The Early Iron Age elite grave from the Kapiteljska njiva barrow cemetery (Novo mesto, Slovenia): an approach to dental profiling through non-invasive and minimally-invasive techniques.

Kapiteljska njiva is a prehistoric cemetery located in the town of Novo mesto in southern Slovenia. There is a long history of archaeological research at this site, as the first investigations date back to the end of the 19th century. In 2004, an Early Iron Age barrow XVI was investigated. The oldest surviving grave in the barrow is the central grave, numbered XVI/34, which according to its position and the richness of the grave goods, belongs to a woman of higher status. Most likely, she was the first member or initiator of a family that continued to bury its dead in the barrow for the next 300 years. There were no preserved skeletal elements; however, in the head part of the grave, remains of human teeth, mostly the tooth crowns and the shells of enamel from the tooth crowns, were found among the scattered amber beads of a necklace. Moreover, these tooth remains are one of the few human biological materials (mostly fragments of skull and long limb bones) unearthed in this cemetery, reflecting the influence of acidic soil from the burial site. This study aimed to create a dental profile of the deceased in a similar way as during a forensic investigation. The remains were examined macroscopically and stereo-microscopically. The morphological traits were scored following the ASUDAS protocol. Ancestry was estimated by entering these scores into a beta version of the web-based application rASUDAS. Brothwell's system was used for age at death estimation from occlusal attrition. Identification of sex was based on the analysis of sex-specific amelogenin isoforms in dental enamel by nanoflow liquid chromatography tandem mass spectrometry (nanoLC-MS/MS). The present study reveals that the dental remains from Kapiteljska njiva belong to the permanent dentition of a single individual and that only the right upper central incisor and third molar are completely absent. The remains of teeth exhibit a simple external morphology, characterised by the absence of morphological dental traits, with a notable exception of the two-rooted left lower canine. The probability of assigning this individual to the Western Eurasian ancestry group is 98%. According to the degree of dentine exposure on the occlusal surfaces of molars, the estimated age range is 17-25 years. Another line of evidence comes from the observation that the first signs of approximal attrition are present in lower third molars but absent in the only preserved upper counterpart, indicating that the age at which the lower third molars entered into occlusion represents a proxy of the individual's age at death. Data on the chronology of the lower third molar development and eruption in present-day European populations from forensic literature (Brkic et al. 2011; Olze et al. 2008; Selmanagic et al. 2013) and The London Atlas confirm the above age-range estimate. Caution is, however, needed in the interpretation of results, because reference data are not based on the population of the individual's origin. Proteomic analysis classified the individual as a female, which is in line with the archaeological evidence. No pathological lesions or indicators of systemic stress were identified; however, the absence of approximal attrition facets in upper anterior teeth indicates interdental spacing. The results of dental profiling are discussed in the context of historical background, today's clinical knowledge, and epidemiological data.

Morphometric and morphologic analysis of the foramen spinosum in the Slovenian population with clinical correlations.

Background: The foramen spinosum (FS) is a pivotal passage for neurovascular structures within the skull base. We performed a detailed morphometric and morphological analysis of the FS to emphasize its clinical relevance. Materials & Methods: The study was performed on dried skull specimens obtained from the anatomical collections of the Institute of Anatomy and Institute of Forensic Medicine of the University of Ljubljana. The morphometric and morphologic features of FS in 126 whole human skulls and 15 skull halves were analyzed, including dimensions, shape, and other anatomical variations, as well as relationships to surrounding structures. Measurements were done with a digital sliding caliper. Results: The mean length and width of the FS were 2.45 ± 0.65 mm and 2.03 ± 0.53 mm on the right side and 2.49 ± 0.61 mm and 2.08 ± 0.48 mm on the left side. The most frequently observed shape was round (56.7%), followed by oval (28.2%), irregular (8.7%) and drop shaped (6.3%). Several anatomical variations were also noted, including FS duplication, confluences with other foramina, and FS obstruction due to marginal bony outgrowths. Conclusion: The FS exhibits notable interindividual differences in anatomical characteristics which should be considered during neurosurgical procedures and radiological interventions in the skull base.

Skeletal muscle myosin heavy chain expression and 3D capillary network changes in streptozotocin-induced diabetic female mice.

It is not well-understood how type 1 diabetes (T1DM) affects skeletal muscle histological phenotype, particularly capillarisation. This study aimed to analyze skeletal muscle myosin heavy chain (MyHC) fibre type changes and 3D capillary network characteristics in experimental T1DM mice. Female C57BL/6J-OlaHsd mice were categorized into streptozotocin (STZ)-induced diabetic (n = 12) and age-matched non-diabetic controls (n =12). The muscle fibre phenotype of the soleus, gluteus maximus, and gastrocnemius muscles were characterized based on the expression of MyHC isoforms, while capillaries of the gluteus maximus were assessed with immunofluorescence staining, confocal laser microscopy and 3D image analysis. STZ-induced diabetic mice exhibited elevated glucose levels, reduced body weight, and prolonged thermal latency, verifying the T1DM phenotype. In both T1DM and non-diabetic mice, the gluteus maximus and gastrocnemius muscles predominantly expressed fast-twitch type 2b fibers, with no significant differences noted. However, the soleus muscle in non-diabetic mice had a greater proportion of type 2a fibers and comparable type 1 fiber densities (26.2 ± 14.6% vs 21.9 ± 13.5%) relative to diabetic mice. T1DM mice showed reduced fiber diameters (P = 0.026), and the 3D capillary network analysis indicated a higher capillary length per muscle volume in the gluteus maximus of diabetic mice compared to controls (P < 0.05). Overall, T1DM induced significant changes in the skeletal muscle, including shifts in MyHC fibre types, decreased fibre diameters, and increased relative capillarisation, possibly due to muscle fibre atrophy. Our findings emphasize the superior detail provided by the 3D analytical method for characterizing skeletal muscle capillary architecture, highlighting caution in interpreting 2D data for capillary changes in T1DM.

Sonographic assessment of the tarsal tunnel compared to cadaveric findings: a pictorial study.

Aim of the study: To present the anatomy of the tarsal tunnel and demonstrate the utility of high-resolution ultrasound for tarsal tunnel examination. Materials and methods: Anatomical dissection was performed on a defrosted cadaveric model to demonstrate relevant anatomical structures of the tarsal tunnel, namely tendons, vessels and nerves. The tibial nerve division was demonstrated; the bifurcation of the tibial nerve into the medial and lateral plantar nerve, two medial calcaneal nerve branches were identified originating from the tibial nerve and the Baxter's nerve was identified as the first branch of the lateral plantar nerve. An ultrasound examination of the tarsal tunnel region was performed on a healthy volunteer. A linear probe was used and sonographic images were obtained at different levels of the tarsal tunnel: the proximal tarsal tunnel, the tibial nerve division into the medial and lateral plantar nerves, the distal tarsal tunnel, the Baxter's nerve branching point and the Baxter's nerve crossing between the abductor hallucis and quadratus plantae muscle. Results: Sonographic images were correlated with anatomical structures exposed during cadaveric dissection. Conclusions: We presented the anatomic-sonographic correlation of the tarsal tunnel and showed that high-resolution ultrasound is a useful imaging modality for tarsal tunnel assessment.

Correlation between diffusion tensor indices and fascicular morphometric parameters of peripheral nerve.

Introduction: Diffusion tensor imaging (DTI) is a magnetic resonance imaging (MRI) technique that measures the anisotropy of water diffusion. Clinical magnetic resonance imaging scanners enable visualization of the structural integrity of larger axonal bundles in the central nervous system and smaller structures like peripheral nerves; however, their resolution for the depiction of nerve fascicular morphology is limited. Accordingly, high-field strength MRI and strong magnetic field gradients are needed to depict the fascicular pattern. The study aimed to quantify diffusion tensor indices with high-field strength MRI within different anatomical compartments of the median nerve and determine if they correlate with nerve structure at the fascicular level. Methods: Three-dimensional pulsed gradient spin-echo (PGSE) imaging sequence in 19 different gradient directions and b value 1,150 s/mm2 was performed on a 9.4T wide-bore vertical superconducting magnet. Nine-millimeter-long segments of five median nerve samples were obtained from fresh cadavers and acquired in sixteen 0.625 mm thick slices. Each nerve sample had the fascicles, perineurium, and interfascicular epineurium segmented. The diffusion tensor was calculated from the region-average diffusion-weighted signals for all diffusion gradient directions. Subsequently, correlations between diffusion tensor indices of segmentations and nerve structure at the fascicular level (number of fascicles, fascicular ratio, and cross-sectional area of fascicles or nerve) were assessed. The acquired diffusion tensor imaging data was employed for display with trajectories and diffusion ellipsoids. Results: The nerve fascicles proved to be the most anisotropic nerve compartment with fractional anisotropy 0.44 ± 0.05. In the interfascicular epineurium, the diffusion was more prominent in orthogonal directions with fractional anisotropy 0.13 ± 0.02. Diffusion tensor indices within the fascicles and perineurium differed significantly between the subjects (p < 0.0001); however, there were no differences within the interfascicular epineurium (p ≥ 0.37). There were no correlations between diffusion tensor indices and nerve structure at the fascicular level (p ≥ 0.29). Conclusion: High-field strength MRI enabled the depiction of the anisotropic diffusion within the fascicles and perineurium. Diffusion tensor indices of the peripheral nerve did not correlate with nerve structure at the fascicular level. Future studies should investigate the relationship between diffusion tensor indices at the fascicular level and axon- and myelin-related parameters.

Sphenoidal Foramen Ovale in the Slovenian Population: An Anatomical Evaluation with Clinical Correlations.

The foramen ovale (FO) is a crucial feature of the skull base, serving as a passage for clinically important neurovascular structures. The present study aimed to provide a comprehensive morphometric and morphologic analysis of the FO and highlight the clinical significance of the anatomical characterization. A total of 267 FO were analyzed in skulls obtained from deceased inhabitants of the Slovenian territory. The anteroposterior (length) and the transverse (width) diameters were measured using a digital sliding vernier caliper. Dimensions, shape, and anatomical variations of FO were analyzed. The mean length and width of the FO were 7.13 and 3.71 mm on the right side and 7.20 and 3.88 mm on the left side. The most frequently observed shape was oval (37.1%), followed by almond (28.1%), irregular (21.0%), D-shaped (4.5%), round (3.0%), pear-shaped (1.9%), kidney-shaped (1.5%), elongated (1.5%), triangular (0.7%), and slit-like (0.7%). In addition, marginal outgrowths (16.6%) and several anatomical variations were noted, including duplications, confluences, and obstruction due to a complete (5.6%) or incomplete (8.2%) pterygospinous bar. Our observations revealed substantial interindividual variation in the anatomical characteristics of the FO in the studied population, which could potentially impact the feasibility and safety of neurosurgical diagnostic and therapeutic procedures.

Comparison of the feasibility and safety of infrazygomatic and suprazygomatic approaches to pterygopalatine fossa using virtual reality.

BACKGROUND: Injections of local anesthetics into pterygopalatine fossa gained popularity for treating acute and chronic facial pain and headaches. Injury of maxillary artery during pterygopalatine fossa injection can result in pseudoaneurysm formation or acute bleeding. We aimed to identify the optimal approach into pterygopalatine fossa by comparing feasibility and safety of suprazygomatic and two infrazygomatic approaches. METHODS: We analyzed 100 diagnostic CT angiographies of cerebral arteries using 3D virtual reality. Each approach was determined as a target point in pterygomaxillary fissure and an array of outermost edges trajectories leading to it. The primary outcomes were feasibility and safety for each approach. The secondary outcome was the determination of maxillary artery position for each approach to identify the safest needle entry point. RESULTS: Suprazygomatic approach was feasible in 96.5% of cases, while both infrazygomatic approaches were feasible in all cases. Suprazygomatic approach proved safe in all cases, posterior infrazygomatic in 73.5%, and anterior infrazygomatic in 38%. The risk of maxillary artery puncture in anterior infrazygomatic approach was 14.7%±26.4% compared to 7.5%±17.2%. in posterior infrazygomatic with the safest needle entry point in the upper-lateral quadrant in both approaches. CONCLUSION: The suprazygomatic approach proved to be the safest, however not always feasible. The posterior infrazygomatic approach was always feasible and predominantly safe if the needle entry point was just anterior to the condylar process. The anterior infrazygomatic approach was always feasible, however least safe even with an optimal needle entry point just anterior to the coronoid process.

Exploration of macromolecular phenotype of human skeletal muscle in diabetes using infrared spectroscopy.

Introduction: The global burden of diabetes mellitus is escalating, and more efficient investigative strategies are needed for a deeper understanding of underlying pathophysiological mechanisms. The crucial role of skeletal muscle in carbohydrate and lipid metabolism makes it one of the most susceptible tissues to diabetes-related metabolic disorders. In tissue studies, conventional histochemical methods have several technical limitations and have been shown to inadequately characterise the biomolecular phenotype of skeletal muscle to provide a holistic view of the pathologically altered proportions of macromolecular constituents. Materials and methods: In this pilot study, we examined the composition of five different human skeletal muscles from male donors diagnosed with type 2 diabetes and non-diabetic controls. We analysed the lipid, glycogen, and collagen content in the muscles in a traditional manner with histochemical assays using different staining techniques. This served as a reference for comparison with the unconventional analysis of tissue composition using Fourier-transform infrared spectroscopy as an alternative methodological approach. Results: A thorough chemometric post-processing of the infrared spectra using a multi-stage spectral decomposition allowed the simultaneous identification of various compositional details from a vibrational spectrum measured in a single experiment. We obtained multifaceted information about the proportions of the different macromolecular constituents of skeletal muscle, which even allowed us to distinguish protein constituents with different structural properties. The most important methodological steps for a comprehensive insight into muscle composition have thus been set and parameters identified that can be used for the comparison between healthy and diabetic muscles. Conclusion: We have established a methodological framework based on vibrational spectroscopy for the detailed macromolecular analysis of human skeletal muscle that can effectively complement or may even serve as an alternative to histochemical assays. As this is a pilot study with relatively small sample sets, we remain cautious at this stage in drawing definitive conclusions about diabetes-related changes in skeletal muscle composition. However, the main focus and contribution of our work has been to provide an alternative, simple and efficient approach for this purpose. We are confident that we have achieved this goal and have brought our methodology to a level from which it can be successfully transferred to a large-scale study that allows the effects of diabetes on skeletal muscle composition and the interrelationships between the macromolecular tissue alterations due to diabetes to be investigated.

Pathophysiological and Therapeutic Roles of Fascial Hyaluronan in Obesity-Related Myofascial Disease.

Myofascial disease is an important complication associated with obesity and one of the leading causes of physical disability globally. In the face of limited treatment options, the burden of myofascial disorders is predicted to increase along with the escalating prevalence of obesity. Several pathological processes in obesity contribute to modifications in fascial extracellular matrix mechanical and biological properties and functions. Changes in adipose tissue metabolism, chronic inflammatory phenotype, oxidative stress, and other mechanisms in obesity may alter the physiochemical and biomechanical properties of fascial hyaluronan. Understanding the pathophysiological importance of hyaluronan and other components of the fascial connective tissue matrix in obesity may shed light on the etiology of associated myofascial disorders and inform treatment strategies. Given its unique and favorable pharmacological properties, hyaluronan has found a broad range of clinical applications, notably in orthopedic conditions such as osteoarthritis and tendinopathies, which share important pathophysiological mechanisms implicated in myofascial diseases. However, while existing clinical studies uniformly affirm the therapeutic value of hyaluronan in myofascial disorders, more extensive studies in broader pharmacological and clinical contexts are needed to firmly validate its therapeutic adaptation.

Application of FTIR Spectroscopy to Detect Changes in Skeletal Muscle Composition Due to Obesity with Insulin Resistance and STZ-Induced Diabetes.

Age, obesity, and diabetes mellitus are pathophysiologically interconnected factors that significantly contribute to the global burden of non-communicable diseases. These metabolic conditions are associated with impaired insulin function, which disrupts the metabolism of carbohydrates, lipids, and proteins and can lead to structural and functional changes in skeletal muscle. Therefore, the alterations in the macromolecular composition of skeletal muscle may provide an indication of the underlying mechanisms of insulin-related disorders. The aim of this study was to investigate the potential of Fourier transform infrared (FTIR) spectroscopy to reveal the changes in macromolecular composition in weight-bearing and non-weight-bearing muscles of old, obese, insulin-resistant, and young streptozotocin (STZ)-induced diabetic mice. The efficiency of FTIR spectroscopy was evaluated by comparison with the results of gold-standard histochemical techniques. The differences in biomolecular phenotypes and the alterations in muscle composition in relation to their functional properties observed from FTIR spectra suggest that FTIR spectroscopy can detect most of the changes observed in muscle tissue by histochemical analyses and more. Therefore, it could be used as an effective alternative because it allows for the complete characterization of macromolecular composition in a single, relatively simple experiment, avoiding some obvious drawbacks of histochemical methods.

The Influence of Diabetic Peripheral Neuropathy on the Duration of Sciatic Nerve Block with 1.3% Liposomal Bupivacaine and 0.25% Bupivacaine Hydrochloride in a Mouse Model.

Little is known regarding the pharmacological properties of extended-release local anesthetics in the setting of diabetic peripheral neuropathy. We investigated and compared the duration of sciatic nerve block following administration of clinically relevant concentrations of liposomal bupivacaine (LB) and bupivacaine hydrochloride (BH) in diabetic mice with peripheral neuropathy. In this prospective, randomized, and double-blind study, twenty-four female C57BL/6J-OlaHsd mice were assigned to a streptozotocin-induced type 1 diabetes group and a control group without diabetes. The presence of peripheral neuropathy was established by assessing the duration of thermal latency of the plantar and tail-flick tests, following which both groups were subdivided into two subgroups in which 35 mg/kg of 1.31% LB and 7 mg/kg of 0.25% BH were respectively administered for sciatic nerve block. The average sensory block duration with BH was 106 min and 117.1 min in the control and diabetic groups, respectively. With LB, the average sensory block duration was 118 min in the control mice, while in mice with diabetic peripheral neuropathy, the average block duration was significantly longer and above the 270 min limit set in our study. Accordingly, sensory block duration was longer with LB compared to BH, and diabetic peripheral neuropathy significantly increased sciatic nerve block duration with LB.

Diffusion Tensor Imaging of a Median Nerve by Magnetic Resonance: A Pilot Study.

The magnetic resonance Diffusion Tensor Imaging (DTI) is a powerful extension of Diffusion Weighted Imaging (DWI) utilizing multiple bipolar gradients, allowing for the evaluation of the microstructural environment of the highly anisotropic tissues. DTI was predominantly used for the assessment of the central nervous system (CNS), but with the advancement in magnetic resonance (MR) hardware and software, it has now become possible to image the peripheral nerves which were difficult to evaluate previously because of their small caliber. This study focuses on the assessment of the human median peripheral nerve ex vivo by DTI microscopy at 9.4 T magnetic field which allowed the evaluation of diffusion eigenvalues, the mean diffusivity and the fractional anisotropy at 35 µm in-plane resolution. The resolution was sufficient for clear depiction of all nerve anatomical structures and therefore further image analysis allowed the obtaining of average values for DT parameters in nerve fascicles (intrafascicular region and perineurium) as well as in the surrounding epineurium. The results confirmed the highest fractional anisotropy of 0.33 and principal diffusion eigenvalue of 1.0 × 10-9 m2/s in the intrafascicular region, somewhat lower values of 0.27 and 0.95 × 10-9 m2/s in the perineurium region and close to isotropic with very slow diffusion (0.15 and 0.05 × 10-9 m2/s) in the epineurium region.

Median and ulnar nerve fascicle imaging using MR microscopy and high-resolution ultrasound.

BACKGROUND AND PURPOSE: Understanding nerve microanatomy is important as different neuropathies and some nerve neoplasms present with fascicle enlargement. The aim of our study was to gain clinically oriented knowledge on nerve fascicular anatomy using imaging modalities. METHODS: On a cadaveric upper extremity, high-resolution ultrasound (HRUS) scan with 22 MHz probe was performed. Sections of the median and ulnar nerves were excised at the level of the distal arm and after magnetic resonance microscopy (MRM), histological cross-sections (HCS) were prepared. Cross-referencing of the MRM and HRUS images with HCS was performed. Fascicle and nerve contouring was performed with morphometric software in order to assess nerve and fascicular cross-sectional area (CSA), fascicle count, and interfascicular distances. Based on fascicle differentiation, factual fascicle (FF) group and fascicular cluster (FC) group were defined. RESULTS: On the cross-referenced imaging material, fascicles were differentiated in 92.7% on MRM and in 57.3% on HRUS. High to very high positive correlation among imaging material was observed for the fascicle CSA. FF depiction was 30.1% on HRUS. In comparison to the FF group, the FC group had significantly larger fascicle CSA and shorter interfascicular distances. DISCUSSION: The findings of our study contribute to understanding of fascicle depiction on imaging modalities. HRUS offers good visualization of fascicles. The capability of differentiating fascicles is modality specific and depends on the fascicle CSA and the amount of interfascicular epineurium.

Skeletal Muscle Microvascular Dysfunction in Obesity-Related Insulin Resistance: Pathophysiological Mechanisms and Therapeutic Perspectives.

Obesity is a worrisomely escalating public health problem globally and one of the leading causes of morbidity and mortality from noncommunicable disease. The epidemiological link between obesity and a broad spectrum of cardiometabolic disorders has been well documented; however, the underlying pathophysiological mechanisms are only partially understood, and effective treatment options remain scarce. Given its critical role in glucose metabolism, skeletal muscle has increasingly become a focus of attention in understanding the mechanisms of impaired insulin function in obesity and the associated metabolic sequelae. We examined the current evidence on the relationship between microvascular dysfunction and insulin resistance in obesity. A growing body of evidence suggest an intimate and reciprocal relationship between skeletal muscle microvascular and glucometabolic physiology. The obesity phenotype is characterized by structural and functional changes in the skeletal muscle microcirculation which contribute to insulin dysfunction and disturbed glucose homeostasis. Several interconnected etiologic molecular mechanisms have been suggested, including endothelial dysfunction by several factors, extracellular matrix remodelling, and induction of oxidative stress and the immunoinflammatory phenotype. We further correlated currently available pharmacological agents that have deductive therapeutic relevance to the explored pathophysiological mechanisms, highlighting a potential clinical perspective in obesity treatment.

Transurethral catheterization of female domestic pigs with a Foley catheter.

This report describes transurethral catheterization with a Foley catheter in 19 anaesthetized, laterally recumbent female pigs with a chest circumference of 72.1 ± 4.2 cm and weighing approximately 40 kg. Catheterization within five minutes was successful in 78.9% of the pigs. Catheterization depth, measured as the distance between the bladder neck and the mucocutaneous junction of the vulva, was 14.5 ± 1.3 cm and correlated with chest circumference but not body length. Measurement of catheterization depth aids in optimal positioning and may prevent inflation of the Foley catheter in the urethra.

Determination of bupivacaine tissue concentration in human biopsy samples using high-performance liquid chromatography with mass spectrometry.

In the present study, we developed a simple and rapid analytical method for the quantification of bupivacaine hydrochloride in human biopsy samples of adipose, muscle, neural, connective and cartilage tissue using liquid chromatography-mass spectrometry. Anesthetics were extracted from the tissue samples using 0.1% formic acid in acetonitrile for protein denaturation and hexane for removal of lipophilic impurities. Analytes were separated adequately on Phenomenex Luna Omega polar C18 column using a gradient mobile phase 0.1% formic acid in water and 0.1% formic acid in acetonitrile. The lower limits of quantification were ≤ 97 ng g-1 tissue for all studied tissues. Intra-day recoveries were between 48.2% and 82.1% with relative standard deviations (RSDs) between 1.47% and 14.28%, whereas inter-day recoveries were between 52.2% and 77.6% with RSDs between 2.98% and 14.79%. The calibration curve showed a linear fit with R2 higher than 0.99 in the concentration range from 0.16 to 100 µg g-1 . Lidocaine hydrochloride was tested as internal standard because its recoveries and matrix effects were comparable to bupivacaine hydrochloride. Post-analytical corrections of measured bupivacaine tissue concentrations can accordingly be made based on recovery of lidocaine as internal standard, with recoveries between 51.2% and 86.9% and RSDs between 1.99% and 16.88%. The developed method could be used to study time-dependent spread of bupivacaine locally or to more distant locations across tissue barriers.

Skeletal muscle and fiber type-specific intramyocellular lipid accumulation in obese mice.

In obesity, accumulation of lipid droplets in skeletal muscle fibers and a shift towards fast muscle fiber types can both contribute to insulin resistance. However, it is not yet clear how intramyocellular lipid accumulation and fiber type changes are associated. Therefore, we investigated to what extent the lipids accumulated in a fiber type-specific manner in the functionally similar fast-, intermediate- and slow-twitch gastrocnemius, plantaris, and soleus muscles, respectively, in high-fat diet-induced obese 54-week-old female C57BL/6JOlaHsd mice (n = 9) compared to control standard-diet-treated lean mice (n = 9). A high-fat diet was administered for 26 weeks. Fiber-type specific intramyocellular lipid content analysis and muscle fiber typing were performed using histochemical analysis of lipids with Sudan Black and immunohistochemical analysis of myosin heavy chains on serial sections of skeletal muscles. Compared to the lean mice, the lipid accumulation was most prominent in types 2a and 2x/d fibers (p < 0.05) of fast-twitch gastrocnemius and intermediate plantaris muscles in the obese mice, while in slow-twitch soleus muscle, there was no significant lipid accumulation in the obese animals. Furthermore, the slow-twitch soleus muscle of the obese mice with no significant change in muscle fiber diameters exhibited the most pronounced shift towards fast-type myosin heavy chain isoform expression (p < 0.05). In contrast, the fast-twitch and intermediate-twitch gastrocnemius and plantaris muscles, respectively, in which the muscle fiber diameters increased (p < 0.05), were more resistant toward myosin heavy chain expression changes. In conclusion, we demonstrated both muscle- and fiber-type specificity in intramyocellular lipid accumulation in obese mice, suggesting that in obesity, similar muscle fiber types in different muscles accumulate lipids differentially.

Horizontal deformation of skeletal muscle thick sections visualised by confocal microscopy.

Certain morphological parameters of the skeletal muscle tissue can be better understood via 3D considerations. Fluorescent confocal microscopy of thick tissue sections is a well-established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, thick tissue sections are prone to deformations which may significantly influence some stereological and morphometric results like muscle fibre diameter and capillary length, but not dimensionless parameters like object number and Euler-Poincaré characteristics. To better understand this phenomenon, we studied the horizontal deformation of thick (100 µm) transverse skeletal muscle sections, by comparing the muscle fibre diameters measured on thick sections to muscle fibre diameters measured on thin (10 µm) sections of the same sample. Diameter changes were further correlated with shrinkage in the Z direction (axial shrinkage) and deviation of the muscle fibre preferential axis from the Z-axis. We showed that the thick sections dilated in horizontal and shrunk in Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in the Z direction. The latter was more pronounced in transversely than obliquely cut tissue sections. The results emphasise that even when shrinkage in the Z direction can be corrected using calibration, it is important to optimise histological protocols to minimise the Z-axis collapse that could cause horizontal dilation. LAY DESCRIPTION: In skeletal muscle research, 3D analysis is especially important for studying the microvasculature. Laser scanning confocal microscopy of skeletal muscle thick tissue sections is a well-established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, such sections are prone to deformations which may significantly influence the study results. To better understand this phenomenon, we studied the horizontal deformation of thick transverse skeletal muscle sections. We compared the average muscle fibre diameters measured on thick skeletal muscle sections, thin fixed skeletal muscle sections and immunohistochemically stained thin skeletal muscle sections with the muscle fibre diameters measured on thin native skeletal muscle sections of the same sample, with the latter condition serving as the standard diameters (ie the control condition). We further studied the association among muscle fibre diameter changes, shrinkage of the thick skeletal muscle sections in the Z direction and their sectioning angle. We showed that the thick skeletal muscle sections dilated in the horizontal direction and shrunk in the Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in Z direction. The shrinkage in the Z direction was more pronounced in transversely than obliquely cut tissue sections. These results emphasise that even when shrinkage in the Z direction can be corrected using Z-axis calibration, it is very important to optimise histological protocols to minimise the Z-axis collapse that could cause horizontal dilation in order to enhance the integrity of study results.

Transversus nuchae muscle fused with risorius muscle: a case report / Músculo transverso de la nuca fusionado con músculo risorius: informe de un caso

SUMMARY: Facial muscles are known to have a variable anatomy. Such anatomical variations can affect facial expressions. A natural smile is produced by contracting several muscles of facial expression, including the muscles invested in the superficial musculoaponeurotic system. Change in the width of the mouth on smiling is a known iatrogenic complication affecting the risorius muscle when using Botulinum Neurotoxin as a non-invasive treatment for masseteric hypertrophy. Also damage to the risorius muscle, with consequent clinical deficits, is a common surgical complication in facial aesthetic, oncologic and other head and neck procedures such as parotidectomy and rhytidectomy. Hence, a thorough knowledge of muscle anatomy, particularly its potential variations, is critical in reducing the associated operative morbidity. Accordingly, we report an unusual variation of the transversus nuchal muscle fused with the risorius muscle noted in an 83-year-old female body cadaver.

RESUMEN: Los músculos faciales tienen una anatomía variable. Tales variaciones anatómicas pueden afectar las expresiones faciales. Una sonrisa natural se produce al contraer varios músculos de la expresión facial, incluidos los músculos invertidos en el sistema musculoaponeurótico superficial. El cambio en el ancho de la boca al sonreír es una complicación iatrogénica conocida, que afecta el músculo risorio cuando se usa la neurotoxina botulínica como un tratamiento no invasivo para la hipertrofia masterérica. Además, el daño al músculo risorio, con los consecuentes déficits clínicos, es una complicación quirúrgica común en la estética facial, oncológica y otros procedimientos de cabeza y cuello, como la parotidectomía y la ritidectomía. Por lo tanto, un conocimiento profundo de la anatomía muscular, particularmente sus posibles variaciones, es crítico para reducir la morbilidad operativa asociada. En consecuencia, informamos una variación inusual del músculo transverso de la nuca fusionado con el músculo risorio, observado en un cadáver de cuerpo femenino de 83 años.