Cardiac Uptake of Minocycline and Mechanisms for In Vivo Cardioprotection
Diego Romero-Perez, BS,
Eduardo Fricovsky, PharmD,
Katrina Go Yamasaki, BS,
Michael Griffin, PhD,
Maraliz Barraza-Hidalgo, BS,
Wolfgang Dillmann, MD and
Francisco Villarreal, MD, PhD*
Department of Medicine, University of California, San Diego, San Diego, California
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Figure 1 Reduction of Myocardial Infarct Size by Minocycline
(A) Morphometric analysis of area at risk (AAR) as a function of left ventricular area (AAR/LV). (B) Infarct area (IA) as a function of AAR (IA/AAR) in ischemia-reperfusion control (solid bars) (n = 12) and minocycline-treated (open bars) (n = 10) hearts. Values are mean ± SEM.
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Figure 2 Reduction of MMP-9 Levels in the Ischemic Region With Minocycline Treatment
(A) Representative gelatin zymography of myocardial matrix metalloproteinase (MMP)-2 and -9 levels in ischemia-reperfusion control (n = 4) and minocycline-treated (n = 5) hearts. (B) Bar graph derived from densitometric analysis of 92-kDa MMP-9 zymographic activity in control (nonischemic and ischemic) and minocycline-treated (nonischemic and ischemic) hearts regions. Values are mean ± SEM. ADU = arbitrary densitometric units.
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Figure 3 Reduction of Tissue Oxidative Stress With Minocycline Treatment
Tissue oxidative stress levels in control (n = 4) and minocycline-treated (n = 5) animals. Decreased ratio of oxidized to reduced glutathione content (glutathione disulfide [GSSG]/ glutathione [GSH]) was observed in ischemic heart tissue homogenates of minocycline-treated animals. Values are mean ± SEM. RFU = relative fluorescence units.
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Figure 4 In Vitro Minocycline Antioxidant Potential
Using an in vitro assay kit, we found that minocycline displays antioxidant activity similar to that of the vitamin E analog Trolox. Values are mean ± SEM of 3 experiments.
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Figure 5 Accumulation of Minocycline in Plasma, Nonischemic, and Ischemic Cardiac Tissue
Normal cardiac tissue displays a 24-fold minocycline accumulation versus plasma, which is further enhanced by 2-fold in response to ischemia. Values are mean ± SEM (n = 5).
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Figure 6 Minocycline Accumulation in Neonatal and Adult Cardiac Fibroblasts
Neonatal (A) or adult (B) cells were incubated in a monolayer of 40 µmol/l minocycline in Hank's balanced salt solution at 4°C (solid diamonds) and 37°C (open diamonds), and uptake was monitored over the indicated time intervals. Values represent mean ± SEM of 3 independent experiments.
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Figure 7 Minocycline Accumulation by Cardiac Myocytes
Cells were incubated in a monolayer of 40 µM minocycline in Hank's balanced salt solution at 4°C and 37°C and uptake was monitored over the indicated time intervals. Values represent mean ± SEM of 3 independent experiments.
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Figure 8 IC50 Values of Minocycline for Human Recombinant MMP-7 and -9
In vitro assays that used active MMPs show inhibition at IC50 = 125 µM for MMP-7 and IC50 = 180 µM for MMP-9. Abbreviations as in Figure 2.
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Figure 9 Global Heart Tissue MMP Activity
Samples used were from control (n = 4) and minocycline-treated (n = 5) animals. Minocycline significantly inhibited global MMP activity in hearts from nonischemic and ischemic regions versus corresponding controls. Values are mean ± SEM. Abbreviations as in Figure 2.
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