Search Results (77)

Morus alba L. (Moraceae), white mulberry, is an ancient, well-known source of several compounds with potent biological activities and beneficial effects on human health. In this study, the juices of three stabilised undifferentiated cell lines, calli maintained in light and dark conditions, and suspensions maintained in dark condition of M. alba were investigated for their phytochemical content and biological activity. The results highlighted the main presence of oxyresveratrol and resveratrol-backbone glucosides, together with benzofuran derivatives. Oxyresveratrol triglucoside was found for the first time in M. alba in vitro cultures, where it represents the main compound, accounting for almost 90 µg/mL in all the juices. The total stilbenoid content resulted significantly higher in calli juices during the logarithmic phase of the growth cycle, and cell suspension juice exhibited the statistically highest total content (313.21 µg/mL of juice). Only cell suspension juice showed ROS reduction in Caco-2 cells, whereas all the juices reduced IL-1β and TNF-α levels in Caco-2 cells stimulated with LPS. These results lay the groundwork for the future exploitation of M. alba dedifferentiated cultures as sustainable resources of stilbenoid compounds to be used in the nutraceutical, cosmetic, and pharmaceutical industries. Full article

Stem cells, both normal somatic (SSC) and cancer stem cells (CSC) exist in minimally two states, i.e., quiescent and activated. Regulation of these two states, including their reliance on different metabolic processes, i.e., FAO and glycolysis in quiescent versus activated stem cells respectively, involves the analysis of a complex array of factors (nutrient and oxygen levels, adhesion molecules, cytokines, etc.) to initiate the epigenetic changes to either depart or enter quiescence. Quiescence is a critical feature of SSC that is required to maintain the genomic integrity of the stem cell pool, particularly in long lived complex organisms. Quiescence in CSC, whether they are derived from mutations arising in SSC, aberrant microenvironmental regulation, or via dedifferentiation of more committed progenitors, is a critical component of therapy resistance and disease latency and relapse. At the beginning of vertebrate evolution, approximately 450 million years ago, a gene duplication generated the two members of the Kat3 family, CREBBP (CBP) and EP300 (p300). Despite their very high degree of homology, these two Kat3 coactivators play critical and non-redundant roles at enhancers and super-enhancers via acetylation of H3K27, thereby controlling stem cell quiescence versus activation and the cells metabolic requirements. In this review/perspective, we discuss the unique regulatory roles of CBP and p300 and how specifically targeting the CBP/β-catenin interaction utilizing small molecule antagonists, can correct lineage infidelity and safely eliminate quiescent CSC. Full article

Dedifferentiated fat (DFAT) cells are adipocyte-derived cells that are able to differentiate into multiple cell lineages such as adipocytes, osteoblasts and chondrocytes, similar to mesenchymal stem cells (MSCs). Despite their great potential for developing novel clinical interventions by using their multipotency, the detailed mechanisms of how adipocytes undergo dedifferentiation into DFAT cells are not completely understood, because useful in vitro tools for studying adipocyte dedifferentiation are missing. In this study, we show that mature adipocytes derived from the MSC cell line C3H10T1/2 underwent dedifferentiation into cells with DFAT cell-like characteristics, when they were cultured in an inverted flask. During the dedifferentiation, expression levels of genes and protein specific to adipocytes were continuously decreased, whereas those for MSC, proliferation and WNT/β-catenin signaling were gradually increased. These DFAT-like cells also underwent differentiation into adipocytes, osteoblasts and chondrocytes with their specific cell morphology and gene expression. We also observed that an individually cultured single adipocyte also underwent dedifferentiation into DFAT-like cells that were able to differentiate into the multiple cell lineages. Our results indicate that C3H10T1/2 cells could be a great tool for determining molecular biological and biochemical mechanisms underlying adipocyte dedifferentiation. Full article

Arteriovenous fistulae (AVF) are the preferred mode of dialysis vascular access but have a maturation failure rate of over 50%. Venous segment stenosis is one of the main reasons. However, the reasons for the prevalence of stenosis in the venous segment, as opposed to the arterial segment, remain unknown. We hypothesize that differences in the protein expression profiles and in the activation of key signaling pathways of the vascular smooth muscle cells (VSMCs) in these two segments may contribute to this difference. In this study, arterial porcine VSMCs (ApSMCs) and venous porcine VSMCs (VpSMCs) were isolated to examine relevant protein expression and cell signaling pathway activation via Western blots and immunofluorescent staining. Our results revealed that growth-medium-stimulated cell proliferation was significantly higher in VpSMCs compared to ApSMCs, but no difference was detected in PDGF-BB-stimulated proliferation. VpSMCs migration was significantly greater than ApSMCs under both the serum-free condition and in the growth medium condition. In addition, VpSMCs appeared more susceptible to PDGF-BB-stimulated cell dedifferentiation. Furthermore, P53, vitronectin, collagen 1A1 and integrin β3 had lower expression in VpSMCs, whereas fibronectin, FAK, VCAM-1, MMP-9, TIMP-2 and TIMP-3 had higher expression as compared to ApSMCs. Meanwhile, PDGF-BB stimulated P38, JNKs and AKT activation, which were more potent in VpSMCs. Our results identify significant differences between VpSMCs and ApSMCs in the expression of functional proteins and the activation of signaling pathways, which may be responsible for more aggressive venous—as compared to arterial—stenosis in the clinical setting of arteriovenous access or other similar procedures. Full article

Diffuse intimal thickening (DIT) is a pre-clinical stage of atherosclerosis characterized by thickened intima. The molecular basis of its susceptibility to atherogenesis is unknown, and mechanistic investigations cannot be performed in commonly used mouse models, in which DIT does not exist. Vascular smooth muscle cells (SMCs) are the predominant cell type that occupies the intima and media of DIT. The molecular differences between these two layers may reveal the earliest phenotypic changes in SMCs to promote atherosclerosis. We benchmarked the RNA quality of human coronary arteries from autopsies (n = 7) and explanted hearts (n = 7) and performed Visium spatial gene expression on tissue sections with DIT. Although autopsy samples met the RNA quality standard for Visium (DV200 ≥ 30%), only arteries from explanted hearts exhibited reliable sequencing performance. Genes enriched in TGF-β-mediated remodeling of the extracellular matrix were overrepresented in the intima. SMCs enriched in the intima are dedifferentiated, but unlike those in the atherosclerotic lesions, they are not pro-inflammatory. Our findings indicate that autopsy samples are not ideal to distinguish subtle differences among cell phenotypes. SMCs in thickened intima may lead to lipid retention but not necessarily the onset of atherosclerosis. Full article

Type 2 diabetes mellitus (T2DM), a prevalent chronic disease affecting over 400 million people globally, is driven by genetic and environmental factors. The pathogenesis involves insulin resistance and β-cell dysfunction, mediated by mechanisms such as the dedifferentiation of β-cells, mitochondrial dysfunction, and oxidative stress. Treatment should be based on non-pharmacological therapy. Strategies such as increased physical activity, dietary modifications, cognitive-behavioral therapy are important in maintaining normal glycemia. Advanced therapies, including SGLT2 inhibitors and GLP-1 receptor agonists, complement these treatments and offer solid glycemic control, weight control, and reduced cardiovascular risk. Complications of T2DM, such as diabetic kidney disease, retinopathy, and neuropathy, underscore the need for early diagnosis and comprehensive management to improve patient outcomes and quality of life. Full article

Inflammation models with the proinflammatory cytokine interleukin-1β (IL-1β) are widely used in the in vitro investigation of new therapeutic approaches for osteoarthritis (OA). The aim of this study was to systematically analyze the influence of IL-1β in a 3D chondral pellet culture model. Bovine articular chondrocytes were cultured to passage 3 and then placed in pellet culture. Titration of IL-1β (100–0.1 ng/mL) was performed with both human and bovine recombinant protein in chondrocyte culture for 2 weeks. Gene expression of anabolic (collagen 2, aggrecan, cartilage oligomeric protein (COMP), proteoglycan-4 (PRG-4)), catabolic matrix metallo proteinases (MMP-3, MMP-13), dedifferentiation (collagen 1) markers and inflammatory cytokines IL-6 and IL-8 was determined. Analysis of the cell culture medium was performed for the inflammatory markers IL-6 and nitric oxide (NO). In general, the influence of IL-1β was shown by a decrease in the expression of anabolic markers (collagen 2, aggrecan, PRG-4), whereas the catabolic markers MMP-3 and MMP-13 as well as the inflammatory markers IL-6 and IL-8 were significantly increased. This was observed both at the early time point (day 4) and at the late time point (day 14). The described inflammatory effects were confirmed by increased concentration-dependent release of NO and IL-6. The threshold concentration for a detectable effect compared to control differed between groups, but was reached earlier by homologous application of IL-1β. This study provides a systematic evaluation of IL-1β-specific effects on chondrocytes in a 3D pellet culture model, which is highly relevant for comparisons of studies in OA-specific drug development. Full article

The limited self-repair capacity of cartilage due to its avascular and aneural nature leads to minimal regenerative ability. Autologous chondrocyte transplantation (ACT) is a popular treatment for cartilage defects but faces challenges due to chondrocyte dedifferentiation in later passages, which results in undesirable fibroblastic phenotypes. A promising treatment for cartilage injuries and diseases involves tissue engineering using cells (e.g., chondrocytes), scaffolds (e.g., Alginate Sulfate (AlgSulf)), and biochemical signals (e.g., Salidroside and TGF-β). This study focuses on investigating the effects of AlgSulf scaffolds with varying degrees of sulfation, Salidroside, and TGF-β on the proliferation, viability, and phenotype maintenance of chondrocytes. The findings demonstrate that AlgSulf films with a degree of sulfation (DS) = 2, treated with a combination of Salidroside and TGF-β, significantly enhanced chondrocyte proliferation (p < 0.001 and p < 0.0001 in P2 and P4, respectively), preserved round cell morphology, and maintained cartilage-specific gene expression (Col2, Aggrecans, and SOX9) while downregulating fibroblastic markers (Col1, MMP13, IL-1β, and IL-6). Our findings suggest the potential of this combination for enhancing cartilage regeneration in tissue engineering applications. Full article

Background/Objectives: This study aims to investigate the effects of 4-methylumbelliferone (4-MU) on islet morphology, cell phenotype and function, and to explore possible mechanisms of β cell regeneration. Methods: The Type 1 diabetes (T1D) model was induced by continuous dose injection of streptozotocin (STZ), and mice were treated with 4-MU for 3 weeks. Plasma insulin level, islet cell phenotype and immune infiltration were determined by IPGTT, ELISA, HE and immunofluorescence. The Ins2^Cre/+/Rosa26-eGFP transgenic mice model was used to detect β identity change. Primary rodent islets were incubated with 4-MU or vehicle in the presence or absence of STZ, AO/PI staining, and a scanning electron microscope (SEM), PCR and ELISA were used to evaluated islet viability, islet morphology, the specific markers of islet β cells and insulin secretion. Results: Treatment with 4-MU significantly decreased blood glucose and increased plasma insulin levels in STZ-induced diabetes. The plasma insulin level in the STZ group was 7.211 ± 2.602 ng/mL, which was significantly lower than the control group level (26.94 ± 4.300 ng/mL, p < 0.001). In contrast, the plasma insulin level in the STZ + 4-MU group was 22.29 ± 7.791 ng/mL, which was significantly higher than the STZ group (p < 0.05). The 4-MU treatment increased islet and β cells numbers and decreased α cell numbers in STZ-induced diabetes. Conclusions: Islet inflammation as indicated by insulin and CD3 was caused by infiltrates, and the β cell proliferation as indicated by insulin and Ki67 was boosted by 4-MU. β cell dedifferentiation was inhibited by 4-MU as assessed by insulin and glucagon double-positive cells and confirmed by Ins2^Cre/+/Rosa26-eGFP mice. In cultured primary rodent islets, 4-MU restored islet viability, protected islet morphology, inhibited β-cell dedifferentiation, and promoted insulin secretion. The benefits of 4-MU in T1D have been proved to be associated with β cells self-replication, dedifferentiation inhibition and immune progression suppression, which help to maintain β cell mass. Full article

Obesity is concurrent with immunological dysregulation, resulting in chronic low-grade inflammation and cellular dysfunction. In pancreatic islets, this loss of function has been correlated with mature β-cells dedifferentiating into a precursor-like state through constant exposure to inflammatory stressors. As mature adipocytes likewise have the capability to dedifferentiate in vitro and in vivo, we wanted to analyze this cellular change in relation to adipose tissue (AT) inflammation and adipose tissue macrophage (ATM) activity. Using our organotypic AT explant culture method combined with a double-reporter mouse model for labeling ATMs and mature adipocytes, we were able to visualize and quantify dedifferentiated fat (DFAT) cells in AT explants. Preliminary testing showed increased dedifferentiation after tamoxifen (TAM) stimulation, making TAM-dependent lineage-tracing models unsuitable for quantification of naturally occurring DFAT cells. The regulatory role of ATMs in adipocyte dedifferentiation was shown through macrophage depletion using Plexxicon 5622 or clodronate liposomes, which significantly increased DFAT cell levels. Subsequent bulk RNA sequencing of macrophage-depleted explants revealed enrichment of the tumor necrosis factor α (TNFα) signaling pathway as well as downregulation of associated genes. Direct stimulation with TNFα decreased adipocyte dedifferentiation, while application of a TNFα-neutralizing antibody did not significantly alter DFAT cell levels. Our findings suggest a regulatory role of resident ATMs in maintaining the mature adipocyte phenotype and preventing excessive adipocyte dedifferentiation. The specific regulatory pathways as well as the impact that DFAT cells might have on ATMs, and vice versa, are subject to further investigation. Full article

Osteoarthritis (OA) is a degenerative joint disease characterized by cartilage breakdown and chronic inflammation in joints. As the most prevalent form of arthritis, OA affects around 600 million people globally. Despite the increasing number of individuals with OA risk factors, such as aging and obesity, there is currently no effective cure for the disease. In this context, this study investigated the therapeutic effects of tamarixetin, a flavonoid with antioxidative and anti-inflammatory properties, against OA pathology and elucidated the underlying molecular mechanism. In interleukin-1β (IL-1β)-treated chondrocytes, tamarixetin inhibited the OA phenotypes, restoring cell viability and chondrogenic properties while reducing hypertrophic differentiation and dedifferentiation. Tamarixetin alleviated oxidative stress via the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway activation and inhibited mitogen-activated protein kinase and nuclear factor-κB (NF-κB). Furthermore, tamarixetin attenuated pyroptosis, a programmed cell death caused by excessive inflammation, by suppressing inflammasome activation. We confirmed that the chondroprotective effects of tamarixetin are mediated by the concurrent upregulation of Nrf2 signaling and downregulation of NF-κB signaling, which are key players in balancing antioxidative and inflammatory responses. Overall, our study demonstrated that tamarixetin possesses chondroprotective properties by alleviating IL-1β-induced cellular stress in chondrocytes, suggesting its therapeutic potential to relieve OA phenotype. Full article

Background: Obesity poses a significant global health challenge, given its association with the excessive accumulation of adipose tissue (AT) and various systemic disruptions. Within the adipose microenvironment, expansion and enrichment with immune cells trigger the release of inflammatory mediators and growth factors, which can disrupt tissues, including bones. While obesity’s contribution to bone loss is well established, the direct impact of obese AT on osteoblast maturation remains uncertain. This study aimed to explore the influence of the secretomes from obese and lean AT on osteoblast differentiation and activity. Methods: SAOS-2 cells were exposed to the secretomes obtained by culturing human subcutaneous AT from individuals with obesity (OATS) or lean patients, and their effects on osteoblasts were evaluated. Results: In the presence of the OATS, mature osteoblasts underwent dedifferentiation, showing an increased proliferation accompanied by a morphological shift towards a mesenchymal phenotype, with detrimental effects on osteogenic markers and the calcification capacity. Concurrently, the OATS promoted the expression of mesenchymal and adipogenic markers, inducing the formation of cytoplasmic lipid droplets in SAOS-2 cells exposed to an adipogenic differentiation medium. Additionally, TGF-β1 emerged as a key mediator of these effects, as the OATS was enriched with this growth factor. Conclusions: Our findings demonstrate that obese subcutaneous AT promotes the dedifferentiation of osteoblasts and increases the adipogenic profile in these cells. Full article

The aim of this study was to investigate the effects of hypoxia-induced phenotype, glucose metabolism, ROS levels, and the PDK1-mediated regulation of TGF-β/Smad signaling in yellow cattles, yaks, and those overexpressing PDK1 PASMCs using growth curves, flow cytometry, scratch experiments, glucose and lactic acid assays, RT-qPCR, and Western blotting. The results showed that hypoxia significantly promoted proliferation, migration, antiapoptosis, ROS levels, glucose consumption, and lactate production in yellow cattle PASMCs (p < 0.05), and the cells were dedifferentiated from the contractile phenotype; conversely, hypoxia had no significant effect on yak PASMCs (p > 0.05). PDK1 overexpression significantly promoted proliferation, antiapoptosis, glucose consumption, and lactate production in yak PASMCs under normoxia and hypoxia (p < 0.05), decreased their migration levels under hypoxia (p < 0.05), and dedifferentiated the contractile phenotype of the cells. Overexpression of PDK1 in yak PASMCs is detrimental to their adaptation to hypoxic environments. Yak PASMCs adapted to the effects of hypoxia on lung tissue by downregulating the expression of genes related to the PDK1 and TGF-β/Smad signaling pathways. Taken together, the regulation of PDK1-mediated TGF-β/Smad signaling may be involved in the process of yaks’ adaptation to the hypoxic environment of the plateau, reflecting the good adaptive ability of yaks. The present study provides basic information to further elucidate the mechanism of PDK1-mediated TGF-β/Smad signaling induced by hypoxia in the lungs of yaks, as well as target genes for the treatment of plateau diseases in humans and animals. Full article

The β-cells within the pancreas play a pivotal role in insulin production and secretion, responding to fluctuations in blood glucose levels. However, factors like obesity, dietary habits, and prolonged insulin resistance can compromise β-cell function, contributing to the development of Type 2 Diabetes (T2D). A critical aspect of this dysfunction involves β-cell dedifferentiation and transdifferentiation, wherein these cells lose their specialized characteristics and adopt different identities, notably transitioning towards progenitor or other pancreatic cell types like α-cells. This process significantly contributes to β-cell malfunction and the progression of T2D, often surpassing the impact of outright β-cell loss. Alterations in the expressions of specific genes and transcription factors unique to β-cells, along with epigenetic modifications and environmental factors such as inflammation, oxidative stress, and mitochondrial dysfunction, underpin the occurrence of β-cell dedifferentiation and the onset of T2D. Recent research underscores the potential therapeutic value for targeting β-cell dedifferentiation to manage T2D effectively. In this review, we aim to dissect the intricate mechanisms governing β-cell dedifferentiation and explore the therapeutic avenues stemming from these insights. Full article

The cancer stem cell (SC) theory proposes that a population of SCs serves as the driving force behind fundamental tumor processes, including metastasis, recurrence, and resistance to therapy. The standard of care for patients with stage III and high-risk stage II colorectal cancer (CRC) includes surgery and adjuvant chemotherapy. Fluoropyrimidines and their combination with oxaliplatin increased the cure rates, being able to eradicate the occult metastatic SC in a fraction of patients. The treatment for unresectable metastatic CRC is based on chemotherapy, antibodies to VEGF and EGFR, and tyrosine-kinase inhibitors. Immunotherapy is used in MSI-H tumors. Currently used drugs target dividing cells and, while often effective at debulking tumor mass, these agents have largely failed to cure metastatic disease. SCs are generated either due to genetic and epigenetic alterations in stem/progenitor cells or to the dedifferentiation of somatic cells where diverse signaling pathways such as Wnt/β-catenin, Hedgehog, Notch, TGF-β/SMAD, PI3K/Akt/mTOR, NF-κB, JAK/STAT, DNA damage response, and Hippo-YAP play a key role. Anti-neoplastic treatments could be improved by elimination of SCs, becoming an attractive target for the design of novel agents. Here, we present a review of clinical trials assessing the efficacy of targeted treatment focusing on these pathways in CRC. Full article