Cytoprotective and antioxidant effects of aged garlic extract against adriamycin-induced cardiotoxicity in adult male rats

Figures

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  Fig. 1. Light micrographs of rat heart. (A, B) Control and AGE-treated group showing normal organization of the heart structure of branching MF, with myocytes having central elongated N, and numerous BV in-between. (C) ADR-treated showing marked tissue necrosis (***), disorganized degenerated MF, apoptotic N, interstitial edema, and dilated congested BV. (D) AGE pre-treated group showing areas of tissue necrosis (***), degenerated MF, and BV. (E) AGE post-treated group showing minimal myofibrillar degeneration (MF) with the wavy organization and congested BV. (F) Combined pre- and post-AGE treated group showing normally organized MF, myocytes with central elongated N and vascular congestion (BV) in-between. H&E stain, Scale bars=20 µm (A–F). ADR, adriamycin; AGE, aged garlic extract; BV, blood vessels; MF, myofibrils; N, nuclei.

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  Fig. 2. Light micrographs of rat heart. (A) Control group showing little collagen fibers (R) between the MF and around the BV. (B) AGE-treated group showing minimal collagen fibers (R) between the MF and around the BV. (C) ADR-treated group showing an excessive amount of collagen fibers (R) around the congested BV and between the MF. (D) AGE pre-treated group showing an excessive amount of collagen fibers (R) between the MF and around the BV. (E) AGE post-treated group showing minimal collagen fibers (R) between the cardiac MF and around the BV. (C–E) Symbol ** represent the degenerated area of myocardium. (F) AGE combined pre- and post-treated group showing a minimal amount of collagen fibers (R) between the MF and around the BV. Masson trichrome stain, Scale bars=20 µm (A–F). ADR, adriamycin; AGE, aged garlic extract; BV, blood vessels; MF, myofibrils; R, positive PAS reaction.

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  Fig. 3. Light micrographs of the rat heart. (A) Control group showing a R at the perimysium of the MF. (B) AGE-treated group showing a minimal R at the perimysium of the MF. (C) ADR-treated group showing minimal R at the perimysium of the remaining MF. (D) AGE pre-treated group showing a R at the perimysium between the MF. (E) AGE post-treated group showing a R at the perimysium between the MF. (C–E) Symbol *** represent the degenerated area of myocardium. (F) AGE combined pre- and post-treated group showing an excessive R at the perimysium between the MF. PAS stain, Scale bars=20 µm (A–F). ADR, adriamycin; AGE, aged garlic extract; MF, myofibrils; PAS, periodic acid Schiff’s; R, positive PAS reaction.

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  Fig. 4. Electron micrograph of control rat heart showing (A–C) normal organization of branching MF with cross striations. Regular S extends between the two Z. White filament (I-band) surrounds the Z and the dark filaments (A-band) in-between. Columns of M are present between the MF and around the N. Tubular SR is present between the MF near to Z. (D) The ICD is present between two myocytes. The disc consists of different types of JC. TEM osmium tetraoxide-Silver nitrate stains (A–D). Scale bars=2 µm (A–C), 500 nm (D). BV, blood vessels; CY, cytoplasm; ICD, intercalated disc; JC, junctional complexes; M, mitochondria; MF, myofibrils; N, nucleus; S, sarcomere; SR, sarcoplasmic reticulum; Z, Z-line.

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  Fig. 5. Electron micrographs of rat heart treated with AGE showing (A–D) normal organization of MF with columns of round M in-between. The MF have cross striations with numerous S between the Z, white filaments (I-band) around the Z and dark filaments (A-band) between the white filaments. The myocyte has a regular SL with subsarcolemmal space (S) underneath. (B) The myocytes have central elongated regular heterochromatic N. Scattered tubular SR is seen between the MF near to the Z. (D) Complex ICD composing of multiple JC are present between the neighboring myocytes. TEM osmium tetraoxide-Silver nitrate stains (A–D). Scale bars=2 µm (A–C), 500 nm (D). AGE, aged garlic extract; ICD, intercalated disc; JC, junctional complexes; M, mitochondria; MF, myofibrils; N, nucleus; S, sarcomere; SL, sarcolemmal membrane; SR, sarcoplasmic reticulum; Z, Z-line.

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  Fig. 6. Electron micrographs of ADR-treated rat heart showing (A–C) marked tissue degeneration and necrosis (**) with MF disorganization. Irregular interrupted Z and SL are seen with cardiac tissue. The myocytes have apoptotic N* with wide electron-lucent perinuclear space. Scattered pleomorphic degenerated M are seen between the atrophic MF. The M show irregular outline, moth-eaten degeneration (*), condensed electron-dense matrix and loss of their cristae. Irregular discontinuing SL is seen limiting the myocytes. (C) Segmented apoptotic N* with peripheral condensed heterochromatin is seen within the myocyte. (C, D) Fragmented ICD composed of loose JC are seen between the myocytes. (D) Short contracted MF and degenerated (*) M are present between the myocytes. TEM osmium tetraoxide-Silver nitrate stains (A–D). Scale bars=2 µm (A–C), 500 nm (D). ADR, adriamycin; ICD, intercalated disc; JC, junctional complexes; M, mitochondria; MF, myofibrils; N, nucleus; SL, sarcolemmal membrane; SR, sarcoplasmic reticulum; Z, Z-line.

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  Fig. 7. Electron micrographs of the rats’ hearts treated with AGE for one week before ADR injection (AGE pre-treated group) showing (A–C) organized MF with cross striations, well-formed Z, white (I-band) and dark (A-band and H-band) filaments. Symbol * represent the moth-eaten degenerated mitochondria. Scattered area of tissue necrosis (**) is present within the cardiac tissue. Many degenerated M with loss of its cristae are seen between the MF. (A) The myocyte exhibits regular central oblong heterochromatic N with electron-dense n and intended nuclear envelope. (D) An ICD with loose JC is seen between the myocytes. Contracted short myofilaments (MF) are seen around the ICD. TEM osmium tetraoxide-Silver nitrate stains (A–D). Scale bars=2 µm (A–C), 500 nm (D). ADR, adriamycin; AGE, aged garlic extract; ICD, intercalated disc; M, mitochondria; MF, myofibrils; N, nucleus; n, nucleolus; SR, sarcoplasmic reticulum; Z, Z-line.

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  Fig. 8. Electron micrographs of the rats’ hearts treated with AGE for one week after ADR injection (AGE post-treated group) showing (A–C) organized branching MF with columns of compact M in-between. The S are present between the regular Z with white filaments (I-band) around and dark filaments (A-band) in-between. H-bands are present at the centre of the dark filaments (A-band). (B) Regular elongated heterochromatic N with wide electron-lucent perinuclear space. Few degenerated M with loss transverse cristae are observed between the MF. (C) Area of myocardium degeneration (**). (D) An ICD consisting of multiple JC is present between the neighboring myocytes. TEM osmium tetraoxide-Silver nitrate stains (A–D). Scale bars=2 µm (A–C), 500 nm (D). ADR, adriamycin; AGE, aged garlic extract; ICD, intercalated disc; JC, junctional complexes; M, mitochondria; MF, myofibrils; N, nucleus; S, sarcomere; SL, sarcolemmal membrane; SR, sarcoplasmic reticulum; Z, Z-line.

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  Fig. 9. Electron micrographs of the rats’ hearts treated with AGE for one week before and one week after ADR injection (combined AGE pre- and post-treated group) showing (A–D) normal well-organized MF having transverse striations and columns of large-sized regular normal M. The MF have normal white (I-band) and dark (A-band and A-band) filaments. Scattered tubular SR is seen between the MF near to the regular Z. (B) The myocytes have central elongated heterochromatic N with electron-dense small n within its nucleoplasm and an intended regular nuclear envelope. The N is surrounded by groups of rounded-shape normal M and tubular SR. (D) Well-formed ICD with different types of the JC are seen between the adjacent myocytes. The ICD are surrounded by well-organized normal MF with light (I-band) and dark (A-band and H-band) filaments. TEM osmium tetraoxide-Silver nitrate stains (A–D). Scale bars=2 µm (A–C), 500 nm (D). ADR, adriamycin; AGE, aged garlic extract; ICD, intercalated disc; JC, junctional complexes; M, mitochondria; MF, myofibrils; N, nucleus; n, nucleolus; S, sarcomere; SR, sarcoplasmic reticulum; Z, Z-line.