Ultrastructural Dynamics of Transendothelial Migration of Lymphocytes Through High Endothelial Venules (HEVs) of the Mucosa Associated Peyer’s Patches

Background 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. Different 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.  </p

No relation between EFHC2 gene polymorphism and Idiopathic generalized epilepsy.

Background: Idiopathic generalized epilepsy (IGE) is an epilepsy form without an underlying brain lesion or neurological indication or symptom. Recent investigations on the genetic origins of IGE and its subtypes report that certain mutations of various ion and non-ion channels genes in the central nervous system may be associated with IGE.Purpose: In this study we evaluated the relation between IGE and S430Y polymorphism in EFHC2 gene in a Turkish population.Material/methods: The study enrolled 96 healthy volunteers (47 male, 49 female), served as controls, and 96 IGE patients (41 male, 55 female), IGE diagnosis was confirmed in the neurology department. DNA extractions were performed. The presence of S430Y polymorphism in the exon 9 of EFHC2 gene were analyzed by Real-Time PCR. The findings obtained from the control and patient groups were compared.Results: In the patient group there was one heterozygous male with 685 T&gt;C mutation. In the control group, there were two objects with 685 T&gt;C mutation; one heterozygous male, one heterozygous female. 662 G&gt;A mutation was determined in neither controls nor patients.Conclusion: In our series of 96 IGE patients and 96 healthy controls, there was no relation between S430Y polymorphism in EFHC2 gene and IGE presence.Keywords: EFHC2 gene polymorphism, idiopathic generalized epilepsy

Non-Penetrating Wounds of the Chest

The heart may be seriously injured by compression of the chest by a steering wheel in automobile accidents or in other injuries by non-penetrating blows to the chest. These injuries vary from a simple bruise of the heart to actual laceration of heart muscle and heart valves. The changes may be identified by electrocardiograph and changes in heart rhythm. Attorneys should make certain that any injury by a blow to the chest is studied by a cardiologist, using not one but a series of electrocardiographs, as even seemingly harmless blows to the chest may result in injuries equivalent to heart attacks

No relation between EFHC2 gene polymorphism and Idiopathic generalized epilepsy.

Background: Idiopathic generalized epilepsy (IGE) is an epilepsy form without an underlying brain lesion or neurological indication or symptom. Recent investigations on the genetic origins of IGE and its subtypes report that certain mutations of various ion and non-ion channels genes in the central nervous system may be associated with IGE. Purpose: In this study we evaluated the relation between IGE and S430Y polymorphism in EFHC2 gene in a Turkish population. Material/Methods: The study enrolled 96 healthy volunteers (47 male, 49 female), served as controls, and 96 IGE patients (41 male, 55 female), IGE diagnosis was confirmed in the neurology department. DNA extractions were performed. The presence of S430Y polymorphism in the exon 9 of EFHC2 gene were analyzed by Real-Time PCR. The findings obtained from the control and patient groups were compared. Results: In the patient group there was one heterozygous male with 685 T&gt;C mutation. In the control group, there were two objects with 685 T&gt;C mutation; one heterozygous male, one heterozygous female. 662 G&gt;A mutation was determined in neither controls nor patients. Conclusion: In our series of 96 IGE patients and 96 healthy controls, there was no relation between S430Y polymorphism in EFHC2 gene and IGE presence

Fare lenfoid organlardaki özel venüllerin (HEV'lerin) ince yapılarının, glikozaminoglikanlarının ve lenfosit göçünün karşılaştırılması

VI ABSTRACT THE COMPARISON OF ULTRASTRUCTURES, GLYCOSAMTNOGLYCANS AND LYMPHOCYTE MIGRATION OF SPECIALIZED VENULES (HEVs) OF THE MOUSE LYMPHOID TISSUES BALCAN Erdal Ph.D. in Biology Supervisor: Prof. Dr. Sabire KARAÇALI January 1999, 87 pages In this study the ultrastmcture of specialized venules (HEVs) within the peripheral lymph node and Peyer's patches, and HEV-like vessels in the thymus of BALB/c type mice were searched in epon sections, stained with uranyl acetate-lead citrate, by electron microscope. All of the examined three HEVs have structuraly similar cytological properties which were composed of cuboidal endothelial cells and cells with cleared cytoplasm. Endothelial cells of peripheral lymph node HEVs are cuboidal shaped. However, endothelial cells of other HEVs were lesser in height. Within the Peyer's patches HEVs, it has been found that the specialized endoplasmic reticulum sacs, contained septate desmosome-like bridges. In addition, characteristically, cells with cleared cytoplasm contained abundant electron lucent sacs. In these cells, spreading of well developed cytoplasmic extensions resembled theVII antigen presenting cells, having migrating abilities, where found outside of HEV's. Lymphocytes have been found within the lumens of all the three HEV's. In addition to lymphocytes polymorphonuclear leukocytes in lumens of peripheral lymph node HEV's and lymphoblast in HEV-like vessels of thymus were found. HEV's were surrounded with an external laminae, containing supporting cells. Glycosaminoglycans (GAG's) were investigated histochemically using Alcian Blue at different MgCİ2 concentrations at pH 5.8 and cytochemically with Ruthenium Red. With the Alcian Blue, luminal surfaces and external laminaes of HEV's were densely stained The results of the staining showed that the sulfated and carboxylated GAG's were mostly found in the peripheral lymph nodes. Peyer's patches and thymus HEV's were stained in lesser extent with Alcian Blue. Generally, in the all of the samples, the carboxylated GAG's were denser than sulphated ones. Ruthenium Red, cytochemically, reacted positively on the surfaces of the mouse peripheral lymph node HEV wall cells and on the luminal lymphocytes. In all three venules, lymphocyte passing from HEV's occurs by same way. Endothelial cells and cells with cleared cytoplasm in the HEV's walls accompanied the migration by surrounding the lymphocytes. During the passage through the HEV's, cells changed their shapes. Finally, lymphocytes also pass through external laminae which is rich in GAG content and emerge outside of HEV. Keywords: Lymphoid tissues, high endothelial venule, glycosaminoglycan, lymphocyte migrationIV ÖZET FARE LENFOID ORGANLARDAKİ ÖZEL VENÜLLERİN (HEV'lerin) İNCE YAPILARININ, GLİKOZAMİNOGLİKANLARININ VE LENFOSİT GÖÇÜNÜN KARŞILAŞTIRILMASI BALCAN, Erdal Doktora Tezi, Biyoloji Bölümü Tez Yöneticisi: Prof. Dr. Sabire KARAÇALI Ocak 1999, 87 sayfa Bu çalışmada, BALB/c tip farelerde, periferal lenf düğümü ve Peyer plaklarmdaki özel venüller (HEV'ler) ile timustaki HEV benzeri venüllerin ince yapılan, uranil asetat-kurşun sitrat ile boyanmış epon kesitlerde elektron mikroskobu ile çalışılmıştır. İncelenen her üç lenfoid dokudaki HEV'ler, yapısal olarak benzer sitolojik özellikler gösterir ve kübik endotel hücreler ile açık renk sitoplazmalı hücrelerden oluşur. Periferal lenf düğümü HEV'lerindeki endotel hücreleri kübik şekillidir. Diğer HEV'lerde bulunan endotel hücrelerinin yüksekliği daha azdır. Peyer plakları HEV'lerin endotel hücrelerinde, içinde septat desmozom benzeri köprülerin şekillendiği özel tip gedoplazmik retikulum keseleri bulunur. Açık renk sitoplazmalı hücreler, karakteristik şekilde bol olarak elektronca az yoğun kesecikler içerirler. Gelişmiş sitoplazmik uzantılarının yayılışı, HEV'lerin dışmda bulunan ve göçetme yeteneğine sahip olan antijen sunan hücrelere benzer. Her üç HEV'in lümeninde de lenfositler bulunur. Lenfositlere ek olarak, periferal lenf düğümü HEV'lerin lümenlerinde polimorfonüklear lökositler, timus HEV benzeri venüllerde ise lenfoblastlar yer alır. HEV ler, içlerinde destek hücrelerinin bulunduğu bir dış tabaka ile kuşatılır. Glikozaminoglikanlar (GAG' lar), pH 5.8 de çeşitli MgCİ2 konsantrasyonlanndaki aldan blue ile histokimyasal olarak, rutenyum red ile sitokimyasal olarak gösterilmiştir. Aldan blue ile HEV'lerin lümene bakan kısımları ve dış tabakaları koyu boyanır. Boyanma sonuçları, sulfatlanmış ve karboksillenmiş GAG'larm en fazla periferal lenf düğümünde olduğunu işaret eder. Peyer plakları ve timus HEV'leri daha az yoğun boyanırlar. Genel olarak hepsinde, karboksillenmiş GAG' lar sulfatlanmış olanlardan daha yoğun boyanır. Sitokimyasal olarak rutenyum red, fare periferal lenf düğümü HEV duvarı hücrelerinin yüzeyinde ve lümen içindeki lenfositlerin yüzeylerinde pozitif reaksiyon verir. HEV'lerden lenfosit geçişi, her üç tip venülde de aynı yolla olmaktadır. HEV duvarındaki endotel hücreleri ve açık renk sitoplazmalı hücreler, lenfositleri kuşatarak göçe yardımcı olurlar. Geçiş sırasında hücreler şekil değiştirirler. Lenfositler, GAG içeriği bakımından zengin olan dış tabakadan da geçerler ve HEV dışına çıkarla

Türkiye’de izole edilen iki farklı toxoplasma gondii suşundan üretilen adjuvanlı takizoit eriyik protein aşılarının uyardığı immün yanıtların karşılaştırılması

Toxoplasma 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-;amp;#947; 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-;amp;#947; level was increased in only TAnkPE-Freund vaccination group, however IL-4 levels were increased in TAnkPE-Freund, TEgePE-Freund and TEge- PE groups. Our data showed that TAnkPE-Freund vaccine led to increase in IgG and IFN-;amp;#947; 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.Toxoplasma gondii, çok geniş konak aralığında, sıcak kanlı hayvanlarda ve kuşlarda enfeksiyon oluşturabilen zorunlu hücre içi bir parazittir. İnsanlarda T.gondii enfeksiyonu, yeni doğanlarda fetal anomalilere yol açan konjenital toksoplazmoz, körlüğe sebep olan retinokoroidit, immün sistem yetmezliği olan kişilerde fatal seyreden toksoplazmik ensefalit ve organ nakli yapılanlarda organ reddi ve ölüme sebep olmaktadır. Farklı T.gondii suşlarının hayvan modellerinde uyardıkları immün yanıta bağlı olarak, oluşturdukları patogenez de değişiklik göstermektedir. Toksoplazmoza karşı korunmada hücresel immün yanıtın daha önemli olduğu belirtilmektedir. Bu çalışmada, ülkemizde izole edilen T.gondii Ankara ve Ege suşlarından üretilen adjuvanlı takizoit protein aşılarının hayvan modellerinde uyardığı hümoral ve hücresel immün yanıt tiplerinin araştırılması amaçlanmıştır. Çalışmada 6-8 haftalık dişi BALB/c fareler kullanılmış ve aşılama için her biri üç adet fare içeren altı grup oluşturulmuştur. Birinci ve ikinci grup T.gondii Ankara ve Ege (TAnkPE; TEgePE) takizoit eriyiği ile, üçüncü ve dördüncü grup Freund adjuvanı ile birleştirilmiş TAnkPE (TAnkPE-Freund) ve TEgePE (TEgePE-Freund) takizoit eriyiği ile, kontrol grupları olan beşinci ve altıncı gruplar ise PBS ve Freund adjuvanı ile aşılanmıştır. Hayvanların immünizasyonu üç hafta aralıklarla iki kez yapılmıştır. Aşılama öncesi ve her aşılama sonrası alınan serum örneklerinde Western blot yöntemiyle IgG yanıtı, ELISA testi ile IgG1 ve IgG2a yanıtları araştırılmıştır. Hücresel immün yanıtları belirlemek için, hücre kültürü ortamında uyarılan dalak hücrelerinin CD8/CD4 oranı, hücre içi IFN-? ve IL-4 sitokinleri akış sitometrisi ile ölçülmüştür. Çalışmamızda, Toxoplasma IgG antikorları sadece TAnkPE-Freund aşısı uygulanan grupta saptanmış; IgG1 ve IgG2a antikor yanıtlarının hiçbir aşılama grubunda artmadığı tespit edilmiş, IgG1 veya IgG2a yönünde belirgin bir polarizasyon olmadığı belirlenmiştir. Aşılanan grupların hiçbirisinde uyarılmış dalak hücrelerinde CD8/CD4 oranında değişiklik saptanmamıştır. IFN-? üretimi sadece TAnkPE-Freund ile aşılanan grupta artarken; IL-4 üretimi TAnkPE-Freund, TEgePE-Freund ve TEgePE ile aşılanan gruplarda artmıştır. Çalışmamızın verileri, TAnkPE-Freund aşısının BALB/c farelerinde IgG ve IFN-? yanıtlarını artırdığını göstermiş, ancak toksoplazmoza karşı geliştirdiğimiz bu eriyik protein aşılarının koruyucu immün yanıtı yeterince uyarmadıkları saptanmıştır. Bu nedenle ileride yapılacak olan aşı çalışmalarında daha özgül proteinlerin kullanılması yönünde kanaat oluşmuştur