Tetrahymena dimorpha sp.nov. (Hymenostomatida: Tetrahymenidae), a new ciliate parasite of Simuliidae (Diptera) with potential as a model for the study of ciliate morphogenesis

1983 ◽  
Vol 301 (1106) ◽  
pp. 345-363 ◽  
Keyword(s):  
Limited Range ◽  
Culture Conditions ◽  
Morphological Transformation ◽  
Sterile Culture ◽  
Proteose Peptone ◽  
Wide Range ◽  
Yeast Extract Medium ◽  
Insect Tissue Culture ◽  
Heavy Burden

A new species of hymenostome ciliate, Tetrahymena dimorpha sp.nov., is described. This ciliate occurs as a parasite in the haemocoel of larval, pupal and adult Simuliidae (Diptera). In larval hosts the total number of parasites never exceeds about 240 and the infection is benign. Within larval hosts the ciliates are large and broadly oval and possess an unusually wide range of somatic kineties, from 30 to 66; moreover a variable proportion of these kineties are characteristically disorganized, being incomplete, meandering or branched. Metamorphosis of the host to the adult fly is accompanied by a dramatic increase in the number of ciliates, which reach pathogenic intensity. Adult hosts may contain up to 19000 ciliates and the flies soon die from this heavy burden. Associated with ciliate population growth during host metamorphosis is a startling morphological transformation of the ciliates themselves. In adult hosts the ciliates are smaller and pyriform in shape and the cortex is greatly modified; the total number of somatic kineties is considerably reduced and has a limited range of 19—22. Most significantly, the kineties are ordered with typical tetrahymenine precision. By application of appropriate culture conditions to ciliates isolated from any host stage, either of the distinctive morphological forms of T. dimorpha may be selectively induced in vitro . In bacterized infusions, ciliates are produced that have the general form and cortical characteristics of those found naturally in adult hosts. Sterile culture in serum-supplemented Mitsuhashi and Maramorosch insect tissue culture medium produces a population showing features characteristic of ciliates from larval hosts. Sterile culture in proteose-peptone-yeast-extract medium results in populations exhibiting concurrent dimorphism, even after cloning. The extreme nature and multiple facets of the dimorphism together with the ease of its manipulation in vitro afford opportunities for the experimental investigation of many problems, particularly those related to cell surface patterning in ciliates, and these possibilities are discussed in relation to current concepts of ciliate morphogenesis.

scholarly journals Activation of the receptor tyrosine kinase RET improves long-term hematopoietic stem cell outgrowth and potency

Blood ◽  
2020 ◽  
Vol 136 (22) ◽  
pp. 2535-2547 ◽  
Author(s):  
W. Grey ◽  
R. Chauhan ◽  
M. Piganeau ◽  
H. Huerga Encabo ◽  
M. Garcia-Albornoz ◽  
...  
Keyword(s):  
Intracellular Signaling ◽  
Culture Conditions ◽  
Cellular Growth ◽  
Great Promise ◽  
Bone Marrow Niche ◽  
Hematopoietic Stem ◽  
Intracellular Signaling Pathway ◽  
Wide Range

Abstract Expansion of human hematopoietic stem cells (HSCs) is a rapidly advancing field showing great promise for clinical applications. Recent evidence has implicated the nervous system and glial family ligands (GFLs) as potential drivers of hematopoietic survival and self-renewal in the bone marrow niche; how to apply this process to HSC maintenance and expansion has yet to be explored. We show a role for the GFL receptor, RET, at the cell surface of HSCs in mediating sustained cellular growth, resistance to stress, and improved cell survival throughout in vitro expansion. HSCs treated with the key RET ligand/coreceptor complex, glial-derived neurotrophic factor and its coreceptor, exhibit improved progenitor function at primary transplantation and improved long-term HSC function at secondary transplantation. Finally, we show that RET drives a multifaceted intracellular signaling pathway, including key signaling intermediates protein kinase B, extracellular signal-regulated kinase 1/2, NF-κB, and p53, responsible for a wide range of cellular and genetic responses that improve cell growth and survival under culture conditions.

scholarly journals Quantitative analysis of tumour spheroid structure

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Alexander P Browning ◽  
Jesse A Sharp ◽  
Ryan J Murphy ◽  
Gency Gunasingh ◽  
Brodie Lawson ◽  
...  
Keyword(s):  
Cell Lines ◽  
Tumour Microenvironment ◽  
Culture Conditions ◽  
Experimental Models ◽  
Seeding Density ◽  
Modelling Framework ◽  
Tumour Spheroid ◽  
Tumour Spheroids ◽  
Wide Range

Tumour spheroids are common in vitro experimental models of avascular tumour growth. Compared with traditional two-dimensional culture, tumour spheroids more closely mimic the avascular tumour microenvironment where spatial differences in nutrient availability strongly influence growth. We show that spheroids initiated using significantly different numbers of cells grow to similar limiting sizes, suggesting that avascular tumours have a limiting structure; in agreement with untested predictions of classical mathematical models of tumour spheroids. We develop a novel mathematical and statistical framework to study the structure of tumour spheroids seeded from cells transduced with fluorescent cell cycle indicators, enabling us to discriminate between arrested and cycling cells and identify an arrested region. Our analysis shows that transient spheroid structure is independent of initial spheroid size, and the limiting structure can be independent of seeding density. Standard experimental protocols compare spheroid size as a function of time; however, our analysis suggests that comparing spheroid structure as a function of overall size produces results that are relatively insensitive to variability in spheroid size. Our experimental observations are made using two melanoma cell lines, but our modelling framework applies across a wide range of spheroid culture conditions and cell lines.

scholarly journals Verification of the Field Productivity of Rehmannia glutinosa (Gaertn.) DC. Developed Through Optimized In Vitro Culture Method

Plants ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 317
Author(s):  
Yong-Goo Kim ◽  
Richard Komakech ◽  
Dae Hui Jeong ◽  
Yun mi Park ◽  
Tae Kyoung Lee ◽  
...  
Keyword(s):  
Near Infrared ◽  
Chemical Constituents ◽  
Culture Method ◽  
Poor Quality ◽  
Culture Conditions ◽  
Rehmannia Glutinosa ◽  
Sterile Culture ◽  
Perennial Plant ◽  
The Family

Rehmannia glutinosa (Gaertn.) DC is a perennial plant belonging to the family Scropulariidae. The root of R. glutinosa is used in oriental medicine and mainly grown using rootstock rather than seed cultivation, which gives rise to several problems including root rot, and results in a low productivity and poor quality. To solve the challenges involved in R. glutinosa seed cultivation, our team previously used the formative features and genetic analysis of R. glutinosa to determine the optimal in vitro tissue culture conditions for producing sterile culture seedlings and rootstocks of R. glutinosa. The aim of the present study was to identify differences between R. glutinosa standard rootstock seedlings (SR), R. glutinosa culture rootstock seedlings (CR), and culture seedlings (CS) under field conditions. The reproductive characteristics of the aerial part were more robust while the area and length of leaves were smaller for SR than those for CR and CS. The characteristic that differed the most in SR was flowering, which did not occur in CR and CS. In addition, the fresh and dry weights of the subterranean parts of CR and CS were two-fold greater than those of SR. Fourier transform near-infrared (FT-NIR) analysis showed only slight differences between the chemical constituents of SR and its culture products, which was confirmed by measuring the content of catalpol, an indexing substance. Catalpol had a reduced content in the culture products compared to SR. However, this difference was not significant. Our findings will be useful for the identification of the best seedling type of R. glutinosa to enable its mass production.

scholarly journals Using Cell Cultures for the Investigation of Treatments for Attention Deficit Hyperactivity Disorder: A Systematic Review

2019 ◽  
Vol 17 (10) ◽  
pp. 916-925
Author(s):  
Danielly Chierrito ◽  
Camila B. Villas-Boas ◽  
Fernanda S. Tonin ◽  
Fernando Fernandez-Llimos ◽  
Andréia C.C. Sanches ◽  
...  
Keyword(s):  
Attention Deficit Hyperactivity Disorder ◽  
Cell Cultures ◽  
Attention Deficit ◽  
Experimental Studies ◽  
Drug Efficacy ◽  
Culture Conditions ◽  
Qualitative Synthesis ◽  
Hyperactivity Disorder ◽  
Wide Range

Background: Advances in basic and molecular biology have promoted the use of cell cultures in a wide range of areas, including the evaluation of drug efficacy, safety and toxicity. Objective: This article aims to provide a general overview of the methodological parameters of cell cultures used to investigate therapeutic options for Attention Deficit Hyperactivity Disorder. Methods: A systematic search was performed in the electronic databases PubMed, Scopus, and DOAJ. In vitro experimental studies using cell cultures were included. Results: A total of 328 studies were initially identified, with 16 included for qualitative synthesis. Seven studies used neuronal cells (SH-SY5Y neuroblastoma and PC12 cell line) and nine used nonneuronal cells. All the studies described the culture conditions, but most studies were inconsistent with regard to reporting results and raw data. Only one-third of the studies performed cell viability assays, while a further 30% conducted gene expression analysis. Other additional tests included electrophysiological evaluation and transporter activity. More than 50% of the studies evaluated the effects of drugs such as methylphenidate and atomoxetine, while plant extracts were assessed in four studies and polyunsaturated fatty acids in one. Conclusion: We suggested a flowchart to guide the planning and execution of studies, and a checklist to be completed by authors to allow the standardized reporting of results. This may guide the elaboration of laboratory protocols and further in vitro studies.

scholarly journals Development of an in vitro tissue culture system for hammer coral (Fimbriaphyllia ancora) ovaries

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yi-Ling Chiu ◽  
Ching-Fong Chang ◽  
Shinya Shikina
Keyword(s):  
Structural Integrity ◽  
Culture Method ◽  
Culture Conditions ◽  
Short Term ◽  
Wide Range ◽  
Fetal Bovine ◽  
Short Term Culture ◽  
Mesenterial Filaments

AbstractIn vitro gonad culture systems have proven useful to investigate intrinsic mechanisms of sexual reproduction in animals. Here we describe development of an in vitro culture method for coral ovaries. Mesenterial tissues containing both ovaries and mesenterial filaments were microscopically isolated from the scleractinian coral, Fimbriaphyllia ancora, and culture conditions were optimized. M199 diluted 10× (10% M199, pH 8.1) and supplemented with 25 mM HEPES and the antibiotics, ampicillin, penicillin and streptomycin, supported oocyte survival and maintained the structural integrity of ovaries during short-term culture (~ 6 days). Addition of a commercial antibiotic–antimycotic solution (Anti–Anti) and fetal bovine serum adversely affected ovary maintenance and caused tissue disintegration. Characterization of cultured ovaries showed that there is no difference in cell proliferation of ovarian somatic cells between culture Days 1 and 6. Moreover, the presence of oogonia and expression of a major yolk protein, vitellogenin, were confirmed in ovaries cultured for 6 days. This system will be useful for studying effects of a wide range of substances on coral oogenesis.

scholarly journals BrainPhys neuronal medium optimized for imaging and optogenetics in vitro

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Michael Zabolocki ◽  
Kasandra McCormack ◽  
Mark van den Hurk ◽  
Bridget Milky ◽  
Andrew P. Shoubridge ◽  
...  
Keyword(s):  
Cell Biology ◽  
Culture Conditions ◽  
Cellular Reprogramming ◽  
Synaptic Activity ◽  
Neuronal Cultures ◽  
Neuronal Viability ◽  
Human Neurons ◽  
Wide Range ◽  
And Function

AbstractThe capabilities of imaging technologies, fluorescent sensors, and optogenetics tools for cell biology are advancing. In parallel, cellular reprogramming and organoid engineering are expanding the use of human neuronal models in vitro. This creates an increasing need for tissue culture conditions better adapted to live-cell imaging. Here, we identify multiple caveats of traditional media when used for live imaging and functional assays on neuronal cultures (i.e., suboptimal fluorescence signals, phototoxicity, and unphysiological neuronal activity). To overcome these issues, we develop a neuromedium called BrainPhys™ Imaging (BPI) in which we optimize the concentrations of fluorescent and phototoxic compounds. BPI is based on the formulation of the original BrainPhys medium. We benchmark available neuronal media and show that BPI enhances fluorescence signals, reduces phototoxicity and optimally supports the electrical and synaptic activity of neurons in culture. We also show the superior capacity of BPI for optogenetics and calcium imaging of human neurons. Altogether, our study shows that BPI improves the quality of a wide range of fluorescence imaging applications with live neurons in vitro while supporting optimal neuronal viability and function.

scholarly journals Effect of Cytokinins on Micropropagation of Moringa Oleifera Lam. and Genomic Stability Assessment Using Flow-Cytometry

2022 ◽  
Author(s):  
Rohit Bharati ◽  
Moses Okao ◽  
Katerina Hamouzová ◽  
Eloy Fernandez-Cusimamani
Keyword(s):  
Flow Cytometry ◽  
Moringa Oleifera ◽  
Genetic Fidelity ◽  
Limited Range ◽  
Surface Sterilization ◽  
Wide Range ◽  
The Family ◽  
Livestock Feeding ◽  
Planting Materials

Abstract Moringa oleifera Lam. is a multipurpose medicinal plant of the family Moringaceae which has been widely utilized as a pharmaceutical remedy to treat a wide range of diseases. In addition, the tree has several applications in human nutrition as well as livestock feeding. M. oleifera is easily multiplied through epigeal germination (recalcitrant) but seed propagated plants are heterogeneous and take longer to reach fruit-bearing age. As an alternative, branch cuttings have been used but their establishment is erratic and often leads to reduced growth of the mother plant. Thus, to produce superior planting materials, in-vitro propagation has become paramount. As a result, several studies using a limited range of cytokinin have been undertaken to multiply M. oleifera through tissue culture. Otherwise, a study was conducted to examine the effect of five different cytokinins on in-vitro regeneration of this tree species. Results showed that nodal explants cultured on Murashige and Skoog (MS) medium supplemented with 1.0 mg/L 6-Benzylaminopurine (BAP) and subsequently rooted on basic MS media was the most optimal treatment. Furthermore, acclimatization of plantlets in sterile soil substrate and perlite (1:3;v/v) under transparent polythene sheet for 7 days resulted in survival rate of 100%. Assessment of genetic fidelity using flow cytometry revealed that surface sterilization alongside cytokinin treatments produced plantlets that were genetically stable regardless of the growth regulator used. Thus, the in-vitro protocol developed in this study can be utilized for in-vitro studies and mass propagation of this imperative plant species.

scholarly journals Quantitative analysis of tumour spheroid structure

2021 ◽  
Author(s):  
Alexander P Browning ◽  
Jesse A Sharp ◽  
Ryan J Murphy ◽  
Gency Gunasingh ◽  
Brodie Lawson ◽  
...  
Keyword(s):  
Cell Lines ◽  
Tumour Microenvironment ◽  
Culture Conditions ◽  
Experimental Models ◽  
Seeding Density ◽  
Modelling Framework ◽  
Tumour Spheroid ◽  
Tumour Spheroids ◽  
Wide Range

Tumour spheroids are common in vitro experimental models of avascular tumour growth. Compared with traditional two-dimensional culture, tumour spheroids more closely mimic the avascular tumour microenvironment where spatial differences in nutrient availability strongly influence growth. We show that spheroids initiated using significantly different numbers of cells grow to similar limiting sizes, suggesting that avascular tumours have a limiting structure; in agreement with untested predictions of classical mathematical models of tumour spheroids. We develop a novel mathematical and statistical framework to study the structure of tumour spheroids seeded from cells transduced with fluorescent cell cycle indicators, enabling us to discriminate between arrested and cycling cells and identify an arrested region. Our analysis shows that transient spheroid structure is independent of initial spheroid size, and the limiting structure can be independent of seeding density. Standard experimental protocols compare spheroid size as a function of time; however, our analysis suggests that comparing spheroid structure as a function of overall size produces results that are relatively insensitive to variability in spheroid size. Our experimental observations are made using two melanoma cell lines, but our modelling framework applies across a wide range of spheroid culture conditions and cell lines.

scholarly journals Failure of in vitro differentiation of Plasmodium falciparum gametocytes into ookinetes arises because of poor gamete fertilisation

2017 ◽  
Author(s):  
Michael J Delves ◽  
Sara R Marques ◽  
Andrea Ruecker ◽  
Ursula Straschil ◽  
Celia Miguel-Blanco ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Culture Conditions ◽  
Transmission Cycle ◽  
Potential Barriers ◽  
Wide Range ◽  
Species Specific ◽  
Transmission Biology ◽  
Avian Malaria Parasites

ABSTRACTA critical step towards malaria elimination will be the interruption of Plasmodium transmission from the human host to the mosquito. At the core of the transmission cycle lies Plasmodium sexual reproduction leading to zygote formation and mosquito midgut colonisation by ookinetes. Whilst in vitro ookinete culture from the murine and avian malaria parasites, Plasmodium berghei and P. gallinaceum, has greatly increased our knowledge of transmission biology; efforts to mimic the process in the human parasite P. falciparum have, to date, had only limited success. Using fluorescence microscopy and flow cytometry with antibodies specific to the male gametocyte and developing ookinetes, we sought to evaluate P. falciparum ookinete production using previously published in vitro protocols. We then compared in vitro versus in vivo ookinete production in both P. falciparum and P. berghei parasites, exploring potential barriers to complete development. Finally, we sought to test a wide range of literature-led culture conditions towards further optimisation of in vitro P. falciparum ookinete production. Despite extensive testing, our efforts to replicate published methods did not produce appreciable quantities of fully formed P. falciparum ookinetes in vitro. In parallel, however, gametocyte cultures that failed to differentiate fully in vitro successfully developed into ookinetes in vivo with an efficiency approximating that of P. berghei. Flow cytometry analysis showed that this disparity likely lies with the poor fertilization of P. falciparum gametes in vitro. Attempts to improve gametocyte fertility or define conditions more permissive to fertilisation/ookinete survival in vitro were also unsuccessful. Current in vitro conditions for P. falciparum ookinete production are not optimal for gamete fertilisation either due to the lack of parasite-species-specific mosquito factors missing from in vitro culture, or non-permissive cues contaminating culture preparations.

scholarly journals In VitroCulture Conditions for Maintaining a Complex Population of Human Gastrointestinal Tract Microbiota

2011 ◽  
Vol 2011 ◽  
pp. 1-10 ◽  
Author(s):  
Bong-Soo Kim ◽  
Jong Nam Kim ◽  
Carl E. Cerniglia
Keyword(s):  
Gastrointestinal Tract ◽  
Bacterial Community ◽  
Intestinal Microbiota ◽  
Brain Heart Infusion ◽  
Culture Conditions ◽  
Human Gastrointestinal Tract ◽  
Gastrointestinal Microbiota ◽  
Wide Range ◽  
The Impact

A stable intestinal microbiota is important in maintaining human physiology and health. Although there have been a number of studies usingin vitroandin vivoapproaches to determine the impact of diet and xenobiotics on intestinal microbiota, there is no consensus for the bestin vitroculture conditions for growth of the human gastrointestinal microbiota. To investigate the dynamics and activities of intestinal microbiota, it is important for the culture conditions to support the growth of a wide range of intestinal bacteria and maintain a complex microbial community representative of the human gastrointestinal tract. Here, we compared the bacterial community in three culture media: brain heart infusion broth and high- and low-carbohydrate medium with different growth supplements. The bacterial community was analyzed using denaturing gradient gel electrophoresis (DGGE), pyrosequencing and real-time PCR. Based on the molecular analysis, this study indicated that the 3% fecal inoculum in low-concentration carbohydrate medium with 1% autoclaved fecal supernatant provided enhanced growth conditions to conductin vitrostudies representative of the human intestinal microbiota.

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