Here, we review current evidence of the impact of cell reprogramming on established aging hallmarks and the underlying mechanisms that mediate these effects. We also provide a critical assessment of the challenges in translating these findings and, overall, cell reprogramming technologies into clinically translatable antiaging interventions. KEY MESSAGES: Cellular reprogramming has the potential to reverse at least partially some key hallmarks of aging. However, further research is necessary to determine the biological significance and duration of such changes and to ensure the safety of cell reprogramming as a rejuvenation approach. With this review, we hope to stimulate new research directions in the quest to extend health span effectively.

Ultrasound-triggered three dimensional hyaluronic acid hydrogel promotes in vitro and in vivo reprogramming into induced pluripotent stem cells.

Cellular reprogramming technologies have been developed with different physicochemical factors to improve the reprogramming efficiencies of induced pluripotent stem cells (iPSCs). Ultrasound is a clinically applied noncontact biophysical factor known for regulating various cellular behaviors but remains uninvestigated for cellular reprogramming. Here, we present a new reprogramming strategy using low-intensity ultrasound (LIUS) to improve cellular reprogramming of iPSCs in vitro and in vivo. Under 3D microenvironment conditions, increased LIUS stimulation shows enhanced cellular reprogramming of the iPSCs. The cellular reprogramming process facilitated by LIUS is accompanied by increased mesenchymal to epithelial transition and histone modification. LIUS stimulation transiently modulates the cytoskeletal rearrangement, along with increased membrane fluidity and mobility to increase HA/CD44 interactions. Furthermore, LIUS stimulation with HA hydrogel can be utilized in application of both human cells and in vivo environment, for enhanced reprogrammed cells into iPSCs. Thus, LIUS stimulation with a combinatorial 3D microenvironment system can improve cellular reprogramming in vitro and in vivo environments, which can be applied in various biomedical fields.

Establishment of Transgene-Free Porcine Induced Pluripotent Stem Cells.

Although protocols to generate authentic transgene-free mouse and human induced pluripotent stem cells (iPSCs) are now well established, standard methods for reprogramming porcine somatic cells still suffer from low efficiency and transgene retention. The Basic Protocol describes reprogramming procedures to establish transgene-free porcine iPSCs (PiPSCs) from porcine fibroblasts. This method uses episomal plasmids encoding POU5F1, SOX2, NANOG, KLF4, SV40LT, c-MYC, LIN28A, and microRNA-302/367, combined with an optimized medium, to establish PiPSC lines. Support protocols describe the establishment and characterization of clonal PiPSC lines, as well as the preparation of feeder cells and EBNA1 mRNA. This optimized, step-by-step approach tailored to this species enables the efficient derivation of PiPSCs in ∼4 weeks. The establishment of transgene-free PiPSCs provides a new and valuable model for studies of larger mammalian species' development, disease, and regenerative biology. © 2024 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Reprogramming of porcine fibroblasts with episomal plasmids Support Protocol 1: Preparation of mouse embryonic fibroblasts for feeder layer Support Protocol 2: Preparation of in vitro-transcribed EBNA1 mRNA Support Protocol 3: Establishment of clonal porcine induced pluripotent stem cell (PiPSC) lines Support Protocol 4: PiPSC characterization: Genomic DNA PCR and RT-PCR Support Protocol 5: PiPSC characterization: Immunostaining.

Generation of Artificial Blastoids Combining miR-200-Mediated Reprogramming and Mechanical Cues.

In vitro-generated blastocyst-like structures are of great importance since they recapitulate specific features or processes of early embryogenesis, thus avoiding ethical concerns as well as increasing scalability and accessibility compared to the use of natural embryos. Here, we combine cell reprogramming and mechanical stimuli to create 3D spherical aggregates that are phenotypically similar to those of natural embryos. Specifically, dermal fibroblasts are reprogrammed, exploiting the miR-200 family property to induce a high plasticity state in somatic cells. Subsequently, miR-200-reprogrammed cells are either driven towards the trophectoderm (TR) lineage using an ad hoc induction protocol or encapsulated into polytetrafluoroethylene micro-bioreactors to maintain and promote pluripotency, generating inner cell mass (ICM)-like spheroids. The obtained TR-like cells and ICM-like spheroids are then co-cultured in the same micro-bioreactor and, subsequently, transferred to microwells to encourage blastoid formation. Notably, the above protocol was applied to fibroblasts obtained from young as well as aged donors, with results that highlighted miR-200's ability to successfully reprogram young and aged cells with comparable blastoid rates, regardless of the donor's cell age. Overall, the approach here described represents a novel strategy for the creation of artificial blastoids to be used in the field of assisted reproduction technologies for the study of peri- and early post-implantation mechanisms.

SOX17/ETV2 improves the direct reprogramming of adult fibroblasts to endothelial cells.

An autologous source of vascular endothelial cells (ECs) is valuable for vascular regeneration and tissue engineering without the concern of immune rejection. The transcription factor ETS variant 2 (ETV2) has been shown to directly convert patient fibroblasts into vascular EC-like cells. However, reprogramming efficiency is low and there are limitations in EC functions, such as eNOS expression. In this study, we directly reprogram adult human dermal fibroblasts into reprogrammed ECs (rECs) by overexpressing SOX17 in conjunction with ETV2. We find several advantages to rEC generation using this approach, including improved reprogramming efficiency, increased enrichment of EC genes, formation of large blood vessels carrying blood from the host, and, most importantly, expression of eNOS in vivo. From these results, we present an improved method to reprogram adult fibroblasts into functional ECs and posit ideas for the future that could potentially further improve the reprogramming process.

In Vivo Reprogramming Using Yamanaka Factors in the CNS: A Scoping Review.

Central nervous system diseases, particularly neurodegenerative disorders, pose significant challenges in medicine. These conditions, characterized by progressive neuronal loss, have remained largely incurable, exacting a heavy toll on individuals and society. In recent years, in vivo reprogramming using Yamanaka factors has emerged as a promising approach for central nervous system regeneration. This technique involves introducing transcription factors, such as Oct4, Sox2, Klf4, and c-Myc, into adult cells to induce their conversion into neurons. This review summarizes the current state of in vivo reprogramming research in the central nervous system, focusing on the use of Yamanaka factors. In vivo reprogramming using Yamanaka factors has shown promising results in several animal models of central nervous system diseases. Studies have demonstrated that this approach can promote the generation of new neurons, improve functional outcomes, and reduce scar formation. However, there are still several challenges that need to be addressed before this approach can be translated into clinical practice. These challenges include optimizing the efficiency of reprogramming, understanding the cell of origin for each transcription factor, and developing methods for reprogramming in non-subventricular zone areas. Further research is needed to overcome the remaining challenges, but this approach has the potential to revolutionize the way we treat central nervous system disorders.

Stage II oesophageal carcinoma: peril in disguise associated with cellular reprogramming and oncogenesis regulated by pseudogenes.

INTRODUCTION: Pseudogenes have been implicated for their role in regulating cellular differentiation and organismal development. However, their role in promoting cancer-associated differentiation has not been well-studied. This study explores the tumour landscape of oesophageal carcinoma to identify pseudogenes that may regulate events of differentiation to promote oncogenic transformation. MATERIALS AND METHOD: De-regulated differentiation-associated pseudogenes were identified using DeSeq2 followed by 'InteractiVenn' analysis to identify their expression pattern. Gene expression dependent and independent enrichment analyses were performed with GSEA and ShinyGO, respectively, followed by quantification of cellular reprogramming, extent of differentiation and pleiotropy using three unique metrics. Stage-specific gene regulatory networks using Bayesian Network Splitting Average were generated, followed by network topology analysis. MEME, STREME and Tomtom were employed to identify transcription factors and miRNAs that play a regulatory role downstream of pseudogenes to initiate cellular reprogramming and further promote oncogenic transformation. The patient samples were stratified based on the expression pattern of pseudogenes, followed by GSEA, mutation analysis and survival analysis using GSEA, MAF and 'survminer', respectively. RESULTS: Pseudogenes display a unique stage-wise expression pattern that characterizes stage II (SII) ESCA with a high rate of cellular reprogramming, degree of differentiation and pleiotropy. Gene regulatory network and associated topology indicate high robustness, thus validating high pleiotropy observed for SII. Pseudogene-regulated expression of SOX2, FEV, PRRX1 and TFAP2A in SII may modulate cellular reprogramming and promote oncogenesis. Additionally, patient stratification-based mutational analysis in SII signifies APOBEC3A (A3A) as a potential hallmark of homeostatic mutational events of reprogrammed cells which in addition to de-regulated APOBEC3G leads to distinct events of hypermutations. Further enrichment analysis for both cohorts revealed the critical role of combinatorial expression of pseudogenes in cellular reprogramming. Finally, survival analysis reveals distinct genes that promote poor prognosis in SII ESCA and patient-stratified cohorts, thus providing valuable prognostic bio-markers along with markers of differentiation and oncogenesis for distinct landscapes of pseudogene expression. CONCLUSION: Pseudogenes associated with the events of differentiation potentially aid in the initiation of cellular reprogramming to facilitate oncogenic transformation, especially during SII ESCA. Despite a better overall survival of SII, patient stratification reveals combinatorial de-regulation of pseudogenes as a notable marker for a high degree of cellular differentiation with a unique mutational landscape.

Effect of Octamer-Binding Transcription Factor 4 Overexpression on the Neural Induction of Human Dental Pulp Stem Cells.

Stem cell-based therapy is a potential alternative strategy for brain repair, with neural stem cells (NSC) presenting as the most promising candidates. Obtaining sufficient quantities of NSC for clinical applications is challenging, therefore alternative cell types, such as neural crest-derived dental pulp stem cells (DPSC), may be considered. Human DPSC possess neurogenic potential, exerting positive effects in the damaged brain through paracrine effects. However, a method for conversion of DPSC into NSC has yet to be developed. Here, overexpression of octamer-binding transcription factor 4 (OCT4) in combination with neural inductive conditions was used to reprogram human DPSC along the neural lineage. The reprogrammed DPSC demonstrated a neuronal-like phenotype, with increased expression levels of neural markers, limited capacity for sphere formation, and enhanced neuronal but not glial differentiation. Transcriptomic analysis further highlighted the expression of genes associated with neural and neuronal functions. In vivo analysis using a developmental avian model showed that implanted DPSC survived in the developing central nervous system and respond to endogenous signals, displaying neuronal phenotypes. Therefore, OCT4 enhances the neural potential of DPSC, which exhibited characteristics aligning with neuronal progenitors. This method can be used to standardise DPSC neural induction and provide an alternative source of neural cell types.

Induction of Transient Morula-Like Cells in Mice Through STAT3 Activation.

Developing in vitro cell models that faithfully replicate the molecular and functional traits of cells from the earliest stages of mammalian development presents a significant challenge. The strategic induction of signal transducer and activator of transcription 3 (STAT3) phosphorylation, coupled with carefully defined culture conditions, facilitates the efficient reprogramming of mouse pluripotent cells into a transient morula-like cell (MLC) state. The resulting MLCs closely mirror their in vivo counterparts, exhibiting not only molecular resemblance but also the ability to differentiate into both embryonic and extraembryonic lineages. This reprogramming approach provides valuable insights into controlled cellular fate choice and opens new opportunities for studying early developmental processes in a dish.

Inducing Pluripotency in Somatic Cells: Historical Perspective and Recent Advances.

Inducing pluripotency in somatic cells is mediated by the Yamanaka factors Oct4, Sox2, Klf4, and c-Myc. The resulting induced pluripotent stem cells (iPSCs) hold great promise for regenerative medicine by virtue of their ability to differentiate into different types of functional cells. Specifically, iPSCs derived directly from patients offer a powerful platform for creating in vitro disease models. This facilitates elucidation of pathological mechanisms underlying human diseases and development of new therapeutic agents mitigating disease phenotypes. Furthermore, genetically and phenotypically corrected patient-derived iPSCs by gene-editing technology or the supply of specific pharmaceutical agents can be used for preclinical and clinical trials to investigate their therapeutic potential. Despite great advances in developing reprogramming methods, the efficiency of iPSC generation remains still low and varies between donor cell types, hampering the potential application of iPSC technology. This paper reviews histological timeline showing important discoveries that have led to iPSC generation and discusses recent advances in iPSC technology by highlighting donor cell types employed for iPSC generation.

Navigating the Landscape of Intestinal Regeneration: A Spotlight on Quiescence Regulation and Fetal Reprogramming.

Tissue-specific adult stem cells are pivotal in maintaining tissue homeostasis, especially in the rapidly renewing intestinal epithelium. At the heart of this process are leucine-rich repeat-containing G protein-coupled receptor 5-expressing crypt base columnar cells (CBCs) that differentiate into various intestinal epithelial cells. However, while these CBCs are vital for tissue turnover, they are vulnerable to cytotoxic agents. Recent advances indicate that alternative stem cell sources drive the epithelial regeneration post-injury. Techniques like lineage tracing and single-cell RNA sequencing, combined with in vitro organoid systems, highlight the remarkable cellular adaptability of the intestinal epithelium during repair. These regenerative responses are mediated by the reactivation of conserved stem cells, predominantly quiescent stem cells and revival stem cells. With focus on these cells, this review unpacks underlying mechanisms governing intestinal regeneration and explores their potential clinical applications.

Metabolic Adaptation and Cellular Stress Response As Targets for Cancer Therapy.

Cancer cells, which divide indefinitely and without control, are frequently exposed to various stress factors but manage to adapt and survive. The mechanisms by which cancer cells maintain cellular homeostasis and exploit stress conditions are not yet clear. Here, we elucidate the roles of diverse cellular metabolism and its regulatory mechanisms, highlighting the essential role of metabolism in cellular composition and signal transduction. Cells respond to various stresses, including DNA damage, energy stress, and oxidative stress, thereby causing metabolic alteration. We provide profound insight into the adaptive mechanisms employed by cancer cells to ensure their survival among internal and external stressors through a comprehensive analysis of the correlation between metabolic alterations and cellular stress. Furthermore, this research establishes a robust framework for the development of innovative therapeutic strategies that specifically target the cellular adaptations of cancer cells.

Partial cellular reprogramming: A deep dive into an emerging rejuvenation technology.

Aging and age-associated disease are a major medical and societal burden in need of effective treatments. Cellular reprogramming is a biological process capable of modulating cell fate and cellular age. Harnessing the rejuvenating benefits without altering cell identity via partial cellular reprogramming has emerged as a novel translational strategy with therapeutic potential and strong commercial interests. Here, we explore the aging-related benefits of partial cellular reprogramming while examining limitations and future directions for the field.