This study explored the expression patterns and interrelationships of myogenesis-related genes in the Semitendinosus and Longissimus dorsi of sheep foetuses at different developmental stages. Haematoxylin and eosin (H&E) staining was used to observe the microstructure of both tissues. The expression and localization of MyoD, MyoG, Myomaker, and Glucocorticoid Receptor (GR) proteins were detected by immunohistochemistry. Quantitative real-time PCR and Western blotting were respectively used to detect the mRNA and protein expression of myogenesis-related genes (MyoD, MyoG, Myf5, Myf6, Pax3, Pax7, Myomaker, Myostatin, and GR) at different developmental stages. The correlations among myogenesis-related genes were analysed by the Pearson correlation coefficient. Finally, the regulatory elements and transcription factor binding sites in the 5' regulatory region of sheep myogenesis-related genes were predicted by bioinformatics analysis. Our results showed that the number and cross-sectional area of muscle fibres in Semitendinosus were lower than those in Longissimus dorsi. The expression patterns and correlations of myogenesis-related genes at the protein and/or mRNA levels were not entirely consistent between the two muscle groups, indicating the complexity of spatiotemporal regulation. The regulatory element analyses suggested that the potential self-, mutual, and cooperative transcriptional regulation relationships, either direct or indirect, existed among the myogenesis-related genes. The study provides a foundation for further exploring the gene regulatory mechanisms and molecular breeding strategies in sheep.

This study provides a detailed anatomical analysis of the distal pelvic limb musculature in the domestic cat (Felis catus), focusing on intrinsic muscles and their relationships with the extrinsic muscle-tendon system. The aim was to document muscle morphology, origin-insertion patterns, tendon courses, and morphometric parameters, while addressing terminological inconsistencies. Ten adult cat cadavers (five males and five females) were dissected in a proximodistal manner under stereomicroscopic guidance, examining the cranial/dorsal and caudal/plantar regions of the crus and pes. The results demonstrated a distinct organization of the distal membrum pelvinum muscles, with notable features in the m. flexor digitorum profundus, m. popliteus, m. soleus, and intrinsic plantar musculature. The digital flexor apparatus, lumbrical attachments, and bipartite structure of the m. abductor digiti V were characterized. No discrete tendinous or connective tissue linkage between the m. flexor digitorum profundus tendon and the manica flexoria (derived from the m. flexor digitorum superficialis tendon) was identified. Morphometric analysis revealed sex-related differences, with males generally exhibiting greater absolute muscle dimensions. However, size-normalized data indicated a more complex pattern, with relatively larger muscle proportions in females for several muscle groups, while short muscles more frequently showed relatively greater dimensions in males. Overall, this study establishes a comprehensive anatomical framework for the distal pelvic limb in the domestic cat and provides baseline morphometric data for future functional and comparative investigations.

The American crocodile (Crocodylus acutus) is a widely distributed Neotropical species for which detailed osteological descriptions of the skull and mandible remain scarce. This study provides an integrative characterization of the cranial and mandibular complex using gross anatomy, descriptive cranial morphometry, radiography, and three-dimensional (3D) scanning. Four skulls and mandibles from post mortem specimens were prepared following standard osteological procedures, and descriptive cranial measurements were recorded. Radiographic projection allowed visualization of bone margins, cranial sutures, dental alveoli, fenestrae, and articular regions. Three-dimensional models generated through a structured-light 3D scanning workflow documented external contours, choanal margins, temporal and mandibular fenestrae, alveolar borders, and sutural patterns. In dorsal, ventral, lateral, and caudal views, the examined skulls displayed an elongated rostrum, dorsally positioned orbits and external nares, paired supratemporal and lateral temporal fenestrae, and a single occipital condyle. The mandibles exhibited elongated and firmly fused symphyses, pronounced retroarticular processes, and well-defined internal and external mandibular fenestrae. Dental preservation varied among specimens, but tooth spacing, conical morphology, and rostrocaudal size variation could be documented in the preserved series. These findings provide a specimen-based reference for the cranial and mandibular complex of C. acutus by integrating morphological, morphometric, radiographic, and three-dimensional (3D) scanning evidence. The digital models constitute an additional resource for anatomical teaching, comparative studies, and long-term digital documentation of osteological morphology.

The South American coati (Nasua nasua) is a mammal of the order Carnivora and family Procyonidae, characterized by scansorial locomotion. Accordingly, several anatomical and functional studies examined its thoracic limbs. Nevertheless, the anatomical descriptions and terminology differ among those studies, and none considered the evolutionary derivations of the brachium and antebrachium muscles based on topology, innervation and anatomical variants. Therefore, the present study aimed to infer the muscle derivations and reconstruct muscle maps of the brachial and craniolateral antebrachial muscles in N. nasua based on the evidence from direct gross dissections and comparisons with literature in embryology and myology of representative mammals from monotremes to procyonids. The caudomedial group will be addressed in a separate manuscript. Seventeen thoracic limbs from eight formaldehyde-fixed specimens and one fresh specimen of N. nasua were dissected. As main results that differed from the available literature, the cranial part of the tensor fasciae antebrachii muscle of non-Nasua procyonids was not lost in N. nasua but fused to the caudal part. The anconeus medialis muscle was not a head of the triceps brachii muscle and may have derived from the caudomedial antebrachial musculature. The caput accessorium of the triceps brachii muscle may have derived mainly from its caput laterale. The extensor carpi radialis muscle was undivided proximally and only separated into two bellies (longus and brevis) from the middle third of the antebrachium. In conclusion, the brachium and craniolateral antebrachium muscles of N. nasua had several differences from those described previously, and most evolutionary derivations could be inferred.

Quantitative cardiac morphology remains poorly documented in crocodilians. Hearts from four commercially reared male Caiman latirostris were examined by gross morphometry, histology, stereology, reticulin staining and Masson's trichrome. Relative heart mass averaged 0.125% of body mass. The left ventricular wall was numerically thicker than the right. Cardiomyocyte volume density and cross-sectional area varied among chambers, whereas collagen-positive area was greater in the right ventricle. Pulmonary arterial walls were markedly thinner than the systemic aortae, and the left pulmonary arterial wall was thicker than the right. These findings extend quantitative cardiac reference data for this species.

The Sunda porcupine (Hystrix javanica) possesses keratin-based quills with exceptional rigidity, necessitating softening porcupine treatments for histological and ultrastructural analyses. This study evaluated the effectiveness of 30% hydrogen peroxide (H2O2) alone and in combination with 20% potassium hydroxide (KOH) on true, flat and rattle quills. Quills from 12 captive Sunda porcupines were subjected to treatments of varying durations and examined using histology and scanning electron microscopy (SEM). Treatment with 30% H2O2 produced moderate softening after 3 days in flat and rattle quills, whereas true quills required 7 days of immersion but remained less effectively softened. In contrast, the combined treatment of 30% H2O2 and 20% KOH achieved effective softening within 25-30 min, resulting in softer texture, increased quill diameter and improved sectioning quality. Histological analyses revealed structural differences among quill types, while SEM showed that 30% H2O2 alone produced surface cracking while retaining discernible interfibrillar boundaries. In contrast, the combined treatment with 30% H2O2 and 20% KOH resulted in a more irregular macrofibril surface, indistinct interfibrillar boundaries and interfibrillar fusion. These findings indicate that the H2O2 + KOH protocol provides a more efficient and rapid method than H2O2 alone, although optimization is necessary to minimise artefacts and preserve structural integrity. Overall, this combination offers a practical approach for preparing porcupine quills for microscopic evaluation, while future studies should investigate milder alternatives and explore the biomedical potential of quill-derived keratin.

Reliable identification of isolated sheep and goat phalanges is challenging because of the close skeletal similarity between these species and the repeated arrangement of homologous digital bones across the forelimbs, hindlimbs, and medial-lateral positions. This study evaluated three-dimensional phalangeal shape for discrimination between 12-month-old male Hamdani sheep and Hair goats and for localization among the sampled anatomical positions. Proximal phalanx (P1), middle phalanx (P2), and distal phalanx (P3) from 30 male Hamdani sheep (Ovis aries) and 30 male Hair goats (Capra hircus), all 12 months of age, were analysed. Bone-specific configurations of 60, 77, and 60 surface pseudolandmarks were used for P1, P2, and P3, respectively. Three-dimensional shape provided substantially greater diagnostic information than centroid size. Shape-only species classification reached 98.3% for P1, 98.3% for P2, and 100% for P3, whereas forelimb-hindlimb classification reached 100%, 98.3%, and 96.2%, respectively. P1 provided the strongest discrimination among the four sampled compound anatomical positions, achieving 91.1% internal classification accuracy. Size-only classification accuracy ranged from 48.5% to 58.6%, compared with 98.3-100.0% for shape-only models. P2 showed the largest adjusted breed-species effect in the animal-mean analysis (partial R[2] = 0.421), whereas P3 yielded the highest internal species-classification accuracy (100.0%) and the largest descriptive sheep-goat mean-shape distance within its own phalanx-specific shape space. Importantly, P3 showed no significant interspecific centroid-size difference despite perfect shape-based species classification, demonstrating that its diagnostic signal was primarily geometric rather than size-dependent. Overall, the results revealed a phalanx-specific diagnostic pattern within the present reference sample: P3 yielded the highest observed breed-species classification accuracy, whereas P1 provided the strongest discrimination of limb region and the four sampled compound anatomical positions. These findings support discrimination within the present modern reference sample; independent validation across other breeds, both sexes, different ages, and archaeological specimens of molecularly confirmed taxon is required before routine application.

The pancreas is a crucial digestive and endocrine organ, yet the histology of the animal pancreas remains incompletely elucidated, particularly in amphibians. This study aimed to elucidate the histology of the American bullfrog (Lithobates catesbeianus) pancreas using histological, immunohistochemical, transmission electron microscopic methods. Histologically, the pancreas displayed typical exocrine acini and scattered endocrine islets. Two types of endocrine cells were distinguished based on their secretory granules: Type I, found diffusely among acini and containing rounded granules (0.28 ± 0.07 μm), and Type II, forming islets and containing rod-shaped granules (major axis 0.37 μm; minor axis 0.12 μm). Importantly, telocytes (TCs) were observed within the pancreatic stroma, characterised by small cell bodies and long telopodes (Tps) with podoms (0.51-2.71 μm) and podomers (0.08-0.14 μm). TCs established close contacts with acinar cells, islets, capillaries and nerve fibres. Most notably, ribosome-associated vesicles (RAV) (mean diameter: 0.27 ± 0.07 μm) were found adjacent to Tp podoms. Immunohistochemistry revealed CD34[+], c-Kit[+] and vimentin[+] interstitial cells with morphological features consistent with TCs, forming a stromal network surrounding acini and islets. This study presents the first ultrastructural and immunohistochemical evidence of TCs and their associated RAV in the pancreas of L. catesbeianus. These findings demonstrate that TCs are structurally integrated interstitial cells in the amphibian pancreas; moreover, RAVs were observed in the vicinity of Tps.

Restoring bone marrow microenvironment (BMM) homeostasis is critical for fracture healing, yet non-invasively tracking physiological events like apoptosis, angiogenesis and matrix remodelling remains challenging. This study evaluated multiparametric 7.0T MRI (T2 mapping and proton density-weighted imaging [PDWI]) for in vivo tracking of BMM spatiotemporal remodelling during natural and pharmacologically accelerated bone repair. Utilizing a rat tibial drill-hole injury model (N = 36), we compared a natural healing control with a Huoxue Qiangji Decoction (HQD) intervention group. Serial high-resolution sagittal MRIs were acquired at days 7, 14 and 21, with signal intensity ratios (SIR) normalized to ipsilateral intact marrow using ITK-SNAP. Distance correlation (dCor) analysis explored non-linear dependencies between MRI metrics and histological features (c-cas3, Masson's trichrome and day-14 CD31/Ki67 expression). Results showed distinct MRI signal trajectories, with earlier homeostatic stabilization in the accelerated healing group. Spatiotemporal mapping revealed closely mirrored non-linear dependencies between MRI and histology. Specifically, T2 mapping SIR mirrored early-stage apoptosis (dCor = 0.527) and mid-term cellular proliferation (Ki67, dCor = 0.732). Concurrently, PDWI reflected dynamic trends of mid-term angiogenesis (CD31, dCor = 0.637) and late-stage collagen deposition (dCor = 0.480). In conclusion, normalized multiparametric MRI provides complementary macroscopic insights into BMM histological progression, highlighting its potential as an objective, non-invasive translational tool for monitoring musculoskeletal healing.

Melanomacrophages (MMs) are pigmented phagocytic cells observed in ectotherms. Investigating MMs is valuable for elucidating the evolutionary background of the vertebrate immune system. However, studies regarding the distribution and function of hepatic MMs remain limited. This study investigated the spatial distribution, histochemical, and ultrastructural characteristics of intracellular inclusions in hepatic MMs in red-eared sliders (Trachemys scripta elegans). MMs are distributed throughout the periphery of the hepatic portal system, with minimal presence elsewhere. MMs contained heterogeneous intracellular inclusions with variable staining characteristics on Masson's trichrome, periodic acid-Schiff, and Berlin blue stains. These findings provide an anatomical basis for comparative immunological investigations of MMs.

The present study describes the gross morphological features and morphometric characteristics of the respiratory apparatus of adult Spur-winged Geese (Plectropterus gambensis), a resident waterfowl species of the Sudano-Sahelian wetlands of Nigeria. Ten ethically obtained adult birds comprising five males and five females were used for this purpose. A standardized gross dissection technique was employed following euthanasia to examine the respiratory viscera. The paired slit-like openings of the nostrils were dorsolateral at the base of the upper beak. The nasal cavity was divided into two equal halves by the nasal septum and consisted of a rostral vestibular region and a broader caudal region containing the nasal conchae, each with a distinct morphological outline. The cranial larynx presented a paired dorsal arytenoid cartilage and a caudal cricoid cartilage capped by the laryngeal mound. The mean length of the trachea in males was 34.72 ± 0.87 cm comprising 138-146 tracheal rings. In females, the trachea measured 31.42 ± 0.74 cm and contained 136-142 tracheal rings. The syrinx was tracheobronchial type. The primary bronchi presented C-shaped cartilages. The paired bright red lungs were triangular shaped, spongy and displayed impressions on their surfaces from contact with adjacent structures. The extrapulmonary primary bronchi and pulmonary vessels entered the lungs through the ventral hilar region. This study establishes the first normal respiratory baseline for the spur-winged goose, filling a critical gap in avian comparative anatomy. The findings also support research on tropical wetland adaptations and provide a practical diagnostic tool for veterinarians.

Comprehensive knowledge of feline vascular anatomy is essential for accurate diagnostic imaging interpretation and safe thoracic cardiovascular surgery. This study aimed to characterize the branching patterns, quantitative morphometry, and precise topographical positions of the aortic arch branches and subclavian arteries in domestic cats (Felis catus). Twenty adult cat cadavers from Northwestern Iran were examined. Coloured latex was injected into the aortic arch, followed by fixation in 10% neutral buffered formalin and subsequent meticulous gross dissection. Topographical positions relative to thoracic skeletal landmarks and morphometric distances were recorded. Non-parametric Mann-Whitney U tests were used for statistical comparisons between identified patterns. Two branching configurations of the brachiocephalic trunk were identified: a bifurcated pattern (85%, n = 17) and a trifurcated pattern (15%, n = 3). The aortic arch was predominantly located at the fourth intercostal space (90%). For the left subclavian artery, three branching patterns were identified, with sequential origin (VT-IT-CCT-SC) being predominant (55%). The right subclavian artery exhibited three distinct arrangements, dominated by sequential (45%) and opposite [IT≡VT] origins (45%). Significant side-dependent variations were observed in the branching order and topography. Morphometric distances (BC + RS, BC-LS, and LCC-RCC) showed no statistically significant differences between bifurcated and trifurcated patterns (p > 0.05). Feline aortic arch branches exhibit substantial topographical variation despite overall metric consistency across branching types. The observed bilateral asymmetry and internal thoracic artery positional mobility likely reflect distinct embryological origin pathways. These findings provide critical baseline anatomical data for improving the accuracy and safety of feline surgical interventions and diagnostic imaging interpretation.

Nedd4-associated E3 ubiquitin ligase 2 (NEDL2) and p97/valosin-containing protein (p97/VCP) are critical regulators of ubiquitin-mediated proteostasis and cellular homeostasis. However, their age-dependent expression patterns in pancreatic tissue during postnatal development and senescent decline remain uncharacterized. This study aimed to investigate the histological alterations alongside the expression profiles of NEDL2 and p97/VCP in rat pancreatic tissue across the postnatal lifespan and ageing. Pancreatic tissues obtained from male Wistar albino rats at postnatal Weeks 1, 4, 10, 36 and 72 were evaluated using histochemical, immunohistochemical and immunofluorescence analyses. Haematoxylin and eosin (H&E) staining were performed to assess morphological changes in endocrine and exocrine pancreas. Immunohistochemistry and semi-quantitative H-score analyses were utilized to evaluate NEDL2 and p97/VCP expression levels. Double immunofluorescence staining was conducted to determine cellular co-localization of NEDL2 with p97/VCP, insulin and glucagon in pancreatic islet cells. Histological analysis showed that the islets of Langerhans progressively enlarged and hypertrophic changes during postnatal ageing, while the exocrine acinar structures maintained its morphological integrity. Immunohistochemical findings revealed dynamic and age-dependent expression of NEDL2 and p97/VCP. Both proteins demonstrated a progressive increase in expression from Week 1 through Week 4. NEDL2 expression peaked significantly at Week 10 (p < 0.001) followed by a gradual decline, whereas p97/VCP expression reached its maximum level at Week 36 (p < 0.001). A significant decrease in expression was observed at Week 72. Immunofluorescence analyses confirmed that NEDL2 strongly co-localized with p97/VCP, insulin-positive β cells and glucagon-positive α cells, particularly during young and mature adulthood (Weeks 10 and 36). Conversely, a marked reduction in fluorescence intensity and co-localization was observed in senescent pancreatic tissue (Week 72). These findings demonstrate that NEDL2 and p97/VCP exhibit coordinated and developmentally regulated expression patterns within pancreatic islet cells. Their elevated expression and prominent co-localization during adulthood suggest a key role for these proteins in pancreatic maturation, endocrine function and cellular proteostasis. Conversely, their down-regulation in senescent tissue may reflect an age-related impairment of protein quality control systems during pancreatic ageing.

The epididymis is a specialized male reproductive organ that undergoes marked postnatal differentiation and segmental specialization to support sperm maturation, transport and storage. Previous studies have demonstrated the presence of epidermal growth factor receptor (EGFR) and epidermal growth factor (EGF) in the epididymis; however, their comparative postnatal localization, together with that of transforming growth factor-α (TGF-α) and amphiregulin (AREG), across specific epididymal segments and epithelial cell types has not been systematically characterized in the rat. This study investigated the postnatal histomorphological development and cell type- and segment-associated localization of EGFR, EGF, TGF-α and AREG in the epididymis of Wistar albino rats. Epididymal tissues were collected from neonatal (PND5), infantile (PND20), postpubertal (PND50) and adult (PND70) rats (n = 6 per group). Histomorphological development was evaluated using Crossmon's modified trichrome, periodic acid-Schiff (PAS) and toluidine blue staining, whereas immunolocalization was assessed semiquantitatively by immunohistochemistry. Histological examination revealed progressive epithelial differentiation and segment-specific morphological maturation during postnatal development. EGFR immunoreactivity was detected in basal cells, clear cells and periductal smooth muscle cells, with prominent staining in clear cells of the cauda epididymis in postpubertal and adult rats. Immunoreactivity for EGF and TGF-α was restricted to principal, apical and basal cells of the caput epididymis and was detected only during the postpubertal and adult stages. AREG immunoreactivity was observed in basal and clear cells as well as in periductal smooth muscle cells, without statistically significant developmental or segmental differences. Semiquantitative analysis demonstrated significant increases in EGF and TGF-α immunoreactivity in the caput epididymis during the postpubertal and adult stages (adjusted p = 0.043 for both), and immunoreactivity in the caput was significantly higher than that in the corpus and cauda segments (adjusted p = 0.01 for both). No statistically significant differences were detected in EGFR or AREG immunoreactivity. These findings reveal distinct postnatal cell type- and segment-associated localization patterns of EGFR and its ligands. The localization of EGFR and AREG in basal and clear cells, together with the caput-restricted distribution of EGF and TGF-α, provides a descriptive framework for future studies investigating their possible involvement in epididymal epithelial maturation and regional specialization.

To describe the structural development of the liver in young guppies (Poecilia reticulata) and complement the adult characterisation, we conducted a histological study at 7, 17, and 28 days post-birth. We used serial sections to trace the organisation of blood vessels, bile ducts, and pancreatic tissue within and around the liver. At all ages, the liver was highly cellular and composed mainly of hepatocytes. Pigmented melanomacrophage centres or aggregates were not observed, whereas rodlet cells were rare and restricted to extrahepatic connective tissue. At 7 days, we identified four distinct venous entry points. This multi-afferent pattern persisted at Days 17 and 28. Arterioles remained restricted to the outer hilar region and were not observed within the liver. The gallbladder and common hepatic duct were recognisable at Day 7. In contrast, intrahepatic bile ducts were not discernible at Day 7, were scarce at Day 17, and were more frequent, although still limited, at Day 28. Exocrine pancreatic tissue was mainly extrahepatic at Day 7; intrahepatic pancreatic acini were detected in one fish at this age and were increasingly observed at Days 17 and 28 in association with afferent venous tracts. No vascular-biliary associations comparable to those described in adult guppies were observed. These findings establish a histological baseline for normal post-hatching liver development in guppies and provide a reference for future studies in comparative anatomy and of environmentally associated changes in liver structure during early life stages.