Multicolour flow cytometry analyses and autofluorescence in chlorophytes: lessons from programmed cell death studies in Chlamydomonas reinhardtii

Apoptosis is defined as a highly regulated form of programmed cell death with typical morphological and biochemical features. A variety of factors, including heavy metals, may influence the intensity of programmed cell death. The aim of the work was to simulate apoptosis in an in vitrosystem under the conditions of stable strontium exposure. The children’s population consuming drinking water with high strontium (Sr2+) content (n = 49) was observed. The level of lymphocyte apoptosis was determined with flow cytometry technique, by means of labeled annexin V-FITC conjugate (AnnV-FITC) and propidium iodide (PI) staining. AnnV-FITC+PI- cells were regarded as early apoptotic forms, whereas late apoptotic and/or necrotic cells were AnnV-FITC+PI+. The isolated leukocytes were incubated with Sr2+ at a concentration of 7.0 mg/l, the maximal permitted concentration (MPC) for water of aqueous objects, for 4 hours at 37 ºC. Expression of CD95 and p53 apoptosis markers was performed by flow cytometry using labeled monoclonal antibodies.In vitroexposure to strontium was associated with significantly decreased expression of apoptosisregulating factors, i.e., membrane marker CD95 and intracellular transcription protein p53, 1.56- and 1.68-fold, respectively. Meanwhile, we revealed a significantly (4.68-fold) decreased amounts of AnnV-FITC+PI--cells, as well as a statistically significant (1.35-fold) increase of the AnnV-FITC+PI+-cells. Moreover, the amounts of AnnV-FITC+ PI--lymphocytes in all samples were below the physiological ranges and control values. The number of samples with higher contents of AnnV-FITC+PI+-lymphocyte exceeding the established standards and control values, was 30.8%. Thus, it has been experimentally proven that strontium, at a concentration corresponding to MPC for water objects may significantly inhibit cell death along apoptotic pathways, with switching to necrotic cell death mechanisms, according to phosphatidylserine contents, as detected by annexin V binding test. The data have revealed an ability of strontium to have a significant effect upon the parameters of regulation and maintenance of cellular homeostasis, by influencing the apoptosis intensity, due to shifting a balance towards necrosis and reducing expression of apoptosis-regulating factors. The results of this study may be used in order to identify some marker indexes of immune disorders potentially induced by external influence of strontium upon human health under specific environmental factors.

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Comparative evaluation of the effectiveness of methods for certification of transplanted cell lines

Veterinaria i kormlenie ◽

10.30917/att-vk-1814-9588-2021-1-9 ◽

2021 ◽

Author(s):

E.V. Markova ◽

V.T. Nochevny ◽

B.L. Manin ◽

I.N. Matveeva

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cell Death ◽

Programmed Cell Death ◽

Cell Lines ◽

Traditional Method ◽

Biological Properties ◽

International Standards ◽

Growth Potential ◽

Wide Range

The article presents the results of certification of two trofovariants of MDVK cell lines with the help of traditional method and flow cytometry. Research object was the test cultures MDBK-E and MDBK-B, which passed 30 and 43 passages, respectively, after cryopreservation. The traditional method of attestation of transplanted cell lines, widely used in practice, is rather laborious and requires significant expenditures of labor, money and time. The flow cytometry method is based on a wide range of cytochemical and fluorescent methods for the analysis of sizes, granularity, phases of the cell cycle, structural components (DNA, RNA, protein), cell apoptosis and a number of other indicators. It was experimentally established that the sublines of MDBK-E and MDBK-B cells differed in cultural, cytomorphological and karyological parameters, as well as in contamination by foreign agents and sensitivity to parainfluenza-3 viruses and infectious rhinotracheitis in cattle. Analysis of histograms of cell distribution depending on the DNA content showed that the studied lines MDBK-E and MDBK-B did not exceed the standard indicator in terms of apoptosis and were at the level of 3,9 and 6,8%, respectively. Cells of the MDBK-E line did not contain viral and mycoplasma contamination, were characterized by a pronounced growth potential, retained the original cell morphology and were the most promising substrate for the production of antigens of parainfluenza-3, infectious rhinotracheitis in cattle. Analysis of granularity distribution results testified to the violation of the division processes and the appearance in the population of the subline MDBK-B of abnormal cells, as well as inadequate conditions for maintaining the test culture. It has been established that the flow cytometry method is objective and quite promising in the selection of culture models that meet the requirements of domestic and international standards. The revealed correlation between the magnitude of apoptosis, cultural properties and parameters of the cell cycle makes it possible to assess the biological properties of the producer culture as one of the leading factors in the change in programmed cell death. Changes the index of programmed cell death underlies a number of important pathological conditions and degenerative processes.

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Temporal and metabolic overlap between lipid accumulation and programmed cell death due to nitrogen starvation in the unicellular chlorophyte Chlamydomonas reinhardtii

Phycological Research ◽

10.1111/pre.12368 ◽

2019 ◽

Vol 67 (3) ◽

pp. 173-183 ◽

Cited By ~ 4

Author(s):

Santosh Sathe ◽

Mónica V. Orellana ◽

Nitin S. Baliga ◽

Pierre M. Durand

Keyword(s):

Cell Death ◽

Programmed Cell Death ◽

Chlamydomonas Reinhardtii ◽

Lipid Accumulation ◽

Nitrogen Starvation

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KCl induces a caspase-independent programmed cell death in the unicellular green chlorophyte Chlamydomonas reinhardtii (Chlorophyceae)

Phycologia ◽

10.2216/15-103.1 ◽

2016 ◽

Vol 55 (4) ◽

pp. 378-392 ◽

Cited By ~ 7

Author(s):

Sirisha L. Vavilala ◽

Mahuya Sinha ◽

Kanak K. Gawde ◽

Seema M. Shirolikar ◽

Jacinta S. D'Souza

Keyword(s):

Cell Death ◽

Programmed Cell Death ◽

Chlamydomonas Reinhardtii

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Acetic acid-induced programmed cell death and release of volatile organic compounds in Chlamydomonas reinhardtii

Plant Physiology and Biochemistry ◽

10.1016/j.plaphy.2011.11.003 ◽

2012 ◽

Vol 51 ◽

pp. 175-184 ◽

Cited By ~ 51

Author(s):

Zhaojiang Zuo ◽

Yerong Zhu ◽

Yanling Bai ◽

Yong Wang

Keyword(s):

Cell Death ◽

Acetic Acid ◽

Volatile Organic Compounds ◽

Programmed Cell Death ◽

Chlamydomonas Reinhardtii ◽

Organic Compounds ◽

Volatile Organic

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Features of lymphocytes during programmed cell death in patients with atopic bronchial asthma

Rossiiskii Allergologicheskii ZHurnal ◽

10.36691/rja870 ◽

2009 ◽

Vol 7 (4) ◽

pp. 32-39

Author(s):

Y V Pinchuk ◽

A C Vodunon ◽

I G Mustafin ◽

Z I Abramova

Keyword(s):

Flow Cytometry ◽

Cell Death ◽

Programmed Cell Death ◽

Morphological Difference ◽

Disease Severity Score ◽

Electronic Microscopy ◽

Disease Pathogenesis ◽

Atopic Bronchial Asthma ◽

The Difference ◽

Number Of Cells

Background. To establish features of programmed cell death of lymphocytes depending on disease severity score. Methods. The morphology of lymphocytes was investigated with a method of electronic microscopy. Apoptotic cells were defined by flow cytometry. Results. We have revealed lymphocytes morphological difference between asthmatics and normal donors. Also we found out the difference between number of cells during the incubation process. Conclusion. These findings can promote deeper understanding of the disease pathogenesis.

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Programmed death in a unicellular organism has species-specific fitness effects

Biology Letters ◽

10.1098/rsbl.2013.1088 ◽

2014 ◽

Vol 10 (2) ◽

pp. 20131088 ◽

Cited By ~ 16

Author(s):

Pierre M. Durand ◽

Rajdeep Choudhury ◽

Armin Rashidi ◽

Richard E. Michod

Keyword(s):

Cell Death ◽

Programmed Cell Death ◽

Chlamydomonas Reinhardtii ◽

Genetic Relatedness ◽

Model Organism ◽

Inhibitory Effect ◽

Unicellular Organism ◽

Fitness Effects ◽

Programmed Death ◽

Species Specific

Programmed cell death (PCD) is an ancient phenomenon and its origin and maintenance in unicellular life is unclear. We report that programmed death provides differential fitness effects that are species specific in the model organism Chlamydomonas reinhardtii . Remarkably, PCD in this organism not only benefits others of the same species, but also has an inhibitory effect on the growth of other species. These data reveal that the fitness effects of PCD can depend upon genetic relatedness.

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