MyoSight—semi-automated image analysis of skeletal muscle cross sections

Background Manual analysis of cross-sectional area, fiber-type distribution, and total and centralized nuclei in skeletal muscle cross sections is tedious and time consuming, necessitating an accurate, automated method of analysis. While several excellent programs are available, our analyses of skeletal muscle disease models suggest the need for additional features and flexibility to adequately describe disease pathology. We introduce a new semi-automated analysis program, MyoSight, which is designed to facilitate image analysis of skeletal muscle cross sections and provide additional flexibility in the analyses. Results We describe staining and imaging methods that generate high-quality images of immunofluorescent-labelled cross sections from mouse skeletal muscle. Using these methods, we can analyze up to 5 different fluorophores in a single image, allowing simultaneous analyses of perinuclei, central nuclei, fiber size, and fiber-type distribution. MyoSight displays high reproducibility among users, and the data generated are in close agreement with data obtained from manual analyses of cross-sectional area (CSA), fiber number, fiber-type distribution, and number and localization of myonuclei. Furthermore, MyoSight clearly delineates changes in these parameters in muscle sections from a mouse model of Duchenne muscular dystrophy (mdx). Conclusions MyoSight is a new program based on an algorithm that can be optimized by the user to obtain highly accurate fiber size, fiber-type identification, and perinuclei and central nuclei per fiber measurements. MyoSight combines features available separately in other programs, is user friendly, and provides visual outputs that allow the user to confirm the accuracy of the analyses and correct any inaccuracies. We present MyoSight as a new program to facilitate the analyses of fiber type and CSA changes arising from injury, disease, exercise, and therapeutic interventions.

In vivo amygdala nuclei volumes in schizophrenia and bipolar disorders

Background: Abnormalities in amygdala volume are well-established in schizophrenia and commonly reported in bipolar disorders. However, the specificity of volumetric differences in individual amygdala nuclei is largely unknown. Methods: Patients with schizophrenia disorders (SCZ, n=452, including schizophrenia, schizoaffective and other psychotic disorders, mean age 30.7±9.2 (SD), females 44.4%), bipolar disorders (BP, n=316, including bipolar I and II, 33.7±11.4, 58.5%) and healthy controls (n=753, 34.1±9.1, 40.9%) underwent T1-weighted magnetic resonance imaging. Total amygdala and nuclei volumes as well as intracranial volume (ICV) were estimated with Freesurfer (v6.0.0). Analysis of covariance and multiple linear regression models, adjusting for age, age2, ICV and sex, were fitted to examine diagnostic group and subgroup differences in volume, respectively. Results: Bilateral total amygdala and all nuclei volumes, except the medial and central nuclei, were significantly smaller in patients relative to controls. The largest effect sizes were found for the basal nucleus, accessory basal nucleus and cortico-amygdaloid transition area (partial η2 > 0.02). The diagnostic subgroup analysis showed that reductions in amygdala nuclei volume were most widespread in schizophrenia, with the lateral, cortical, paralaminar and central nuclei being solely reduced in this disorder. The right accessory basal nucleus was marginally smaller in SCZ relative to BP (t = 2.32, p = 0.05). Conclusions: Our study is the first to demonstrate distinct patterns of amygdala nuclei volume reductions in a well-powered sample of patients with schizophrenia and bipolar disorders. Volume differences in the basolateral complex (lateral, basal, accessory basal nuclei) may be putative neuroimaging markers for differentiating schizophrenia and bipolar disorders.

Diffuse Cutaneous Mastocytosis (Pigmented Maculopapular Cutaneous Mastocytosis) in a Cat

Diffuse cutaneous mastocytosis was diagnosed in a 6-year-old, indoor, neutered female domestic European shorthair cat. Marked pruritus located mainly on the head and neck was noticed in the cat and in this area the animal had developed alopecia, crusts, and plaques. Histologically, monomorphic mast cells were found in the superficial dermis and around the hair follicles. Mast cells were well differentiated, with central nuclei and granular cytoplasm, with metachromatic granules which stained positively with Toluidine blue stain. The animal was successfully treated with oclacitinib at a dose of 1 mg/kg, twice a day per os.

Displaced Myonuclei in Cancer Cachexia Suggest Altered Innervation

An idiopathic myopathy characterized by central nuclei in muscle fibers, a hallmark of muscle regeneration, has been observed in cancer patients. In cancer cachexia skeletal muscle is incapable of regeneration, consequently, this observation remains unaccounted for. In C26-tumor bearing, cachectic mice, we observed muscle fibers with central nuclei in the absence of molecular markers of bona fide regeneration. These clustered, non-peripheral nuclei were present in NCAM-expressing muscle fibers. Since NCAM expression is upregulated in denervated myofibers, we searched for additional makers of denervation, including AchRs, MUSK, and HDAC. This last one being also consistently upregulated in cachectic muscles, correlated with an increase of central myonuclei. This held true in the musculature of patients suffering from gastrointestinal cancer, where a progressive increase in the number of central myonuclei was observed in weight stable and in cachectic patients, compared to healthy subjects. Based on all of the above, the presence of central myonuclei in cancer patients and animal models of cachexia is consistent with motor neuron loss or NMJ perturbation and could underlie a previously neglected phenomenon of denervation, rather than representing myofiber damage and regeneration in cachexia. Similarly to aging, denervation-dependent myofiber atrophy could contribute to muscle wasting in cancer cachexia.

Connectional heterogeneity in the mouse auditory corticocollicular system

The auditory cortex (AC) sends long-range projections to virtually all subcortical structures important for hearing. One of the largest and most complex of these - the projection between AC and inferior colliculus (IC, the corticocollicular pathway) - has attracted attention due to its potential to alter IC response properties. The corticocollicular pathway comprises a component originating from layer 5, but recent evidence suggests a significant contribution from deep layer 6, constituting 25% of corticocollicular neurons in mouse. The functions of layer-specific corticocollicular projections are poorly understood. Here, using a combination of tracers and in vivo imaging, we observed that layer 5 and layer 6 corticocollicular neurons differ in their cortical areas of origin, as well as IC termination patterns. Layer 5 corticocollicular neurons are concentrated in primary AC areas while layer 6 corticocollicular neurons emanate from broad auditory and non-auditory areas of temporal cortex. In addition, layer 5 projects to three IC subdivisions with axo-somatic terminals in the central nucleus, while layer 6 projects to non-central nuclei and targets the most superficial layers. These findings suggest that layer 5 corticocollicular neurons form a direct connection between primary AC and IC while the layer 6 projection is more diffusely organized and carries non-auditory information to modulate IC.

Resveratrol Decreases Oxidative Stress by Restoring Mitophagy and Improves the Pathophysiology of Dystrophin-Deficient mdx Mice

We previously showed that treatment with resveratrol (3,5,4′-trihydroxy-trans-stilbene), an activator of the NAD+-dependent deacetylase SIRT1 at 4 g/kg food for 32 weeks, significantly decreased the muscular reactive oxygen species (ROS) levels and ameliorated the pathology of mdx mice, an animal model of Duchenne muscular dystrophy (DMD). Here, we treated mdx mice with various doses of resveratrol (0.04, 0.4, and 4 g/kg food) for 56 weeks and examined the effects on serum creatine kinase levels and physical activities. Because resveratrol promotes autophagy, we also investigated whether autophagy including mitochondrial autophagy (mitophagy) is involved in resveratrol’s effects. Autophagy/mitophagy-related genes and autophagic flux were downregulated in the muscle of mdx mice, and these phenomena were reversed by resveratrol with significant ROS reduction. Resveratrol at 4 g/kg food reduced the number of immature myofibers containing central nuclei and fine fibers < 400 μm2 and increased that of thicker myofibers in the quadriceps, suggesting that resveratrol decreased myofiber wasting and promoted muscular maturation. Accordingly, resveratrol at 0.4 g/kg food reduced the creatine kinase levels to one-third of those in untreated mdx mice and significantly increased the animals’ physical activities. In C2C12 myoblast cells, resveratrol promoted mitophagy and eliminated mitochondria containing high superoxide levels. The clearance of damaged mitochondria and ROS reduction by resveratrol was completely suppressed by an autophagy inhibitor (chloroquine) and by knocking down Atg5 or Pink1, essential genes for autophagy and mitophagy, respectively. Thus, resveratrol is a potential therapeutic agent for DMD, and the clearance of damaged mitochondria probably contributes to its action.

Meanings attributed to the perception of to be-ostomized-in-the-world

Objective: To understand the meanings of the experience of to be-ostomized-in-the-world through Merleau-Ponty phenomenology. Method: Descriptive study with a qualitative approach, in the merleau-ponty phenomenological perspective, conducted through interviews with ten people with intestinal stomas assisted by the Association of Ostomized of the State of Ceara. The IRaMuTeQ software was used. Similarity analysis and word cloud were adopted. Results: The words ‘person’, ‘much’, ‘how’, ‘see’ and ‘bag’ was identified by discourse analysis, which gave rise to ‘say’, as central nuclei. The ‘person’, as be-ostomized-in-the-world, perceives itself as’ being-sick ‘that’ reluctantly ‘senses-motor sensations (feces and odors) that cause’ repulsiveness’, marked by the presence of the ‘ (stoma and bag). The words most often in the cloud were ‘bag’, ‘life’, ‘God’, ‘people’ and ‘surgery’. The collector bag evidences ‘abnormality’ and bodily imperfection. Conclusion: The perception of be-ostomized-in-the-world is marked by the presence of the collector bag of stoma, being perceived as new element of its body.