scholarly journals Explant-Like Passaging of Cells Growing on Portable Substrata Permits the Avoidance of Enzyme Application and Facilitates the Passage Procedure

2019 ◽  
Vol 67 (4) ◽  
pp. 169-176
Author(s):  
Damian RYSZAWY ◽  
Justyna PODULKA ◽  
Anna TWORZYDŁO ◽  
Justyna TOMASSY ◽  
Michał JARMULSKI ◽  
...  
Keyword(s):  
Cell Culture ◽  
Fluorescence Microscopy ◽  
Cell Viability ◽  
Glass Fiber ◽  
Proteolytic Enzymes ◽  
Culture Vessel ◽  
Cell Rounding ◽  
Tissue Explants ◽  
Numerous Cell ◽  
Enzyme Application

The experiments presented in this paper show how growing cells on portable substrats can be useful to facilitate and accelerate the passaging (subculture) of anchorage-dependent cells. Experiments have shown that portable substrats are cheap, commercially available, and transparent. They are easily cut into various shapes and sizes, and are easy to sterilize. Portable substrats are also friendly to cells and permit faster than usual cell passaging procedures. Anchorage-dependent cells growing on the bottom of a culture vessel made of glass or polystyrene can be quickly passaged with previously-cut small fragments of glass fiber or nylon meshes or small fragments of polyester foil as well as nylon fishing lines and biodegradable surgeon threads that have been inserted into the vessel. The surfaces of such fragments of portable supports are quickly overgrown with cells and can be easily transferred to a new culture vessel. As with tissue explants, cells migrate and grow over the bottom of the new culture vessels. Using cell viability tests, analyses of proliferation and fluorescence microscopy, we confirmed the utility of the investigated substrats for cell culture. In addition, the passaging cells, together with a portable support (like explants), eliminate the need for an application of proteolytic enzymes which modify numerous cell properties and activities and would keep the cell from detaching from the substratum which would lead to the cell rounding and changes in the cell's cytoskeleton architecture.

IN VITRO METHODS FOR THE STUDY OF THE ADENOHYPOPHYSIAL FUNCTIONS TO SECRETE GONADOTROPHIN

1971 ◽  
Vol 68 (1_Suppl) ◽  
pp. S27-S40 ◽  
Author(s):  
T. Kobayashi ◽  
T. Kigawa ◽  
M. Mizuno ◽  
T. Watanabe
Keyword(s):  
Cell Culture ◽  
Organ Culture ◽  
Cultured Cells ◽  
Cell Sheet ◽  
Short Term ◽  
Hypothalamic Extract ◽  
Numerous Cell ◽  
Single Cell Culture ◽  
Almost All

ABSTRACT There are several in vitro methods to analyse the function of the adenohypophysis or the mechanisms of its regulation. The present paper deals with single cell culture, organ culture and short term incubation techniques by which the morphology and gonadotrophin-secreting function of the adenohypophysis were studied. In trypsin-dispersed cell culture, the adenohypophysial cells showed extensive propagation to form numerous cell colonies and finally develop into a confluent monolayer cell sheet covering completely the surface of culture vessels. Almost all of the cultured cells, however, became chromophobic, at least at the end of the first week of cultivation, when gonadotrophin was detectable neither in the culture medium nor in the cells themselves. After the addition of the hypothalamic extract, gonadotrophin became detectable again, and basophilic or PAS-positive granules also reappeared within the cells, suggesting that the gonadotrophs were stimulated by the extract to produce gonadotrophin. In organ culture and short term incubation, the incorporation of [3H] leucine into the adenohypophysial cells in relation to the addition of hypothalamic extract was examined. It was obvious that the ability to incorporate [3H] leucine into the gonadotrophs in vitro was highly dependent upon the presence of the hypothalamic extract.

scholarly journals Zinc, Zinc Transporters, and Cadmium Cytotoxicity in a Cell Culture Model of Human Urothelium

Toxics ◽  
2021 ◽  
Vol 9 (5) ◽  
pp. 94
Author(s):  
Soisungwan Satarug ◽  
Scott H. Garrett ◽  
Seema Somji ◽  
Mary Ann Sens ◽  
Donald A. Sens
Keyword(s):  
Dna Methylation ◽  
Cell Culture ◽  
Cell Viability ◽  
Zinc Transporters ◽  
Cell Culture Model ◽  
Induced Expression ◽  
Glutathione Biosynthesis ◽  
Culture Model ◽  
A Cell ◽  
Cd Exposure

We explored the potential role of zinc (Zn) and zinc transporters in protection against cytotoxicity of cadmium (Cd) in a cell culture model of human urothelium, named UROtsa. We used real-time qRT-PCR to quantify transcript levels of 19 Zn transporters of the Zrt-/Irt-like protein (ZIP) and ZnT gene families that were expressed in UROtsa cells and were altered by Cd exposure. Cd as low as 0.1 µM induced expression of ZnT1, known to mediate efflux of Zn and Cd. Loss of cell viability by 57% was seen 24 h after exposure to 2.5 µM Cd. Exposure to 2.5 µM Cd together with 10–50 µM Zn prevented loss of cell viability by 66%. Pretreatment of the UROtsa cells with an inhibitor of glutathione biosynthesis (buthionine sulfoximine) diminished ZnT1 induction by Cd with a resultant increase in sensitivity to Cd cytotoxicity. Conversely, pretreatment of UROtsa cells with an inhibitor of DNA methylation, 5-aza-2’-deoxycytidine (aza-dC) did not change the extent of ZnT1 induction by Cd. The induced expression of ZnT1 that remained impervious in cells treated with aza-dC coincided with resistance to Cd cytotoxicity. Therefore, expression of ZnT1 efflux transporter and Cd toxicity in UROtsa cells could be modulated, in part, by DNA methylation and glutathione biosynthesis. Induced expression of ZnT1 may be a viable mechanistic approach to mitigating cytotoxicity of Cd.

A New Technology for Three-Dimensional Cell Culture: The Hydrodynamic Focusing Bioreactor

1999 ◽  
Author(s):  
Yow-Min D. Tsao ◽  
Steve R. Gonda
Keyword(s):  
Cell Culture ◽  
New Technology ◽  
Three Dimensional ◽  
Culture Vessel ◽  
Air Bubbles ◽  
Hydrodynamic Focusing ◽  
Shaped Cell ◽  
Fluid Environment ◽  
Dimensional Cell ◽  
Low Shear

Abstract The Hydrodynamic Focusing Bioreactor (HDFB) developed by NASA at the Johnson Space Center provides a unique hydrofocusing capability that simultaneously enables a low-shear culture environment and a unique hydrofocusing-based “herding” of suspended cells, cell aggregates, and air bubbles. The HDFB is a rotating dome-shaped cell culture vessel with a centrally located sampling port and an internal rotating viscous spinner attached to a rotating base. The vessel and viscous spinner can rotate at different speeds and in either the same or different directions. Adjusting the differential rotation rate between the vessel and spinner results in a controllable hydrodynamic focusing force. The resultant hydrodynamic force suspends the cells in a low-shear fluid environment that supports the formation of delicate three-dimensional tissue assemblies. Both suspension and anchorage-dependent cells have been successfully cultured.

scholarly journals Is Oxytocin Proper for Cancer Adjuvant Therapy?

Proceedings ◽  
2018 ◽  
Vol 2 (25) ◽  
pp. 1582
Author(s):  
Ali Taghizadehghalehjoughi ◽  
Betul Cicek ◽  
Ahmet Hacimuftuoglu ◽  
Mustaf Gul
Keyword(s):  
Cell Culture ◽  
Cell Viability ◽  
Cell Line ◽  
Culture Media ◽  
Neuroblastoma Cell ◽  
Sympathetic Ganglia ◽  
Control Group ◽  
Human Neuroblastoma ◽  
Lactation Stage ◽  
Cortex Cell

Neuroblastomas are solid tumors and mostly seen in the adrenal medulla and sympathetic ganglia. It is known that neuroblastoma cell proliferation is inhibited by cisplatin and vincristine. The aim of this study was to investigate the effect of oxytocin on cell viability in human neuroblastoma SH-SY5Y cell line and primary cerebral cortex cell culture exposed to cisplatin and vincristine. In this direction, SH-SY5Y cell line and cortex neurons were obtained from the medical pharmacology department, Ataturk University. Both cells were grown in the appropriate cell culture media. Cisplatin (5, 10, 15 μg), vincristine (0.5, 1 and 2 ng) and oxytocin (1 μM) were applied to SH-SY5Y cell line and primary cortex cell culture for 24 h. MTT and TAC-TOS tests were performed 24 h after the application. As a result of the MTT assay, the combination of cisplatin and vincristine reduced cell viability in both cultures approximately 25% and 22%, respectively, compared to the control group. It appears that oxytocin increases neuroblastoma and cortex neuron viability, 112% and 95%, respectively. In this relation, we need to investigate why oxytocin increases cell viability and what are the possible implications in women in lactation stage.

Integrated paper-based 3D platform for long-term cell culture and in situ cell viability monitoring of Alzheimer's disease cell model

Talanta ◽  
2021 ◽  
Vol 223 ◽  
pp. 121738
Author(s):  
Meng-Meng Liu ◽  
Hui Liu ◽  
Shan-Hong Li ◽  
Yu Zhong ◽  
Yao Chen ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Cell Culture ◽  
Cell Viability ◽  
Cell Model ◽  
Disease Cell

Primary Acinar Cell Culture: Is Cell Viability Enough for Research of Pancreatic Pathophysiologic Events?

2011 ◽  
Vol 140 (5) ◽  
pp. S-858
Author(s):  
Maria Luaces-Regueira ◽  
Margarita Castiñeira-Alvariño ◽  
Julio Iglesias-Garcia ◽  
Enrique Dominguez-Munoz
Keyword(s):  
Cell Culture ◽  
Cell Viability ◽  
Acinar Cell ◽  
Acinar Cell Culture

Long-term In Vitro Toxicity Models: Comparisons Between a Flow-cell Bioreactor, a Static-cell Bioreactor and Static Cell Cultures

2002 ◽  
Vol 30 (5) ◽  
pp. 515-523 ◽  
Author(s):  
Patricia Pazos ◽  
Salvador Fortaner ◽  
Pilar Prieto
Keyword(s):  
Cell Culture ◽  
Cell Viability ◽  
Flow Cell ◽  
Model Systems ◽  
In Vitro Toxicity ◽  
Protein Determination ◽  
Efficient System ◽  
Cytotoxicity Studies

In vitro long-term toxicity testing is becoming an important issue in the field of toxicology, and there is a need to develop new model systems that mimic human chronic exposure and its effects. The aim of this work was to test two long-term in vitro toxicity systems which are available, a flow-cell bioreactor (Tecnomouse) and a static cell bioreactor system (CELLine CL 6-well), and to compare them with the use of conventional cell culture flasks. A human cell line, Int 407, was exposed to cadmium chloride (CdCl2; 10–7–10–8M) for 4 weeks. Cell numbers and cell viabilities were determined by the trypan blue (TB) exclusion assay and from exclusion of propidium iodide (PI) as determined by flow cytometry; and cell viability and metabolic activity were determined by the MTT assay. In addition, total protein determination and cadmium uptake measurements were performed. The results obtained with TB and PI exclusion did not show clear differences in cell viability with increasing CdCl2 concentration. However, in the static cell-culture systems, an increase in MTT reduction was found at low concentrations of CdCl2. Expression of heat-shock protein (Hsp27 and Hsp70) increased differently, depending on the CdCl2 concentration applied and the system used. In summary, of the two bioreactors, the CELLine CL 6-well bioreactor was shown to be the more efficient system for performing long-term cytotoxicity studies. It is easy to handle, it permits the assessment of several endpoints, and sufficient replicates can be made available.

Biocompatibility of a Bicarbonate/Lactate-Buffered PD Fluid Tested with a Double-Chamber Cell Culture System

2005 ◽  
Vol 25 (4) ◽  
pp. 387-393 ◽  
Author(s):  
Andreas Fusshoeller ◽  
Jessica Baehr ◽  
Bernd Grabensee ◽  
Joerg Plum
Keyword(s):  
Cell Culture ◽  
Cell Viability ◽  
Culture System ◽  
Mesothelial Cell ◽  
In Vitro Studies ◽  
Cell Culture System ◽  
And Function ◽  
Tgf Β1 ◽  
Double Chamber

Objective In peritoneal dialysis (PD), neutrally buffered PD fluids with lower concentrations of glucose degradation products (GDP) have tested superior to conventional fluids in terms of biocompatibility. However, conventional in vitro studies provoke debate because, due to the lack of subsequent equilibration with the blood, they do not resemble the true intraperitoneal situation of PD. Methods We established a double-chamber cell culture system with peritoneal mesothelial cells seeded on top of a permeable membrane, with a physiological buffer below. Thus adequately reflecting the in vivo equilibration pattern, we compared a conventional fluid with a neutral bicarbonate/lactate-buffered PD solution. Using an exchange pattern adapted from an 8-hour continuous ambulatory PD regimen, cell viability was assessed with an MTT assay, and cell function via constitutive and stimulated interleukin (IL)-6 release. As an indicator of potential induction of fibrosis and as a parameter of mesothelial cell integrity, respectively, transforming growth factor-beta 1 (TGF-β1) generation and cancer antigen 125 (CA125) release were measured. Results The conventional solution significantly compromised mesothelial cell viability and function in terms of mitochondrial activity ( p < 0.05) and stimulated IL-6 release ( p < 0.05). The bicarbonate/lactate fluid had no effect on cell viability or IL-6 release and turned out to be equivalent to the properties of the growth medium. Whereas lactate-incubated cells did not respond to IL-1β stimulation, bicarbonate/lactate-treated cells adequately increased IL-6 release after stimulation ( p < 0.0005). Release of TGF-β1 and CA125 did not differ between the different fluids and the control. Conclusions Due to the sustained equilibration process, the double-chamber cell culture model allows a more realistic insight into mesothelial cell viability and function in terms of PD. As in classic in vitro studies, an adverse effect of conventional PD solutions on mesothelial cells was overt in the present cell culture system. The neutral bicarbonate/lactate-buffered fluid with low GDP content, however, did not interfere with mesothelial cell vitality or function, indicating superior biocompatibility.

scholarly journals Lipodendriplexes mediated enhanced gene delivery: a cellular to pre-clinical investigation

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Imran Tariq ◽  
Muhammad Yasir Ali ◽  
Muhammad Farhan Sohail ◽  
Muhammad Umair Amin ◽  
Sajid Ali ◽  
...  
Keyword(s):  
Cell Culture ◽  
Gene Delivery ◽  
Cell Viability ◽  
3D Cell Culture ◽  
Serum Biochemistry ◽  
Cellular Protein ◽  
Ros Generation ◽  
Gfp Expression

AbstractClinical success of effective gene therapy is mainly hampered by the insufficiency of safe and efficient internalization of a transgene to the targeted cellular site. Therefore, the development of a safe and efficient nanocarrier system is one of the fundamental challenges to transfer the therapeutic genes to the diseased cells. Polyamidoamine (PAMAM) dendrimer has been used as an efficient non-viral gene vector (dendriplexes) but the toxicity and unusual biodistribution induced by the terminal amino groups (–NH2) limit its in vivo applications. Hence, a state of the art lipid modification with PAMAM based gene carrier (lipodendriplexes) was planned to investigate theirs in vitro (2D and 3D cell culture) and in vivo behaviour. In vitro pDNA transfection, lactate dehydrogenase (LDH) release, reactive oxygen species (ROS) generation, cellular protein contents, live/dead staining and apoptosis were studied in 2D cell culture of HEK-293 cells while GFP transfection, 3D cell viability and live/dead staining of spheroids were performed in its 3D cell culture. Acute toxicity studies including organ to body index ratio, hematological parameters, serum biochemistry, histopathological profiles and in vivo transgene expression were assessed in female BALB/c mice. The results suggested that, in comparison to dendriplexes the lipodendriplexes exhibited significant improvement of pDNA transfection (p < 0.001) with lower LDH release (p < 0.01) and ROS generation (p < 0.05). A substantially higher cellular protein content (p < 0.01) and cell viability were also observed in 2D culture. A strong GFP expression with an improved cell viability profile (p < 0.05) was indicated in lipodendriplexes treated 3D spheroids. In vivo archives showed the superiority of lipid-modified nanocarrier system, depicted a significant increase in green fluorescent protein (GFP) expression in the lungs (p < 0.01), heart (p < 0.001), liver (p < 0.001) and kidneys (p < 0.001) with improved serum biochemistry and hematological profile as compared to unmodified dendriplexes. No tissue necrosis was evident in the animal groups treated with lipid-shielded molecules. Therefore, a non-covalent conjugation of lipids with PAMAM based carrier system could be considered as a promising approach for an efficient and biocompatible gene delivery system.

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