Methyl Green-pyronin Research Articles

Pathological alterations and molecular identification of Marek’s disease in chicken

Marek’s Disease (MD) in chickens, caused by the oncogenic avian herpesvirus known as Marek’s Disease Virus (MDV), poses significant challenges to the poultry industry. This study was aimed at the molecular detection of MDV in afflicted chickens focussing on the gross and histopathological changes. The diagnosis was confirmed by cytological analysis of touch impression smear of the visceral organs with tumour like lesions. Employing polymerase chain reaction (PCR) technique, the presence of the virus was detected in various tissues. Pathological examination revealed characteristic lesions such as lymphoproliferative tumours in various organs including the spleen, liver, kidney, proventriculus, heart and ovary. On microscopic examination, visceral organs revealed densely populated parenchyma with infiltration of pleomorphic lymphoid cells. Out of 70 samples, 17 cases tested positive for MD. The findings provide valuable insights into the occurrence, pathology and diagnosis of MD in chickens, contributing to better disease management strategies and control measures of this economically important poultry disease. Keywords: Marek`s disease, cytology, lymphocytes, methyl green-pyronine

A case of plasmacytoma with widespread metastasis in a dog

In this study, a case of plasmacytoma with widespread metastasis in a nine-year-old male Rottweiler dog was described by histopathological and immunohistochemical methods. Macroscopic examination revealed multifocal white or pink-red colored masses ranging in size from 0,5 to 3 cm in the lung, liver, mediastinal and mesenterial lymph nodes, spleen, heart, pancreas, intestine and kidneys. In microscopic examination, tumor foci consisting mostly of atypical plasma cells were observed in the lung, liver, spleen, pancreas, mediastinal and mesenteric lymph nodes, intestine, and kidneys. Tumor-type giant cells were observed in tumor foci in the lung, mitotic figures in tumor foci in other organs, including the lung, and abundant tumor cells in capillary vessels. In Methyl Green Pyronin staining of lung, liver, spleen, pancreas, mediastinal and mesenterial lymph nodes, intestine and kidney sections, the cytoplasm of plasma cells was observed to be stained pink. Congo Red staining applied to liver and spleen sections showed amyloid deposits stained in brick red color. In addition, in the immunohistochemical staining of lung, liver, spleen, pancreas, mediastinal and mesenterial lymph nodes, intestine and kidney sections with Proliferating Cell Nuclear Antigen (PCNA) antibody, severe immunoreactivity was detected in tumor cells. As a result, due to the presence of rarely reported widespread metastases in the lung, liver, spleen, pancreas, mediastinal and mesenterial lymph nodes, intestine and kidneys of the diagnosed plasmacytoma case, it was deemed appropriate to publish the case in order to contribute to the literature by attracting the attention of veterinarians.

Direct specimen collection during routine operation improves nucleic acid quality in genetic analysis samples for head and neck tumors: A retrospective study

The nucleic acid integrity of head and neck squamous cell carcinoma (HNSCC) samples is poor, and the material available for genetic analysis is limited. Therefore, to expand the effectiveness of personalized medicine in patients with HNSCC, a new sampling method is needed. In total, 128 samples from 44 patients with HNSCC were studied: 32 genetic analysis samples (GASs) collected as 5 × 5 × 5 mm tissue fragments from resected large tumors and immediately embedded in a small formalin bottle within 10 min (i.e., the ischemic time), 43 primary tumor components (primary), 14 decalcified tumor (DC) samples, 32 metastatic tumors in lymph nodes (LNs), and 7 parakeratinized components (PKCs). The nucleic acid quality in the GAS, primary, DC, LN, and PKC groups was compared and next-generation sequencing (NGS) was performed. DNA integrity number and percentage of RNA fragments with > 200 nucleotides were significantly higher in the GAS group than those in the other groups. RNA integrity number decreased first in LN, followed by GAS, primary, and DC. No significant differences were observed in DIN, RIN and DV200 among the PKC, primary and LN. Following methyl green-pyronin staining, preserved DNA and RNA were not visualized in DC samples. Most NGS metrics did not differ significantly among primary, LN, and PKC samples. In conclusion, GASs should be collected during routine hospital activities. When the volume of viable materials is limited, PKCs should be considered for genetic analysis.

Immune synergistic mechanism of recombinant plasmid adjuvant containing chicken IL-4 and IL-2 fusion genes on chicken coccidia live vaccine

The recombinant plasmid pCI-IL-4-IL-2-EGFP containing fusion genes of chicken IL-4 and IL-2 can be used as an adjuvant to enhance the anticoccidiosis effect of the chicken coccidia live vaccine. The chickens were divided into 3 groups: blank control group, vaccine+pCI-IL-4-IL-2-EGFP adjuvant coimmunization group, and vaccine-only group to investigate the immune synergy mechanism of recombinant plasmid adjuvant pCI-IL-4-IL-2-EGFP. The expressions of IL-2, IL-4, TNF-α, and IFN-γ in chicken sera and tissues were detected by ELISA and RT-qPCR, and the proliferation of T and B lymphocytes and antigen presenting cells (APC) in chicken immune organs and intestines were detected by acid alpha-naphthalase (ANAE) staining, methyl green pyronine (MGP) staining, and immunofluorescence (IF) staining, respectively. Results showed that the mRNA expression of IL-2, IL-4, IFN-γ and the number of activated T and B lymphocytes were significantly upregulated in the spleen and cecum tonsils of chickens in vaccine+pCI-IL-4-IL-2-EGFP group compared with the vaccine-only group on 7 d after vaccination (P < 0.05). Protein contents of IL-2, IL-4 and TNF-α in vaccine+pCI-IL-4-IL-2-EGFP group were significantly increased compared to vaccine-only group on 28 d of inoculation (P < 0.05). The number of T and B lymphocytes and APC in chickens of the vaccine+ pCI-IL-4-IL-2-EGFP group was significantly higher than that of the vaccine-only group in cecum tonsils, thymus and spleen after 14 and 28 d of inoculation (P < 0.05). All results revealed that pCI-IL-4-IL-2-EGFP adjuvant enhanced the immune response of chicken coccidia live vaccine by upregulating the expression of IL-2, IL-4, TNF-α, and IFN-γ and promoting the proliferation of T, B lymphocytes and APCs in chicken intestines and immune organ sites. Moreover, our study provides a theoretical basis for the clinical application of cytogenic plasmids as adjuvants.

COMPREHENSIVE ASSESSMENT OF MORPHOLOGICAL CHANGES IN THE LIVER OF RABBITS EXPOSED TO WATER-SOLUBLE SILICON FOR THREE MONTHS

The article presents the results obtained when studying the liver of rabbits who were in an experiment with the intake of silicon with drinking water at a concentration of 20 mg / l expressed in terms of silicon. The aim of the study was a comprehensive assessment of liver changes in laboratory rabbits under the conditions of the experiment with the intake of water-soluble silicon for three months. Material and methods. Rabbits were kept in a vivarium on a standard diet with free access to drinking water for three months. Animals of the control group (n = 3) received bottled drinking water, rabbits of the experimental group (n = 3) received the same water, but with the addition of sodium metasilicate enneahydrate at a concentration of 20 mg /l expressed in terms of silicon. After the animals were removed from the experiment, the liver was extracted. A part of the liver was sent to a cryostat for freezing with further preparation of fresh slices with a thickness of 10 microns and producing a Cross, Evan and Rost reaction to detect histamine luminescent structures, the other part was placed in a 10% formalin solution for subsequent paraffin embedding. Paraffin blocks were made; sections of 5 microns thick after deparaffination were stained with hematoxylin and eosin, methyl green-pyronine by Brachet method, as well as by Van Gieson method to identify connective tissue. The measurement results were considered statistically significant at p &lt; 0.05. Results and their discussion. When stained with hematoxylin and eosin in the liver preparations of rabbits of the experimental group who received silicon with drinking water at a concentration of 20 mg/l ad libitum for three months, statistically significant increases in the average area of hepatocytes, the average area of hepatocyte nuclei, and the average diameter of sinusoidal capillaries were revealed in comparison with the preparations in the control group. An increase in the area of central veins was noted, but these changes were not statistically significant. A change in the shape of hepatocytes' nuclei was observed, which was manifested by a decrease in the shape factor of the nucleus and eccentricity and an increase in the index of the nucleus contour, as well as leukocytic infiltration of the portal tract area, formation of areas of hepatocytes' focal necrosis, proliferation of connective tissue with the formation of portal fibrosis and portoportal septa. In rabbits of the experimental group, a decrease in the DNA optical density was determined in liver histopreparations stained by Brush method in hepatocyte nuclei. We found an increase in the intensity of histamine luminescence in the luminescent granular liver cells of rabbits containing it. Conclusions. The intake of silicon at a concentration of 20 mg/l for three months results in pronounced morphological changes in the liver of rabbits.

Comparison of the Development and Involution Periods of Bursa of Fabricius with Histological and Histochemical Methods

The aim of this study is to histologically and histochemically determine the developmental and involutional stages of bursa of Fabricius of henna partridge (Alectoris chukar). In the study, bursa of Fabricius of 12 3-month-old (6 males, 6 females), 12 6-month-old (6 males, 6 females) henna partridges purchased from a private farm were used. It was observed that bursa of Fabricius was surrounded by a connective tissue capsule and consisted of tunica serosa, tunica muscularis, and tunica mucosa layers from the outside to the inside. It was seen that the tunica muscularis consisted of outer longitudinal and inner circular smooth muscle fibers. It was observed that the tunica mucosa made plicae towards the lumen of the organ and consisted of 10-15 plicae. It was seen that lamina epithelialis and lymph follicles were present in each plica. It was determined that the lamina epithelialis consisted of two parts called FAE (Follicle-Associated Epithelium) and IFE (İnter Follicular Epithelium). It was noted that the lymph follicles contained cortex and medulla sections and were separated locally by capillaries together with cortical medullary boundry cells. In the Methyl Green-pyronin staining method, plasma cells were found in the bursa of Fabricius of the henna partridge, in the connective tissue surrounding the organ, around the blood vessels and inside the follicles. In AB pH=2.5 staining, AB-positive reaction was seen only in the apical part of the epithelial cells forming FAE and IFE in the pre- and post-involution period. In PAS staining, PAS-positive reaction was observed only in the apical part of the epithelial cells forming FAE and IFE in the pre- and post-involution period. In PAS/AB pH=2.5 combined staining method, AB-positive reaction was observed only in the apical part of epithelial cells in the pre- and post-involution period. As a result of this study, it was found that bursa of Fabricius of the henna partridge did not undergo any histochemical changes despite its histologically large differences after involution.

Nuclear and Nucleolar Morphometric Changes in MGP Stained Oral Epithelial Cells In Cigarette Smokers and Non Smokers

Aim: To compare nuclear and nucleolar morphometric changes in methyl green pyronin stained oral epithelial cells in cigarette smokers, passive smokers and non smokers Study design: A comparative cross sectional study Place and duration: Carried out in Postgraduate Medical Institute, Lahore. Duration was 6 months, March 2022 to August 2022. Methods: 120 subjects were equally divided into three main groups, cigarette smokers, passive smokers and non smokers, each fulfilling the inclusion and exclusion criteria. Buccal smears were collected by exfoliative cytology of mucosa and then stained by methyl green pyronin stain. The quantitative nuclear morphometric parameters i.e., nuclear and nucleolar diameter and area, and number of nucleoli, were measured. Results: Themedian of all nuclear parameters of smokers was significantly higher as compared to passive smokers and non-smokers in MGP stain (p ≤ 0.05). Further, effect of duration of smoking on morphometric variables showed significant (p&lt; 0.05) results amongst the smokers in MGP stain. However, no significant differences were seen in all morphometric parameters with respect to frequency of cigarette smoking. Conclusion: The study confirms nuclear morphometric changes in oral epithelial cells of cigarette smokers indicating that smoking does influence cellular alterations. The simplicity of cytology technique with the use of methyl green pyronin stain is valuable screening tool for detecting early oral mucosal changes in high risk patients. Keywords: smoking, exfoliative cytology, nuclear morphometric parameters, methyl green pyronin stain.

Evaluation of Nuclear Parameters in Relation to Regional Lymph Node Involvement in Oral Squamous Cell Carcinoma: A Cytomorphometric Study

Background: Up-regulation of ribosome biogenesis encodes the factors related to carcinogenesis. It has been shown that the nucleus diameter and number of nucleoli increase from normal mucosa to oral squamous cell carcinoma (OSCC). The relationship between nuclear parameters and lymph node involvement in OSCC has not been established, yet. The aim of this study was to evaluate the nuclear parameters comprising nucleoli count and nucleus: nucleoli ratio in relation to regional lymph node involvement in OSCC. Methods: Thirty-four formalin-fixed, paraffin embedded sections from different histopathologic grades of OSCC were stained with methyl green-pyronin. Mean number of nucleoli and nucleus: nucleoli ratio were calculated in 100 tumor cells from 10 random selected fields and compared based on lymph node involvement. Results: Nucleoli count in cases with metastasis to regional lymph nodes was not significantly different from that in cases without regional lymph nodes involvement (P=0.29). The difference of nucleus: nucleoli ratio in cases with and without lymph nodes involvement were not significant (P=0.52). No significant correlation was found between the nucleoli count and lymph node involvement (r=0.08, P=0.78). The correlation between nucleus: nucleoli ratio and lymph node involvement was significant (r=0.58, P=0.02). Conclusion: The nucleus: nucleoli ratio of tumoral cells in OSCC was correlated to lymph node involvement. Based on the results, nucleus: nucleoli ratio can potentially be a useful tool to determine the lymph node involvement in OSCC.

Revisiting Pyronin Y, A Classical Photostable Xanthene Dye useful for Cost Effective Multimodal Fluorescence Microscopy

In search of cost-effective alternatives for developing high-throughput imaging methods for NETosis, several classical histology dyes were revisited. It was found that the second component of the marketed premix Methyl Green-Pyronin Y (Gurr’s Certistain) has several properties compatible with this aim. Therefore, a simple purification method was developed, and the obtained Pyronin Y fraction, dried and redissolved in a biocompatible medium, was found to be photostable, selectively internalized by non-viable cells, and had spectroscopic features that make it compatible with many common fluorophores for multicolor imaging. Under certain conditions, Pyronin Y has a maximum absorption at 548nm, so it can be excited by 514nm and 561nm Laser lines, with an emission peak at 571nm. Experiments with human polymorphonuclear leukocytes (PMN) enzymatically induced to NETosis confirmed the usefulness of Pyronin Y in detecting early NETotic nuclei. By super-resolution microscopy, the compound enhances the visualization of nuclear subdomains, enabling the reduction of assay duration by detecting morphological alterations indicative of initial NETosis steps. Also, in chemically fixed resting PMN, azurophilic granules were stained by Pyronin Y, suggesting a previously unappreciated nucleic acid-independent staining mechanism. In addition, fluorescence lifetime imaging microscopy (FLIM) followed by time domain phasor analysis demonstrates the utility of the dye in differentiating between NETs released into the extracellular space and the genetic material still remaining inside the cell.

Apoptotic investigation of brain tissue cells in dogs naturally infected by canine distemper virus

BackgroundCanine distemper caused by canine distemper virus that belongs to the Morbillivirus genus of the Paramyxoviridae family is still a global epidemic significant infectious disease, especially in pet dogs in China and serious harm to the development of the dog industry. It has been known that apoptosis caused by the canine distemper virus can show in culture cells, lymphoid tissues, and the cerebellum. However, its occurrence in brain tissue cells remains unclear. To investigate the relationship among canine distemper infecting brain tissues, apoptosis in brain tissue cells, and demyelinating pathogenesis was investigated.Methods16 naturally infected dogs that exhibited clinical signs of CD and tested positive for the anti-CDV monoclonal antibody and six healthy dogs that served as the control, were used in the research. Brain specimens were divided into the cerebrum, brain stem, and cerebellum embedded in paraffin and made the sections respectively. Approximately 5 µm-thick sections were stained by hematoxylin–eosin, methyl green pyronin, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling technique, and immunohistochemistry. CDV nucleocapsid protein was detected by immune streptavidin-biotinylated peroxidase complex.ResultsAlterations in the brain tissues of CDV-infected dogs involved both various cells and nerve fibers. CDV had varying degrees of cytotropism to all brain tissue cells; apoptosis also occurred in all brain cells, especially in the endothelia of cerebral vessels, astrocytes, oligodendrocytes, and ependymal cells, the more serious infection, the more obvious apoptosis. Serious infections also involved the pyramidal and Purkinje cells. The nervous fibers exhibited demyelinating lesions (showed small multifocal vacuole), and some axonal neuron atrophy gradually disappeared (formed large vacuole).ConclusionsApoptosis in brain tissue cells was mainly related to the propagation path and cytotropism of CDV. The apoptosis of astrocytes, oligodendrocytes, and some neurons may play a significant role in the demyelinating pathogenesis in dogs with acute canine distemper. A lot of diverse nervous signs shown in the clinic may be related to different neuron apoptosis.

Brain Morphological Changes in COVID-19

The aim of the study was to identify the pathomorphology of brain damage in patients who died of COVID-19.Material and methods. Autopsy reports and autopsy brain material of 17 deceased patients with pre-mortem confirmed COVID-19 infection were analyzed. Fatal cases in which COVID19 was the major cause of death were included in the study. Five people were diagnosed with cerebral infarction. Organ samples were taken for histological examination during autopsy. Sections were stained with hematoxylin and eosin and by Nissl to assess brain histopathology. To study the vascular basal membranes the PAS reaction was used, to detect fibrin in vessels phosphotungstic acid-hematoxylin (PTAH) staining was used, to determine DNA in nuclei sections were stained according to Feulgen, to detect RNA in neuronal nuclei and cytoplasm sections were stained with methyl green-pyronin. Immunohistochemical study of a neuronal marker, nuclear protein NeuN, was performed to assess neuronal damage.Results. The signs of neuronal damage found in patients who died of COVID-19 included nonspecific changes of nerve cells (acute swelling, retrograde degeneration, karyolysis and cytolysis, ‘ghost' cells, neuronophagia and satellitosis) and signs of circulatory disorders (perivascular and pericellular edema, diapedesis, congested and engorged microvasculature).Conclusion. Brain histopathological data indicate damage to the central nervous system in COVID-19 patients. Ischemic stroke in patients with COVID-19 is mostly caused by a combination of hypoxia resulting from respiratory failure and individual risk factors, including cerebrovascular atherosclerosis and hypertension.

Comparative study for identification of fixatives and their influences on vertebral CNS tissues of five species stained with methyl green-pyronin

Comparative study for identification of fixatives and their influences on vertebral CNS tissues of five species stained with methyl green-pyronin

Detection of Apoptosis in Leukoplakia and Oral Squamous Cell Carcinoma using Methyl Green Pyronin and Hematoxylin and Eosin.

Background & Objective:Cell population and turnover are controlled by a balance between cell proliferation and apoptosis. Detection of apoptosis in oral cancer contributes to its better prognosis and improved management. This study aimed to quantify apoptotic cells in leukoplakia and oral squamous cell carcinoma (OSCC) using methyl green-pyronin (MGP) and hematoxylin and eosin (H & E) staining.Methods:The sample included a total of 130 subjects (comprising 108 males and 22 females). Formalin fixed and paraffin embedded tissues were used and categorized into three groups of normal oral mucosa (n=10), leukoplakia with dysplasia (n=60), and OSCC (n=60). The number of apoptotic cells and apoptotic index (AI) were calculated after staining with MGP and routine H & E stained slides.Results:MGP stained the condensed chromatin of apoptotic cells. Statistically significant difference (P≤0.001) was observed among various study groups in terms of numbers of AI and apoptotic cells. Also, AI increased with increasing grades of dysplasia, and it was the highest in well differentiated OSCC. Results were statistically significant in both H & E and MGP stained sections (P≤0.001). A good correlation was found between MGP and H & E staining results.Conclusion:MGP is more specific and can lead to intense staining for chromatin in apoptotic cells. Accordingly, it can provide a good alternative to H&E in identifying apoptotic cells.

Insights into myxozoan composition and physiology revealed by histochemical properties of myxospores.

Myxozoa (phylum Cnidaria) are a diverse group of metazoan parasites that predominately infect fish. Little is known regarding the composition and physiology of their myxospore life stage. The objective of this work was to investigate the composition of myxospores and extrasporogonic stages of nine myxozoan species infecting various teleost fish using histochemical staining techniques. Thirty histochemical stains were applied to formalin-fixed, paraffin-embedded tissues processed routinely for light microscopic evaluation. The polar capsules were the most consistent stain target across the taxa examined. Polar capsule staining with Alizarin red, von Kossa and methyl green-pyronin suggests the presence of intracapsular calcium and phosphate, which may contribute to polar filament discharge or pathogenesis of host invasion. The shell valves and suture lines of most myxozoans were stained with Luna and phosphotungstic acid haematoxylin stains, consistent with the presence of chitin and microfibrils, respectively. Vacuoles were consistently highlighted by diastase-susceptible periodic acid-Schiff and Grocott's methenamine silver staining, indicating glycogen. Other histochemical stains exhibited inconsistent staining across the taxa, suggesting differences in myxospore composition potentially reflective of physiologic variations and tissue tropisms. This work provides some information on conserved features and taxa-associated composition of myxospores and lends insight into myxozoan physiology and host-parasite interactions.

Comparative analysis of nucleomorphometric parameters in methyl green-pyronin-stained sections of oral epithelial dysplasia, oral submucous fibrosis and oral squamous cell carcinoma.

Context:The diagnosis and grading of epithelial dysplasia is based on a combination of architectural and cytological changes. A gradual increase in quantitative DNA aberrations has been found to correlate with increasing degree of dysplasia in oral mucous membranes.Aims:The aim of this study is to assess nuclear parameters in potentially malignant and malignant lesions of the oral cavity and to assess cytomorphometric changes in the nucleus and nucleolus in oral epithelial dysplasia (OED), oral submucous fibrosis (OSMF), oral squamous cell carcinoma (OSCC) and normal oral mucosa using methyl green-pyronin staining to determine its suitability for detecting potentially malignant lesions and the stage of carcinogenesis.Methods:Forty-five archival sections of OED, OSMF and OSCC and 5 cases of normal oral mucosa as the control group were stained according to methyl green-pyronin-staining protocol. Cytomorphometric parameters such as nuclear diameter, nucleolar diameter, number of nucleoli and cytoplasmic RNA were assessed.Statistical Analysis Used:The study was subjected to statistical analysis to evaluate the association between morphometric parameters using analysis of variance test, followed by Bonferroni's post hoc analysis.Results:A progressive increase in the nuclear parameters as well as cytoplasmic RNA content was observed between normal mucosa through dysplasia and OSMF to OSCC.Conclusion:This study serves as an effective diagnostic aid in assessing nuclear parameters in potentially malignant and malignant epithelial lesions affecting oral cavity.

Pathomorphology of cats with myocardial infectious peritonitis

The article presents the results of macroscopic and microscopic examinations of myocardial cats in wet and mixed forms of FIP. A pathoanatomical study of 19 cat carcasses, aged from 3 months to 7 years, was diagnosed with infectious peritonitis during life (on the basis of anamnesis, clinical features, morphological and biochemical blood test, ultrasound, Rivalt test and FCVetx rapid test VetE.) All animals were kept at home. For the microstructural study, samples of cats' hearts were selected, which were fixed in 10% aqueous formalin neutral solution, Carnua, Buen solutions and 96 ° ethyl alcohol. Histogram sections were stained with hematoxylin and eosin staining, picrofuxin (Van Gizon), PAS reaction (McManus), methyl green pyronin (Brache), Malory, and examined under a microscope. Histological examination of the cardiac muscle of cats in various forms of infectious peritonitis revealed changes of non-inflammatory and inflammatory nature. In the exudative form, non-inflammatory processes prevailed. In the myocardium, the most severe changes occurred in the capillaries, the walls of the arterial vessels and the stroma, which were characterized by diapedic hemorrhage, mucoid and fibrinoid swelling and necrosis of the walls of the arterioles. Disorganization of connective tissue was accompanied by stratification of connective tissue fibers and impregnation of weakly oxyphilic, PAS-positive compounds of the intermuscular lumen, which was combined with dystrophic changes in cardiomyocytes. In the mixed form, proliferative-destructive vasculitis, diffuse or focal lymphoid-histiocytic infiltrates in the myocardium prevail. The revealed optical changes in the structural elements of the heart indicated a sharp weakening of the contractile function of cardiomyocytes and heart failure. In addition, it should be noted that the characteristic morphological manifestation for immunocomplex diseases is the development of vasculitis, which is preceded by fibrinoid necrosis of the walls of the arterial vessels and intensive infiltration of their circulatory elements, and these changes occurred in infectious peritonitis of cats.

Histological and immunohistochemical studies of the proximal caecum and caecal tonsils of quail (Coturnix coturnix japonica).

The proximal caecum in quails consists of lymphoid and non-lymphoid structures. The caecal tonsils in the proximal part of the caecum are units of gut-associated lymphoid tissue in poultry. This study aimed to examine the histological characteristics of the proximal caecum, as well as compositions of dendritic cells (DCs) and antigen-presenting cells (APCs) in the caecal tonsil of quails. Tissue sections were stained with Crossman's triple, periodic acid-Schiff, Gordon and Sweet's silver, Congo red and methyl green-pyronin dyes, as well as immunohistochemically by the streptavidin-biotin-peroxidase complex method. Caecal lymphoid tissue was located in the lamina propria and submucosa. Germinative centres were observed within the lymphoid tissue. Reticular fibres were mainly distributed in the border area of the germinal centre with only a few fibres scattered in the centre. Plasma cells were observed in the subepithelial region and germinal centres. Eosinophil granulocytes were prevalent in the lymphoid tissue. Additionally, CD83-immunoreactive DCs and MHC class II immunoreactive APCs were present in the subepithelial area and diffuse lymphoid tissue. While DCs were seen in the germinal centres of tonsillar units, APCs were rarely present in the germinal centres, but they were noticed around the germinal centres. In conclusion, the histological structure of the proximal caecum in quails and the distributions of some immunological cells in the caecal tonsils were revealed. Therefore, the defensive role of the caecal tonsils in the digestive system may be better understood, and comparative studies may be carried out.

The Diagnostic Role of Methyl Green-Pyronin Y Staining in Oral Leukoplakia and Oral Squamous Cell Carcinoma: An Exfoliative Cytology-Based Cytomorphometric Analysis

Background: Oral exfoliative cytology is a noninvasive and nonpainful technique for early diagnosis of oral potentially malignant disorders and oral cancer, and the use of cytomorphometry ameliorates its diagnostic reliability. The objective of the present study was to analyze methyl green-pyronin Y (MGP)-stained oral exfoliated cells (OECs) of oral leukoplakia (OL) and oral squamous cell carcinoma (OSCC) by cytomorphometry. Materials and Method: An observational study was conducted on 150 individuals equally divided into three groups: normal mucosa, OL, and OSCC. Smears were prepared from OECs and stained with MGP. Cytomorphometry was done for 100 cells per subject, and various cell and nuclear parameters were measured and calculated. Results: The Kruskal-Wallis test with post hoc correlation showed significant differences in nucleus and cell diameter (ND, CD), nucleus and cell area (NA, CA), nucleus and cell perimeter (NP, CP), and nucleus to cytoplasmic (N:C) ratio for diameter, perimeter, and area. Spearman’s ρ correlation of various N:C ratio methods showed good correlation between N:C perimeter and diameter ratio, N:C diameter and ellipse ratio, and N:C area and ellipse ratio. Additional morphological factors showed significant relations for both cell and nuclear regularity factor, shape factor, and nuclear contour index. Discussion: MGP-based cytomorphometry showed a significant decrease in CD, CA, and CP and increase in ND, NA, NP, and N:C ratio from normal mucosa to OL and OSCC. MGP proved its worth as an effective stain for OECs, despite its strict standardization.

Visualization of viral DNA assembly using nonfluorescent staining

ABSTRACTWe present an easy test for rapid visualization of viral DNA assemblies in infected cell cytoplasm. We selected the best stains for nuclear staining: Nile blue A, Bismarck brown, gallocyanin chrome alum, methyl green pyronin and azure II. None of the staining techniques is fluorescent, which facilitates their use in everyday experiments. Methyl green is most promising for routine detection of viral DNA assemblies in the cytoplasm; the procedure enables ready detection of viral DNA accumulation in the cytoplasm.

Гістоструктурні показники перешийку яйцепроводу гусок у продуктивному періоді першого циклу яйцекладки

The oviducts of the geese of large grey breed of 8 - 13 months of age were investigated. The macroscopic parameters of the organ were determined in each group. For histological studies, the samples of oviduct isthmus were fixed in a 10% aqueous formalin solution. The histological slides were staining by the following methods: hematoxylin and eosin, tricolor method (by Mallory), methyl green-pyronin (by Brashe), PAS-reaction and alcian blue. It was established that the period of egg-laying in the goose of a large grey breed began at the age of 10 months with the peak of egg-laying at the 11th - 12th months of age. Oviducts of geese of the 8th and 9th month of age are not macroscopically differentiated into divisions. Structural features of the wall of the oyiduct isthmus were determined at the microscopic level. The thickening of the isthmus wall occurred proportionately to the mucous membrane and muscle membranes. The beginning of the formation of the secretory apparatus of the isthmus came from the age of 8 to 9 months of the geese. Since the age of 10 months, the parameters of the thickness of the mucous membrane and the height of folds have not veraciously changed. It is a feature of the completion of the relief formation of mucous membrane and the secretory apparatus. There was a maximum increasing of the diameter of the glands of the mucous membrane for geese of this age group. It is an indicator of the completion of the differentiation of the glandular apparatus of the isthmus at the time of the start of the egg-laying. The morphometric indices of lumen epithelium and glands decreased by almost 1.4 times when egg, surrounded by membranes, is presented in the isthmus. It is caused by the extrusion of the secret and the formation of the material of the subshell membranes. The height of the folds of the mucous membrane and the diameter of the tubular glands in the geese of 13 months of age did not veraciously change, with a decreasing of egg productivity on 35%. Wherein, the thickness of the mucous membrane even increased. Such dynamics of morphometric indices is the result of accumulation and stagnation of the secret in the cytoplasm of glandular cells.