List of Titles

Apoptosis and beyond: cytometry in studies of programmed cell death

- Wlodkowic, Donald, Telford, W, Skommer, Joanna, Darzynkiewicz, Zbigniew

Authors: Wlodkowic, Donald , Telford, W , Skommer, Joanna , Darzynkiewicz, Zbigniew
Date: 2011
Type: Text , Book chapter
Relation: Methods in Cell Biology p. 55-98
Full Text: false
Reviewed:
Description: A cell undergoing apoptosis demonstrates multitude of characteristic morphological and biochemical features, which vary depending on the inducer of apoptosis, cell type and the “time window” at which the process of apoptosis is observed. Because the gross majority of apoptotic hallmarks can be revealed by flow and image cytometry, the cytometric methods become a technology of choice in diverse studies of cellular demise. Variety of cytometric methods designed to identify apoptotic cells, detect particular events of apoptosis and probe mechanisms associated with this mode of cell death have been developed during the past two decades. In the present review, we outline commonly used methods that are based on the assessment of mitochondrial transmembrane potential, activation of caspases, DNA fragmentation, and plasma membrane alterations. We also present novel developments in the field such as the use of cyanine SYTO and TO-PRO family of probes. Strategies of selecting the optimal multiparameter approaches, as well as potential difficulties in the experimental procedures, are thoroughly summarized

MicroRNAs in a hypertrophic heart : From foetal life to adulthood

- Sadiq, Shahzad, Crowley, Tamsyn, Charchar, Fadi, Sanigorski, Andrew, Lewandowski, Paul

Authors: Sadiq, Shahzad , Crowley, Tamsyn , Charchar, Fadi , Sanigorski, Andrew , Lewandowski, Paul
Date: 2017
Type: Text , Journal article
Relation: Biological Reviews Vol. 92, no. 3 (2017), p. 1314-1331
Full Text:
Reviewed:
Description: The heart is the first organ to form and undergoes adaptive remodelling with age. Ventricular hypertrophy is one such adaptation, which allows the heart to cope with an increase in cardiac demand. This adaptation is necessary as part of natural growth from foetal life to adulthood. It may also occur in response to resistance in blood flow due to various insults on the heart and vessels that accumulate with age. The heart can only compensate to this increase in workload to a certain extent without losing its functional architecture, ultimately resulting in heart failure. Many genes have been implicated in cardiac hypertrophy, however none have been shown conclusively to be responsible for pathological cardiac hypertrophy. MicroRNAs offer an alternative mechanism for cellular regulation by altering gene expression. Since 1993 when the function of a non-coding DNA sequence was first discovered in the model organism Caenorhabditis elegans, many microRNAs have been implicated in having a central role in numerous physiological and pathological cellular processes. The level of control these antisense oligonucleotides offer can often be exploited to manipulate the expression of target genes. Moreover, altered levels of microRNAs can serve as diagnostic biomarkers, with the prospect of diagnosing a disease process as early as during foetal life. Therefore, it is vital to ascertain and investigate the function of microRNAs that are involved in heart development and subsequent ventricular remodelling. Here we present an overview of the complicated network of microRNAs and their target genes that have previously been implicated in cardiogenesis and hypertrophy. It is interesting to note that microRNAs in both of these growth processes can be of possible remedial value to counter a similar disease pathophysiology.

Multivariate analysis of apoptotic markers versus cell cycle phase in living human cancer cells by microfluidic cytometry

- Akagi, Jin, Skommer, Joanna, Matuszek, Anna, Takeda, Kazuo, Fujimura, Yuu, Khoshmanesh, Khashayar, Kalantar-zadeh, Kourosh, Mitchell, Arnan, Errington, Rachel, Smith, Paul, Darzynkiewicz, Zbigniew, Wlodkowic, Donald

Authors: Akagi, Jin , Skommer, Joanna , Matuszek, Anna , Takeda, Kazuo , Fujimura, Yuu , Khoshmanesh, Khashayar , Kalantar-zadeh, Kourosh , Mitchell, Arnan , Errington, Rachel , Smith, Paul , Darzynkiewicz, Zbigniew , Wlodkowic, Donald
Date: 2013
Type: Text , Conference paper
Relation: Microfluidics, BioMEMS, and Medical Microsystems XI
Full Text: false
Reviewed:
Description: Measurement of apoptotic markers in tumors can be directly correlated with the cell cycle phase using flow cytometry (FCM). The conventional DNA content analysis requires cell permeabilization to stain nuclei with fluorescent probes such as propidium iodide or use of a costly UV-excitation line for Hoechst 33342 probe. The access to FCM is also still limited to centralized core facilities due to its inherent high costs and complex operation. This work describes development and proof-of-concept validation of a portable and user-friendly microfluidic flow cytometer (μFCM) that can perform multivariate real time analysis on live cells using sampling volumes as small as 10 microliters. The μFCM system employs disposable microfluidic cartridges fabricated using injection molding in poly(methylmethacrylate) transparent thermoplastic. Furthermore, the dedicated and miniaturized electronic hardware interface enables up to six parameter detection using a combination of spatially separated solid-state 473 (10 mW) and 640 nm (20 mW) lasers and x-y stage for rapid laser alignment adjustment. We provide new evidence that a simple 2D flow focusing on a chip is sufficient to measure cellular DNA content in live tumor cells using a far-red DNA probe DRAQ5. The feasibility of using the μFCM system for a dose-response profiling of investigational anti-cancer agents on human hematopoietic cancer cells is also demonstrated. The data show that μFCM can provide a viable novel alternative to conventional FCM for multiparameter detection of caspase activation and dissipation of mitochondrial inner membrane potential (ΔΨm) in relation to DNA content (cell cycle phase) in live tumor cells.
Description: Measurement of apoptotic markers in tumors can be directly correlated with the cell cycle phase using flow cytometry (FCM). The conventional DNA content analysis requires cell permeabilization to stain nuclei with fluorescent probes such as propidium iodide or use of a costly UV-excitation line for Hoechst 33342 probe. The access to FCM is also still limited to centralized core facilities due to its inherent high costs and complex operation. This work describes development and proof-of-concept validation of a portable and user-friendly microfluidic flow cytometer (

Cardiac telomere length in heart development, function, and disease

- Booth, Scott, Charchar, Fadi

Authors: Booth, Scott , Charchar, Fadi
Date: 2017
Type: Text , Journal article , Review
Relation: Physiological Genomics Vol. 49, no. 7 (2017), p. 368-384
Relation: http://purl.org/au-research/grants/nhmrc/1034371
Full Text: false
Reviewed:
Description: Telomeres are repetitive nucleoprotein structures at chromosome ends, and a decrease in the number of these repeats, known as a reduction in telomere length (TL), triggers cellular senescence and apoptosis. Heart disease, the worldwide leading cause of death, often results from the loss of cardiac cells, which could be explained by decreases in TL. Due to the cell-specific regulation of TL, this review focuses on studies that have measured telomeres in heart cells and critically assesses the relationship between cardiac TL and heart function. There are several lines of evidence that have identified rapid changes in cardiac TL during the onset and progression of heart disease as well as at critical stages of development. There are also many factors, such as the loss of telomeric proteins, oxidative stress, and hypoxia, that decrease cardiac TL and heart function. In contrast, antioxidants, calorie restriction, and exercise can prevent both cardiac telomere attrition and the progression of heart disease. TL in the heart is also indicative of proliferative potential and could facilitate the identification of cells suitable for cardiac rejuvenation. Although these findings highlight the involvement of TL in heart function, there are important questions regarding the validity of animal models, as well as several confounding factors, that need to be considered when interpreting results and planning future research. With these in mind, elucidating the telomeric mechanisms involved in heart development and the transition to disease holds promise to prevent cardiac dysfunction and potentiate regeneration after injury. © 2017 the American Physiological Society.

Quick View

A meta-analysis of gene expression signatures of blood pressure and hypertension

- Charchar, Fadi

Authors: Charchar, Fadi
Date: 2015
Type: Text , Journal article
Relation: PLoS Genetics Vol. 11, no. 3 (2015), p. 1-29
Full Text:
Reviewed:
Description: Genome-wide association studies (GWAS) have uncovered numerous genetic variants (SNPs) that are associated with blood pressure (BP). Genetic variants may lead to BP changes by acting on intermediate molecular phenotypes such as coded protein sequence or gene expression, which in turn affect BP variability. Therefore, characterizing genes whose expression is associated with BP may reveal cellular processes involved in BP regulation and uncover how transcripts mediate genetic and environmental effects on BP variability. A meta-analysis of results from six studies of global gene expression profiles of BP and hypertension in whole blood was performed in 7017 individuals who were not receiving antihypertensive drug treatment. We identified 34 genes that were differentially expressed in relation to BP (Bonferroni-corrected p<0.05). Among these genes, FOS and PTGS2 have been previously reported to be involved in BP-related processes; the others are novel. The top BP signature genes in aggregate explain 5%–9% of inter-individual variance in BP. Of note, rs3184504 in SH2B3, which was also reported in GWAS to be associated with BP, was found to be a trans regulator of the expression of 6 of the transcripts we found to be associated with BP (FOS, MYADM, PP1R15A, TAGAP, S100A10, and FGBP2). Gene set enrichment analysis suggested that the BP-related global gene expression changes include genes involved in inflammatory response and apoptosis pathways. Our study provides new insights into molecular mechanisms underlying BP regulation, and suggests novel transcriptomic markers for the treatment and prevention of hypertension. **Please note that there are multiple authors for this article therefore only the name of the Federation University Australia affiliate is provided in this record**

MicroRNAs mediate the cardioprotective effect of angiotensin-converting enzyme inhibition in acute kidney injury

- Rana, Indrajeetsinh, Velkoska, Elena, Patel, Sheila, Burrell, Louise, Charchar, Fadi

Authors: Rana, Indrajeetsinh , Velkoska, Elena , Patel, Sheila , Burrell, Louise , Charchar, Fadi
Date: 2015
Type: Text , Journal article
Relation: American Journal of Physiology - Renal Physiology Vol. 309, no. 11 (2015), p. F943-F954
Full Text: false
Reviewed:
Description: Cardiovascular disease, including cardiac hypertrophy, is common in patients with kidney disease and can be partially attenuated using blockers of the renin-angiotensin system (RAS). It is unknown whether cardiac microRNAs contribute to the pathogenesis of cardiac hypertrophy or to the protective effect of RAS blockade in kidney disease. Using a subtotal nephrectomy rat model of kidney injury, we investigated changes in cardiac microRNAs that are known to have direct target genes involved in the regulation of apoptosis, fibrosis, and hypertrophy. The effect of treatment with the angiotensin-converting enzyme (ACE) inhibitor ramipril on cardiac microRNAs was also investigated. Kidney injury led to a significant increase in cardiac microRNA-212 and mi- croRNA-132 expression. Ramipril reduced cardiac hypertrophy, attenuated the increase in microRNA-212 and microRNA-132, and significantly increased microRNA-133 and microRNA-1 expression. There was altered expression of caspase-9, B cell lymphoma-2, transforming growth factor-Î², fibronectin 1, collagen type 1A1, and forkhead box protein O3, which are all known to be involved in the regulation of apoptosis, fibrosis, and hypertrophy in cardiac cells while being targets for the above microRNAs. ACE inhibitor treatment increased expression of microRNA-133 and microRNA-1. The inhibitory action of ACE inhibitor treatment on increased cardiac NADPH oxidase isoform 1 expression after subtotal nephrectomy surgery suggests that inhibition of oxidative stress is also one of mechanism of ACE inhibitor-mediated cardioprotection. These finding suggests the involvement of microRNAs in the cardioprotective action of ACE inhibition in acute renal injury, which is mediated through an inhibitory action on profibrotic and proapoptotic target genes and stimulatory action on antihypertrophic and antiapoptotic target genes. © 2015 the American Physiological Society. Funding: APP1048285; NHMRC; National Health and Medical Research Council

Quick View

Characterisation of apoptosis in thye1M6

- Hay, Stewart

Authors: Hay, Stewart
Date: 2003
Type: Text , Thesis , PhD
Full Text:
Description: "Interest surrounds apoptosis-inducing factors for use in treatment of proliferative disorders and for the elucidation of cell death pathways. Here, a murine SCID thymic lymphoma cell line (ThyE1M6) is described which produces a cytotoxic factor that acts on specific human and murine cancer cell lines, including some lymphomas and hepatomas. The cytotoxic property of the lymphoma was first noted in co-culture experiments that revealed increased apoptosis in some cell types. This attribute was thought to be unusual and as a result a project was initiated with the aim to characterise the factor responsible for cell death."
Description: Doctor of Philosophy

Quick View

Australian ethnomedicinal plant extracts promote apoptosis-mediated cell death In human hepatocellular carcinoma in vitro

- Kiran, Sudha, Chew, Guat Siew, Johnson, Joel Benjamin, Mani, Janice Sandra, Wakeling, Lara, Portman, Andrew, Naiker, Mani

Authors: Kiran, Sudha , Chew, Guat Siew , Johnson, Joel Benjamin , Mani, Janice Sandra , Wakeling, Lara , Portman, Andrew , Naiker, Mani
Date: 2021
Type: Text , Journal article
Relation: Pharmacognosy communications Vol. 11, no. 4 (2021), p. 210-213
Full Text:
Reviewed:
Description: Introduction: Hepatocellular carcinoma (HCC) Is the leading cause of primary liver cancer with Its prevalence continuing to rise. Although the number of cases continues to rise In both developing and developed countries, prognosis remains poor due to a lack of successful treatments. Inspired by the prospect of developing complementary medicines for this condition, we explore several native Australian plants for anti-carcinogenic activity, especially against HCC. Methods: Cytotoxicity assays against HCC cell lines were conducted using various plant extracts. Biochemical profiling of the plant species was conducted for total phenolics and antioxidant capacity, while reverse transcription-polymerase chain reaction (RT-PCR) was used to determine the active apoptotic pathways. Results: Westringia fruticosa and Prostanthera ovalifolia (small-leaved variety) had high antioxidant (410 and 227 mg/g, respectively) and phenolic contents (72.7 and 42.7 mg/g, respectively). P ovalifolia (small-leaved variety) demonstrated the greatest cytotoxic activity against HepG2 cells (IC50 4.61 ± 0.98 pg/mL) followed by Solanum laciniatum leaves (11.79 ± 0.43 pg/mL) and fruit extracts (ripe, unripe) (14.85 ± 1.80 and 19 ± 1.32 pg/mL, respectively). RT-PCR results confirmed apoptotic events in HepG2 cells, exposed to ripe and unripe S. laciniatum fruit extracts, via caspase-3 pathway. The highest apoptotic induction occurred after 8 hr. Compared to unripe fruits, ripe fruits induced a greater level of apoptosis, as evidenced by a 73 % higher level of caspase-3 mRNA expression and 22 % lower IC50 value. Conclusion: With further investigation, these fruits may provide a valuable source of anticarcinogenic compounds for use as chemotherapeutic or complementary therapies.

Showing items 1 - 8 of 8