Levels Of Snail Research Articles

MiR-455-3p inhibits gastric cancer progression by repressing Wnt/β-catenin signaling through binding to ARMC8

BackgroundGlobally, gastric cancer (GC) is one of the world’s most widespread malignancies, with persistent high mortality and morbidity rates. Increasing evidence now suggests that microRNAs (miRNAs) participate in many biological processes, with miR-455-3p having key roles in the progression of diverse cancers. Nevertheless, miR-455-3p function and expression in GC remain unclear.MethodsWe explored miR-455-3p expression in GC using quantitative polymerase chain reaction (qPCR). To further examine the effect of miR-455-3p in GC, after transfecting miR-455-3p mimics or inhibitors into GC cells, 5-ethynyl-2’-deoxyuridine (EdU) incorporation and colony formation assays were performed to examine cell proliferation. Flow cytometry was used to detect apoptosis, and expression levels of Bax, Bcl-2, Snail, N-cadherin, E-cadherin, and Caspase-3 were assessed by western blotting (WB). Using online databases and luciferase assays, we identified armadillo repeat-containing protein 8 (ARMC8) as a promising target of miR-455-3p. A mouse tumor model was established to investigate actions of miR-455-3p in vivo. Expression levels of C-myc, cyclinD1, and β-catenin were examined using WB and immunofluorescence.ResultsMiR-455-3p expression was attenuated in GC tissue and cell lines. MiR-455-3p overexpression inhibited GC cell proliferation, epithelial-mesenchymal transition (EMT), as well as facilitated apoptosis, while suppression of miR-455-3p had the opposite effects. From luciferase assays, we confirmed that ARMC8 was a novel and direct downstream target gene of miR-455-3p, and that the tumor suppressive role of miR-455-3p was in part reversed due to ARMC8 overexpression. Moreover, miR-455-3p inhibited GC growth in vivo via ARMC8. We also observed that miR-455-3p repressed canonical Wnt pathway activation by binding to ARMC8.ConclusionsMiR-455-3p exerted tumor inhibitory effects in GC by targeting ARMC8. Therefore, intervening in the miR-455-3p/ARMC8/Wnt/βcatenin axis could be a promising novel treatment strategy for GC.

LINC01117 inhibits invasion and migration of lung adenocarcinoma through influencing EMT process.

Studying the mechanism of action of LncRNAs in lung adenocarcinoma (LUAD) is of great importance for an in-depth understanding of the molecular mechanism of lung adeno carcinogenesis and development. The aim is to identify a long non-coding RNA LINC01117 that is specifically and highly expressed in LUAD cells and to investigate its biological functions and molecular mechanisms in LUAD cells, providing a new potential target for targeting LUAD therapy. This study used publicly available data downloaded from The Cancer Genome Atlas (TCGA) database. Construction of siRNA and overexpression plasmid-packed lentiviral constructs were used to knock down and increase the expression of LINC01117 in LUAD cells. The effect of LINC01117 on LUAD cell migration and invasion was verified by scratch assays and Transwell assays. Western blot assays were performed to verify the effect of knocking down LINC01117 expression on key proteins of the EMT process. The effect of overexpression and knockdown LINC01117 expression on key proteins of the EMT process and the nuclear and cytoplasmic distribution of YAP1, a key effector molecule of the Hippo pathway, was verified by Western blot assays. LINC01117 expression was upregulated in LUAD tissues and cell lines. Clinical correlation and prognostic analyses showed that LINC01117 was associated with poorer clinical features (staging and N classification) and poorer prognosis and could be analyzed as an independent prognostic factor. Cell migration and invasion were significantly inhibited in the knockdown group compared to the control group; in contrast, cell migration and invasion were promoted in the overexpression group. Overexpression of LINC01117 resulted in down-regulation of E-cadherin expression and increased expression levels of N-cadherin, vimentin, ZEB1, snail and slug; in contrast, knockdown of LINC01117 appeared to have the opposite effect. Furthermore, knockdown of LINC01117 increased the enrichment of YAP1 protein in the cytoplasm and reduced its level in the nucleus; overexpression of LINC01117 produced the opposite intracellular distribution results. LINC01117 was highly expressed in LUAD, and knockdown of LINC01117 significantly inhibited the migration and invasion of LUAD cells, while overexpression of LINC01117 significantly promoted the migration and invasion of LUAD cells, and affected the EMT process, and was able to alter the distribution of YAP1 in the nucleus and cytoplasm. This suggests that LINC01117 may regulate the activity of the Hippo pathway by altering the nuclear and cytoplasmic distribution of YAP1, which in turn induces the EMT process in lung adenocarcinoma cells and thus exerts a pro-cancer effect. It suggests that LINC01117 may play a key role in the occurrence and development of LUAD.

Mesenchymal stem cells facilitate gastric cancer cell growth and oxaliplatin resista nce via upregulation of SMARCA5

Purpose: To evaluate the role of mesenchymal stem cells (MSCs) in gastric cancer (GC), and reveal its underlying mechanisms.
 Methods: Expression of SMARCA5 was quantified in GES-1, AGS and MKN45 cell lines. In MSCs-GC co-cultured cells, cell viability was assessed using cell counting kit-8 (CCK-8) assay, and colony formation were performed to determine cell proliferation. The invasion of the MSCs-GC cells was evaluated by Transwell assay. The levels of vimentin, snail and slug, as well as Wingless-Int (Wnt)/β-catenin related proteins β-catenin, Axin, c-myc, and matrix metalloproteinase (MMP)-7 were determined using immunoblotting. Oxaliplatin was added to GC cells, and the effect of siSMARCA5 on cell sensitivity to oxaliplatin was investigated after transfection with SMARCA5 siRNA into MSCs-GC cells.
 Results: SMARCA5 was highly expressed after co-culture with MSCs (p < 0.001). The MSCs facilitated the proliferation of GC cells, and enhanced cell invasion as well as migration (p < 0.001) compared with untreated cells. MSCs also promoted epithelial–mesenchymal transition (EMT) by increasing production of vimentin, snail, and slug (p < 0.001). The MSCs decreased the sensitivity of GC cells to oxaliplatin, while the knockdown of SMARCA5 reversed the effect (p < 0.001). Furthermore, MSCs up-regulated β-catenin, c-myc, and MMP-7 levels, but downregulated Axin expression compared with untreated cells (p < 0.001). However, siSMARCA5 blocked these processes compared with si-NC group (p < 0.001).
 Conclusion: MSCs facilitate GC cell growth and oxaliplatin resistance by upregulating SMARCA5 and activating Wnt/β-catenin signaling. This may provide an alternative treatment target for patients with GC.

The sub-molecular characterization identification for cervical cancer

BackgroundThe efficacy of therapy in cervical cancer (CESC) is blocked by high molecular heterogeneity. Thus, the sub-molecular characterization remains primarily explored for personalizing the treatment of CESC patients. MethodsDatasets with 741 CESC patients were obtained from TCGA and GEO databases. The NMF algorithm, random forest algorithm, and multivariate Cox analysis were utilized to construct a classifier for defining the sub-molecular characterization. Then, the biological characteristics, genomic variations, prognosis, and immune landscape in molecular subtypes were explored. The significance of classifier genes was validated by quantitative Real-Time PCR, cell transfection, cell colony formation assay, wound healing assay, cell proliferation assay, and Western blot. ResultsThe CESC patients were classified into two subtypes, and the high classifier-score patients with significant differences in ECM-receptor interaction, PI3K-Akt signaling pathway, and MAPK signaling pathway showed a poorer prognosis in OS (p < 0.001), DFI (p = 0.016), PFI (p < 0.001) and DSS (p < 0.001), and with high the M0 Macrophage and resting Mast cells infiltration and low HLA family gene expression. Moreover, the constructed classifier owns a high identified accuracy in the tumor/normal groups (AUC: 0.993), the tumor/CIN1–CIN3 groups (AUC: 0.963), and normal/CIN1–CIN3 groups (AUC: 0.962), and the total prediction performance is better than currently published signatures in CESC (C−index: 0,763). The combined prediction performance further indicated that Nomogram (AUC = 0.837) is superior to the classifier (AUC = 0.835) and Stage (AUC = 0.568), and the C−index of calibration curves is 0.784. The potential biological function of classifier genes indicated that silencing GALNT2 inhibited the cancer cell's proliferation, migration, and colony formation; Conversely, the cancer cell's proliferation, migration, and colony formation were increased after the upregulation of GALNT2. The Epithelial-Mesenchymal Transition Experiment showed that GALNT2 knockdown might reduce the levels of Snail and Vimentin proteins and increase E-cadherin; Conversely, the levels of Snail and Vimentin proteins were increased, E-cadherin was reduced by GALNT2 upregulation. ConclusionThe classifier we constructed may help improve our understanding of subtype characteristics and provide a new strategy for developing CESC therapeutics. Remarkably, GALNT2 may be an option to directly target drivers in CESC cancer therapy.

TMEM211 Promotes Tumor Progression and Metastasis in Colon Cancer.

Colon cancer is the third most important cancer type, leading to a remarkable number of deaths, indicating the necessity of new biomarkers and therapeutic targets for colon cancer patients. Several transmembrane proteins (TMEMs) are associated with tumor progression and cancer malignancy. However, the clinical significance and biological roles of TMEM211 in cancer, especially in colon cancer, are still unknown. In this study, we found that TMEM211 was highly expressed in tumor tissues and the increased TMEM211 was associated with poor prognosis in colon cancer patients from The Cancer Genome Atlas (TCGA) database. We also showed that abilities regarding migration and invasion were reduced in TMEM211-silenced colon cancer cells (HCT116 and DLD-1). Moreover, TMEM211-silenced colon cancer cells showed decreased levels of Twist1, N-cadherin, Snail and Slug but increased levels of E-cadherin. Levels of phosphorylated ERK, AKT and RelA (NF-κB p65) were also decreased in TMEM211-silenced colon cancer cells. Our findings indicate that TMEM211 regulates epithelial-mesenchymal transition for metastasis through coactivating the ERK, AKT and NF-κB signaling pathways, which might provide a potential prognostic biomarker or therapeutic target for colon cancer patients in the future.

Asiatic acid exhibits antimetastatic activity in human prostate cancer cells by modulating the MZF-1/Elk-1/Snail signaling axis

Prostate cancer metastasis is associated with poor prognosis and is difficult to treat clinically. Numerous studies have shown that Asiatic Acid (AA) has antibacterial, anti-inflammatory, and antioxidant effects. However, the effect of AA on prostate cancer metastasis is still unclear. This purpose of this study is to investigate the effect of AA on prostate cancer metastasis and to better understand its molecular mechanisms of action. Our results indicate that AA ≤ 30 μM did not influence cell viability and cell cycle distribution in PC3, 22Rv1 and DU145 cells. AA inhibited the migratory and invasive capabilities of three prostate cancer cells to be due to its effects on Snail, but did not have activity on Slug. We observed that AA inhibited the Myeloid zinc finger 1 (MZF-1) and ETS Like-1 (Elk-1) protein interaction and affected the complex's binding capacity to the Snail promoter region, ultimately blocking Snail transcription activity. Kinase cascade analysis revealed that phosphorylation of MEK3/6 and p38MAPK was inhibited by AA treatment. Moreover, knockdown of p38MAPK enhanced AA-suppressed protein levels of MZF-1, Elk-1, and Snail, suggesting that p38MAPK influences prostate cancer cell metastasis. These results provide promise for AA as a future candidate in the development of drug therapies to prevent or treat prostate cancer metastasis.

ALDH2 promotes cancer stemness and metastasis in colorectal cancer through activating β-catenin signaling.

Colorectal cancer (CRC) is the primary cause of death from gastrointestinal cancers. Aldehyde dehydrogenase 2 (ALDH2), a crucial mitochondrial enzyme for the oxidative pathway of alcohol metabolism, plays a dual role in cancer progression. In some cancers, it is tumor suppressive; in others, it drives cancer progression. However, whether targeting ALDH2 has any therapeutic implications or prognostic value in CRC is still unclear. Here, we investigated the role of ALDH2 in CRC progression by targeting its enzymatic activity rather than gene expression. We found that inhibiting ALDH2 by CVT-10216 and daidzein significantly decrease migration and stemness properties of both DLD-1 and HCT 116 cells, whereas activating ALDH2 by Alda-1 enhances migration rate. Concomitantly, ALDH2 inhibition by both CVT-10216 and daidzein downregulates the mRNA levels of fibronectin, snail, twist, MMP7, CD44, c-Myc, SOX2, and OCT-4, which are oncogenic in the advanced stage of CRC. Furthermore, Gene Set Enrichment Analysis (GSEA) on ALDH2 co-expressed genes from The Cancer Genome Atlas (TCGA) revealed that MYC target gene sets are upregulated. We found that ALDH2 inhibition decreased the nuclear protein levels of pGSK3β serine 9 and c-Myc. This suggests that ALDH2 probably targets β-catenin signaling in CRC cells. Together, our results demonstrate the prognostic value of ALDH2 in CRC as it regulates both CRC stemness and migration. Our findings also propose that the plant-derived isoflavone daidzein could be a potential chemotherapeutic drug targeting ALDH2 in CRC.

Snail inhibits metastasis via regulation of E‑cadherin and is associated with prognosis in colorectal cancer.

The overall survival (OS) rate of patients with colorectal cancer (CRC) remains low due to the lack of clear prognostic markers. Therefore, the identification of valuable prognostic markers is urgently required. Snail and E-Cadherin (E-Cad) are important protein molecules in the EMT process and play a crucial role in tumor invasion and metastasis. The present study investigated the clinical significance of Snail and E-cad expression in CRC. Compared with those in adjacent tissue, the expression levels of Snail and E-cad were significantly increased and decreased, respectively, in CRC. Moreover, low Snail and high E-cad expression were associated with clinicopathological features and longer OS time. Furthermore, Snail and E-cad could predict the prognosis of patients with CRC. Reverse transcription-qPCR, Western blotting, Wound scratch assay, High content cell migration experiment, which showed that low Snail or high E-cad expression inhibited invasion and metastasis of CRC. In conclusion, Snail can promote CRC invasion and metastasis by regulating E-cad. Snail and E-cad expression constitute a novel prognostic marker for CRC, and the present study revealed a greater combined effect of Snail and E-cad as effective prognostic markers in CRC for the first time.

ING5 overexpression upregulates miR-34c-5p/Snail1 to inhibit EMT and invasion of lung cancer cells.

ING5 belongs to the inhibitor of growth (ING) candidate tumor suppressor family, which is involved in multiple cellular functions, such as cell cycle regulation, apoptosis, and chromatin remodelling. Previously, we reported that ING5 overexpression inhibits EMT by regulating EMT-related molecules, including Snail1, at the mRNA and protein levels. However, the mechanisms remain unclear. In the current study, we identify that ING5 overexpression induces the upregulation of miR-34c-5p. The expression levels of both ING5 and miR-34c-5p in NSCLC tissues from the TCGA database are decreased compared with that in adjacent tissues. Higher expression levels of both ING5 and miR-34c-5p predict longer overall survival (OS). Snail1 is the target gene of miR-34c-5p, as predicted by an online database, which is further verified by a dual-luciferase reporter assay. The expression level of Snail1 in NSCLC cells is markedly reduced following miR-34c-5p overexpression, leading to the inactivation of the Snail1 downstream TGF-β/Smad3 signaling pathway. The TGF-β signaling-specific inhibitor LY2157299 reverses the enhanced EMT, proliferation, migration, and invasion abilities induced by the miR-34c-5p inhibitor. Furthermore, tail vein injection of miR-34c-5p agomir inhibits xenografted tumor metastasis. Overall, this study concludes that miR-34c-5p, induced by ING5 overexpression, is a tumor suppressor that targets Snail1 and mediates the inhibitory effects of ING5 on the EMT and invasion of NSCLC cells. These results provide a novel mechanism mediating the antitumor effects of ING5.

Sustained Shugoshin 1 downregulation reduces tumor growth and metastasis in a mouse xenograft tumor model of triple-negative breast cancer

BackgroundTriple-negative breast cancer (TBNC) is an aggressive breast cancer subtype with a poor prognosis. Shugoshin-1 (SGO1) protects chromatids from early separation. Previous studies from our group have demonstrated that transient SGO1 downregulation suppresses early stages of metastasis (the epithelial-to-mesenchymal transition, or EMT, cell invasion, and cell migration) in TNBC cells. Thus, the inhibition of SGO1 activity may represent a potential therapeutic intervention against cancers that progress to metastasis. Therefore, we aimed to investigate the effects of sustained shRNA-mediated SGO1 downregulation on tumor growth and metastasis in TBNC. To that end, female NOD-SCID Gamma (NSG) mice were injected with 2.5 × 106 shRNA Control (n = 10) or shRNA SGO1 (n = 10) MDA-MB-231 cells. After eight weeks, the number of mice with metastasis to the lymph nodes was calculated. Primary and metastatic tumors, as well as lung and liver tissue, were harvested, measured, sectioned, and stained with hematoxylin and eosin (H&E) stain.ResultsTumor growth and metastasis to the lymph nodes and lungs were significantly reduced in the shRNA SGO1-treated mice group, while metastasis to the liver tends to be lower in cells with downregulated SGO1, but it did not reach statistical significance. Furthermore, sustained SGO1 downregulation significantly reduced cell proliferation, cell migration, and invasion which correlated with lower levels of Snail, Slug, MMP2, MMP3, and MMP9.ConclusionThe supression of SGO1 activity in TNBC harboring dysregulated expression of SGO1 may be a potential target for preventing breast cancer growth and metastasis.

Biochemical and histopathological responses of Biomphalaria alexandrina to RIPEX (plant growth regulator)

BackgroundPlant growth regulators are widely used in agriculture for increasing the growth and ripening of plants, but they endanger the aquatic ecosystem. The current study assessed the effect of sublethal exposure to RIPEX 48% EC concentrations (8 and 16 µL/L) on oxidative stress parameters, sex hormones, immune potential enzymes, differential hemocyte counts, and the histopathology of digestive glands and ovotestis in Biomphalaria alexandrina snails.ResultsRIPEX exposure caused an overall increase in Superoxide dismutase and Glutathione-S-Transferase activities in B. alexandrina. However, extreme RIPEX exposure inhibits SOD activity in snails. Malondialdehyde activity showed an increase in B. alexandrina exposed to both concentrations after all exposure periods. RIPEX also caused a significant increase in testosterone in snails exposed to 16 µL/L, it did, however, reduce hormone levels in snails exposed to 8 µL/L at 7 days. Regarding estradiol, there was a significant increase after 3 days of exposure to 16 µl/L and 7 days of exposure to 8 µL/L. RIPEX exposure also increased the activities of Myeloperoxidase and Adenosine deaminase enzymes in the digestive glands of snails. It increased the total hemocyte count of exposed snails as well as the number of granulocytes. Snail digestive glands and ovotestis showed pathological alterations after 7 days of RIPEX exposure.ConclusionsThese findings suggest that RIPEX is toxic to B. alexandrina and that this snail can be used as a bioindicator for environmental contamination with plant growth regulators.

Eriocalyxin B mediates the migration and inflammation of TGF β2 induced human lens epithelial cells by inhibiting JAK/STAT3 pathway

Purpose: To study the role of Eriocalyxin B (EriB) in the migration and inflammation of TGF-β2-induced human lens epithelial cells (hLECs), and to elucidate the molecular mechanisms involved. Methods: The hLECs cultured in vitro were divided into 5 groups, viz, control, TGFβ2, TGFβ2+2 μM EriB, TGFβ2+4 μM EriB, and TGFβ2+8 μM EriB groups. CCK-8, clone formation and Edu labeling assays were performed to assess the effect of EriB on the proliferation of hLECs cells. To determine the role of EriB in cell migration, Transwell and wound healing assays were used. The levels of vimentin, α-SMA, snail, TNF-α，IL-1β，IL-6, P65, p-P65, p-JAK2, JAK2, p-STAT3, STAT3, and β-catenin in hLECs cells were evaluated by enzyme-linked immunosorbent assay (ELISA) and western blot analysis in order ascertain the signaling pathways involved. Results: The rate of cell proliferation significantly decreased in TGFβ2+2μM EriB, TGFβ2+4μM and TGFβ2+8μM groups compared with TGFβ2 group (p &lt; 0.001). In addition, the migration of hLECs cells and epithelial mesenchymal transition were inhibited by EriB in a dose-dependent way (p &lt; 0.001). ELISA results showed that compared to TGFβ2 group, TNF-α, IL-1β and IL-6 levels in EriB group significantly decreased (p &lt; 0.001). The levels of TNF-α, IL-1β, IL-6, p-P65/P65, p-JAK2/JAK2, p-STAT3/STAT3 and metastasis-associated proteins (α-SMA and snail) in hLECs cells were downregulated by EriB (p &lt; 0.001). Furthermore, vimentin level was increased by EriB (p &lt; 0.001). Conclusion: The results show that EriB inhibits the growth, metastasis and inflammation of hLECs cells by inhibiting JAK/STAT3 pathway, thus indicating that this pathway is a potential therapeutic target for treating cataract

Downregulation of desmoglein 2 promotes EMT progression in gallbladder cancer.

To explore the correlation between the expression level of Desmoglein 2 (DSG2) and the epithelial-mesenchymal transition (EMT) progression in gallbladder cancer (GBC). 106 GBC tissue specimens and corresponding clinical information were collected to make a tissue microarray. Immunohistochemical method was used to test the expression level of DSG2 in GBC tissues. DSG2 was knocked down in the GBC cell line GBC-SD to detect the change of its invasion and metastasis ability. Then RT-qPCR and Western Blot were applied on the DSG2-knocked down GBC-SD cells to detect the expression level change of genes associated with EMT. The high expression rate of DSG2 was significantly correlated with the N, M and TNM staging of patients (P<0.05). Survival analysis identified that GBC patients with high DSG2 expression level had significantly better survival (P<0.05). To further investigate the potential mechanism of DSG2 on regulating GBC tumor progression, we used knockdown DSG2 on GBC-SD cell lines. The results showed that GBC-SD cell lines with DSG2 knockdown showed a promotion of cell invasion and metastatic ability. The mRNA levels of EMT-related genes E-Cadherin, Snail, Twist, ZEB1, and β-catenin, which is a key protein in the Wnt signaling pathway, were also significantly altered. Besides, protein levels of E-cadherin and Snail showed consistent results. The downregulation of DSG2 in gallbladder cancer is hypothesized to be associated with the invasion and metastasis progression of gallbladder cancer cells by regulating EMT-related pathways. Its expression level can be a novel biomarker for gallbladder cancer, providing new perspectives for diagnosis and treatment strategies.

MP04-16 EPITHELIAL TO MESENCHYMAL TRANSITION PROMOTES PD-L1 EXPRESSION IN RENAL CELL CARCINOMA

You have accessJournal of UrologyCME1 Apr 2023MP04-16 EPITHELIAL TO MESENCHYMAL TRANSITION PROMOTES PD-L1 EXPRESSION IN RENAL CELL CARCINOMA Allison May, Jonathan Jiang, Tyler Robinson, and Evan Keller Allison MayAllison May More articles by this author , Jonathan JiangJonathan Jiang More articles by this author , Tyler RobinsonTyler Robinson More articles by this author , and Evan KellerEvan Keller More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003215.16AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Sarcomatoid dedifferentiation in renal cell carcinoma (RCC) is thought to occur when the parental cell type, such as clear cell RCC (ccRCC), undergoes an epithelial mesenchymal transition (EMT). The mesenchymal sarcomatoid cells have increased ability to invade and metastasize, and thus sarcomatoid RCC (sRCC) has a very poor prognosis. Interestingly however, there is encouraging evidence that sRCC responds at higher rates to PD-1/PD-L1 directed immunotherapy than ccRCC. We aim to test the hypothesis that EMT directly upregulates PD-L1 expression in sRCC and that PD-L1 may have positive feedback on induction of EMT. METHODS: Baseline expression of EMT markers, EMT transcription factors and PD-L1 were measured using Western Blot and qPCR in two clear cell RCC lines (Caki-1 and 786-O) and five sarcomatoid RCC lines (RCJ-41T1, RCJ-41T2, RCJ-41M, UOK-276, UOK-127). EMT was induced in the cells through treatment with multiple agents including TGFB, IFNg, HGF, SPP1, and CoCl2 and by over expression of Snail or Zeb1. Effect of EMT on PD-L1 expression was measured. To evaluate the role of PD-L1 on EMT, PD-L1 expression was decreased using siRNA and its effect on EMT related protein expression was measured using Western Blot. RESULTS: At baseline, sRCC cell lines generally had higher expression of EMT related markers and transcription factors than ccRCC lines. Baseline PD-L1 expression positively correlated with increasing EMT state in all cell lines. Treatment of cells with known EMT inducers, TGFB, IFNg, and hypoxia led to increased levels of mesenchymal marker N-cadherin, decreased epithelial marker E-cadherin, and increased levels of EMT transcription factors Zeb1, Snail, and Slug. Treatment of cells with HGF and SPP1, two molecules associated with sRCC based on our prior work, also induced EMT. Finally, over expression of known EMT transcription factors Snail and Zeb1 led to EMT as well. All methods of EMT induction concomitantly increased PD-L1 expression. Knockdown of PD-L1 was tested in UOK-276 and 786-O cells and led to decreased expression levels of EMT transcription factors and increased epithelial markers. CONCLUSIONS: These data support the theory that sRCC arises from EMT and that EMT directly leads to upregulation of PD-L1. Furthermore, these results suggest that PD-L1 maintains the mesenchymal state. These findings may help explain the enhanced response to PD-1/PD-L1 directed therapy in sRCC and suggest that markers of EMT state could represent a biomarker for immunotherapy response in RCC. Source of Funding: Clarke Family Fellowship for Kidney Cancer Research © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e39 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Allison May More articles by this author Jonathan Jiang More articles by this author Tyler Robinson More articles by this author Evan Keller More articles by this author Expand All Advertisement PDF downloadLoading ...

OTX1 silencing suppresses ovarian cancer progression through inhibiting the JAK/STAT signaling

BackgroundThe aim of our study was to investigate the roles and the underlying mechanisms of orthodenticle homolog 1 (OTX1) in ovarian cancer. MethodsOTX1 expression was obtained from TCGA database. OTX1 expression in ovarian cancer cells was detected using qRT-PCR and western blot assay. The cell viability and proliferation was detected by CCK-8 and EdU assays. Cell invasion and migration were detected by transwell assay. Flow cytometry was utilized to determine cell apoptosis and cycle. In addition, western blot assay was used to detect the expression of cell cycle related protein (Cyclin D1 and p21), epithelial-mesenchymal transition (EMT) related protein (E-cadherin, N-cadherin, Vimentin, and Snail), apoptosis related protein (Bcl-2, Bax, and cleaved caspase-3), and JAK/STAT pathway related protein (p-JAK2, JAK2, STAT3, and p-STAT3). ResultsOTX1 was highly expressed in ovarian cancer tissues and cells. OTX1 silencing blocked the cell cycle and repressed cell viability, proliferation, invasion, and migration, while OTX1 silencing facilitated the apoptosis of OVCAR3 and Caov3 cells. OTX1 silencing increased the protein levels of p21, E-cadherin, Bax, and cleaved caspase-3, while the protein levels of Cyclin D1, Bcl-2, N-cadherin, Vimentin, and Snail were decreased by OTX1 silencing. Furthermore, OTX1 silencing suppressed the protein levels of p-JAK2/JAK2 and p-STAT3/STAT3 in OVCAR3 and Caov3 cells. Moreover, overexpression of OTX1 promoted cell proliferation and invasion and inhibited apoptosis in Caov3 cells, but AG490 (an inhibitor of JAK/STAT pathway) reversed the influences on cell biological behavior induced by overexpression of OTX1. ConclusionsOTX1 silencing repressed ovarian cancer cell proliferation, invasion, and migration and induced cell apoptosis, which might be involved in JAK/STAT signaling pathway. OTX1 may be considered as a novel therapeutic target for ovarian cancer.

PLAGL2 promotes Snail expression and gastric cancer progression via UCA1/miR-145-5p/YTHDF1 axis.

Although great progress has made in gastric cancer (GC) in the past years, the overall 5-year survival rate remains to be low for advanced GC patients. A recent study showed that PLAGL2 was increased in GC and enhanced the proliferation and metastasis of GC. Nevertheless, the underlying mechanism still needs to be investigated. Gene and protein expressions were assessed using RT-qPCR and western blot. The migration, proliferation and invasion of GC cells were examined using scratch assay, CCK-8 assay and Transwell assay, respectively. ChIP-PCR, dual-luciferase assay, RIP-qPCR and CoiP were utilized to confirm the interaction among PLAGL2, UCA1, miR-145-5p and YTHDF1 as well as METTL3, YTHDF1 and eEF-2. A mouse xenograft model was used utilized to further confirm the regulatory network. PLAGL2 bound to the upstream promoter of UCA1, which regulated YTHDF1 by sponging miR-145-5p. METTL3 can mediate the m6A modification level of Snail. YTHDF1 recognized m6A-modified Snail by interacting with eEF-2 and thus promoted Snail expression, which eventually induced epithelial-mesenchymal transition (EMT) in GC cells and metastasis of GC. Overall, our study demonstrates that PLAGL2 enhances Snail expression and GC progression via the UCA1/miR-145-5p/YTHDF1 axis, suggesting that PLAGL2 may become a therapeutic target for GC treatment.

Bufalin-loaded vitamin E succinate-grafted chitosan oligosaccharide/RGD-conjugated TPGS mixed micelles inhibit intraperitoneal metastasis of ovarian cancer

BackgroundIntraperitoneal metastasis is one of the major causes of the high mortality rate of ovarian cancer. Bufalin (BU) is an effective component of the traditional Chinese medicine Chansu that exerts antitumor effects, including metastasis inhibition. In our previous studies, we found that BU inhibited the migration and invasion of ovarian cancer cells. However, the application of BU is limited due to its insolubility, toxicity and imprecise targeting. The aim of this study was to use vitamin E succinate (VES)-grafted chitosan oligosaccharide (CSO)/arginine-glycine-aspartic acid peptide (RGD)-conjugated d-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS) mixed micelles (VeC/T-RGD MMs) to deliver BU to ovarian cancer cells to inhibit intraperitoneal metastasis. Moreover, the toxicity of BU was reduced by coating it with the mixed micelles to increase its biocompatibility for practical applications.ResultsThe BU-loaded VeC/T-RGD MMs (BU@MMs) had an average diameter of 161 ± 1.4 nm, a zeta potential of 4.49 ± 1.54 mV and a loading efficiency of 2.54%. The results showed that these micelles inhibited cell proliferation, induced apoptosis, and reduced the migration and invasion of A2780 and SKOV3 cells. Further studies indicated that BU@MMs enhanced the levels of e-cadherin and decreased the expression levels of N-cadherin, vimentin and Snail in vitro. In addition, the mixed micelles effectively enhanced the anticancer effect and inhibited intraperitoneal metastasis in intraperitoneal metastatic models. The BU@MMs exhibited fewer toxic side effects than BU, indicating better biocompatibility and biosafety for in vivo applications.ConclusionsOur studies show that BU@MMs are a potential multifunctional nano-drug delivery system that can effectively inhibit the intraperitoneal metastasis of ovarian cancer.

Tumor malignancy by genetic transfer between cells forming cell-in-cell structures

Cell-in-cell structures (CICs) refer to a type of unique structure with one or more cells within another one, whose biological outcomes are poorly understood. The present study aims to investigate the effects of CICs formation on tumor progression. Using genetically marked hepatocellular cancer cell lines, we explored the possibility that tumor cells might acquire genetic information and malignant phenotypes from parental cells undergoing CICs formation. The present study showed that the derivatives, isolated from CICs formed between two subpopulations by flow cytometry sorting, were found to inherit aggressive features from the parental cells, manifested with increased abilities in both proliferation and invasiveness. Consistently, the CICs clones expressed a lower level of E-cadherin and a higher level of Vimentin, ZEB-1, Fibronectin, MMP9, MMP2 and Snail as compared with the parental cells, indicating epithelial-mesenchymal transition. Remarkably, the new derivatives exhibited significantly enhanced tumorigenicity in the xenograft mouse models. Moreover, whole exome sequencing analysis identified a group of potential genes which were involved in CIC-mediated genetic transfer. These results are consistent with a role of genetic transfer by CICs formation in genomic instability and malignancy of tumor cells, which suggest that the formation of CICs may promote genetic transfer and gain of malignancy during tumor progression.

The significance of spread through air spaces in the prognostic assessment model of stage I lung adenocarcinoma and the exploration of its invasion mechanism.

Spread through air spaces (STAS) is a crucial invasive mode of lung cancer and has been shown to be associated with early recurrence and metastasis. We aimed to develop a prognostic risk assessment model for stage I lung adenocarcinoma based on STAS and other pathological features and to explore the potential relationship between CXCL-8, Smad2, Snail, and STAS. 312 patients who underwent surgery at Harbin Medical University Cancer Hospital with pathologically diagnosed stage I lung adenocarcinoma were reviewed in the study. STAS and other pathological features were identified by H&E staining, and a prognostic risk assessment model was established. The expression levels of CXCL8, Smad2, and Snail were determined by immunohistochemistry. The nomogram was established based on age, smoking history, STAS, tumor lymphocyte infiltration, tissue subtype, nuclear grade, and tumor size. The C-index for DFS was (training set 0.84 vs validation set 0.77) and for OS was (training set 0.83 vs validation set 0.78). Decision curve analysis showed that the model constructed has a better net benefit than traditional reporting. The prognostic risk score validated the risk stratification value for stage I lung adenocarcinoma. STAS was an important prognostic factor associated with stronger invasiveness and higher expression of CXCL8, Smad2, and Snail. CXCL8 was associated with poorer DFS and OS. We developed and validated a survival risk assessment model and the prognostic risk score formula for stage I lung adenocarcinoma. Additionally, we found that CXCL8 could be used as a potential biomarker for STAS and poor prognosis, and its mechanism may be related to EMT.

RUTİNİN EPİTELYAL VE MEZENKİMAL GEÇİŞ ÜZERİNE OLAN ETKİSİNİN PROSTAT KANSERİ HÜCRELERİNDE BELİRLENMESİ

Objective: The difficulties experienced in the treatment of prostate cancer and the excess of side effects due to chemotherapy have brought the search for alternative treatment strategies. In recent studies, it is known that Rutin (RUT) has an anti-cancer effect on cancer cells. Our study aimed to determine the effects of RUT on epithelial-mesenchymal transition (EMT) in prostate cancer cells, for the first time.&#x0D; Methods: The anticancer activity of RUT in prostate cancer cells (PC-3) was determined by WST-1, Annexin V ELISA, DAPI and Acridine Orange staining, and the anti-cancer and anti-metastatic properties of RUT were evaluated with the Scratch Assay test. The mRNA expression level of Bax, Bcl-2, Snail, Twist, Vimentin and E-cadherin genes was determined by RT-PCR.&#x0D; Results: PC-3 cells were treated with RUT (500, 750, 1000, 1500 µM) for 24 and 48 hours. The viability rates decreased with increasing RUT concentration depending on dose and time (p