Comparative Study of Histological and Histochemical Image Processing in Muscle Fibre Sections of Broiler Chicken

Image processing of muscular fibre has significantly improved over the last decades, from light microscopy acquisition to virtual microscopy and from manual segmentation and fibre typing to automatic. The study aims to discuss main tools used in different histoenzymological image processing ‎phases of muscle fibre, which are consecutively, acquisition, segmentation and fibre typing and theirs ‎efficacy in morphometric parameters’ determination. Firstly, the acquisition, optical microscopic image with different magnifications ‎‎(x100, x200, x250 and x400) were compared with virtual slides digitized by Slide Scanner, in ‎muscle fibre sections, for cell number counting. Secondly, the segmentation, three software ‎‎(Fiji «Digitizing pen, Mouse», Image Pro Plus 10 -semi-automatic- and Cytoinformatics LLC -automatic-) were compared ‎for image segmentation quality and fibbers number determination. Thirdly, manual fibre typing with Fiji of segmented images using three software cited previously were performed to calculate the accuracy of morphometric parameter (CSA, perimeter and DMF). Results of acquisition showed that scanner slide have ‎a better resolution and detected a higher number of cells compared to optical microscopic with ‎different magnifications. For this latter, the processing can be performed only with an ‎acceptable resolution, where the number of cells is lesser, which requires unfortunately several ‎repetitions and exhausting work. Our findings regarding segmentation indicate that ‎Cytoinformatics LLC showed the best processing time and the highest quality followed by IP ‎and Fiji. For the last step, morphometric parameter calculation showed the best accuracy using Fiji followed by Cytoinformatics and finally by IP. ‎ The findings of this study suggest that Fiji (semi-automatic) showed the best quality/price ratio (open access software) for segmentation and fibre typing, but time consuming compared to ‎ Image Pro Plus 10 (semi-automatic) and Cytoinformatics LLC‎ (automatic) which are a paid service.

Immunohistochemical markers of reactive skeletal muscle fibres

Although most patients undergo muscle biopsies to elucidate the cause of muscle symptoms (weakess, cramping, etc.), many muscle biopsies show relatively few specific alterations on routine staining. Immunohistochemical methods for muscle fibre typing and characterisation of inflammatory cell infiltrates are now well established but the value of other markers is less well documented. A preliminary study of other potentially useful immunohistochemical markers revealed that muscle biopsies in our hospital often contain CD56 and/or D2-40 positive myofibres. This study was extended to a series of 32 biopsies from adult patients (age 21–81, 12 males 20 females), 11 of which showed only minor changes on routine examination. Most cases contained CD56 positive mature fibres; D2-40 positive muscle fibres were more common in cases of inflammatory myopathy. Five cases with minor changes on routine examination showed CD56 and D2-40 staining of otherwise unremarkable myofibres, which might represent reactive changes.LEARNING OBJECTIVESThis presentation will enable the learner to:1.Describe patterns of immunohistochemical staining in reactive muscle fibres2.Discuss the underlying physiology of reactive muscle fibres

Variation in instrumental meat quality among 15 muscles from 14-month-old sheep and its relationship with fibre typing

The aims of the present study were to describe intermuscular differences in meat-quality traits in 15 young-sheep muscles, and to study the associations between meat quality and fibre typing across all (pooled) muscles as well as in previously selected homogeneous contractile–metabolic groups of muscles (slow-oxidative, intermediate and fast-glycolytic muscles). Meat-quality traits (pH, colour, expressed juice, cooking losses, tenderness and sarcomere length) and fibre typing were evaluated after 24 h of slaughter in 15 muscles from five cross-bred young sheep. Across all the studied muscles, intermuscular differences in some meat-quality traits (pH24, a* and expressed juice) seemed to be mainly explained by muscle oxidative activity, while intermuscular variation in other meat-quality traits (L*, b* and Warner–Bratzler shear force) were mainly explained by differences in fibre sizes. Within fast-glycolytic muscles, larger fast-glycolytic fibres and reduced oxidative activity were generally associated with lower ultimate pH, higher L* values, lower a* values and longer sarcomeres. Within intermediate muscles, larger fast-glycolytic fibres and reduced oxidative activity were generally associated with lower ultimate pH, higher L* values, shorter sarcomeres and reduced meat tenderness. Within slow-oxidative muscles, larger fast-glycolytic fibres and reduced oxidative activity were generally associated with lower amounts of expressed juice, lower a* values and reduced meat tenderness. The present study has contributed to a better understanding of the influence of muscle fibre types on intermuscular meat-quality variation, suggesting that although muscle fibre diversity may explain, at least in part, intermuscular differences in meat quality, these associations can also slightly vary among muscle contractile–metabolic groups.

Deep UV excited muscle cell autofluorescence varies with the fibre type

DUV autofluorescence microspectroscopy allows label free fibre typing in muscles.

Early Changes in Costameric and Mitochondrial Protein Expression with Unloading Are Muscle Specific

We hypothesised that load-sensitive expression of costameric proteins, which hold the sarcomere in place and position the mitochondria, contributes to the early adaptations of antigravity muscle to unloading and would depend on muscle fibre composition and chymotrypsin activity of the proteasome. Biopsies were obtained from vastus lateralis (VL) and soleus (SOL) muscles of eight men before and after 3 days of unilateral lower limb suspension (ULLS) and subjected to fibre typing and measures for costameric (FAK and FRNK), mitochondrial (NDUFA9, SDHA, UQCRC1, UCP3, and ATP5A1), and MHCI protein and RNA content. Mean cross-sectional area (MCSA) of types I and II muscle fibres in VL and type I fibres in SOL demonstrated a trend for a reduction after ULLS (0.05≤P<0.10). FAK phosphorylation at tyrosine 397 showed a 20% reduction in VL muscle (P=0.029). SOL muscle demonstrated a specific reduction in UCP3 content (-23%;P = 0.012). Muscle-specific effects of ULLS were identified for linear relationships between measured proteins, chymotrypsin activity and fibre MCSA. The molecular modifications in costamere turnover and energy homoeostasis identify that aspects of atrophy and fibre transformation are detectable at the protein level in weight-bearing muscles within 3 days of unloading.

Statin-Induced Muscle Necrosis in the Rat: Distribution, Development, and Fibre Selectivity

Simvastatin and cerivastatin have been used to investigate the development of statin-induced muscle necrosis in the rat. This was similar for both statins and was treatment-duration dependent, only occurring after 10 days had elapsed even if the dose was increased, and still occurring after this time when dosing was terminated earlier as a result of morbidity. It was then widespread and affected all areas of the muscular system. However, even when myotoxicity was severe, particular individual muscles and some types of fibres within affected muscles were spared consistently. Fibre typing of spared muscles and of acutely necrotic fibres within affected muscles indicated a differential fibre sensitivity to statin-induced muscle necrosis. The fibres showed a necrotic response to statin administration that matched their oxidative/glycolytic metabolic nature: Least sensitive →I ↔ IIA ↔ IID ↔ IIB ← most sensitive. Type I and IIB fibres represent metabolic extremes of a continuum of metabolic properties through the fibre types with type I fibres most oxidative in metabolism and type IIB fibres most glycolytic. In addition, in some (nonnecrotic) glycolytic fibres from muscles showing early multifocal single fibre necrosis the only subcellular alterations present in isolation of any other changes were mitochondrial. These changes were characterised by an increased incidence of vacuolation and the formation of myelinoid vesicular bodies that accumulated in the subsarcolemmal areas. These findings suggest an important early involvement of mitochondria in selective glycolytic muscle fibre necrosis following inhibition of the enzyme HMG-CoA reductase.