Light and scanning electron microscopy of the eye of Siganus luridus (Rüppell, 1828).

Introduction: The morphological characteristics of eyes in fishes are highly diverse and have evolved to meet the specific visual requirements as per their habitats. These morphological features of eyes are important for researchers and ecologists. The dusky spinefoot (Siganus luridus) is a tropical teleost fish with a laterally flattened body which lives in the Mediterranean Sea. Currently, there are no histological data relating to the Siganus luridus eye. Methods: In this study, the morphology of the Siganus luridus eye was examined to enhance our understanding of its structure and its relationship to fish ecology. Detailed gross and microscopic features were recorded using light and scanning microscopy. Results: The key observations describe the main structural features of the eye of Siganus luridus, specifically, the diameter of the orbit, architecture of three tunics of eye and detailed lens description. The choroid was divided into four layers, and had a rete mirabile, consisting of numerous small blood vessels in the choroidal gland. The tapetum lucidum was observed, which is interesting since Siganus luridus is herbivore and herbivores typically lack a tapetum lucidum. Discussion: These observations shed new light on the intricate eye structure of Siganus luridus and provide valuable insights into its visual abilities and adaptations to the aquatic environment and feeding behavior.

A new insight for investigating the prenatal and postnatal ossification centers of pelvic and femur bones in white New Zealand rabbits (Oryctolagus cuniculus) using 3D CT, double stain technique, and morphometry.

BACKGROUND: The ossification centers in rabbit limbs are related to fetal age and bone maturation. OBJECTIVE: To address the limited studies on ossification in the hind limbs of New Zealand rabbits, we investigated the prenatal and postnatal development of the pelvic and femur bones. METHODS: Double staining with Alcian Blue and Alizarin Red, computed tomography (CT), and 3D reconstruction were employed to visualize and analyze ossification centers in detail. RESULTS: Using double staining, we observed these patterns: At prenatal days 18 and 21, ossification centers appeared in the ilium. By prenatal days 23 and 25, ossification began in the ischium. On postnatal day 1, ilium ossification centers spread across most of the ilium wings, except for the iliac crest, and new centers appeared in the pubis and cotyloid bones. Most bones had ossified by the third week and one month postnatal, except for the iliac crest and ischial tuberosity. At 1.5 months, both were fully ossified. On day 18 post coitum, an ossification center was visible in the middle of the femur shaft. By day 28 post coitum, ossification extended through the shaft, and postnatally, new ossification spots appeared at the extremities by day one and week one. By the third week, complete ossification of the femur head, lesser trochanter, third trochanter, medial condyle, and lateral condyle was observed. At 1.5 months, the entire proximal extremity was ossified. CONCLUSION: 3D CT provided clear imaging of ossification progression in the pelvic and femur bones. This study enhances our understanding of vertebrate skeletal development.

A novel immunofluorescence study of Lingual Salivary Glands in the Egyptian Tortoise (Testudo kleinmanni) and its ecological significance.

The Egyptian tortoise (Testudo kleinmanni) is remarkably adapted to its harsh desert environment, a characteristic that is crucial for its survival under extreme conditions. This study was aimed at providing a deeper understanding of the lingual salivary gland structures in the Egyptian tortoise and examining how these structures help the tortoise manage hydration and nutrition in arid conditions. Utilizing a combination of light microscopy and immunofluorescence, this research introduced pioneering methods involving seven different antibodies, marking a first in the study of reptilian salivary glands. Our investigations categorized the tortoise's salivary glands into papillary and non-papillary types. The papillary glands were further classified into superficial, deep, interpapillary, and intraepithelial salivary glands, while non-papillary glands included superficial and deep lingual types. Structurally, these glands are organized into lobules, delineated by interlobular septa, and are equipped with a duct system comprising interlobular, intercalated, and main excretory ducts with gland openings on the tongue's surface and the papillae surfaces. Notably, the superficial glands displayed both tubuloalveolar and acinar configurations, whereas the deep lingual glands were exclusively acinar. Immunofluorescence results indicated that α-smooth muscle actin (α-SMA) was prevalent in myoepithelial cells, myofibroblasts, and blood vessels, suggesting their integral role in glandular function and support. E-cadherin was predominantly found in epithelial cells, enhancing cell adhesion and integrity, which are critical for efficient saliva secretion. Importantly, Mucin 1 (MUC1) and Mucin 5B (MUC5B) staining revealed that most glands were mucous in nature, with MUC5B specifically marking mucin within secretory cells, confirming their primary function in mucous secretion. PDGFRα and CD34 highlighted the presence of telocytes and stromal cells within the glandular and interlobular septa, indicating a role in structural organization and possibly in regenerative processes. Cytokeratin 14 expression was noted in the basal cells of the glands, underscoring its role in upholding the structural foundation of the epithelial barrier. In conclusion, this detailed morphological and immunological characterization of the Egyptian tortoise's salivary glands provides new insights into their complex structure and essential functions. These findings not only enhance our understanding of reptilian physiology but also underline the critical nature of salivary glands in supporting life in arid environments. This study's innovative use of a broad range of immunofluorescence markers opens new avenues for further research into the adaptive mechanisms of reptiles.

Scanning electron microscopy and morphometric analysis of the hair in dromedaries with SEM-EDX in relation to age.

BACKGROUND: Hair characterization is critical for determining animal individuality throughout life. This study aimed to assess the morphological features of dromedary camel hair in relation to age. MATERIALS AND METHODS: Hair samples were obtained from the camel humps of 30 dromedary camels separated into three groups: G1 (n:10) aged one-year, G2 (n:10) aged 3-5 years, and G3 (n:10) at the age of 8-10 years. The hair was examined using light microscopy, SEM, and SEM-EDX. RESULTS: The Maghrebi camel had varied medulla patterns and structures across the ages. In the G1 group 75% had continuous medulla patterns and amorphous medulla structures, compared to 70% in G2, and 90% in G3. The medulla index increased with age, rising from 0.3 to 0.77%. The shaft width grew in size from G1 to G2, then fell to approximately one-third of the G2 size at G3. The cortex and cuticle widths were also determined by age, and they increased in the G1 compared to G3 camels. The shape of the cuticle scales in G1 camels was wavy, like mountain tops with irregular edges, within G2 camels the scales were particularly long, oval-shaped scales with smooth, wavy borders. The scales of the older G3 camels were quite long and rectangular. SEM-EDX spectra recognized carbon, oxygen, nitrogen, sulfur, calcium, aluminum, silicon, and potassium at the medulla and cortex. Sulfur levels were highest in the G2 samples but lowest in the G1 samples. CONCLUSION: The dromedary camel's hair structure and mineral content, particularly carbon and nitrogen, differed as camels aged.

Novel Insights Into the Architecture of Macro and Microstructures in Cattle Ossa Cordis.

Ossa cordis, bones located within the heart trigones, are often classified as heterotopic or ectopic bones. Despite their high prevalence in cattle and some other bovids, little is known about their structure or development. Scanning electron microscopy, X-ray microtomography, gross dissections, and measurements showed the anatomical locations, prevalence, shapes, and measurements of the cardiac bones in both Egyptian Baladi cattle and Holstein-Friesians. All cattle (n = 12) had an Ossa cordis dextrum (average = 50.70 × 20.91 × 5.40 mm). Additionally, 80% Egyptian Baladi and 57% Holstein-Friesian had a smaller Ossa cordis sinistrum (average = 24.94 × 12.75 × 4.12 mm). Egyptian Baladi Ossa cordis were smaller than observed in Holstein-Friesians. Energy-dispersive X-ray analysis showed the elemental constitution (carbon, oxygen, calcium, nitrogen, phosphorus, sodium, and magnesium) of Ossa cordis and Cartilago cordis. These imaging techniques, plus four histological stains (hematoxylin and eosin, Crossman's trichrome, Alcian blue with Van Gieson, and Sirius Red) and microscopy, demonstrated osteoblasts, osteocytes, osteoclasts, astrocytes, blood vessels, bone marrow, lamellar and woven bone, cortical bone, trabeculations with pores and canaliculi, and fibrous components including collagen in the Ossa cordis dextrum and sinistrum. Hyaline cartilage and fibrocartilage (chondrocytes and cartilage matrix) were found within and surrounding the Ossa cordis. These findings were additionally compared against other cattle breeds and species.

Macroscopic, microscopic, and immunofluorescent characterization of the Greek tortoise (Testudo graeca graeca) oropharyngeal floor with concern to its feed adaptation as a herbivorous land reptile.

The current investigation focuses on gross anatomy, light, and scanning electron microscopy (SEM) of the Testudo graeca oropharyngeal floor, with particular reference to the immunofluorescence technique to examine its tongue. The T. graeca oropharyngeal floor showed many anatomical structures: the lower rhamphotheca, paralingual ridge, lower alveolar ridge, tongue, laryngeal mound, and glottis. The lower rhamphotheca appeared as a V-shaped jaw line with a highly serrated edge and a median tomium (beak). SEM observations of the lingual apex and the lingual body showed rectangular and conical filiform papillae with porous surfaces and taste pores. Meanwhile, the lingual root had two wings that carried papillae with different shapes: dagger-shaped, conical, bifurcated, and leaf-like papillae, and these papillae lacked taste pores. The laryngeal mound had openings for the laryngeal mucus gland and its secretions. Light microscopy findings showed mucous glands in the propria submucosa and near the mucosal surface of the lingual apex. The lingual root had lingual papillae and two hyaline cartilaginous skeletons between skeletal muscles, and the lingual papillae were elongated filiform, rectangular filiform papillae, and fungiform papillae. The lamina propria constituted the core of the lingual papillae and the mucous gland, they had a positive reaction with the periodic acid schiff (PAS) reagent. The apical surface of the fungiform papillae had taste pores. Under immunofluorescence, the vimentin was detected in taste bud cells, and synaptophysin reacted to the taste buds and nerve bundles. The current study of the Greek tortoise oropharyngeal floor investigated its herbivorous eating habits using its serrated lower rhamphotheca, a large tongue with differently shaped papillae, and numerous mucous glands. RESEARCH HIGHLIGHTS: The Greek tortoise (T. graeca graeca) oropharyngeal floor showed many anatomical structures: lower rhamphotheca, paralingual ridge, lower alveolar ridge, tongue, laryngeal mound, and glottis. SEM and light microscopy observations of the tongue revealed varied types and shapes of lingual papillae with a porous surface on the tongue apex (rectangular or conical filiform papillae), on the tongue body (filiform and fungiform papillae), and on the tongue root (dagger-shaped, conical, bifurcated, and leaf-like papillae). Light microscopy findings: the lamina propria constituted the core of the lingual papillae and had numerous mucous glands that had a slightly magenta-red color with PAS reagent. The apical surface of the fungiform papillae had taste pores. Vimentin and synaptophysin gave a reaction to the taste buds.

Comparative morphological analysis of telson and uropods in Penaeus canaliculatus (Olivier, 1811), Penaeus semisulcatus (De Haan, 1844), and Metapenaeus stebbingi (Nobili, 1904) using scanning electron microscopy and EDX analysis.

The telson and uropods collectively form the tail fan, playing crucial roles in locomotion, buoyancy, defense, and respiration. We aimed to compare telson and uropod structures in three shrimp species-Penaeus canaliculatus, Penaeus semisulcatus, and Metapenaeus stebbingi-to identify the species with the most robust telson for its environment. Our analysis involved morphological measurements and scanning electron microscopy (SEM), supplemented by a novel approach-Energy-Dispersive X-ray (EDX) spectroscopy, a technique not previously utilized in studies on these three species. M. stebbingi exhibited the longest telson length, whereas P. semisulcatus had the longest uropod. P. canaliculatus featured a single pair of fixed spines, while P. semisulcatus had evenly spaced small conical spines along the sides of the median elevation and groove. A distinctive feature of M. stebbingi was the telson, which had three pairs of large spines. Diverse setae on telsons included simple, unipennate, and plumose setae. Notably, specialized branched tubular setae on uropods' endopods may aid in grooming or swimming behavior. EDX spectroscopy revealed that the telson cuticle primarily consists of carbon, nitrogen, and oxygen, with significantly high concentrations alongside comparatively lower calcium and phosphorous concentrations. P. semisulcatus exhibited the highest calcium and phosphorus content among the three species. In conclusion, M. stebbingi's telson is structurally robust, emphasizing the importance of morphology, while P. semisulcatus demonstrated a hard telson through EDX analysis. Our study underscores not solely relying on morphology for telson strength assessment but considering telson composition. These variations among species may be attributed to diverse ecological and physiological adaptations.

The tongue of the red-eared slider (Trachemys scripta elegans): morphological characterization through gross, light, scanning electron, and immunofluorescence microscopic examination.

The red-eared slider (Trachemys scripta elegans) is renowned for its remarkable adaptations, yet much of its complex biology remains unknown. In this pioneering study, we utilized a combination of gross anatomy, scanning electron microscopy (SEM), light microscopy, and immunofluorescence techniques to examine the tongue's omnivorous adaptation in this species. This research bridges a critical knowledge gap, enhancing our understanding of this intriguing reptile. Gross examination revealed a unique arrowhead-shaped tongue with a median lingual fissure and puzzle-piece-shaped tongue papillae. SEM unveiled rectangular filiform, conical, and fungiform papillae, with taste pores predominantly on the dorsal surface and mucous cells on the lateral surface of the papillae. Histologically, the tongue's apex featured short rectangular filiform and fungiform papillae, while the body exhibited varying filiform shapes and multiple taste buds on fungiform papillae. The tongue's root contained lymphatic tissue with numerous lymphocytes surrounding the central crypt, alongside lingual skeletal musculature, blood and lymph vessels, and Raffin corpuscles in the submucosa. The lingual striated muscle bundles had different orientations, and the lingual hyaline cartilage displayed a bluish coloration of the ground substance, along with a characteristic isogenous group of chondrocytes. Our research represents the first comprehensive application of immunofluorescence techniques to investigate the cellular intricacies of the red-eared slider's tongue by employing seven distinct antibodies, revealing a wide array of compelling and significant findings. Vimentin revealed the presence of taste bud cells, while synaptophysin provided insights into taste bud and nerve bundle characteristics. CD34 and PDGFRα illuminated lingual stromal cells, and SOX9 and PDGFRα shed light on chondrocytes within the tongue's cartilage. CD20 mapped B-cell lymphocyte distribution in the lingual tonsil, while alpha smooth actin (α-SMA) exposed the intricate myofibroblast and smooth muscle network surrounding the lingual blood vessels and salivary glands. In conclusion, our comprehensive study advances our knowledge of the red-eared slider's tongue anatomy and physiology, addressing a significant research gap. These findings not only contribute to the field of turtle biology but also deepen our appreciation for the species' remarkable adaptations in their specific ecological niches.

A comprehensive exploration of diverse skin cell types in the limb of the desert tortoise (Testudo graeca) through light, transmission, scanning electron microscopy, and immunofluorescence techniques.

The Greek tortoise, inhabiting harsh desert environments, provides a compelling case for investigating skin adaptations to extreme conditions. We have utilized light microscopy, scanning electron microscopy (SEM), transmission electron microscopy (TEM), and immunofluorescence analysis to describe the structure of the arid-adapted limb skin in the Greek tortoise. Our aim was to identify the cell types that reflect the skin adaptation of this tortoise to arid conditions. Utilizing seven antibodies, we localized and elucidated the functions of various skin cells, shedding light on how the tortoise adapts to adverse environmental conditions. Our findings unveiled numerous scales on the limbs, varying in size and color, acting as protective armor against abrasions, bites, and other potential threats in their rocky habitats. The epidermis comprises four layers: stratum basalis, stratum spinosum, peri-corneous layer, and stratum corneum. Cytokeratin 14 (CK14) was explicitly detected in the basal layer of the epidermis, suggesting a role in maintaining epidermal integrity and cellular function. Langerhans cells were observed between epidermal cells filled with ribosomes and Birbeck granules. Numerous dendritic-shaped Langerhans cells revealed through E-Cadherin signify strong immunity in tortoises' skin. Melanophores were identified using the Melan-A antibody, labeling the cytoplasm, and the SOX10 antibody, labeling the nucleus, providing comprehensive insights into melanophores morphology and distribution. Two types of melanophores were found: dendritic below the stratum basalis of the epidermis and clustered oval melanophores in the deep dermal layer. Varied melanophores distribution resulted in a spotted skin pattern, potentially offering adaptive camouflage and protection against environmental challenges. Numerous myofibroblasts were discerned through alpha-smooth actin (α-SMA) expression, indicating that the Greek tortoise's skin possesses a robust tissue repair and remodeling capacity. B-cell lymphocytes detected via CD20 immunostaining exhibited sporadic distribution in the dermis, concentrating in lymphoid aggregates and around vessels, implying potential roles in local immune responses and inflammation modulation. Employing Tom20 to identify skin cells with abundant mitochondria revealed a notable presence in melanophores and the basal layer of the epidermis, suggesting high metabolic activity in these cell types and potentially influencing cellular functions. These findings contribute to our comprehension of tortoise skin anatomy and physiology, offering insights into the remarkable adaptations of this species finely tuned to their specific environmental habitats.

Investigating the role of Purkinje fibers and synaptic connectivity in balance regulation through comprehensive ultrastructural and immunohistochemical analysis of the donkey's (Equus asinus) cerebellum.

The donkey's extraordinary capacity to endure substantial loads over long distances while maintaining equilibrium suggests a distinctive cerebellar architecture specialized in balance regulation. Consequently, our study aims to investigate the intricate histophysiology of the donkey's cerebellum using advanced ultrastructural and immunohistochemical methodologies to comprehend the mechanisms that govern this exceptional ability. This study represents the pioneering investigation to comprehensively describe the ultrastructure and immunohistochemistry within the donkey cerebellum. Five adult donkeys' cerebella were utilized for the study, employing stains such as hematoxylin, eosin, and toluidine blue to facilitate a comprehensive histological examination. For immunohistochemical investigation, synaptophysin (SP), calretinin, and glial fibrillary acidic protein were used and evaluated by the Image J software. Furthermore, a double immunofluorescence staining of SP and neuron-specific enolase (NSE) was performed to highlight the co-localization of these markers and explore their potential contribution to synaptic function within the donkey cerebellum. This investigation aims to understand their possible roles in regulating neuronal activity and synaptic connectivity. We observed co-expression of SP and NSE in the donkey cerebellum, which emphasizes the crucial role of efficient energy utilization for motor coordination and balance, highlighting the interdependence of synaptic function and energy metabolism. The Purkinje cells were situated in the intermediate zone of the cerebellum cortex, known as the Purkinje cell layer. Characteristically, the Purkinje cell's bodies exhibited a distinct pear-like shape. The cross-section area of the Purkinje cells was 107.7 ± 0.2 µm2 , and the Purkinje cell nucleus was 95.7 ± 0.1 µm2 . The length and diameter of the Purkinje cells were 36.4 × 23.4 µm. By scanning electron microscopy, the body of the Purkinje cell looked like a triangular or oval with a meandrous outer surface. The dendrites appeared to have small spines. The Purkinje cells' cytoplasm was rich with mitochondria, rough endoplasmic reticulum, ribosomes, Golgi apparatus, multivesicular bodies, and lysosomes. Purkinje cell dendrites were discovered in the molecular layer, resembling trees. This study sheds light on the anatomical and cellular characteristics underlying the donkey's exceptional balance-maintaining abilities.

Unveiling Cellular Diversity in the Buffalo Corneal Stroma: Insights into Telocytes and Keratocytes Using Light Microscope, Transmission Electron Microscope, and Immunofluorescence Analysis.

Telocytes and keratocytes are important cells that maintain the structure and function of the cornea. The buffalo cornea, known for its resilience in harsh conditions, has not been extensively studied regarding the presence and role of telocytes and keratocytes. We used light microscopy, transmission electron microscopy (TEM), and immunofluorescence assays with platelet-derived growth factor receptor alpha (PDGFRα), CD34, and Vimentin markers to investigate their expression and localization in the cornea. TEM analysis confirmed the presence of spindle-shaped keratocytes with intercellular connections, while telocytes exhibited small spindle-shaped bodies with long, thin branches connecting to corneal keratocytes. Immunofluorescence findings showed that CD34 was more abundant near the endothelium, Vimentin was prominently expressed near the epithelium, and PDGFRα was uniformly distributed throughout the corneal stroma. Co-expression of CD34 and Vimentin, PDGFRα and Vimentin, as well as CD34 and PDGFRα, was observed in keratocytes and telocytes within the stroma, indicating the potential presence of mesenchymal cells. These results suggest the involvement of telocytes and keratocytes in corneal wound healing, transparency maintenance, and homeostasis. The co-expression of these markers highlights the critical role of telocytes and keratocytes in regulating corneal physiological functions, further enhancing our understanding of corneal biology in the buffalo model.

Calbindin Has a Potential Spatiotemporal Correlation with Proliferation and Apoptosis in the Postnatal Rat Kidney.

The protein calbindin-D28k modulates calcium reabsorption in the kidney. Here, we aimed to study the influence of proliferation and apoptosis in different compartments of the kidney on the developmental function of calbindin. Using immunohistochemistry, we investigated the postnatal development of rats' kidneys by using calbindin, proliferative cell nuclear antigen (PCNA), and apoptotic single-stranded DNA (ssDNA). In the neonatal stage (1-day and 1-week-old rats), calbindin showed a positive reaction in the distal convoluted tubule (DCT), a short nephron segment between the macula densa, collecting ducts, and tubules. Moreover, the localization of calbindin was restricted to immature nephrons and mesenchymal tissues. Furthermore, PCNA immunoreactivity was moderate in early-developed podocytes with no reactivity in other renal tubules. The ssDNA immunoreactivity was moderate in the undifferentiated nephron. Then, in the mature stage (3 and 6 weeks old), there was an intense calbindin reaction in DCT but a moderate reaction to PCNA and ssDNA in podocytes. A more intense calbindin reactivity was found in the adult stage (2- and 3-month-old rats) in DCT and collecting tubules. Therefore, in this study, calbindin localization showed an inverse relationship with PCNA and ssDNA of the nephron compartments, which might reflect the efficiency of bone-building and muscle contraction during animal development.