Dedifferentiation Of Cells Research Articles

Epithelial plasticity enhances regeneration of committed taste receptor cells following nerve injury

Taste receptor cells are taste bud epithelial cells that are dependent upon the innervating nerve for continuous renewal and are maintained by resident tissue stem/progenitor cells. Transection of the innervating nerve causes degeneration of taste buds and taste receptor cells. However, a subset of the taste receptor cells is maintained without nerve contact after glossopharyngeal nerve transection in the circumvallate papilla in adult mice. Here, we revealed that injury caused by glossopharyngeal nerve transection triggers the remaining differentiated K8-positive taste receptor cells to dedifferentiate and acquire transient progenitor cell-like states during regeneration. Dedifferentiated taste receptor cells proliferate, express progenitor cell markers (K14, Sox2, PCNA) and form organoids in vitro. These data indicate that differentiated taste receptor cells can enter the cell cycle, acquire stemness, and participate in taste bud regeneration. We propose that dedifferentiated taste receptor cells in combination with stem/progenitor cells enhance the regeneration of taste buds following nerve injury.

The Metabolic Changes between Monolayer (2D) and Three-Dimensional (3D) Culture Conditions in Human Mesenchymal Stem/Stromal Cells Derived from Adipose Tissue

One of the key factors that may influence the therapeutic potential of mesenchymal stem/stromal cells (MSCs) is their metabolism. The switch between mitochondrial respiration and glycolysis can be affected by many factors, including the oxygen concentration and the spatial form of culture. This study compared the metabolic features of adipose-derived mesenchymal stem/stromal cells (ASCs) and dedifferentiated fat cells (DFATs) cultivated as monolayer or spheroid culture under 5% O2 concentration (physiological normoxia) and their impact on MSCs therapeutic abilities. We observed that the cells cultured as spheroids had a slightly lower viability and a reduced proliferation rate but a higher expression of the stemness-related transcriptional factors compared to the cells cultured in monolayer. The three-dimensional culture form increased mtDNA content, oxygen consumption rate (OCR) and extracellular acidification rate (ECAR), especially in DFATs-3D population. The DFATs spheroids also demonstrated increased levels of Complex V proteins and higher rates of ATP production. Moreover, increased reactive oxygen species and lower intracellular lactic acid levels were also found in 3D culture. Our results may suggest that metabolic reconfiguration accompanies the transition from 2D to 3D culture and the processes of both mitochondrial respiration and glycolysis become more active. Intensified metabolism might be associated with the increased demand for energy, which is needed to maintain the expression of pluripotency genes and stemness state.

ECM proteins and cationic polymers coating promote dedifferentiation of patient-derived mature adipocytes to stem cells.

Reprogramming of mature adipocytes is an attractive research area due to the plasticity of these cells. Mature adipocytes can be reprogrammed in vitro, transforming them into dedifferentiated fat cells (DFATs), which are considered a new type of stem cell, and thereby have a high potential for use in tissue engineering and regenerative medicine. However, there are still no reports or findings on in vitro controlling the dedifferentiation. Although ceiling culture performed in related studies is a relatively simple method, its yield is low and does not allow manipulation of mature adipocytes to increase or decrease the dedifferentiation. In this study, to understand the role of physicochemical surface effects on the dedifferentiation of patient-derived mature adipocytes, the surfaces of cell culture flasks were coated with extracellular matrix, basement membrane proteins, and cationic/anionic polymers. Extracellular matrix such as fibronectin and collagen type I, and basement membrane proteins such as collagen type IV and laminin strongly promoted dedifferentiation of mature adipocytes, with laminin showing the highest effect with a DFAT ratio of 2.98 (±0.84). Interestingly, cationic polymers also showed a high dedifferentiation effect, but anionic polymers did not, and poly(diallyl dimethylammonium chloride) showed the highest DFAT ratio of 2.27 (±2.8) among the cationic polymers. Protein assay results revealed that serum proteins were strongly adsorbed on the surfaces of the cationic polymer coating, including inducing high mature adipocyte adhesion. This study demonstrates for the first time the possibility of regulating the transformation of mature adipocytes to DFAT stem cells by controlling the physicochemical properties of the surface of conventional cell culture flasks.

Plant <i>in situ</i> tissue regeneration: dynamics, mechanisms and implications for forestry research

Plants possess remarkable developmental plasticity and regenerative ability to reshape themselves in response to external stimulations. After local injuries, they can initiate cellular reprogramming at the wound sites to repair or regrow structures that could substitute the damaged or lost parts functionally. This way of regeneration in plants is called plant <italic>in situ</italic> tissue regeneration. Upon wounding like excision, incision or girdling, the original tissue patterns are completely or partially destroyed, the remanent tissues could perceive the wounding signals and thereby initiate cell de-differentiation, trans-differentiation or re-differentiation to reconstruct the lost or damaged tissues. In this review, we summarize the regenerative dynamics and regulatory mechanisms during the major <italic>in situ</italic> tissue regeneration processes in plants, including secondary vascular tissue (SVT) regeneration after girdling, apex regeneration after excision and tissue reunion after incision. In addition, we compare the features of SVT regeneration, the most relevant system for forestry, with other plant <italic>in situ</italic> tissue regeneration systems. We further discuss the unsolved issues and the potential applications of plant <italic>in situ</italic> regeneration for forestry research, aiming to provide new insights for the study of woody plant development.

Sodium/Iodide Symporter Analysis for Prognostication and Management in Thyroglobulin-elevated Negative Iodine Scintigraphy Syndrome: Current Advances and Future Prospects

ABSTRACT Well-differentiated thyroid cancers are typically managed with surgery followed by radioiodine ablation. These cancers often retain the ability to concentrate intracellular iodine. However, cancer cell dedifferentiation frequently confers radioiodine resistance, termed thyroglobulin-elevated negative iodine scintigraphy (TENIS) syndrome. Elucidation of the sodium/iodide symporter (NIS) mechanism that mediates active iodide transport has enabled advances in the pathophysiologic understanding and clinical management of TENIS syndrome. This review summarizes molecular cloning of the NIS gene and implications for thyroid biology. It highlights studies of NIS gene regulation across tissues and utilization of NIS as a reporter gene to restore radioiodine uptake in advanced thyroid cancers. Quantitative NIS expression analyses and correlations with radioiodine uptake are also discussed. Overall, this review underscores the vital role of NIS-mediated iodide transport in thyroid pathophysiology and modern radioiodine therapeutic paradigms for advanced thyroid cancers, including TENIS syndrome. Further research into mechanisms underlying differential NIS expression and therapeutic modulation of NIS activity is warranted.

Astroglial differentiation of fibronectin‑positive human "glia-like" cells.

Fibronectin (Fn) is a glycoprotein of extracellular matrix produced by a variety of mesenchymal and neoplastic cell types. In adult brain tissue, Fn is restricted to blood vessels. However, adult human brain cultures are almost entirely comprised of flat or spindle‑shaped Fn-positive cells usually referred to as "glia-like" cells. Since Fn is primarily present in fibroblasts, these cultures may be considered to be of non-glial origin. Cells gained by long-term culturing of adult human brain tissue derived from brain biopsies obtained from 12 patients with non-malignant diagnoses were examined by immunofluorescence methods. Primary cultures contained GFAP-/Vim+/Fn+ "glia-like" cells (95-98 %) and GFAP+/Vim+/Fn- astrocytes (0.1 %) which disappeared by passage number 3. The formation of cell processes and enlargement of cell bodies was observed in 9 of 12 cultures with decreased cell growth during passages 12 to 17. It is remarkable that during this period, all "glia-like" cells became GFAP+/Vim+/Fn+. Herein, we confirm our previously published hypothesis about the origin of adult human "glia-like" cells, which we consider to be precursor cells scattered through the brain cortex and subcortical white matter. Cultures were comprised entirely of GFAP-/Fn+ "glia-like" cells and showed morphological and immunochemical astroglial differentiation with spontaneously decelerated growth during prolonged passaging. We propose that the adult human brain tissue contains a "dormant population" of undefined glial precursor cells. Under culture, these cells show to have a high proliferative capacity and different stages of cell dedifferentiation (Fig. 2, Ref. 21).

PDGF-BB is involved in HIF-1α/CXCR4/CXCR7 axis promoting capillarization of hepatic sinusoidal endothelial cells

BackgroundThe activation of HIF-1α/CXCR4 pathway in liver sinusoidal endothelial cells (LSECs) could downregulate CXCR7, leading to the capillarization of LSECs to promote hepatic fibrosis. However, the mechanism between CXCR4 and CXCR7 is still undefined. The aim is to investigate the role of PDGF-BB in the dedifferentiation of LSECs and hepatic stellate cells (HSCs) activation. MethodsThe activation of HIF-1α/CXCR4 pathway in two kinds of liver fibrosis models were observed. The effects of HIF-1α, CXCR4, PDGF-BB on the dedifferentiation of LSECs were investigated by using the inhibitors of HIF-1α, CXCR4 or PDGFR-β separately or transfecting with a CXCR4 knockdown lentiviral vector. In addition, the relationship between LSECs and HSCs was demonstrated by co-culture of LSECs and HSCs using the transwell chamber. ResultsCXCR4 upregulation and CXCR7 downregulation were accompanied by LSECs capillarization and HSCs activation both in CCl4-induced and BDL-induced fibrotic liver. In vitro, downregulation of HIF-1α significantly descreased CXCR4 and CD31 expression, and enhanced the expressions of CXCR7, CD44 and LYVE1. Downregulation of CXCR4 in LSECs significantly downregulated PDGF-BB, PDGFR-β and CD31, and enhanced CXCR7, CD44 and LYVE1 expression, while the expression of HIF-1α did not change significantly. STI571, a PDGF receptor inhibitor, could significantly downregulate PDGFR-β and increase the expression of CXCR7 to inhibit the dedifferentiation of LSECs. In addition, alleviateion the dedifferentiation of LSECs could decrease the expression of PDGFR-β of HSCs, then inhibiting the activation of HSCs. ConclusionsThis study revealed that HIF-1α/CXCR4/PDGF-BB/CXCR7 axis promoted the dedifferentiation of LSECs, consequently triggering HSCs activation and liver fibrosis.

Single-Cell Transcriptomics Unveils the Dedifferentiation Mechanism of Lung Adenocarcinoma Stem Cells

Lung adenocarcinoma (LUAD) is a major subtype of lung cancer, and its prognosis is still poor due to therapy resistance, metastasis, and recurrence. In recent years, increasing evidence has shown that the existence of lung cancer stem cells is responsible for the propagation, metastasis, therapy resistance, and recurrence of the tumor. During their transition to cancer stem cells, tumor cells need to inhibit cell differentiation and acquire invasive characteristics. However, our understanding of the property and role of such lung cancer stem cells is still limited. In this study, lung adenocarcinoma cancer stem cells (LCSCs) were enriched from the PC-9 cell line in a serum-free condition. PC-9 cells grew into spheres and showed higher survival rates when exposed to gefitinib: the drug used for the treatment of LUAD. Additionally, we found that the canonical stemness marker protein CD44 was significantly increased in the enriched LCSCs. Then, LCSCs were inoculated into the groin of nude mice for 1.5 months, and tumors were detected in the animals, indicating the strong stemness of the cells. After that, we performed single-cell RNA sequencing (scRNA-seq) on 7320 LCSCs and explored the changes in their transcriptomic signatures. We identified cell populations with a heterogeneous expression of cancer stem marker genes in LCSCs and subsets with different degrees of differentiation. Further analyses revealed that the activation of the FOXM1 (oncoprotein) transcription factor is a key factor in cell dedifferentiation, which enables tumor cells to acquire an epithelial-mesenchymal transition phenotype and increases the LCSC surface marker CD44. Moreover, we found that the combination of CD44, ABCG2, and ALCAM was a specific marker for LCSCs. In summary, this study identified the potential factors and molecular mechanisms underlying the stemness properties of LUAD cancer cells; it could also provide insight into developing novel and effective therapeutic approaches.

Diminished schwann cell repair responses play a role in delayed diabetes-associated wound healing.

Diabetes mellitus is the most common metabolic disease associated with impaired wound healing. Recently, Schwann cells (SCs), the glia of the peripheral nervous system, have been suggested to accelerate normal skin wound healing. However, the roles of SCs in diabetic wound healing are not fully understood. In this study, Full-thickness wounds were made in the dorsal skin of C57/B6 mice and db/db (diabetic) mice. Tissue samples were collected at different time points, and immunohistochemical and immunofluorescence analyses were performed to detect markers of de-differentiated SCs, including myelin basic protein, Sox 10, p75, c-Jun, and Ki67. In addition, in vitro experiments were performed using rat SC (RSC96) and murine fibroblast (L929) cell lines to examine the effects of high glucose conditions (50mM) on the de-differentiation of SCs and the paracrine effects of SCs on myofibroblast formation. Here, we found that, compared with that in normal mice, wound healing was delayed and SCs failed to rapidly activate a repair program after skin wound injury in diabetic mice. Furthermore, we found that SCs from diabetic mice displayed functional impairments in cell de-differentiation, cell-cycle re-entry, and cell migration. In vitro, hyperglycemia impaired RSC 96 cell de-differentiation, cell-cycle re-entry, and cell migration, as well as their paracrine effects on myofibroblast formation, including the secretion of TGF-β and Timp1. These results suggest that delayed wound healing in diabetes is due in part to a diminished SC repair response and attenuated paracrine effects on myofibroblast formation.

Induction of Somatic Embryogenesis in Plants: Different Players and Focus on WUSCHEL and WUS-RELATED HOMEOBOX (WOX) Transcription Factors.

In plants, other cells can express totipotency in addition to the zygote, thus resulting in embryo differentiation; this appears evident in apomictic and epiphyllous plants. According to Haberlandt's theory, all plant cells can regenerate a complete plant if the nucleus and the membrane system are intact. In fact, under in vitro conditions, ectopic embryos and adventitious shoots can develop from many organs of the mature plant body. We are beginning to understand how determination processes are regulated and how cell specialization occurs. However, we still need to unravel the mechanisms whereby a cell interprets its position, decides its fate, and communicates it to others. The induction of somatic embryogenesis might be based on a plant growth regulator signal (auxin) to determine an appropriate cellular environment and other factors, including stress and ectopic expression of embryo or meristem identity transcription factors (TFs). Still, we are far from having a complete view of the regulatory genes, their target genes, and their action hierarchy. As in animals, epigenetic reprogramming also plays an essential role in re-establishing the competence of differentiated cells to undergo somatic embryogenesis. Herein, we describe the functions of WUSCHEL-RELATED HOMEOBOX (WOX) transcription factors in regulating the differentiation-dedifferentiation cell process and in the developmental phase of in vitro regenerated adventitious structures.

A pro B cell population forms the apex of the leukemic hierarchy in Hoxa9/Meis1-dependent AML

Relapse is a major challenge to therapeutic success in acute myeloid leukemia (AML) and can be partly associated with heterogeneous leukemic stem cell (LSC) properties. In the murine Hoxa9/Meis1-dependent (H9M) AML model, LSC potential lies in three defined immunophenotypes, including Lin−cKit+ progenitor cells (Lin−), Gr1+CD11b+cKit+ myeloid cells, and lymphoid cells (Lym+). Previous reports demonstrated their interconversion and distinct drug sensitivities. In contrast, we here show that H9M AML is hierarchically organized. We, therefore, tracked the developmental potential of LSC phenotypes. This unexpectedly revealed a substantial fraction of Lin− LSCs that failed to regenerate Lym+ LSCs, and that harbored reduced leukemogenic potential. However, Lin− LSCs capable of producing Lym+ LSCs as well as Lym+ LSCs triggered rapid disease development suggestive of their high relapse-driving potential. Transcriptional analyses revealed that B lymphoid master regulators, including Sox4 and Bach2, correlated with Lym+ LSC development and presumably aggressive disease. Lentiviral overexpression of Sox4 and Bach2 induced dedifferentiation of H9M cells towards a lineage-negative state in vitro as the first step of lineage conversion. This work suggests that the potency to initiate a partial B lymphoid primed transcriptional program as present in infant AML correlates with aggressive disease and governs the H9M LSC hierarchy.

Ferroptotic stress facilitates smooth muscle cell dedifferentiation in arterial remodelling by disrupting mitochondrial homeostasis.

Smooth muscle cell (SMC) phenotypic switch from a quiescent 'contractile' phenotype to a dedifferentiated and proliferative state underlies the development of cardiovascular diseases (CVDs); however, our understanding of the mechanism is still incomplete. In the present study, we explored the potential role of ferroptosis, a novel nonapoptotic form of cell death, in SMC phenotypic switch and related neointimal formation. We found that ferroptotic stress was triggered in cultured dedifferentiated SMCs and arterial neointimal tissue of wire-injured mice. Moreover, pro-ferroptosis stress was activated in arterial neointimal tissue of clinical patients who underwent carotid endarterectomy. Blockade of ferroptotic stress via administration of a pharmacological inhibitor or by global genetic overexpression of glutathione peroxidase-4 (GPX4), a well-established anti-ferroptosis molecule, delayed SMC phenotype switch and arterial remodelling. Conditional SMC-specific gene delivery of GPX4 using adreno-associated virus in the carotid artery inhibited ferroptosis and prevented neointimal formation. Conversely, ferroptosis stress directly triggered dedifferentiation of SMCs. Transcriptomics analysis demonstrated that inhibition of ferroptotic stress mainly targets the mitochondrial respiratory chain and oxidative phosphorylation. Mechanistically, ferroptosis inhibition corrected the disrupted mitochondrial homeostasis in dedifferentiated SMCs, including enhanced mitochondrial ROS production, dysregulated mitochondrial dynamics, and mitochondrial hyperpolarization, and ultimately inhibited SMC phenotypic switch and growth. Copper-diacetyl-bisN4-methylthiosemicarbazone (CuATSM), an agent used for clinical molecular imaging and that potently inhibits ferroptosis, prevented SMC phenotypic switch, neointimal formation and arterial inflammation in mice. These results indicate that pro-ferroptosis stress is likely to promote SMC phenotypic switch during neointimal formation and imply that inhibition of ferroptotic stress may be a promising translational approach to treat CVDs with SMC phenotype switch.

Abstract B009: A SOX9-B7x axis safeguards dedifferentiated tumor cells from immunosurveillance to enable DCIS progression

Abstract Accumulating evidence has demonstrated a central role of dedifferentiation in carcinogenesis. In basal-like breast cancer mouse models, upregulation of SOX9 drives dedifferentiation of luminal progenitors to cells with luminal-basal hybrid features resembling embryonic multipotent mammary stem cells. Additionally, genetic inactivation of SOX9 inhibits dedifferentiation and blocks the progression of in situ tumors to malignant carcinoma. Whether the host immune system can recognize these dedifferentiated cells and how they eventually evade immunosurveillance are unclear. Here, we showed that SOX9 was necessary for limiting infiltrating T lymphocytes to protect dedifferentiated cells from immune surveillance in premalignant lesions of basal-like breast cancer. SOX9 deletion in spontaneous BLBC tumor models caused massive T lymphocyte infiltration and elimination of dedifferentiated cells. Antibody-mediated T cell depletion rescued the accumulation of dedifferentiated tumor cells in SOX9-deficient DCIS and its progression to invasive tumors. Mechanistically, through both stimulation of the STAT3 pathway and direct transcriptional activation, SOX9 upregulated the expression of the immune checkpoint B7x, which mediated the action of SOX9 in immunosuppression. In advanced tumors, B7x blockade reactivated antitumor immunity and effectively inhibited tumor growth. Furthermore, B7x depletion overcame the resistance to anti-PD-L1 immunotherapy. Collectively, these findings uncovered a dedifferentiation-associated immune evasion mechanism and demonstrated the therapeutic potential of targeting the SOX9-B7x pathway to eradicate dedifferentiated tumor cells for intercepting breast cancer progression. Citation Format: Yu Liu, Peter John, John R Christin, Maja Oktay, XingXing Zang, Wenjun Guo. A SOX9-B7x axis safeguards dedifferentiated tumor cells from immunosurveillance to enable DCIS progression [abstract]. In: Proceedings of the AACR Special Conference on Rethinking DCIS: An Opportunity for Prevention?; 2022 Sep 8-11; Philadelphia, PA. Philadelphia (PA): AACR; Can Prev Res 2022;15(12 Suppl_1): Abstract nr B009.

Cyclin G1 induces maladaptive proximal tubule cell dedifferentiation and renal fibrosis through CDK5 activation.

Acute kidney injury (AKI) occurs in approximately 13% of hospitalized patients and predisposes patients to chronic kidney disease (CKD) through the AKI-to-CKD transition. Studies from our laboratory and others have demonstrated that maladaptive repair of proximal tubule cells (PTCs), including induction of dedifferentiation, G2/M cell cycle arrest, senescence, and profibrotic cytokine secretion, is a key process promoting AKI-to-CKD transition, kidney fibrosis, and CKD progression. The molecular mechanisms governing maladaptive repair and the relative contribution of dedifferentiation, G2/M arrest, and senescence to CKD remain to be resolved. We identified cyclin G1 (CG1) as a factor upregulated in chronically injured and maladaptively repaired PTCs. We demonstrated that global deletion of CG1 inhibits G2/M arrest and fibrosis. Pharmacological induction of G2/M arrest in CG1-knockout mice, however, did not fully reverse the antifibrotic phenotype. Knockout of CG1 did not alter dedifferentiation and proliferation in the adaptive repair response following AKI. Instead, CG1 specifically promoted the prolonged dedifferentiation of kidney tubule epithelial cells observed in CKD. Mechanistically, CG1 promotes dedifferentiation through activation of cyclin-dependent kinase 5 (CDK5). Deletion of CDK5 in kidney tubule cells did not prevent G2/M arrest but did inhibit dedifferentiation and fibrosis. Thus, CG1 and CDK5 represent a unique pathway that regulates maladaptive, but not adaptive, dedifferentiation, suggesting they could be therapeutic targets for CKD.

Comparative Study of the Osteogenic Differentiation Potential of Adipose Tissue-Derived Stromal Cells and Dedifferentiated Adipose Cells of the Same Tissue Origin under Pro and Antioxidant Conditions.

Adipose tissue-derived stromal cells (ASCs) play an important role in various therapeutic approaches to bone regeneration. However, such applications become challenging when the obtained cells show a functional disorder, e.g., an impaired osteogenic differentiation potential (ODP). In addition to ASCs, human adipose tissue is also a source for another cell type with therapeutic potential, the dedifferentiated fat cells (DFATs), which can be obtained from mature adipocytes. Here, we for the first time compared the ODPs of each donors ASC and DFAT obtained from the same adipose tissue sample as well as the role of oxidative stress or antioxidative catalase on their osteogenic outcome. Osteogenic potential of ASC and DFAT from nine human donors were compared in vitro. Flow cytometry, staining for calcium accumulation with alizarin red, alkaline phosphatase assay and Western blots were used over an osteogenic induction period of up to 14 days. H2O2 was used to induce oxidative stress and catalase was used as an antioxidative measure. We have found that ASC and DFAT cultures' ODPs are nearly identical. If ASCs from an adipose tissue sample showed good or bad ODP, so did the corresponding DFAT cultures. The inter-individual variability of the donor ODPs was immense with a maximum factor of about 20 and correlated neither with the age nor the sex of the donors of the adipose tissue. Oxidative stress in the form of exogenously added H2O2 led to a significant ODP decrease in both cell types, with this ODP decrease being significantly lower in DFAT cultures than in the corresponding ASC cultures. Regardless of the individual cell culture-specific ODP, however, exogenously applied catalase led to an approx. 2.5-fold increase in osteogenesis in the ASC and DFAT cultures. Catalase appears to be a potent pro-osteogenic factor, at least in vitro. A new finding that points to innovative strategies and therapeutic approaches in bone regeneration. Furthermore, our results show that DFATs behave similarly to ASCs of the same adipose tissue sample with respect to ODPs and could therefore be a very attractive and readily available source of multipotent stem cells in bone regenerative therapies.

Transcriptome Dynamics of Rooting Zone and Leaves during In Vitro Adventitious Root Formation in Eucalyptus nitens.

Wood properties and agronomic traits associated with fast growth and frost tolerance make Eucalyptus nitens a valuable forest alternative. However, the rapid age-related decline in the adventitious root (AR) formation (herein, meaning induction, initiation, and expression stages) limits its propagation. We analyzed transcriptomic profile variation in leaves and stem bases during AR induction of microcuttings to elucidate the molecular mechanisms involved in AR formation. In addition, we quantified expressions of candidate genes associated with recalcitrance. We delimited the ontogenic phases of root formation using histological techniques and Scarecrow and Short-Root expression quantification for RNA sequencing sample collection. We quantified the gene expressions associated with root meristem formation, auxin biosynthesis, perception, signaling, conjugation, and cytokinin signaling in shoots harvested from 2- to 36-month-old plants. After IBA treatment, 702 transcripts changed their expressions. Several were involved in hormone homeostasis and the signaling pathways that determine cell dedifferentiation, leading to root meristem formation. In part, the age-related decline in the rooting capacity is attributable to the increase in the ARR1 gene expression, which negatively affects auxin homeostasis. The analysis of the transcriptomic variation in the leaves and rooting zones provided profuse information: (1) To elucidate the auxin metabolism; (2) to understand the hormonal and signaling processes involved; (3) to collect data associated with their recalcitrance.

The Revelation of Continuously Organized, Co-Overexpressed Protein-Coding Genes with Roles in Cellular Communications in Breast Cancer

Many human cancers, including breast cancer, are polygenic and involve the co-dysregulation of multiple regulatory molecules and pathways. Though the overexpression of genes and amplified chromosomal regions have been closely linked in breast cancer, the notion of the co-upregulation of genes at a single locus remains poorly described. Here, we describe the co-overexpression of 34 continuously organized protein-coding genes with diverse functions at 8q.24.3(143437655-144326919) in breast and other cancer types, the CanCord34 genes. In total, 10 out of 34 genes have not been reported to be overexpressed in breast cancer. Interestingly, the overexpression of CanCord34 genes is not necessarily associated with genomic amplification and is independent of hormonal or HER2 status in breast cancer. CanCord34 genes exhibit diverse known and predicted functions, including enzymatic activities, cell viability, multipotency, cancer stem cells, and secretory activities, including extracellular vesicles. The co-overexpression of 33 of the CanCord34 genes in a multivariant analysis was correlated with poor survival among patients with breast cancer. The analysis of the genome-wide RNAi functional screening, cell dependency fitness, and breast cancer stem cell databases indicated that three diverse overexpressed CanCord34 genes, including a component of spliceosome PUF60, a component of exosome complex EXOSC4, and a ribosomal biogenesis factor BOP1, shared roles in cell viability, cell fitness, and stem cell phenotypes. In addition, 17 of the CanCord34 genes were found in the microvesicles (MVs) secreted from the mesenchymal stem cells that were primed with MDA-MB-231 breast cancer cells. Since these MVs were important in the chemoresistance and dedifferentiation of breast cancer cells into cancer stem cells, these findings highlight the significance of the CanCord34 genes in cellular communications. In brief, the persistent co-overexpression of CanCord34 genes with diverse functions can lead to the dysregulation of complementary functions in breast cancer. In brief, the present study provides new insights into the polygenic nature of breast cancer and opens new research avenues for basic, preclinical, and therapeutic studies in human cancer.

Prolyl-hydroxylase inhibitor-induced regeneration of alveolar bone and soft tissue in a mouse model of periodontitis through metabolic reprogramming.

Bone injuries and fractures reliably heal through a process of regeneration with restoration to original structure and function when the gap between adjacent sides of a fracture site is small. However, when there is significant volumetric loss of bone, bone regeneration usually does not occur. In the present studies, we explore a particular case of volumetric bone loss in a mouse model of human periodontal disease (PD) in which alveolar bone surrounding teeth is permanently lost and not replaced. This model employs the placement a ligature around the upper second molar for 10 days leading to inflammation and bone breakdown and faithfully replicates the bacterially-induced inflammatory etiology of human PD to induce bone degeneration. After ligature removal, mice are treated with a timed-release formulation of a small molecule inhibitor of prolylhydroxylases (PHDi; 1,4-DPCA) previously shown to induce epimorphic regeneration of soft tissue in non-regenerating mice. This PHDi induces high expression of HIF-1α and is able to shift the metabolic state from OXPHOS to aerobic glycolysis, an energetic state used by stem cells and embryonic tissue. This regenerative response was completely blocked by siHIF1a. In these studies, we show that timed-release 1,4-DPCA rapidly and completely restores PD-affected bone and soft tissue with normal anatomic fidelity and with increased stem cell markers due to site-specific stem cell migration and/or de-differentiation of local tissue, periodontal ligament (PDL) cell proliferation, and increased vascularization. In-vitro studies using gingival tissue show that 1,4-DPCA indeed induces de-differentiation and the expression of stem cell markers but does not exclude the role of migrating stem cells. Evidence of metabolic reprogramming is seen by the expression of not only HIF-1a, its gene targets, and resultant de-differentiation markers, but also the metabolic genes Glut-1, Gapdh, Pdk1, Pgk1 and Ldh-a in jaw periodontal tissue.

Metabolic Changes Associated With Cardiomyocyte Dedifferentiation Enable Adult Mammalian Cardiac Regeneration.

Cardiac regeneration after injury is limited by the low proliferative capacity of adult mammalian cardiomyocytes (CMs). However, certain animals readily regenerate lost myocardium through a process involving dedifferentiation, which unlocks their proliferative capacities. We bred mice with inducible, CM-specific expression of the Yamanaka factors, enabling adult CM reprogramming and dedifferentiation in vivo. Two days after induction, adult CMs presented a dedifferentiated phenotype and increased proliferation in vivo. Microarray analysis revealed that upregulation of ketogenesis was central to this process. Adeno-associated virus-driven HMGCS2 overexpression induced ketogenesis in adult CMs and recapitulated CM dedifferentiation and proliferation observed during partial reprogramming. This same phenomenon was found to occur after myocardial infarction, specifically in the border zone tissue, and HMGCS2 knockout mice showed impaired cardiac function and response to injury. Finally, we showed that exogenous HMGCS2 rescues cardiac function after ischemic injury. Our data demonstrate the importance of HMGCS2-induced ketogenesis as a means to regulate metabolic response to CM injury, thus allowing cell dedifferentiation and proliferation as a regenerative response.

Transection injury differentially alters the proteome of the human sural nerve.

Regeneration after severe peripheral nerve injury is often poor. Knowledge of human nerve regeneration and the growth microenvironment is greatly lacking. We aimed to identify the regenerative proteins in human peripheral nerve by comparing the proteome before and after a transection injury. In a unique study design, we collected closely matched samples of naïve and injured sural nerve. Naïve and injured (two weeks after injury) samples were analyzed using mass spectrometry and immunoassays. We found significantly altered levels following the nerve injury. Mass spectrometry revealed that injury samples had 568 proteins significantly upregulated and 471 significantly downregulated compared to naïve samples (q-value ≤ 0.05 and Z ≥ |2| (log2)). We used Gene Ontology (GO) pathway overrepresentation analysis to highlight groups of proteins that were significantly upregulated or downregulated with injury-induced degeneration and regeneration. Significant protein changes in key pathways were identified including growth factor levels, Schwann cell de-differentiation, myelination downregulation, epithelial-mesenchymal transition (EMT), and axonal regeneration pathways. The proteomes of the uninjured nerve compared to the degenerating/regenerating nerve may reveal biomarkers to aid in the development of repair strategies such as infusing supplemental trophic factors and in monitoring neural tissue regeneration.