Sphere Formation Research Articles

Antimicrobial agent chloroxylenol targets β‑catenin‑mediated Wnt signaling and exerts anticancer activity in colorectal cancer.

Chloroxylenol is the active ingredient of the antibacterial agent Dettol. The anticancer effect and underlying mechanisms of this compound and other common antimicrobial agents have not been clearly elucidated. In the present study, the effects of chloroxylenol, benzalkonium chloride, benzethonium chloride, triclosan and triclocarban on β‑catenin‑mediated Wnt signaling in colorectal cancer were evaluated using the SuperTOPFlash reporter assay. It was demonstrated that chloroxylenol, but not the other antimicrobial agents tested, inhibited the Wnt/β‑catenin signaling pathway by decreasing the nuclear translocation of β‑catenin and disrupting β‑catenin/T‑cell factor 4 complex, which resulted in the downregulation of the Wnt target genes Axin2, Survivin and Leucine‑rich G protein‑coupled receptor‑5. Chloroxylenol effectively inhibited the viability, proliferation, migration and invasion, and sphere formation, and induced apoptosis in HCT116 and SW480 cells. Notably, chloroxylenol attenuated the growth of colorectal cancer in the MC38 cell xenograft model and inhibited organoid formation by the patient‑derived cells. Chloroxylenol also demonstrated inhibitory effects on the stemness of colorectal cancer cells. The results of the present study demonstrated that chloroxylenol could exert anti‑tumor activities in colorectal cancer by targeting the Wnt/β‑catenin signaling pathway, which provided an insight into its therapeutic potential as an anticancer agent.

Polyphenol modification of graphene-stabilized emulsions to form electrically conductive polymer spheres

HypothesisPolyphenols, specifically tannic acid, should increase the hydrophilicity of graphene when added during the interfacial exfoliation through π-π stacking. Following Bancroft’s rule, increasing the hydrophilicity of graphene will result in a phase inversion of water-in-oil emulsions stabilized by graphene. Polymerization of the oil phase will then lead to graphene-coated spheres rather than graphene-stabilized polyHIPEs. ExperimentsOptical particle sizing, microscopy, contact angle, and electrical conductivity measurements were performed to determine the mechanism of sphere formation in graphene-stabilized emulsions modified with tannic acid. Studies focused on the effect of graphite flake size, graphite concentration, tannic acid concentration, and oil phase composition. Particle sizing and scanning electron microscopy examined the spheres' size, shape, and surface morphology. Contact angle measurements gave insight into the change in graphene surface energy. Conductivity studies examined the graphene shell surrounding the spheres. FindingsAdding tannic acid to graphene-stabilized emulsions induced a phase change from water-in-oil to oil-in-water. Contact angle measurements confirmed greater hydrophilicity of graphene in the presence of tannic acid. However, very high tannic acid concentrations led to a decrease in the stability of the emulsion. Varying the graphite flake size and concentration resulted in morphology and conductivity changes. Dilution of the monomer phase produced hollow microcapsules.

Ionizing radiation normalizes the features of active breast cancer stromal fibroblasts and suppresses their paracrine pro-carcinogenic effects

BackgroundRadiotherapy is an important therapeutic strategy for breast cancer patients through reducing the chances of recurrence and metastasis, which are fueled by cancer-associated fibroblasts (CAFs). Thereby, we addressed here the effect of various doses of X-rays on breast CAFs and their adjacent counterparts. MethodsWe have used WST1 and annexin V-associated with flow cytometry to test the cytotoxic effects of X-rays. Immunoblotting and ELISA was used to assess the expression/secretion of various proteins. Immunohistochemistry was utilized to determine the level of β-galactosidase and Ki-67. Sphere formation assay was used to test the ability of breast cancer cells to form tumorspheres. Orthotopic tumor xenografts were also used to evaluate the effect of X-ray-treated breast stromal fibroblasts on breast cancer tumor growth in vivo. ResultsBreast stromal fibroblasts showed high resistance to X-rays. While the low dose (5 Gy) inhibited cell proliferation and the active features of CAFs, the higher doses (16 and 50 Gy) promoted senescence. However, this was not accompanied by the senescence-associated secretory phenotype (SASP), but rather a reduction in the synthesis/secretion of various cancer-associated cytokines. Additionally, X-rays suppressed the features of active breast stromal fibroblasts, and their paracrine pro-carcinogenic effects. The ablative dose (16 Gy) inhibited the capacity of active stromal fibroblasts to promote the pro-metastatic processes epithelial-to-mesenchymal transition, the formation of cancer stem cells, as well as the growth of humanized orthotopic breast tumor xenografts. ConclusionTogether, these findings indicate that X-rays can normalize the features of active breast stromal fibroblasts through promoting senescence without SASP.

Regulating POLR3G by MicroRNA-26a-5p as a promising therapeutic target of lung cancer stemness and chemosensitivity.

Cancer stem cells (CSCs) identified in lung cancer exhibit resistance to chemotherapy, radiotherapy, and targeted therapy. Therefore, a technology for controlling CSCs is needed to overcome such resistance to cancer therapy. Various evidences about the association between epithelial-mesenchymal transition related transcriptomic alteration and acquisition of CSC phenotype have been proposed recently. Down-regulated miR-26a-5p is closely related to mesenchymal-like lung cancer cell lines. These findings suggest that miR-26a-5p might be involved in lung cancer stemness. RNA polymerase III subunit G (POLR3G) was selected as a candidate target of miR-26a-5p related to cancer stemness. It was found that miR-26a-5p directly regulates the expression of POLR3G.Overexpression of miR-26a-5p induced a marked reduction of colony formation and sphere formation. Co-treatment of miR-26a-5p and paclitaxel decreased cell growth, suggesting that miR-26a-5p might play a role as a chemotherapy sensitizer. In the cancer genome atlas data, high miR-26a-5p and low POLR3G expression were also related to higher survival rate of patients with lung adenocarcinoma. These results suggest that miR-26a-5p can suppress lung cancer stemness and make cancer cell become sensitive to chemotherapy. This finding provides a novel insight into a potential lung cancer treatment by regulating stemness.

LHPP promotes the intracellular reactive oxygen species accumulation and sensitivity of gastric cancer to cisplatin via JNK and p38 MAPK pathways.

Cisplatin is the first-line chemotherapy drug for the treatment of gastric cancer (GC) patients. However, GC patients who are resistant to cisplatin often do not benefit from it. Therefore, finding a key molecule that affects cisplatin sensitivity is expected to enhance the efficacy of cisplatin in GC treatment. The human GC cell lines SGC-7901 and BGC-823 were used. The protein chip array was used to screen the cisplatin-resistance genes from the complete response and non-complete response GC patients' tissues, then, the differential gene expression analysis, GO function annotation analysis, and KEGG pathway enrichment analysis were performed. The GC tissue chip in the GEO database was analyzed to screen the target gene. Flow cytometry, Hoechst 33342 staining assay, Western Blot, MTT, tumor sphere formation, cell cycle, and apoptosis assays were performed to explore the effect of Phospholysine Phosphohistidine Inorganic Pyrophosphate Phosphatase (LHPP) on the apoptosis, stemness, and reactive oxygen species (ROS) accumulation of cisplatin-resistant GC cells treated with cisplatin. In vivo, the cisplatin-resistant GC cell lines transfected with pcDNA-LHPP or si-LHPP were injected subcutaneously into mice to construct GC subcutaneous xenograft GC models. Based on protein chip array and bioinformatics analysis, it was found that LHPP is the core molecule in the cisplatin resistance regulatory network in GC, and its expression is down-regulated in GC cisplatin-resistant tissues and cells. In vitro and in vivo experimental results show that the up-regulated expression of LHPP is closely related to the increase in sensitivity of GC to cisplatin. Mechanically, we found that overexpression of LHPP may inhibit the activation of the JNK and p38 MAPK pathways, promote cisplatin-induced ROS accumulation, suppress stemness, and enhance the sensitivity of GC to cisplatin. Up-regulation of LHPP may inhibit the activation of the JNK and p38 MAPK pathways, attenuate stemness, and enhance the accumulation of intracellular ROS, thereby promoting cisplatin-mediated GC cell apoptosis and enhancing cisplatin sensitivity.

Fabrication of tunable band gap carbon based zinc nanocomposites for enhanced capacitive behaviour

This manuscript highlights the tunable properties of carbon nanospheres by controlling the concentration of zinc nitrate in them. Zinc nitrate has converted the phase of carbon spheres, which has also altered the optical, structural and electrochemical properties of carbon nanospheres by forming nanocomposites. Carbon nanospheres and their nanocomposites have been synthesized by using a two-step hydrothermal method. X-ray diffraction analysis of the as synthesized material revealed the formation of carbon spheres and their nanocomposites. It is also observed that the crystallinity of the as synthesized material increases as the concentration of Zn(NO3)2.6H2O increases. UV- visible measurements revealed a blue shift in the as-synthesized samples. With the increase in the concentration of zinc, the band gap was also found to increase from 0.6 eV to 4.7 eV. The morphological and microstructural analysis of the as-synthesized samples showed the formation of nanospheres for as-synthesized carbon, and nano flakes for carbon nanocomposites. Fourier-Transform Infrared Spectroscopy (FTIR) measurement provided the information about the molecular structure and vibrational bands present in the samples. Electrochemical analysis of the thin film revealed the capacitive behaviour of the material. The aerial capacitance and Nyquist plot represents the capacitive properties of the material. The present study on carbon nanospheres and their nanocomposites showed that the material is a potential candidate for the application in capacitors, supercapacitors and energy storage devices.

FORMATION OF EDUCATIONAL INDEPENDENCE AS A CONDITION FOR ACADEMIC PERFORMANCE

Introduction. Improving the academic performance of students is one of the urgent problems of modern higher education, since academic activity is a determining indicator of the readiness of university graduates for professional activities. Academic achievement depends on many factors, among which the authors emphasize the ability and desire to study as the basis of educational independence.
 The purpose of the study. The purpose of the study is to examine the degree of influence of the formation of the motivational sphere and skills of independent work, determining the educational independence, on the academic performance.
 Materials and methods. The following methods were used to achieve the research objectives: questioning, testing, comparative and correlation analysis. The results were processed by methods of mathematical statistics in SPSS Statistics 26.
 Results. The results obtained allow us to conclude that the formation of independent work skills has a relationship with academic performance, while strong positive relationships were identified between the indicators for all five blocks of independent work skills, as well as internal and external positive motivation. Practical recommendations on maintaining the optimal motivation of students are presented.
 Practical implications. The results can be used to further investigate the factors that influence academic performance and find ways to improve it.
 Conclusion. After analyzing the results, the authors concluded that the important conditions for academic performance are the formation of the structure of independent educational activities of students, as well as the leading role of their internal and external positive motivation. Higher educational achievements are possible only with high levels of formation of the structure of students’ own educational activities and their positive motivation. From here follow two main directions related to the prevention of academic failure and dropout of students: the formation of optimal ways for them to independently solve educational, and in the future, professional problems and, in addition, high positive motivation at all its hierarchical levels.

YY1-regulated lncRNA SOCS2-AS1 suppresses HCC cell stemness and progression via miR-454-3p/CPEB1

BackgroundCancer stem cells are one fundamental reason for the high recurrence rate of hepatocellular carcinoma (HCC) and its resistance to treatment. This study explored the mechanism by which SOCS2-AS1 affects HCC cell stemness. MethodsStem cells of HCC cell lines Huh7 and SNU-398 were sorted as NANOG-positive by flow cytometry. Stem cell sphere formation ability was detected. Stem cell viability, migration, invasion, and apoptosis were assessed by colony formation assays, Transwell assays, wound-healing assays, and TUNEL assays, respectively. The binding sites for SOCS2-AS1, miR-454-3p, miR-454-3p, and CPEB1 mRNA were assessed by dual-luciferase reporter assays. Quantitative real-time PCR (qPCR) and Western blot studies were performed to evaluate gene expression levels. ChIP and EMSA assays were conducted to confirm that YY1 binds with the SOCS2-AS1 promoter. A subcutaneous xenograft model was used to verify results in vivo. Tumor tissues were analyzed by H&E and TUNEL staining. ResultsSOCS2-AS1 was expressed at low levels in NANOG+ HCC stem cells, and HCC patients with a high level of SOCS2-AS1 expression had a higher survival rate. SOCS2-AS1 inhibited HCC cell stemness, migration, and invasion, and increased the cisplatin sensitivity of HCC cells by regulating miR-454-3p/CPEB1. YY1 was confirmed as a transcription factor of SOCS2-AS1, and served to inhibit SOCS2-AS1 transcription. YY1 knockdown suppressed HCC stemness via SOCS2-AS1. The role of SOCS2-AS1 was confirmed in a subcutaneous xenograft model, and SOCS2-AS1 overexpression enhanced the inhibitory effect of cisplatin on HCC in vivo. ConclusionsYY1-regulated lncRNA SOCS2-AS1 suppresses HCC cell stemness and progression via miR-454-3p/CPEB1.

Transcriptional factor BRD4 promotes the stemness of esophageal cancer by activating the nuclear PD-L1/RelB axis.

Esophageal cancer (EC) is a prevalent malignancy associated with therapeutic resistance and poor prognosis. This study investigates the role of programmed death-ligand 1 (PD-L1) in esophageal cancer stem cell (ECSC) formation. ECSCs were enriched and characterized using various assays. We found that both PD-L1 and bromodomain-containing protein 4 (BRD4) were upregulated in ECSCs, promoting their stemness. Inhibiting BRD4 suppressed ECSC markers expression and sphere formation. Furthermore, BRD4 inhibitors downregulated membrane and nuclear PD-L1 levels, with knockdown of PD-L1 inhibiting ECSC formation. PD-L1 degraders also affected PD-L1 and its downstream effector RelB expression. Moreover, inhibiting RelB influenced sphere formation through interleukin-6 expression. This study reveals the critical role of the BRD4/nuclear PD-L1/RelB axis in ECSC formation, highlighting nuclear PD-L1 as a potential immunotherapeutic target for refractory EC.

Influence of Bruton's Tyrosine Kinase (BTK) on Epithelial-Mesenchymal Transition (EMT) Processes and Cancer Stem Cell (CSC) Enrichment in Head and Neck Squamous Cell Carcinoma (HNSCC).

Constitutively active kinases play a crucial role in carcinogenesis, and their inhibition is a common target for molecular tumor therapy. We recently discovered the expression of two oncogenic isoforms of Bruton's Tyrosine Kinase (BTK) in head and neck squamous cell carcinoma (HNSCC), Btk-p80 and BTK-p65. However, the precise role of BTK in HNSCC remains unclear. Analyses of a tissue microarray containing benign and malignant as well as inflammatory tissue samples of the head and neck region revealed the preferential expression of BTK-p80 in malignant tissue, whereas BTK-p65 expression was confirmed in over 80% of analyzed metastatic head and neck tumor cases. Therefore, processes associated with metastasis, like cancer stem cell (CSC) enrichment and the epithelial-mesenchymal transition (EMT), which in turn depend on an appropriate cytokine milieu, were analyzed. Treatment of HNSCC-derived cell lines cultured under 3D conditions with the BTK inhibitor AVL-292 caused reduced sphere formation, which was accompanied by reduced numbers of ALDH1A1+ CSCs as well as biological changes associated with the EMT. Moreover, we observed reduced NF-κB expression as well as altered NF-κB dependent pro-tumorigenic and EMT-associated cytokine release of IL-6, IFNγ, and TNFα when BTK activity was dampened. Therefore, an autocrine regulation of the oncogenic BTK-dependent process in HNSCC can be suggested, with BTK inhibition expected to be an effective treatment option for HNSCC.

Metformin inhibits self-renewal of colorectal cancer stem cells by inhibiting mitochondrial oxidative phosphorylation

To investigate the mechanism of metformin for inhibiting self-renewal of colorectal cancer stem cells (CSCs). CSCs were sorted from Wnt reporter- transfected colorectal cancer patient-derived organoids (PDOs) by fluorescence-activated cell sorting (FACS) and treated with metformin. The changes in self-renewal of the cells were assessed using sphere formation, colony formation and limiting dilution assays. The mRNA expressions of genes related with stemness and differentiation and Wnt target genes was detected by qRT-PCR. Wnt activity was assessed using flow cytometry in the CSCs. Seahorse analysis was used to evaluate cellular oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) after metformin treatment. Mitochondrial membrane potential levels were detected with TMRE staining, and reactive oxygen species (ROS) levels were detected using MitoSOX staining. Galactose (10 mmol/L), metformin (10 μmol/L), NAC (5 mmol/L), and galactose+metformin were used to modulate ROS levels in the CSCs, and sphere-formation assay and flow cytometry were used to assess the changes in self- renewal capacity and Wnt activity. The effect of lentiviral transfection of yeast NADH dehydrogenase NDI1 on TMRE staining, MitoSOX staining and Wnt activity in the CSCs were analyzed with flow cytometry. Metformin significantly decreased the capacities of CSCs to form spheres, colonies and xenografts and reduced Wnt activity in the cells (P < 0.01). The mRNA levels of stemness-related genes and Wnt target genes decreased significantly while those of differentiation-related genes increased in metformin-treated CSCs (P < 0.05), which also showed significantly decreased OCR, TMRE and ROS levels with enhanced ECAR (P < 0.001). Galactose significantly increased sphereforming capacity, ROS levels and Wnt activity of the cells, and these effects were significantly inhibited by metformin (P < 0.05). Transfection of the CSCs with NDI1 significantly attenuated the inhibitory effects of metformin on proportion of CSCs and Wnt signaling pathway activity. Metformin reduces mitochondrial oxidative phosphorylation and ROS levels by inhibiting mitochondrial complex Ⅰ, thereby suppressing Wnt signaling pathway to reduce selfrenewal ability of colorectal CSCs.

OLR1 Is a Pan-Cancer Prognostic and Immunotherapeutic Predictor Associated with EMT and Cuproptosis in HNSCC.

Metabolism plays a critical role in cancer. OLR1 has been implicated in cardiovascular and metabolic disorders, while its association with tumorigenesis and tumor immunity remains poorly defined in the literature. We conducted comprehensive pan-cancer analyses based on the TCGA database to examine OLR1 expression and its prognostic implications. Correlations between OLR1 expression level and tumor immunity and immunotherapy were investigated by immune infiltration, enrichment, and TIDE analysis methods. Immunohistochemistry detected OLR1 expression in HNSCC. We used the GSEA method to explore the potential signaling pathways in which OLR1 is involved, and a correlation analysis to investigate the relationships between OLR1 and epithelial-mesenchymal transition (EMT) and cuproptosis. In addition, the effects of OLR1 knockdown on the EMT process, invasion, stemness, and cuproptosis of HNSCC cells were examined by scratch, Transwell, CCK8, sphere formation, and flow cytometry, while changes in related proteins were detected using the immunoblotting method. OLR1 is highly expressed in most cancers, and it is associated with patient prognosis. OLR1 expression positively correlates with immunosuppressive cell infiltration and immune checkpoint molecules, while being negatively associated with effector T cells. Moreover, significant correlations are observed between OLR1 expression and tumor mutation burden (TMB) and microsatellite instability (MSI) in some cancers. In HNSCC, OLR1 expression is related to advanced clinicopathological factors and unfavorable outcomes. Patients with high OLR1 expression levels are prone to experience immune escape and benefit less from immune checkpoint inhibitor (ICI) therapy. Moreover, OLR1 expression may affect EMT, stemness, and cuproptosis resistance outcomes. OLR1 is an immune-related prognostic biomarker with potential as a prognostic indicator for immunotherapy, and it may also be involved in regulating the EMT process and cuproptosis in HNSCC.

Improvement on the Radiation Stability of Multi-scale Supramolecular Assembly in Ionic Liquid-Based Extraction.

The multi-scale supramolecular assembly (MSSA)-based extraction strategy with hydroxyl-functionalized ionic liquid (IL) is promising in the separation of metal ions from radioactive environments for which a comprehensive understanding toward the radiation stability of the MSSA system is necessary. Herein, we report on the analyses of the radiation stability of MSSA in extraction, especially the adopted ILs 1-(2-hydroxyethyl)-3-methylimidazolium bis(trifluoromethylsulfonyl) imide (IL-1) and 1-(2-hydroxyethyl)-2,3-dimethylimidazolium bis(trifluoromethylsulfonyl) imide (IL-2), by UV-vis, 1H NMR, and ESI-HRMS. It was found that the macroscopic assembly (MA) sphere could not be formed after γ irradiation on the extraction system with IL-1. On the contrary, the trisubstituted IL-2 instead of the disubstituted IL-1 remarkably improved the radiation stability of the MSSA system to guarantee the formation of the MA sphere. The high extraction efficiency could be kept, and the mechanism of such an improvement was revealed.

CD58 acts as a tumor promotor in hepatocellular carcinoma via activating the AKT/GSK-3β/β-catenin pathway

BackgroundHepatocellular carcinoma (HCC) is one of the most prevalent malignancies worldwide because of rapid progression and high incidence of metastasis or recurrence. Accumulating evidence shows that CD58-expressing tumor cell is implicated in development of various cancers. The present study aimed to reveal the functional significance of CD58 in HCC progression and the underlying mechanisms.MethodsImmunohistochemical staining (IHC), and western blotting were used to detect the expression of CD58 in HCC tissues and cells. The levels of sCD58 (a soluble form of CD58) in the cell supernatants and serum were assessed by ELISA. CCK-8, colony formation, and xenograft assays were used to detect the function of CD58 on proliferation in vitro and in vivo. Transwell assay and sphere formation assay were performed to evaluate the effect of CD58 and sCD58 on metastasis and self-renewal ability of HCC cells. Western blotting, immunofluorescence (IF), TOP/FOP Flash reporter assay, and subcellular fractionation assay were conducted to investigate the molecular regulation between CD58/sCD58 and AKT/GSK-3β/β-catenin axis in HCC cells.ResultsCD58 was significantly upregulated in HCC tissues. Elevation of CD58 expression correlated with more satellite foci and vascular invasion, and poorer tumor-free and overall survival in HCC patients. Higher sCD58 levels were in HCC patients' serum compared to healthy individuals. Functionally, CD58 promotes the proliferation of HCC cells in vitro and in vivo. Meanwhile, CD58 and sCD58 induce metastasis, self-renewal and pluripotency in HCC cells in vitro. Mechanistically, CD58 activates the AKT/GSK-3β/β-catenin signaling pathway by increasing phosphorylation of AKT or GSK3β signaling, promoting expression of Wnt/β-catenin target proteins and TCF/LEF-mediated transcriptional activity. Furthermore, AKT activator SC-79 or inhibitor LY294002 abolished the inhibitory effect of CD58 silencing on the proliferation, metastasis, and stemness of HCC cells.ConclusionsTaken together, CD58 promotes HCC progression and metastasis via activating the AKT/GSK-3β/β-catenin pathway, suggesting that CD58 is a novel prognostic biomarker and therapeutic target for HCC.Graphical

AXIOLOGICAL DIMENSION OF HIGHER EDUCATION IN UKRAINE AND THE IMPACT IT MAKES ON ITS DEVELOPMENT

The article presents the main principles of the axiological approach to the formation of the content of higher education in the context of current global trends and challenges caused by the modernization of the educational environments in Ukraine and the EU. The socio-economic, political, technological and cultural transformations that have been taking place in the world over the recent decades have had a significant impact on the development of education. In 2018, the EU adopted the key competencies for lifelong learning, including digital competence and entrepreneurship, which were introduced for the first time. This demonstrates that in order for the graduates to be employed universities have to respond quickly to the new labour market requirements and promote the formation of a competitive personality that has well-developed life, career and study skills. The EU-Ukraine Association Agreement was signed in 2014, and since then the cooperation between Ukrainian and European educational institutions has got intensified. In this respect, it is vital to consider the axiological aspects of higher education and the determination of value components of the content and criteria of high quality education. By definition, the axiological approach proclaims the human being as the highest value of society and allows integrating the development of universal, national, professional and personal values in the educational process. The university is able to be the environment where the formation and consolidation of the value sphere of the individual takes place. The content of the value component of education is formed at the state level within regulatory documents and at the university level within the framework of educational and professional programs, training courses, compulsory and elective disciplines. However, formal copying of the value-based education of other countries cannot bring the desired results, as the factors that determine which values are the basis for the content of higher education in European countries differ from those in Ukraine. A comparative analysis of the value component of the content of university education in Ukraine and the EU countries shows that national traditions of lifestyle, production practice, politics and culture are essential the development of education

Biomimetic Microenvironmental Stiffness Boosts Stemness of Pancreatic Ductal Adenocarcinoma via Augmented Autophagy.

Pancreatic ductal adenocarcinoma (PDAC) features high recurrence rates and intensified lethality, accompanied by stiffening of the extracellular matrix (ECM) microenvironment, which is mainly due to the deposition, remodeling, and cross-linking of collagen. Boosted stemness plays an essential role during occurrence and progression, which indicates a poor prognosis. Therefore, it is of great importance to understand the effect of the underlying interaction of matrix stiffness and stemness on PDAC. For this purpose, a methacrylated gelatin (GelMA) hydrogel with tunable stiffness was applied for incubating MIA PaCa-2 and PANC-1 cells. The results demonstrated that compared to the soft group (5% GelMA, w/v), the expression of stemness-related genes (SOX2, OCT4, and NANOG) in the stiff group (10% GelMA, w/v) displayed pronounced elevation as well as sphere formation. Intriguingly, we also observed that matrix stiffness regulated autophagy of PDAC, which played a momentous role in stemness promotion. In order to clarify the underlying relationship between matrix stiffness-mediated cell autophagy and stemness, rescue experiments with rapamycin and chloroquine were conducted with transmission electron microscopy, immunofluorescence staining, sphere formation, and qRT-PCR assays to evaluate the level of stemness and autophagy. For exploring the molecular mechanism in depth, RNA-seq and differential expression of miRNAs were carried out, which may sensor and respond to matrix stiffness during the regulation of stemness and autophagy. In conclusion, we validated that blocking autophagy repressed the stemness induced by matrix stiffness in PDAC and provided a potential therapeutic strategy for this aggressive cancer.

AR antagonists develop drug resistance through TOMM20 autophagic degradation-promoted transformation to neuroendocrine prostate cancer

BackgroundProstate cancer(PCa) is the most commonly occurring male cancer in the USA. Abiraterone or Enzalutamide have been approved for the treatment of metastatic castration-resistant prostate cancer (CRPC). However, the treatment-emergent neuroendocrine PCa (t-NEPC) may develop, resulting in drug resistance in about 10–17% CRPC patients. The detailed mechanisms remain unclear..MethodsThe expression correlation of TOMM20 and AR in PCa was determined by analyzing publicly available datasets, or by IHC staining in tumor specimens. The protein interaction of TOMM20 and AR was validated by co-immunoprecipitation or GST pull-down assay. The impact of TOMM20 depletion on drug sensitivity were elucidated by assays of cell proliferation, invasion, sphere formation, xenograft growth and intravenous metastasis. The intracellular ROS level was measured by flow cytometry, and the NEPC transdifferentiation and characteristics of cancer stem-like cells were validated by RNA-seq, RT-PCR and western blotting.ResultsThe protein level of TOMM20 is positively correlated with AR in PCa cells and specimens. TOMM20 protein physically interacts with AR. AR antagonists induced the protein degradation of TOMM20 through autophagy-lysosomal pathway, thereby elevating the intracellular ROS level and activating PI3K/AKT signaling pathway. When TOMM20 was depleted, PCa cells underwent EMT, acquired the characteristics of cancer stem-like cells, and developed resistance to AR antagonists. The stable depletion of TOMM20 promoted the transdifferentiation of PCa adenocarcinoma into NEPC and metastasis. Conversely, the rescue of TOMM20 re-sensitized the resistant PCa cells to AR antagonists.ConclusionsTOMM20 protein degradation induced by AR antagonists promoted the transdifferentiation of PCa to NEPC, thereby revealing a novel molecular mechanism by which AR antagonists develop drug resistance through mitochondrial outer membrane-mediated signaling pathway. These findings suggested that the decreasing or loss of TOMM20 expression in PCa tissues might become a useful predictor of PCa resistance to AR antagonists.

Ex vivo expansion of lung cancer-derived disseminated cancer cells from lymph nodes identifies cells associated with metastatic progression.

The cellular basis of the apparent aggressiveness in lung cancer is poorly understood but likely associated with functional or molecular features of disseminated cancer cells (DCCs). DCCs from epithelial cancers are mostly detected by antibodies directed against histogenetic markers such as cytokeratin or EpCAM. It has been argued that marker-negative metastatic founder cells might escape detection. We therefore used ex vivo sphere formation for functional detection of candidate metastasis founders. We generated cell suspensions from 199 LN samples of 131 lung cancer patients and placed them into non-adherent cell culture. Sphere formation was associated with detection of DCCs using EpCAM immunocytology and with significantly poorer prognosis. The prognostic impact of sphere formation was strongly associated with high numbers of EpCAM-positive DCCs and aberrant genotypes of expanded spheres. We also noted sphere formation in patients with no evidence of lymphatic spread, however such spheres showed infrequent expression of signature genes associated with spheres from EpCAM-positive samples and displayed neither typical lung cancer mutations (KRAS, TP53, ERBB1) nor copy number variations, but might be linked to disease progression >5 years post curative surgery. We conclude that EpCAM identifies relevant disease-driving DCCs, that such cells can be expanded for model generation and that further research is needed to clarify the functional and prognostic role of rare EpCAM-negative sphere forming cells.

Подготовка тетраэтоксисилана для получения сферических частиц кремнезема. Часть 2. Примеси и их влияние на размеры формирующихся глобул

Monodisperse spherical silica particles, as a basis for creating supramolecular 3D matrices, are currently of increasing interest due to the prospects for their wide application in the synthesis of new nanocomposite materials. At the same time, one of main problems of their large-scale synthesis is associated with the unstable behavior of tetraethoxysilane (TEOS) during hydrolysis, which leads both to the deviation of the particles from a given size and their shape from a spherical one. In this paper, on the basis of the study of tetraethoxysilane from various manufacturers, we continued to work on determining factors that can influence the process of hydrolysis of tetraethoxysilane and, as a result, the monodispersity and size of the formed particles, and also other factors. To solve this task, all TEOS samples were studied by various physicochemical methods of analysis, including Fourier IR and Raman spectroscopy, gas chromatography-mass spectrometry. As a result of the studies, we showed that the presence of di- and trisiloxanes in the system did not significantly affect the size of the formed silica particles, while significantly accelerating the rate of formation of silica spheres. Moreover, the presence of methoxyl groups in the initial silane, and an insignificant content of ethanol in the system did not affect the size stability of the formed particles. At the same time, the replacement of part of the ethoxy groups in the initial TEOS by methyl or ethyl groups, as they are not capable of participating in the hydrolysis reaction, largely contribute to the deviation of sizes of the formed silica particles from each other. The results obtained on the effect of impurities on the size of the formed silica globules are important, among other things, for understanding the processes of formation of natural supramolecular structures of silica.

LANCL1, a cell surface protein, promotes liver tumor initiation through FAM49B-Rac1 axis to suppress oxidative stress.

HCC is an aggressive cancer with a poor clinical outcome. Understanding the mechanisms that drive tumor initiation is important for improving treatment strategy. This study aimed to identify functional cell membrane proteins that promote HCC tumor initiation. Tailor-made siRNA library screening was performed for all membrane protein-encoding genes that are upregulated in human HCC (n = 134), with sphere formation as a surrogate readout for tumor initiation. Upon confirmation of membranous localization by immunofluorescence and tumor initiation ability by limiting dilution assay in vivo, LanC-like protein-1 (LANCL1) was selected for further characterization. LANCL1 suppressed intracellular reactive oxygen species (ROS) and promoted tumorigenicity both in vitro and in vivo. Mechanistically, with mass spectrometry, FAM49B was identified as a downstream binding partner of LANCL1. LANCL1 stabilized FAM49B by blocking the interaction of FAM49B with the specific E3 ubiquitin ligase TRIM21, thus protecting FAM49B from ubiquitin-proteasome degradation. The LANCL1-FAM49B axis suppressed the Rac1-NADPH oxidase-driven ROS production, but this suppression of ROS was independent of the glutathione transferase function of LANCL1. Clinically, HCCs with high co-expression of LANCL1 and FAM49B were associated with more advanced tumor stage, poorer overall survival, and disease-free survival. In addition, anti-LANCL1 antibodies targeting the extracellular N-terminal domain were able to suppress the self-renewal ability, as demonstrated by the sphere formation ability of HCC cells. Our data showed that LANCL1 is a cell surface protein and a key contributor to HCC initiation. Targeting the LANCL1-FAM49B-Rac1-NADPH oxidase-ROS signaling axis may be a promising therapeutic strategy for HCC.