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Background: Smooth muscle cells (SMCs) play significant roles in atherosclerosis via phenotypic switching, a pathological process in which SMC dedifferentiation, migration, and transdifferentiation into other cell types. Yet how SMCs contribute to the pathophysiology of atherosclerosis remains elusive. Methods: To reveal the trajectories of SMC transdifferentiation during atherosclerosis and to identify molecular targets for disease therapy, we combined SMC fate mapping and single-cell RNA sequencing of both mouse and human atherosclerotic plaques. We also performed cell biology experiments on isolated SMC-derived cells, conducted integrative human genomics, and used pharmacological studies targeting SMC-derived cells both in vivo and in vitro. Results: We found that SMCs transitioned to an intermediate cell state during atherosclerosis, which was also found in human atherosclerotic plaques of carotid and coronary arteries. SMC-derived intermediate cells, termed “SEM” cells (stem cell, endothelial cell, monocyte), were multipotent and could differentiate into macrophage-like and fibrochondrocyte-like cells, as well as return toward the SMC phenotype. Retinoic acid (RA) signaling was identified as a regulator of SMC to SEM cell transition, and RA signaling was dysregulated in symptomatic human atherosclerosis. Human genomics revealed enrichment of genome-wide association study signals for coronary artery disease in RA signaling target gene loci and correlation between coronary artery disease risk alleles and repressed expression of these genes. Activation of RA signaling by all-trans RA, an anticancer drug for acute promyelocytic leukemia, blocked SMC transition to SEM cells, reduced atherosclerotic burden, and promoted fibrous cap stability. Conclusions: Integration of cell-specific fate mapping, single-cell genomics, and human genetics adds novel insights into the complexity of SMC biology and reveals regulatory pathways for therapeutic targeting of SMC transitions in atherosclerotic cardiovascular disease.

Context.— Dedifferentiated chondrosarcoma is a rare adult bone tumor with a dismal prognosis and is composed of a conventional chondrosarcoma juxtaposed to high-grade nonchondrogenic sarcoma. Dedifferentiated chondrosarcomas may represent tumor progression from a differentiated to a primitive histotype. Objective.— To determine the genetic and molecular events that drive progression from a conventional chondrosarcoma to high grade nonchondrogenic sarcoma. Design.— We analyzed the genomic landscape of paired conventional and dedifferentiated components of 11 dedifferentiated chondrosarcoma using targeted next-generation DNA sequencing with immunohistochemical validation. Clinical, radiographic, and pathologic features of tumors were reviewed. Capture-based DNA sequencing targeting the coding regions of 479 cancer genes and select introns was performed. Results.— The tumors arose in the femur (n = 4; 36%), scapula (n = 3; 27%), pelvis (n = 3; 27%), and humerus (n = 1; 9%) of 7 men (64%) and 4 women (36%; median age, 61 years). DNA was adequate for sequencing from all 11 dedifferentiated components (100%) and 9 paired conventional chondrosarcoma components (82%). All tumors (100%) harbored either IDH1 p.R132 or IDH2 p.R172S hotspot mutations. Seven tumors (64%) displayed COL2A1 alterations. TERT promoter mutations were present in 5 of 9 pairs (56%) and 2 (22%) additional unpaired dedifferentiated components. IDH1/2, COL2A1, and TERT mutations were identical in both components of the paired samples. Pathogenic missense or truncating mutations in TP53 and large-scale copy number alterations were more common in dedifferentiated components than in those of matched conventional components. Conclusions.— The results support IDH1/2, COL2A1, and TERT promoter mutations being common in dedifferentiated chondrosarcoma and as likely early events in progression, whereas inactivating mutation of TP53 and high-level copy number alterations may be later events in the dedifferentiated phenotype.

PDA is a major cause of US cancer-related deaths. Oncogenic Kras presents in 90% of human PDAs. Kras mutations occur early in pre-neoplastic lesions but are insufficient to cause PDA. Other contributing factors early in disease progression include chronic pancreatitis, alterations in epigenetic regulators, and tumor suppressor gene mutation. GPCRs activate heterotrimeric G-proteins that stimulate intracellular calcium and oncogenic Kras signaling, thereby promoting pancreatitis and progression to PDA. By contrast, Rgs proteins inhibit Gi/q-coupled GPCRs to negatively regulate PDA progression. Rgs16::GFP is expressed in response to caerulein-induced acinar cell dedifferentiation, early neoplasia, and throughout PDA progression. In genetically engineered mouse models of PDA, Rgs16::GFP is useful for pre-clinical rapid in vivo validation of novel chemotherapeutics targeting early lesions in patients following successful resection or at high risk for progressing to PDA. Cultured primary PDA cells express Rgs16::GFP in response to cytotoxic drugs. A histone deacetylase inhibitor, TSA, stimulated Rgs16::GFP expression in PDA primary cells, potentiated gemcitabine and JQ1 cytotoxicity in cell culture, and Gem + TSA + JQ1 inhibited tumor initiation and progression in vivo. Here we establish the use of Rgs16::GFP expression for testing drug combinations in cell culture and validation of best candidates in our rapid in vivo screen.

Interactions between tissues such as epicardial adipose (EAT), and myocardial tissues is important in the pathogenesis of heart failure. Changes in adipose tissues in obesity or diabetes impair preadipocyte differentiation. Furthermore, proinflammatory cytokine secretion is higher in preadipocytes than in mature adipocytes in diabetes and obesity. However, how undifferentiated cells committed to the adipose lineage directly influence cardiomyocytes is not yet understood. We used human-derived dedifferentiated fat (DFAT) cells as models of undifferentiated cells committed to an adipose lineage. Here, we evaluated the effects of soluble factor interactions in indirect cocultures of DFAT cells and induced pluripotent stem cell-derived cardiomyocytes. Our RNA sequencing findings showed that these interactions were predominantly inflammatory responses. Furthermore, proinflammatory cytokines secreted by DFAT cells reduced myocardial functions such as contraction frequency and catecholamine sensitivity, and simultaneously increased apoptosis, decreased antioxidative stress tolerance, and reduced oxygen consumption rates in cardiomyocytes. These adverse effects might be attributable to monocyte chemoattractant protein-1, chemokine (C-X-C motif) ligands 1 (CXCL1), and 12, granulocyte colony-stimulating factor, interleukins 6 and 8, macrophage migration inhibitory factor (MIF), and plasminogen activator inhibitor 1-A among the proinflammatory mediators secreted by DFAT cells. Our results could be useful for understanding the pathogenesis of EAT-related heart failure in terms of the involvement of undifferentiated cells committed to the adipose lineage. Furthermore, we suggest the importance of focusing on surrounding adipose tissues as a strategy with which to maximize the survival and function of transplanted stem cell-derived cardiomyocytes.

Immunotherapy has revolutionized cancer treatment, yet most patients do not respond. While tumor antigens are needed for effective immunotherapy, a favorable tumor immune microenvironment is also critical. In this review, we discuss emerging evidence that tumor cells exploit cellular plasticity and dedifferentiation programs to avoid immune surveillance, which in turn drives metastatic dissemination and resistance to immunotherapy. A deeper understanding of these programs may provide novel opportunities to enhance the efficacy of existing immunotherapies.

Small intestinal neuroendocrine tumors (SI-NET) are serotonin-secreting well-differentiated neuroendocrine tumors of putative enterochromaffin (EC) cell origin. Recent studies recognize a subset of EC cells that is label-retaining at the +4 position in the crypt and functions as a reserve intestinal stem cell. Importantly, this +4 reserve EC cell subset not only contributes to regeneration of the intestinal epithelium during injury and inflammation but also to basal crypt homeostasis at a constant rate. The latter function suggests that the +4 EC cell subset serves as an active reserve stem cell via a constant rate of dedifferentiation. Characterization of early tumor formation of SI-NET, observed as crypt-based EC cell clusters in many cases of familial SI-NETs, suggests that the +4 active reserve EC cell subset is the cell of origin. This newly discovered active reserve stem cell property of EC cells can account for unique biological mechanisms and processes associated with the genesis and development of SI-NETs. The recognition of this property of the +4 active reserve EC cell subset may provide novel opportunities to explore NETs in the gastrointestinal tract and other organs.

Molecular profiling was performed in 50 problematic ovarian sex cord-stromal tumours (SCSTs) most of which were seen in consultation. Following analysis, 17 were classified as adult granulosa cell tumour (AGCT), 16 of which showed a FOXL2 sequence variant (mutation); the initial favoured diagnosis in five of the cases was benign thecoma/fibrothecoma. Thirteen tumours ultimately classified as cellular fibroma or thecoma were FOXL2 sequence variant negative which was helpful in excluding AGCT. All six Sertoli-Leydig cell tumours (SLCTs) demonstrated DICER1 'hot spot' sequence variants, and one case each of AGCT and SLCT showed high grade histological transformation associated with a concurrent TP53 sequence variant. All eight unclassified SCSTs were negative for FOXL2 mutations and the six tested cases were DICER1 wild type; however, three tumours demonstrated MET, CTNNB1 or TP53 sequence variants. Four cases were classified as juvenile granulosa cell tumour, and one of these harboured a GNAS sequence variant. The single gynandroblastoma and microcystic stromal tumours in the series demonstrated FOXL2 and CTNNB1 alterations, respectively. In summary, molecular analysis aids in accurate classification of challenging ovarian SCSTs and sometimes leads to revision of the favoured provisional diagnosis. TP53 sequence variants may be associated with dedifferentiation in both SLCTs and AGCTs.

Switch/sucrose nonfermenting complex subunits, such as BRG1, INI1, and ARID1B, are inactivated in a subset of endometrial undifferentiated carcinoma and dedifferentiated carcinoma (DC). Limited information is currently available on their prevalence in other subtypes or the nosological status of endometrial carcinoma with their deficiencies. This study immunohistochemically examined the expression status of BRG1, INI1, and ARID1B using 570 archived cases of endometrial carcinoma and carcinosarcoma resected at a single institution. We identified 1 BRG1-deficient undifferentiated carcinoma, 8 BRG1/INI1/ARID1B-deficient DC, and 3 BRG1-deficient clear-cell carcinomas. None of the cases of endometrioid and serous carcinomas or carcinosarcoma showed deficiencies of these subunits. We then compared 8 BRG1/INI1/ARID1B-deficient DC with 6 BRG1/INI1/ARID1B-intact DC and 28 carcinosarcomas, the latter of which was often confused with DC. Histologically, BRG1/INI1/ARID1B-intact and BRG1/INI1/ARID1B-deficient DC shared a monotonous solid appearance with rhabdoid and epithelioid cells and a myxoid stroma; however, abrupt keratinization and cell spindling was absent in BRG1/INI1/ARID1B-deficient tumors. The median overall survival of patients with BRG1/INI1/ARID1B-deficient DC was 3.8 months, which was worse than those with BRG1/INI1/ARID1B-intact DC (P=0.008) and with carcinosarcoma (P=0.004). BRG1/INI1/ARID1B-deficient DC may be a separate entity with an aggressive behavior to be distinguished from BRG1/INI1/ARID1B-intact DC and carcinosarcoma. Regarding clear-cell carcinoma (n=12), BRG1 deficiency appeared to be mutually exclusive with abnormal ARID1A, BRM, and p53 expression. Further studies are needed to clarify whether BRG1 deficiency plays a role in the pathogenesis of clear-cell carcinoma.

Pygopus 2 (Pygo2) is a coactivator of Wnt/β-catenin signaling that can bind bi- or trimethylated lysine 4 of histone-3 (H3K4me2/3) and participate in chromatin reading and writing. It remains unknown whether the Pygo2–H3K4me2/3 association has a functional relevance in breast cancer progression in vivo. To investigate the functional relevance of histone-binding activity of Pygo2 in malignant progression of breast cancer, we generated a knock-in mouse model where binding of Pygo2 to H3K4me2/3 was rendered ineffective. Loss of Pygo2–histone interaction resulted in smaller, differentiated, and less metastatic tumors, due, in part, to decreased canonical Wnt/β-catenin signaling. RNA- and ATAC-sequencing analyses of tumor-derived cell lines revealed downregulation of TGFβ signaling and upregulation of differentiation pathways such as PDGFR signaling. Increased differentiation correlated with a luminal cell fate that could be reversed by inhibition of PDGFR activity. Mechanistically, the Pygo2–histone interaction potentiated Wnt/β-catenin signaling, in part, by repressing the expression of Wnt signaling antagonists. Furthermore, Pygo2 and β-catenin regulated the expression of miR-29 family members, which, in turn, repressed PDGFR expression to promote dedifferentiation of wild-type Pygo2 mammary epithelial tumor cells. Collectively, these results demonstrate that the histone binding function of Pygo2 is important for driving dedifferentiation and malignancy of breast tumors, and loss of this binding activates various differentiation pathways that attenuate primary tumor growth and metastasis formation. Interfering with the Pygo2–H3K4me2/3 interaction may therefore serve as an attractive therapeutic target for metastatic breast cancer. Significance: Pygo2 represents a potential therapeutic target in metastatic breast cancer, as its histone-binding capability promotes β-catenin–mediated Wnt signaling and transcriptional control in breast cancer cell dedifferentiation, EMT, and metastasis.

The aryl hydrocarbon receptor (AHR) is a ligand binding-transcription factor of the basic helix-loop-helix family regulating multiple cellular functions such as differentiation, cell cycle, apoptosis, and inflammatory reactions. In neoplastic diseases, the AHR has been described to modulate proliferation and differentiation in dichotomous ways, either inhibiting or augmenting the growth of tumors. The precise role of AHR in melanoma is mostly unknown. Here, we report a functional effect of AHR activation on inflammation-induced melanoma cell dedifferentiation and the development of lung metastases in a mouse model. Via in silico analyses of "The Cancer Genome Atlas" human melanoma cohort, we detected a correlation between AHR expression levels and a dedifferentiated melanoma cell phenotype with an invasive gene signature, which we were able to functionally recapitulate in a panel of human melanoma cell lines. Both human and mouse melanoma cell lines upregulated AHR expression after inflammatory stimulation with tumor necrosis factor-α (TNF-α). Activation of AHR in human and mouse melanoma cell lines with the endogenous ligand formylindolo(3,2-b)carbazole (FICZ) promoted inflammation-induced dedifferentiation in vitro. Importantly, mouse melanoma cells with CRISPR/Cas9-mediated disruption of the AHR gene showed impaired in vivo tumor growth after transplantation in the skin as well as decreased numbers of spontaneous lung metastases. Taken together, our results demonstrate a functional role for AHR expression in melanoma development and metastatic progression. This provides a scientific basis for future experiments that further dissect the underlying molecular mechanisms and assess the potential for AHR inhibition as part of multimodal melanoma treatment strategies.

Solitary fibrous tumor/hemangiopericytoma is a mesenchymal tumor that originates from a common NAB2-STAT6 fusion gene and is known to very rarely demonstrate dedifferentiation in the pattern of local recurrence or distant metastasis. Here we describe for the first time a rare case of intracranial dedifferentiated solitary fibrous tumor/hemangiopericytoma with osteosarcoma components that developed in an 84-year-old man after frequent gamma knife radiosurgery over a 14-year period. We performed tumor-debulking and gamma knife radiosurgery, but unfortunately the patient died shortly after the development of dedifferentiation. There is no established treatment for dedifferentiated cases due to the rare histology and limited published data, and therefore further accumulation of histological and genetic profiles is necessary to develop novel target gene therapies.

Restenosis, re-narrowing of arterial lumen following intervention for cardiovascular disease, remains a major issue limiting the long-term therapeutic efficacy of treatment. The signaling molecules, TGFβ (transforming growth factor-beta) and Smad3, play important roles in vascular restenosis, but very little is yet known about the down-stream dynamics in global protein expression and phosphorylation. Here, we develop a highly multiplexed quantitative proteomic and phosphoproteomic strategy employing 12-plex N,N-dimethyl leucine (DiLeu) isobaric tags and The DiLeu Tool software to globally assess protein expression and phosphorylation changes in smooth muscle cells (SMCs) treated with TGFβ/Smad3 and/or SDF-1α (stromal cell-derived factor). A total of 4086 proteins were quantified in the combined dataset of proteome and phosphoproteome across 12-plex DiLeu-labeled SMC samples. 2317 localized phosphorylation sites were quantified, corresponding to 1193 phosphoproteins. TGFβ/Smad3 induced up-regulation of 40 phosphosites and down-regulation of 50 phosphosites, and TGFβ/Smad3-specific SDF-1α exclusively facilitated up-regulation of 27 phosphosites and down-regulation of 47 phosphosites. TGFβ/Smad3 inhibited the expression of contractile-associated proteins including smooth muscle myosin heavy chain, calponin, cardiac muscle alpha-actin, and smooth muscle protein 22α. Gene ontology and pathway enrichment analysis revealed that elevated TGFβ/Smad3 activated cell proliferation and TGFβ signaling pathway, sequentially stimulating phosphorylation of CXCR4 (C-X-C chemokine receptor 4). SDF-1α/CXCR4 activated extracellular signal-regulating kinase signaling pathway and facilitated the expression of synthetic marker, osteopontin, which was validated through targeted analysis. These findings provide new insights into the mechanisms of TGFβ regulated SMC dedifferentiation, as well as new avenues for designing effective therapeutics for vascular disease.

BRAFV600E confers poor prognosis and is associated with a distinct subtype of colorectal cancer (CRC). Little is known, however, about the genetic events driving the initiation and progression of BRAFV600E mutant CRCs. Recent genetic analyses of CRCs indicate that BRAFV600E often coexists with alterations in the WNT- and p53 pathways, but their cooperation remains ill-defined. Therefore, we systematically compared small and large intestinal organoids from mice harboring conditional BraffloxV600E, Trp53LSL-R172H, and/or Apcflox/flox alleles. Using these isogenic models, we observe tissue-specific differences toward sudden BRAFV600E expression, which can be attributed to different ERK-pathway ground states in small and large intestinal crypts. BRAFV600E alone causes transient proliferation and suppresses epithelial organization, followed by organoid disintegration. Moreover, BRAFV600E induces a fetal-like dedifferentiation transcriptional program in colonic organoids, which resembles human BRAFV600E-driven CRC. Co-expression of p53R172H delays organoid disintegration, confers anchorage-independent growth, and induces invasive properties. Interestingly, p53R172H cooperates with BRAFV600E to modulate the abundance of transcripts linked to carcinogenesis, in particular within colonic organoids. Remarkably, WNT-pathway activation by Apc deletion fully protects organoids against BRAFV600E-induced disintegration and confers growth/niche factor independence. Still, Apc-deficient BRAFV600E-mutant organoids remain sensitive toward the MEK inhibitor trametinib, albeit p53R172H confers partial resistance against this clinically relevant compound. In summary, our systematic comparison of the response of small and large intestinal organoids to oncogenic alterations suggests colonic organoids to be better suited to model the human situation. In addition, our work on BRAF-, p53-, and WNT-pathway mutations provides new insights into their cooperation and for the design of targeted therapies.

Damage to the intestinal stem cell niche can result from mechanical stress, infections, chronic inflammation or cytotoxic therapies. Progenitor cells can compensate for insults to the stem cell population through dedifferentiation. The microenvironment modulates this regenerative response by influencing the activity of signaling pathways, including Wnt, Notch, and YAP/TAZ. For instance, mesenchymal cells and immune cells become more abundant after damage and secrete signaling molecules that promote the regenerative process. Furthermore, regeneration is influenced by the nutritional state, microbiome, and extracellular matrix. Here, we review how all these components cooperate to restore epithelial homeostasis in the intestine after injury.

Apatinib (YN968D1) is a small‐molecule tyrosine kinase inhibitor（TKI）which can inhibit the activity of vascular endothelial growth factor receptor‐2 (VEGFR‐2). It has been reported that apatinib has anti‐tumour effect of inhibiting proliferation and inducing apoptosis of a variety of solid tumour cells, whereas its effect on vascular smooth muscle cells (VSMC) remains unclear. This study investigated the effect of apatinib on phenotypic switching of arterial smooth muscle cells in vascular remodelling. Compared to the vehicle groups, mice that were performed carotid artery ligation injury and treated with apatinib produced a reduction in abnormal neointimal area. For in vitro experiment, apatinib administration inhibited VSMC proliferation, migration and reversed VSMC dedifferentiation with the stimulation of platelet‐derived growth factor type BB (PDGF‐BB).In terms of mechanism, with the preincubation of apatinib, the activations of PDGF receptor‐β (PDGFR‐β) and phosphoinositide‐specific phospholipase C‐γ1 (PLC‐γ1) induced by PDGF‐BB were inhibited in VSMCs. With the preincubation of apatinib, the phosphorylation of PDGFR‐β, extracellular signal‐related kinases (ERK1/2) and Jun amino‐terminal kinases (JNK) induced by PDGF‐BB were also inhibited in rat vascular smooth muscle cell line A7r5. Herein, we found that apatinib attenuates phenotypic switching of arterial smooth muscle cells induced by PDGF‐BB in vitro and vascular remodelling in vivo. Therefore, apatinib is a potential candidate to treat vascular proliferative diseases.

Vascular smooth muscle cells (VSMCs) shift from a physiological contractile phenotype to an adverse proliferative or synthetic state, which is a major event leading to aortic disease. VSMCs are exposed to multiple mechanical signals from their microenvironment including vascular extracellular matrix (ECM) stiffness and stretch which regulate VSMC contraction. How ECM stiffness regulates the function and phenotype of VSMCs is not well understood. In this study, we introduce in vitro and in vivo models to evaluate the impact of ECM stiffnesses on VSMC function. Through unbiased transcriptome sequencing analysis, we detected upregulation of synthetic phenotype-related genes including osteopontin, matrix metalloproteinases, and inflammatory cytokines in VSMCs cultured using soft matrix hydrogels in vitro, suggesting VSMC dedifferentiation toward a synthetic phenotype upon ECM softening. For the in vivo model, the lysyl oxidase inhibitor β-aminopropionitrile monofumarate (BAPN) was administrated to disrupt the cross-linking of collagen to induce ECM softening. Consistently, decreased ECM stiffnesses promoted VSMC phenotypic switching to a synthetic phenotype as evidenced by upregulation of synthetic phenotype-related genes in the aortas of mice following BAPN treatment. Finally, BAPN-treated mice showed severe expansion and developed aortic dissection. Our study reveals the pivotal role of ECM softening in regulating the VSMC phenotype switch and provides a potential target for treating VSMC dysfunction and aortic dissection disease.

Purpose Our previous studies demonstrated that mature adipocyte-derived dedifferentiated fat (DFAT) cells possess similar multipotency as mesenchymal stem cells. Here, we examined the immunoregulatory potential of DFAT cells in vitro and the therapeutic effect of DFAT cell transplantation in a mouse inflammatory bowel disease (IBD) model. Methods The effect of DFAT cell co-culture on T cell proliferation and expression of immunosuppression-related genes in DFAT cells were evaluated. To create IBD, CD4+CD45RBhigh T cells were intraperitoneally injected into SCID mice. One week later, DFAT cells (1 × 105, DFAT group) or saline (Control group) were intraperitoneally injected. Subsequently bodyweight was measured every week and IBD clinical and histological scores were evaluated at 5 weeks after T cell administration. Results The T cell proliferation was inhibited by co-cultured DFAT cells in a cell density-dependent manner. Gene expression of TRAIL, IDO1, and NOS2 in DFAT cells was upregulated by TNFα stimulation. DFAT group improved IBD-associated weight loss, IBD clinical and histological scores compared to Control group. Conclusion DFAT cells possess immunoregulatory potential and the cell transplantation promoted recovery from colon damage and improved clinical symptoms in the IBD model. DFAT cells could play an important role in the treatment of IBD.

Spermatogonial stem cells (SSCs) are defined as the tissue-specific stem cells in the testes that produce sperm and support life-long spermatogenesis. A culture microenvironment could convert mouse SSCs from unipotent to pluripotent states; however, the underlying mechanism is unclear. NANOG has been considered the decisive transcriptional factor for pluripotency transition of stem cells, but NANOG is not expressed in SSCs. Here, we investigated whether NANOG overexpression could result in SSCs being converted into a pluripotent state. We found that rare NANOG-positive cells could be detected in spermatogonia, pachytene spermatocytes, and even round spermatids in mouse testes, and that the induction of NANOG could promote the proliferation of cultured SSCs in vitro and partially compensate for the role of the growth factor GDNF. In vivo allogeneic transplantation of NANOG-overexpressing germ cells did not yield any teratoma-like tissues, but regenerated normal colonies of spermatogenesis in the testes of recipient mice. Collectively, our data showed that overexpression of the pluripotency factor NANOG along did not dedifferentiate testis germline stem cells into a pluripotent state, suggesting that other genetic or epigenetic factors are involved in SSC reprogramming.

Smooth muscle cells (SMCs) are the major type of cells constituting arterial walls and play a role to maintain stiffness via producing extracellular matrix. Here, the loss and degenerative changes of SMCs become the major histopathological features of an intracranial aneurysm (IA), a major cause of subarachnoid hemorrhage. Considering the important role of SMCs and the loss of this type of cells in IA lesions, we in the present study subjected rats to IA models and examined how SMCs behave during disease progression. We found that, at the neck portion of IAs, SMCs accumulated underneath the internal elastic lamina according to disease progression and formed the intimal hyperplasia. As these SMCs were positive for a dedifferentiation marker, myosin heavy chain 10, and contained abundant mitochondria and rough endoplasmic reticulum, SMCs at the intimal hyperplasia were dedifferentiated and activated. Furthermore, dedifferentiated SMCs expressed some pro-inflammatory factors, suggesting the role in the formation of inflammatory microenvironment to promote the disease. Intriguingly, some SMCs at the intimal hyperplasia were positive for CD68 and contained lipid depositions, indicating similarity with atherosclerosis. We next examined a potential factor mediating dedifferentiation and recruitment of SMCs. Platelet derived growth factor (PDGF)-BB was expressed in endothelial cells at the neck portion of lesions where high wall shear stress (WSS) was loaded. PDGF-BB facilitated migration of SMCs across matrigel-coated pores in a transwell system, promoted dedifferentiation of SMCs and induced expression of pro-inflammatory genes in these cells in vitro. Because, in a stenosis model of rats, PDGF-BB expression was expressed in endothelial cells loaded in high WSS regions, and SMCs present nearby were dedifferentiated, hence a correlation existed between high WSS, PDGFB and dedifferentiation in vivo. In conclusion, dedifferentiated SMCs presumably by PDGF-BB produced from high WSS-loaded endothelial cells accumulate in the intimal hyperplasia to form inflammatory microenvironment leading to the progression of the disease.

A physical, noninvasive, and reversible means of controlling the nanoscale presentation of bioactive ligands is highly desirable for regulating and investigating the time-dependent responses of cells, including stem cells. Herein we report a magnetically actuated dynamic cell culture platform consisting of a soft hydrogel substrate conjugated with RGD-bearing magnetic nanoparticle (RGD-MNP). The downward/upward magnetic attraction conceals/promotes the presentation of the RGD-MNP in/on the soft hydrogel matrix, thereby inhibiting/enhancing the cell adhesion and mechanosensing-dependent differentiation. Meanwhile, the lateral magnetic attraction promotes the unidirectional migration of cells in the opposite direction on the hydrogel. Furthermore, cyclic switching between the "Exposed" and "Hidden" conditions induces the repeated cycles of differentiation/dedifferentiation of hMSCs which significantly enhances the differentiation potential of hMSCs. Our design approach capitalizes on the bulk biomaterial matrix as the macroscopic caging structure to enable dynamic regulation of cell-matrix interactions reversibly, which is hard to achieve by using conventional cell culture systems.

Liver cancer stem cells (CSCs) are involved in tumorigenesis, progression, drug resistance and recurrence of hepatocellular carcinoma (HCC). However, the underlying mechanism for the propagation of liver cancer stem cells was unclear. Herein, we observed miR-206 expression was reduced in both chemoresistant HCCs and recurrent HCCs from patients. A dramatically decrease of miR-206 was detected in cluster of differentiation 133 (CD133) or epithelial cell adhesion molecule (EpCAM)–positive liver CSCs and in CSC-enriched hepatoma spheres. Functional studies revealed that a forced expression of miR-206 inhibited liver CSCs expansion by suppressing the dedifferentiation of hepatoma cells and attenuating the self-renewal of liver CSCs. Mechanistically, bioinformatic and luciferase reporter analysis identified epidermal growth factor receptor (EGFR) as a direct target of miR-206. Moreover, miR-206 downregulated the expression of EGFR in liver CSCs. There was a significant inverse correlation between miR-206 and EGFR mRNA expression in HCC samples. Special EGFR inhibitor Gefitinib abolished the discrepancy in liver CSC proportion and the self-renewal capacity between miR-206 overexpression hepatoma cells and control cells, which further confirmed that EGFR was required in miR-206-inhibited liver CSCs expansion. Conclusion: miR-206 could suppress HCC cell dedifferentiation and liver CSCs expansion by targeting EGFR signaling.

Papillary thyroid carcinoma (PTC) tall cell variant (TCV) with squamous dedifferentiation is a rare entity. We present a case of 90-year-old woman who initially had a 2.8 cm conventional PTC in right lobe of thyroid who, couple decades later, had metastatic dedifferentiated PTC to right neck lymph nodes level II and IV with tall cell features; to right level IV and V lymph nodes with tall cell and squamous components, which recently presented exclusively as squamous cell carcinoma (SCC) metastasizing to lung. The squamous component in the lymph node and SCC in the lung were both positive for squamous marker p63 and PTC markers TTF1, PAX-8 and BRAF V600E while negative for thyroglobulin and p16. The papillary component was positive for TTF-1, BRAF V600E and P63 (majority); negative for thyroglobulin and p16. Final diagnoses were rendered based on combination of cytological features and immunohistochemical profiles. This report highlights the utilization of current biomarkers to distinguish between metastatic dedifferentiated PTC with squamous features and primary lung SCC, as well as the importance of recognizing this rare entity.

Introduction Establishment of hemochorial placenta is associated with development and remodelling of uterine vasculature at the maternal fetal interface. This results in calibration of high resistance uterine arteries to flaccid low resistance vessels resulting in increased blood flow to the placenta and fetus in humans and rodents. Mechanisms underlying these remodelling events are poorly understood. In this report, we examine regulation of vascular remodelling using vascular smooth muscle cell (VSMC) phenotype switching as a primary parameter. Methods Cellular dynamics was assessed by Immunofluorescence, qRT-PCR, western blotting in timed pregnant rat tissue. In vitro co-culture of trophoblast cells with vascular smooth muscle cells was used to understand regulation mechanism. Results Analysis of cellular dynamics on days 13.5, 16.5 and 19.5 of gestation in the rat metrial gland, the entry point of uterine arteries, revealed that invasion of trophoblast cells preceded disappearance of VSMC α-SMA, a contractile state marker. Co-culture of VSMCs with trophoblast cells in vitro recapitulated trophoblast-induced de-differentiation of VSMCs in vivo. Interestingly, co-culturing with trophoblast cells activated PDGFRβ signalling in VSMCs, an effect mediated by secreted PDGF-BB from trophoblast cells. Trophoblast cells failed to elicit its effect on VSMC de-differentiation upon inhibition of PDGFRβ signalling using a selective inhibitor. Moreover, co-culturing with trophoblast cells also led to substantial increase in Akt activation and a modest increase in Erk phosphorylation in VSMCs and this effect was abolished by PDGFRβ inhibition. Discussion Our results highlight that trophoblast cells direct VSMC phenotype switching and trophoblast derived PDGF-BB is one of the modulator.

Purpose Ligand-dependent corepressor (LCoR) and receptor-interacting protein 140 (RIP140/NRIP1) play an important role in the regulation of multiple oncogenic signaling pathways and the development of cancer. LCoR and RIP140 form a nuclear complex in breast cancer cells and are of prognostic value in further prostate and cervical cancer. The purpose of this study was to analyze the regulation of these proteins in the development of cervical intraepithelial neoplasia (CIN I–III). Methods Immunohistochemical analysis was obtained to quantify RIP140 and LCoR expression in formalin-fixed paraffin embedded tissue sections of cervical intraepithelial neoplasia samples. Tissue (n = 94) was collected from patients treated in the Department of Gynecology and Obstetrics, Ludwig-Maximilians-University of Munich, Germany, between 2002 and 2014. Correlations of expression levels with clinical outcome were carried out to assess for prognostic relevance in patients with CIN2 progression. Kruskal–Wallis test and Mann–Whitney U test were used for data analysis. Results Nuclear LCoR overexpression correlates significantly with CIN II progression. Nuclear RIP140 expression significantly increases and nuclear LCoR expression decreases with higher grading of cervical intraepithelial neoplasia. Cytoplasmic RIP140 expression is significantly higher in CIN III than in CIN I or CIN II. Conclusion A decrease of nuclear LCoR expression in line with an increase of dedifferentiation of CIN can be observed. Nuclear LCoR overexpression correlates with CIN II progression indicating a prognostic value of LCoR in cervical intraepithelial neoplasia. Nuclear and cytoplasmic RIP140 expression increases significantly with higher grading of cervical intraepithelial neoplasia underlining its potential role in the development of pre-cancerous lesions. These findings support the relevance of LCoR and RIP140 in the tumorigenesis indicating a possible role of LCoR and RIP140 as targets for novel therapeutic approaches in cervical intraepithelial neoplasia and cervical cancer.

The progression of cancer is accompanied by the acquisition of stemness features. Many stemness evaluation methods based on transcriptional profiles have been presented to reveal the relationship between stemness and cancer. However, instead of absolute stemness index values—the values with certain range—these methods gave the values without range, which makes them unable to intuitively evaluate the stemness. Besides, these indices were based on the absolute expression values of genes, which were found to be seriously influenced by batch effects and the composition of samples in the dataset. Recently, we have showed that the signatures based on the relative expression orderings (REOs) of gene pairs within a sample were highly robust against these factors, which makes that the REO-based signatures have been stably applied in the evaluations of the continuous scores with certain range. Here, we provided an absolute REO-based stemness index to evaluate the stemness. We found that this stemness index had higher correlation with the culture time of the differentiated stem cells than the previous stemness index. When applied to the cancer and normal tissue samples, the stemness index showed its significant difference between cancers and normal tissues and its ability to reveal the intratumor heterogeneity at stemness level. Importantly, higher stemness index was associated with poorer prognosis and greater oncogenic dedifferentiation reflected by histological grade. All results showed the capability of the REO-based stemness index to assist the assignment of tumor grade and its potential therapeutic and diagnostic implications.

Reduced expression of the key regulator of cardiac metabolism, transcription factor PPARα, in surgical samples of the auricles from patients with coronary heart disease and heart failure was detected by real-time quantitative PCR. These changes indicate reduced activity of this factor and a shift of energy metabolism from oxidative phosphorylation to glycolysis typical of dedifferentiated cells. Electron microscopy revealed dedifferentiated cardiomyocytes with disassembled contractile apparatus and disorganized sarcomeres. In the examined specimens from patients with heart failure, severe myocardial fibrosis was revealed.

Hypoxia affects the function of pancreatic β cells, and the molecular mechanism underlying hypoxia-related β cell dysfunction in human type 2 diabetes mellitus (T2DM) remains to be elucidated. In this study, by comparing the gene expression profiles of islets from nondiabetic and T2D subjects using gene chip array, we aimed to elucidate that hypoxia signaling pathways are activated in human T2DM islets. CoCl2 treatment, which was employed to mimic hypoxic stimulation in human islets, decreased insulin secretion, insulin content, and the functional gene expression of human islets. In parallel, the expression of mature β cell-disallowed genes was upregulated by CoCl2, including progenitor cell marker NGN3, β cell differentiation marker ALDH1A3, and genes that are typically inhibited in mature β cells, namely, GLUT1 and LDHA, indicating that CoCl2-mimicked hypoxia induced β cell dedifferentiation of human islets. This finding in human islets was confirmed in mouse β cell line NIT-1. By using Dimethyloxalylglycine (DMOG) to activate hypoxia-inducible factor-1α (HIF-1α) or siRNAs to knockdown HIF-1α, we found that HIF-1α was a key regulator of hypoxia-induced dedifferentiation of β cells by upregulating mature β cell-disallowed genes. Our findings suggested that HIF-1α activation might be an important contributor to β cell dedifferentiation in human T2DM islets, and HIF-1α-targeted therapies may have the potential to reverse β cell dedifferentiation of human T2DM islets.

Summary Primary gastrointestinal liposarcoma is rare, and information regarding this entity is largely based on single case studies. We report on 8 patients with primary liposarcoma of the gastrointestinal tract and review the pertinent literature. The cohort includes 6 men and 2 women who ranged in age from 51 to 81 years (median 68.5). Two tumors arose in the stomach, 4 in the small intestine, and 2 in the large intestine. Tumors ranged in size from 2.5 to 14.5 cm (median 7 cm), originated in the submucosa or muscularis propria of the intestinal wall, and frequently protruded into the bowel lumen, resulting in mucosal ulceration and luminal obstruction. Six tumors were dedifferentiated liposarcomas, and 2 were well-differentiated liposarcoma. Surgical excision was performed on all tumors except for 1 case of dedifferentiated liposarcoma. On follow-up, 1 patient with dedifferentiated liposarcoma developed a lytic sacral lesion suspicious for metastasis 4 months after resection of the primary, and another underwent marginal resection and presented with recurrence 4 years later, had tumor re-resection, and was considered disease-free at 6 weeks postsurgery. A third patient with dedifferentiated liposarcoma was alive with unknown disease status at 17 months following surgery, and another patient with dedifferentiated liposarcoma was alive without evidence of disease at 30 months following surgery. No follow-up information on the remaining patients is available. Overall, liposarcomas of the intestinal tract are most frequently high-grade dedifferentiated tumors that are biologically aggressive and require surgical excision with widely negative margins to help reduce the risk of local recurrence and dissemination. Important in the differential diagnosis is malignant gastrointestinal stromal tumor. Care must be taken not to misdiagnose one entity for the other because the correct diagnosis carries important therapeutic implications.

As a novel class of noncoding RNAs, microRNAs (miRNAs) can effectively silence their target genes at the posttranscriptional level. Various biological processes, such as cell proliferation, differentiation, and motility, are regulated by miRNAs. In different diseases and different stages of disease, miRNAs have various expression patterns, which makes them candidate prognostic markers and therapeutic targets. Abnormal miRNA expression has been detected in numerous neoplastic diseases in humans, which indicates the potential role of miRNAs in tumorigenesis. Previous studies have indicated that miRNAs are involved in nearly the entire process of tumor development. MicroRNA‐302a, miR‐302b, miR‐302c, miR‐302d, and miR‐367 are members of the miR‐302/367 cluster that plays various biological roles in diverse neoplastic diseases by targeting different genes. These miRNAs have been implicated in several unique characteristics of cancer, including the evasion of growth suppressors, the sustained activation of proliferative signaling, the evasion of cell death and senescence, and the regulation of angiogenesis, invasion, and metastasis. This review provides a critical overview of miR‐302/367 cluster dysregulation and the subsequent effects in cancer and highlights the vast potential of members of this cluster as therapeutic targets and novel biomarkers., The microRNA (miR)‐302/367 cluster has been implicated in several peculiarities of cancer including evading growth suppressors, sustaining proliferative signaling, and evading cell death and senescence, angiogenesis, invasion and metastasis. This review provides a critical overview of miR‐302/367 cluster dysregulation and the subsequent effects in cancer and demonstrates the indispensable roles of this cluster as therapeutic targets and novel biomarkers at current state researches.

Over a decade of research has confirmed the critical role of cancer stem-like cells (CSCs) in tumor initiation, chemoresistance, and metastasis. Increasingly, CSC hierarchies have begun to be defined with some recurring themes. This includes evidence that these hierarchies are 'flexible,' with both cell state transitions and dedifferentiation events possible. These findings pose therapeutic hurdles and opportunities. Here, we review cancer stem cell hierarchies and their interactions with the tumor microenvironment. We also discuss the current therapeutic approaches designed to target CSC hierarchies and initial clinical trial results for CSC targeting agents. While cancer stem cell targeted therapies are still in their infancy, we are beginning to see encouraging results that suggest a positive outlook for CSC-targeting approaches.

During hematogenous metastasis, cancer cells escape from primary lesions and enter into the circulatory system, and only a few can colonize distant organs. However, the mechanism of cell survival and metastasis in the hematopoietic environment remains unclear. Angiorrhea is the character of pathological neovascularization in malignant tumors and commonly detected in osteosarcoma (OS), a bone tumor that prefers circulatory metastasis. In the present study, we focused on the notable role of serum albumin, the highest content in blood plasma, on OS progression. Our results indicated that serum albumin might act as a barrier against exogenous cancer cells during hematogenous metastasis. OS cells with high metastatic potential could gradually obtain strong viability through dedifferentiation under the effect of serum albumin in the angiorrhea region. Further exploration showed that serum albumin could increase the intracellular calcium concentration by activating the voltage-dependent calcium channel Cav2.1 in OS cells to affect the cytoskeleton, sequentially leading to dedifferentiation. Dedifferentiated OS cells with increased FAS apoptosis inhibitory molecule 2 (FAIM2) expression would gradually acquire survival ability, whereas knockdown of FAIM2 caused apoptosis in serum albumin. Moreover, FAIM2 overexpression rescued the viability of OS cells with low metastatic potential in serum albumin. In clinical specimens, OS cells showed markedly stronger positive staining of FAIM2 in the angiorrhea area. Taken together, our findings indicate that serum albumin in the angiorrhea region is a critical substance during pulmonary metastasis of OS cells. Angiorrhea is a nonnegligible prognostic element and FAIM2 might serve as a promising therapeutic target.

The clinical evolution of solitary fibrous tumor (SFT) remains unclear. Although various clinical, morphological and molecular criteria may indicate increased risk of malignancy, some SFT can still progress despite having a clearly benign appearance. Various risk stratification systems have been proposed, but unfortunately they are not sufficient to precisely determine the malignant potential. In this review, we discuss current knowledge on SFT, focusing on the following controversial issues: (i) the diverse morphologic spectrum: 'the great simulator;' (ii) malignant transformation or dedifferentiation; (iii) current risk stratification systems; and (iv) molecular factors associated with clinical evolution. The morphological spectrum of SFT and the list of differential diagnoses continue to expand. Both have increased considerably since the first descriptions of specific molecular alterations. A classification of malignant SFT should not be based on histology alone. The correlation of all pathological and molecular factors is recommended; its inclusion in risk stratification systems may help to improve diagnosis and prognosis.

Insulin plays an important role during adipogenic differentiation of animal preadipocytes and the maintenance of mature phenotypes. However, its role and mechanism in dedifferentiation of adipocyte remains unclear. This study investigated the effects of insulin on dedifferentiation of mice adipocytes, and the potential mechanisms. The preadipocytes were isolated from the subcutaneous white adipose tissue of wild type (WT), TNFα gene mutant (TNFα-/-), leptin gene spontaneous point mutant (db/db) and TNFα-/-/db/db mice and were then induced for differentiation. Interestingly, dedifferentiation of these adipocytes occurred once removing exogenous insulin from the adipogenic medium. As characteristics of dedifferentiation of the adipocytes, downregulation of adipogenic markers, upregulation of stemness markers and loss of intracellular lipids were observed from the four genotypes. Notably, dedifferentiation was occurring earlier if the insulin signal was blocked. These dedifferentiated cells regained the potentials of the stem cell-like characteristics. There is no significant difference in the characteristics of the dedifferentiation between the adipocytes. Overall, the study provided evidence that insulin plays a negative regulatory role in the dedifferentiation of adipocytes. We also confirmed that both dedifferentiation of mouse adipocytes, and effect of the insulin on this process were independent of the cell genotypes, while it is a widespread phenomenon in the adipocytes.

Liposarcomas including atypical lipomatous tumors (ALT)/well-differentiated liposarcomas (WDLPS) and dedifferentiated liposarcomas (DDLPSs) display a histomorphological spectrum with their several diagnostic mimics. Murine double minute 2(MDM2) gene amplification characterizes ALT/WDLPS and DDLPS. Presently, there is no documented study from our subcontinent on the validation of MDM2 gene testing in these tumors.Twenty-eight cases, diagnosed as ALT/WDLPS (n = 5) and DDLPSs (n = 23), along with 10 other tumors were tested for MDM2 gene amplification, using fluorescence in situ hybridization (FISH) on tissue microarrays (TMAs). Fourteen cases, diagnosed as ALT/WDLPS and DDLPS, along with 49 other tumors were tested for MDM2 immunostaining. Twenty tumors were tested for p16FISH was interpretable in 25 (89.2%) cases. Among the 20 cases diagnosed as DDLPSs, 19 displayed MDM2 gene amplification. Among the 5 cases diagnosed as ALT/WDLPS, four showed MDM2 gene amplification. Finally, 19 cases were confirmed as DDLPS and 4 as ALT/WDLPS. Furthermore, 7/19 cases confirmed as DDLPS and all 4 cases as ALT/WDLPS tested for MDM2 immunostaining, displayed its diffuse immunoexpression, while a single case of DDLPS showed its focal immunostaining. None of the 49 control cases displayed diffuse MDM2 immunoexpression. ALL 16 DDLPSs and 4 cases of ALT/WDLPS displayed p16This constitutes the first sizable study on MDM2 testing in ALT/WDLPS and DDLPS from our subcontinent using TMAs. MDM2 gene amplification testing continues as the diagnostic gold standard for ALTs/WDLPSs and DDLPSs and is useful in cases of diagnostic dilemmas. Diffuse MDM2 (IF2 clone) and p16

Background Fat transplantation is a common method employed to treat soft-tissue defects. The dedifferentiation of mature adipocytes has been well documented, but whether it occurs after fat transplantation remains unclear. Objectives The major purpose of this project was to investigate the dedifferentiation of mature adipocytes after fat transplantation. Methods Human lipoaspirate tissue was obtained from 6 female patients who underwent esthetic liposuction. Mature adipocytes were extracted and labeled with PKH26, mixed with lipoaspirate, and injected into nude mice. In addition, PKH26+ adipocytes were subjected to a ceiling culture. Grafted fat was harvested from nude mice, and stromal vascular fragment cells were isolated. The immunophenotype of PKH26+ cells was detected by flow cytometry analysis at 2 days and 1 week. The PKH26+ cells were sorted and counted at 2 and 4 weeks to verify their proliferation and multilineage differentiation abilities. Results Two days after transplantation, almost no PKH26+ cells were found in the stromal vascular fragment cells. The PKH26+ cells found 1 week after transplantation showed a positive expression of cluster of differentiation (CD) 90 (CD90) and CD105 and a negative expression of CD45. This indicates that the labeled adipocytes were dedifferentiated. Its pluripotency was further demonstrated by fluorescent cell sorting and differentiation culture in vitro. In addition, the number of live PKH26+ cells at week 4 [(6.83 ± 1.67) × 104] was similar with that at week 2 [(7.11 ± 1.82) × 104]. Conclusions Human mature adipocytes can dedifferentiate into stem cell-like cells in vivo after fat transplantation.

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.

To discuss the current understanding of “β cell identity” and factors underlying altered identity of pancreatic β cells in diabetes, especially in humans. Altered identity of β cells due to dedifferentiation and/or transdifferentiation has been proposed as a mechanism of loss of β cells in diabetes. In dedifferentiation, β cells do not undergo apoptosis; rather, they lose their identity and function. Dedifferentiation is well characterized by the decrease in expression of key β cell markers such as genes encoding major transcription factors, e.g., MafA, NeuroD1, Nkx6.1, and Foxo1, and an increase in atypical or “disallowed” genes for β cells such as lactate dehydrogenase, monocarboxylate transporter MCT1, or progenitor cell genes (Neurog3, Pax4, or Sox9). Moreover, altered identity of mature β cells in diabetes also involves transdifferentiation of β cells into other islet hormone producing cells. For example, overexpression of α cell specific transcription factor Arx or ablation of Pdx1 resulted in an increase of α cell numbers and a decrease in β cell numbers in rodents. The frequency of α-β double-positive cells was also prominent in human subjects with T2D. These altered identities of β cells likely serve as a compensatory response to enhance function/expand cell numbers and may also camouflage/protect cells from ongoing stress. However, it is equally likely that this may be a reflection of new cell formation as a frank regenerative response to ongoing tissue injury. Physiologically, all these responses are complementary. In diabetes, (1) endocrine identity recapitulates the less mature/less-differentiated fetal/neonatal cell type, possibly representing an adaptive mechanism; (2) residual β cells may be altered in their subtype proportions or other molecular features; (3) in humans, “altered identity” is a preferable term to dedifferentiation as their cellular fate (differentiated cells losing identity or progenitors becoming more differentiated) is unclear as yet.Other Information Published in: Current Diabetes Reports License: https://creativecommons.org/licenses/by/4.0See article on publisher's website: http://dx.doi.org/10.1007/s11892-019-1194-6

Background Despite the frequent use of protoplast-to-plant system in in vitro cultures of plants, the molecular mechanisms regulating the first and most limiting stages of this process, i.e., protoplast dedifferentiation and the first divisions leading to the formation of a microcallus, have not been elucidated. Results In this study, we investigated the function of miRNAs in the dedifferentiation of A. thaliana mesophyll cells in a process stimulated by the enzymatic removal of the cell wall. Leaf cells, protoplasts and CDPs (cells derived from protoplasts) cultured for 24, 72 and 120 h (first cell division). In protoplasts, a strong decrease in the amount of AGO1 in both the nucleus and the cytoplasm, as well as dicing bodies (DBs), which are considered to be sites of miRNA biogenesis, was shown. However during CDPs division, the amounts of AGO1 and DBs strongly increased. MicroRNA transcriptome studies demonstrated that lower amount of differentially expressed miRNAs are present in protoplasts than in CDPs cultured for 120 h. Then analysis of differentially expressed miRNAs, selected pri-miRNA and mRNA targets were performed. Conclusion This result indicates that miRNA function is not a major regulation of gene expression in the initial but in later steps of dedifferentiation during CDPs divisions. miRNAs participate in organogenesis, oxidative stress, nutrient deficiencies and cell cycle regulation in protoplasts and CDPs. The important role played by miRNAs in the process of dedifferentiation of mesophyll cells was confirmed by the increased mortality and reduced cell division of CDPs derived from mutants with defective miRNA biogenesis and miR319b expression.

Beim Tissue Engineering mit prim��ren humanen Chondrozyten ist die Dedifferenzierung w��hrend Zellproliferationsphase in vitro ein limitierender Prozess um eine ausreichende Zellzahl zu generieren f��r die Besiedelung eines klinisch relevanten Produkts. Obwohl Chondrozyten partiell redifferenzieren sobald sie in eine dreidimensionale (3D) Kultur ��berf��hrt werden, entspricht eine neu gebildete Knorpelmatrix nicht der Qualit��t eines Nativknorpels. Es wurde der Einfluss von Hypothermie Bedingungen auf die Dedifferenzierung von humanen Septumchondrozyten untersucht in einer Monolayer Zellkultur zur Proliferation, sowie in einer 3D Pelletkultur. Jede Zellkultur wurde w��hrend Zellproliferation entweder bei 32,2 ��C oder bei 37 ��C kultiviert. Weiter wurden Zellen, die bei diesen Temperaturen proliferiert wurden, in einer Pelletkultur bei 32,2 ��C und bei 37�� C untersucht. Hinwei��gebend auf den zellul��ren Differenzierungsgrad war die Bestimmung der mRNA knorpelspezifischer Zielgene mittels Polymerase Kettenreacktion (PCR) und hieraus gebildete Quotienten (Kollagen II/Kollagen I; Aggrecan/Versican). Ein jeweils h��herer Quotient zeigte einen h��heren Differenzierungsgrad an. Histologische und Immunhistochemische F��rbungen dienten zur Objektivierung vorhandener Proteine in Pelletkultur. In Monolayer wurde die Zellproliferation mittels 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) kolorimetrisch bestimmt. In Monolayer zeigte sich unter dem Einfluss von 32,2 ��C eine signifikant langsamere Zellproliferation verglichen zur Kultur bei 37 ��C, ein Dedifferenzierungsprozess verlief bei 32,2 ��C ebenso langsamer. In Pelletkultur konnte ein Redifferenzierungsprozess bei beiden Temperaturen nachgewiesen werden, dieser zeigte sich in den Kulturen bei 37 ��C ausgepr��gter. M��gliche Vorteile durch langsamere Dedifferenzierung in Monolayer unter 32, 2 ��C werden durch effektivere Redifferenzierungsprozesse bei 37 ��C aufgehoben. Es konnte gezeigt werden dass eine gezielte Verlangsamung der Zellproliferation und Dedifferenzierung bei 32,2 ��C in Monolayer zur Zellproliferation keinen Nachteil hat wenn diese Zellen anschlie��end in eine Pelletkultur bei 37 ��C ��berf��hrt werden.

Primary cutaneous and mucosal melanoma shows a wide histological spectrum. The correct diagnosis depends upon the demonstration of melanocytic differentiation by recognition of an associated in-situ component or immunohistochemical evidence of a melanocytic phenotype using conventional melanocytic markers, such as S-100, SOX10, Melan-A and HMB-45. Exceptionally, melanomas lose their melanocytic phenotype, at least focally, and show differentiation towards other lineages. Review of the literature shows that de- and trans-differentiation in melanoma is rare but probably under-recognised and under-reported. These often large and frequently ulcerated tumours affect adults and show a wide anatomical distribution, including mucosal sites, although there is a predilection for sun-damaged skin of the head and neck. Histologically, the tumours are biphasic and contain a pre-existing conventional melanoma. The de-differentiated component closely resembles atypical fibroxanthoma, both morphologically and immunohistochemically. Trans-differentiated melanoma may show rhabdomyosarcomatous or spindle cell carcinomatous features. Undifferentiated melanomas are similar tumours in which the conventional melanoma component is absent. Their diagnosis depends entirely upon the clinical context and identification of a classical melanoma driver gene mutation, i.e. BRAF V600E. The diagnosis of these rare and unusual tumours is challenging, and requires thorough tumour sampling and recognition of the background of a pre-existing but often focal conventional melanoma together with molecular analysis.

In the retina of zebrafish, Müller glia have the ability to reprogram into stem cells capable of regenerating all classes of retinal neurons and restoring visual function. Understanding the cellular and molecular mechanisms controlling the stem cell properties of Müller glia in zebrafish may provide cues to unlock the regenerative potential in the mammalian nervous system. Midkine is a cytokine/growth factor with multiple roles in neural development, tissue repair, and disease., In the retina of zebrafish, Müller glia have the ability to reprogram into stem cells capable of regenerating all classes of retinal neurons and restoring visual function. Understanding the cellular and molecular mechanisms controlling the stem cell properties of Müller glia in zebrafish may provide cues to unlock the regenerative potential in the mammalian nervous system. Midkine is a cytokine/growth factor with multiple roles in neural development, tissue repair, and disease. In midkine-a loss-of-function mutants of both sexes, Müller glia initiate the appropriate reprogramming response to photoreceptor death by increasing expression of stem cell-associated genes, and entering the G1 phase of the cell cycle. However, transition from G1 to S phase is blocked in the absence of Midkine-a, resulting in significantly reduced proliferation and selective failure to regenerate cone photoreceptors. Failing to progress through the cell cycle, Müller glia undergo reactive gliosis, a pathological hallmark in the injured CNS of mammals. Finally, we determined that the Midkine-a receptor, anaplastic lymphoma kinase, is upstream of the HLH regulatory protein, Id2a, and of the retinoblastoma gene, p130, which regulates progression through the cell cycle. These results demonstrate that Midkine-a functions as a core component of the mechanisms that regulate proliferation of stem cells in the injured CNS. SIGNIFICANCE STATEMENT The death of retinal neurons and photoreceptors is a leading cause of vision loss. Regenerating retinal neurons is a therapeutic goal. Zebrafish can regenerate retinal neurons from intrinsic stem cells, Müller glia, and are a powerful model to understand how stem cells might be used therapeutically. Midkine-a, an injury-induced growth factor/cytokine that is expressed by Müller glia following neuronal death, is required for Müller glia to progress through the cell cycle. The absence of Midkine-a suspends proliferation and neuronal regeneration. With cell cycle progression stalled, Müller glia undergo reactive gliosis, a pathological hallmark of the mammalian retina. This work provides a unique insight into mechanisms that control the cell cycle during neuronal regeneration.

BACKGROUND Angiopoietin-like proteins (ANGPTL) are a family of secretory glycoproteins that are involved in many pathophysiological processes. ANGPTL7 is a newly-discovered member of the ANGPTL family and plays a role in corneal morphogenesis, angiogenesis, glaucoma, and cancer. To date, whether ANGPTL7 is involved in osteoporosis is unknown. Therefore, to discover the effects of ANGPTL7 on osteoporosis, we explored the expression of ANGPTL7 in preosteoblasts and assessed the mechanism underlying its effects on proliferation and differentiation abilities of preosteoblasts. MATERIAL AND METHODS Mouse MC3T3-E1 cells were cultured in osteogenic medium for osteogenic differentiation. The expression levels of ANGPTL7 were detected by RT-qPCR and Western blot assays. Moreover, the overexpressed plasmid of ANGPTL7 pMSCV-ANGPTL7 was transfected into MC3T3-E1 cells. CCK-8 was used to evaluate cell proliferation. ALP activity detection and alizarin red staining were performed to measure the effect of ANGPTL7 on osteogenic differentiation. The expression levels bone morphogenetic proteins (BMPs) and osteogenic markers ALP, runt-related transcription factor 2 (Runx2), osteocalcin (OCN), and collagen I (Col I) were analyzed by Western blot. RESULTS When MC3T3-E1 cells were exposed to osteogenic medium, there was a significant increase in ANGPTL7, and overexpression of ANGPTL7 markedly promoted cell proliferation, ALP activity, and mineralization. Moreover, ANGPTL7 upregulated the levels of BMPs, especially BMP2/7, and the osteogenic markers ALP, Runx2, OCN, and Col I. CONCLUSIONS The results suggest that by regulating the expression of BMPs, ANGPTL7 directly promotes proliferation, differentiation, and mineralization of osteoblasts.

Summary: Hepatocytes are the primary functional cells of the liver, performing its metabolic, detoxification, and endocrine functions. Functional hepatocytes are extremely valuable in drug discovery and evaluation, as well as in cell therapy for liver diseases. However, it has been a long-standing challenge to maintain the functions of hepatocytes in vitro. Even freshly isolated hepatocytes lose essential functions after short-term culture for reasons that are still not well understood. In the present study, we find that mechanical tension-induced yes-associated protein activation triggers hepatocyte dedifferentiation. Alleviation of mechanical tension by confining cell spreading is sufficient to inhibit hepatocyte dedifferentiation. Based on this finding, we identify a small molecular cocktail through reiterative chemical screening that can maintain hepatocyte functions over the long term and in vivo repopulation capacity by targeting actin polymerization and actomyosin contraction. Our work reveals the mechanisms underlying hepatocyte dedifferentiation and establishes feasible approaches to maintain hepatocyte functions. : It has been a long-standing challenge to maintain the functions of hepatocytes in vitro. Sun et al. find that mechanical tension-induced Yap activation triggers hepatocyte dedifferentiation. Alleviation of mechanical tension by confining cell spreading or treatment with a small molecule cocktail targeting actin/actomyosin dynamics could maintain hepatocyte functions. Keywords: hepatocytes, dedifferentiation, small molecules, mechanical tension

Background Lysophosphatidic acid (LPA) is a pleiotropic lipid messenger that addresses at least six specific G-protein coupled receptors. Accumulating evidence indicates a significant involvement of LPA in immune cell regulation as well as Schwann cell physiology, with potential relevance for the pathophysiology of peripheral neuroinflammation. However, the role of LPA signaling in inflammatory neuropathies has remained completely undefined. Given the broad expression of LPA receptors on both Schwann cells and cells of the innate and adaptive immune system, we hypothesized that inhibition of LPA signaling may ameliorate the course of disease in experimental autoimmune neuritis (EAN). Methods We induced active EAN by inoculation of myelin protein 2 peptide (P255–78) in female Lewis rats. Animals received the orally available LPA receptor antagonist AM095, specifically targeting the LPA1 receptor subtype. AM095 was administered daily via oral gavage in a therapeutic regimen from 10 until 28 days post-immunization (dpi). Analyses were based on clinical testing, hemogram profiles, immunohistochemistry and morphometric assessment of myelination. Results Lewis rats treated with AM095 displayed a significant improvement in clinical scores, most notably during the remission phase. Cellular infiltration of sciatic nerve was only discretely affected by AM095. Hemogram profiles indicated no impact on circulating leukocytes. However, sciatic nerve immunohistochemistry revealed a reduction in the number of Schwann cells expressing the dedifferentiation marker Sox2 paralleled by a corresponding increase in differentiating Sox10-positive Schwann cells. In line with this, morphometric analysis of sciatic nerve semi-thin sections identified a significant increase in large-caliber myelinated axons at 28 dpi. Myelin thickness was unaffected by AM095. Conclusion Thus, LPA1 signaling may present a novel therapeutic target for the treatment of inflammatory neuropathies, potentially affecting regenerative responses in the peripheral nerve by modulating Schwann cell differentiation.