Basic Colorimetric Proliferation Assays: MTT, WST, and Resazurin

Background: Biosynthesis of gold nanoparticles from medicinal plants has become an interesting strategy in biomedical research due to its exclusive properties including less toxic cellular level through its ecofriendly biological function. Objective: To examine the anti-lipid accumulation effect of spherical gold nanoparticles (size 10-20 nm) synthesized from Dendropanax morbifera Léveille (D-AuNPs) in both 3T3-L1 and HepG2 cells. Method: 3T3-L1 preadipocytes and HepG2 hepatocytes were stimulated with cocktail media to generate obese and fatty liver disease models. Cell cytotoxicity and cell proliferation assays were performed in adipocytes at different stages of growth. An anti-lipid accumulation assay was performed in 3T3-L1 obese and HepG2 fatty liver models using different doses of D-AuNPs. Expression of adipogenic genes of PPARγ, CEBPα, Jak2, STAT3, and ap2 and hepatogenic genes PPARα, FAS, and ACC was measured by real-time PCR. In addition, protein expression of PPARγ and CEBPα was evaluated by immunoblotting assay. Result: We found that D-AuNPs (size 10–20 nm) at concentrations up to 100 µg/ml were nontoxic to 3T3-L1 and HepG2 at post-confluent and mature stages. In addition, pretreatment of D-AuNPs at post-confluent stage reduced triglyceride content. In addition, the adipogenesis process was negatively controlled by D-AuNPs, with downregulated PPARγ, CEBPα, Jak2, STAT3, and ap2 expression in 3T3-L1 cells and FAS and ACC levels in HepG2 cells. Conclusion: These data indicated that D-AuNPs exert antiadipogenic properties. We hypothesize that Dendropanax contains a large amount of phenolic compound that coats the surface of gold nanoparticles and has the ability to reduce the excess amount of lipid in both cell lines.

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Effects of an Acetic Acid and Acetone Mixture on the Characteristics and Scaffold–Cell Interaction of Electrospun Polycaprolactone Membranes

Applied Sciences ◽

10.3390/app9204350 ◽

2019 ◽

Vol 9 (20) ◽

pp. 4350 ◽

Cited By ~ 4

Author(s):

Minh Hieu Ho ◽

Thien Bui-Thuan Do ◽

Nhi Ngoc-Thao Dang ◽

An Nguyen-My Le ◽

Hanh Thi-Kieu Ta ◽

...

Keyword(s):

Acetic Acid ◽

Cell Interaction ◽

Fibrous Materials ◽

Cell Adherence ◽

Physical Strength ◽

Membrane Morphology ◽

Cell Scaffold ◽

Different Types ◽

Proliferation Assays ◽

Cell Adhesion And Proliferation

Green electrospinning has attracted great interest since non-toxic solvents were shown to be applicable in the fabrication of fibrous materials while ensuring health safety and environmental protection. Less harmful reagents such as acetone (AC) and acetic acid (AA) have been employed in this field in recent years. However, research in this area is still rare, yielding only preliminary results. In this study, two different types of solvents (pure AC and an AA/AC mixture) were used to fabricate electrospun polycaprolactone (PCL) membranes. Sample morphology, wettability, tensile strength, and chemical composition were compared between two types of membranes. Cell–scaffold interaction was also examined by cell adhesion and proliferation assays. The results demonstrate that the two types of solvents had significant effects on membrane morphology, physical strength, and cell adherence behaviors, which should be considered for different application purposes.

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Functional consequences of the first reported mutations of the proto-oncogene PTTG1IP/PBF

Endocrine Related Cancer ◽

10.1530/erc-16-0340 ◽

2017 ◽

Vol 24 (9) ◽

pp. 459-474 ◽

Cited By ~ 2

Author(s):

W Imruetaicharoenchoke ◽

A Fletcher ◽

W Lu ◽

R J Watkins ◽

B Modasia ◽

...

Keyword(s):

Colony Formation ◽

Human Cancer ◽

Soft Agar ◽

Cellular Migration ◽

3T3 Fibroblasts ◽

Wide Range ◽

Functional Consequences ◽

Altered Cell ◽

Proliferation Assays ◽

Nih 3T3

Pituitary tumor-transforming gene 1-binding factor (PTTG1IP; PBF) is a multifunctional glycoprotein, which is overexpressed in a wide range of tumours, and significantly associated with poorer oncological outcomes, such as early tumour recurrence, distant metastasis, extramural vascular invasion and decreased disease-specific survival. PBF transforms NIH 3T3 fibroblasts and induces tumours in nude mice, while mice harbouring transgenic thyroidal PBF expression show hyperplasia and macrofollicular lesions. Our assumption that PBF becomes an oncogene purely through increased expression has been challenged by the recent report of mutations in PBF within the Catalogue of Somatic Mutations in Cancer (COSMIC) database. We therefore sought to determine whether the first 10 PBF missense substitutions in human cancer might be oncogenic. Anisomycin half-life studies revealed that most mutations were associated with reduced protein stability compared to wild-type (WT) PBF. Proliferation assays narrowed our interest to two mutational events which significantly altered cell turnover: C51R and R140W. C51R was mainly confined to the endoplasmic reticulum while R140W was apparent in the Golgi apparatus. Both C51R and R140W lost the capacity to induce cellular migration and significantly reduced cell invasion. Colony formation and soft agar assays demonstrated that, in contrast to WT PBF, both mutants were unable to elicit significant colony formation or anchorage-independent growth. However, C51R and R140W retained the ability to repress radioiodide uptake, a functional hallmark of PBF. Our data reveal new insight into PBF function and confirm that, rather than being oncogenic, mutations in PBF are likely to be passenger effects, with overexpression of PBF the more important aetiological event in human cancer.

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Anti-IL-6 Receptor Antibody Causes Less Promotion of Tuberculosis Infection than Anti-TNF- Antibody in Mice

Clinical and Developmental Immunology ◽

10.1155/2011/404929 ◽

2011 ◽

Vol 2011 ◽

pp. 1-9 ◽

Cited By ~ 14

Author(s):

Masaji Okada ◽

Yoko Kita ◽

Noriko Kanamaru ◽

Satomi Hashimoto ◽

Yasushi Uchiyama ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Spleen Cell ◽

Tuberculosis Infection ◽

Specific Response ◽

Tnf Receptor ◽

Spleen Cells ◽

Histopathological Findings ◽

Receptor Antibody ◽

Cell Responses ◽

Proliferation Assays

Objective. Our aim was to investigate the effects of IL-6 blockade on the progression ofMycobacterium tuberculosis(TB) and compare them with those of TNF-αblockade in mice.Methods. Mice were intravenously infected with TB and injected with antibodies. Survival was monitored and histological and immunological studies were carried out.Results. All anti-IL-6R Ab-treated mice and 8 of 10 control mice survived until sacrificed 224 days after TB challenge, whereas anti-TNF-αAb-treated mice all died between 120 and 181 days. Anti-IL-6R Ab-treated mice exhibited no significant differences in TB CFU in organs, including the lungs, and no deterioration in histopathology compared to control mice at 4 weeks. In contrast, anti-TNF-αAb-treated mice exhibited increased TB CFU and greater progression of histopathological findings in organs than control mice. Spleen cells from anti-TNF-αAb-treated mice had decreased antigen-specific response in IFN-γrelease and proliferation assays. The results in anti-IL-6R Ab-treated mice suggest that spleen cell responses were decreased to a lesser degree. Similar results were obtained in IL-6 knockout (KO) mice, compared with TNF receptor 1 (TNFR1) KO and TNFR1/IL-6 double KO (DKO) mice.Conclusion. IL-6R blockade promotes the progression of TB infection in mice far less than TNF-αblockade.

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SALL4 is a key regulator of survival and apoptosis in human leukemic cells

Blood ◽

10.1182/blood-2007-11-126326 ◽

2008 ◽

Vol 112 (3) ◽

pp. 805-813 ◽

Cited By ~ 88

Author(s):

Jianchang Yang ◽

Li Chai ◽

Chong Gao ◽

Taylor C. Fowles ◽

Zaida Alipio ◽

...

Keyword(s):

Stem Cell ◽

Leukemia Cell ◽

Annexin V ◽

Leukemic Cell ◽

Leukemic Cells ◽

Immunodeficient Mice ◽

Leukemia Cell Lines ◽

Human Leukemic Cells ◽

Cell Phenotypes ◽

Proliferation Assays

Abstract Increasing studies suggest that SALL4 may play vital roles in leukemogenesis and stem cell phenotypes. We have mapped the global gene targets of SALL4 using chromatin immunoprecipitation followed by microarray hybridization and identified more than 2000 high-confidence, SALL4-binding genes in the human acute promyelocytic leukemic cell line, NB4. Analysis of SALL4-binding sites reveals that genes involved in cell death, cancer, DNA replication/repair, and cell cycle were highly enriched (P < .05). These genes include 38 important apoptosis-inducing genes (TNF, TP53, PTEN, CARD9, CARD11, CYCS, LTA) and apoptosis-inhibiting genes (Bmi-1, BCL2, XIAP, DAD1, TEGT). Real-time polymerase chain reaction has shown that expression levels of these genes changed significantly after SALL4 knockdown, which ubiquitously led to cell apoptosis. Flow cytometry revealed that reduction of SALL4 expression in NB4 and other leukemia cell lines dramatically increased caspase-3, annexin V, and DNA fragmentation activity. Bromodeoxyuridine-incorporation assays showed decreased numbers of S-phase cells and increased numbers of G1- and G2-phase cells indicating reduced DNA synthesis, consistent with results from cell proliferation assays. In addition, NB4 cells that express low levels of SALL4 have significantly decreased tumorigenecity in immunodeficient mice. Our studies provide a foundation in the development of leukemia stem cell–specific therapy by targeting SALL4.

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Evaluation of the T Cell Workshop Monoclonal Antibodies in In Vitro Lymphocyte Proliferation Assays

Leukocyte Typing II ◽

10.1007/978-1-4613-8587-5_15 ◽

1986 ◽

pp. 173-187

Author(s):

Nicolas L. von Jeney ◽

Katja Olas ◽

Walter Knapp

Keyword(s):

Monoclonal Antibodies ◽

T Cell ◽

Lymphocyte Proliferation ◽

Proliferation Assays

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Cell Proliferation Assays

In Vitro Bioassay Techniques for Anticancer Drug Discovery and Development ◽

10.1201/9781315191256-5 ◽

2017 ◽

pp. 33-40

Keyword(s):

Cell Proliferation ◽

Proliferation Assays

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Genetic diversity and immunogenicity of the merozoite surface protein 1 C-terminal 19-kDa fragment of Plasmodium ovale imported from Africa into China

Parasites & Vectors ◽

10.1186/s13071-021-05086-6 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Qinwen Xu ◽

Sihong Liu ◽

Kokouvi Kassegne ◽

Bo Yang ◽

Jiachen Lu ◽

...

Keyword(s):

Genetic Diversity ◽

Immune Responses ◽

Expression System ◽

Surface Protein ◽

Merozoite Surface Protein ◽

Enzyme Linked Immunosorbent Assay ◽

High Avidity ◽

Plasmodium Ovale ◽

Merozoite Surface Protein 1 ◽

Proliferation Assays

Abstract Background Merozoite surface protein 1 (MSP1) plays an essential role in erythrocyte invasion by malaria parasites. The C-terminal 19-kDa region of MSP1 has long been considered one of the major candidate antigens for a malaria blood-stage vaccine against Plasmodium falciparum. However, there is limited information on the C-terminal 19-kDa region of Plasmodium ovale MSP1 (PoMSP119). This study aims to analyze the genetic diversity and immunogenicity of PoMSP119. Methods A total of 37 clinical Plasmodium ovale isolates including Plasmodium ovale curtisi and Plasmodium ovale wallikeri imported from Africa into China and collected during the period 2012–2016 were used. Genomic DNA was used to amplify P. ovale curtisi (poc) msp119 (pocmsp119) and P. ovale wallikeri (pow) msp119 (powmsp119) genes by polymerase chain reaction. The genetic diversity of pomsp119 was analyzed using the GeneDoc version 6 programs. Recombinant PoMSP119 (rPoMSP119)-glutathione S-transferase (GST) proteins were expressed in an Escherichia coli expression system and analyzed by western blot. Immune responses in BALB/c mice immunized with rPoMSP119-GST were determined using enzyme-linked immunosorbent assay. In addition, antigen-specific T cell responses were assessed by lymphocyte proliferation assays. A total of 49 serum samples from healthy individuals and individuals infected with P. ovale were used for the evaluation of natural immune responses by using protein microarrays. Results Sequences of pomsp119 were found to be thoroughly conserved in all the clinical isolates. rPoMSP119 proteins were efficiently expressed and purified as ~ 37-kDa proteins. High antibody responses in mice immunized with rPoMSP119-GST were observed. rPoMSP119-GST induced high avidity indexes, with an average of 92.57% and 85.32% for rPocMSP119 and rPowMSP119, respectively. Cross-reactivity between rPocMSP119 and rPowMSP119 was observed. Cellular immune responses to rPocMSP119 (69.51%) and rPowMSP119 (52.17%) induced in rPocMSP119- and rPowMSP119-immunized mice were found in the splenocyte proliferation assays. The sensitivity and specificity of rPoMSP119-GST proteins for the detection of natural immune responses in patients infected with P. ovale were 89.96% and 75%, respectively. Conclusions This study revealed highly conserved gene sequences of pomsp119. In addition, naturally acquired humoral immune responses against rPoMSP1 were observed in P. ovale infections, and high immunogenicity of rPoMSP119 in mice was also identified. These instructive findings should encourage further testing of PoMSP119 for rational vaccine design. Graphical abstract

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