Cardiac troponin T N-domain variant destabilizes the actin interface resulting in disturbed myofilament function.

Missense variant Ile79Asn in human cardiac troponin T (cTnT-I79N) has been associated with hypertrophic cardiomyopathy and sudden cardiac arrest in juveniles. cTnT-I79N is located in the cTnT N-terminal (TnT1) loop region and is known for its pathological and prognostic relevance. A recent structural study revealed that I79 is part of a hydrophobic interface between the TnT1 loop and actin, which stabilizes the relaxed (OFF) state of the cardiac thin filament. Given the importance of understanding the role of TnT1 loop region in Ca2+ regulation of the cardiac thin filament along with the underlying mechanisms of cTnT-I79N-linked pathogenesis, we investigated the effects of cTnT-I79N on cardiac myofilament function. Transgenic I79N (Tg-I79N) muscle bundles displayed increased myofilament Ca2+ sensitivity, smaller myofilament lattice spacing, and slower crossbridge kinetics. These findings can be attributed to destabilization of the cardiac thin filament's relaxed state resulting in an increased number of crossbridges during Ca2+ activation. Additionally, in the low Ca2+-relaxed state (pCa8), we showed that more myosin heads are in the disordered-relaxed state (DRX) that are more likely to interact with actin in cTnT-I79N muscle bundles. Dysregulation of the myosin super-relaxed state (SRX) and the SRX/DRX equilibrium in cTnT-I79N muscle bundles likely result in increased mobility of myosin heads at pCa8, enhanced actomyosin interactions as evidenced by increased active force at low Ca2+, and increased sinusoidal stiffness. These findings point to a mechanism whereby cTnT-I79N weakens the interaction of the TnT1 loop with the actin filament, which in turn destabilizes the relaxed state of the cardiac thin filament.

Antimicrobial peptide hLF1-11 directs granulocyte-macrophage colony-stimulating factor-driven monocyte differentiation toward macrophages with enhanced recognition and clearance of pathogens.

The human lactoferrin-derived peptide hLF1-11 displays antimicrobial activities in vitro and is effective against infections with antibiotic-resistant bacteria and fluconazole-resistant Candida albicans in animals. However, the mechanisms underlying these activities remain largely unclear. Since hLF1-11 is ineffective in vitro at physiological salt concentrations, we suggested modulation of the immune system as an additional mechanism of action of the peptide. We investigated whether hLF1-11 affects human monocyte-macrophage differentiation and determined the antimicrobial activities of the resulting macrophages. Monocytes were cultured for 7 days with GM-CSF in the presence of hLF1-11, control peptide, or saline for various intervals. At day 6, the cells were stimulated with lipopolysaccharide (LPS), lipoteichoic acid (LTA), or heat-killed C. albicans for 24 h. Thereafter, the levels of cytokines in the culture supernatants, the expression of pathogen recognition receptors, and the antimicrobial activities of these macrophages were determined. The results showed that a short exposure of monocytes to hLF1-11 during GM-CSF-driven differentiation is sufficient to direct differentiation of monocytes toward a macrophage subset characterized by both pro- and anti-inflammatory cytokine production and increased responsiveness to microbial structures. Moreover, these macrophages are highly effective against C. albicans and Staphylococcus aureus. In conclusion, hLF1-11 directs GM-CSF-driven differentiation of monocytes toward macrophages with enhanced effector functions.

Synthetic peptides derived from human antimicrobial peptide ubiquicidin accumulate at sites of infections and eradicate (multi-drug resistant) Staphylococcus aureus in mice.

The presence and antimicrobial activity of antimicrobial peptides (AMPs) has been widely recognized as an evolutionary preserved part of the innate immune system. Based on evidence in animal models and humans, AMPs are now positioned as novel anti-infective agents. The current study aimed to evaluate the potential antimicrobial activity of ubiquicidin and small synthetic fragments thereof towards methicillin resistant Staphylococcus aureus (MRSA), as a high priority target for novel antibiotics. In vitro killing of MRSA by synthetic peptides derived from the alpha-helix or beta-sheet domains of the human cationic peptide ubiquicidin (UBI 1-59), allowed selection of AMPs for possible treatment of MRSA infections. The strongest antibacterial activity was observed for the entire peptide UBI 1-59 and for synthetic fragments comprising amino acids 31-38. The availability, chemical synthesis opportunities, and size of these small peptides, combined with their strong antimicrobial activity towards MRSA make these compounds promising candidates for antimicrobial therapy and detection of infections in man.