PIF's targeting of microglia in a cell line and neurons both in vitro and in vivo was demonstrated [26 (link), 27 (link)]. However, whether PIF targets astrocytes, which emerged as an important cell type in neurodegenerative diseases such as MS, has not been tested [54 (link)]. Astrocyte cultures were prepared from 2-day old C57BL/6 mouse neonates. Cortices were isolated, stripped of their meninges and mechanically dissociated in ice-cold HBSS. The resulting cell suspension was then incubated with 0.05% trypsin for 25 min at 37° C followed by rinsing and filtration through a nylon mesh (70-μm pore size). The cells were then plated on collagen-coated plates and were maintained in astrocyte medium (ThermoFisher Scientific). Cells were studied after 2 weeks in culture. The effect of PIF 100 or 200nM on BDNF, recognized as key for MS therapy, was tested after a 48 hour culture [54 (link)]. In addition, three genes identified in the brain array were also validated using RT-qPCR, namely SLC2A1 (glucose transporter) and HSP90ab1 plus E2f5 related to oxidative stress and cell cycle control, respectively). At the end of the experiments cells were rinsed, then RNA was extracted and processed for RT-qPCR. Fold change was determined and compared to control. Data was generated in triplicate in three different experiments, setting significance at p<0.05.
Astrocyte medium
Manufactured by Thermo Fisher Scientific
Sourced in United States
Astrocyte medium is a specialized cell culture medium designed to support the growth and maintenance of astrocytes, which are a type of glial cell found in the central nervous system. The medium provides the necessary nutrients and growth factors required for the optimal culture of astrocytes in a laboratory setting.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (3 mentions)
U87mg, by National Collection of Authenticated Cell Cultures (1 mentions)
Shg44, by National Collection of Authenticated Cell Cultures (1 mentions)
Shg 44, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (1 mentions)
U87mg, by Thermo Fisher Scientific (1 mentions)
Roswell park memorial institute 1640 medium, by Thermo Fisher Scientific (1 mentions)
Minimum eagle s medium, by Thermo Fisher Scientific (1 mentions)
Ln229, by National Collection of Authenticated Cell Cultures (1 mentions)
Fetal bovine serum (fbs), by ScienCell (1 mentions)
Hek293t, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle medium dmem, by Thermo Fisher Scientific (1 mentions)
Hek293t, by Cell Resource Center (1 mentions)
Ln229, by Cell Resource Center (1 mentions)
Snb19, by American Type Culture Collection (1 mentions)
Ha cell line, by ScienCell (1 mentions)
Rs 2000 biological irradiator, by Rad Source (1 mentions)
Low glucose dmem, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
13 protocols using astrocyte medium
1
Investigating PIF's Effects on Astrocytes
2
Astrocyte and Glioma Cell Culture
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Normal human astrocyte cell line (NHA) and glioma cell lines (A172 and SHG-44) were purchased from The Cell Bank of Type Culture Collection of the Chinese Academy of Sciences. U87 MG was obtained from the China Center for Type Culture Collection (cat. no. 3111C0001CCC000208). NHA, A172, U87-MG and SHG-44 cells were cultured in Astrocyte medium, Dulbecco's modified Eagle's medium, minimum Eagle's medium and Roswell Park Memorial Institute 1640 medium (Invitrogen; Thermo Fisher Scientific, Inc.), respectively, with the addition of 10% fetal bovine serum (FBS; Invitrogen; Thermo Fisher Scientific, Inc.) in a 5%CO2 atmosphere at 37°C. It has been observed that U87 MG from ATCC is most probably a glioblastoma whose origin is unknown. The U87 MG was used in this study as a reference with SHG-44 and A172.
3
Cell Culture Protocols for Neuroblastoma, Glioma, and More
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
SH-SY5Y neuroblastoma cells (ATCC CRL-2266™; RRID:CVCL_0019) were cultured in a 1:1 mixture of HAMS and Minimal Essential Media, containing 10% fetal bovine serum (FBS). C6 glioma cells (ATCC CCL-107™; RRID:CVCL_0194) were cultured in HAMS and 15% horse serum (ATCC) and 2.5% FBS. HEK293 cells (ATCC CRL-1573™; RRID:CVCL_0045) were cultured in DMEM and 10% FBS. CHO745 cells and CHO cells (ATCC CRL-9618™; RRID:CVCL_0214) were cultured in 10% FBS and HAMS (ATCC). Rat primary astrocytes (iXCells Biotechnologies 10RA-005) were cultured in astrocyte medium (Thermo Fisher Scientific, USA), 2% FBS and 100 U/mL Penicillin–Streptomycin. Cell cultures were maintained in a humidified atmosphere of 5% CO2 at 37 °C and cultured in CELLSTAR 25-cm2 culture flasks with filters.
4
Establishing Glioma Cell Line Cultures
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The glioma cell lines (LN229, T98G, A172, U87 and U251) were purchased from the Chinese Academy of Sciences Cell Bank (Shanghai, China), while the normal human astrocytes (NHAs) were acquired from ScienCell Research Laboratories (Carlsbad, CA, USA). The glioma cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Gibco, USA) supplemented with 10% fetal bovine serum (ScienCell, USA), and the NHAs were cultured in astrocyte medium (Life Technologies MA, USA). All cells were cultured in an incubator containing 5% CO2 at 37°C.
5
Cytotoxicity Assays for Glioblastoma
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Ink4a/arf EGFR-VIII mouse cells and U87 human GBM cells were cultured in DMEM medium with 10% FBS. GBM4 and GBM8 patient-derived tumor neurosphere lines were cultured in stem cell medium supplemented with FGF and EGF. Primary normal human astrocytes (NHA) were cultured in astrocyte medium (Life Technologies, Grand Island, N.Y.) with EGF. Cytotoxicity of the compounds was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay (for Ink4a/arf EGFR-VIII cells) as described earlier [42 (link)] or by Alamar Blue assay (for all other cell types).
For GBM4, GBM8, U87, and NHA cell lines, viability was quantified by Alamar Blue Assay. In this assay, cells were treated with inhibitor compounds as described and Alamar Blue added after 72 h. Emission values at 590 nm were measured after the addition of Alamar Blue. Dose-response curves for MTT assays and Alamar Blue Assays were plotted and IC50 values were calculated by using GraphPad Prism (GraphPad Prism Software, Inc., La Jolla, Calif.).
For GBM4, GBM8, U87, and NHA cell lines, viability was quantified by Alamar Blue Assay. In this assay, cells were treated with inhibitor compounds as described and Alamar Blue added after 72 h. Emission values at 590 nm were measured after the addition of Alamar Blue. Dose-response curves for MTT assays and Alamar Blue Assays were plotted and IC50 values were calculated by using GraphPad Prism (GraphPad Prism Software, Inc., La Jolla, Calif.).
6
Cultivation of Human Glioma and Astrocyte Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human glioma cell lines (U251, LN229, H4, SW1783, and A172) and human embryonic kidney cell line HEK-293 T were purchased from Shanghai Institutes for Biological Sciences Cell Resource Center (Shanghai, China), while normal human astrocytes (NHAs) were purchased from Sciencell Research Laboratories (Carlsbad, CA, USA). HEK-293 T cells were cultured in Dulbecco’s modified eagle medium (DMEM, Life Technologies, Carlsbad, CA) supplemented with glucose and 10% fetal bovine serum (FBS; Life Technologies), glioma cells were cultured in Dulbecco’s modified eagle medium/F12 mixed medium supplemented with 10% FBS, and NHAs were cultured in astrocyte medium (Life Technologies). All cells were cultured at 37 °C in a humidified incubator containing 5% CO2.
7
Cell Culture and Radiation Exposure
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
U251 and SNB19 cells were purchased from American Type Culture Collection (ATCC) were cultured in Eagle’s Minimum Essential Medium (EMEM) (Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 2 mM glutamine, 1% nonessential amino acids, 1 mM sodium pyruvate and 10% fetal bovine serum. HA cell line was purchased from ScienCell™ Research Laboratories and was cultured in the astrocyte medium (Gibco) with 10% fetal bovine serum and 1% penicillin/streptomycin and was maintained at 37°C in an air atmosphere. Radiation was performed using an RS-2000 biological irradiator (Rad Source Technologies, Inc., Suwanee, GA, USA) with 160 kV X-rays with a 0.3 mm copper filter at a dose rate of ~1.2 Gy/min.
8
Mesenchymal Stromal Cells Astrocyte Differentiation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Isolation of mesenchymal stromal cells (MenSCs) derived from wild-type human menstrual blood were obtained from tissue bank code (TBC# 69308)-UdeA [29 (link)]. For astrocyte differentiation, 1 × 104 WT MenSCs/cm2 were seeded in 25 cm2 culture flasks in regular culture medium (RCm, DMEM low glucose Sigma cat# D6046 media supplemented with 10% FBS) until reach 40% of confluence. Then, the medium was replaced, and cells were incubated either in DMEM low glucose media supplemented with 2% FBS (minimal culture medium, thereafter MCm) or Astrocyte medium® (GIBCO®, cat#A1261301, 1204 N Western St., Suite C, Amarillo, TX, USA) for 7 days.
9
Bacterial and Human Cell Cultures
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bacterial strains of E. coli (KCCM 11234), P. aeruginosa (ATCC 9027), B. subtilis (ATCC 6051), S. epidermis (ATCC 12228), S. aureus (KCCM 11335), and methicillin-resistant S. aureus (MRSA, ATCC 33591) were used in this study. Every strain was maintained in Mueller-Hinton broth (MHB, Difco Laboratories, Detroit, MI, United States) under shaking conditions at 37°C. In addition, human cell lines of A549 (ATCC CCL-185), MCF7 (ATCC HTB-22), HaCaT (CLS 300493), NHA (Sciencell 1800), and NHDF (ATCC PCS-201–012) were used. The cells were cultured under a humidified atmosphere with 5% CO2 at 37°C. A549 cells were cultured in RPMI-1640 (Gibco, Grand Island, NY, United States) supplemented with 10% fetal bovine serum (FBS, Gibco) and 1% penicillin and streptomycin (PS, Gibco). HaCaT and NHDF cells were cultured in Dulbecco’s modified Eagle medium supplemented with 10% FBS and 1% PS. MCF7 cells were cultured in minimum essential media (Gibco) supplemented with 10% FBS, 1% PS, and 1 × non-essential amino acids solution (Gibco). Finally, NHA cells were cultured in astrocyte medium (Gibco).
10
GBM and Astrocyte Cell Cultivation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
We obtained GBM cell line U87 and astrocytic cell line HA from the Institute of Biochemistry and Cell Biology, Chinese Academy of Science. U87 cells were cultured in DMEM (Gibco) supplemented with 10% fetal bovine serum (Gibco). HA cells were cultured in Astrocyte Medium (Gibco). All cell lines were cultured at 37 °C and 5% CO2.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!