Duchenne muscular dystrophy (DMD) is a devastating disease affecting about 1 out of 5000 male births and caused by mutations in the dystrophin gene. Genome editing has the potential to restore ...expression of a modified dystrophin gene from the native locus to modulate disease progression. In this study, adeno-associated virus was used to deliver the clustered regularly interspaced short palindromic repeats (CRISPR)–Cas9 system to the mdx mouse model of DMD to remove the mutated exon 23 from the dystrophin gene. This includes local and systemic delivery to adult mice and systemic delivery to neonatal mice. Exon 23 deletion by CRISPR-Cas9 resulted in expression of the modified dystrophin gene, partial recovery of functional dystrophin protein in skeletal myofibers and cardiac muscle, improvement of muscle biochemistry, and significant enhancement of muscle force. This work establishes CRISPR-Cas9–based genome editing as a potential therapy to treat DMD.
Duchenne muscular dystrophy (DMD) is a monogenic disorder and a candidate for therapeutic genome editing. There have been several recent reports of genome editing in preclinical models of Duchenne ...muscular dystrophy
, however, the long-term persistence and safety of these genome editing approaches have not been addressed. Here we show that genome editing and dystrophin protein restoration is sustained in the mdx mouse model of Duchenne muscular dystrophy for 1 year after a single intravenous administration of an adeno-associated virus that encodes CRISPR (AAV-CRISPR). We also show that AAV-CRISPR is immunogenic when administered to adult mice
; however, humoral and cellular immune responses can be avoided by treating neonatal mice. Additionally, we describe unintended genome and transcript alterations induced by AAV-CRISPR that should be considered for the development of AAV-CRISPR as a therapeutic approach. This study shows the potential of AAV-CRISPR for permanent genome corrections and highlights aspects of host response and alternative genome editing outcomes that require further study.
Preexisting neutralizing antibodies (NAbs) against adeno-associated viruses (AAVs) pose a major, unresolved challenge that restricts patient enrollment in gene therapy clinical trials using ...recombinant AAV vectors. Structural studies suggest that despite a high degree of sequence variability, antibody recognition sites or antigenic hotspots on AAVs and other related parvoviruses might be evolutionarily conserved. To test this hypothesis, we developed a structure-guided evolution approach that does not require selective pressure exerted by NAbs. This strategy yielded highly divergent antigenic footprints that do not exist in natural AAV isolates. Specifically, synthetic variants obtained by evolving murine antigenic epitopes on an AAV serotype 1 capsid template can evade NAbs without compromising titer, transduction efficiency, or tissue tropism. One lead AAV variant generated by combining multiple evolved antigenic sites effectively evades polyclonal anti-AAV1 neutralizing sera from immunized mice and rhesus macaques. Furthermore, this variant displays robust immune evasion in nonhuman primate and human serum samples at dilution factors as high as 1:5, currently mandated by several clinical trials. Our results provide evidence that antibody recognition of AAV capsids is conserved across species. This approach can be applied to any AAV strain to evade NAbs in prospective patients for human gene therapy.
The glymphatic system is a brain-wide clearance pathway; its impairment contributes to the accumulation of amyloid-β. Influx of cerebrospinal fluid (CSF) depends upon the expression and perivascular ...localization of the astroglial water channel aquaporin-4 (AQP4). Prompted by a recent failure to find an effect of
knock-out (KO) on CSF and interstitial fluid (ISF) tracer transport, five groups re-examined the importance of AQP4 in glymphatic transport. We concur that CSF influx is higher in wild-type mice than in four different
KO lines and in one line that lacks perivascular AQP4 (
KO). Meta-analysis of all studies demonstrated a significant decrease in tracer transport in KO mice and rats compared to controls. Meta-regression indicated that anesthesia, age, and tracer delivery explain the opposing results. We also report that intrastriatal injections suppress glymphatic function. This validates the role of AQP4 and shows that glymphatic studies must avoid the use of invasive procedures.
While recent forays to bring postcolonial studies in conversation with American Indian studies and to frame indigenous Latinx communities within a settler colonial paradigm gesture toward new ...engagements with the global South, these efforts remain focused on North America.1 The term itself is difficult to translate.4 We are left with the quandary of debating who is a settler....few scholars in Latin America are familiar with this body of work because of a lack of Spanish and Portuguese translations.Efforts to distinguish regimes of colonialism in the Americas by their method of dispossession, as rooted in either land or labor expropriation, ends up reproducing binaries (land/labor, settler/native, Latinx/Latin American) that mask articulations spanning imperial and colonial regimes.5 The emphasis on binaries risks reproducing a monolithic, self-contained theory of settler colonialism lacking historical and relational specificity, the very project initially challenged by Patrick Wolfe.6 We advance an analytic project that acknowledges the multiple iterations of settler colonial projects that have been instantiated within and beyond postcolonizing societies.7 Scholars of Latin America have relied on theories of “coloniality” to account for the production of racialized power as a hegemonic and historical project.8 Coloniality is a cognitive mode of power based on a “new perspective of knowledge within which non-Europe was the past, and because of that inferior, if not always primitive.”To challenge the “brute binary division” of models of coloniality, scholars like Lisa Lowe and Evelyn Nakano Glenn propose a comparative and relational model that illuminates colonialism’s spatial and temporal projects.12 Cognizant of the diverse experiences and histories that have shaped indigenous communities in the Americas, the essays connect these distinctive geographies, histories, and intimacies by identifying and historicizing settler colonial projects in Mexico, Guatemala, Honduras, and the United States.
Treatment for peripheral nerve injuries includes the use of autografts and nerve guide conduits (NGCs). However, outcomes are limited, and full recovery is rarely achieved. The use of nerve scaffolds ...as a platform to surface immobilize neurotrophic factors and deliver locally is a promising approach to support neurite and nerve outgrowth after injury. We report on a bioactive surface using functional amine groups, to which heparin binds electrostatically. X‐ray photoelectron spectroscopy analysis was used to characterize the presence of nitrogen and sulfur. Nerve growth factor (NGF) and brain‐derived neurotrophic factor (BDNF) were bound by electrostatic interaction to heparin, and the release profile evaluated by enzyme‐linked immunosorbent assay, which showed that ca. 1% of NGF was released from each of the bioactive surface within 7 days. Furthermore, each surface showed a maximum release of 97% of BDNF. Neurotrophin release on neurite outgrowth was evaluated by primary dorsal root ganglion with a maximum neurite growth response in vitro of 1,075 µm detected for surfaces immobilized with NGF at 1 ng/ml. In summary, the study reports on the design and construction of a biomimetic platform to deliver NGF and BDNF using physiologically low concentrations of neurotrophin. The platform is directly applicable and scalable for improving the regenerative ability of existing NGCs and scaffolds.
Biomaterial surfaces immobilized with nerve growth factor (NGF; 1 ng/ml) stimulate neurite outgrowth. Biomimetic surfaces were fabricated using positively charged amine groups to which heparin was bound electrostatically. NGF and brain‐derived neurotrophic factor (BDNF) were immobilized to heparin electrostatically. Both neurotrophins stimulated neurite outgrowth from primary dorsal root ganglion (DRG) neuronal cells after 7 days of culture, with maximum responses from NGF surfaces preloaded at 1 ng/ml.
The objective is to conduct a longitudinal analysis of the effects of the pandemic and alarm situation on the mental health of the general population at three points in time: two weeks after ...beginning the confinement, after a month, and after two months, when the lockdown was lifted and the country returned to the new normality.
The evaluations were carried out by means of an online survey, with a sample of 3480 persons in the first data collection and 1041 and 569 persons in the successive evaluation periods. The presence of depressive symptoms, anxiety and posttraumatic stress disorder (PTSD) was evaluated by means of screening tests. Sociodemographic data, Covid-19 variables, loneliness, psychological well-being, social support, discrimination and a sense of belonging, were collected.
Depressive symptoms increased significantly throughout the confinement, decreasing at the last assessment but not dropping to previous levels. In anxiety, there are no significant changes between the three evaluations, but a downward trend can be seen over time. Regarding the symptomatology of PTSD, a downward trend is observed throughout the three evaluations, with significantly lower scores between the first and third assessments. The different regression models developed reveal the importance of perceived loneliness and spiritual well-being as the main predictors of mental health, as well as the importance of the lower age for depression and the female gender for anxiety and PSTD.
This research shows that the pandemic has had a negative impact on our mental health, which still does not seem to be at pre-crisis levels, although it has improved as the emergency situation subsides. These results underline the importance of paying greater attention to mental health, and reveal key variables such as spiritual well-being and perceived loneliness in which to intervene from different care services, as well as younger people and women as vulnerable groups on which to focus more attention.
•Covid-19 had a strong impact on the mental health throughout the confinement.•Depression decreased over time, although it did not reach previous levels.•Anxiety remains but a downward trend is observed, and PSTD decreases significantly.•Women and youth have been most affected by the pandemic.•Loneliness and well-being are key variables in predicting mental health impact.
Summary
Background
Frontal fibrosing alopecia (FFA) is a chronic cicatricial alopecia with an increasing incidence and unknown aetiology.
Aim
To identify possible environmental and hormonal factors ...related to FFA.
Methods
We conducted a multicentre case–control study paired by sex and age, and recruited 664 women (335 cases and 329 controls) and 106 men (20 cases and 86 controls). Study subjects completed an exhaustive questionnaire enquiring about pharmacological, environmental, hormonal, social, job exposure, lifestyle, drugs and diet factors to which they were exposed at least 5 years prior to the onset of the disease.
Results
For women, there was a statistical association between alopecia and history of pregnancy (OR = 1.6; 95% CI 1.06–2.41), use of facial sunscreen (OR = 1.6; 95% CI 1.06–2.41) and hormone replacement therapy (HRT) (OR = 1.76; 95% CI 1.11–2.8) or raloxifene (no controls exposed therefore OR was not calculated), exposure to alkylphenolic compounds (OR = 1.48; 95% CI 1.05–2.08), and presence of rosacea (OR = 1.91; 95% CI 1.07–3.39), lichen planus pigmentosus (LPP) (OR = 5.14; 95% CI 1.11–23.6) or hypothyroidism (OR = 1.73; 95% CI 1.11–2.69). For men, there was a statistical association between alopecia and use of facial sunscreens (OR = 11.6; 95% CI 1.7–80.9) or antiageing creams (OR = 1.84; 95% CI 1.04–3.23).
Conclusions
FFA seems to be associated with hormonal exposure (pregnancy, HRT and raloxifene), comorbidities (hypothyroidism, LPP and rosacea) and environmental factors (facial sunscreens, antiageing creams and occupational exposure). Further research is required to analyse the exact mechanism in which these environmental factors participate in the development of this alopecia.
Travelling across several time zones requires a fast adjustment of the circadian system and the differential adjustment speeds of organs and systems results in what is commonly referred as jet lag. ...During this transitory state of circadian disruption, individuals feel discomfort, appetite loss, fatigue, disturbed sleep and deficient performance of multiple tasks. We have demonstrated that after a 6‐h phase advance of the light–dark cycle (LD) scheduled food in phase with the new night onset can speed up re‐entrainment. In this study, we explored the possible mechanisms underlying the fast re‐entrainment due to the feeding schedule. We focused on first‐ and second‐order structures that provide metabolic information to the suprachiasmatic nucleus (SCN). We compared (i) control rats without change in LD cycle; (ii) rats exposed to a 6‐h phase advance of the LD cycle with food ad libitum; and (iii) rats exposed to the 6‐h phase advance combined with food access in phase with the new night. We found an immediate synchronizing effect of food on stomach distention and on c‐Fos expression in the nucleus of the solitary tract, arcuate nucleus of the hypothalamus, dorsomedial hypothalamic nucleus and paraventricular nucleus. These observations indicate that in a model of jet lag, scheduled feeding can favour an immediate shift in first‐ and second‐order relays to the SCN and that by keeping feeding schedules coupled to the new night, a fast re‐entrainment may be achieved by shifting peripheral and extra‐SCN oscillations.
Individual exposed to a sudden shift of the light cycle requires 7–10 cycles for re‐entrainment. Scheduled food coinciding with the new night onset speeds up re‐entrainment. Here, we show that scheduled food in phase with the new night produced an immediate shift in brain areas that receive and process food‐elicited signals (red symbols), contrasting with brain regions that receive direct light input from the retina (IGL and SCN). These observations indicate that effects of scheduled feeding are mediated by first and second‐order relays to the SCN.
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