研究目的
To demonstrate whether the progesterone retinal protection in retinitis pigmentosa (RP) is related to its possible anti-inflammatory properties.
研究成果
Progesterone reduces photoreceptor cell death, reactive gliosis, microglial activation, and lipid peroxidation in rd10 mice, suggesting its anti-inflammatory and antioxidant properties. It could be a therapeutic option for retinal diseases like RP associated with inflammation and oxidative stress.
研究不足
The study is limited to an animal model (rd10 mice) and may not fully translate to human retinal diseases. The dose and timing of progesterone administration were specific to this model, and long-term effects were not extensively studied. The ex vivo lipid peroxidation experiment used liver tissue instead of retina due to tissue amount constraints, which might not perfectly replicate retinal conditions.
1:Experimental Design and Method Selection:
Used rd10 mice as an animal model of RP. Administered oral progesterone (150 mg/kg weight dissolved in olive oil) on alternate days from post-natal day 15 (P15) to P
2:Euthanized mice five hours after the last dose. Included control groups with vehicle (olive oil) only. Sample Selection and Data Sources:
C57BL/6J and rd10 mice housed under controlled conditions. Both male and female populations were used.
3:List of Experimental Equipment and Materials:
Progesterone, olive oil, gastric tube, cryostat (Leica CM 1850 UV Ag protect), microscope (Leica DM 2000), camera (Nikon DS-Fi1), software (Leica application Suite version
4:0 R1, Image J 45s, SPSS version 0), antibodies (anti-GFAP, anti-Iba1, anti-nNOS, anti-iNOS), secondary antibodies (Alexa Fluor 488), TUNEL assay kit (Roche Diagnostics), western blot equipment, homogenizer (ultraturrax T25), HPLC for MDA determination. Experimental Procedures and Operational Workflow:
Performed histological studies (TUNEL assay, immunofluorescence for GFAP, Iba1, nNOS), biochemical studies (western blot for iNOS, MDA concentration determination using HPLC, ex vivo lipid peroxidation induction in liver tissue with Fe(II) salts and ascorbic acid). Data analyzed using ANOVA and post-hoc tests.
5:Data Analysis Methods:
Statistical analysis with SPSS software, including Levene's test, ANOVA, Fisher's LSD or Games-Howell tests. Densitometry for western blots, image processing with Image J.
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cryostat
CM 1850 UV Ag protect
Leica
Used to obtain retinal sections for histological studies.
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microscope
DM 2000
Leica
Used for imaging retinal sections in immunofluorescence studies.
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software
Leica application Suite version 2.7.0 R1
Leica
Used for image acquisition and analysis.
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camera
DS-Fi1
Nikon
Attached to microscope for capturing images of retinal sections.
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software
Image J 1.45s
NIH
Used for image processing and measurements in retinal studies.
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software
SPSS version 15.0
IBM
Used for statistical analysis of experimental data.
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TUNEL assay kit
in situ cell death detection kit
Roche Diagnostics
Used to detect apoptotic cells in retinal sections.
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antibody
anti-GFAP
Dako cytomation
Used for immunofluorescent staining to detect glial fibrillary acidic protein.
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antibody
anti-Iba1
Abcam
Used for immunofluorescent staining to detect microglial cells.
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antibody
anti-nNOS
Santacruz Biotechnology
Used for immunofluorescent staining to detect neuronal nitric oxide synthase.
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secondary antibody
Alexa Fluor 488
Invitrogen
Fluorescence-conjugated secondary antibody used in immunofluorescence.
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homogenizer
ultraturrax T25
Labortech
Used for homogenizing retinal and liver tissues for biochemical studies.
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western blot equipment
GE Healthcare
Used for protein analysis, including iNOS expression.
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HPLC
Used for determining malondialdehyde (MDA) concentrations.
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