The degree of HIV-1 amino acid variability is strictly related to different disease progression rates

The aim of this study is to evaluate the amino acid variability of HIV-1 Gp41, C2–V3, and Nef in a group of patients characterized by different disease progression rates. HIV-1 sequences were collected from 19 Long term non progressor patients (LTNPs), 9 slow progressors (SPs), and 11 rapid progressors (RPs). Phylogenetic trees were estimated by MEGA 6. Differences in amino acid variability among sequences belonging to the 3 groups have been evaluated by amino acid divergence, Shannon entropy analysis, and the number of amino acid mutations (defined as amino acid variations compared with HxB2). The involvement of amino acid mutations on epitope rich regions was also investigated. The population was mainly composed of males (74.3%) and HIV-1 subtype B strains (B: 92.32%, CRF_12BF, A1, C: 2.56% each). Viral load (log₁₀ copies/mL) and CD4⁺T cell count (cells/mm³) were 3.9 (3.5–4.2) and 618 (504–857) in LTNPs, 3.3 (2.8–4.7) and 463 (333–627) in SPs, and 4.6 (4.3–5.3) and 201 (110–254) in RPs. Gp41 and C2–V3 amino acid divergence was lower in LTNP and SP strains compared to RPs (median value: 0.085 and 0.091 vs. 0.114, p?=?0.005 and 0.042) and a trend of lower variability was observed for Nef (p?=?0.198). A lower entropy value was observed at 10, 3, and 7 positions of Gp41, C2–V3, and Nef belonging to LTNPs and at 7, 3, and 1 positions of Gp41, C2–V3, and Nef belonging to SPs compared with RPs (p?<?0.05). Focusing on epitope rich regions, again a higher degree of conservation was observed in Gp41 and C2–V3 sequences belonging to LTNPs and SPs compared to those belonging to RPs. This study shows that the extent of amino acid variability correlates with a different HIV-1 progression rate. This variability also involves CTL epitope rich regions, thus suggesting its involvement in the immune escape process modulation.     

Anatomical features of tibia and femur: Influence on laxity in the anterior cruciate ligament deficient knee

Background

Until now, there has been a lack of in vivo analysis of the correlation between bony morphological features and laxity values after an anterior cruciate ligament (ACL) injury.

A Comparison of the Conditioning Regimens BEAM and FEAM for Autologous Hematopoietic Stem Cell Transplantation in Lymphoma: An Observational Study on 1038 Patients From Fondazione Italiana Linfomi

BEAM (carmustine [bis-chloroethylnitrosourea (BCNU)]-etoposide-cytarabine-melphalan) chemotherapy is the standard conditioning regimen for autologous stem cell transplantation (ASCT) in lymphomas. Owing to BCNU shortages, many centers switched to fotemustine-substituted BEAM (FEAM), lacking proof of equivalence. We conducted a retrospective cohort study in 18 Italian centers to compare the safety and efficacy of BEAM and FEAM regimens for ASCT in lymphomas performed from 2008 to 2015. We enrolled 1038 patients (BEAM =?607, FEAM =?431), of which 27% had Hodgkin lymphoma (HL), 14% indolent non-Hodgkin lymphoma (NHL), and 59% aggressive NHL. Baseline characteristics including age, sex, stage, B-symptoms, extranodal involvement, previous treatments, response before ASCT, and overall conditioning intensity were well balanced between BEAM and FEAM; notable exceptions were median ASCT year (BEAM?=?2011 versus FEAM?=?2013, P?<?.001), Sorror score ≥3 (BEAM?=?15% versus FEAM?=?10%, P?=?.017), and radiotherapy use (BEAM?=?18% versus FEAM?=?10%, P?<?.001). FEAM conditioning resulted in higher rates of gastrointestinal and infectious toxicities, including severe oral mucositis grade ≥3 (BEAM?=?31% versus FEAM?=?44%, P?<?.001), and sepsis from Gram-negative bacteria (mean isolates/patient: BEAM?=?.1 versus FEAM?=?.19, P?<?.001). Response status at day 100 post-ASCT (overall response: BEAM?=?91% versus FEAM?=?88%, P?=?.42), 2-year overall survival (83.9%; 95% confidence interval [CI], 81.5% to 86.1%) and progression-free survival (70.3%; 95% CI, 67.4% to 73.1%) were not different in the two groups. Mortality from infection was higher in the FEAM group (subhazard ratio, 1.99; 95% CI, 1.02 to 3.88; P?=?.04). BEAM and FEAM do not appear different in terms of survival and disease control. However, due to concerns of higher toxicity, fotemustine substitution in BEAM does not seem justified, if not for easier supply.     

Mossy fiber-evoked subthreshold responses induce timing-dependent plasticity at hippocampal CA3 recurrent synapses

Dentate granule cells exhibit exceptionally low levels of activity and rarely elicit action potentials in targeted CA3 pyramidal cells. It is thus unclear how such weak input from the granule cells sustains adequate levels of synaptic plasticity in the targeted CA3 network. We report that subthreshold potentials evoked by mossy fibers are sufficient to induce synaptic plasticity between CA3 pyramidal cells, thereby complementing the sparse action potential discharge. Repetitive pairing of a CA3–CA3 recurrent synaptic response with a subsequent subthreshold mossy fiber response induced long-term potentiation at CA3 recurrent synapses in rat hippocampus in vitro. Reversing the timing of the inputs induced long-term depression. The underlying mechanism depends on a passively conducted giant excitatory postsynaptic potential evoked by a mossy fiber that enhances NMDA receptor-mediated current at active CA3 recurrent synapses by relieving magnesium block. The resulting NMDA spike generates a supralinear depolarization that contributes to synaptic plasticity in hippocampal neuronal ensembles implicated in memory.The CA3 area of the hippocampus exhibits a distinctive, highly recurrent circuitry proposed to support autoassociative memory representation (1, 2). This prediction has been confirmed by experimental work demonstrating the pattern completion capabilities of CA3 networks (3), as well as their roles in the spatial tuning of CA1 pyramidal cells, in one-trial contextual learning (4) and in certain forms of memory consolidation (5). CA3 pyramidal cells receive, via the mossy fibers, information processed by granule cells important for both pattern separation (6, 7) and pattern completion functions (7). The faithful transmission of mossy fiber input appears to be ensured by giant synapses composed of presynaptic boutons with up to 45 release sites (8) that target massive spines, the thorny excrescences, on the apical dendrite of CA3 pyramidal cells. Thus, the mossy fiber synapse is often referred to as a detonator synapse (9). In fact, mossy fiber signaling is more compatible with a gatekeeper function than a high-throughput data relay. Although high-frequency bursts of action potentials in a hippocampal granule cell can discharge a targeted CA3 pyramidal cell, the majority of responses evoked by granule cells in CA3 pyramidal cells do not attain the firing threshold (10). Nevertheless, mossy fibers generate powerful signals evoking subthreshold responses that are much larger than typical synaptic events in the brain, with excitatory postsynaptic potentials (EPSPs) and excitatory postsynaptic currents (EPSCs) reaching amplitudes of 10 mV and 1 nA, respectively (11). Here we examined in rat slice cultures how EPSPs generated at mossy fiber synapses are processed in CA3 pyramidal cell dendrites, and evaluated whether subthreshold synaptic responses evoked by mossy fiber stimulation can act as instructive signals to induce plasticity at the pyramidal cell synapses forming the CA3 recurrent network.     

Comparison of the isomerization mechanisms of human melanopsin and invertebrate and vertebrate rhodopsins

Comparative modeling and ab initio multiconfigurational quantum chemistry are combined to investigate the reactivity of the human nonvisual photoreceptor melanopsin. It is found that both the thermal and photochemical isomerization of the melanopsin 11-cis retinal chromophore occur via a space-saving mechanism involving the unidirectional, counterclockwise twisting of the =C11H-C12H= moiety with respect to its Lys340-linked frame as proposed by Warshel for visual pigments [Warshel A (1976) Nature 260(5553):679–683]. A comparison with the mechanisms documented for vertebrate (bovine) and invertebrate (squid) visual photoreceptors shows that such a mechanism is not affected by the diversity of the three chromophore cavities. Despite such invariance, trajectory computations indicate that although all receptors display less than 100 fs excited state dynamics, human melanopsin decays from the excited state ∼40 fs earlier than bovine rhodopsin. Some diversity is also found in the energy barriers controlling thermal isomerization. Human melanopsin features the highest computed barrier which appears to be ∼2.5 kcal mol⁻¹ higher than that of bovine rhodopsin. When assuming the validity of both the reaction speed/quantum yield correlation discussed by Warshel, Mathies and coworkers [Weiss RM, Warshel A (1979) J Am Chem Soc 101:6131–6133; Schoenlein RW, Peteanu LA, Mathies RA, Shank CV (1991) Science 254(5030):412–415] and of a relationship between thermal isomerization rate and thermal activation of the photocycle, melanopsin turns out to be a highly sensitive pigment consistent with the low number of melanopsin-containing cells found in the retina and with the extraretina location of melanopsin in nonmammalian vertebrates.For a long time it was assumed that the human retina contains only two types of photoreceptor cells: the rods and cones responsible for dim-light and daylight vision, respectively. However, recent studies have revealed the existence of a small number of intrinsically photosensitive retinal ganglion cells (ipRGCs) that regulate nonvisual photoresponses (1). ipRGCs express an atypical opsin-like protein named melanopsin (2, 3) which plays a role in the regulation of unconscious visual reflexes and in the synchronization of endogenous physiological responses to the dawn/dusk cycle (circadian rhythms) (4, 5).Melanopsins are unique among vertebrate photoreceptors because their amino acid sequence shares greater similarity to invertebrate than vertebrate rhodopsin (i.e., the photoreceptor of rods) (6, 7). Like rhodopsins, melanopsins feature an up–down bundle architecture of seven transmembrane α-helices incorporating the 11-cis isomer of retinal as a covalently bound protonated Schiff base (PSB11 in Fig. 1A). Light-induced (i.e., photochemical) isomerization of PSB11 to its all-trans isomer (PSBAT) triggers an opsin conformational change that, ultimately, activates the receptor and signaling cascade (8, 9). However, similar to invertebrate and in contrast to vertebrate rhodopsins, melanopsins are bistable (10). Indeed, although vertebrate rhodopsins need a retinoid cycle (11) to regenerate PSB11, melanopsins have an intrinsic light-driven chromophore regeneration function via PSBAT back-isomerization. Furthermore, past studies have shown that melanopsins use an invertebrate-like signal transduction cascade (12).Open in a separate windowFig. 1.PSB11 chromophore reactivity. (A) Chromophore structure and isomerization to PSBAT. (B) Schematic representation of the photochemical (full arrows) and thermal (dashed arrows) isomerization paths. The CI is located energetically above the TS, features a different geometrical structure, and drives a far-from-equilibrium process. ΔE_S1-S0, τ_cis→trans, and Ea^T (in red) are the fundamental quantities computed in the present work.Melanopsins are held responsible for photoentrainment, using the changes of irradiance and spectral composition to adjust the circadian rhythm (13). The different studies carried out so far on melanopsin light sensitivity do not lead to consistent results. Although Do et al. (14) argue that ipRGCs work at extremely low irradiation intensities showing a single-photon response larger than rods, Ferrer et al. (15) conclude that the melanopsin has a reduced sensitivity relative to visual pigments. On the other hand, these photoreceptors would be expected to display high light sensitivity (14). In the vertebrate retina their density is 10⁴ times lower than that of rhodopsins. Moreover, the receptor is not confined in a dedicated cellular domain such as the outer segment of rods and cones, resulting in a ipRGCs photon capture more than 10⁶-fold lower than that of rods and cones per unit of retina illumination. A high sensitivity of melanopsins would also be consistent with their presence in extraretina locations such as in pineal complex, deep brain, and derma of nonmammalian vertebrates (e.g., amphibian) (16–18). The amount of light that can penetrate into such regions is limited and enriched in the red component due to light scattering by the surrounding tissues (14).The molecular-level understanding of the primary light response of melanopsin is a prerequisite for the comprehension of more complex properties such as its activation and sensitivity. Despite numerous studies carried out since its discovery (16), there is presently little information on the molecular mechanism of melanopsin activation. The common PSB11 chromophore of melanopsins and rhodopsins does not guarantee that the same mechanism operates in both photoreceptors. This not only concerns light-induced activation but also thermal activation: a process whose rate limits the photoreceptor light sensitivity and that is currently associated with thermal, rather than photochemical, PSB11 isomerization (19–24).The mechanism of light-induced PSB11 isomerization in vertebrate rhodopsins has been extensively investigated. Spectroscopic studies have shown that in bovine rhodopsin (Rh) the isomerization occurs on a subpicosecond timescale (25–27). Moreover, the observation of ground state (S₀) vibrational coherence (28) is consistent with a direct transfer of the excited state (S₁) population to the photoproduct (Fig. 1B) passing through a conical intersection (CI). Such a path has been located along the S₁ potential energy surface by constructing a multiconfigurational quantum chemistry (MCQC) based computer model of the photoreceptor (29–31) and spectroscopically supported by probing in the infrared (31). More recently (32), the same computer model has been used to map the Rh thermal isomerization path (Fig. 1B) providing information on the transition states controlling the reaction.Here we present a computational study focusing on the mechanism of photochemical and thermal isomerization of human melanopsin (hMeOp). This would require the construction of a computer model of hMeOp starting from the receptor crystal structure. However, the lack of hMeOp crystallographic data does not allow the use of the protocol previously applied in Rh studies. The significant sequence similarity between squid rhodopsin (sqRh), whose crystal structure is available (PDB code: 2Z73) (33), and hMeOp (40%, SI Appendix, Fig. S1) provides the fundamentals for constructing a structural model of hMeOp at a significant atomic resolution. Building on a study by Batista and coworkers (34) on murine melanopsin, we combine comparative modeling of hMeOp with MCQC to construct a quantum mechanics/molecular mechanics (QM/MM) computer model capable of simulating the photochemical and thermal isomerization reactions of hMeOp. The results are then compared with those found using Rh and sqRh models constructed using the same protocol. Such a comparison is expected to provide information on the differences in spectral and functional properties of these evolutionary distant pigments. As we will show below, the models indicate that hMeOp has a faster photochemical isomerization dynamics and a higher thermal isomerization barrier than both Rh and sqRh.     

Reliability and Responsiveness of NutriQoL<Superscript>®</Superscript> Questionnaire

Introduction

NutriQoL^® (Nestlé Health Science, Vevay, Switzerland) is a questionnaire developed to assess the health-related quality-of-life (HRQoL) of patients with home enteral nutrition (HEN) irrespective of their underlying condition and route of administration. The aim of this work is assessing the questionnaire’s reliability and responsiveness to change.

Methods

Two cohorts of patients with HEN and their primary caregivers were enrolled to assess reliability and responsiveness, respectively. All participants had to be 18 years of age or older, without mental deterioration (≤3 or 4 errors in the Pfeiffer’s test) and with sufficient functional status (>40 points on Karnovsky’s performance status scale). When the patients’ ability to respond to the questionnaire was impaired due to underlying disease, their caregivers answered on their behalf. NutriQoL was administered in two and three visits to reliability and responsiveness cohorts, respectively. Test–retest reliability and internal consistency were assessed by the intra-class correlation coefficient (ICC) and the Cronbach’s α, respectively. Responsiveness was evaluated by standardized effect size and standardized response mean between basal visit and third visit. Finally, the minimal clinically important difference (MCID) was estimated.

Results

A total of 54 and 86 participants were recruited to the reliability and responsiveness cohort, respectively. Thirty-five caregivers were selected to assess the inter-observer reliability. ICC values confirmed the good reproducibility level (ICC >0.75) of the questionnaire in both “physical functioning and activities of daily living” and “social life” domains and total score. The assessment of internal consistency in both domains of the questionnaire showed good internal consistency in visit 2. ICC showed the excellent agreement level between caregiver and patient in the global NutriQoL score. Finally, patients classified as having a minimal change in their health reported a mean (standard deviation) MCID in NutriQoL score of 0.63 (11.51).

Conclusion

NutriQoL is a reliable and unique instrument to measure the HRQoL in HEN patients. NutriQoL detects changes in the health status of the patient. Nevertheless, further research is needed to determine the full extent of the questionnaire responsiveness.

     

Brain activation of lower extremity movement in chronically impaired stroke survivors

Lower extremity paresis poses significant disability to chronic stroke survivors. Unlike for the upper extremity, cortical adaptations in networks controlling the paretic leg have not been characterized after stroke. Here, the hypotheses are that brain activation associated with unilateral knee movement in chronic stroke survivors is abnormal, depends on lesion location, and is related to walking ability. Functional magnetic resonance imaging of unilateral knee movement was obtained in 31 patients 26.9 months (mean, IQ range: 11.3-68.1) after stroke and in 10 age-matched healthy controls. Strokes were stratified according to lesion location. Locomotor disability (30 ft walking speed) did not differ between patient groups (9 cortical, 12 subcortical, 10 brainstem lesions). Significant differences in brain activation as measured by voxel counts in 10 regions of interest were found between controls and patients with brainstem (P = 0.006) and cortical strokes (P = 0.002), and between subcortical and cortical patients (P = 0.026). Statistical parametric mapping of data per group revealed similar activation patterns in subcortical patients and controls with recruitment of contralateral primary motor cortex (M1), supplementary motor area (SMA), and bilateral somatosensory area 2 (S2). Cortical recruitment was reduced in brainstem and cortical stroke. Better walking was associated with lesser contralateral sensorimotor cortex activation in brainstem, but stronger recruitment of ipsilateral sensorimotor and bilateral somatosensory cortices in subcortical and cortical patients, respectively. A post hoc comparison of brainstem patients with and without mirror movements (50%) revealed lesser recruitment of ipsilateral cerebellum in the latter. Subcortical patients with mirror movements (58%) showed lesser bilateral sensorimotor cortex activation. No cortical patient had mirror movements. The data reveal adaptations in networks controlling unilateral paretic knee movement in chronic stroke survivors. These adaptations depend on lesion location and seem to have functional relevance for locomotion.