A neutrophil-biomimic platform for eradicating metastatic breast cancer stem-like cells by redox microenvironment modulation and hypoxia-triggered differentiation therapy

Metastasis accounts for 90% of breast cancer deaths, where the lethality could be attributed to the poor drug accumulation at the metastatic loci. The tolerance to chemotherapy induced by breast cancer stem cells (BCSCs) and their particular redox microenvironment further aggravate the therapeutic dilemma. To be specific, therapy-resistant BCSCs can differentiate into heterogeneous tumor cells constantly, and simultaneously dynamic maintenance of redox homeostasis promote tumor cells to retro-differentiate into stem-like state in response to cytotoxic chemotherapy. Herein, we develop a specifically-designed biomimic platform employing neutrophil membrane as shell to inherit a neutrophil-like tumor-targeting capability, and anchored chemotherapeutic and BCSCs-differentiating reagents with nitroimidazole (NI) to yield two hypoxia-responsive prodrugs, which could be encapsulated into a polymeric nitroimidazole core. The platform can actively target the lung metastasis sites of triple negative breast cancer (TNBC), and release the escorted drugs upon being triggered by the hypoxia microenvironment. During the responsiveness, the differentiating agent could promote transferring BCSCs into non-BCSCs, and simultaneously the nitroimidazole moieties conjugated on the polymer and prodrugs could modulate the tumor microenvironment by depleting nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) and amplifying intracellular oxidative stress to prevent tumor cells retro-differentiation into BCSCs. In combination, the BCSCs differentiation and tumor microenvironment modulation synergistically could enhance the chemotherapeutic cytotoxicity, and remarkably suppress tumor growth and lung metastasis. Hopefully, this work can provide a new insight in to comprehensively treat TNBC and lung metastasis using a versatile platform.

Regulation of tumor suppressor microRNA-218 on malignant progression and cancer stem cell properties of prostate cancer / 现代泌尿外科杂志

【Objective】 To investigate the role of microRNA-218 (miR-218) in regulating prostate cancer (PCa) cell stemness and epithelial-mesenchymal transition (EMT). 【Methods】 PCa cell line stably overexpressing miR-218 was constructed with lentivirus transfection. The expression of miR-218 was detected with real-time fluorescence quantitative polymerase chain reaction (q-PCR). The migration ability was detected with Transwell assay. The expression of EMT related proteins were detected with Western blot. The properties of cells were determined with colony formation and tumor sphere formation assays. 【Results】 The results of q-PCR showed that the mRNA level of miR-218 was significantly lower in PCa cell lines LNCaP and C4-2 than in BPH-1. Transwell assay showed that miR-218 inhibited the migration of PCa cells. Western blot showed that the expression of EMT related proteins were inhibited by miR-218. Colony formation and tumor sphere formation assays showed that overexpression of miR-218 significantly inhibited the properties of cells. 【Conclusion】 The expression of miR-218 is downregulated in PCa cell lines. miR-28 can inhibit cell migration, EMT and cancer stem cell properties.

Casticin Attenuates Stemness in Cervical Cancer Stem-Like Cells by Regulating Activity and Expression of DNMT1 / 中国结合医学杂志

OBJECTIVE@#To explore whether casticin (CAS) suppresses stemness in cancer stem-like cells (CSLCs) obtained from human cervical cancer (CCSLCs) and the underlying mechanism.@*METHODS@#Spheres from HeLa and CaSki cells were used as CCSLCs. DNA methyltransferase 1 (DNMT1) activity and mRNA levels, self-renewal capability (Nanog and Sox2), and cancer stem cell markers (CD133 and CD44), were detected by a colorimetric DNMT activity/inhibition assay kit, quantitative real-time reverse transcription-polymerase chain reaction, sphere and colony formation assays, and immunoblot, respectively. Knockdown and overexpression of DNMT1 by transfection with shRNA and cDNA, respectively, were performed to explore the mechanism for action of CAS (0, 10, 30, and 100 nmol/L).@*RESULTS@#DNMT1 activity was increased in CCSLCs compared with HeLa and CaSki cells (P<0.05). In addition, HeLa-derived CCSLCs transfected with DNMT1 shRNA showed reduced sphere and colony formation abilities, and lower CD133, CD44, Nanog and Sox2 protein expressions (P<0.05). Conversely, overexpression of DNMT1 in HeLa cells exhibited the oppositive effects. Furthermore, CAS significantly reduced DNMT1 activity and transcription levels as well as stemness in HeLa-derived CCSLCs (P<0.05). Interestingly, DNMT1 knockdown enhanced the inhibitory effect of CAS on stemness. As expected, DNMT1 overexpression reversed the inhibitory effect of CAS on stemness in HeLa cells.@*CONCLUSION@#CAS effectively inhibits stemness in CCSLCs through suppression of DNMT1 activation, suggesting that CAS acts as a promising preventive and therapeutic candidate in cervical cancer.

The expression and function of PD-L1 in CD133(+) human liver cancer stem-like cells / 中华肿瘤杂志

Objective: To investigate the expression of programmed death protein-ligand 1 (PD-L1) in liver cancer stem-like cells (LCSLC) and its effect on the characteristics of tumor stem cells and tumor biological function, to explore the upstream signaling pathway regulating PD-L1 expression in LCSLC and the downstream molecular mechanism of PD-L1 regulating stem cell characteristics, also tumor biological functions. Methods: HepG2 was cultured by sphere-formating method to obtain LCSLC. The expressions of CD133 and other stemness markers were detected by flow cytometry, western blot and real-time quantitative polymerase chain reaction (RT-qPCR) were used to detect the expressions of stemness markers and PD-L1. The biological functions of the LCSLC were tested by cell function assays, to confirm that the LCSLC has the characteristics of tumor stem cells. LCSLC was treated with cell signaling pathway inhibitors to identify relevant upstream signaling pathways mediating PD-L1 expression changes. The expression of PD-L1 in LCSLC was down regulated by small interfering RNA (siRNA), the expression of stem cell markers, tumor biological functions of LCSLC, and the changes of cell signaling pathways were detected. Results: Compared with HepG2 cells, the expression rate of CD133 in LCSLC was upregulated [(92.78±6.91)% and (1.40±1.77)%, P<0.001], the expressions of CD133, Nanog, Oct4A and Snail in LCSLC were also higher than those in HepG2 cells (P<0.05), the number of sphere-formating cells increased on day 7 [(395.30±54.05) and (124.70±19.30), P=0.001], cell migration rate increased [(35.41±6.78)% and (10.89±4.34)%, P=0.006], the number of transmembrane cells increased [(75.77±10.85) and (20.00±7.94), P=0.002], the number of cloned cells increased [(120.00±29.51) and (62.67±16.77), P=0.043]. Cell cycle experiments showed that LCSLC had significantly more cells in the G(0)/G(1) phase than those in HepG2 [(54.89±3.27) and (32.36±1.50), P<0.001]. The tumor formation experiment of mice showed that the weight of transplanted tumor in LCSLC group was (1.32±0.17)g, the volume is (1 779.0±200.2) mm(3), were higher than those of HepG2 cell [(0.31±0.06)g and (645.6±154.9)mm(3), P<0.001]. The expression level of PD-L1 protein in LCSLC was 1.88±0.52 and mRNA expression level was 2.53±0.62, both of which were higher than those of HepG2 cells (P<0.05). The expression levels of phosphorylation signal transduction and transcription activation factor 3 (p-STAT3) and p-Akt in LCSLC were higher than those in HepG2 cells (P<0.05). After the expression of p-STAT3 and p-Akt was down-regulated by inhibitor treatment, the expression of PD-L1 was also down-regulated (P<0.05). In contrast, the expression level of phosphorylated extracellular signal-regulated protein kinase 1/2 (p-ERK1/2) in LCSLC was lower than that in HepG2 cells (P<0.01), there was no significant change in PD-L1 expression after down-regulated by inhibitor treatment (P>0.05). After the expression of PD-L1 was knockdown by siRNA, the expressions of CD133, Nanog, Oct4A and Snail in LCSLC were decreased compared with those of siRNA-negative control (NC) group (P<0.05). The number of sphere-formating cells decreased [(45.33±12.01) and (282.00±29.21), P<0.001], the cell migration rate was lower than that in siRNA-NC group [(20.86±2.74)% and (46.73±15.43)%, P=0.046], the number of transmembrane cells decreased [(39.67±1.53) and (102.70±11.59), P=0.001], the number of cloned cells decreased [(57.67±14.57) and (120.70±15.04), P=0.007], the number of cells in G(0)/G(1) phase decreased [(37.68±2.51) and (57.27±0.92), P<0.001], the number of cells in S phase was more than that in siRNA-NC group [(30.78±0.52) and (15.52±0.83), P<0.001]. Tumor formation in mice showed that the tumor weight of shRNA-PD-L1 group was (0.47±0.12)g, the volume is (761.3±221.4)mm(3), were lower than those of shRNA-NC group [(1.57±0.45)g and (1 829.0±218.3)mm(3), P<0.001]. Meanwhile, the expression levels of p-STAT3 and p-Akt in siRNA-PD-L1 group were decreased (P<0.05), while the expression levels of p-ERK1/2 and β-catenin did not change significantly (P>0.05). Conclusion: Elevated PD-L1 expression in CD133(+) LCSLC is crucial to maintain stemness and promotes the tumor biological function of LCSLC.

Advances in understanding leukemia bone marrow microenvironments / 药学学报

The bone marrow microenvironment, also known as the bone marrow niche, plays a critical role in maintaining the functions of hematopoietic stem cells. Under physiological conditions, various bone marrow cells regulate each other to sustain hematopoietic homeostasis. However, bone marrow cells gain abnormal function under pathological conditions to cause and promote the occurrence of leukemia and induce drug resistance. Recent findings indicate that abnormal proliferation and differentiation are not the sole reason to cause leukemia. Different types of bone marrow cells also induce intercellular adhesion, abnormally secrete cytokines and chemokines, accelerating leukemia's progress. This article reviews the multiple signaling pathways that regulate the formation and progress of leukemia bone marrow niche, such as C-X-C motif chemokine ligand 12/C-X-C motif chemokine receptor 4 signaling pathway, et al. It emphasizes that targeting leukemia bone marrow niche is a vital strategy for improving the leukemia treatment.

Expression and regulation of keratin 6B in CD44+ bladder cancer stem cells / 中国组织工程研究

BACKGROUND: Bladder cancer stem cells could promote the recurrence and drug resistance of bladder cancer. Numerous studies have shown that keratin 6B (KRT6B) is involved in the production and progression of tumors, and is closely related to the prognosis of tumors. OBJECTIVE: To observe the expression of keratin 6B in CD44+ bladder cancer stem cells and to show the influence of keratin 6B on proliferation, migration, and self-renewal of bladder cancer stem cells, and to further explore the effect of keratin 6B expression on the prognosis of bladder cancer patients. METHODS: (1) CD44+ 5637 bladder cancer stem cells were isolated by magnetic active cell sorting. Cancer stem cell-related gene expression of SOX2, OCT4, and NANOG was detected via real-time polymerase chain reaction. The spheroid formation assay was used to detect the ability of self-renewal of cancer stem cells in CD44+ cells. Keratin 6B expression was detected in CD44+ bladder cancer stem cells by real-time polymerase chain reaction. (2) The CD44+5637 bladder cancer stem cells were divided into two groups. In the keratin 6B siRNA group, keratin 6B small interfering RNA was transfected into CD44+ bladder cancer stem cells. Untransfected CD44+ bladder cancer stem cells were used as the black control group. Cells were collected at 2 days post-transfection. The proliferation, migration, and self-renewal capacity of keratin 6B siRNA CD44+ bladder cancer stem cells were detected by the colony and wound healing assay and spheroid formation respectively. (3) Totally 24 bladder cancer tissues were used by immunohistochemistry to analyze the expression of CD44v6 and keratin 6B. (4) ONCOMINE database was used to analyze the effect of keratin 6B expression on the overall survival of bladder cancer. RESULTS AND CONCLUSION: (1) Cancer stem cell-related genes (SOX2, OCT4, NANOG) and keratin 6B expression was higher in CD44+ cells isolated by magnetic active cell sorting compared with CD44- cells (P < 0.05). Cell proliferation, migration, and in vitro spheroid formation were significantly increased (P < 0.05). Keratin 6B small interfering RNA down-regulated the expression of keratin 6B in CD44+ bladder cancer stem cells (P < 0.05). (2) Compared with the blank control group, the proliferation and migration of CD44+ bladder cancer stem cells after transfection of keratin 6B small interfering RNA (P < 0.05), and the number of tumorsphere significantly diminished (P < 0.05); the expression of Notch1 and Hes1 mRNA increased (P < 0.05). (3) Keratin 6B and CD44v6 were significantly different in bladder cancer tissue (P=0.006). The overall survival rate of bladder cancer patients with high expression of keratin 6B was lower than that of patients with low expression of keratin 6B. (4) The results showed that keratin 6B was highly expressed in CD44+ bladder cancer stem cells, and could promote the proliferation, migration, and self-renewal capacity of bladder cancer stem cells. The high expression of keratin 6B contributes to improving the survival of bladder cancer patients.

Capilliposide from Lysimachia capillipes promotes terminal differentiations and reverses paclitaxel resistance in A2780T cells of human ovarian cancer by regulating Fos/Jun pathway / 中草药·英文版

Objective: To investigate the potential effect of Lysimachia capillipes capilliposide (LCC) on the chemo sensitivity and the stemness of human ovarian cancer cells. Methods: Cell Counting Kit-8 (CCK8) was used to measure the IC

Interaction and Targeting Strategies of Colorectal Cancer Stem Cells and Tumor Microenvironment / 肿瘤防治研究

Colorectal cancer stem cells are undifferentiated tumorigenic cells with malignant phenotypic characteristics in colorectal cancer and considered as the main cause of tumor recurrence, drug resistance, and metastasis through self-renewal and differentiated cloning. Colorectal cancer stem cells and various components in the tumor microenvironment are interdependent and influence one another. The relationship between tumor stem cells and other components in the tumor microenvironment may play an important role in the treatment of colorectal cancer. This review summarizes this relationship.

The origin and function of thyroid cancer stem cells: A brief introduction / 中华内分泌代谢杂志

Thyroid cancer is highly heterogeneous and has three main types of histopathology: low-malignant papillary thyroid cancer, follicular cancer, and high-malignant undifferentiated cancer. It has recently been hypothesized that thyroid cancer stem cell (TCSC) plays a major role in both cancer initiation and metastasis, but the origin of TCSC and underlying mechanism in the development of thyroid cancer are still unclear. In this article, we introduced the origin theory of thyroid cancer from TCSC and the relation of TCSC to immune system and tumor microenvironment as well as TCSC-based treatment strategy. This will provide a new direction for the treatment of thyroid cancer, so as to improve the prognosis of patients with refractory thyroid cancer.

Cancer stem cell-targeted chimeric antigen receptor (CAR)-T cell therapy: Challenges and prospects

Cancer stem cells (CSCs) with their self-renewal ability are accepted as cells which initiate tumors. CSCs are regarded as interesting targets for novel anticancer therapeutic agents because of their association with tumor recurrence and resistance to conventional therapies, including radiotherapy and chemotherapy. Chimeric antigen receptor (CAR)-T cells are engineered T cells which express an artificial receptor specific for tumor associated antigens (TAAs) by which they accurately target and kill cancer cells. In recent years, CAR-T cell therapy has shown more efficiency in cancer treatment, particularly regarding blood cancers. The expression of specific markers such as TAAs on CSCs in varied cancer types makes them as potent tools for CAR-T cell therapy. Here we review the CSC markers that have been previously targeted with CAR-T cells, as well as the CSC markers that may be used as possible targets for CAR-T cell therapy in the future. Furthermore, we will detail the most important obstacles against CAR-T cell therapy and suggest solutions.

Anti-ENO1 antibody combined with metformin reverses the resistance of human non-small cell lung cancer A549 cells to cetuximab by targeting cancer stem cells / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To explore the effect of anti-ENO1 (enolase 1) antibody and metformin (MET) treatment on the proliferation, migration, invasion and stemness of cetuximab (CTX) -resistant non-small cell lung cancer (NSCLC) cells through targeting cancer stem cells and the possible mechanism. Methods: 10 mmol/L MET combined with 40 μg/ml anti-ENO1 antibody was used to treat CTX(35 µg/ml)-resistant NSCLC A549 cells for 4 d, and the effects of combined treatment on A549 cells were detected with proliferation experiment, colony formation assay, migration and invasion experiments and methylcellulose ball formation experiment. In the meanwhile, FCM was used to detect the effects of CTX, MET and anti-ENO1 antibody single-drug treatment as well as the three-drug combination treatment on ALDH+ and CD44+ lung cancer stem cell subsets. Results: CTX combined with MET and anti-ENO1 antibody treatment significantly inhibited the proliferation, migration, invasion and self-renewal capacity of A549 cells. FCM analysis found that MET could significantly inhibit ALDH+ stem cell subpopulations, while anti-ENO1 antibody could significantly inhibit CD44+ stem cell subpopulations, and the three-drug combination treatment could simultaneously suppress ALDH+ and CD44+ stem cell subpopulations. Conclusion: MET and anti-ENO1 antibody respectively target ALDH+ and CD44+ cancer stem cell subsets, and the combined treatment of MET and anti-ENO1 antibody can effectively reverse the resistance of A549 cells to CTX, and thereby more effectively inhibiting stemness, proliferation, metastasis of A549 cells and tumor recurrence.

The upregulation mechanism of OCT4 signaling by ID1 in colorectal cancer / 药学学报

Inhibitor of DNA binding 1 (ID1) has an aberrantly high expression in multiple cancer tissues, including colon cancer, lung cancer, breast cancer, and so on, which is closely related to cancer aggressiveness and poor clinical outcomes in cancer patients. It has been reported that ID1 maintains colorectal cancer cells (CRCs) stemness traits and contributes to the CRC drug resistance. While, the biological molecular mechanisms have not been fully elucidated. In this research, we found that ID1 upregulates octamer binding transcription factor (OCT4) protein level as well as OCT4 signaling pathway via Western blot, gene set enrichment analysis (GSEA), dual-luciferase reporter assay, and real-time PCR. Through the in vitro sphere formation assay, we found that overexpression of OCT4 reverses the inhibitory effect of knocking down ID1 on CRC sphere formation ability. With the help of JASPAR and GEPIA database, we predicted a novel transcriptional repressor—forkhead box D3 (FOXD3) of OCT4. Finally, by using co-immunoprecipitation (Co-IP), confocal and real-time PCR, we demonstrated that ID1 interacts with FOXD3 to inhibit its transcriptional repression activity and therefore to upregulate OCT4 transcription and OCT4 signaling pathway. In conclusion, this study provides a new theoretical basis for the regulation mechanism of colon cancer stem cells, and the newly found protein-protein interaction of ID1-FOXD3 provides a potential drug target for the therapy of CRC.

Dysregulation of MAD2L1/CAMK2A/PTTG1 Gene Cluster Maintains the Stemness Characteristics of Uterine Corpus Endometrial Carcinoma / 中国医学科学院学报

Objective To study the stemness characteristics of uterine corpus endometrial carcinoma(UCEC)and its potential regulatory mechanism.Methods Transcriptome sequencing data of UCEC was obtained from The Cancer Genome Atlas.Gene expression profile was normalized by edgeR package in R3.5.1.A one-class logistic regression machine learning algorithm was employed to calculated the mRNA stemness index(mRNAsi)of each UCEC sample.Then,the prognostic significance of mRNAsi and candidate genes was evaluated by survminer and survival packages.The high-frequency sub-pathways mining approach(HiFreSP)was used to identify the prognosis-related sub-pathways enriched with differentially expressed genes(DEGs).Subsequently,a gene co-expression network was constructed using WGCNA package,and the key gene modules were analyzed.The clusterProfiler package was adopted to the function annotation of the modules highly correlated with mRNAsi.Finally,the Human Protein Atlas(HPA)was retrieved for immunohistochemical validation.Results The mRNAsi of UCEC samples was significantly higher than that of normal tissues(

Putative stemness markers octamer-binding transcription factor 4, sex-determining region Y-box 2, and NANOG in non-small cell lung carcinoma: A clinicopathological association

Background: The promising improvement in the clinical outcome of lung cancer can be possibly achieved by identification of the molecular events that underlie its pathogenesis. Cancer stem cell (CSC) being one of the subsets of tumor majorly participates in drug resistance and treatment failure because of the moderate cell cycle, lower proliferation, and increased expression of DNA repair and anti-apoptosis genes. Although many putative CSC markers exist, a precise characterization for non-small cell lung cancer is of utmost importance due to increased mortality rate and lack of targeted therapies. Hence, the article focuses on the expression of stemness-associated markers, namely octamer-binding transcription factor 4 (OCT4), NANOG, and sex-determining region Y-box 2 (SOX2) in non-small cell lung cancer (NSCLC) patients. Methods: The expression of OCT4, NANOG, and SOX2 were evaluated in 32 histopathologically confirmed NSCLC tissues using real-time polymerase chain reaction. The obtained expression was correlated with clinical and pathological manifestations using the statistical test such as Student's t-test and Pearson correlation in varied statistical software. Results: Results showed a significantly higher expression of OCT4 and NANOG compared to SOX2 in the tumor tissues. When the expression of these markers was correlated with the clinical parameters, higher expression was seen in males, patients with age above 60 years, and in adenocarcinoma subtype. In correlation with the habit, higher expression of OCT4 and SOX2 was observed in habituated patients. Expression of NANOG and OCT4 was higher even in patients with poor differentiation. Conclusion: The expression and prognostic significance of CSC markers obviously vary depending on histological NSCLC subtype. Importantly, our findings suggest that OCT4, SOX2, and NANOG network together may be promising for ongoing targeted therapies in specific NSCLC subgroups

Monoclonal antibody 18H12 suppresses the self-renewal and invasion of PAMC-82 gastric cancer stem cells / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To investigate the effect of 18H12, a functional monoclonal antibody that can target gastric cancer stem cells, on the self-renewal and invasion ability of gastric cancer cells. Methods: The gastric cancer cell line PAMC-82 was used as cell model, the expression of ENO1 (enolase-1) on the membrane surface of its parental cells and enriched stem cells by sphere culture was detected by Flow cytometry. Flow cytometry was used to separate ENO1+ cells and ENO1- cells to detect their self-renewal ability and invasion ability. With the commercial ENO1 antigen and antibody as the samples, CoIP (co-immunoprecipitation) was used to verify whether 18H12 antibody targeting ENO1 could able to accurately recognize ENO1. After being treated with 18H12 for 12 h, 24 h and 48 h, the selfrenewal and invasion ability of PAMC-82 cells were detected by methylcellulose pelletization experiment and Transwell chamber assay, respectively. Results: Flow cytometry showed that the expression of ENO1 on the membrane surface of PAMC-82 sphere cells was significantly higher than that of its parental cells (P<0.01), so ENO1 could be a potential target for targeting gastric cancer stem cells. The self-renewal ability and invasion ability of the sorted ENO1+ cells were significantly stronger than those of the ENO1- cells and the parental cells (P<0.05 or P<0.01). 18H12 antibody could accurately recognize ENO1, which was consistent with the commercial antibody recognition band. 18H12 could significantly inhibit self-renewal ability and invasion ability of PAMC-82 cells (P<0.01). Conclusion: Monoclonal antibody 18H12 can significantly inhibit the self-renewal and invasion of gastric cancer stem cells and is expected to be a candidate antibody drug targeting gastric cancer stem cells.

The role of CD133+ stem cells in radiotherapy resistance of pancreatic cancer / 西安交通大学学报(医学版)

Objective: To study the role of pancreatic cancer stem cells in radiotherapy resistance of pancreatic cancer. Methods: CD133+ pancreatic cancer stem cells were obtained by using magnetic cell sorting. MTT and flow cytometry were used to observe the proliferation activity and apoptosis of different cell subsets of pancreatic cancer after radiation treatment. Then flow cytometry, Western blot and qRT-PCR were used to observe the expression changes of CD133 in pancreatic cancer cells after radiation treatment. Finally, the cell cycle distribution of different subgroup cells was observed by flow cytometry. Results: Compared with unselected cells and CD133- cells, CD133+ stem cells had the highest survival rate and the lowest apoptosis rate after radiation treatment. The proportion of CD133+ stem cells increased and the expressions of CD133 protein and RNA increased in pancreatic cancer cells. CD133+ stem cells had the highest ratio of G0/G1 phase compared with unsorted cells and CD133- cells, and the difference was statistically significant. Conclusion: CD133+ pancreatic cancer stem cells are one of the important causes of radiotherapy resistance in pancreatic cancer cells, and the increased proportion of G0/G1 phase may be one of the mechanisms.

Research hotspots and trends of breast cancer stem cells / 上海交通大学学报（医学版）

Objective: To give a retrospective bibliometric analysis of documents about the breast cancer stem cell (BCSC) and reveal the hotspots and trends of global output. Methods: Articles about the research of BCSC between 2014 and 2019 were retrieved in PubMed and Scopus databases, and SciVal was used to evaluate the global scholarly output and identify the most active factors, such as publications, countries, institutions, and top journal percentile, from the indicators of Field-Weighted Citation Impact (FWCI), CiteScore (CS), keywords and topic prominence percentile. Research hotspots and trends were discussed in detail. Results: A total of 4 700 publications on the research of BCSC from 2014 to 2019 were retrieved in this study with FWCI 1.73. The USA was the top country with a total of 1 742 publications and National Institutes of Health was the top institution both in total citation and FWCI. Non-coding RNA (ncRNA), mesenchymal stem cells, and immunotherapy were the most frequently used topics in BCSC. Conclusion: Researches on the correlation of BCSC and ncRNA, tumor microenvironment, and immunotherapy are the hotspots and trends in BCSC.

Cisplatin increases self-renewal ability of oral squamous cell carcinoma stem cells through Notch4 signaling pathway / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To study the effect of Notch4 signaling pathway on the self-renewal capacity of cancer stem cells (CSCs) of oral squamous cell carcinoma PJ15 and PJ41 cells and its mechanism. Methods: The expression of Notch4 gene in head and neck tumors was analyzed using TCGA database. 2 μmol/L cisplatin was used to treat oral squamous cell carcinoma PJ15 and PJ41 cells for 48 h, following with the treatment of Notch pathway inhibitor DAPT for 24 h. Flow cytometry was used to detect the ratio of CSCs, Western blotting and qPCR were used to detect the expressions of CSCs markers (Sox2, Bmi-1, Oct4, Nanog) and downstream genes in Notch4 signaling pathway (JAG1, HEY1, HEY2, HES1, HES2, DLL4, etc.). Notch4 was knocked down by siRNA technology. Notch4 was silenced with siRNA technology. Western blotting and qPCR were used to detect the effect of si-Notch4 and cisplatin on the expression level of CSCs markers. The stem cell pelletingtestwasusedtodetecttheself-renewalabilityof CSCs. Results: Notch4 gene was highly expressed in head and neck tumor tissues (P=0.046). After cisplatin treatment, compared with the control group, expressions of NICD4 and CSCs markers (Sox2, Bmi-1, Oct4, Nanog) as well as downstream genes of Notch4 signaling pathway (JAG1, HEY1, HEY2, HES1,HES2, DLL4, etc.) increased significantly (all P<0.05), and the proportion of ALDH1 positive cells increased significantly (P<0.05); with the further addition of DAPT, the expressions of the above genes decreased significantly (P<0.05 or P<0.01). After knocking down Notch4, compared with the control group, the size and number of spheres of PJ15 cells [1.33±0.47] vs [8.00±0.82], P<0.01) and PJ41 cells ([1.00±0.82] vs [7.67±1.25], P<0.01) were significantly lower than that of the control group; after the addition of 2 μmol/L DDP for 24 h, there was no significant difference in the expressions of Nanog and Bmi-1 gene between si-Notch4 group and control group. Conclusion:Activation of Notch4 signaling pathway can enhance the self-renewal ability of CSCs in oral squamous cell carcinoma PJ15 and PJ41 cells.

HistonedemethylaseJMJD3promotesthestemnessofdiffuselargeBcelllymphoma cells / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To investigate the effect of histone demethylase JMJD3 (jumonji domain-containing protein 3) on the stemness of diffuse large B-cell lymphoma (DLBCL) cells. Methods: The relationship between the expression of JMJD3 and the overall survival of DLBCL patients was analyzed using the clinical data of DLBCL patients in TCGA database. The control plasmid (pCMV) and JMJD3 expression plasmid(pCMV-JMJD3)weretransfectedintoDLBCLcellsofABCandGCBsubtype via lipofectamine transfection. Then, the mRNAlevels of JMJD3,ALDH1, OCT4 and SOX2 were detected by RT-PCR and qPCR; the activity ofALDH1 enzyme was detected by Flow cytometry; the protein expressions of OCT4 and SOX2 were detected by Western blotting. Gene enrichment in DLBCL patients with high JMJD3 expression was analyzed by gene set enrichment analysis (GSEA). Results: The result of prognosis analysis showed that high expression of JMJD3 was related with poor prognosis in DLBCL patients (P<0.05); however, multivariate analysis showed that the expression of JMJD3 was not the independent factor affecting the prognosis of DLBCL patients (all P>0.05). The expression of JMJD3 was remarkably increased in DLBCL cells transfected with pCMV-JMJD3, which led to significantly increased mRNA level and enzyme activity of ALDH1 as well as up-regulated mRNA and protein expressions of OCT4 and SOX2 (P<0.05 or P<0.01). GSEA analysis showed that enrichment of Wnt/β-catenin signaling pathway related gene set was observed in DLBCL patients with high JMJD3 expression (P<0.05). Conclusion: JMJD3 promotes the stemness of DLBCL cells, which may be a potential therapeutic target for DLBCL patients.

Effect and molecular mechanism of Lin28 on 5-Fu sensitivity of hepatocellular carcinoma HepG2 cells / 中国肿瘤生物治疗杂志

@# Objective: To investigate the effect and mechanism of RNA binding protein Lin28 on the 5-fluorouracil (5-Fu) sensitivity of HepG2 cells. Methods: HepG2 cells were transfected with plasmid pcDNA3.1-Lin28 or si-Lin28 (small interfering RNA of Lin28). qPCR and Western blotting were used to detect the expression of Lin28 in HepG2 cells after transfection. Changes of cell proliferation in transfected cells after 5-Fu treatment was detected by CCK8 assay and the 50% inhibitory concentration (IC50) was calculated. Flow cytometry was used to detect apoptotic rate after 5-Fu treatment and the expression of apoptosis-related protein was assayed by Western blotting. The mRNA expressions of drug-resistant miRNAs (let-7a and let-7b), as well as cancer stem cell markers (Oct4, Nanog and Sox2) after transfection were detected by qPCR. Results: As compared to the HepG2/Vector cells, the mRNA and protein expressions of Lin28 were significantly up-regulated in HepG2/Lin28 cells (P<0.05 or P<0.01). Over-expression of Lin28 significantly suppressed the sensitivity of HepG2 cells to 5-Fu (IC50elevated obviously, P<0.05) and significantly increased cell proliferation while decreased apoptotic rate and expression of apoptotic-related protein caspase-3 (all P<0.01). As compared to si-control group, expression of Lin28 in HepG2/si-Lin28 cells was significantly down-regulated (P<0.01). Lin28 knockdown significantly reduced cell proliferation and IC50 of 5-Fu (all P<0.01) but increased apoptotic rate and expression of apoptosis-related protein (P<0.01). Compared with HepG2/Vector group, expressions of let-7a and let-7b, as well as cancer stem cell markers (Oct4, Nanog and Sox2) were significantly increased in HepG2/Lin28 cells (all P<0.01); while these molecules were significantly decreased in HepG2/si-Lin28 cells as comparing to si-control group (all P<0.01). Conclusion: Lin28 can modulate the chemosensitivity of HepG2 cells by regulating the expression of miRNAs and the formation of cancer stem cells. Targeting Lin28 might be a promising approach to improve the chemotherapy efficacy in HCC.