An Automatic Cell Counting Method for a Microscopic Tissue Image from Breast Cancer

Abstract Background Recent studies have indicated that serpin peptidase inhibitor, clade A, member 3 (SERPINA3) is a potential marker associated with tumor progression, which connoted that SERPINA3 is related to malignant phenotypes in cancer. However, the biological function of SERPINA3 in breast cancer (BC) remains unclear. Methods Bioinformatics data were downloaded from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Immunohistochemical staining (IHC) was conducted to determine SERPINA3 expression. With strong aggressive abilities, triple-negative breast cancer (TNBC) cell lines (MDA-MB-231, BT549 and MDA-MB-436) were obtained to examine SERPINA3 expression and functions. Wound healing and Transwell assays were performed to measure cell migration and invasion. Cell Counting Kit-8 (CCK-8) assay was conducted to detect cell proliferation abilities and cell viabilities. Results SERPINA3 was upregulated in BC tissues. Functional assays suggested that overexpression of SERPINA3 significantly promoted cell proliferation, where migration and invasion of TNBC cells were accelerated. Knockdown of SERPINA3 had the opposite effects. These results causing by overexpression of SERPINA3 were also confirmed in non-TNBC cell lines. Overexpression of SERPINA3 remarkably enhanced the epithelial–mesenchymal transition (EMT) by upregulating the EMT markers and EZH2. In addition, the overexpression of SERPINA3 reduced the sensitivity of TNBC cells to cisplatin. Conclusion SERPINA3 can regulate the migration, invasion and EMT of TNBC cells and increased expression of SERPINA3 confers resistance to cisplatin in TNBC cells. We discern it is required for the regulation of BC progression and is a critical target for the clinical treatment of BC.

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Adaptive and automatic red blood cell counting method based on microscopic hyperspectral imaging technology

Journal of Optics ◽

10.1088/2040-8986/aa95d7 ◽

2017 ◽

Vol 19 (12) ◽

pp. 124014 ◽

Cited By ~ 3

Author(s):

Xi Liu ◽

Mei Zhou ◽

Song Qiu ◽

Li Sun ◽

Hongying Liu ◽

...

Keyword(s):

Blood Cell ◽

Hyperspectral Imaging ◽

Red Blood Cell ◽

Cell Counting ◽

Counting Method ◽

Imaging Technology ◽

Blood Cell Counting

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Methylseleninic Acid Suppresses Breast Cancer Growth via the JAK2/STAT3 Pathway

Reproductive Sciences ◽

10.1177/1933719118815582 ◽

2018 ◽

Vol 26 (6) ◽

pp. 829-838 ◽

Cited By ~ 4

Author(s):

Changwei Qiu ◽

Tao Zhang ◽

Xinying Zhu ◽

Jinxia Qiu ◽

Kangfeng Jiang ◽

...

Keyword(s):

Breast Cancer ◽

Tumor Growth ◽

Western Blot ◽

Cancer Cell ◽

Anticancer Drug ◽

Cancer Cell Line ◽

Janus Kinase ◽

Cell Counting ◽

Stat3 Pathway ◽

Methylseleninic Acid

Previous studies show that methylseleninic acid (MSA), which is the most common selenium derivative used as a drug in humans, exerts specific cytotoxic effects in several cancer cell types. However, the complex mechanism of these effects has not been fully elucidated. Here, we demonstrate by Cell Counting Kit-8 in mouse breast cancer cell line 4T1 that MSA inhibits cell viability in a concentration-dependent (5, 10, 20 μmol/L) and time-dependent (6, 12, 24 hours) manner. Flow cytometry, Western blot, and Reverse Transcription-Polymerase Chain Reaction (RT-PCR) analyses indicated that MSA inhibits cancer cell invasion and induces apoptosis by the activation of caspase-3, poly ADP ribose polymerase 1 (PARP1), and BCL2-associated X. Furthermore, MSA demonstrated anticancer activity by inhibiting the Janus kinase 2/signal transducers and activators of transcription 3 (JAK2/STAT3) pathway. The MSA treatment for 24 hours decreased the phosphorylation of JAK2 and STAT3 in 4T1 cells by Western blot. We also confirmed this with the use of a JAK2 chemical inhibitor, AG490, as a positive control. In a 4T1 orthotopic allograft model, morphological and TdT-mediated dUTP nick-end labeling analyses showed that MSA treatment (1.5 mg/kg/weight) for 28 days inhibits tumor growth consistent with the clinical anticancer drug cyclophosphamide. Our observations demonstrate that MSA is a potent anticancer drug in breast cancer and uncovered a key role of the JAK2/STAT3 pathway in modulating tumor growth.

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Anlotinib Inhibits the Growth of Breast Cancer Cells by Promoting Autophagy and Apoptosis via the Akt/GSK-3α Signalling Pathway

10.21203/rs.3.rs-97756/v1 ◽

2020 ◽

Author(s):

shuyi chen ◽

Ping Zhu ◽

Xue Wang ◽

Youping Jin ◽

Xiuling Zhi ◽

...

Keyword(s):

Breast Cancer ◽

Cell Cycle ◽

Cell Lines ◽

Kinase Inhibitor ◽

Underlying Mechanism ◽

Cell Counting ◽

Tunel Staining ◽

Western Blots ◽

Qrt Pcr ◽

Mcf 7

Abstract Background: Anlotinib, a multi-target tyrosine kinase inhibitor, has already been indicated to have significant anticancer effects on lung cancer, colon cancer and ovarian cancer in a phase II clinical trial, but its effect on breast cancer (BC) has not been adequately investigated. Methods: The proliferation activity of BC cell lines MCF-7 and MDA-MB-231 with the treatment of anlotinib was tested by Cell Counting Kit-8 (CCK-8) assay and immunocytochemistry (ICC) staining. We investigated the alteration of cell cycle and apoptosis and autophagy level and the underlying mechanism in the cell lines by quantitative real-time reverse-transcription polymerase chain reaction (qRT-PCR), Western blots, ICC and TUNEL staining and flow cytometry. Further, AT-3 cells were subcutaneously injected into C57BL/6 mice, followed by anlotinib intragastrically. The extracted tumours were assessed by qRT-PCR, Western blots and immunohistochemistry.Results: We found that anlotinib suppressed the cell viability and proliferation of MCF-7 and MDA-MB-231 cell lines and tumour growth in BC xenografts in mice, likely due to abnormal cell cycle arrest and induction of autophagy and apoptosis. Then, we further examined the underlying mechanism of anlotinib, and the results indicated that anlotinib induced apoptosis by promoting autophagy in MCF-7 and MDA-MB-231 cells by regulating the Akt/GSK-3α pathway. The analysis of data from patients with BC collected in TCGA revealed that increased VEGFA expression was related to BC.Conclusions: Our study demonstrated that anlotinib inhibited the growth of BC cells via promoting apoptosis through autophagy mediated by Akt/GSK-3α signalling and may be an effective new drug for BC treatment.

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A novel, rapid, cell-counting method for unfixed frozen brains comprehensively quantifies at least four neural cell populations

Molecular Psychiatry ◽

10.1038/mp.2011.152 ◽

2011 ◽

Vol 16 (12) ◽

pp. 1155-1155 ◽

Cited By ~ 29

Author(s):

Y Hayashi ◽

N Nihonmatsu-Kikuchi ◽

S-I Hisanaga ◽

Y Tatebayashi

Keyword(s):

Neural Cell ◽

Cell Counting ◽

Cell Populations ◽

Counting Method

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Chemotherapy-elicited exosomal miR-378a-3p and miR-378d to promote breast cancer stemness and chemoresistance via the activation of EZH2/STAT3 signaling.

Journal of Clinical Oncology ◽

10.1200/jco.2021.39.15_suppl.e12615 ◽

2021 ◽

Vol 39 (15_suppl) ◽

pp. e12615-e12615

Author(s):

Jin Zhang

Keyword(s):

Breast Cancer ◽

Drug Resistance ◽

Neoadjuvant Chemotherapy ◽

Pathologic Complete Response ◽

Chemotherapeutic Agents ◽

Cell Counting ◽

Chemotherapy Response ◽

Mouse Tumor ◽

Response To Chemotherapy ◽

Serum Exosomes

e12615 Background: Not all breast cancer (BC) patients who receive neoadjuvant chemotherapy achieve a pathologic complete response (pCR), but the reasons for this are unknown. Previous studies have shown that exosomes produced in the tumor microenvironment in response to chemotherapy promote a chemotherapy-resistant phenotype in tumors. However, the role of BC chemo-elicited exosomes in regulating chemoresistance is poorly understood. Methods: Using commercial kits, serum exosomes were extracted from patients before neoadjuvant chemotherapy, after one cycle of chemotherapy and after four cycles of chemotherapy consisting of doxorubicin (DOX) and paclitaxel (PTX). Their miRNAs were sequenced, and the correlation between the sequencing results and chemotherapy effects was further verified by RT-qPCR using patient serum exosomes. Cell Counting Kit-8 (CCK-8) was used to detect chemosensitivity. Stemness was assessed by CD44+/CD24- population analysis and mammosphere formation assays. Chromatin immunoprecipitation (ChIP) experiments were performed to verify the binding of signal transducer and activator of transcription 3 (STAT3) to the promoter of miRNAs. Results: Here, we provide clinical evidence that chemotherapy-elicited exosomal miR-378a-3p and miR-378d are closely related to the chemotherapy response and that exosomes produced by BC cells after stimulation with DOX or PTX deliver miR-378a-3p and miR-378d to neighboring cells to activate WNT and NOTCH stemness pathways and induce drug resistance by targeting Dickkopf 3 (DKK3) and NUMB. In addition, STAT3, which is enhanced by zeste homolog 2 (EZH2), bound to the promoter regions of miR-378a-3p and miR-378d, thereby increasing their expression in exosomes. More importantly, chemotherapeutic agents combined with the EZH2 inhibitor tazemetostat reversed chemotherapy-elicited exosome induced drug resistance in a nude mouse tumor xenograft model. Conclusions: This study revealed a novel mechanism of acquired chemoresistance whereby chemotherapy activates the EZH2/STAT3 axis in BC cells, which then secrete chemotherapy-elicited exosomes enriched in miR-378a-3p and miR-378d. These exosomes are absorbed by chemotherapy-surviving BC cells, leading to activation of WNT and NOTCH stem cell pathways via the targeting of DKK3 and NUMB and subsequently resulting in drug resistance. Therefore, blocking this adaptive mechanism during chemotherapy may reduce the development of chemotherapy resistance and maximize the therapeutic effect.

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Long Noncoding RNA TTC39A-AS1 Promotes Breast Cancer Tumorigenicity by Sponging MicroRNA-483-3p and Thereby Upregulating MTA2

Pharmacology ◽

10.1159/000515909 ◽

2021 ◽

pp. 1-15

Author(s):

Zhaohui Zhou ◽

Ping Yang ◽

Binming Zhang ◽

Maohui Yao ◽

Yali Jia ◽

...

Keyword(s):

Breast Cancer ◽

Noncoding Rna ◽

Flow Cytometric Analysis ◽

The Cancer Genome Atlas ◽

Cell Counting ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Rna Immunoprecipitation ◽

Anticancer Treatment ◽

High Level

Introduction: In recent years, the regulatory activities of long noncoding RNAs have received increasing attention as an important research focus. This study aimed to characterize the expression and detailed roles of TTC39A antisense RNA 1 (TTC39A-AS1) in breast cancer (BC), in addition to concentrating on its downstream mechanisms. Methods: Quantitative RT-PCR was performed to determine the expression levels of TTC39A-AS1, microRNA-483-3p (miR-483-3p), and metastasis-associated gene 2 (MTA2). Further, the detailed functions of TTC39A-AS1 in BC cells were confirmed using the Cell Counting Kit 8 assay, flow cytometric analysis, and Transwell cell migration and invasion assays. The targeting relationship between TTC39A-AS1, miR-483-3p, and MTA2 in BC was predicted via bioinformatics analysis and further confirmed by performing the luciferase reporter assay and RNA immunoprecipitation. Results: TTC39A-AS1 was present in high levels in BC; this result was confirmed in our sample cohort and The Cancer Genome Atlas database. Patients with BC with a high level of TTC39A-AS1 had a shorter overall survival than those with a low level of TTC39A-AS1. Functionally, the absence of TTC39A-AS1 accelerated cell apoptosis but retained cell proliferation, migration, and invasion. Mechanistically, TTC39A-AS1 functioned as a competing endogenous RNA in BC by sponging miR-483-3p and thereby indirectly increasing MTA2 expression. Finally, rescue experiments revealed that the tumor-inhibiting actions of TTC39A-AS1 knockdown on the malignant characteristics of BC cells could be reversed by inhibiting miR-483-3p or upregulating MTA2. Conclusion: The newly identified TTC39A-AS1/miR-483-3p/MTA2 pathway was revealed to be a critical regulator in the tumorigenicity of BC, possibly offering a novel therapeutic direction for the anticancer treatment of BC.

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A Novel Automated Blood Cell Counting Method Based on Deconvolution and Convolution and Its Application to Neural Networks

Advances in Intelligent Systems and Computing - Advanced Computing and Systems for Security ◽

10.1007/978-981-15-2930-6_6 ◽

2020 ◽

pp. 67-78

Author(s):

Joydeep Chatterjee ◽

Semanti Chakraborty ◽

Kanik Palodhi

Keyword(s):

Neural Networks ◽

Blood Cell ◽

Cell Counting ◽

Counting Method ◽

Blood Cell Counting

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MiR-142-5p Acts as a Significant Regulator Through Promoting Proliferation, Invasion, and Migration in Breast Cancer Modulated by Targeting SORBS1

Technology in Cancer Research & Treatment ◽

10.1177/1533033819892264 ◽

2019 ◽

Vol 18 ◽

pp. 153303381989226 ◽

Cited By ~ 3

Author(s):

Weixuan Yu ◽

Dongwei Li ◽

Yunda Zhang ◽

Cheukfai Li ◽

Chuanzhao Zhang ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Expression Patterns ◽

Sh3 Domain ◽

Cell Counting ◽

Luciferase Reporter ◽

Invasion And Migration ◽

And Migration ◽

Low Expression

Background: Numerous researches have demonstrated that miR-142-5p plays significant roles in several cancers, although the functional characteristic of miR-142-5p in breast cancer has not been determined. This study is designed to explore the biological significance of miR-142-5p in breast cancer clinical implication and mechanism of action. Methods: The differential expression patterns of miR-142-5p and Sorbin and SH3 domain-containing protein 1 and correlations between them and clinical significances were analyzed based on data from database. The expression levels of miR-142-5p in breast cancer cells were detected using quantitative real-time polymerase chain reaction. Cell counting kit-8, transwell, and wound healing assays were used to explore the potential functions of miR-142-5p in breast cancer cells. In addition, bioinformatics prediction analysis and luciferase reporter assay were utilized to predict and identify the potential target gene of miR-142-5p. A rescue experiment was conducted by transfecting miR-142-5p inhibitors and si-Sorbin and SH3 domain-containing protein 1 into cells to explore miR-142-5p/Sorbin and SH3 domain-containing protein 1 pairs on breast cancer cells behaviors. Results: The analysis results showed that miR-142-5p was highly expressed in patients with breast cancer, while Sorbin and SH3 domain-containing protein 1 presented a trend of low expression. The clinical significances analysis suggested that the overexpression of miR-142-5p is closely correlated with metastasis, while low expression of Sorbin and SH3 domain-containing protein 1 is correlated with clinicopathological characteristics and poor overall survival in patients with breast cancer. In vitro exploration, the expression of miR-142-5p was upregulated in breast cancer cells and inhibition of miR-142-5p expression significantly reduced the proliferation, invasion, and migration of breast cancer cells. Through rescue experiments, breast cancer cells proliferation, invasion, and migration reduction induced by silencing of miR-142-5p were reversed via knockdown Sorbin and SH3 domain-containing protein 1. Conclusion: Our findings insinuate that miR-142-5p functions as a positive regulator of promoting breast cancer cells biological behaviors and clinical metastasis, possibly regulated by targeting Sorbin and SH3 domain-containing protein 1, thus providing valuable information in the development of preventive or even therapeutic strategies for utilizing miR-142-5p as a promising target.

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