Pierre Joanne

Lipid reorganization induced by membrane-active peptides probed using differential scanning calorimetry.

The overlapping biological behaviors between some cell penetrating peptides (CPPs) and antimicrobial peptides (AMPs) suggest both common and different membrane interaction mechanisms. We thus explore the capacity of selected CPPs and AMPs to reorganize the planar distribution of binary lipid mixtures by means of differential scanning calorimetry (DSC). Additionally, membrane integrity assays and circular dichroism (CD) experiments were performed. Two CPPs (Penetratin and RL16) and AMPs belonging to the dermaseptin superfamily (Drs B2 and C-terminal truncated analog [1-23]-Drs B2 and two plasticins DRP-PBN2 and DRP-PD36KF) were selected. Herein we probed the impact of headgroup charges and acyl chain composition (length and unsaturation) on the peptide/lipid interaction by using binary lipid mixtures. All peptides were shown to be alpha-helical in all the lipid mixtures investigated, except for the two CPPs and [1-23]-Drs B2 in the presence of zwitterionic lipid mixtures where they were rather unstructured. Depending on the lipid composition and peptide sequence, simple binding to the lipid surface that occur without affecting the lipid distribution is observed in particular in the case of AMPs. Recruitments and segregation of lipids were observed, essentially for CPPs, without a clear relationship between peptide conformation and their effect in the lipid lateral organization. Nonetheless, in most cases after initial electrostatic recognition between the peptide charged amino acids and the lipid headgroups, the lipids with the lowest phase transition temperature were selectively recruited by cationic peptides while those with the highest phase transition were segregated. Membrane activities of CPPs and AMPs could be thus related to their preferential interactions with membrane defects that correspond to areas with marked fluidity. Moreover, due to the distinct membrane composition of prokaryotes and eukaryotes, lateral heterogeneity may be differently affected by cationic peptides leading to either uptake or/and antimicrobial activities.

Long-term functional benefits of human embryonic stem cell-derived cardiac progenitors embedded into a fibrin scaffold.

BACKGROUND Cardiac-committed cells and biomimetic scaffolds independently improve the therapeutic efficacy of stem cells. In this study we tested the long-term effects of their combination. METHODS Eighty immune-deficient rats underwent permanent coronary artery ligation. Five to 7 weeks later, those with an echocardiographically measured ejection fraction (EF) ≤55% were re-operated on and randomly allocated to receive a cell-free fibrin patch (n = 25), a fibrin patch loaded with 700,000 human embryonic stem cells (ESC) pre-treated to promote early cardiac differentiation (SSEA-1(+) progenitors [n = 30]), or to serve as sham-operated animals (n = 25). Left ventricular function was assessed by echocardiography at baseline and every month thereafter until 4 months. Hearts were then processed for assessment of fibrosis and angiogenesis and a 5-component heart failure score was constructed by integrating the absolute change in left ventricular end-systolic volume (LVESV) between 4 months and baseline, and the quantitative polymerase chain reaction (qPCR)-based expression of natriuretic peptides A and B, myosin heavy chain 7 and periostin. All data were recorded and analyzed in a blinded manner. RESULTS The cell-treated group consistently yielded better functional outcomes than the sham-operated group (p = 0.002 for EF; p = 0.01 for LVESV). Angiogenesis in the border zone was also significantly greater in the cell-fibrin group (p = 0.006), which yielded the lowest heart failure score (p = 0.04 vs sham). Engrafted progenitors were only detected shortly after transplantation; no grafted cells were identified after 4 months. There was no teratoma identified. CONCLUSIONS A fibrin scaffold loaded with ESC-derived cardiac progenitors resulted in sustained improvement in contractility and attenuation of remodeling without sustained donor cell engraftment. A paracrine effect, possibly on innate reparative responses, is a possible mechanism for this enduring effect.

Comparative Study of Two Plasticins: Specificity, Interfacial Behavior, and Bactericidal Activity

A comparative study was designed to evaluate the staphylococcidal efficiency of two sequence-related plasticins from the dermaseptin superfamily we screened previously. Their bactericidal activities against Staphylococcus aureus as well as their chemotactic potential were investigated. The impact of the GraS/GraR two-component system involved in regulating resistance to cationic antimicrobial peptides (CAMPs) was evaluated. Membrane disturbing activity was quantified by membrane depolarization assays using the diS-C3 probe and by membrane integrity assays measuring beta-galactosidase activity with recombinant strain ST1065 reflecting compromised membranes and cytoplasmic leakage. Interactions of plasticins with membrane models composed of either zwitterionic lipids mimicking the S. aureus membrane of CAMP-resistant strains or anionic lipids mimicking the negative charge-depleted membrane of CAMP-sensitive strains were analyzed by jointed Brewster angle microscopy (BAM), polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS), and differential scanning calorimetry (DSC) to yield detailed information about the macroscopic interfacial organization, in situ conformation, orientation of the peptides at the lipid-solvent interface, and lipid-phase disturbance. We clearly found evidence of distinct interfacial behaviors of plasticins we linked to the distribution of charges along the peptides and structural interconversion properties at the membrane interface. Our results also suggest that amidation might play a key role in GraS/GraR-mediated CAMP sensing at the bacterial surface.

Antimicrobial Peptides and Viral Fusion Peptides : How Different They Are?

Similarly to antimicrobial peptides (AMPs), viral fusion peptides (FPs) are membrane-active peptides. This minireview emphasizes the common properties of AMPs and FPs with a special focus on the intrinsic flexibility and structural adaptability of these peptides that are responsible for different mode of interaction with the membrane bilayers. The potential use of AMPs as multifunctional drugs with both antibacterial and antiviral properties is discussed.

Synemin acts as a regulator of signalling molecules during skeletal muscle hypertrophy

ABSTRACT Synemin, a type IV intermediate filament (IF) protein, forms a bridge between IFs and cellular membranes. As an A-kinase-anchoring protein, it also provides temporal and spatial targeting of protein kinase A (PKA). However, little is known about its functional roles in either process. To better understand its functions in muscle tissue, we generated synemin-deficient (Synm−/−) mice. Synm−/− mice displayed normal development and fertility but showed a mild degeneration and regeneration phenotype in myofibres and defects in sarcolemma membranes. Following mechanical overload, Synm−/− mice muscles showed a higher hypertrophic capacity with increased maximal force and fatigue resistance compared with control mice. At the molecular level, increased remodelling capacity was accompanied by decreased myostatin (also known as GDF8) and atrogin (also known as FBXO32) expression, and increased follistatin expression. Furthermore, the activity of muscle-mass control molecules (the PKA RII&agr; subunit, p70S6K and CREB1) was increased in mutant mice. Finally, analysis of muscle satellite cell behaviour suggested that the absence of synemin could affect the balance between self-renewal and differentiation of these cells. Taken together, our results show that synemin is necessary to maintain membrane integrity and regulates signalling molecules during muscle hypertrophy.

Intrinsic flexibility and structural adaptability of Plasticins membrane-damaging peptides as a strategy for functional versatility.

The Plasticins are a family of antimicrobial, 23–29-residue Gly-Leu-rich ortholog peptides from the frog skin that have very similar amino acid sequences, hydrophobicities, and amphipathicities but differ markedly in their conformational plasticity and spectrum of activity. The intrinsic flexibility and structural malleability of Plasticins modulate their ability to bind to and disrupt the bilayer membranes of prokaryotic and eukaryotic cells, and/or to reach intracellular targets, therefore, triggering functional versatility. The discussion is opened herein on several examples of other membrane-active peptides, like viral fusion peptides, cell-penetrating peptides, that are able to display antimicrobial activity. Hence, Plasticins could be regarded as models of multipotent membrane-active peptides guided by structural plasticity.

Investigating the role of GXXXG motifs in helical folding and self-association of plasticins, Gly/Leu-rich antimicrobial peptides☆

Plasticins (PTC) are dermaseptin-related antimicrobial peptides characterized by a large number of leucine and glycine residues arranged in GXXXG motifs that are often described to promote helix association within biological membranes. We report the structure and interaction properties of two plasticins, PTC-B1 from Phyllomedusa bicolor and a cationic analog of PTC-DA1 from Pachymedusa dacnicolor, which exhibit membrane-lytic activities on a broad range of microorganisms. Despite a high number of glycine, CD and NMR spectroscopy show that the two plasticins adopt mainly alpha-helical conformations in a wide variety of environments such as trifluoroethanol, detergent micelles and lipid vesicles. In DPC and SDS, plasticins adopt well-defined helices that lie parallel to the micelle surface, all glycine residues being located on the solvent-exposed face. Spectroscopic data and cross-linking experiments indicate that the GXXXG repeats in these amphipathic helices do not provide a strong oligomerization interface, suggesting a different role from GXXXG motifs found in transmembrane helices.

Dermaseptin DA4, although closely related to dermaseptin B2, presents chemotactic and Gram‐negative selective bactericidal activities

Antimicrobial peptides participate in innate host defense by directly eliminating pathogens as a result of their ability to damage the microbial membrane and by providing danger signals that will recruit innate immune cells to the site of infection. Dermaseptin DA4 (DRS‐DA4), a new antimicrobial peptide of the dermaseptin superfamily, was identified based on its chemotactic properties, contrasting with the currently used microbicidal properties assessment. The peptide was isolated and purified by size exclusion HPLC and RP‐HPLC from the skin of the Mexican frog, Pachymedusa dacnicolor. MS and amino acid sequence analyses were consistent with the structure GMWSKIKNAGKAAKAAAKAAGKAALGAVSEAM. CD experiments showed that, unlike most antimicrobial peptides of the dermaseptin superfamily, DRS‐DA4 is not structured in the presence of zwitterionic lipids. DRS‐DA4 is a potent chemoattractant for human leukocytes and is devoid of hemolytic activity; in addition, bactericidal tests and membrane perturbation assays on model membranes and on Escherichia coli and Staphylococcus aureus strains have shown that the antibacterial effects of DRS‐DA4 and permeabilization of the inner membrane are exclusively selective for Gram‐negative bacteria. Interestingly, despite high sequence homology with dermaseptin S4, dermaseptin B2 was not able to induce directional migration of leukocytes, and displayed a broader bactericidal spectrum. A detailed structure–function analysis of closely related peptides with different capabilities, such as DRS‐DA4 and dermaseptin B2, is critical for the design of new molecules with specific attributes to modulate immunity and/or act as microbicidal agents.

Effect of voluntary physical activity initiated at age 7 months on skeletal hindlimb and cardiac muscle function in mdx mice of both genders.

Introduction: The effects of voluntary activity initiated in adult mdx (C57BL/10ScSc‐DMDmdx/J) mice on skeletal and cardiac muscle function have not been studied extensively. Methods: We studied the effects of 3 months of voluntary wheel running initiated at age 7 months on hindlimb muscle weakness, increased susceptibility to muscle contraction−induced injury, and left ventricular function in mdx mice. Results: We found that voluntary wheel running did not worsen the deficit in force‐generating capacity and the force drop after lengthening contractions in either mdx mouse gender. It increased the absolute maximal force of skeletal muscle in female mdx mice. Moreover, it did not affect left ventricular function, structural heart dimensions, cardiac gene expression of inflammation, fibrosis, or remodeling markers. Conclusion: These results indicate that voluntary activity initiated at age 7 months had no detrimental effects on skeletal or cardiac muscles in either mdx mouse gender. Muscle Nerve 52: 788–794, 2015

Myofiber Androgen Receptor Promotes Maximal Mechanical Overload-Induced Muscle Hypertrophy and Fiber Type Transition in Male Mice

The first aim of this study was to examine the role of myofiber androgen receptor (AR) in male mice on muscle performance gain and remodeling-induced muscle mechanical overloading (OVL) that mimics resistance training. The response of OVL in mice in which AR is selectively ablated in myofibers (AR(skm-/y)) was compared with that of wild-type (WT) mice. In addition, we determined whether the synthetic anabolic androgen nandrolone administration affects the OVL response. We found that OVL increased absolute maximal force and fatigue resistance in both mouse genotypes (P < .05). However, the absolute maximal force increased more in AR(skm-/y) mice as compared with WT mice (+88% vs +63%) (P < .05). Muscle weight increased less in response to OVL in AR(skm-/y) mice (+54%) than in WT mice (+115%) (P < .05). The fiber number per cross-section similarly increased in both mouse genotypes after OVL (P < .05). In contrast to WT mice, the diameter of the fibers expressing myosin heavy chain (MHC)-2x decreased after OVL in AR(skm-/y) mice (P < .05). The MHC-2b to MHC-2a fiber type transition in response to OVL was reduced in AR(skm-/y) mice as compared with WT mice (P < .05). Finally, nandrolone administration during OVL did not further improve absolute maximal force and fatigue resistance and markedly alter muscle remodeling in both mouse genotypes. Together, our results indicate that myofiber AR is required for a complete response to OVL and that exogenous androgens do not increase muscle performance during intensive remodeling in male mice.