Raw data about modulation of astrocyte responses to catechol-induced cytotoxicity by microglia prior to isolation of astrocyte-enriched cultures
Julita Maria Pereira Borges a,b; Lívia Bacelar de Jesus a; Cleide dos Santos Souza a; Victor Diogenes Amaral da Silva a; Silvia Lima Costa a; Maria de Fátima Dias Costa a and Ramon Santos El-Bachá a*
Author’s Institutional Affiliations
a Department of Biochemistry and Biophysics, Institute of Health Sciences, Federal University of Bahia (UFBA), Salvador, Bahia (BA), 40.110-902, Brazil.
b Department of Science and Technology, Southwest Bahia State University (UESB), Jequié, BA, 45.208-409, Brazil.
- Corresponding author: ramon@ufba.br
The raw data contained in this repository were analyzed and originated a manuscript that was submitted to the Journal of Neuroscience Research.
You will find in this repository:
1) Raw data obtained from the MTT test and quinones formation of astrocyte-enriched cultures in which microglia were early removed will be found in the folder "Early MTT and Quinones". Eight independent 96-well plates were treated with 10 different catechol concentrations between 10 and 2000 μM (n = 8 wells for every concentration including control cells treated only with the vehicle 2 × 10^-5 M HCl; n = 7 for the control group treated only with medium, which was used for normalization of data, one well was used as a blank without cells) for 72 hours. Files containing MTT data are: Early 1,2.xls; Early 3,4,5.xls; Early 6,7,8.xls. Files containing data about quinones formation are: Early 1,2 quinones.xls; Early 3,4,5 quinones.xls; Early 6,7,8 quinones.xls.
2) Raw data obtained from the MTT test and quinones formation of astrocyte-enriched cultures in which microglia were late removed will be found in the folder "Late MTT and Quinones". Eight independent 96-well plates were treated with 10 different catechol concentrations between 10 and 2000 μM (n = 8 wells for every concentration including control cells treated only with the vehicle 2 × 10^-5 M HCl; n = 7 for the control group treated only with medium, which was used for normalization of data, one well was used as a blank without cells) for 72 hours. Files containing MTT data are: Late 1,2,3.xls; Late 4,5,6.xls; Late 7 e 8.xls. Files containing data about quinones formation are: Late 1,2,3, quinones.xls; Late 4,5,6 quinones.xls; Late 7 e 8 quinones. xls. 3) Photomicrographs of astrocyte-enriched cultures obtained after an early or late removal of microglia will be found in the folder "Immunocytochemistry", subfolder "Early and late". Astrocytes were labeled with rabbit polyclonal antibody against GFAP (Red). Mouse monoclonal antibody against CD11b/c was used to investigate the presence of microglia, but no cell labeled in green was seen. The nuclear chromatin was stained with DAPI. Files Early control 1.tif and Early control 2.tif show control astrocyte-enriched cultures obtained after an early removal of microglia. Files Late control 1.tif and Late control 2.tif show astrocyte-enriched cultures obtained after a late removal of microglia. Files Early Catechol 1.tif and Early Catechol 2.tif show astrocyte-enriched cultures obtained after an early removal of microglia treated with 300 μM catechol for 72 h. Files Late catechol 1.tif and Late catechol 2.tif show astrocyte-enriched cultures obtained after a late removal of microglia treated with 300 μM catechol for 72 h. Files Early Catechol + GSH 1.tif and Early Catechol + GSH 2.tif show astrocyte-enriched cultures obtained after an early removal of microglia treated with 300 μM catechol and 3.5 μM reduced glutathione for 72 h. Files Late catechol + GSH 1.tif and Late catechol + GSH 2.tif show astrocyte-enriched cultures obtained after a late removal of microglia treated with 300 μM catechol and 3.5 μM GSH for 72 h. 4) In the folder "Immunocytochemistry", subfolder "Morphological Analysis" there are two files: "Results - Number processes - Early and Late.xlsx" contains the number of astrocyte processes counting, and "Results Cell area - Early and Late.xlsx" contains areas of cells. ImageJ was used for these measurements. 5) There are two files in the folder "MTT and Quinones 20% MCM" containing the raw data of three independent experiments in which astrocyte-enriched cultures obtained after an early removal of microglia were incubated in DMEM with 20 % (v/v) microglial-conditioned medium (MCM) and treated with 10 different catechol concentrations between 10 and 2000 μM (n = 8 wells for every concentration including control cells treated only with the vehicle 2 × 10^-5 M HCl; n = 7 for the control group treated only with medium, which was used for normalization of data, one well was used as a blank without cells) for 72 hours. The file MTT 20% MCM 1,2,3.xls contains the raw data of the MTT test, meanwhile the file Quinones 20% MCM 1,2,3.xls contains the raw data of quinones formation. 6) You will find two files in the folder "MTT and quinones 50% MCM". These files contain the raw data of three independent experiments in which astrocyte-enriched cultures obtained after an early removal of microglia were incubated in DMEM with 50 % (v/v) MCM and challenged with 10 different catechol concentrations between 10 and 2000 μM (n = 8 wells for every concentration including control cells treated only with the vehicle 2 × 10^-5 M HCl; n = 7 for the control group treated only with medium, which was used for normalization of data, one well was used as a blank without cells) for 72 hours. The file MTT 50% MCM 1,2,3.xls contains the raw data of the MTT test, meanwhile the file Quinones 50% MCM 1,2,3.xls contains the raw data of quinones formation. 7) The folder "Immunocytochemisty MCM" contains the subfolder "Early MCM" in which you will find photomicrographs of astrocyte-enriched cultures obtained after an early removal of microglia incubated in the absence or in the presence of 20 % (v/v) MCM. Both cultures were treated with 100 μM or 300 μM catechol for 72 hours. Cells were immunolabeled as described in topic 3. Files Early control 1.tif and Early control 2.tif show control cells incubated in the absence of MCM. Files Early control MCM 1.tif and Early control MCM 2.tif show cells incubated in the presence of MCM. Files Early 100 µM catechol 1.tif, Early 100 µM catechol 2.tif, Early 300 µM catechol 1.tif, and Early 300 µM catechol 2.tif show cells incubated with these catechol concentrations in the absence of MCM. Files Early 100 µM Catechol + MCM 1.tif, Early 100 µM Catechol + MCM 2.tif, Early 300 µM Catechol + MCM 1.tif, and Early 300 µM Catechol + MCM 2.tif show cells incubated with these catechol concentrations in the presence of MCM. 8) In the subfolder " Morphological Analysis" of the folder "Immunocytochemisty MCM" there are two files. The file Results - Number processes - Early MCM.xlsx. contains the number of astrocyte processes counting in experiments described in the anterior topic, and the file Results Cell area -Early Catechol MCM.xlsx contains areas of cells. ImageJ was used for these measurements. 9) The folder "qRT-PCR Analysis" contains two files. The file Template neurotrophic factor.xlsx contains data about qRT-PCR for beta-actin (ACTB), hypoxanthine phosphoribosyl transferase (HPRT), glial-derived neurotrophic factor (GDNF), and nerve growth factor (NGF), in astrocyte-enriched cultures in which microglia were early removed (P1) and in those in which microglia were late removed (P2). These cultures were treated only with vehicle (ctl) or with 100 μM or 300 μM catechol for 72 hours. The file qRTPCR cytokines and BDNF.xlsx contains data about qRT-PCR for Hprt, ACTB, interleukin-10 (IL10), interleukin-1beta (IL1B), and brain derived neurotrophic factor (Bdnf) in an independent experiment as the same described for the previous file.
