Cell Renewal Research Articles

Vismodegib, a sonic hedgehog signalling blockade, ameliorates ovalbumin and ovalbumin/lipopolysaccharide-induced airway inflammation and asthma phenotypical models

AimsAsthmatics exhibit clinical fluctuations between manageable and treatment-resistant phenotypes as a worldwide socioeconomic health burden. Sonic Hedgehog (Shh) genes mediate regulatory pulmonary cell renewal in adults and contribute to the pathogenesis of high phenotypic asthma which depends mainly on T helper-2 (Th-2) cells and related cytokines. However, the exact pathophysiological roles of Shh molecular signalling in the Th-17-dependent low phenotypic allergic airway inflammation and asthma are not evidenced previously. Main methodsOvalbumin (OVA) and OVA/lipopolysaccharide (LPS)-sensitized and challenged BALB/c mice were enrolled currently to assess the Shh signalling proteins. Furthermore, the effects of vismodegib, a Smo inhibitor, on the modulation of Shh signalling were compared to dexamethasone. The asthma phenotypes were confirmed by serum total immunoglobulin-E (IgE), bronchoalveolar lavage (BAL) fluid white blood cell counts, lung interleukins, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β1, and histopathological changes, and scoring. Key findingsMice challenged with OVA or OVA/LPS showed upregulated lung Shh, patched (Ptch1), smoothened (Smo), and Gli1 proteins. Vismodegib in the two experimental phenotypes of asthma showed reduced airway inflammation and remodelling. Additionally, vismodegib reduced the eosinophilia and neutrophilia reported in high and low asthma types, respectively. Moreover, vismodegib and dexamethasone exhibited negative feedback control throughout the enhanced Shh signalling cascades, including Shh, Ptch1, and Gli1 in several asthma models. SignificanceIn conclusion, Shh signalling partially elucidates the OVA/LPS-challenged mice with severe asthma, which proposes a new promising molecular therapeutic target. Furthermore, Smo inhibition by vismodegib has therapeutic potential in both experimental eosinophilic and neutrophilic allergic airway diseases.

Localization and characterization of SSCs from pre-pubertal bovine testes

Self renewal and proliferation ability of spermatogonial stem cells (SSCs) support spermatogenesis during adult life. Theoretically, these stem cells can be utilized for transmission of genetic information to descendants via testicular transplantation. However, lack of knowledge in methodologies for identification of SSCs limits the application of SSCs transplantation in domestic animals. Accumulated studies have shown that SSCs specific markers (DBA, UCHL1) and stem cell marker (Sox2, Oct4) are useful to screen SSCs that able to be used for transplantation. However, in cattle, less information is available on the expression status of these markers till date. Therefore, a study was carried out in 2019 at Tsukuba University, Japan where testes from 3, 5 and 7 months old calves were utilized to examine testicular localization and in vitro propogation of stem cell markers. SSCs were isolated by enzymatic digestion combined with centrifugal separation on discontinuous Percoll density gradient. Cell propagation and SSCs marker expression were determined at 5, 10 and 15 days post-culture. Immunostaining in conjunction with Western Blot analysis of cultured cells showed that stem cell markers (UCHL1, Oct4 and Sox2) were expressed in SSCs suggesting that differentiation of gonocyte started by 3 months and SSCs differentiation begins after 5 months of age. Taken together, these results demonstrated marker expression and localization of bull SSCs and showed that in vitro culturing of bull SSCs is implementable.

LYL1 facilitates AETFC assembly and gene activation by recruiting CARM1 in t(8;21) AML

Transcription factors (TFs) play critical roles in hematopoiesis, and their aberrant expression can lead to various types of leukemia. The t(8;21) leukemogenic fusion protein AML1-ETO (AE) is the most common fusion protein in acute myeloid leukemia and can enhance hematopoietic stem cell renewal while blocking differentiation. A key question in understanding AE-mediated leukemia is what determines the choice of AE to activate self-renewal genes or repress differentiation genes. Toward the resolution of this problem, we earlier showed that AE resides in the stable AETFC complex and that its components colocalize on up- or down-regulated target genes and are essential for leukemogenesis. In the current study, using biochemical and genomic approaches, we show that AE-containing complexes are heterogeneous, and that assembly of the larger AETFC (containing AE, CBFβ, HEB, E2A, LYL1, LMO2, and LDB1) requires LYL1. Furthermore, we provide strong evidence that the LYL1-containing AETFC preferentially binds to active enhancers and promotes AE-dependent gene activation. Moreover, we show that coactivator CARM1 interacts with AETFC and facilitates gene activation by AETFC. Collectively, this study describes a role of oncoprotein LYL1 in AETFC assembly and gene activation by recruiting CARM1 to chromatin for AML cell survival.

The Role of MARCKS in Metastasis and Treatment Resistance of Solid Tumors.

Simple SummaryCancer metastasis is a critical event in the progression of solid tumors and is invariably associated with adverse outcomes and mortality. Understanding novel mechanisms or molecules that promote cancer metastasis will facilitate the development of new strategies for cancer treatment. Recently, MARCKS has been studied extensively in several cancers and has been implicated in tumor progression and metastasis. This review summarizes recent advances in the understanding of MARCKS on cancer metastasis, stemness, and therapeutic resistance and provides prospects on targeting MARCKS therapeutically. Specifically, we review the molecular mechanisms and multiple signaling pathways by which MARCKS contributes to the progression and metastasis in solid tumors.The myristoylated alanine-rich C-kinase substrate (MARCKS) is a membrane-associated protein kinase C (PKC) substrate ubiquitously expressed in eukaryotic cells. MARCKS plays important roles in multiple cellular processes, including cell adhesion and motility, mucin secretion, exocytosis, and inflammatory response. Aberrant MARCKS signaling has been observed in the development and progression of multiple cancer types. In addition, MARCKS facilitates cancer metastasis through modulating cancer cell migration and invasion. Moreover, MARCKS contributes to treatment resistance, likely by promoting cancer stem cell renewal as well as immunosuppression. In this review, we describe MARCKS protein structure, cellular localization, and biological functions. We then discuss the role of MARCKS in cancer metastasis as well as its mechanisms of action in solid tumors. Finally, we review recent advances in targeting MARCKS as a new therapeutic strategy in cancer management.

Biomarker profiles in heart failure with preserved vs. reduced ejection fraction: results from the DIAST-CHF study.

Chronic heart failure (HF) is a common disease and one of the leading causes of death worldwide. Heart failure with preserved ejection fraction (HFpEF) and with reduced ejection fraction (HFrEF) are different diseases with distinct as well as comparable pathophysiologies and diverse responses to therapeutic agents. We aimed to identify possible pathobiochemical signalling pathways and biomarkers in HFpEF and HFrEF by using a broad proteomic approach. A total of 180 biomarkers in the plasma of a representative subgroup (71years old) of HFpEF (70% female) with a left ventricular ejection fraction (LVEF)≥50% and HFrEF (18% female) with an LVEF≤40% patients (n=127) from the Prevalence and Clinical Course of Diastolic Dysfunction and Diastolic Heart Failure (DIAST-CHF) trial were examined and compared with a healthy control group (n=40; 48% female). We were able to identify 35 proteins that were expressed significantly different in both HF groups compared with the control group. We determine 29 unique proteins expressed in HFpEF and 33 unique proteins in HFrEF. Significantly up-regulated trefoil factor 3 (TFF3) and down-regulated contactin-1 could be identified as previously unknown biomarkers for HF. However, TFF3 is also a predictive factor for the occurrence of a cardiovascular event in HFpEF patients. In HFpEF, serine protease 27 was found at reduced levels for the first time, which could offer a new therapeutic target. Additionally, network analyses showed a special role of platelet-derived growth factor subunit A, Dickkopf-related protein 1, and tumour necrosis factor receptor superfamily member 6 in HFpEF patients, whereas perlecan and junctional adhesion molecule A stood out in the HFrEF group. Overall, signalling pathways of metabolic processes, cellular stress, and iron metabolism seemed to be important for HFrEF, whereas for HFpEF, oxygen stress, haemostasis, cell renewal, cell migration, and cell proliferation are in the foreground. The identified proteins and signalling pathways offer new therapeutic and diagnostic approaches for patients with chronic HF.

Biological Effects of Yeast-fermented Plant Root Extract Mixture on Skin Cells

Purpose: To verify the biological effects including anti-aging and anti-inflammatory effects, of fermented plant root extract mixtures that were measured <i>in vitro</i>.Method: To select the most effective plant root extract, the effects of DPPH free radical scavenging were measured. The mixture (MC) of selected plant roots was fermented with <i>Saccharomyces cerevisiae</i> (<i>S. cerevisiae</i>) (MF). Reverse transcription (RT)-PCR was performed on human dermal fibroblast to measure the effects of MC and MF on the mRNA expression of <i>COL1A1</i> and hyaluronic acid synthase 2 (<i>HAS2</i>) and on the anti-inflammatory-related gene expressions such as cyclooxygenase-2 (<i>COX-2</i>), interleukin-6 (<i>IL-6</i>), and tumor-necrosis factor (<i>TNF</i>)-α. Additionally, we also tested the cell renewal and proliferation for skin cell migration. Student's t-test was performed for statistical analysis, and results were expressed as mean ± standard deviation.Results: Among 15 root extracts, those of <i>Taraxacum officinale</i> (<i>T. officinale</i>) rhizome, <i>Arctium lappa</i> (<i>A. lappa</i>), <i>Anemarrhena asphodeloides</i> (<i>A. asphodeloides</i>), <i>Pueraria lobata</i> (<i>P. lobata</i>), and <i>Nelumbo nucifera</i> (<i>N. nucifera</i>) were selected based on the effect of DPPH free radical scavenging. The mRNA expression levels of <i>COL1A1</i> and <i>HAS2</i> were increased by MC and MF in a concentration-dependent manner. MC and MF showed cell renewal and proliferation. Inflammation-related genes were inhibited by MC or MF in a concentration-dependent manner (<i>TNF-α</i>, <i>IL-6</i>, and <i>COX-2</i>). MF showed significantly better efficacy than MC.Conclusion: Mixed root extracts of <i>T. officinale rhizome/root</i>, <i>A. lappa</i>, <i>A. asphodeloides</i>, <i>P. lobata</i>, and <i>N. nucifera</i> fermented with <i>S. cerevisiae</i> enable cell renewal, have anti-aging and anti-inflammatory effects, and can be used as an active cosmetic raw materials.

The Musashi proteins direct post-transcriptional control of protein expression and alternate exon splicing in vertebrate photoreceptors

The Musashi proteins, MSI1 and MSI2, are conserved RNA binding proteins with a role in the maintenance and renewal of stem cells. Contrasting with this role, terminally differentiated photoreceptor cells express high levels of MSI1 and MSI2, pointing to a role for the two proteins in vision. Combined knockout of Msi1 and Msi2 in mature photoreceptor cells abrogated the retinal response to light and caused photoreceptor cell death. In photoreceptor cells the Musashi proteins perform distinct nuclear and cytoplasmic functions. In the nucleus, the Musashi proteins promote splicing of photoreceptor-specific alternative exons. Surprisingly, conserved photoreceptor-specific alternative exons in genes critical for vision proved to be dispensable, raising questions about the selective pressures that lead to their conservation. In the cytoplasm MSI1 and MSI2 activate protein expression. Loss of Msi1 and Msi2 lead to reduction in the levels of multiple proteins including proteins required for vision and photoreceptor survival. The requirement for MSI1 and MSI2 in terminally differentiated photoreceptors alongside their role in stem cells shows that, depending on cellular context, these two proteins can control processes ranging from cell proliferation to sensory perception.

Inhibition of GABAAR or Application of Lactobacillus casei Zhang Alleviates Ulcerative Colitis in Mice: GABAAR as a Potential Target for Intestinal Epithelial Renewal and Repair.

Emerging evidence indicates that the gamma−aminobutyric acid type A receptor (GABAAR) and Lactobacillus casei Zhang regulate colitis in a variety of ways, such as by participating in host immune and inflammatory responses, altering the gut microbiota, and influencing intestinal barrier function. However, not much is known about the mechanisms by which GABAAR and L. casei affect colon epithelial cell renewal and the interaction between GABAAR and L. casei during this process. To elucidate this, we established a dextran sulfate sodium (DSS)−induced model and measured the mouse body weights, colon length, the disease activity index (DAI), and histological scores. Our results indicated that inhibition of GABAAR alleviated the DSS−induced colitis symptoms, resulting in less weight loss and more intact colon tissue. Moreover, treatment with bicuculline (Bic, a GABAAR inhibitor) increased the levels of PCNA, β−catenin, and TCF4 in mice with colitis. Interestingly, open field test performances showed that inhibition of GABAAR also attenuated colitis−related anxiety−like behavior. By 16S RNA gene sequencing analysis, we showed that inhibition of GABAAR partially reversed the gut dysbacteriosis of DSS−induced mice and increased the abundance of beneficial bacteria. Additionally, L. casei Zhang supplementation inhibited the expression of GABAAR in mice with colitis, promoted the proliferation and renewal of colon epithelial cells, and alleviated anxiety−like behavior and intestinal microflora disorder in mice. Thus, GABAAR plays a key role in the beneficial effects of L. casei on DSS−induced colitis in mice.

Transcriptome Profile Analysis of Intestinal Upper Villus Epithelial Cells and Crypt Epithelial Cells of Suckling Piglets.

Simple SummaryTo investigate the genetic reprogramming that drives intestinal epithelial cells (IECs) maturation along the crypt-villus axis, entCerocytes were sequentially isolated from the villus tip to the crypts of porcine small intestine. The present study obtained the intestinal upper villus epithelial cells (F1) and crypt epithelial cells (F3) of 21-day suckling piglets using the divalent chelation and precipitation technique. By mRNA high-throughput sequencing analysis of F1 and F3, it was shown that a total of 672 unigenes were differentially expressed between F1 and F3, including 224 highly expressed and 448 minimally expressed unigenes. The greatest number of differentially expressed genes enriched in signal transduction, e.g., Wnt, Hippo, TGF-beta, mTOR, PI3K-Akt, and MAPK signaling pathways, were closely related to the differentiation, proliferation, maturation and apoptosis of IECs. Our results provide important information for identifying new regulators of IECs differentiation and maturation. Moreover, they can also provide insights into the regulatory mechanisms underlying intestinal epithelial cell renewal and the rapid repair of intestinal mucosal oxidative damage.It is well known that the small intestinal epithelial cells of mammals rapidly undergo differentiation, maturation, and apoptosis. However, few studies have defined the physiological state and gene expression changes of enterocytes along the crypt-villus axis in suckling piglets. In the present study, we obtained the intestinal upper villus epithelial cells (F1) and crypt epithelial cells (F3) of 21-day suckling piglets using the divalent chelation and precipitation technique. The activities of alkaline phosphatase, sucrase, and lactase of F1 were significantly higher (p < 0.05) than those of F3. To explore the differences at the gene transcription level, we compared the global transcriptional profiles of F1 and F3 using RNA-seq analysis technology. A total of 672 differentially expressed genes (DEGs) were identified between F1 and F3, including 224 highly expressed and 448 minimally expressed unigenes. Functional analyses indicated that some DEGs were involved in the transcriptional regulation of nutrient transportation (SLC15A1, SLC5A1, and SLC3A1), cell differentiation (LGR5, HOXA5 and KLF4), cell proliferation (PLK2 and TGFB3), transcriptional regulation (JUN, FOS and ATF3), and signaling transduction (WNT10B and BMP1), suggesting that these genes were related to intestinal epithelial cell maturation and cell renewal. Gene Ontology (GO) enrichment analysis showed that the DEGs were mainly associated with binding, catalytic activity, enzyme regulator activity, and molecular transducer activity. Furthermore, KEGG pathway analysis revealed that the DGEs were categorized into 284 significantly enriched pathways. The greatest number of DEGs enriched in signal transduction, some of which (Wnt, Hippo, TGF-beta, mTOR, PI3K-Akt, and MAPK signaling pathways) were closely related to the differentiation, proliferation, maturation and apoptosis of intestinal epithelial cells. We validated the expression levels of eight DEGs in F1 and F3 using qRT-PCR. The present study revealed temporal and regional changes in mRNA expression between F1 and F3 of suckling piglets, which provides insights into the regulatory mechanisms underlying intestinal epithelial cell renewal and the rapid repair of intestinal mucosal damage.

Expression of extracellular matrix components in the meibomian gland.

PurposeExtracellular matrix (ECM) is a key component of the stem cell local microenvironment. Our study aims to explore the periglandular distribution of major components of ECM in the Meibomian gland (MG).MethodsHuman eyelids and mouse eyelids were collected and processed for immunofluorescence staining.ResultsHuman MG tissues stained positive for collagen IV α1, collagen IV α2, collagen IV α5, and collagen IV α6 around the acini and duct, but negative for collagen IV α3 and collagen IV α4. The mouse MG were stained positive for the same collagen IV subunits as early as postnatal day 15. Laminin α2, laminin β1 and perlecan stained the regions surrounding the acini and the acinar/ductal junction in the human MG, but not the region around the duct. Tenascin-C was found specifically located at the junctions between the acini and the central ducts. Neither agrin nor endostatin was found in the human MG tissues.ConclusionThe ECM expresses specific components in different regions around the MG, which may play a role in MG stem cell regulation, renewal, and regeneration.

Vitamin А in pregnancy and breast feeding: a current view on the problem

For the normal development of pregnancy, a sufficient amount of vitamin A is required. Retinol is involved in the formation of the skeleton of the fetus, provides renewal of epithelial cells of the skin and mucous membranes, is necessary for the growth and development of cells, so that the normalization of vitamin A levels in the body helps reduce the risk of birth defects. Vitamin A deficiency at the stage of intrauterine development creates the prerequisites for the formation of insulin resistance, sensorineural hearing loss, schizophrenia spectrum disorders, disorders of the structure and function of the kidneys in the postnatal period. Vitamin A is involved in many metabolic processes, having an anabolic effect, and its deficiency can lead to the development of fetal malnutrition. No increase in the risk of birth defects caused by vitamin A has been observed at doses below 3,000 mcg/day (10,000 IU/day) of this micronutrient. Postpartum supplementation of vitamin A-rich foods or supplemental sources of vitamin A to the mother’s diet may improve breast milk composition and the infant’s vitamin A stores. Vitamin A affects the metabolism of lipids, iron; its metabolism, in turn, depends on the availability of protein, zinc and other substances. The absorption of vitamin A requires the presence of fats, vitamin E and proteins. With a lack of any of these components, the absorption of vitamin A is reduced, even if the food is enriched with it.

A dual role of lola in Drosophila ovary development: regulating stem cell niche establishment and repressing apoptosis

In Drosophila ovary, niche is composed of somatic cells, including terminal filament cells (TFCs), cap cells (CCs) and escort cells (ECs), which provide extrinsic signals to maintain stem cell renewal or initiate cell differentiation. Niche establishment begins in larval stages when terminal filaments (TFs) are formed, but the underlying mechanism for the development of TFs remains largely unknown. Here we report that transcription factor longitudinals lacking (Lola) is essential for ovary morphogenesis. We showed that Lola protein was expressed abundantly in TFCs and CCs, although also in other cells, and lola was required for the establishment of niche during larval stage. Importantly, we found that knockdown expression of lola induced apoptosis in adult ovary, and that lola affected adult ovary morphogenesis by suppressing expression of Regulator of cullins 1b (Roc1b), an apoptosis-related gene that regulates caspase activation during spermatogenesis. These findings significantly expand our understanding of the mechanisms controlling niche establishment and adult oogenesis in Drosophila.

Hepatic cytochrome P450 8B1 and cholic acid potentiate intestinal epithelial injury in colitis by suppressing intestinal stem cell renewal.

Although disrupted bile acid (BA) homeostasis is implicated in inflammatory bowel disease (IBD), the role of hepatic BA metabolism in the pathogenesis of colitis is poorly understood. Here, we found that cholic acid (CA) levels were increased in patients and mice. Cytochrome P450 8B1 (CYP8B1), which synthesizes CA, was induced in livers of colitic mice. CA-treated or liver Cyp8b1-overexpressing mice developed more severe colitis with compromised repair of the mucosal barrier, whereas Cyp8b1-knockout mice were resistant to colitis. Mechanistically, CA inhibited peroxisome proliferator-activated receptor alpha (PPARα), resulting in impeded fatty acid oxidation (FAO) and impaired Lgr5+ intestinal stem cell (ISC) renewal. A PPARα agonist restored FAO and improved Lgr5+ ISC function. Activation of the farnesoid X receptor (FXR) suppressed liver CYP8B1 expression and ameliorated colitis in mice. This study reveals a connection between the hepatic CYP8B1-CA axis and colitis via regulating intestinal epithelial regeneration, suggesting that BA-based strategies might be beneficial in IBD treatment.

Modelling realistic 3D deformations of simple epithelia in dynamic homeostasis

The maintenance of tissue and organ structures during dynamic homeostasis is often not well understood. In order for a system to be stable, cell renewal, cell migration and cell death must be finely balanced. Moreover, a tissue’s shape must remain relatively unchanged. Simple epithelial tissues occur in various structures throughout the body, such as the endothelium, mesothelium, linings of the lungs, saliva and thyroid glands, and gastrointestinal tract. Despite the prevalence of simple epithelial tissues, there are few models which accurately describe how these tissues maintain a stable structure.Here, we present a novel, 3D, deformable, multilayer, cell-centre model of a simple epithelium. Cell movement is governed by the minimisation of a bending potential across the epithelium, cell–cell adhesion, and viscous effects. We show that the model is capable of maintaining a consistent tissue structure while undergoing self renewal. We also demonstrate the model’s robustness under tissue renewal, cell migration and cell removal. The model presented here is a valuable advancement towards the modelling of tissues and organs with complex and generalised structures.

Akkermansia muciniphila Reduces Peritonitis and Improves Intestinal Tissue Wound Healing after a Colonic Transmural Defect by a MyD88-Dependent Mechanism.

Anastomotic leakage is a major complication following colorectal surgery leading to peritonitis, complications, and mortality. Akkermansia muciniphila has shown beneficial effects on the gut barrier function. Whether A. muciniphila reduces peritonitis and mortality during colonic leakage is unknown. Whether A. muciniphila can directly modulate the expression of genes in the colonic mucosa in humans has never been studied. We investigated the effects of a pretreatment (14 days) with live A. muciniphila prior to surgical colonic perforation on peritonitis, mortality, and wound healing. We used mice with an inducible intestinal-epithelial-cell-specific deletion of MyD88 (IEC-MyD88 KO) to investigate the role of the innate immune system in this context. In a proof-of-concept pilot study, healthy humans were exposed to A. muciniphila for 2 h and colonic biopsies taken before and after colonic instillation for transcriptomic analysis. Seven days after colonic perforation, A.-muciniphila-treated mice had significantly lower mortality and severity of peritonitis. This effect was associated with significant improvements of wound histological healing scores, higher production of IL22, but no changes in the mucus layer thickness or genes involved in cell renewal, proliferation, or differentiation. All these effects were abolished in IEC-MyD88 KO mice. Finally, human subjects exposed to A. muciniphila exhibited an increased level of the bacterium at the mucus level 2 h after instillation and significant changes in the expression of different genes involved in the regulation of cell cycling, gene transcription, immunity, and inflammation in their colonic mucosa. A. muciniphila improves wound healing during transmural colonic wall defect through mechanisms possibly involving IL22 signaling and requiring MyD88 in the intestinal cells. In healthy humans, colonic administration of A. muciniphila is well tolerated and changes the expression of genes involved in the immune pathways.

Genome Maintenance in Mammalian Stem Cells.

Various stem cells in the body are tasked with maintaining tissue homeostasis throughout the life of an organism and thus must be resilient to intrinsic and extrinsic challenges such as infection and injury. Crucial to these challenges is genome maintenance because a high mutational load and persistent DNA lesions impact the production of essential gene products at proper levels and compromise optimal stem cell renewal and differentiation. Genome maintenance requires a robust and well-regulated DNA damage response suited to maintaining specific niches and tissues. In this review, we explore the similarities and differences between diverse stem cell types derived from (or preceding) all germ layers, including extraembryonic tissues. These cells utilize different strategies, including implementation of robust repair mechanisms, modulation of cell cycle checkpoints best suited to eliminating compromised cells, minimization of cell divisions, and differentiation in response to excessive damage.

A non-metabolic function of hexokinase 2 in small cell lung cancer: promotes cancer cell stemness by increasing USP11-mediated CD133 stability.

BackgroundMaintenance of cancer stem‐like cell (CSC) stemness supported by aberrantly regulated cancer cell metabolism is critical for CSC self‐renewal and tumor progression. As a key glycolytic enzyme, hexokinase 2 (HK2) plays an instrumental role in aerobic glycolysis and tumor progression. However, whether HK2 directly contribute to CSC stemness maintenance in small cell lung cancer (SCLC) is largely unclear. In this study, we aimed to investgate whether HK2 independent of its glycolytic activity is directly involved in stemness maintenance of CSC in SCLC.MethodsImmunoblotting analyses were conducted to determine the expression of HK2 in SCLC CSCs and their differentiated counterparts. CSC‐like properties and tumorigenesis of SCLC cells with or without HK2 depletion or overexpression were examined by sphere formation assay and xenograft mouse model. Immunoprecipitation and mass spectrometry analyses were performed to identify the binding proteins of CD133. The expression levels of CD133‐associated and CSC‐relevant proteins were evaluated by immunoblotting, immunoprecipitation, immunofluorescence, and immunohistochemistry assay. RNA expression levels of Nanog, POU5F1, Lin28, HK2, Prominin‐1 were analyzed through quantitative reverse transcription PCR. Polyubiquitination of CD133 was examined by in vitro or in vivo ubiquitination assay. CD133+ cells were sorted by flow cytometry using an anti‐CD133 antibody.ResultsWe demonstrated that HK2 expression was much higher in CSCs of SCLC than in their differentiated counterparts. HK2 depletion inhibited CSC stemness and promoted CSC differentiation. Mechanistically, non‐mitochondrial HK2 directly interacted with CD133 and enhanced CD133 expression without affecting CD133 mRNA levels. The interaction of HK2 and CD133 promoted the binding of the deubiquitinase ubiquitin‐specific protease 11 (USP11) to CD133, thereby inhibiting CD133 polyubiquitylation and degradation. HK2‐mediated upregulation of CD133 expression enhanced the expression of cell renewal regulators, SCLC cell stemness, and tumor growth in mice. In addition, HK2 expression was positively correlated with CD133 expression in human SCLC specimens, and their expression levels were associated with poor prognosis of SCLC patients.ConclusionsThese results revealed a critical non‐metabolic function of HK2 in promotion of cancer cell stemness. Our findings provided new insights into the multifaceted roles of HK2 in tumor development.

Ozone Pollution, Oxidative Stress, Regulatory T Cells and Antioxidants.

Ozone pollution, is a serious health problem worldwide. Repeated exposure to low ozone doses causes a loss of regulation of the oxidation–reduction systems, and also induces a chronic state of oxidative stress. This fact is of special importance for the regulation of different systems including the immune system and the inflammatory response. In addition, the oxidation–reduction balance modulates the homeostasis of these and other complex systems such as metabolism, survival capacity, cell renewal, and brain repair, etc. Likewise, it has been widely demonstrated that in chronic degenerative diseases, an alteration in the oxide-reduction balance is present, and this alteration causes a chronic loss in the regulation of the immune response and the inflammatory process. This is because reactive oxygen species disrupt different signaling pathways. Such pathways are related to the role of regulatory T cells (Treg) in inflammation. This causes an increase in chronic deterioration in the degenerative disease over time. The objective of this review was to study the relationship between environmental ozone pollution, the chronic state of oxidative stress and its effect on Treg cells, which causes the loss of regulation in the inflammatory response as well as the role played by antioxidant systems in various pathologies.

Dietary butyrate, lauric acid and stearic acid improve gut morphology and epithelial cell turnover in weaned piglets.

This study was to evaluate the effects of the supplementation of saturated fatty acids with different chain lengths on growth performance, intestinal morphology, epithelial cell proliferation, differentiation and apoptosis in weaned piglets. Thirty-two weaned piglets (Duroc × Landrace × Yorkshire, BW = 7.81 ± 0.26 kg) were weaned at 21 d and randomly assigned to 1 of 4 experimental treatments: (1) a basal diet (control); (2) control + 0.3% butyrate (BT); (3) control + 0.3% lauric acid (LA); (4) control + 0.3% stearic acid (SA). All piglets were then slaughtered for tissue sampling after having been fed experimental diets for 28 d after weaning. Supplementation of BT increased the gain-to-feed ratio (G:F) (P < 0.05) compared to piglets fed the control diet from 14 to 28 d. In addition, the villus height (VH) to crypt depth (CD) ratio (VH:CD ratio) of the ileum were higher in the BT and LA diets than that of the control diet (P < 0.05). The SA-supplemented diet increased ileal VH (P < 0.05), whereas the BT-supplemented diet increased jejunal CD (P < 0.05). Compared to the control, diets supplemented with BT, LA, or SA all tended to increase jejunal proliferation (Ki67/crypt positive cells) (P = 0.190); diets supplemented with BT or SA significantly increased the number of ki67-positive cells in the ileal crypt (P < 0.05). Furthermore, in the jejunum, the protein expression of activated caspase 3 and villin were increased in piglets fed BT, LA, or SA diets compared to those on the control diet (P < 0.05). In the ileum, compared with the control diet, the BT diet tended to increase the protein level of mammalian phosphorylation target of rapamycin (p-mTOR, P < 0.10); LA or SA diets significantly increased p-mTOR protein expression (P < 0.05). These results show that dietary supplementation of BT, LA, or SA promotes jejunal cell renewal in weaned piglets. At the same time, increased proliferation of ileal crypt cells by promoting p-mTOR expression has beneficial effects on ileal morphology in weaned piglets.

BSCI-05 REPURPOSING PROPOFOL FOR THE TREATMENT OF BRAIN METASTASES

BACKGROUNDRecent clinical studies suggest beneficial effects of propofol anesthesia on tumor progression and patient survival in solid tumors but reported benefits are modest. One potential reason is the relatively short, single exposure to propofol, limited to the surgical period. Brain metastases (BM) are the most common brain tumors in adults. Metastatic tumors develop following infiltration of the brain from primary tumors such as lung, breast, melanoma, and colorectal cancers. BM are treated with combination therapies, including surgery, radiotherapy, chemotherapy, and immunotherapy, however the prognosis of most patients with BM remains dismal. In this report we investigated the effects of propofol plus radiation on cancer stem cells derived from human lung cancer brain metastases (BM-CSCs) and their cross-talk with microglia.OBJECTIVESOur hypothesis is that propofol can be repurposed as a treatment of BM in addition to its anesthetic uses. To test this, we first examined the cytotoxic effects of propofol on cancer stem cells established from BM-CSCs alone and with radiation. Also, we studied the effects of propofol on the cross-talk of BM-CSCs and microglia.RESULTSWe found that propofol 1) exerted inhibitory effect on BM-CSCs self-renewal, stemness and cell proliferation; 2) increased cell death of cancer cells but not normal neural elements; 3) sensitized BM-CSCs to radiation; 4) inhibited the pro-tumorigenic BM-CSCs/ microglia cross-talk by promoting M1 phenotypes of co-cultured microglia.CONCLUSIONSPropofol exerted anti-tumor effects on BM-CSCs including inhibition of cell renewal, proliferation, and mesenchymal transition. Propofol at sensitized BM-GSCs to radiation and at higher concentrations induced cell death. Propofol exerted anti-tumor cytotoxicity also by inhibiting the pro-tumorigenic CSC-microglia cross-talk via secreted extracellular vesicles (EVs). Propofol effects can be exploited as a general anesthetic of choice during tumor resection and should be examined as an anti-tumor agent in sub-anesthetic doses either alone or in combination with radiation.