Semithin Sections Research Articles

Ultrastructure of the mammary gland in lactating Saanen goats

The secretory cells of the mammary gland are very diverse in their size, shape, and structure. These are highly specialized cells adapted to the synthesis, accumulation, storage, and excretion of milk. The main feature of glandular cells is the presence of secreted substances. Electron microscopy shows that substances of different secretory cells types have various electron densities. Occasionally, two or more types of secretory granules differing in size and composition can be found in one cell. The material for the morphological study was small (2‐4 mm) tissue samples of goat lactating mammary glands. Pieces were taken from deeper areas of the breast parenchyma. Ultrathin sections with a 50‐70 nm thickness were obtained for electron microscopic analysis using Leica UC7 ultramicrotome. Electron microscopy was performed using a JEOL JEM 1011 microscope. Micrographs were obtained using a Morada camera (Digital Imaging Solutions Inc.). The secretion of the mammary gland occurs according to the apocrine type. Picture of the lactocytes' cytoplasm apical part separation into the lumen of the alveoli is often observed on semi‐thin sections. Thus, numerous droplets of material secreted by lactocytes are found in the lumen of the alveoli. The shape of lactocytes in the lactating mammary gland is close to cubic. The height of the epithelial layer, generally, is 10‐12 microns (µm). The milk ducts, like the alveoli, are dilated. Most of them contain a significant amount of osmiophilic material secreted by the cells of the mammary alveoli. The cells of the epithelium of the single‐layered ducts have a flattened or cuboidal shape. The diameter of the ducts varies noticeably depending on location in the gland parenchyma. The intralobular milk ducts are 10‐20 µm in diameter. At the same time, the larger interlobular ducts reach a diameter of 40‐50 microns and are located in groups in the connective tissue stroma of the mammary gland. Notably, an increase in the mammary alveoli mass in the lactating gland is associated with a decrease in connective tissue volume and the almost complete disappearance of fat cells groups from it. The milk ducts of the teat zone in the lactating gland are strongly expanded, round, or slightly oval‐shaped in cross‐section with a diameter of 30‐60 microns. In contrast to the deeper zones of the mammary gland, we were unable to detect secreted substances in the dilated milk ducts near the teat area. The milk ducts we studied are located close to the most terminal sections of the mammary gland excretory system (gland cistern and teat canal). The milk is inevitably washed out from the ducts during chemical fixation and subsequent stages of objects preparation for histological analysis.

OsCPK29 interacts with MADS68 to regulate pollen development in rice

Pollen development and its germination are obligatory for the reproductive success of flowering plants. Calcium-dependent protein kinases (CPKs, also known as CDPKs) regulate diverse signaling pathways controlling plant growth and development. Here, we report the functional characterization of a novel OsCPK29 from rice, which is mainly expressed during pollen maturation stages of the anther. OsCPK29 exclusively localizes in the nucleus, and its N-terminal variable domain is responsible for retaining it in the nucleus. OsCPK29 knockdown rice plants exhibit reduced fertility, set fewer seeds, and produce collapsed non-viable pollen grains that do not germinate. Cytological analysis of anther semi-thin sections during different developmental stages suggested that pollen abnormalities appear after the vacuolated pollen stage. Detailed microscopic study of pollen grains further revealed that they were lacking the functional intine layer although exine layer was present. Consistent with that, downregulation of known intine development-related rice genes was also observed in OsCPK29 silenced anthers. Furthermore, it has been demonstrated that OsCPK29 interacts in vitro as well as in vivo with the MADS68 transcription factor which is a known regulator of pollen development. Therefore, phenotypic observations and molecular studies suggest that OsCPK29 is an important regulator of pollen development in rice.

STRUCTURAL FEATURES OF LINGUAL MINOR SALIVARY GLANDS UNDER THE EXPERIMENTAL COMBINED EFFECT OF FOOD ADDITIVES

Under certain conditions, some food additives may have a negative effect on the state of the organs of the digestive system. Morphofunctional changes in the minor salivary glands of different types induced by consumed food have been insufficiently studied. The purpose of this experimental study is to investigate the structural features of the mucous and serous lingual minor salivary glands after combined exposure to monosodium glutamate, sodium nitrite, Ponceau 4R in the dynamics for 4 weeks. Matherials and Methods: 30 outbred albino rats were involved into the experiment. The standard ration for the rats of the experimental group was supplemented with complex food additives: monosodium glutamate, Ponceau 4R, sodium nitrite for 1 and 4 weeks. The rats were killed under thiopentone anesthesia overdose. Conventional histological microslides and semi-thin sections were made from the tongue specimens according to the standard technique. Results. The average diameter of the acini of the mucous lingual salivary glands of intact animals was 55.7±2.89 μm. In the glands of this type, the parenchyma and the stroma was accounted for 70% and 30%, respectively. The average diameter of the secretory parts of the serous lingual salivary glands of intact animals was 32.25±3.59 μm. In the serous lingual salivary glands the parenchyma and the stroma accounted for 75.9% and 24.1%, respectively. Following 4 weeks after taking complex food additives with standard ration for the laboratory animals, the relative amount of parenchymatous component in the mucous salivary glands significantly increased (91,0±0,30)% (p=0,01), whereas the stromal component decreased (9,09±0,26)% (p=0,01). The increase in the average diameter of acini (62.44±0.96 μm (p=0.01) was detected. There was a similar tendency in serous glands. The parenchymatous and stromal component accounted for (86.3±1.13) % and (13.67±1.13)%, respectively. The average diameter of acini was 29.63 ± 0.96 (p = 0.03) μm. Conclusion. Four week-long consumption of complex food additives (monosodium glutamate, Ponceau 4R, sodium nitrate) supplemented with standard ration by the laboratory animals leads to an increase in the relative amount of parenchymatous component in the lingual salivary glands, which occurs due to hypertrophy of the acini. The described processes are more pronounced in the mucous lingual salivary glands compared to the serous ones.

Influence of novel readthrough agents on myelin protein zero translation in the peripheral nervous system

Translational readthrough-inducing agents have been developed for the treatment of nonsense mutations in hereditary diseases. The clinical effectiveness of readthrough agents has been reported, although newly developed agents are still desired because of their toxicities or limited clinical effectiveness. Recently, novel negamycin-derived readthrough agents without antimicrobial activity have been developed. Our aim was to evaluate the activities of these readthrough agents by monitoring the production of large myelin protein zero (L-MPZ), the programmed translational readthrough isoform of myelin protein zero (P0, MPZ) mRNA, and to clarify the influence of these agents on the sciatic nerve in vivo. First, we examined the readthrough activities of novel negamycin-derived agents using cell-free and cell culture systems using plasmids encoding human MPZ (hP0) cDNA. Three of the negamycin derivatives, TCP-112, TCP-169, and TCP-1109, suppressed the canonical stop codon to induce readthrough. Direct injection of TCP-1109, which showed higher readthrough activity for Mpz in mouse sciatic nerves, exhibited a 1.3-fold increase in the L-MPZ/P0 ratio compared to that with the vehicle control on western blotting. The nerve conduction velocity and beam walk test showed abnormalities in the classical readthrough agent G418-treated group, but not in the TCP-1109-treated group. Immunofluorescence analysis showed that TCP-1109 caused less damage to the sciatic nerve than G418. In the semi-thin sections, a lower g-ratio and more tomacula-like structures were observed in TCP-1109-treated nerves. Thus, the present results indicate that negamycin-derived readthrough agents enhance programmed translational readthrough, and the management of readthrough activities using canonical stop codons may be important.

Analysis of the extranucleolar ribonucleoprotein particles of Cycas revoluta Thunb. (Cycadaceae) and Ceratozamia mexicana Brongn. (Zamiaceae)

Background: Nuclear ribonucleoprotein particles play a key role in RNA processing and in the gene expression pathway. Interchromatin granules (GICs) involved in the metabolism of pre-messenger RNA (pre-mRNA) were described in Allium cepa and Chiranthodendron pentadactylon . Other particles as Lacandonia granules (LGs) were found in Lacandonia schismatica as well as Ginkgo biloba and Welwitschia mirabilis . LGs are structures equivalent to perichromatin granules (PCGs) described in mammals and to Balbiani ring granules (BRGs) described in the midge Chironomus tentans . PCGs and BRGs are involved in the metabolism of messenger RNA (mRNA). Here, we analyze the extranucleolar particles from Cycas revoluta and Ceratozamia mexicana and compare them to GICs and LGs using conventional electron microscopy and atomic force microscopy.
 Species study: Cycas revoluta (Cycadaceae) and Ceratozamia mexicana (Zamiaceae)
 Hypothesis: The extranucleolar ribonucleoprotein particles in the nuclei of C. revoluta and C. mexicana are equivalent to GICs or GLs.
 Methods: Fragments of young leaves of C. revoluta and C. mexicana were processed for standard transmission electron microscopy. Thin sections were stained with the EDTA technique preferential for ribonucleoproteins and osmium amine specific for DNA. From the semithin sections the samples were studied with the AFM and images of them were obtained.
 Results: Ribonucleoprotein particles 32 nm in diameter are present in the interchromatin and perichromatin space in C. revoluta and C. mexicana .
 Conclusion: Ribonucleoprotein particles present in the cell nuclei of C. mexicana and C. revoluta are ultrastructurally equivalent to LGs .
 
 
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Phylogenetic position of Atriophallophorus minutus (Trematoda: Microphallidae), the type-species of the genus Atriophallophorus Deblock & Rosé, 1964, based on partial 28S rDNA gene sequence.

We found metacercariae of a microphallid trematode Atriophallophorus minutus in freshwater snails Bithynia tentaculata. In this study, we provide a morphological description of whole-mount specimens and semi-thin sections of experimentally grown adults of this species. Our morphological examination supports the idea that the adults of Atriophallophorus spp. have a ventral sucker with a sinistral interruption of the outer edge. In the 28S rDNA gene-based phylogenetic analyses, our specimens of A. minutus were grouped into Atriophallophorus spp. clade, as a sister taxon to Atriophallophorus winterbourni+Atriophallophorus sp. Analysis of the pairwise genetic distances between coI mtDNA gene sequences revealed a low divergence between the two specimens of A. minutus (1.1%) and a greater divergence (up to 16.6%) between them and A. winterbourni. Since other Atriophallophorus spp. are known to have a strict specificity to the polyvalent intermediate host, we suggest that A. minutus reported from different snail species may represent a complex of cryptic species.

High-quality, large-scale, semi-thin, & ultra-thin sections of the optic nerve in large animals: An optimized procedure

Large animal model of optic nerve (ON) injury is an essential tool for translational medicine. Perfusion fixation with paraformaldehyde is mainly used for preparing the semi-thin (1–2 μm thick) and ultra-thin (<0.5 μm thick) sections of the ON tissues. However, this conventional fixation technique in large animals needs a large volume of fixatives, which increases the risk of toxic exposure and is environmentally unfriendly. Additionally, fixed residual ON cannot be used for other tests that require fresh tissue samples. Although conventional immersion fixation is feasible for preparing a semi-thin section of the ON in small animals (0.2–0.6 mm in diameter), it faces technical challenges when fixing the ON of large animals (3 mm in diameters), as increased diameter limits the permeability of the fixatives into deeper tissue. Therefore, we optimized the immersion-fixation method to obtain high-quality, large-scale, semi-thin, and ultra-thin sections for the ON of goat and rhesus macaques. Using this optimized technique, the ON microstructure was well preserved throughout the entire area of 1.5*1.5 square millimeters, allowing confident quantification of axon density/diameter on semi-thin section and identification of specific organelles and glial cells on ultra-thin sections.Furthermore, the optimized technique is a quick, simple, and environmentally friendly fixation method. Notably, the ON regions of large animals with or without an intact neurovascular system can be prepared for light and electron microscopy. In contrast, the residual unfixed ON from the same animal can be further utilized for experiments such as tissue culture and biomolecular tests.

Functional Adaptation of Vocalization Revealed by Morphological and Histochemical Characteristics of Sonic Muscles in Blackmouth Croaker (Atrobucca nibe).

Simple SummarySound production is common in numerous fish species. Some species can emit calls through the contraction of specialized muscles called sonic or drumming muscles. The sonic muscles of fish are among the fastest muscles in vertebrates. Although numerous studies have investigated the mechanism underlying sound production in fish, only the distinct features of the sonic muscles of a few species have been investigated. We demonstrated that the sonic muscles have functionally adapted for fast twitching and fatigue resistance, which support vocalization in the blackmouth croaker (Atrobucca nibe).Sound production in the blackmouth croaker (Atrobucca nibe) was characterized using acoustic, morphological, and histochemical methods. Their calls consisted of a train of two to seven pulses; the frequency ranged from 180 to 3000 Hz, with a dominant frequency of 326 ± 40 Hz. The duration of each call ranged from 80 to 360 ms. Male A. nibe possess a pair of bilaterally symmetric sonic muscles attached to the body wall adjacent to the swim bladder. The average diameter of the sonic muscle fibers was significantly shorter than that of the abdominal muscle fibers. Semithin sections of the sonic muscle fibers revealed a core-like structure (central core) and the radial arrangement of the sarcoplasmic reticulum and myofibrils. Numerous mitochondria were distributed within the central core and around the periphery of the fibers. Most of the fibers were identified as Type IIa on the basis of their myosin adenosine triphosphatase activities, but a few were identified as Type IIc fibers. All sonic muscle fibers exhibited strong oxidative enzyme activity and oxidative and anaerobic capabilities. The features suggest that the sonic muscles of A. nibe are morphologically and physiologically adapted for fast twitching and fatigue resistance, which support fish vocalization.

A unique pre-endothelial layer at the posterior peripheral cornea: ultrastructural study

This study was conducted to investigate the ultrastructure of a unique structures at the anterior side of the endothelium of the posterior peripheral cornea and compare their inner fibers to those of the limbus and sclera. The unique structures at the anterior side of endothelium was referred as a pre-endothelial (PENL) structures in the present manuscript. Ten anonymous-donor human corneoscleral rims (leftover after corneal transplants) were processed for electron microscopy. Semi-thin sections were examined using an Olympus BX53 microscope, and ultrathin sections were studied using a JOEL 1400 transmission electron microscope. A unique PENL structures was identified at the posterior peripheral cornea at a radial distance of approximately 70–638 µm, from the endpoint of Descemet’s membrane. The PENL thinned out gradually and disappeared in the center. The contained an electron-dense sheath with periodic structures (narrow-spacing fibers), wide-spacing fibers, and numerous microfibrils. Typical elastic fibers were present in the sclera and limbus but were not observed in the PENL. This study revealed the existence of a new acellular PENL, containing unique fibrillar structures that were unseen in the corneal stroma. From the evidence describe in this paper we therefore suggest that PENL is a distinct morphological structure present at the corneal periphery.

Novel protocol to observe the intestinal tuft cell using transmission electron microscopy.

ABSTRACTThe tuft cell is a chemosensory cell, a specific cell type sharing the taste transduction system with a taste cell on the tongue, of which the existence has been discovered in various tissues including the gastrointestinal tract, gall bladder, trachea and pancreatic duct. To date, electron microscopic approaches have shown various morphological features of the tuft cell, such as long and thick microvilli, tubulovesicular network at the apical side and prominent skeleton structures. Recently, it has been reported that the small intestinal tuft cell functions to initiate type-2 immunity in response to helminth infection. However, the mechanisms by which such distinguished structures are involved with the physiological functions are poorly understood. To address this question, a combination of physiological study of tuft cells using genetic models and its morphological study using electron microscopy will be required. However, it is a challenge to observe tuft cells by electron microscopy due to their extremely low frequency in the epithelium. Therefore, in this paper, we suggest an advanced protocol to observe the small intestinal tuft cell efficiently by transmission electron microscopy using serial semi-thin sections on Aclar film. This article has an associated First Person interview with the first author of the paper.

Imaging the Kidney with an Unconventional Scanning Electron Microscopy Technique: Analysis of the Subpodocyte Space in Diabetic Mice.

Transmission electron microscopy (TEM) remains the gold standard for renal histopathological diagnoses, given its higher resolving power, compared with light microscopy. However, it imposes several limitations on pathologists, including longer sample preparation time and a small observation area. To overcome these, we introduced a scanning electron microscopy (SEM) technique for imaging resin-embedded semi-thin sections of renal tissue. We developed a rapid tissue preparation protocol for experimental models and human biopsies which, alongside SEM digital imaging acquisition of secondary electrons (SE–SEM), enables fast electron microscopy examination, with a resolution similar to that achieved by TEM. We used this unconventional SEM imaging approach to investigate the subpodocyte space (SPS) in BTBR ob/ob mice with type 2 diabetes. Analysis of semi-thin sections with secondary electrons revealed that the SPS had expanded in volume and covered large areas of the glomerular basement membrane, forming wide spaces between the podocyte body and the underlying filtering membrane. Our results show that SE–SEM is a valuable tool for imaging the kidney at the ultrastructural level, filling the magnification gap between light microscopy and TEM, and reveal that in diabetic mice, the SPS is larger than in normal controls, which is associated with podocyte damage and impaired kidney function.

Multi-target and fast DNA-paint imaging of neuronal tissue using exchange paint and artificial neural networks

The dense nanoscale morphology and arrangement of proteins within the synapse of neuronal tissue was difficult to visualize until the development of superresolution microscopy. Here, we show the application of multi-target DNA-PAINT in semi-thin (400 nm) brain sections of the medial nucleus of the trapezoid body (MNTB) from rat tissue. Four targets were imaged in situ to visualize the organization of calyces of Held presynapses, proteins within the active zone, and the postsynaptic principal cells with its postsynaptic density.

Comparative Morphology of the Symbiont Cultivation Glands in the Antennae of Female Digger Wasps of the Genus Philanthus (Hymenoptera: Crabronidae).

Females of the solitary digger wasp tribe Philanthini, called the beewolves (Hymenoptera, Crabronidae), cultivate strains of symbiotic bacteria that belong to the genus Streptomyces in unique and highly specialized glands in their antennae. The glands consist of large reservoirs that are surrounded by numerous gland cell complexes (class III). The symbionts are cultivated inside the reservoirs and are probably provisioned with nutrients secreted from the surrounding glands and/or sequestered from the hemolymph. The wasp female delivers the bacteria into the subterranean brood cell prior to oviposition. Fully grown larvae take up the bacteria and apply them to their cocoon. There the bacteria produce several antibiotics that protect the wasp offspring against fungus infestation. Hitherto Streptomyces bacteria were detected in the antennae of 38 species of the Philanthini. However, a detailed morphological analysis of the antennal glands is only available for a few species. In order to shed light on the evolutionary history of the association between beewolf wasps and bacteria, we investigated the morphology of the antennal glands of another 14 Philanthus species from the Palearctic, Paleotropic, and Nearctic. We generated 3D-models of the glands based on serial semithin sections and/or micro-CT (μCT). Despite broad similarities in number and structure of antennal glands, the results revealed interspecific differences with regard to overall shape, complexity, and relative size of the reservoirs as well as the number of the surrounding gland cell units. Mapping the morphology of all species studied so far on the phylogeny (that parallels geographical distribution) revealed that related species share similarities in gland morphology, but there are notable differences between lineages. In particular, compared to the North American species the European and African species possess more complex gland structures with a higher number of gland cells. We discuss morphological, ecological, and physiological aspects and provide scenarios for the evolution of the antennal glands of the Philanthini as symbiont cultivation organs.

A Comparison of the Information Provided by Paraffin, Epon Semi-Thin and Thin Sections on the Examination of Prepubertal Rat Testicular Tissue

A Comparison of the Information Provided by Paraffin, Epon Semi-Thin and Thin Sections on the Examination of Prepubertal Rat Testicular Tissue

THE ISSUE OF HISTOLOGICAL IDENTIFICATION OF М-CELLS IN THE PEYER'S PATCHES OF ALBINO RAT SMALL INTESTINE.

The aim: Based on the above cytological signs of M-cells, we set the goal of more detailed clarification of some of their topological relationships with other enterocytes in the follicle-associated epithelium of Peyer's patches of albino rat small intestine. Materials and methods: 10 mature albino male rats weighted 200,0±20,0 g were involved into the study. Anatomical dissection with the sampling of the sections of the small intestine containing Peyer's patches was carried out with subsequent embedment of the latter into paraffin blocks and making of serial histological sections of 4 μm thick in the cross-section of the small intestine, followed with hematoxylin-eosin staining. The specimens were studied and documented on the "Konus" light microscope equipped. Morphometric characteristics of the specimen tissue structures were studied using the Sigeta X 1 mm/100 Div.x0.01mm stage micrometer. Results: The findings of the study revealed enterocytes with phagocytic properties found in the lymphoid-associated epithelium of Peyer's patches of the small intestine of albino rats. Moreover, if they are clearly visualized at the light-optical level, then M-cells are poorly recognizable, which is consistent with a similar assessment made by other authors. Conclusions: Given this, the issue on the topology and functional purpose of M-cells remains uncertain to date and, thereby, the prospect of further research is being outlined, which, in our opinion, can be successful using the method of stereomorphological analysis. For this purpose, multilayer plastic reconstruction methods can be used for serial semi-thin sections of Peyer's patches embedded in epoxy resin, according to the requirements of transmission electron microscopy.

Ultrastructure characteristics of primordial germ cells in stage X of pheasant (Phasianus colchicus) embryo.

Stage X is one of the formation stages in birds at which the blastoderm area is distinguished by two areas of area pellucida being responsible for formation of embryonic tissues and primordial germ cells, and area opaca forming the extra-embryonic tissues. Primordial germ cells are multi-potent stem cells giving rise to spermatogonia or oogonia. The present study was carried out to describe the characteristics of primordial germ cells in stage X of pheasants' embryo using a transmission electron microscope. The blastoderm was dissected out from embryos which were already incubated for 12 hr. Toluidine blue was used for staining semi-thin sections; lead citrate and uranyl acetate were also used to stain ultra-thin sections. Images of primordial germ cells elucidated that the nucleus was situated eccentrically and had a compact spherical structure. Moreover, the nucleolus appeared elongated and was located eccentrically. The cytoplasm was composed of yolk granules and glycogen particles. Mitochondria were observed as round structures in the cytoplasm. The most important finding was that the primordial germ cells contained yolk granules, mitochondria and small amount of glycogen at this stage.

Extragenital Pathology and Immunocompetent Cells Relations of Lactating Breast Gland and Offspring Jejunum

Researched the effect of chronic toxic maternal hepatitis on the cytometric features of immunocompetent cells (ICCls) of mammary gland tissues of the female and the jejunum of the offspring during breastfeeding. For morphological researches, pieces of the right inguinal mammary gland of female outbred rats with heliotrine hepatitis and the jejunum of the lean intestine of the offspring on days 1, 3, 7, 15 and 21 of lactation were used. Cytometric researches were carried out on semi-thin sections stained with methylene blue and pyronin G. It was established that in the last stages of pregnancy and in the first week after the onset of lactation, there is an increase in migration to the terminal secretory sections of the mammary gland of the ICCls, which indicates the participation of the mammary glands in the transfer of adoptive immunity to offspring and in maintaining the immune homeostasis of the small intestine, which decreases as the immune apparatus of the small intestine develops. A decrease in the number of ICCls migrating to the lactating mammary gland in case of mother's CTH negatively affects both immune adaptation and the structural and functional development and maturation of the immune apparatus of the jejunum of the offspring during breastfeeding.

Тканевый и ультраструктурный анализ печени крыс препубертатного возраста при субтоксическом воздействии кадмия и свинца

Using light and electron microscopy, the effect of intoxication with cadmium sulfate (Cd) and lead acetate (Pb) for 3 weeks on the architectonics of the liver parenchyma and ultrastructural features of hepatocytes of 40 prepubertal male Wistar rats was studied, taking into account the model of the classical hepatic lobule. Animals were divided into 4 groups: 1st – control; 2nd – cadmium sulfate solution at a dose of 0.5 mg/kg; 3rd – lead acetate solution 10 mg/kg; 4th – a combination of these solutions. According to stereological analysis of semi-thin sections, statistically significant indicators of periportal zones reflect an increase in the volume of hepatocytes when exposed to Cd (2nd and 4th groups). In all groups, with the exception of the 3rd one (Pb), an increase in the parenchymal compartment in the porto-central gradient was revealed, indicating the formation of hepatocellular ballooning. In all experimental groups, for the periportal zone of the hepatic lobule, a characteristic feature of the ultrastructure of hepatocytes was a statistically significant decrease in the volumetric density of the endoplasmic reticulum; for the centrilobular zone – of mitochondria; for the perivenular zone – mitochondria and nuclei of hepatocytes. The formation of autophagosomes with signs of predominantly mitochondrial utilization attracted attention in groups exposed to Pb. Pathological changes are compensated by increased transsinusoidal metabolism and complexes of biosynthesis organelles.

МОРФОЛОГИЧЕСКИЙ МОНИТОРИНГ ЭКСПЕРИМЕНТАЛЬНОГО ФИБРОЗА ПЕЧЕНИ У КРЫС

Background. Though thioacetamide (TAA)-induced liver fibrosis (LF) is recognized as a classical model of toxic liver damage, there is no literature data on the description of its successive stages of histological and ultrastructural changes in various cell populations involved in fibrosis. Objective. To conduct morphological monitoring of fibrosis formation in the liver of rats using the TAA model of LF based on histological and ultrastructural changes in hepatocytes and perisinusoidal lipocytes (HSC). Material and methods. The experiment was carried out on 18 sexually mature male rats. LF was modeled by intraperitoneal injection of 2% TAA solution at a dose of 10 ml / kg every other day. Light microscopy of semi-thin sections of the liver was performed, as well as electron microscopy of ultrathin sections. Results. The study of semi-thin sections of rat liver tissue from the control group showed a normal architecture of the parenchyma, a large number of HSCs containing large lipid droplets ("resting" phenotype), a very small amount of cytoplasmic matrix poor in membrane organelles. In the animals that were receiving TAA for 4 weeks, a mesenchymalepithelial transition of HSCs from the "resting" type to a fibrogenic state (fibrogenic phenotype) was recorded, that was accompanied by a gradual decrease in the number of retinol-containing drops and the appearance of fibroblastlike cells (FLC) in HSCs. In the animals, that were receiving TAA for 12 weeks, the pool of fibrogenic cells in the liver increased, a mesothelial-mesenchymal transition occurred, characterized by the mesothelial cell migration deeper into the parenchyma and their acquisition of a mesenchymal phenotype. Lipid containing activated FLC were also found in fibrous tissue around the central vein. Foci of hepatic tissue destruction caused by necrosis and apoptosis of hepatocytes were much more common. Conclusions. Administration of TAA induces liver fibrosis while histological and ultrastructural monitoring of the state of hepatocytes and HSCs allows to monitor all stages of fibrosis, clarifying the mechanisms of damage to intracellular organelles and variants of hepatocyte death. This model of LF in rats can be used to test new antifibrotic drugs.

Стан нейронально-астроцитарного апарату гіпокампу на тлі тривалого введення ізоніазиду та рифампіцину за умов корекції мікрофлори товстого кишечника

The purpose of the study was to determine the morphometric and ultrastructural features of neurons and glial cells, as well as the levels of glial fibrillar acid protein in the hippocampus of rats under long-term administration of isoniazid and rifampicin during experimental therapy with a combination of pro/prebiotics. Materials and methods. The studies were conducted on three groups of white Wistar male rats with drug-induced hepatitis, reproduced by intragastric administration of isoniazid and rifampicin for 28 days. In the last 14 days of the intragastric experiment, rats of the research group were administered probiotic and prebiotic. The content of cytosolic glial fibrillar acid protein in the hippocampus was determined by competitive enzyme-linked immunosorbent assay. Semi-thin sections of CA1 sections of the rat hippocampus were analyzed using the Image J. analysis program. The ultrastructural characteristics were studied using a PEM-100-01 transmission electron microscope (Selmi, Ukraine). Results and discussion. Long-term administration of isoniazid and rifampicin was associated with changes in the content of the cytosolic fraction of glial fibrillar acid protein obtained from the hippocampus. These changes were characterized by a tendency to increase the level of this protein by 17.8% (p=0.2) compared with intact animals. In contrast to the group of animals reproduced in the experiment drug-induced liver injury, in the experimental group, the level of glial fibrillar acid protein was significantly lower by 25.0% compared to the drug-induced liver injury group (p &lt;0.05). Morphological analysis revealed a decrease in neuronal density in rats with drug-induced liver injury compared with intact control. At the same time, the specific number of degeneratively altered neurons in comparison with intact control in the group of drug-induced liver injury animals increased by 8.57 times. The number of degeneratively altered neurons in the group receiving yogurt / lactulose was 55.87 ± 4.23%, which was significantly higher than intact control rats, but was 19.4% (p &lt;0.05) lower levels in rats with MIUP. Electron microscopic examination of rat hippocampal neuroglia was based on the study of changes in the astrocytic and oligodendroglial components. Changes in rat hippocampal neuroglia under experimental conditions were mainly related to the astrocytic link. Astrocytes were characterized by significantly lower cytoplasmic edema and adaptive-compensatory changes in the cell. Conclusion. Course administration of pro/prebiotics reduces the severity of manifestations of neurodegeneration, improves the state of astroglia in the hippocampus, and reduces the cytoplasmic levels of glial acidic fibrillar protein in animals with drug-induced liver injury