Preliminary Results of an Open Label Study of Heart Rate Variability Biofeedback for the Treatment of Major Depression

Major depressive disorder (MDD) is a common mood disorder that can result in significant discomfort as well as interpersonal and functional disability. A growing body of research indicates that autonomic function is altered in depression, as evidenced by impaired baroreflex sensitivity, changes in heart rate, and reduced heart rate variability (HRV). Decreased vagal activity and increased sympathetic arousal have been proposed as major contributors to the increased risk of cardiovascular mortality in participants with MDD, and baroreflex gain is decreased. STUDY OBJECTIVES: To assess the feasibility of using HRV biofeedback to treat major depression. DESIGN: This was an open-label study in which all eleven participants received the treatment condition. Participants attended 10 weekly sessions. Questionnaires and physiological data were collected in an orientation (baseline) session and Treatment Sessions 1, 4, 7 and 10. MEASUREMENTS AND RESULTS: Significant improvements were noted in the Hamilton Depression Scale (HAM-D) and the Beck Depression Inventory (BDI-II) by Session 4, with concurrent increases in SDNN, standard deviation of normal cardiac interbeat intervals) an electrocardiographic estimate of overall measure of adaptability. SDNN decreased to baseline levels at the end of treatment and at follow-up, but clinically and statistically significant improvement in depression persisted. Main effects for task and session occurred for low frequency range (LF) and SDNN. Increases in these variables also occurred during breathing at one's resonant frequency, which targets baroreflex function and vagus nerve activity, showing that subjects performed the task correctly. CONCLUSIONS: HRV biofeedback appears to be a useful adjunctive treatment for the treatment of MDD, associated with large acute increases in HRV and some chronic increases, suggesting increased cardiovagal activity. It is possible that regular exercise of homeostatic reflexes helps depression even when changes in baseline HRV are smaller. A randomized controlled trial is warranted.     

An overview of malaria transmission from the perspective of Amazon Anopheles vectors

In the Americas, areas with a high risk of malaria transmission are mainly located in the Amazon Forest, which extends across nine countries. One keystone step to understanding the Plasmodium life cycle in Anopheles species from the Amazon Region is to obtain experimentally infected mosquito vectors. Several attempts to colonise Ano- pheles species have been conducted, but with only short-lived success or no success at all. In this review, we review the literature on malaria transmission from the perspective of its Amazon vectors. Currently, it is possible to develop experimental Plasmodium vivax infection of the colonised and field-captured vectors in laboratories located close to Amazonian endemic areas. We are also reviewing studies related to the immune response to P. vivax infection of Anopheles aquasalis, a coastal mosquito species. Finally, we discuss the importance of the modulation of Plasmodium infection by the vector microbiota and also consider the anopheline genomes. The establishment of experimental mosquito infections with Plasmodium falciparum, Plasmodium yoelii and Plasmodium berghei parasites that could provide interesting models for studying malaria in the Amazonian scenario is important. Understanding the molecular mechanisms involved in the development of the parasites in New World vectors is crucial in order to better determine the interaction process and vectorial competence.     

Eukaryote-to-eukaryote gene transfer gives rise to genome mosaicism in euglenids

Background  

Euglenophytes are a group of photosynthetic flagellates possessing a plastid derived from a green algal endosymbiont, which was incorporated into an ancestral host cell via secondary endosymbiosis. However, the impact of endosymbiosis on the euglenophyte nuclear genome is not fully understood due to its complex nature as a 'hybrid' of a non-photosynthetic host cell and a secondary endosymbiont.     

Reduced transforming growth factor-beta signaling in cartilage of old mice: role in impaired repair capacity

Osteoarthritis (OA) is a common joint disease, mainly effecting the elderly population. The cause of OA seems to be an imbalance in catabolic and anabolic factors that develops with age. IL-1 is a catabolic factor known to induce cartilage damage, and transforming growth factor (TGF)-beta is an anabolic factor that can counteract many IL-1-induced effects. In old mice, we observed reduced responsiveness to TGF-beta-induced IL-1 counteraction. We investigated whether expression of TGF-beta and its signaling molecules altered with age. To mimic the TGF-beta deprived conditions in aged mice, we assessed the functional consequence of TGF-beta blocking. We isolated knee joints of mice aged 5 months or 2 years, half of which were exposed to IL-1 by intra-articular injection 24 h prior to knee joint isolation. Immunohistochemistry was performed, staining for TGF-beta1, -2 or -3, TGF-betaRI or -RII, Smad2, -3, -4, -6 and -7 and Smad-2P. The percentage of cells staining positive was determined in tibial cartilage. To mimic the lack of TGF-beta signaling in old mice, young mice were injected with IL-1 and after 2 days Ad-LAP (TGF-beta inhibitor) or a control virus were injected. Proteoglycan (PG) synthesis (³⁵S-sulfate incorporation) and PG content of the cartilage were determined. Our experiments revealed that TGF-beta2 and -3 expression decreased with age, as did the TGF-beta receptors. Although the number of cells positive for the Smad proteins was not altered, the number of cells expressing Smad2P strongly dropped in old mice. IL-1 did not alter the expression patterns. We mimicked the lack of TGF-beta signaling in old mice by TGF-beta inhibition with LAP. This resulted in a reduced level of PG synthesis and aggravation of PG depletion. The limited response of old mice to TGF-beta induced-IL-1 counteraction is not due to a diminished level of intracellular signaling molecules or an upregulation of intracellular inhibitors, but is likely due to an intrinsic absence of sufficient TGF-beta receptor expression. Blocking TGF-beta distorted the natural repair response after IL-1 injection. In conclusion, TGF-beta appears to play an important role in repair of cartilage and a lack of TGF-beta responsiveness in old mice might be at the root of OA development.     

Solid-state NMR investigation of the selective perturbation of lipid bilayers by the cyclic antimicrobial peptide RTD-1

RTD-1 is a cyclic beta-hairpin antimicrobial peptide isolated from rhesus macaque leukocytes. Using (31)P, (2)H, (13)C, and (15)N solid-state NMR, we investigated the interaction of RTD-1 with lipid bilayers of different compositions. (31)P and (2)H NMR of uniaxially oriented membranes provided valuable information about how RTD-1 affects the static and dynamic disorder of the bilayer. Toward phosphatidylcholine (PC) bilayers, RTD-1 causes moderate orientational disorder independent of the bilayer thickness, suggesting that RTD-1 binds to the surface of PC bilayers without perturbing its hydrophobic core. Addition of cholesterol to the POPC membrane does not affect the orientational disorder. In contrast, binding of RTD-1 to anionic bilayers containing PC and phosphatidylglycerol lipids induces much greater orientational disorder without affecting the dynamic disorder of the membrane. These correlate with the selectivity of RTD-1 for anionic bacterial membranes as opposed to cholesterol-rich zwitterionic mammalian membranes. Line shape simulations indicate that RTD-1 induces the formation of micrometer-diameter lipid cylinders in anionic membranes. The curvature stress induced by RTD-1 may underlie the antimicrobial activity of RTD-1. (13)C and (15)N anisotropic chemical shifts of RTD-1 in oriented PC bilayers indicate that the peptide adopts a distribution of orientations relative to the magnetic field. This is most likely due to a small fraction of lipid cylinders that change the RTD-1 orientation with respect to the magnetic field. Membrane-bound RTD-1 exhibits narrow line widths in magic-angle spinning spectra, but the sideband intensities indicate rigid-limit anisotropies. These suggest that RTD-1 has a well-defined secondary structure and is likely aggregated in the membrane. These structural and dynamical features of RTD-1 differ significantly from those of PG-1, a related beta-hairpin antimicrobial peptide.     

Distances between tropomyosin sites across the muscle thin filament using luminescence resonance energy transfer: evidence for tropomyosin flexibility

To obtain information about the interaction of tropomyosin (Tm) with actin associated with the regulatory states of the muscle thin filament, we used luminescence resonance energy transfer (LRET) between Tb(3+) as a donor and rhodamine as an acceptor. A novel Tb(3+) chelator, S-(2-nitro-5-thiobenzoate)cysteaminyl-DTPA-Cs124, was synthesized, which specifically labels Cys groups in proteins. With the Tb chelate as the donor and tetramethylrhodamine-5-maleimide as the acceptor, both bound to specific Cys groups of Tm, we obtained 67 A as the distance between Tm's across the actin filament, a much shorter value than that obtained from structural studies (72-86 A). The difference appears to be due to submillisecond motion associated with Tm flexibility, which brings the probes closer during the millisecond lifetime of the donor. Ca(2+) did not change the energy transfer with the reconstituted thin filament, but myosin subfragment 1 decreased the transfer, consistent with either a 5-6 A increase in distance or, more likely, a decrease in flexibility.     

Functional homodimers and heterodimers of recombinant smooth muscle tropomyosin

Skeletal and smooth muscle tropomyosin (Tm) require acetylation of their N-termini to bind strongly to actin. Tm containing an N-terminal alanine-serine (AS) extension to mimic acetylation has been widely used to increase binding. The current study investigates the ability of an N-terminal AS extension to mimic native acetylation for both alpha alpha and beta beta smooth Tm homodimers. We show that (1) AS alpha-Tm binds actin 100-fold tighter than alpha-Tm and 2-fold tighter than native smooth alphabeta-Tm, (2) beta-Tm requires an AS extension to bind actin, and (3) AS beta-Tm binds actin 10-fold weaker than AS alpha-Tm. Tm is present in smooth muscle tissues as >95% heterodimer; therefore, we studied the binding of recombinant alphabeta heterodimers with different AS extensions. This study shows that recombinant Tm requires an AS extension on both alpha and beta chains to bind like native Tm and that the alpha chain contributes more to actin binding than the beta chain. Once assembled onto an actin filament, all smooth muscle Tm's regulate S1 binding to actin Tm in the same way, irrespective of the presence of an AS extension.     

Insertion selectivity of antimicrobial peptide protegrin-1 into lipid monolayers: Effect of head group electrostatics and tail group packing

The ability to selectively target the harmful microbial membrane over that of the host cell is one of the most important characteristics of the antimicrobial peptides (AMPs). This selectivity strongly depends on the chemical and structural properties of the lipids that make up the cell membrane. A systematic study of the initial membrane selectivity of protegrin-1 (PG-1), a β-sheet AMP, was performed using Langmuir monolayers. Constant pressure insertion assay was used to quantify the amount of PG-1 insertion and fluorescence microscopy was employed to observe the effect of PG-1 on lipid ordering. Charge and packing properties of the monolayer were altered by using lipids with different head groups, substituting saturated with unsaturated lipid tail group(s) and incorporating spacer molecules. PG-1 inserted most readily into anionic films composed of phosphatidylglycerol (PG) and lipid A, consistent with its high selectivity for microbial membranes. It also discriminated between zwitteranionic phospholipids, inserting more readily into phosphatidylcholine (PC) monolayers than those composed of phosphatidylethanolamine, potentially explaining why PG-1 is hemolytic for PC-rich human erythrocytes and not for the PE-rich erythrocytes of ruminants. Increased packing density of the monolayer by increased surface pressure, increased tail group saturation or incorporation of dihydrocholesterol diminishes the insertion of PG-1. Fluorescence microscopy shows that lipid packing is disordered upon PG-1 insertion. However, the presence of PG-1 can still affect lipid morphology even with no observed PG-1 insertion. These results show the important role that lipid composition of the cell membrane plays in the activity of AMPs.     

Clavaspirin, an antibacterial and haemolytic peptide from Styela clava.

We cloned the precursor of a novel peptide from a cDNA library prepared from pharyngeal tissues of the tunicate, Styela clava. Its sequence predicted a histidine-rich, amidated 23-residue peptide (FLRF(IG)SVIHGIGHLVHHIGVAL-NH2) that we named clavaspirin. A synthetic clavaspirin was prepared and it was found that it killed Gram-positive and Gram-negative bacteria, permeabilized the outer and inner membranes of Escherichia coli, lysed phosphatidylglycerol (POPG) liposomes, and was potently haemolytic towards human and bovine erythrocytes. Each of these activities was performed more effectively at an acidic pH. Circular dichroism measurements of synthetic clavaspirin revealed a largely alpha-helical structure and polarized and residue-specific FTIR spectrometry showed that its association with phospholipid membranes was influenced by pH. Peptides such as clavaspirin may equip tunicate haemocytes to mediate cytotoxicity and participate in antimicrobial defence.     

Overexpression and structural study of the cathelicidin motif of the protegrin-3 precursor.

Numerous precursors of antibacterial peptides with unrelated sequences share a similar prosequence of 96-101 residues, referred to as the cathelicidin motif. The structure of this widespread motif has not yet been reported. The cathelicidin motif of protegrin-3 (ProS) was overexpressed in Escherichia coli as a His-tagged protein to facilitate its purification. The His tag was then removed by thrombin cleavage. In addition, the complete proprotegrin-3 (ProS-PG-3) (120 residues) was overexpressed in baculovirus-infected insect cells. As it contained the antibacterial peptide protegrin-3 in its C-terminal part, ProS-PG-3 contained four disulfide bonds. At neutral pH, ProS and ProS-PG-3 adopted two slowly exchanging conformations that existed in a ratio of 55/45. This ratio was progressively modified at acidic pH to reach a 90/10 value at pH 3.0, suggesting that electrostatic interactions are involved in such a conformational change. Therefore, the structural study of the main conformer was undertaken at pH 3.0 by circular dichroism, mass spectrometry, and homo- and heteronuclear NMR. In parallel, a model for the ProS structure was built from the X-ray structure of the chicken cystatin. ProS and the chicken cystatin share two conserved disulfide bonds as well as a high conservation of hydrophobic residues. The ProS model features the conservation of a hydrophobic core made of the interface between the N-terminal helix and the wrapping beta-sheet. Although the full assignment of the main conformer of ProS could not be obtained, available NMR data validated the presence of the N-terminal helix and of a four-stranded beta-sheet, in agreement with the cystatin fold. Moreover, we clearly demonstrated that ProS and ProS-PG-3 share the same global structure, suggesting that the presence of the highly constrained beta-hairpin of protegrin does not significantly modify the structure of the cathelicidin motif of the protegrin precursor.