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Astrocyte growth media

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Astrocyte Growth Media is a specialized cell culture medium designed to support the growth and maintenance of astrocytes, a type of glial cell found in the central nervous system. The media is formulated to provide the necessary nutrients and growth factors required for optimal astrocyte proliferation and differentiation, enabling researchers to study these important cells in vitro.

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Lab products found in correlation

Trizol, by Thermo Fisher Scientific (2 mentions) Papain, by Worthington (2 mentions) Hhsec, by ScienCell (2 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Egm 2, by Lonza (2 mentions) Egm 2 growth medium, by Lonza (2 mentions) Ha1800, by ScienCell (1 mentions) Cucurbitacin 1, by Merck Group (1 mentions) L glutamine, by Lonza (1 mentions) Insulin, by Lonza (1 mentions) Normal human astrocyte cells, by Lonza (1 mentions) Hek293t, by National Collection of Authenticated Cell Cultures (1 mentions) U87mg, by National Collection of Authenticated Cell Cultures (1 mentions) Normal human astrocytes nhas, by Lonza (1 mentions) Bulletkits, by Lonza (1 mentions)

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Growth media (5 citations) Provided astrocyte growth media (2 citations)

10 protocols using astrocyte growth media

1

Cytokine-Induced Astrocyte Response

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Cultured primary human cortical astrocytes (HA1800) were purchased from ScienCell (Carlsbad, CA) and treated with cytokines: IL-1α, TNF- α, and C1q for 24 hrs21 (link). Astrocytes were treated with LXB4 (Cayman Chemicals, Ann Arbor, MI) (1μM) or vehicle for 20 mins before adding cytokines. After 24 hrs, the cells were collected in Trizol (Invitrogen, Waltham, MA) for RNA isolation or ice-cold methanol for lipidomic analysis. For primary retinal astrocytes, four mice retinas were pooled and digested using papain (Worthington, Lakewood, NJ) and plated on Poly-D-Lysine coated flasks with Astrocyte Growth Media (Lonza, Basel, Switzerland), changing the media every 3 days.
Karnam S., Maurya S., Ng E., Choudhary A., Thobani A., Flanagan J.G, & Gronert K. (2023). Dysregulation of Neuroprotective Lipoxin Pathway in Astrocytes in Response to Cytokines and Ocular Hypertension. bioRxiv.
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2

Glioma Cell Lines and Inhibitor Treatments

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Human glioma cell lines T98G and U87 were purchased from the ATCC (American Type Culture Collection) and were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% of heat-inactivated fetal bovine serum (FBS) and cultured at 37°C, 5% carbon dioxide in a humidified chamber. NHAs were purchased from Lonza (Lonza, Walkersville, MD) and maintained in Astrocyte Growth Media (Lonza) con-taining 2.5% FBS, 0.1% ascorbic acid, 0.5% recombinant human epidermal growth factor, 0.1% GA-1000, 0.25% insulin, and 1% L-glutamine (Lonza). LIMK inhibitor was purchased from SYNKINASE (SYN-1024 BMS-5) and Cucurbitacin I was purchased from Sigma-Aldrich (C-4493). Both drugs were dissolved in DMSO making a master stock of 1 mM and used at doses and times points listed in the Results section.
Park J.B., Agnihotri S., Golbourn B., Bertrand K.C., Luck A., Sabha N., Smith C.A., Byron S., Zadeh G., Croul S., Berens M, & Rutka J.T. (2014). Transcriptional profiling of GBM invasion genes identifies effective inhibitors of the LIM kinase-Cofilin pathway. Oncotarget, 5(19), 9382-9395.
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3

Astrocyte Response to Inflammatory Stimuli

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Cultured primary human cortical astrocytes (HA1800) were purchased from ScienCell (Carlsbad, CA) and treated with the cytokines: IL-1α, TNF- α, and C1q for 24 h [20 ]. Astrocytes were treated with LXB4 (Cayman Chemicals, Ann Arbor, MI) (1 µM) or vehicle for 20 min before adding cytokines. After 24 h, the cells were collected in TRIzol (Invitrogen, Waltham, MA) for RNA isolation or ice-cold methanol for lipidomic analysis. For primary retinal astrocytes and optic nerve astrocytes, four mouse retinas were pooled and digested using papain (Worthington, Lakewood, NJ) and plated on poly-D-lysine coated flasks with astrocyte growth media (Lonza, Basel, Switzerland), changing the media every 3 days. Over 95% of the cultured cells were identified as astrocytes through immunofluorescence staining for GFAP and vimentin and were negative for RGCs, microglia, and oligodendrocyte markers.
Karnam S., Maurya S., Ng E., Choudhary A., Thobani A., Flanagan J.G, & Gronert K. (2024). Dysregulation of neuroprotective lipoxin pathway in astrocytes in response to cytokines and ocular hypertension​. Acta Neuropathologica Communications, 12, 58.
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4

Establishment of Primary Cell Cultures

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Freshly isolated human umbilical vein endothelial cells (HUVECs) were provided kindly by Dr. Andrew Putnam (University of Michigan) and propagated in fully supplemented EGM-2 growth medium (Lonza). Human hepatic sinusoidal endothelial cells (HHSEC, ScienCell#5000) were cultured in EGM-2 (Lonza). Human brain vascular adventitial fibroblasts (HBVAFs, #1110) and human lung fibroblast (LFs,#3300) were obtained commercially (ScienCell) and propagated in high-glucose DMEM supplemented with 10% fetal bovine serum (FBS, ThermoFisher). Human astrocytes (ScienCell#1800) were cultured in Astrocyte Growth Media (Lonza). Bone marrow-derived human mesenchymal stem cells (MSCs, ScienCell#7500) were cultured and propagated in low-glucose DMEM supplemented with 10% FBS. All primary human cells were used in experiments from passages eight to ten.
T4-2 cells were kind gift of Dr. Mina Bissell at Lawrence Berkeley National Laboratory, University of California, Berkeley, and grown as described previously62 . MDA-MB-231 were purchased from ATCC (#HTB26) and its brain-metastatic derivative line MDA-MB-231-BR7 was generously provided by Dr. Patricia Steeg (NCI) 64 . Both MDA-MB-231 and MDA-MB-231-BR7 cells were grown in high-glucose DMEM supplemented with 10% FBS. Triple-negative murine mammary tumor line E0771 (Ch3 Biosystems#94A001) were grown in high-glucose DMEM supplemented with 10% FBS.
Dai J., Cimino P.J., Gouin KH I.I.I., Grzelak C.A., Barrett A., Lim A.R., Long A., Weaver S., Saldin L.T., Uzamere A., Schulte V., Clegg N., Pisarsky L., Lyden D., Bissell M.J., Knott S., Welm A.L., Bielas J.H., Hansen K.C., Winkler F., Holland E.C, & Ghajar C.M. (2021). Astrocytic laminin-211 drives disseminated breast tumor cell dormancy in brain. Nature cancer, 3(1), 25-42.
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5

Culturing Glioblastoma and Astrocyte Cells in Hydrogels

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U-87 MG primary human glioblastoma cells (ATCC), D1TNC1 rat astrocyte cells (ATCC), and C6 rat glioblastoma cells (ATCC) were all cultured in Dulbecco’s Modified Eagle Medium (DMEM) containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin (PS). Normal Human Astrocyte (NHA) cells (Lonza) were cultured in Astrocyte Growth Media (Lonza). PC12 undifferentiated rat neurons (ATCC) were cultured in DMEM containing 5% horse serum, 5% calf serum and 1% PS. DBTRG human glioblastoma cells (ATCC) were culture in RPMI medium containing 10% FBS, 2 mM L-glutamine, 1% PS and 0.1 mM non-essential amino acids. All cells were grown in culture at 37 °C in 5% CO2 in a humidified incubator. Cells were seeded in hydrogels at a density of 1 × 106 cells/mL. The hydrogels were submerged in appropriate growth media for the cell type at 37 °C in 5% CO2 in a humidified incubator and cell viability was maintained within hydrogels for up to 7 days (Fig. 2a).
Ivey J.W., Latouche E.L., Sano M.B., Rossmeisl J.H., Davalos R.V, & Verbridge S.S. (2015). Targeted cellular ablation based on the morphology of malignant cells. Scientific Reports, 5, 17157.
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6

Culturing Normal Human Astrocytes and Glioblastoma Cell Lines

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Normal human astrocytes (NHAs) were obtained from Lonza (Basel, Switzerland) and cultured in astrocyte growth media (Lonza) containing 3% fetal bovine serum (FBS), 4.5% glucose and reagents from BulletKits (Lonza) in a humidified incubator at 37°C and 5% CO2. Human GBM cell lines U87-MG, U251 and HEK-293T were purchased from Chinese Academy of Sciences Cell Bank (Shanghai, People’s Republic of China), and were cultured in DMEM (Hyclone, Logan, UT, USA) supplemented with 100 units of penicillin/mL, 100 ng of streptomycin/mL as well as 10% FBS. All the cells were maintained at 37°C in a humidified atmosphere with 5% CO2.
Zhang C., Yang M., Li Y., Tang S, & Sun X. (2018). FOXA1 is upregulated in glioma and promotes proliferation as well as cell cycle through regulation of cyclin D1 expression. Cancer Management and Research, 10, 3283-3293.
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7

Astrocyte Differentiation from Neural Progenitors

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Astrocytes were differentiated from the NPCs following a protocol detailed elsewhere (Freitas, et al, 2016 Neuron - in press). Briefly, a confluent 100 mm diameter NPC plate was scraped forming neurospheres in a six-well plate by keeping at constant shaking (95 rpm). The media was changed on the day after cells were suspended once the neurospheres were well formed using the NPC media containing FGF. After efficient formation of spheres around 48 hrs post scrapping, the rock inhibitor was added to a final concentration of 5 µM for 48 hrs concomitant with the removal of FGF from the media in the next media change. Cells were kept in constant shaking with neuronal inducing media for a week. Next, the astrocyte growth media (Lonza, Allendale, NJ) was added to the spheres for two weeks still under 95 rpm’s. Following two weeks on the astrocyte media, spheres are plated in laminin coated plates and the astrocytes grow out of the sphere spreading on the plate to form a multilayer cell formation. After the first passage, cells surrounding the neurospheres were dissociated enzymatically using accutase (Cellgro) and plated. The neurospheres were removed manually by vacuum suction using a Pasteur pipet and the result was a confluent and homogeneous plate of GFAP and S100β positive astrocytes.
Yao H., Azad P., Zhao H.W., Wang J., Poulsen O., Freitas B.C., Muotri A.R, & Haddad G.G. (2016). The Na+/HCO3− co-transporter is protective during ischemia in astrocytes. Neuroscience, 339, 329-337.
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8

Establishment of Primary Cell Cultures

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Freshly isolated human umbilical vein endothelial cells (HUVECs) were provided kindly by Dr. Andrew Putnam (University of Michigan) and propagated in fully supplemented EGM-2 growth medium (Lonza). Human hepatic sinusoidal endothelial cells (HHSEC, ScienCell#5000) were cultured in EGM-2 (Lonza). Human brain vascular adventitial fibroblasts (HBVAFs, #1110) and human lung fibroblast (LFs,#3300) were obtained commercially (ScienCell) and propagated in high-glucose DMEM supplemented with 10% fetal bovine serum (FBS, ThermoFisher). Human astrocytes (ScienCell#1800) were cultured in Astrocyte Growth Media (Lonza). Bone marrow-derived human mesenchymal stem cells (MSCs, ScienCell#7500) were cultured and propagated in low-glucose DMEM supplemented with 10% FBS. All primary human cells were used in experiments from passages eight to ten.
T4-2 cells were kind gift of Dr. Mina Bissell at Lawrence Berkeley National Laboratory, University of California, Berkeley, and grown as described previously62 . MDA-MB-231 were purchased from ATCC (#HTB26) and its brain-metastatic derivative line MDA-MB-231-BR7 was generously provided by Dr. Patricia Steeg (NCI) 64 . Both MDA-MB-231 and MDA-MB-231-BR7 cells were grown in high-glucose DMEM supplemented with 10% FBS. Triple-negative murine mammary tumor line E0771 (Ch3 Biosystems#94A001) were grown in high-glucose DMEM supplemented with 10% FBS.
Dai J., Cimino P.J., Gouin KH I.I.I., Grzelak C.A., Barrett A., Lim A.R., Long A., Weaver S., Saldin L.T., Uzamere A., Schulte V., Clegg N., Pisarsky L., Lyden D., Bissell M.J., Knott S., Welm A.L., Bielas J.H., Hansen K.C., Winkler F., Holland E.C, & Ghajar C.M. (2021). Astrocytic laminin-211 drives disseminated breast tumor cell dormancy in brain. Nature cancer, 3(1), 25-42.
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9

SIV Infection of Primary Macaque Astrocytes

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Primary macaque astrocytes were purchased from Cambrex and cultured in astrocyte growth media with supplements (Lonza) as described previously41 (link). Twenty-four hours prior to infection, astrocyte medium was changed to DMEM with 10% fetal bovine serum. Cells were infected at moi = 0.05 with SIV/17E-Fr59 (link). At six hours post-infection, cells were washed five times with PBS and re-fed with fresh medium. The next day, cells were treated or not with 100 units/mL of recombinant macaque IFN-beta (PBL Assay Science, Piscataway, NJ). Cells were half re-fed twice weekly with medium containing the cytokine as appropriate. Astrocyte lysates at the specified time points were collected in mirVana lysis buffer and stored at −80 °C until RNA purification.
Buchanan E.L., Espinoza D.A., McAlexander M.A., Myers S.L., Moyer A, & Witwer K.W. (2016). SAMHD1 transcript upregulation during SIV infection of the central nervous system does not associate with reduced viral load. Scientific Reports, 6, 22629.
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10

Differentiation of Astrocytes from NPCs

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Astrocytes were differentiated from the NPCs following a protocol detailed previously (Yao et al., 2016 (link)). Briefly, a confluent 100 mm diameter NPC plate was scraped forming neurospheres in a six-well plate by keeping at constant shaking (95 rpm). Media was changed on the day after cells were suspended once the neurospheres were well formed using NPC media containing FGF. After efficient formation of spheres around 48 hrs post scrapping, rock inhibitor was added to a final concentration of 5 μM for 48 hrs concomitant with the removal of FGF from the media in the next media change. Cells were kept in constant shaking with neuronal inducing media for a week. Next, astrocyte growth media (Lonza, Allendale, NJ) was added to the spheres for two weeks still under 95 rpm’s. Following two weeks on astrocyte media, spheres were plated in laminin coated plates and the astrocytes grew out of the sphere spreading on the plate to form a monolayer cell formation. After the first passage, cells surrounding the neurospheres were dissociated enzymatically using accutase and plated. The neurospheres were removed manually by vacuum suction using a Pasteur pipet and the result was a confluent and homogeneous plate of GFAP and S100β positive astrocytes.
Yao H., Zhao H., Wang J, & Haddad G.G. (2018). Intracellular pH regulation in iPSCs-derived astrocytes from subjects with Chronic Mountain Sickness. Neuroscience, 375, 25-33.
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