Antioxidative and Anti-Inflammatory Activities of Chrysin and Naringenin in a Drug-Induced Bone Loss Model in Rats
Abstract
:1. Introduction
2. Results
2.1. Retinoic Acid Is an Effective Drug for Induction of Osteoporosis in Rats
2.2. Body Weight Change
2.3. Relative Weight Changes of Uterus, Liver, Kidney, and Spleen
2.4. Relative Bone Weight, Length, and Diameter
2.5. Ca and P Levels in the Femur and Serum
2.6. Bone Mineral Content, Bone Mineral Density, and Serum Markers of Bone Turnover
2.7. Changes in Biochemical and Haematological Parameters and Pro-Inflammatory Cytokines
2.8. Effect of Naringenin, Chrysin, and Alendronate on the Oxido-Reduction Status in Liver, Kidney, Spleen, and Ovary Tissues
2.9. DNA Damage of Peripheral Blood Cells
2.10. Histological Changes in Bone Tissue
3. Discussion
4. Material and Methods
4.1. Reagents
4.2. Chrysin and Naringenin
4.3. Experimental Animals, Study Design, and Organ Processing
4.4. Monitoring Weight Changes of Animals and Selected Organs
4.5. Bone Harvesting and Analysis of Bone Physical Parameters
4.6. Serum Markers of Bone Turnover
4.7. Analysis of Haematological and Biochemical Parameters
4.8. Histological Analysis
4.9. Tissue Preparations for Quantitative Analysis of Oxidative Stress Biomarkers
4.10. Analysis of Inflammatory Cytokines
4.11. The Alkaline Comet Assay
4.12. Statistical Analyses
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Groups a | Femur Physical Characteristics (cm/100 g) (X ± SE) | ||||||
---|---|---|---|---|---|---|---|
AP ϕ Proximal Epiphysis | ML ϕ Proximal Epiphysis | AP ϕ Mid-Diaphysis | ML ϕ Mid-Diaphysis | AP ϕ Distal Epiphysis | ML Distal Epiphysis | Femur Length Trochanter-Condyle | |
Control | 0.153 ± 0.004 | 0.223 ± 0.010 | 0.129 ± 0.004 | 0.172 ± 0.021 ◊ | 0.154 ± 0.009 | 0.207 ± 0.015 | 1.405 ± 0.050 |
13cRA | 0.137 ± 0.009 | 0.214 ± 0.011 | 0.118 ± 0.005 | 0.141 ± 0.010 * | 0.150 ± 0.008 | 0.213 ± 0.009 | 1.325 ± 0.044 |
Naringenin | 0.152 ± 0.004 | 0.209 ± 0.009 | 0.136 ± 0.009 | 0.148 ± 0.010 | 0.153 ± 0.010 | 0.207 ± 0.016 | 1.489 ± 0.065 ◊ |
Chrysin | 0.140 ± 0.010 | 0.198 ± 0.012 | 0.120 ± 0.009 | 0.145 ± 0.019 | 0.144 ± 0.017 | 0.193 ± 0.011 | 1.389 ± 0.102 |
Alendronate | 0.146 ± 0.004 | 0.222 ± 0.005 | 0.128 ± 0.005 | 0.149 ± 0.004 | 0.169 ± 0.015 | 0.228 ± 0.001 | 1.390 ± 0.036 |
Groups a | Biochemical Parameters-Enzymes (X ± SE) | |||||
---|---|---|---|---|---|---|
AST (U/L) | ALT (U/L) | ALP (U/L) | GGT (U/L) | LDH (U/L) | Amylase (U/L) | |
Control | 78.33 ± 7.73 | 46.40 ± 9.52 | 170.13 ± 24.54 ◊ | 0.00 ± 0.00 | 464.50 ± 130.97 | 1751.00 ± 609.96 |
13cRA | 96.16 ± 13.34 | 43.16 ± 4.66 | 224.00 ± 15.01 * | 0.66 ± 0.51 | 725.66 ± 157.59 * | 2046.16 ± 213.69 |
Naringenin | 74.00 ± 5.25 ◊ | 41.00 ± 3.89 | 147.50 ± 30.09 ◊ | 0.33 ± 0.51 | 201.50 ± 97.65 ◊◊◊ | 1897.00 ± 311.35 |
Chrysin | 83.00 ± 4.00 | 41.00 ± 4.19 | 186.33 ± 45.60 | 0.16 ± 0.40 | 325.16 ± 121.79 ◊ | 1912.16 ± 510.88 |
Alendronate | 87.66 ± 7.50 | 42.00 ± 2.64 | 115.00 ± 12.00 ◊◊ | 0.33 ± 0.57 | 402.00 ± 122.74 | 2184.00 ± 419.06 |
Groups a | Leukocytes (×109 L−1) | Differential Blood Count (%) (X ± SE) | ||||
---|---|---|---|---|---|---|
Lymphocytes | Monocytes | Neutrophils | Basophils | Eosinophils | ||
Control | 2.70 ± 1.17 | 79.06 ± 7.99 | 0.42 ± 0.21 | 13.46 ± 2.97 | 0.38 ± 0.21 | 0.61 ± 0.27 |
13cRA | 3.88 ± 1.12 | 83.96 ± 2.74 | 0.45 ± 0.16 | 15.18 ± 2.28 | 0.55 ± 0.17 | 0.73 ± 0.36 |
Naringenin | 3.85 ± 0.47 | 86.95 ± 2.02 | 0.38 ± 0.21 | 11.45 ± 2.37 | 0.36 ± 0.23 | 0.85 ± 0.38 |
Chrysin | 4.51 ± 1.33 | 79.55 ± 6.86 | 0.36 ± 0.22 | 15.33 ± 3.29 | 0.46 ± 0.15 | 1.05 ± 0.25 |
Alendronate | 5.35 ± 1.36 * | 70.16 ± 3.29 | 0.86 ± 0.20 * | 28.00 ± 4.33 * | 1.23 ± 0.50 | 0.73 ± 0.41 |
Groups a | MDA (nmol/mg Proteins) | |||
---|---|---|---|---|
Liver | Spleen | Kidney | Ovary | |
Control | 10.22 ± 0.56 ◊ | 16.24 ± 1.05 | 4.09 ± 0.60 ◊◊ | 12.12 ± 1.63 ◊◊ |
13cRA | 15.58 ± 0.63 * | 17.61 ± 1.56 | 12.33 ± 0.72 ** | 35.78 ± 3.45 ** |
Naringenin | 13.36 ± 0.33 | 13.66 ± 0.92 ♦ | 3.90 ± 0.46 ◊◊ | 20.06 ± 2.01 ◊ |
Chrysin | 14.04 ± 0.42 | 19.11 ± 1.70 | 4.81 ± 0.53 ◊◊ | 28.99 ± 1.33 *◊ |
Alendronate | 18.05 ± 0.83 ** | 23.62 ± 2.15 | 9.84 ± 0.87 * | 37.41 ± 2.14 ** |
GSH (µg/mg proteins) | ||||
Control | 8.90 ± 0.71 | 14.78 ± 2.25 | 8.46 ± 0.11 ◊ | 0.95 ± 0.15 ◊ |
13cRA | 5.65 ± 0.54 | 8.39 ± 0.38 | 3.61 ± 0.32 * | 0.20 ± 0.02 * |
Naringenin | 7.08 ± 1.05 | 15.11 ± 3.18 | 10.07 ± 0.54 ◊ | 0.56 ± 0.10 |
Chrysin | 7.64 ± 1.63 | 13.38 ± 1.77 | 6.75 ± 0.69 | 1.01 ± 0.09 ◊ |
Alendronate | 5.09 ± 1.20 | 12.69 ± 1.27 | 10.63 ± 0.61 ◊◊ | 0.37 ± 0.20 |
SOD (U/mg proteins) | ||||
Control | 3.67 ± 0.69 ◊ | 5.38 ± 0.52 | 4.07 ± 0.48 | 15.46 ± 1.81 ♦♦ |
13cRA | 1.66 ± 0.13 * | 4.22 ± 1.14 | 3.94 ± 0.52 | 7.44 ± 0.69 |
Naringenin | 4.61 ± 0.35 ◊◊♦ | 9.03 ± 1.80 ◊ | 4.56 ± 0.86 | 9.28 ± 1.01 ♦ |
Chrysin | 2.50 ± 0.22 | 4.93 ± 1.06 | 4.06 ± 1.40 | 11.54 ± 1.81 ♦ |
Alendronate | 1.77 ± 0.38 | 5.29 ± 1.37 | 7.74 ± 1.75 | 2.46 ± 0.43 ** |
CAT (U/mg proteins) | ||||
Control | 4.98 ± 0.42 | 4.22 ± 0.28 | 21.99 ± 1.58 | 14.03 ± 0.06 |
13cRA | 4.67 ± 0.61 | 2.76 ± 0.40 | 8.94 ± 0.73 | 11.64 ± 1.73 |
Naringenin | 4.45 ± 0.62 | 4.09 ± 0.87 | 43.77 ± 5.71 ◊◊ | 15.92 ± 1.85 |
Chrysin | 4.70 ± 0.34 | 2.82 ± 0.33 ♦ | 41.24 ± 4.87 ◊◊ | 24.53 ± 0.24 **◊ |
Alendronate | 4.08 ± 0.86 | 6.99 ± 0.56 ◊ | 66.45 ± 4.21 ◊◊◊ | 20.92 ± 1.10 *◊ |
Parameter | Method | |
---|---|---|
Serum biochemical parameters | Aspartate aminotransferase (AST), Alanine aminotransferase (ALT), Alkaline phosphatase (ALP), Glutamyl transferase (GGT), Amylase Urea, creatinine, Blood glucose levels (glucose), Lactate dehydrogenase (LDH), Total protein and serum Ca and P levels | Colorimetric method |
Hematological parameters | Erythrocytes (E), The average cellular volume of erythrocytes (MCV), Haemoglobin (Hgb), Haematocrit (Hct), Mean cell haemoglobin (MCH), Mean cell Haemoglobin concentration (MCHC), Total leukocyte count (L), and The total number of platelets (Plt) | Colorimetric method |
Serum Inflammatory cytokines | IL1α, IL1β, IL2, IL4, IL6, IL10, IL12, IL17A, IFNγ, TNFα, GM-CSF, and RANTES | ELISA method |
Biochemical bone parameters | Ca and P analyses | Atomic absorption spectrophotometry |
Bone mineral density (BMD) | Dual-energy X-ray absorptiometry (DXA). | |
Serum markers of bone turnover Osteocalcin (OC) β-CrossLaps | Electrochemiluminescence “ECLIA” method | |
Bone physical parameters | Relative bone weight index, Length of the bone (large trochanter-condyle), The distal and proximal epiphyseal diameters of the femur were measured in mediolateral (ML) and anteroposterior directions (AP) | Digital scale Small bone caliper |
Histological parameters | Histological analyses | Haematoxylin and eosin (HE) methods |
Oxidative stress parameters | Lipid peroxidation level (MDA), Glutathione level (GSH) Superoxide dismutase (SOD) activity Catalase (CAT) activity | Colorimetric method |
DNA damage | Alkaline Comet Assay |
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Oršolić, N.; Nemrava, J.; Jeleč, Ž.; Kukolj, M.; Odeh, D.; Jakopović, B.; Jazvinšćak Jembrek, M.; Bagatin, T.; Fureš, R.; Bagatin, D. Antioxidative and Anti-Inflammatory Activities of Chrysin and Naringenin in a Drug-Induced Bone Loss Model in Rats. Int. J. Mol. Sci. 2022, 23, 2872. https://doi.org/10.3390/ijms23052872
Oršolić N, Nemrava J, Jeleč Ž, Kukolj M, Odeh D, Jakopović B, Jazvinšćak Jembrek M, Bagatin T, Fureš R, Bagatin D. Antioxidative and Anti-Inflammatory Activities of Chrysin and Naringenin in a Drug-Induced Bone Loss Model in Rats. International Journal of Molecular Sciences. 2022; 23(5):2872. https://doi.org/10.3390/ijms23052872
Chicago/Turabian StyleOršolić, Nada, Johann Nemrava, Željko Jeleč, Marina Kukolj, Dyana Odeh, Boris Jakopović, Maja Jazvinšćak Jembrek, Tomica Bagatin, Rajko Fureš, and Dinko Bagatin. 2022. "Antioxidative and Anti-Inflammatory Activities of Chrysin and Naringenin in a Drug-Induced Bone Loss Model in Rats" International Journal of Molecular Sciences 23, no. 5: 2872. https://doi.org/10.3390/ijms23052872
APA StyleOršolić, N., Nemrava, J., Jeleč, Ž., Kukolj, M., Odeh, D., Jakopović, B., Jazvinšćak Jembrek, M., Bagatin, T., Fureš, R., & Bagatin, D. (2022). Antioxidative and Anti-Inflammatory Activities of Chrysin and Naringenin in a Drug-Induced Bone Loss Model in Rats. International Journal of Molecular Sciences, 23(5), 2872. https://doi.org/10.3390/ijms23052872