Browsing by Author "Balcan, E"
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Item Regressing amphibian tail as a model for the cadherin/β-catenin complex disruption and glycosylation alteration during epithelial apoptosisSahin, M; Balcan, EEpidermis is one of the many tissues that are resorbed during metamorphosis in the regressing tail of amphibian tadpoles. Apoptotic mechanisms play an important rote in this process. In this study, loss of intercellular contacts and alterations in plasma membrane glycosylation were observed during apoptosis. The cadherin/beta-catenin complex represents one of the major adhesive systems in multiple epithelial. tissues. Here, we analysed the fate of cadherin/beta-catenin complex and alterations of plasma membrane glycoconjugate compositions in apoptotic epithelial cells. Our results showed that the cadherin molecules were cleaved into extracellular and beta-catenin associated cytosolic domains by an intracellular mechanism. However, the extracellular domains were probably removed completely by matrix metalloproteinases. Lectin histochemistry studies suggested that mannose and alpha(2 --> 6) linked (but not alpha(2 --> 3) linked) sialic acids were major sugar motifs in plasma membranes of apoptotic tadpole epithelial cells. Although previous studies indicated reduced levels of sialic acid residues during apoptosis, elevated Sambucus nigra agglutinin (SNA) reactivity might be due to the degradation of high molecular weight glycoproteins (probably including cadherin) that masked the SNA-binding residues of the plasma membrane prior to apoptosis. (C) 2008 Elsevier GmbH. All rights reserved.Item The glycosylation status of murin postnatal thymusBalcan, E; Gümüs, A; Sahin, MDuring the intrathymic development, the fate of the thymocytes depends largely on variable expression of CD4/CD8 markers and T cell receptor protein expressions. In addition, changes of cell surface glycosylation status also affect the thymocyte maturation. In this study the glycosylation alterations in thymic tissues from 1, 9, 13 and 16 days old mice were evaluated by histochemical and lectin blotting techniques. With alcian blue (AB) at pH 5.7/periodic acid-Schiff (PAS) stainings, it was shown that thymic microenvironments contained carboxlylated and sulfated glycosaminoglycans (GAGs). Strong positivity to AB at pH 2.5, which specific for sialomucins, was seen in some medullary thymocytes. Similarly, it was shown that with Maackia amurensis agglutinin (MAL) medullary thymocytes, but not cortical ones, contained alpha(2 -> 3) linked sialic acid structures. On the other hand, while reaction with peanut agglutinin (PNA), which specific for core disaccharide galactose beta(1 -> 3) N-acetylgalactosamine, was only seen in cortical thymocytes, reaction with Galanthus nivalis agglutinin (GNA), which specific for terminal mannose residues, was seen in both cortex and medulla. However, Datura stramonium agglutinin (DSA), which recognizes galactose beta(1 -> 4) N-acetylglucosamine, was not only cell-specific, but it was bound some thymic vessels. With lectin blotting studies, five glycoprotein bands of molecular weights similar to 39, similar to 54, 100, similar to 110 and similar to 212 were found which reacted with MAL, PNA and DSA as well as GNA. These results suggest that glycosylation patterns of cell surface glycoconjugates are modified during thymocyte selection processes of postnatal days.Item The Effect of Tunicamycin on Embryonic and Newborn Murine Spleen TissuesBalcan, E; Arslan, Ö; Gümüs, A; Sahin, MTunicamycin is an antibiotic that widely used in cell biology for its ability to inhibit N-linked glycosylation of asparagine residues on proteins and to induce endoplasmic reticulum stress. In the present study, the effects of tunicamycin on murine splenic tissues at 17th embryonic day and 1st and 3rd postnatal days were evaluated with three structural and physiological parameters: 1) alterations in glycosaminoglycan molecules, 2) apoptosis and 3) alterations in laminin molecules. It was shown that splenic microenvironments of control groups contain carboxylated glycosaminoglycans, but their content slightly decreased in all tunicamycin treated groups by alcian blue-periodic acid-Schiff method. On the other hand, there was an increase in the interstitial space among the cells of tunicamycin treated groups. In addition, it was shown by immunoblotting analyses that expression levels of laminin molecules were decreased by tunicamycin tratment in developing spleen tissues. In order to determine apoptotic effects of tunicamycin, TUNEL assay was performed and it was found that the number of apoptotic cells in developing spleen was strongly increased with tunicamycin treatments. These results suggest that, during the spleen development, the alterations of glycosaminoglycan contents in the extracellular matrix and the glycosylation status of extracellular glycoproteins (e. g. laminins) that mediate cell-extracellular matrix interactions are very important factors that seal the fate of cell physiology and morphogenesis.Item Effect of Tunicamycin on Glycosaminoglycans and Laminins in Embryonic and Postnatal Thymic TissuesBalcan, E; Arslan, ÖOBJECTIVE: To compare histological and molecular alterations in the embryonic and neonatal thymi following exposure to tunicamycin. STUDY DESIGN: Mouse embryos at gestational days 17 (n=7) and 18 (n=7) and newborn animals at postnatal days 1 (n=5) and 3 (n=5) were divided into two groups: control and tunicamycin-treated. Combined Alcian blue and Periodic acid Schiff sequences immunohistochemistry and immunoblotting were performed to determine glycosaminoglycan (GAG) accumulation and laminin expression in control and tunicamycin-treated embryonic and postnatal thymi. The apoptotic effect of tunicamycin was evaluated by TUNEL assay. RESULTS: In the control group acidic GAGs first appeared in medullary cells at postnatal day 3, whereas treatment with tunicamycin promoted the accumulation of acidic GAGs in all treated groups as of embryonic day 17. However, tunicamycin slightly decreased the laminin expression, and the number of apoptotic cells was considerably increased after tunicamycin treatment. CONCLUSION: Results suggest that carboxylated and acidic GAGs are two presumptive candidates to establish the thymic microenvironment during the late fetal development and postnatal periods of mice and that tunicamycin would be implicated in this establishment by increasing the acidic GAG accumulation and by reducing the laminin expression and the thymic stromal cell population.Item Acute responses of strength-related gene expressions to maximum strength and force sense acuityAltun, M; Balcan, E; Batir, S; Gökmen, MH; Özgünes, S; Öztel, ZBackground/aim: Although high muscle strength worsens the sense of force, it is unknown whether there is a relationship between this deterioration and the underlying molecular mechanisms. This study examined the relationship between decreased force sense (FS) acuity and strength -related gene expressions. Materials and methods: Maximal voluntary isometric contraction (MVIC) and FS (50% MVIC) tests were performed on the knee joints of twenty-two subjects. The expression analyses were evaluated by qRT-PCR in blood samples taken before, after MVIC, after 50% MVIC, and 15 min after the test. Results: MVIC and FS error values were significantly correlated with each other (r = .659, p = .001). The qRT-PCR analyses demonstrated that the expressed mRNAs of the interleukin 6 (IL -6), alpha-actinin 3 (ACTN3), angiotensin-converting enzyme (ACE), brain -derived neurotrophic factor (BDNF), and ciliary neurotrophic factor receptor (CNTFR) genes dramatically increased until 50% MVIC and subsequently decreased 15 min after the exercise (p < .05). The muscle -specific creatine kinase (CKMM), myosin light chain kinase (MLCK), and G -protein beta 3 subunit (GNB3) genes reached their peak expression levels 30 min after MVIC (p < .05). ACE and ACTN3 gene expression increased significantly in parallel with the increased FS error (p < .05). These gene expression fluctuations observed at 50% MVIC and after the rest could be related to changes in cellular metabolism leading to fatigue. Conclusion: The time points of gene expression levels during exercise need to be considered. The force acuity of those whose maximal force develops too much may deteriorate.Item Quantitative approach to lectin-based glycoprofiling of thymic tissues in the control- and the dexamethasone-treated miceBalcan, EDexamethason (DEX) is the most commonly used synthetic glucocorticoid in treatment of various inflammatory conditions. Here we focused on evaluating the effect of DEX on apoptosis and glycan profile in the mouse thymic tissues. Histological examinations revealed that the DEX treatment cause severe alterations in thymus, such as disruption of thymic capsule, impaired epithelial cell-thymocyte contacts, cellular loss and increased apoptosis. The identification of thymic glycans in the control- and the DEX-treated mice was carried out by using a panel of five plant lectins, Maackia amurensis agglutinin (MAA), peanut agglutinin (PNA), Sambucus nigra agglutinin (SNA), Concanavalin A (ConA) and wheat germ agglutinin (WGA). Lectin histochemistry results showed that glycosylation pattern of thymus changes upon DEX treatment. For further detailed quantitative analyses of the binding intensities for each lectin, histochemical data were scored as high positive (HP), mild positive (MP) and low positive (LP) and differences among signaling densities were investigated. The staining patterns of thymic regions observed with lectin histochemistry suggest that DEX can affect the thymic glycan profile as well as thymocyte apoptosis. These results are consistent with the opinion that not only sialic acid, but also other sugar motifs maybe responsible for thymocyte development. (C) 2016 Elsevier Ltd. All rights reserved.Item Cell surface glycosylation diversity of embryonic thymic tissuesBalcan, E; Tuglu, I; Sahin, M; Toparlak, PIn the thymus, glycosylation status of many cell surface molecules changes during the thymocyte maturation and selection processes. In this study, we evaluated the glycosylation changes and possible relationships with programmed cell death in the thymic tissues from mouse embryos at the days 14 (E14), 15 (E15), 16 (E16), 17 (E17) and 18 (E18) of embryonic development. In order to determine glycosylation changes we used three different plant Lectins: peanut agglutinin (PNA), Moackia amurensis leucoagglutinin (MAL or MAAI) and Sombucus nigra agglutinin (SNA), which recognize core disaccharide galactose (1-3) N-acetylgalactosamine [Gal beta(1 -> 3)GalNAc], siatic acid linked (2 -> 3) to galactose [SA alpha(2 -> 3)Gal] and siatic acid linked to galactose [SA)alpha(2 -> 6)Gal] structures, respectively. Our lectin histochemistry and lectin blotting studies indicated that glycosylation pattern was modified in thymocytes at the embryonic developmental. stages analyzed. The immature cortical thymocytes were labeled by PNA, whereas medullary thymocytes were positive for MAL and SNA binding. Many medullary thymocytes exhibited alpha(2 -> 6)-linked siatic acid on their surface and this increased throughout the gestational stages. In the lectin blotting studies, different protein bands of various molecular weights were identified in thymocytes. Two of them were putatively identified as CD43 and CD45 glycoproteins. In addition, TUNEL (deoxynucleotdyltransferase-mediated dUDP nick end labeling) indicated that only PNA-positive cortical thymocytes were deleted in all embryonic stages. These results indicate that the glycosylation pattern was modified in thymocytes at all embryonic developmental stages, and these modifications can affect the T cell deletion, probably via the galectin-1 molecule in the embryonic thymus. (C) 2007 Elsevier GmbH. All. rights reserved.Item Glycoconjugate histochemistry of mucous glands in the skin of metamorphosing Bufo viridisSahin, M; Gumus, A; Balcan, EMucins are the major glycoprotein secretions of mucous glands and display important functions in amphibian skin such as regulation of water homeostasis and mechanical and chemical protection. In the present study, we evaluated the glycoconjugate contents of developing mucous glands on dorsal regions of metamorphosing Bufo viridis (Amphibia: Anura) tadpoles using an alcian blue-PAS panel and lectin histochemistry. All the conical cells of mucous glands showed weak positivity for alcian blue in 0.025 M MgCl2 at pH 5.7 but only a few cells were positive for 0.3 M MgCl2 at the same pH. In addition, all the conical cells of mucous glands were negative for alcian blue at pH 2.5. In lectin histochemistry, conical cells reacted strongly with Galanthus nivalis agglutinin (GNA), Datura stramonium agglutinin (DSA) and peanut agglutinin (PNA), weakly with Maackia amurensis leucoagglutinin (MAL). These results suggest that they express predominantly mannose, galactose and partially alpha(2-->3)-linked sialic acid containing glycoconjugates. We concluded that dorsal mucous glands of metamorphosing Bufo viridis tadpoles contain at least two different conical cell types and glycoconjugate heterogeneity of mucous glands may be related with different functions of mucins.Item Tunicamycin induced endoplasmic reticulum stress in the small intestineÖztel, Z; Gazan, S; Balcan, EBecause the small intestine is exposed to variety of foreign substances, it participates in host immune response. We investigated whether the expression levels of intestinal MAdCAM-1, PECAM-1 (CD31) and CAV-1 are affected by endoplasmic reticulum (ER) stress following brief treatment with tunicamycin (TN). We administered a single dose of TN intraperitoneally. Twenty-four hours later, MAdCAM-1, PECAM-1 and CAV-1 expression levels in Peyer's patches and villi were examined using immunohistochemistry (IHC), immunofluorescence (IF) and western blotting. Immunostaining of MAdCAM-1 and CAV-1 in control and TN treated Peyer's patches and villi exhibited similar staining patterns. The immunoreactivity of PECAM-1 was similar for the control and TN treated Payer's patches, whereas staining was decreased significantly in TN treated villi. Our findings suggest that short term TN treatment did not affect leukocyte movement to lymphoid compartments of the small intestine, but it altered villus architecture due to decreased PECAM-1 expression.Item The Role of Endoplasmic Reticulum Stress in Gastroesophageal Reflux Disease Symptoms and TreatmentKasap, E; Buran, T; Avcu, AT; Hasdemir, PS; Balcan, E; Aktan, Ç; Korkmaz, MBackground: Gastroesophageal reflux disease is a common condition worldwide. There is no curative treatment for gastroesophageal reflux disease. Endoplasmic reticulum stress leads to the activation of the unfolded protein response and has an important role in inflammation. The aim is to determine the role of endoplasmic reticulum stress in the follow-up of individuals with gastroesophageal reflux disease and the temporal changes of endoplasmic reticulum stress markers with treatment. Methods: Twenty-four subjects in total were recruited prospectively, of whom 15 had nonerosive reflux disease. Two biopsies from 2 cm above the esophagogastric junction, 2 biopsies from gastric antrum mucosa, and 2 biopsies from gastric corpus mucosa were taken. Simultaneously, 2 tubes of venous blood samples were drawn from each individual (1 tube for studying the genetic markers and 1 tube for analyzing the CYP2C19 polymorphism). Results: The mean age was 42.3 +/- 17.6 for women and 34.66 +/- 11.2 for men. Pantoprazole, esomeprazole, rabeprazole, and lansoprazole preparations were used for treatment. There was no significant difference between tissue and blood samples for panel genes ATF-6, XBP-1, DDIT-3, DNAJC-10, and EIF-2-AK before treatment. There was a significant decrease in the level of ATF-6, XBP-1, DNAJC-9, EIF2-AK, and NF-2L-2 genes in blood after treatment. In the comparison of proton pump inhibitors, significant decreases in the expression of the ATF-6, XBP-1, and DNAJC-9 mRNAs were detected in blood from individuals after treatment. Conclusion: Endoplasmic reticulum stress can be for evaluating the clinical improvement and the effectiveness of treatment in gastroesophageal reflux disease.Item Ultrastructural Dynamics of Transendothelial Migration of Lymphocytes Through High Endothelial Venules (HEVs) of the Mucosa Associated Peyer's PatchesBalcan, E; Karaçali, SBackground and Purpose: Although the pivotal role of high endothelial venules (HEVs) in the migration of leukocytes from the blood into the secondary lymphoid parenchyma is well established, conflicting ideas concerning the cellular dynamics both of leukocytes and endothelial cells throughout the migratory processes have been present. Here we focused specifically on the cellular dynamics of HEVs from Peyer's patches in an ultrastructural perspective. Materials and Methods: In order to determine the microstructural organization of transendothelial migration we used conventional methods for transmission electron microscopy. Results: Our results indicate that both lymphocytes and endothelial cells are highly active in the processes of transmigration steps, and a series of morphological and cellular alterations can occur depending upon their activity. Various types of cellular protrusions provide a direct contact between luminal lymphocytes and the endothelial cells at the initial phases of the migration. The endothelial protrusions subsequently embrace the lymphocytes and guide them into lymphoid stroma during the transcellular migration. Meanwhile, different sizes of vesicles show different cellular localizations according to their roles. The vesicles which are clustered near the lateral borders and the stand alone ones found only in the abluminal surfaces of endothelial cells might be involved in the paracellular migration. Concurrently, the other types of vesicles were smaller and appeared in the lateral border of the endothelial cells. Differ from the clustered and abluminally localized vesicles, they were closely related with plasma membranes. Conclusions: These results indicated that not only adhesion molecules, but also cellular dynamics of leukocytes and endothelial cells regulate the leukocyte traffic into lymphoid stroma or vice versa.Item Endoplasmic reticulum stress influences the localization of prion protein in the small intestine and mesenteric lymph nodesBalcan, E; Öztel, Z; Polevshchikov, ATunicamycin is an endoplasmic reticulum (ER) stressor that inhibits protein glycosylation and promotes ER stress. To better understand the localization and traffic of prion protein (PrP) in both basal and ER stress conditions, we evaluated the presence and relative expression of PrP in the intestinal compartments of normal and tunicamycin-treated mice. After tunicamycin treatment, the level of PrP was significantly increased in enterocytes and blind-ended villous lymphatic vessels (lacteals), but was decreased in M cells. These results suggested that intake from the gut and transfer into lymphoid compartments of basal PrP occurs largely through the M cell-Peyer's patch-mesenteric lymph node axis, and also alternatively through the enterocyte-lacteal-mesenteric lymph node axis. In ER stress, the enterocyte-lacteal-mesenteric lymph node is the sole axis for PrP transmission. Results also indicated that germinal centers and high endothelial venules (HEVs) are the most prominent portal for entry of PrP in both basal and ER stressed conditions. We speculated that PrP may use alternative routes for entry into intestinal compartments according to the pathophysiological state and that the mechanism managing the routes of PrP could contribute to the development of new therapeutic strategies against prion diseases as well as ER stress-related intestinal disorders.Item Determination of histopathological effects and myoglobin, periostin gene-protein expression levels in Danio rerio muscle tissue after acaricide yoksorrun-5EC (hexythiazox) applicationKoca, YB; Koca, S; Öztel, Z; Balcan, EAlthough pesticides are essential agrochemicals to annihilate harmful organisms in agriculture, their uncontrolled use has become an important threat to environmental health. Exposure to pesticides can affect many biological systems including immune system, endocrine system, and nervous system. However, the potential side effects of pesticides to skeletal muscle system remain unclear. Present study has focused on the evaluation of this issue by using an acaricide, yoksorrun-5EC (hexythiazox), in an aquatic model organism, Danio rerio. The histological analyses revealed that increased concentrations of the acaricide cause degradation of skeletal muscle along with increased necrosis and atrophy in myocytes, intercellular edema, and increased infiltrations between perimysium sheaths of muscle fibers. The effects of acaricide on myoglobin and periostin, which are associated with oxygen transport and muscle regeneration, respectively, were investigated at the gene and protein levels. RT-PCR results suggested that high concentration yoksorrun-5EC (hexythiazox) can induce myoglobin and periostin genes. Similar results were also obtained in the protein levels of these genes by western blotting analysis. These results suggested that yoksorrun-5EC (hexythiazox)-dependent disruption of skeletal muscle architecture is closely associated with the expression levels of myoglobin and periostin genes in Danio rerio model.Item Do prion protein gene polymorphisms induce apoptosis in non-mammals?Birkan, T; Sahin, M; Öztel, Z; Balcan, EGenetic variations such as single nucleotide polymorphisms (SNPs) in prion protein coding gene, Prnp, greatly affect susceptibility to prion diseases in mammals. Here, the coding region of Prnp was screened for polymorphisms in redeared turtle, Trachemys scripta. Four polymorphisms, L203V, N205I, V225A and M237V, were common in 15 out of 30 turtles; in one sample, three SNPs, L203V, N205I and M237V, and in the remaining 14 samples, only L203V and N205I polymorphisms, were investigated. Besides, C658T, C664T, C670A and C823A SNPs were silent mutations. To elucidate the relationship between the SNPs and apoptosis, TUNEL assays and active caspase-3 immunodetection techniques in brain sections of the polymorphic samples were performed. The results revealed that TUNEL-positive cells and active caspase-3-positive cells in the turtles with four polymorphisms were significantly increased compared with those of the turtles with two polymorphisms (P<0.01 and P<0.05, respectively). In conclusion, this study provides preliminary information about the possible relationship between SNPs within the Prnp locus and apoptosis in a non-mammalian species, Trachemys scripta, in which prion disease has never been reported.Item Prion protein-dependent regulation of p53-MDM2 crosstalk during endoplasmic reticulum stress and doxorubicin treatments might be essential for cell fate in human breast cancer cell line, MCF-7Tugrul, B; Balcan, E; Öztel, Z; Çöllü, F; Gürcü, BIn this study, we investigated the effect of doxorubicin and tunicamycin treatment alone or in combination on MDM-, Cul9-and prion protein (PrP)-mediated subcellular regulation of p53 in the context of apoptosis and autophagy. MTT analysis was performed to determine the cytotoxic effect of the agents. Apoptosis was monitorized by ELISA, flow cytometry and JC-1 assay. Monodansylcadaverine assay was performed for autophagy. Western blotting and immunofluorescence were performed to determine p53, MDM2, CUL9 and PrP levels. Doxorubicin increased p53, MDM2 and CUL9 levels in a dose-dependent manner. Expression of p53 and MDM2 was higher at the 0.25 & mu;M concentration of tunicamycin compared to the control, but it decreased at 0.5 & mu;M and 1 & mu;M concentrations. CUL9 expression was significantly decreased only after treatment of tunicamycin at 0.25 & mu;M. According to its glycosylation status, the upper band of PrP increased only in combination treatment. In combination treatment, p53 expression was higher than control, whereas MDM2 and CUL9 expressions were decreased. Combination treatments may make MCF-7 cells more susceptible to apoptosis rather than autophagy. In conclusion, PrP may be important in determining the fate of cell death through crosstalk between proteins such as p53 and MDM2 under endoplasmic reticulum (ER) stress conditions. Further studies are needed to obtain in-depth information on these potential molecular networks.Item Determination of Glycoconjugate Residues of Erythrocytes at Different Age Groups of RatsGumus, A; Balcan, EAging is a time-dependent process that contains cell injury caused by molecular damages and eventually functional impairment of tissues and organs. The possible roles of cell surface carbohydrates, which are very important molecules in cell to cell and/or cell to extracellular matrix recognition, on aging process are not yet clear. In this study, glycoconjugate alterations of membrane glycoproteins of erythrocytes in aging organism were evaluated with lectin histochemistry and lectin blotting studies in 1, 4 and 7 months old rats. Lectin histochemistry results indicated that alpha(2 -> 3) and alpha(2 -> 6)-linked sialic acids are intensively found in erythrocyte membranes, but this intensity of sialic acids decreases with age. Similar evidences were obtained from lectin blotting studies which performed with same lectins. These results suggest that sialic acid reactivity alters with the age of organism. We thought that (a) sialic acid containing glycoconjugates altered not only with erythrocyte senescence, but also aging process of organism or (b) the number of sialic acid containing erythrocytes decreased by age.Item Comparison of Immune Responses Elicited by Adjuvanted Tachyzoite Lysate Vaccines Developed from Two Different Toxoplasma gondii Strains Isolated in TurkeyPolat, C; Gülçe Iz, S; Döskaya, M; Can, H; Caner, A; Degirmenci, A; Balcan, E; Gürüz, YToxoplasma gondii the causative agent of toxoplasmosis is an obligate intracellular parasite with a wide host range including all warm-blooded animals and birds. T.gondii infection causes congenital toxoplasmosis in newborns and this may lead to fetal anomalies, retinochoroiditis leading to blindness, lethal toxoplasmic encephalitis in immune compromised patients, and organ failure in transplantation patients. The pathogenesis of toxoplasmosis change due to differences in the specific immune response elicited by diverse T.gondii strains. The protective immunity against toxoplasmosis is conferred by cellular immune responses. In the present study, two different strains isolated from Turkey named T.gondii Ankara and Ege were used to evaluate the types of humoral and cellular immune responses elicited by adjuvanted tachyzoite protein vaccines in an animal model. In the study, 6-8 weeks old female BALB/c mice were used and six study groups (each contains three mice) were composed for vaccination. The first and second groups were vaccinated with T.gondii Ankara and Ege (TAnkPE and TEgePE, respectively) tacyhzoite lysates, the third and fourth groups were vaccinated by tacyhzoite lysates adjuvanted with Freund's adjuvant (TAnkPE-Freund; TEgePE-Freund, respectively). The fifth and sixth groups were vaccinated with PBS and Freund's adjuvant as controls. Immunization of the animals was performed two times at three weeks intervals. The serum samples were collected before vaccination and after each vaccination to determine the IgG response by Western blotting, and IgG1 and IgG2a responses by ELISA. To determine the cellular immune response, CD8/CD4 cell ratio, intracellular IFN-gamma and IL-4 levels were determined in stimulated spleen cells grown in cell culture systems by flow cytometry. Toxoplasma IgG antibodies were only detected in TAnkPE-Freund group. IgG1 and IgG2a responses did not increase in any vaccination groups and there was not any polarization towards IgG1 or IgG2a. There was no significant increase in CD8/CD4 ratio of stimulated spleen cells. IFN-gamma level was increased in only TAnkPE-Freund vaccination group, however IL-4 levels were increased in TAnkPE-Freund, TEgePE-Freund and TEgePE groups. Our data showed that TAnkPE-Freund vaccine led to increase in IgG and IFN-gamma responses in BALB/c mice, however, tachyzoite lysate vaccines developed in this study did not induce sufficient protective immune response against toxoplasmosis. Thus, use of specific immunogenic proteins must be taken into consideration in the future vaccine development studies against toxoplasmosis.Item Cold pressed Pinus halepensis Mill. seed oil for potential health applications: Analgesic, anti-inflammatory effects, and assessment of inflammatory mediators by RT-qPCR in skin wound healingAtmane, SA; Batir, MB; Özbek, ZA; Ergönül, PG; Balcan, E; Eldjoudi, DA; Özkale, E; Bribi, N; Khettal, BEthnopharmacological relevance: Pinus halepensis Mill. seed (Pinaceae), commonly known as Zgougou, is widely consumed in the Mediterranean region and has long been used to treat diseases including bronchitis, rheumatism, infection, and inflammation.Aim of the study: The present study was focused on the investigation of some pharmacological activities, such as analgesic, anti-inflammatory, and wound healing effects, of cold-pressed oil of Pinus halepensis Mill. seed (COPHS).Materials and methods: The fixed oil of Pinus halepensis seed was extracted by a cold pressing process. The analgesic activity of COPHS was examined by acetic acid and formalin-induced contortion and pain tests. The anti-inflammatory effects were evaluated in carrageenan and formalin induced paw and ear edema models. Wound healing potential was assessed on an experimental skin wound, and the expression levels of inflammation mediators were determined by RT-qPCR in skin wound healing. Results: The results showed analgesic efficacy through significant inhibition of abdominal cramps (59.15%) and pain (75.91%). In addition, this oil exerted an anti-inflammatory effect by inhibiting ear (62.25%) and paw (70.00%) edema. The COPHS stimulated wound contraction in experimental skin wound healing with a contraction rate of 89.23% with notable reduction of TNF-alpha and NF-kB expression levels in the treated groups. Conclusions: This study provided for the first time the pharmacological profile, particularly the analgesic, antiinflammatory, and healing effects, of fixed oil extracted by cold pressing from the seed of Pinus halepensis Mill.