CRISPR-based gene editing enables FOXP3 gene repair in IPEX patient cells.

The prototypical genetic autoimmune disease is immune dysregulation polyendocrinopathy enteropathy X-linked (IPEX) syndrome, a severe pediatric disease with limited treatment options. IPEX syndrome is caused by mutations in the forkhead box protein 3 (FOXP3) gene, which plays a critical role in immune regulation. As a monogenic disease, IPEX is an ideal candidate for a therapeutic approach in which autologous hematopoietic stem and progenitor (HSPC) cells or T cells are gene edited ex vivo and reinfused. Here, we describe a CRISPR-based gene correction permitting regulated expression of FOXP3 protein. We demonstrate that gene editing preserves HSPC differentiation potential, and that edited regulatory and effector T cells maintain their in vitro phenotype and function. Additionally, we show that this strategy is suitable for IPEX patient cells with diverse mutations. These results demonstrate the feasibility of gene correction, which will be instrumental for the development of therapeutic approaches for other genetic autoimmune diseases.

Pinealectomy causes hippocampal CA1 and CA3 cell loss: reversal by melatonin supplementation.

This experiment determined if the loss of endogenous melatonin via pinealectomy affected rat CA1 and CA3 pyramidal neuron numbers over a 20-month span. Since pinealectomy eliminates many neurohormones, some rats received daily melatonin supplementation to determine if this would reverse its effects. CA1 pyramidal cells were significantly reduced between 2 and 4 months after pinealectomy. CA3 loss was evident at 2 months post-pinealectomy. Melatonin replacement in the drinking water prevented these effects and seemingly promoted the genesis of CA1 cells. Analyses of hippocampal thiobarbituric acid reactive substances (TBARS) levels at 3 and 20 months post-surgery, revealed no significant group differences in lipid peroxidation. However, hippocampal TBARS were higher at 20 than at 3 months in all groups. Pinealectomized rats exhibited a significantly higher ratio of reduced to oxidized glutathione at 3 months but not 20 months, when compared to the sham and melatonin-supplemented rats. This suggests that pinealectomy caused oxidative stress and a subsequent compensatory change in the glutathione system. These results indicate that endogenous melatonin is neuroprotective.

Pinealectomy: behavioral and neuropathological consequences in a chronic cerebral hypoperfusion model.

This experiment determined if pinealectomy (PX) affects the consequences of chronic, moderate brain ischemia. Rats were pinealectomized at 25 days of age and trained at 9 months on a tactile radial maze. They then underwent permanent occlusion of the common carotid arteries (2VO) or sham surgery, followed by maze retraining and then neurohistological assessment at 16 months. Combined PX + 2VO rats committed more working memory errors on the maze. 2VO itself caused a 10% reduction in hippocampal CA1 pyramidal cell number. PX alone caused a 21% reduction. Combined PX and 2VO caused the greatest reduction (32%) of CA1 cells. Similar results were seen for CA4. PX also increased glial fibrillary acidic protein immunoreactivity in both CA1 and CA4. Thus PX not only augmented the consequences of chronic brain ischemia but notably, PX itself caused hippocampal damage. These effects seemed not to result from the small cortical lesion caused by the PX procedure. The results are consistent with the hypothesis that endogenous melatonin is a neuroprotectant in the aging brain.

Neurotrophic factor structures reveal clues to evolution, binding, specificity, and receptor activation.

The neutrophin family, the glial-derived neurotrophic factor family, and the ciliary neurotrophic factor are the best described growth factors specific for developing neurons and neutral crest cells. As might be expected for regulatory molecules of the complex central and peripheral nervous system, these factors show considerable receptor specificity and cross-talk. Thanks to a decade of intense research by numerous laboratories, the structures of many of these factors are now available. This review discusses the structural bases of receptor binding, specificity, and activation in each of these systems. Using structure-based sequence alignments, the evolutionary implications of these molecules and their receptors are discussed, followed by suggestions for further directions for research on the structure and function of these neurotrophic factors.

Functional studies of the BTB domain in the Drosophila GAGA and Mod(mdg4) proteins.

The BTB/POZ (BTB) domain is an approximately 120 residue sequence that is conserved at the N-terminus of many proteins in both vertebrates and invertebrates. We found that the protein encoded by a lethal allele of the Drosophila modifier of mdg4 [mod(mdg4)] gene has two mutated residues in its BTB domain. The identities of the residues at the positions of these mutations are highly conserved in the BTB domain family of proteins, and when the corresponding mutations were engineered into the BTB domain-containing GAGA protein, the activity of GAGA as a transcription activator in a transient transfection assay was severely reduced. The functional equivalence of the BTB domains was established by showing that the BTB domain of the mod(mdg4) protein can effectively substitute for that of GAGA.

Pediatric myocardial infarction after racemic epinephrine administration.

Myocardial infarction is a previously unreported complication of treatment with racemic epinephrine that is used commonly in the emergency department for severe respiratory distress in bronchiolitis or croup syndrome. We describe a pediatric patient who presented with the croup syndrome and severe respiratory distress that required multiple doses of nebulized racemic epinephrine in the emergency department. The patient developed ventricular tachycardia and mild chest discomfort during one treatment, which resolved spontaneously on discontinuation of the nebulization. Persistently abnormal electrocardiograms and elevated creatine phosphokinase MB isoenzyme (CPK-MB) levels suggested a myocardial infarction had occurred. Subsequent echocardiography, cardiac catheterization, and angiography revealed an anatomically normal heart with normal coronary circulation; however, a stress nuclear study showed a small myocardial infarct. The significance of this previously unreported complication of racemic epinephrine is discussed, along with recommendations for proper use in the emergency department.

Crystal structure of neurotrophin-3 homodimer shows distinct regions are used to bind its receptors.

Neurotrophin-3 (NT-3) is a cystine knot growth factor that promotes the survival, proliferation, and differentiation of developing neurons and is a potential therapeutic for neurodegenerative diseases. To clarify the structural basis of receptor specificity and the role of neurotrophin dimerization in receptor activation, the structure of the NT-3 homodimer was determined using X-ray crystallography. The orthorhombic crystals diffract to 2.4 A, with dimer symmetry occurring about a crystallographic 2-fold axis. The overall structure of NT-3 resembles that of the other neurotrophins, NGF and BDNF; each protomer forms a twisted four-stranded beta sheet, with three intertwined disulfide bonds. There are notable differences, however, between NT-3 and NGF in the surface loops and in three functionally important regions, shown in previous mutagenesis studies to be critical for binding. One such difference implies that NT-3's binding affinity and specificity depend on a novel hydrogen bond between Gln 83, a residue important for binding specificity with TrkC, and Arg 103, a residue crucial for binding affinity with TrkC. NT-3's extensive dimer interface buries much of the otherwise solvent-accessible hydrophobic surface area and suggests that the dimeric state is stabilized through the formation of this hydrophobic core. A comparison of the dimer interface between the NT-3 homodimer and the BDNF/NT-3 heterodimer reveals similar patterns of hydrogen bonds and nonpolar contacts, which reinforces the notion that the evolutionarily conserved neurotrophin interface resulted from the need for receptor dimerization in signal initiation.

Topological and phenomenological classification of bursting oscillations.

We describe a classification scheme for bursting oscillations which encompasses many of those found in the literature on bursting in excitable media. This is an extension of the scheme of Rinzel (in Mathematical Topics in Population Biology, Springer, Berlin, 1987), put in the context of a sequence of horizontal cuts through a two-parameter bifurcation diagram. We use this to describe the phenomenological character of different types of bursting, addressing the issue of how well the bursting can be characterized given the limited amount of information often available in experimental settings.

Circadian rhythms of beta-endorphin in the aged.

A circadian rhythm for beta-endorphin was observed by means of the cosinor method in aged subjects. The rhythm mesor is a little lower but its amplitude is more extended than in young subjects. The confidence intervals of the acrophases of the two groups are superimposable with a timing in the morning.

[Arrhythmias in the elderly]. / Le aritmie nell'anziano.

A survey was made on a population of 6059 subjects aged more than 60 years with the aim to assess 1. the prevalence of heart arrhythmias and 2. the relationships between arrhythmias and some other ecg alterations. Arrhythmias resulted present in 29.0% of the whole population with a significantly higher prevalence among males (30.7% vs 28.1%, P less than 0.05) and among subjects over 75 years of age (33.2% vs 23.9%, P less than 0.001). Supraventricular extrasystoles (SE, 11.55%), atrial fibrillation (AF, 10.44%) and ventricular extrasystoles (VE, 8.91%) were the most frequent arrhythmias, followed by sinus bradycardia (SB, 2.04%), sinus arrhythmia (SA, 1.35%), atrial flutter (AFL, 1.09%) and junctional rhythms (JR, 0.20%). AF and AFL resulted significantly more frequent among females, whilst SE, VE and SB were more frequent among males. All the above arrhythmias, with the exception of AFL and JR resulted significantly more frequent among subjects over 75. A significantly higher prevalence of ecg signs of left ventricular hypertrophy, ischemia, previous myocardial infarction (MI) and of the so-called "minor" T-wave changes (MTC) was found among the subjects with arrhythmia as compared with those free from rhythm disturbances. Ecg signs of MI and MTC were significantly more frequent among males and MTC were more frequent among females and among subjects over 75. It is concluded that in an old person the presence of an arrhythmia should lead to a careful evaluation of the general and cardiological clinical situation in order to avoid 1. to prescribe an unnecessary and potentially dangerous antiarrhythmic treatment, and 2. to misdiagnose an underlying clinical condition liable to a decisive improvement under adequate treatment.