Avidin-biotin Peroxidase Complex Research Articles

Determination of M1/M2 Macrophage Polarization in Ipsilateral and Contralateral Rat Testis Tissue Following Unilateral Torsion/Detorsion

The present study investigates the changes in M1/M2 macrophage polarization resulting from unilateral testicular torsion in the bilateral testis. The study sample included 63 male Sprague–Dawley rats, which were randomly divided into nine groups (n = 7): Control, Sham (4 h (4 h), 24 h, 7 days (7d), 14d), and Torsion/Detorsion (T/D) (4 h, 24 h, 7d, 14d). Histopathological evaluations revealed no changes in the Sham groups, while T/D was noted to cause edema, vascular occlusion, disruption of seminiferous tubule epithelial organization, germ cell abnormalities and structural anomalies in the experimental rats, the severity and extent of which increased from 4 h to 14d after T/D. The Cosentino scores used to determine the degree of histological damage were consistent with the histopathological findings in all groups, while the Johnsen scores, as a marker of spermatogenesis, were lower in the T/D groups. Seminiferous tubule diameters and germinal epithelial thickness decreased significantly in parallel with increased tubule damage in the ipsilateral testicles. Testicular torsion significantly affected sperm motility, with significant reductions observed in the T/D 7d and T/D 14d groups. A hormone profile analysis revealed decreased testosterone levels in both the Sham and T/D groups when compared to the Controls. CD68 and CD163 immunoreactivities, as M1 and M2 macrophage surface markers, were determined in the testicular tissue using the avidin–biotin-peroxidase complex method. T/D interventions caused M1/M2 macrophage polarization changes and increased M1 macrophages, particularly in contralateral testicular tissue. The increase in M1 macrophages in contralateral testicular tissue following T/D in the present study suggests that cell processes, including macrophages, may play an important role in contralateral testicular injury.

Investigation of Immunohistochemical Localization of Oxytocin Receptor in Diabetic and Non-Diabetic Mouse Heart

The aim of this study was to investigate the immunohistochemical localization of oxytocin receptor (OTR) in diabetic and non-diabetic mouse heart tissue. Eighteen male Balb-c adult (8-12 week) mice were used in the study. Animals were divided into three groups; control, sham and diabetes. The diabetes group was given STZ by intraperitoneally (i.p) injections and diabetes was induced. Sham group was again treated with sodium citrate solution by i.p. The animals in the control group did not receive any treatment. After 30 days of STZ application, mice were cervical dislocated under ether anesthesia and their heart tissues were removed. Each heart tissue was vertically divided into two parts and routine histological procedures were applied and then tissues were blocked in paraffin and sections were taken. For histological examination, Haematoxylin&Eosin (H&E), Crossman’s Triple staining and Periodic Acid Schiff (PAS) were applied to the sections. Immunoreactivity of OTR was determined by Avidin-Biotin-Peroxidase Complex (ABC) method. At the end of the study period; the body weight of the groups, blood glucose level, tissue weights and immunohistochemical localization of OTR in heart tissue samples and histological structure of tissue were compared. When weights of heart tissue were compared between the groups, there was no statistically significant difference between the groups (p>0.05). As a result of histological examinations, it was found that there was more degeneration in the cells in the myocardium of the heart in the diabetes group compared to the other groups. Immunohistochemical examinations showed that OTR showed similar immunoreactivity in sham and control groups. In the diabetic group, the immunoreactivity of OTR was similar in endothelial and capillary areas, but less in cell membrane, cytoplasm and purkinje cells. In conclusion, the results of this study showed that there is a significant relationship between the OTR, diabetes and heart tissue. As a result, it is thought that diabetes may have an effect on the cardiovascular system through the OTR (p

The effects of hypoxia-inducible factors-1α and -2α and erythroferrone on hepcidin in patients with chronic kidney disease stages 3–5 and renal anemia

Objective This study aimed to investigate the effects of hypoxia-inducible factor-1α (HIF-1α), hypoxia-inducible factor-2α (HIF-2α), and erythroferrone (ERFE) on hepcidin in patients with chronic kidney disease (CKD) stages 3–5 and renal anemia. Methods A total of 90 patients with CKD stages 3–5 and renal anemia were selected for the study at the Nephrology Department of Fujian Provincial People’s Hospital and divided into three groups, according to CKD stage, while another 30 healthy subjects who underwent a physical examination at the hospital during the same period were selected as the normal group. The serum levels of hepcidin, HIF-1α, HIF-2α, ERFE, and furin were measured using an avidin biotin peroxidase complex enzyme-linked immunosorbent assay to compare the differences between the groups in the related indicators. Results ① Serum HIF-2α, HIF-1α, ERFE, and furin levels increased gradually in the patients with CKD stages 3–5 ( p < 0.05, p < 0.01). ②Simple correlation analysis:Serum hepcidin was positively correlated with HIF-2α, ERFE, and HIF-1α in the CKD patients ( p < 0.01). ③Serum hepcidin was positively correlated with HIF-2α, HIF-1α, and ERFE in the CKD patients injected with erythropoietin (EPO) ( p < 0.01), while serum hepcidin was positively correlated with HIF-2α and HIF-1α ( p < 0.01) in the patients not injected with EPO. ④ Multivariate linear regression analysis showed that HIF-1α, (β = 4.36, p < 0.01), serum ferritin(SF) (β = 0.13, p < 0.01), and HIF-2α (β = 0.66, p < 0.01) were significantly correlated with hepcidin. Conclusion HIF-1α, HIF-2α, and SF are factors which have an effect on hepcidin in patients with CKD stages 3–5 and renal anemia. The increase of HIF-1α, HIF-2α, and ERFE does not seem to inhibit the increase of hepcidin.

Prognostic significance of Ezrin expression in colorectal carcinoma

Objectives This study aimed to evaluate the immunohistochemical expression of Ezrin in various histopathological types of colorectal carcinoma, and to examine its association with various clinicopathological features and so evaluate its prognostic significance. Background Colorectal cancer is considered one of the most common cancers worldwide, it developed because of the cumulative effect of sequential genetic alterations. Thus, more extensive studies are needed to demonstrate the molecular mechanism of colorectal cancer initiation and progression. Ezrin gene has an important role in cell movement, migration, mitosis, and other physiological functions, and is involved in maintaining cell structure and cell motility. Its expression correlates with many human malignancies. Patients and methods Ezrin antibody immunostaining was performed using the avidin–biotin–peroxidase complex method in 204 cases of primary colorectal carcinomas, including 118 (57.8%) cases of adenocarcinoma, 56 (27.5%) cases of a mucoid carcinoma, and 30 (14.7%) cases of signet ring cell carcinoma. Results Cytoplasmic Ezrin expression was significantly higher in lymph-node-positive cases (P Conclusion High Ezrin expression is considered a poor prognostic factor for colorectal adenocarcinoma.

Evaluation of MMP-9 and iNOS expressions in sheep with encephalitic listeriosis

This study aimed to correlate Matrix Metalloproteinase-9 and iNOS expressions with the severity of histopathological findings in tissue samples taken from sheep with encephalitic listeriosis. Thus, the role of these molecules in the pathogenesis of the disease can be elucidated. After systemic necropsy, tissue samples of adult sheeps with meningoencephalitis were investigated by the culture, histopathological and immunohistochemical methods in the presence of Listeria spp. isolation from tissues was performed in accordance with the USDA-FSIS method with some modifications. Tissue samples were fixed in a 10% buffered formaldehyde solution. Following routine procedures, tissue sections at 5 μm were stained with Hematoxylin and Eosin, investigated under light microscope and photographed. Immunohistochemical staining was performed on the tissues using the avidin-biotin immune peroxidase complex method. Listeria spp. were obtained in 20 (83.3%) of 24 tissue samples with the presence of bright grey-black centred smooth colonies on Listeria Selective Agar and identified as Listeria monocytogenes through the phenotypically supportive tests. Liquefaction necrosis, purulent meningoencephalitis, perivascular cuffing, microabscesses and glial nodules were the most important histopathological findings. MMP-9 immunpositive reactions were observed in the cytoplasm of microglial cells and neurons in areas where inflammatory and necrotic areas are concentrated in medulla oblongata and pons. In perivascular cuffing areas, immune reactions in endothelial cells were detected. We detected iNOS positive reactions in the medulla oblangata and pons, especially in inflammatory cells in the microabscesses. Consequently, a positive correlation (p < 0.05) was found between MMP-9 expression and the severity of histopathological findings in sheep with encephalitic listeriosis. expression. In addition, we found that iNOS expression increased in parallel with the increase in MMP-9

Expression of PCNA, MMP-9, p53, Bax and Bcl-2 in canine transmissible venereal tumors

In this study, we aimed to evaluate the proliferative, metastatic and apoptotic capacities of TVT cases, which are in various phases of development, by using immunohistochemical markers. The material of this study consisted of twelve female and six male dogs diagnosed with TVT brought to our department between 2007 and 2020 years. Diff-quick staining was applied to the smear taken from tumoral masses for cytological examinations. Tumoral tissues from dogs were fixed in a 10% neutral buffered formaldehyde solution. After routine tissue procedures Hematoxylin & Eosin stain was applied to the sections. Tissue sections were investigated under a light microscope and photographed. Immunohistochemical staining was performed on the tissues using the avidin-biotin immune peroxidase complex method. As a result of macroscopic, cytological and histopathological examinations, TVT positive cases were divided into three according to their developmental stages. While the expression of PCNA, MMP-9, mutant p53 and Bcl-2 increased significantly in progressive cases compared to regressive and stable cases, Bax expression increased significantly in regressive cases compared to progressive and stable cases. In conclusion, we thought that the mentioned markers are very useful for understanding the prognosis of TVT, the tumor aggressiveness and the survival of the malignant cells.

Comparison of renal lesions in cats and dogs using pathomorphological and ımmunohistochemical methods

ABSTRACTWe investigated the pathogenesis of chronic renal fibrosis in cats and dogs using immunohistochemistry. We used the avidin-biotin complex peroxidase (ABC-P) method with antibodies against transforming growth factor-β1, cytokeratin, E-cadherin, S100A4, alpha-smooth muscle actin, vimentin and nestin to determine whether tubule epithelial cells had undergone epithelial-mesenchymal transformation (EMT) that resulted in loss of epithelial cells and an increased number of mesenchymal cells. Although nephrosis was more common in dogs, nephritis was more common in cats; these pathologies developed in both kidneys. We found that EMT participated in the pathogenesis of renal fibrosis in both dogs and cats.

GLP-1 Localisation and Proglucagon Gene Expression in Healthy and Diabetic Mouse Ileum.

IntroductionGlucagon-like peptide-1 (GLP-1) is a polypeptide that is mainly produced by intestinal L cells and is encoded by the proglucagon gene. In this study, GLP-1 localisation was investigated in the ileum of healthy and diabetic mice by immunohistochemistry and proglucagon gene expression was assayed by reverse transcription-polymerase chain reaction.Material and MethodsThis study included 18 male Balb/c mice that were divided into diabetic, sham, and control groups. Mice in the diabetic group received 100 mg/kg of streptozotocin. Immunohistochemical expression of GLP-1 was determined using the avidin–biotin–peroxidase complex technique, and proglucagon gene expression was determined by RT-PCR.ResultsAnalysis of GLP-1 immunohistochemical localisation showed that GLP-1-immunopositive cells (L cells) were present between epithelial cells in the intestinal crypts. The intensity and localisation of GLP-1 immunoreactivity were similar among the mice in all the groups. Proglucagon gene expression levels were also statistically similar among the mice in all the groups.ConclusionNo difference was demonstrated among the mice in the diabetic, sham, or control groups with respect to proglucagon gene expression and GLP-1 localisation in the ileum, suggesting that diabetes does not affect proglucagon gene expression in the ileum.

Comparison of Peroxidase‐Anti‐Peroxidase and Avidin‐Biotin‐Peroxidase Complex (ABC) Immunohistochemical Staining Procedures for Tryptophan Hydroxylase in Rat Brain

The neurotransmitter serotonin (5‐hydroxytryptamine; 5‐HT) plays a key role in sleep, feeding behavior, motor activity and, importantly, mood and the mechanism of action of antidepressant drugs. Immunohistochemistry for tryptophan hydroxylase (TpOH), the synthetic enzyme for 5‐HT, can be used to identify 5‐HT neurons in the central nervous system. As part of a project examining 5‐HT in brain function in the rat, we compared 2 indirect methods of visualizing primary antibodies against TpOH, a peroxidase‐anti‐peroxidase (PAP) method and the more sensitive avidin‐biotin‐peroxidase (ABC) procedure.A cryostat microtome was used to cut 40 micron sections from brains from rats transcardially perfused with 0.9% saline and 4% p‐formaldehyde in 0.1 M phosphate buffer (PB). Brains were post‐fixed 24 h in p‐formaldehyde and cryoprotected in 20% sucrose in 0.1 M PB. Sections were washed in phosphate‐buffered saline (PBS) before and between all steps. Tissues were treated with 0.4% H2O2 in methanol for 20 min to quench endogenous peroxidase activity and 1% sodium borohydride in 0.1 M PB for 30 min to enhance permeability of the tissue and improve staining quality. Sections were then treated with 1% normal donkey (PAP method) or horse (ABC method) serum in PBS with 0.2% Triton X‐100 detergent for 1 h and then 0.8% bovine serum albumin in PBS for 40 min. Tissues were incubated for 48 h in mouse monoclonal anti‐TpOH antibody (Sigma, 1:1000) in PBS with 0.3% Triton X‐100 and serum. Some sections were incubated without the primary antibody (omit negative controls). For the PAP protocol, tissues were treated with donkey anti‐mouse antibody (Jackson; 1:100) and then mouse PAP (Jackson; 1:400), both in PBS with 0.3% Triton X‐100 for 1 h. For the ABC protocol, tissues were treated with biotinylated horse anti‐mouse antibody and then avidin‐biotin‐peroxidase complex (Vector Elite kit), both in PBS with 0.3% Triton X‐100 and serum for 1 h. All sections were reacted with 0.0005% diaminobenzidine with 0.0001% H2O2 for 45 – 120 sec and mounted on gelatin‐coated slides that were coverslipped with DPX. Stained and negative control sections from both methods were examined using standard light microscopy.Staining using the PAP method was faint, homogenous and only slightly darker than negative controls. Individual neurons could not be identified. Increasing concentrations (1:500, 1:250, 1:100) of anti‐TpOH antibody had no effect on specific labeling but slightly increased background staining. This was surprising because the exact same reagents and PAP procedure has produced outstanding staining of tyrosine hydroxylase in brainstem catecholamine neurons (e.g., Muma et al., Exp. Neurology, 168:135–143, 2001). In contrast, labeling of TpOH using the ABC method was excellent. TpOH immunoreactivity was highly visible in the somata and primary dendrites of neurons in the dorsal and median raphe nuclei. These results add to and amplify the enhanced staining quality of the ABC procedure over the PAP method in formaldehyde‐fixed rat brain sections.Support or Funding InformationSupported by the School of Biological Sciences and the Department of Psychology, Illinois State University.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Cultural and immunohistochemical evaluation of bacterial agents in caprine pneumonia in Nigeria

Goats are important protein-producing animals but pneumonia, in spite of the huge investments in small ruminant production, remains a major limitation. This study evaluates the prevalence of important bacterial pathogens associated with caprine pneumonia in Nigeria using cultural and immunohistochemical techniques. One hundred and fifty goat lungs were randomly examined from macroscopic, cultural isolation and microscopic changes using standard techniques. The common bacteria identified were formalin inactivated for bacterine and polyclonal antibody production. Immunohistochemical staining was performed against the bacteria antigens using the avidin-biotin peroxidase complex technique. The data were presented in frequencies and percentages, and compared using non-parametric statistics at α = 0.05. The pathology in the caprine lungs included broncho-interstitial pneumonia (41), interstitial pneumonia (27), and bronchopneumonia (72). Ten of the lungs were normal. Bacterial isolation yielded Mannheimia haemolytica (40%), Pasteurella multocida (20%), Escherichia coli (13%), Klebsiella pneumoniae (10%), Streptococcus sp. (8%), Baccillus sp. (5%), and Pseudomonas sp. (3%). Mannheimia haemolytica (Mh) and Pasturella multocida (Pm) were the most common isolated bacteria. The bacterial antigens were detected on the surface of respiratory bronchiolar, and alveolar epithelia, within inflammatory exudate and in cytoplasm of alveolar macrophages. One hundred twenty (80%) of the caprine lungs were positive for the Mh and Pm antigens; 47 for Mh, 59 for Pm, 14 for both, and 30 negative for either antigens. Significant differences were observed in the staining intensities of the antigens. This study validated Mannheimia haemolytica and Pasteurella multocida as most important bacterial pathogens in caprine pneumonia in Nigeria. Thus, adequate control of caprine pneumonia should consider bacterine or multivalent vaccines incorporating these important bacteria to curb morbidity and mortality of goats.

Immunohistochemical expression of Mum-1, Oct-2 and Bcl-6 in systemic anaplastic large cell lymphomas.

Several transcription factors predominantly used for B-cell lineage identification are also expressed in a small percentage of T cells within germinal centers and interfollicular areas. The aim of the study was to evaluate the expression of Mum-1, Oct-2 and Bcl-6 in systemic anaplastic large cell lymphoma. Thirty cases of anaplastic large cell lymphoma were retrieved from our archives and tissue microarray constructed. Immunohistochemistry was carried out using an avidin-biotin peroxidase complex method. A predominance of nuclear staining was observed for all transcription factors. Mum-1 was positive in all but one case (96.7%). Half of the cases displayed Oct-2 expression (15/30 cases). A considerable number of cases also had Bcl-6 expression (9/30). Bcl-6 staining was noted to be more common in ALK positive cases. Our findings emphasize that these markers are not restricted to B-cell lineage and that extensive expression can be observed in anaplastic large cell lymphoma of T/null cell phenotype.

148 Inmunolocalization of β-Nerve Growth Factor (NGF) in Male Reproductive Tract and NGF Levels in Serum and Seminal Plasma at Puberty and Adulthood in Rabbit

β-Nerve growth factor (β-NGF) is a neurotrophin with different roles in reproduction that could be regulated by sexual hormone concentrations. The objective of the present study was to characterise β-NGF in the accessory glands and epididymis of male rabbit and to determine whether its expression differentially changed near puberty (week 22) and adulthood (week 37) according to serum testosterone levels and NGF levels in both serum and seminal plasma (SP). Semen and blood were collected every week from 6 males during 3 months and analysed at 3 time points (weeks 22, 30, and 37) by ELISA. The SP was separated from sperm cells by centrifugation (3,000 × g for 15 min at 4°C) and stored at –20°C; blood was collected in EDTA tubes, centrifuged 15 min at 700 × g at 4°C and stored at –20°C too. Reproductive tissues (prostate, bulbouretral gland, and caput and cauda of epididymis) were collected at the beginning (week 22; n = 4) and at the end (week 37; n = 4) of the experiment. For tissue recovery, males were killed and glands and epididymis were dissected, fixed in modified Bouin’s fluid, and mounted in paraffin. The ELISA for β-NGF (abx259154, Rabbit NGF ELISA kit, Abbexa, Cambridge, UK) and testosterone (DE1559, Demeditec Diagnostics, Kiel, Germany) were performed following the kit protocols. For immunohistochemistry (IHC), goat polyclonal anti-NGF antibody was used in a dilution 1:100 (N8773, Sigma Aldrich, St. Louis, MO, USA). The avidin-biotin-peroxidase complex (ABC) method was performed with the Vectastain ABC kit (Vector Laboratories, Burlingame, CA, USA) and then slides were contrasted with hematoxylin. Results of serum testosterone levels revealed a significant decrease (P < 0.05) at week 37 (0.57 ± 0.18 ng mL−1) from week 30 (4.68 ± 0.86 ng mL−1) and week 22 (3.38 ± 0.99 ng mL−1). However, β-NGF levels at week 37 in serum (299.32 ± 60.22 pg mL−1) and in SP (947.29 ± 249.45 pg mL−1) also decreased but there were no significant differences. Also, we found no correlation between β-NGF in serum and SP and testosterone levels. β-Nerve growth factor was immunolocated in prostate of both ages where epithelial cells were highly stained. At 22 weeks old, stain was located mostly in the apical zone of the cytoplasm, whereas at 37 weeks old, the protein was localised in the entire cytoplasm of the cell. Furthermore, the interstitial tissue in adult males had a moderate stain in contrast with younger males, which did not show signal in that tissue. The content in the prostate lumen was stained too. Bulbourethral glands had a low signal in interstitial tissue that seems to be greater at week 37. Caput and cauda of epididymis of both ages were not stained. These results suggest that β-NGF has different immunolocation in the rabbit male accessory glands depending on the age of animal that could be related to changes in serum testosterone levels. However, these differences were not correlated with β-NGF levels neither in blood serum nor in SP. Research funded by AGL2015-65572-C2-2-R Grant and Predoctoral Contract UCM-Santander.

Studi Distribusi Glukosa Transporter 4 pada Otot Skelet Ayam Kedu Cemani

Glucose transporter (GLUT 4) is glucose transporter protein regulated by insulin, found in adipose tissue and striated muscle (skeletal and cardiac muscle). Kedu cemani chicken is one of Indonesia endemic animal, found in Kedu, Temanggung regency, Central Java. This study was required to complete microscopic documentation of Indonesia’s native biodiversity. The objective of this study was to clarify GLUT 4 distribution in skeletal muscle fibers of kedu cemani chicken by using avidin-biotin-peroxidase complex (ABC) immunohistochemistry method. This study was conducted by using pectorales major, biceps brachii, and biceps femoris muscle tissue from 5 kedu cemani chicken. The result showed that GLUT 4 immunoreactivity were detected in sarcolemma and myofibrils component of pectorales major, biceps brachii, and biceps femoris muscle tissue. Intensity of GLUT 4 immunorectivites increased from weak intensity in pectorales major muscle tissue, moderate intensity in biceps brachii muscle tissue, then strong intensity in biceps femoris muscle tissue. This result might motivate to further exploration about the other kedu cemani chicken specific features to complete microscopic documentation of Indonesia’s native biodiversity.

Substance P and Calcitonin Gene-Related Peptide in the Glands of External Auditory Canal Skin.

ObjectivesThe earwax (cerumen) that covers external auditory canal (EAC) skin contains a mixture of ceruminous and sebaceous gland substances, such as lipids, peptides, and proteins. The components secreted from the ceruminous gland that is a modified sweat gland form cerumen and contain several antimicrobial factors. Since substance P (SP) and calcitonin gene-related peptide (CGRP), known as a secretagogue, have been found in sweat glands, our purpose was to determine the expression of SP and CGRP in the glands of EAC skin.Methods Sections of normal human EAC skins were immunostained for the presence of SP and CGRP using polyclonal antibodies. Immunoreactivity was detected using an avidin-biotin peroxidase complex method.Results SP staining was found in ceruminous gland acini and myoepithelial cells. But the SP staining was not found in the sebaceous glands and epidermal region. CGRP was strongly stained in the ceruminous gland and weakly in the sebaceous gland cells. Interestingly, most prominent staining of SP and CGRP was noted in the myoepithelial cells of the ceruminous gland.Conclusion The findings in this study suggest that SP and CGRP are expressed in the glands of the EAC skin and secreted in the process of ceruminous gland secretion.

Identification of CD146 as a novel molecular actor involved in systemic sclerosis

We highlight for the first time that CD146/sCD146 is involved in fibrotic process during SSc. sCD146 could thus constitute a new biomarker to assess disease activity, and potentially a new target for therapeutic applications.

COMPARATIVE STUDIES FOR DETECTION OF BRUCELLA MICRO-ORGANISM IN ABORTED BOVINE FOETI USING CONVENTIONAL, IMMUNOHISTOCHEMICAL AND MOLECULAR METHODS

The present study was carried out on a total number of 78 bovine (cows48- Buffaloes 30) The obtained sera were investigated for detection of brucella-antibody titer using serological tests Rose Bengal plate Test (RBPT), Tube agglutination test (TAT), indirect ELISA (iELISA) and Complement Fixation Test (CFT). Abomasal content were collected from 78 aborted fetuses for bacteriological and Polymerase Chain Reaction (PCR) assays. The results of serological analysis revealed that the positive reactors were 60 (76.92 %), 55 (70.51 %), 58 (74.36 %) and 57 (73.08 %) using RBPT, TAT, iELISA and CFT, respectively. RBPT and iELISA tests showed the highest seropositivity. Meanwhile, the lowest ones were obtained by TAT and CFT tests. Brucellaem elitensis biovar3 was isolated from 31 out of 48cows (abomasal content) aborted fetuses and21 out of 30buffaloes (abomasal content) aborted fetuses. PCR assay for detection of Brucella in aborted fetuses were 33 out of 48 in cow and 22 out of 30 in buffaloes. Pathological examination in organs of aborted fetuses infected by Brucella melitensis showed significantly pathological lesions in lungs, liver, spleen and placenta. Immunohistochemistry revealed the presence of positive immunostaining brucella antigen in formalin-fixed, paraffin imbedded tissue sections of lung, liver, spleen, and placenta by using avidin–biotin complex peroxidase technique. Electron microscopical finding of lung, liver and spleen showed that the cytoplasm of neutrophil and macrophages containing dark bodies of cocobacilli. TEM of placenta revealed that the Trophoblasts filled with degenerated organisms.

Expression of epidermal growth factor-like domain 7 may be a predictive marker of the effect of neoadjuvant chemotherapy for locally advanced uterine cervical cancer.

Neoadjuvant chemotherapy (NAC) followed by hysterectomy may be effective for the treatment of locally advanced uterine cervical cancer and improve patient prognosis. It is important to identify markers that are able to predict whether NAC may be successful. Epidermal growth factor-like domain 7 (EGFL7) regulates vascular sprouting in blood vessel formation. In numerous types of human cancer, EGFL7 is upregulated and inhibits endothelial cell adhesion molecules, decreasing vascular tightness and, thus, increasing vascular permeability. It is considered that the overexpression of EGFL7 is able to inhibit drug delivery, resistance to apoptosis and the epithelial-to-mesenchymal transition. In the current study, 63 patients with locally advanced uterine cervical cancer were reviewed and classified as stage IIIA-IIIB using the International Federation of Gynecology and Obstetrics criteria. These patients (aged <70 years) were treated at Osaka City University Medical School Hospital, Japan, between 1995 and 2010. Tumor tissue samples were obtained by biopsy prior to NAC. The tissue samples were classified as group 1 or 2 depending on the efficacy of NAC. Surgery and radiotherapy were administered in group 1 (n=36), for which NAC was effective. In the patients of group 2 NAC was not effective, and radiotherapy alone was administered (n=27). The expression of EGFL7 and Snail was examined in paraffin-embedded tissue sample sections using the avidin-biotin peroxidase complex method. The results indicated that EGFL7 expression levels were significantly higher in group 2, as compared with group 1 (P<0.001). A similar result was observed for the expression levels of Snail (P=0.001). Group 1 exhibited significantly longer overall survival times compared with group 2 (P=0.001). The patients were also classified into a low EGFL7 expression level group (weighted score of ≤6) and a high EGFL7 expression level group (weighted score of ≥8). NAC was observed to be significantly more effective in the low EGFL7 expression level group (P<0.001), as compared with the high expression level group. The results suggest that the expression levels of EGFL7 may be a potential predictive marker of the efficacy of NAC for the treatment of locally advanced uterine cervical cancer.

Repeated asenapine treatment does not participate in the mild stress induced FosB/ΔFosB expression in the rat hypothalamic paraventricular nucleus neurons

Effect of repeated asenapine (ASE) treatment on FosB/ΔFosB expression was studied in the hypothalamic paraventricular nucleus (PVN) of male rats exposed to chronic mild stress (CMS) for 21days. Our intention was to find out whether repeated ASE treatment for 14days may: 1) induce FosB/ΔFosB expression in the PVN; 2) activate selected PVN neuronal phenotypes, synthesizing oxytocin (OXY), vasopressin (AVP), corticoliberin (CRH) or tyrosine hydroxylase (TH); and 3) interfere with the impact of CMS. Control, ASE, CMS, and CMS+ASE treated groups were used. CMS included restraint, social isolation, crowding, swimming, and cold. From the 7th day of CMS, rats received ASE (0.3mg/kg) or saline (300μl/rat) subcutaneously, twice a day for 14days. They were sacrificed on the day 22nd (16–18h after last treatments). FosB/ΔFosB was visualized with avidin biotin peroxidase complex and OXY, AVP, CRH or TH antibodies by fluorescent dyes. Saline and ASE did not promote FosB/ΔFosB expression in the PVN. CMS and CMS+ASE elicited FosB/ΔFosB-expression in the PVN, whereas, ASE did not augment or attenuate FosB/ΔFosB induction elicited by CMS. FosB/ΔFosB-CRH occurred after CMS and CMS+ASE treatments in the PVN middle sector, while FosB/ΔFosB-AVP and FosB/ΔFosB-OXY after CMS and CMS+ASE treatments in the PVN posterior sector. FosB/ΔFosB-TH colocalization was rare. Larger FosB/ΔFosB profiles, running above the PVN, did not show any colocalizations. The study provides an anatomical/functional knowledge about an unaccented nature of prolonged ASE treatment at the level of PVN and excludes its positive or negative interplay with CMS effect. Data indicate that long-lasting ASE treatment might not act as a stressor acting at the PVN level.

Apoptosis in natural rabies virus infection in dogs

Abstract Introduction: In the present study apoptosis was investigated in the cornu ammonis and cerebellum of 10 dogs naturally infected with rabies virus. Diagnosis of rabies was based on the results of fluorescent antibody staining and experimental inoculation. Material and Methods: The paraffin tissue sections were stained with haematoxylin and eosin, avidin-biotin complex peroxidase (ABC-P), and terminal deoxynucleotidyl transferase biotin-dUTP nick end-labelling (TUNEL) methods. Results: Histopathological examination revealed encephalomyelitis of varying severity and the presence of Negri bodies. Dense rabies antigens were determined in the motor neurons with ABC-P method. On the other hand, Bcl-2 protein and Bax protein gave positive reaction in seven and five cases, respectively. TUNEL staining demonstrated very marked apoptotic changes in the nuclei of neurons localised deep in the substantia alba of the cerebellum. Similar changes were also determined in perivascular mononuclear cells and glia cells within the substantia alba. No apoptopic changes were found in the motor neurons of the cornu ammonis. Conclusion: The absence of apoptotic changes in the neurons was considered to be the consequence of the necrotic changes that developed in these neurons.

Impact of repeated asenapine treatment on FosB/ΔFosB expression in the forebrain structures under normal conditions and mild stress preconditioning in the rat

Long-term effect of asenapine (ASE), an atypical antipsychotic drug, on FosB/ΔFosB quantitative variations in the striatum, septum, nucleus accumbens, and prefrontal cortex, was light microscopically evaluated in normal rats and rats preconditioned with chronic unpredictable mild stress (CMS). CMS included restraint, social isolation, crowding, swimming, and cold. The rats were exposed to CMS for 21 days. From the 7th day of CMS, the rats were injected subcutaneously with saline (300μl/rat) or ASE (0.3mg/kg b.w.), twice a day for 14 days. On the 22nd day, i.e. 16-18h after the last treatment, the animals were perfused with fixative and the brains cut into 30μm thick coronal sections. FosB/ΔFosB protein was immunohistochemically visualized by avidin-biotin peroxidase complex (ABC). Four groups of animals were investigated: control+vehicle, control+ASE, CMS+vehicle, and CMS+ASE. Repeated ASE treatment significantly increased the amount of FosB/ΔFosB immunostained cell nuclei in the dorsolateral and dorsomedial striatum and the shell of the nucleus accumbens, followed by strVM and coACC, as assessed by numerical analysis in both total (different size for each structure) and unified (equal size for each structure) brain sectors. The effect of ASE was significantly lowered by CMS preconditioning only in the dorsolateral striatum, dorsomedial striatum, and the shell of the nucleus accumbens, indicated by both total and unified calculations. Although, highest FosB/ΔFosB expression was seen in the prefrontal cortex and lowest in the dorsolateral and ventrolateral septum, no differences between the groups occurred. CMS itself did not affect FosB/ΔFosB expression level. These findings demonstrate for the first time that repeated administration of ASE may result in eliciting of long-lasting FosB/ΔFosB-like transcription factors that could mediate some of the persistent and region-specific changes in brain function, interconnected with chronic drug exposure. However, it cannot be excluded that the impact of repeated ASE exposure might be influenced by an ambient stressogen leverage.