Real-Time Cell Counting in Unlabeled Microscopy Images

The good result reliability of regular analyzes of milk composition could improve the health monitoring of dairy cows and herd management. The aim of this study was the analysis of measurement of abilities and properties of RT (Real Time) system (AfiLab = AfiMilk (NIR measurement unit (near infrared spectroscopy) and electrical conductivity (C) of milk by conductometry) + AfiFarm (calibration and interpretation software)) for the analysis of individual milk samples (IMSs). There were 2 × 30 IMSs in the experiment. The reference values (RVs) of milk components and properties (fat (F), proteins (P), lactose (L), C and the somatic cell count (SCC)) were determined by conventional (direct and indirect: conductometry (C); infrared spectroscopy 1) with the filter technology and 2) with the Fourier transformations (F, P, L); fluoro-opto-electronic cell counting (SCC) in the film on the rotation disc (1) and by flow cytometry (2)) methods. AfiLab method (alternative) showed less close relationships as compared to the RVs as relationships between reference methods. This was expected. However, these relationships (r) were mostly significant: F from .597 to .738 (P ≤ 0.01 and ≤ 0.001); P from .284 to .787 (P > 0.05 and P ≤ 0.001); C .773 (P ≤ 0.001). Correlations (r) were not significant (P > 0.05): L from −.013 to .194; SCC from −.148 to −.133. Variability of the RVs explained the following percentages of variability in AfiLab results: F to 54.4 %; P to 61.9 %; L only 3.8 %; C to 59.7 %. Explanatory power (reliability) of AfiLab results to the animal is increasing with the regularity of their measurements (principle of real time application). Correlation values r (x minus 1.64 × sd for confidence interval (one-sided) at a level of 95 %) can be used for an alternative method in assessing the calibration quality. These limits are F 0.564, P 0.784 and C 0.715 and can be essential with the further implementation of this advanced technology of dairy herd management.

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CircRNA profiling identifies circRNF180 as a tumor suppressor in hepatocellular carcinoma

Epigenomics ◽

10.2217/epi-2020-0385 ◽

2021 ◽

Vol 13 (7) ◽

pp. 513-530

Author(s):

Xi Zeng ◽

Chao Tan ◽

Meile Mo ◽

Xiaoling Qin ◽

Xiaoyun Ma ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Tumor Suppressor ◽

Real Time ◽

Real Time Pcr ◽

Expression Profiles ◽

Cell Counting ◽

Migration And Invasion ◽

Quantitative Real Time Pcr ◽

Potential Biomarker ◽

Hcc Cells

Aim: To explore the expression profiles and functions of circRNAs in hepatocellular carcinoma (HCC). Materials & methods: We obtained circRNA expression profiles through RNA sequencing. Expression levels of circRNAs were confirmed by quantitative real-time PCR. The effects on HCC progression were determined using Cell Counting Kit 8, clone formation and transwell assays. Results: We identified 114 upregulated and 144 downregulated circRNAs in HCC tissues. The results of quantitative real-time PCR showed that circGNAO1, circRNF180 and circMERTK were significantly downregulated in HCC tissues, whereas circSNX6 was significantly upregulated. CircRNF180 was associated with microvascular invasion. Overexpression of circRNF180 inhibits the proliferation, colony formation, migration and invasion of HCC cells. Conclusion: CircRNF180 may function as a tumor suppressor and could serve as a potential biomarker and therapeutic target in HCC.

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MiR-887 Promotes the Progression of Hepatocellular Carcinoma via Targeting VHL

Technology in Cancer Research & Treatment ◽

10.1177/1533033820940425 ◽

2020 ◽

Vol 19 ◽

pp. 153303382094042

Author(s):

Wei Zou ◽

Jun Cheng

Keyword(s):

Hepatocellular Carcinoma ◽

Polymerase Chain Reaction ◽

Real Time ◽

Luciferase Activity ◽

Cell Counting ◽

Luciferase Reporter ◽

Chain Reaction ◽

Gene Assay ◽

Proliferation And Metastasis ◽

Polymerase Chain

Background: MiR-887 has been proved to promote the tumorigenesis in diverse cancers, but its function and downstream mechanism in hepatocellular carcinoma remain obscure. Methods: Quantitative real-time polymerase chain reaction was performed to detect the expression levels of miR-887 in hepatocellular carcinoma tissues and cell lines. MiR-887 mimics and miR-887 inhibitor were transfected into Huh7 and MHCC97H to establish miR-887 overexpression or inhibition models. Cell Counting Kit-8 and colony formation experiment were conducted to monitor cell proliferation. Subcutaneous xenotransplanted tumor model and tail vein injection model in mice were also established to further verify the effect of miR-887 on hepatocellular carcinoma in vivo. The targeting relationship between miR-887 and von Hippel-Lindau tumor suppressor (VHL) was determined by quantitative real-time polymerase chain reaction, Western blot, and luciferase reporter gene assay. Results: miR-887 expression in hepatocellular carcinoma tissues was significantly upregulated. Compared with the control cells, the proliferation and metastasis of cancer cells were enhanced by miR-887 mimics and suppressed by miR-887 inhibitor. Compared with control mice, the volume and weight of subcutaneous tumors of mice in the miR-887 mimics group were significantly elevated, and the significant increase was found in the occurrence of lung metastasis. Moreover, bioinformatics tools showed that miR-887 and VHL had 2 binding sites. Luciferase activity assay demonstrated that miR-887 can inhibit the luciferase activity of VHL, and miR-887 mimics could reduce the expressions of VHL at both messenger RNA and protein levels to increase hypoxia-inducible factor α expression. Conclusion: The upregulation of miR-887 could facilitate the proliferation and metastasis of hepatocellular carcinoma cells via targeting VHL.

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Real-time robust line detection in microscopy images

10.1117/12.290321 ◽

1997 ◽

Cited By ~ 1

Author(s):

Ashit Talukder ◽

David P. Casasent

Keyword(s):

Real Time ◽

Line Detection ◽

Microscopy Images

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Static real-time capture of 3D microscopy images

SPIE Newsroom ◽

10.1117/2.1201503.005832 ◽

2015 ◽

Author(s):

Manuel Martínez-Corral ◽

Ana Doblas ◽

Emilio Sánchez-Ortiga ◽

Jorge Sola-Pikabea ◽

Genaro Saavedra

Keyword(s):

Real Time ◽

3D Microscopy ◽

Microscopy Images

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Assessment of the synergic effect of Avastin and VEGF siRNA on inhibition of cell proliferation and angiogenic factor expression of human breast cancer cells

10.21203/rs.3.rs-889004/v1 ◽

2021 ◽

Author(s):

Abdolkhalegh Deezagi ◽

Bahar Ghorbani

Keyword(s):

Breast Cancer ◽

Cell Proliferation ◽

Cell Death ◽

Real Time ◽

Real Time Pcr ◽

Angiogenic Factor ◽

Cell Counting ◽

Synergic Effect ◽

Vegf Expression ◽

Cell Growth And Migration

Abstract Angiogenesis is an important process in tumor growth and metastasis and vascular endothelial growth factor (VEGF) plays an important role in this process. Several VEGF inhibitors have been developed as anticancer agents including humanized monoclonal antibodies, and various small molecules. The aim of this work was to investigate the effect of combination of VEGF siRNA and Avastin on breast cancer MCF-7 cell line behavior. For this purpose, the cells were treated with different concentrations of Avastin and/or VEGF siRNA and their combination. The cell survival and cell proliferation were assayed by cell counting, trypan blue and MTT tests. The cell migration was assayed by scratching test. VEGF expression was assayed by RT and real-time PCR and ELISA methods. Results indicated the significant increase in cell death following treatment with Avastin (50% cell death at 100 µg/ml). Cell death with VEGF siRNA transfection was lower than Avastin, however, it was significant. This result in VEGF siRNA + Avastin (100 µg/ml) treatment was greater compared to treatment with each of these compounds alone (47%). Scratching results also showed the synergic effect of VEGF siRNA and Avastin (57% decrease). Real-time PCR results showed that Avastin at concentrations of ≥ 50 µg/ml led to 2.5 to 7.5-fold decrease in VEGF expression levels. Also, treatment with VEGF siRNA led to 15.5-fold decrease in VEGF expression. Finally, VEGF expression following VEGF siRNA + Avastin treatment led to a significant 47.5-fold decrease in VEGF expression. It could be concluded that combination of VEGF siRNA and Avastin have a more significant impact on the inhibition of cell growth and migration and it can probably be used as an effective therapeutic approach.

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Effect of DMSA Modified Fe3O4 and Multidrug Resistance Modulator HZ08 on the Reversal of Multidrug Resistance in K562/A02 Cell Line

Blood ◽

10.1182/blood.v124.21.5235.5235 ◽

2014 ◽

Vol 124 (21) ◽

pp. 5235-5235

Author(s):

Fei Shen ◽

Mingfei Zhou ◽

Xuzhang Lu ◽

Yanping Chen ◽

Baoan Chen

Keyword(s):

Multidrug Resistance ◽

Cell Line ◽

Real Time ◽

Caspase 3 ◽

Conflicts Of Interest ◽

Morphological Changes ◽

Leukemia Cell ◽

Cell Counting ◽

Leukemia Cell Line ◽

Gene And Protein Expression

Abstract The objective of the present study was to investigate the reversible effect of HZ08 and daunorubicin(DNR) combined with dimercaptosuccinic acid modified iron oxide (DMSA-Fe3O4) magnetic nanoparticles (MNPs) in human chronic leukemia cell line K562/A02, and the mechanism potentially involved. The growth inhibition rate of K562/A02 cells was determined by Cell Counting Kit-8 (CCK-8) assay. Cell apoptosis and intracellular concentration of DNR were detected by flow cytometry (FCM). DAPI staining was used to view apoptotic cellular morphology. Subsequently, transcription levels of MDR1 mRNA and expression levels of P-glycoprotein (P-gp) and caspase-3 were determined by real time polymerase chain reaction (real-time PCR) and Western blotting analysis, respectively. group clearly exhibited more morphological changes (severe structural alterations) than other groups. In addition, transcription of MDR1 gene and protein expression of P-gp and caspase-3 of K562/A02 cells were regulated at the most remarkable extent in DNR-HZ08-MNPs group when compared with other groups. These findings suggest that the remarkable effects of the novel DNR-HZ08-MNPs on multidrug resistant K562/A02 leukemia cells would be a promising strategy for overcoming multidrug resistance. Disclosures No relevant conflicts of interest to declare.

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Serine/Threonine Kinase 35, a Target Gene of STAT3, Regulates the Proliferation and Apoptosis of Osteosarcoma Cells

Cellular Physiology and Biochemistry ◽

10.1159/000487172 ◽

2018 ◽

Vol 45 (2) ◽

pp. 808-818 ◽

Cited By ~ 8

Author(s):

Zhong Wu ◽

Jie Liu ◽

Siyuan Hu ◽

Yuchang Zhu ◽

Shaohua Li

Keyword(s):

Mrna Expression ◽

Real Time ◽

Real Time Pcr ◽

Gene Set Enrichment Analysis ◽

Cell Counting ◽

Luciferase Reporter ◽

Osteosarcoma Cells ◽

Serine Threonine Kinase ◽

Threonine Kinase ◽

Proliferation And Apoptosis

Background/Aims: Serine/threonine kinase 35 (STK35) may be associated with Parkinson disease and human colorectal cancer, but there have been no reports on the expression levels or roles of STK35 in osteosarcoma. Methods: STK35 mRNA expression was determined in osteosarcoma and bone cyst tissues by real-time PCR. Cell proliferation and apoptosis were assessed by Cell Counting Kit-8 (CCK-8) assay and flow cytometry analysis, respectively. Results: STK35 was up-regulated in osteosarcoma tissues as indicated by analyzing publicly available expression data (GEO dataset E-MEXP-3628) and real-time PCR analysis on our own cohort. We subsequently investigated the effects of STK35 knockdown on two osteosarcoma cell lines, MG63 and U2OS. STK35 knockdown inhibited the growth of osteosarcoma cells in vitro and in xenograft tumors. Meanwhile, STK35 knockdown enhanced apoptosis. Expression of the active forms and the activity of two major executioner caspases, caspase 3 and caspase 7, were also increased in osteosarcoma cells with STK35 silenced. Additionally, Gene Set Enrichment Analysis (GSEA) identified that the JAK/STAT signaling pathway was positively correlated with STK35 expression. The mRNA expression of STK35 was repressed by STAT3 small interfering RNA (siRNA), but not by siRNA of STAT4, STAT5A or STAT6. A luciferase reporter assay further demonstrated that STAT3 transcriptionally regulated STK35 expression. A chromatin immunoprecipitation (ChIP) assay confirmed the direct recruitment of STAT3 to the STK35 promoter. The promotion effects of STAT3 knockdown on cell apoptosis were partially abolished by STK35 overexpression. Furthermore, STK35 mRNA expression was positively correlated with STAT3 mRNA expression in osteosarcoma tissues by Pearson correlation analysis. Conclusions: These results collectively reveal that STAT3 regulates the transcription of STK35 in osteosarcoma. STK35 may exert an oncogenic role in osteosarcoma.

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