Protective effects of myocardium-targeted nanoparticles loaded L-arginineon on sepsis-induced myocardial injury

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To prepare primary cardiomyocyte (PCM) specific peptide-conjugated mesoporous silicon nanoparticles (MSN) with L-arginine (LA) as a core (PCM-MSN@LA), and evaluate its specific protective effect on septic myocardium. PCM-MSN@LA was prepared by condensation reaction, the characterization of PCM-MSN@LA, the amount of LA modification and release was detected, and the phagocytosis of PCM-MSN@LA and its affinity to myocardial tissue was observed. (1) Experiment one: SD neonatal rat cardiomyocytes were divided into control group (Con group), lipopolysaccharide (LPS) group, MSN@LA/LPS group and PCM-MSN@LA/LPS group. The LPS group was stimulated with 5 mg/L LPS for 16 hours, while the MSN@LA/LPS group and PCM-MSN@LA/LPS group were treated with 5 mg/L LPS and 25 mg/L LA-containing nanoparticles (MSN@LA and PCM-MSN@LA) for 16 hours. Cell viability and reactive oxygen species (ROS) production levels were detected. Apoptosis was observed via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling method (TUNEL). Western Blot was used to detect the changes in endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) proteins. (2) Experiment two: 64 healthy male C57BL/6 mice were divided into Sham group, LPS group, MSN@LA/LPS group and PCM-MSN@LA/LPS group by random number table method, 16 mice in each group. LPS group were injected 50 mg/kg LPS intraperitoneally. MSN@LA/LPS group and PCM-MSN@LA/LPS group were injected with 0.5 mg/kg MSN@LA and PCM-MSN@LA via tail vein immediately after intraperitoneal injection of LPS. Eight animals in each group were used to observe the 24-hour survival rate, and the other 8 mice were used to detect cardiac function by echocardiography at 12 hours after operation; mRNA expressions of interleukin (IL-1, IL-6) and tumor necrosis factor-α (TNF-α) were measured by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR). PCM-MSN@LA was spherical, with particle size of about 180 nm, Zeta potential of about -21 mV, with LA loaded. The amount of LA modification and release rate were 12.3% and 24.3%, respectively. Cell phagocytosis experiments showed that PCM-MSN@LA had the targeting ability of cardiomyocytes and myocardial tissue. Experiment one: after LPS stimulation of myocardial cells, cell viability decreased, while ROS generation, apoptosis, eNOS and iNOS protein expressions increased. Compared with LPS group, MSN@LA/LPS group and PCM-MSN@LA/LPS group had higher cell viability, reduced ROS levels and apoptosis, increased expressions of eNOS and iNOS. PCM-MSN@LA/LPS group changed the above effect further than MSN@LA/LPS group [cell viability (A value): 0.51±0.08 vs. 0.41±0.03, ROS (relative fluorescence intensity): 28 450±1 941 vs. 35 628±2 551, TUNEL positive cells/total cells: 0.27±0.03 vs. 0.35±0.04, eNOS/β-Tubulin: 1.467±0.046 vs. 1.201±0.131, iNOS/β-Tubulin: 1.700±0.033 vs. 1.577±0.068, all P < 0.05]. Experiment two: the number of 24-hour survive in MSN@LA/LPS group and PCM-MSN@LA/LPS group were higher than LPS group (number: 2, 4 vs. 1, P values were 0.36 and 0.03 respectively). Compared with Sham group, the cardiac function of LPS group was significantly inhibited and the mRNA expression of inflammatory factors increased. The PCM-MSN@LA/LPS group had higher left ventricular ejection fraction (LVEF) and left ventricular short-axis shortening rate (LVFS) than LPS group, and lower mRNA expressions of IL-1, IL-6, and TNF-α mRNA [LVEF: 0.456±0.019 vs. 0.337±0.017, LVFS: (21.97±1.78)% vs. (15.53±1.67)%, IL-1 mRNA (2-ΔΔCT): 169.22±8.95 vs. 189.79±6.79, IL-6 mRNA (2-ΔΔCT): 19.90±1.60 vs. 23.74±1.45, TNF-α mRNA (2-ΔΔCT): 8.21±0.81 vs. 11.00±1.48, all P < 0.05]. There was no significant difference in each index between the MSN@LA/LPS group and LPS group. PCM-MSN@LA with myocardial targeting characteristic significantly increased the activity of myocardial cells, down-regulated the expression of inflammatory factors and the production of ROS, alleviated cardiac insufficiency in sepsis, and achieved the targeted treatment of myocardial injury in sepsis.