Sperm kriyoprezervasyonunda DNA fragmantasyonunun araştırılması:TUNEL yöntemi

Kriyoprezervasyon, bozulmamış hücre ve dokuların yanı sıra çeşitli hayvan türlerinin spermlerinin düşük sıcaklıklarda düşük maliyetle uzun süre muhafaza edildiği bir depolama tekniğidir. Sperm kriyoprezervasyonu; evcil ve yabanıl tür populasyonlarında genetik çeşitliliğin korunması, "genetik olarak üstün" olan yerel türlerin dağıtımının kolaylaştırılması, iatrojenik infertilitenin tedavisi ve insan sağlığı ve hastalıkları için genetiği değiştirilmiş hayvan modellerinin gen bankacılığında kullanılması nedeniyle oldukça önemlidir. Ancak, kriyoprezervasyonda uygulanan dondurma-çözme işlemi spermlere çeşitli zararlar vermesi muhtemel bir işlem olarak kabul edilmektedir. Bu işlemin spermlerde geri dönüşümsüz hasarlara yol açtığı ve spermlerin membran bütünlüğü, morfolojisi ve hareketliliğini etkilediği bilinmektedir. Kriyoprezervasyon esnasında spermde meydana gelen hasarlar arasında en önemlisi DNA fragmantasyonudur. DNA fragmantasyonunun belirlenmesi için en sık kullanılan yöntem TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) yöntemidir. Bu derlemede DNA fragmantasyonu belirlenmesinde TUNEL yönteminin kullanımı ve bu yöntemin kullanıldığı güncel çalışmalar hakkında bilgiler sunulmaktadır

Histochemical demonstration of mucosubstances in the mouse gastrointestinal tract treated with Origanum hypericifolium O. Schwartz and P.H. Davis extract

Glycoconjugate-containing mucous in the gastrointestinal tract has lubricant and cytoprotective effects. This study was performed to demonstrate the intensity of mucous in the stomach, small and large intestine tissues of female BALB/c mice injected intraperitoneally with 0.2 ml of the Origanum hypericifolium extract for six weeks. The staining procedures employed were Alcian Blue (AB) at pH 1 and 2.5, periodic acid-Schiff (PAS) and lectins including Galanthus nivalis agglutinin (GNA), Arachis hypogaea (PNA, Peanut Agglutinin) and Maackia amurensis leucoagglutinin (MAL). The mucous glands in the large intestine of the experimental groups were found to show stronger positive reaction for AB at pH 2.5 and PAS when compared with the controls. In lectin histochemistry, the mucous glands which reacted with lectins exhibited less density for PNA in the stomach and for MAL and PNA in the small intestine in the experimental groups than the controls. In the large intestine of the experimental groups, less staining pattern of GNA was observed on the surface epithelium of villi when compared with the controls. The results suggest that O. hypericifolium extract increases acidic and neutral mucosubstances in the large intestine and also changes some glycan structures in the gastrointestinal tract

The investigation of the effects of Crataegus aronia var. dentata Browicz extract on spleen: a histochemical study

Aim: Spleen is an organ of immune and hematopoietic system and involves extensive cellular interactions. Cell surface and extracellular matrix carbohydrates are important in cellular interactions (cell-cell and cell-matrix). Spleen is affected by the exposure to toxic agents. Crataegus (Hawthorn) species, which contains mainly flavonoid compounds, have such effects as antioxidant, cardioprotective, antiinflammatory and anticarcinogenic. In this study, the effects of Crataegus aronia var. dentata Browicz extracts on spleen were investigated via light microscopy. Materials and methods: The plant extract at concentration of 1% was given orally to rats for 6 weeks. At the end of the experimental period, tissues were fixed, processed and embedded in paraffin. 5 μm sections were stained with Hematoxylin and Eosin, Periodic acid-Schiff stain for histomorphology and Perl’s Prussian blue stain for hemosiderin. For lectin histochemistry, frozen sections were treated with three digoxigenin labelled lectins; peanut agglutinin (PNA) specific for galactose (1→3) N-acetylgalactosamine [Galβ(1→3)GalNAc]; Maackia amurensis leucoagglutinin (MAL) specific for sialic acidα(2→3)Galactose; Galanthus nivalis agglutinin (GNA) specific for terminal mannose residues. Results: In general, the histomorphology of the spleens and the accumulation of hemosiderin in macrophages were similar for both control and experimental groups. Lectins showed positive reactions in some cell cytoplasms and on the cell surfaces of both groups. A little more intensive GNA-positive reactions were observed on the cell surfaces. Conclusions: The plant extract at concentration of 1% did not lead to any alterations in some carbohydrate molecules and hemosiderin in the spleen

Immunohistochemical investigation of galectin-3 in the skin of mice applied with Origanum hypericifolium essential oil and irradiated with ultraviolet B

Aim: To demonstrate galectin-3-immunoreactivity in the undiluted essential oil of Origanum hypericifolium when applied to the ultraviolet B (UVB) irradiated skin of mice. Materials and methods: Female BALB/c mice were allocated to 4 groups, each comprising 6 mice (Group 1: control; Group 2: UVB irradiated control; Group 3: undiluted O. hypericifolium essential oil applied; Group 4: undiluted O. hypericifolium essential oil applied before UVB irradiation). One week prior to UVB irradiation, the undiluted O. hypericifolium essential oil was applied to the shaved dorsal skin of mice 3 times a week. Subsequently, the mice were irradiated 3 times per week with UVB (week 1: 50 mJ/cm2, week 2: 70 mJ/cm2, and weeks 3 and 4: 80 mJ/cm2) for 4 weeks. At the end of this period, immunohistochemical staining for galectin-3 was performed on frozen sections of skin specimens, and then they were photographed. Results: Numerous galectin-3-immunoreactive cells, which were considered to be immune system cells, were observed in the dermis of Group 3. Conclusion: It is suggested that undiluted O. hypericifolium essential oil may cause an increase in the galectin-3-immunoreactive cells. However, there is a need to research these findings with further molecular analyses. © TÜBİTAK

Glycohistochemistry of the lateral tympanic membrane in the syrinx of the denizli cock

The Denizli cock is a special race that has harmonious long crowing. In this study, the syrinx of the Denizli cock was studied by using glycohistochemical analyses. Paraffin sections were stained with Alcian blue (AB) at pH 1 and pH 2.5 for demonstration of sulfated and carboxylated carbohydrates, respectively. At AB-pH 1, the lateral tympanic membrane (LTM) and also the medial tympanic membrane (MTM) exhibited a weak reactivity, whereas they were stained intensely at AB-pH 2.5. The frozen sections were incubated with 5 digoxigenin-labeled lectins for lectin histochemistry: peanut agglutinin, Maackia amurensis leucoagglutinin, Galanthus nivalis agglutinin, Datura stramonium agglutinin, and Sambucus nigra agglutinin. In LTM, positive stainings were observed for the 5 lectins. These results suggest that LTM exhibits an intense reactivity with AB-pH 2.5, indicating carboxylated carbohydrates, and it also has terminal saccharide moieties consisting of β(1→3) linked galactose, α(2→3) linked sialic acid, mannose, β(1→4) linked galactose, and α(2→6) linked sialic acid. © Tübİtak

Histochemical investigation of the effects of Cyclamen graecum extracts on the rat lower gastrointestinal system

Aim: It is known that saponins, triterpenoid or steroidal glycosides, have effects on gastrointestinal system (GIS) and mucus. Cyclamen graecum , a tuberous plant belonging to the Primulaceae family, contains cyclamin as the main saponin. In the present study, it was aimed to investigate the effects of C. graecum tubers ethanolic extracts at the concentrations of 0.1 and 0.3 g/L on the rat lower GIS histomorphology and mucus intensity. Material-Method: In this study, rats were allocated to 3 groups, each comprising 5 rats [Group I: Control group, physiologic saline was administered; Group II: Experimental group, 0.1 g/L(aq) C. graecum extract was administered; Group III: Experimental group, 0.3 g/L(aq) C. graecum extract was administered]. C. graecum tubers ethanolic extracts were given orally to the rats during 10 weeks. At the end of the experimental period, Hematoxylin and Eosin (H&E), Periodic Acid-Schiff (PAS), Alcian Blue (AB) pH 1 and AB pH 2.5 histochemical staining techniques were employed on the tissue samples of small and large intestines in order to demonstrate the histomorphological structures, the neutral, sulphated and acidic mucosubstances, respectively. Results: In both experimental groups, small and large intestine histomorphology, and AB and PAS positive reaction intensities of neutral, acidic, and sulphated mucosubstances were similar to the control group. Discussion: It can be suggested that the C. graecum ethanolic extract has no effect on the rat lower GIS histomorphology and mucus intensity at the administered doses in our study

The effects of Origanum hypericifolium essential oil application and ultraviolet B irradiation on mouse skin: An ultrastructural study

Exposure to UV radiation can cause histopathological and ultrastructural changes in the skin. Origanum hypericifolium, an endemic Turkish plant,essential oil is mainly composed of monoterpenes. The effects of undiluted O. hypericifolium oil on the ultrastructural characteristics of the UVB-irradiated dorsal skin of mice were investigated using transmission electron microscopy. The BALB/c mice were shaved of dorsal hair and randomly housed into 4 groups: 1: control; 2: UVB-irradiated; 3: oil applied; and 4: oil applied and UVB-irradiated. The oil was applied topically to the dorsal skins of the mice on alternate days for 1 week prior to UVB exposure. The skins were irradiated for a total dose of 3.5 J/cm 2 . The sections were stained with hematoxylin and eosin, semithin sections were stained with toluidine blue and ultrathin sections were contrasted with uranyl acetate/lead citrate. There were histopathological changes such as parakeratosis and squamous hyperplasia in the epidermal cell layers (Groups 3 and 4). There were also ultrastructural changes including lacunae formations throughout the stratum corneum layer (Groups 2, 3, and 4), enlargement of intercellular spaces (Groups 2 and 3), reduced desmosomes, narrow and elongated interdigitations, shortened, relatively indistinct and electron dense intermediate keratin filament bundles (Group 3). There were various sizes of cytoplasmic and perinucleolar vacuoles (Groups 3 and 4) and apoptotic bodies phagocytized by keratinocytes (Group 4). I conclude that undiluted oil has side-effects and the potential to inflict injury to the skin. The oil does not ameliorate the negative effects of UVB on epidermal skin cells. © 2016 Elsevier B.V. All rights reserved

The effects of origanum hypericifolium essential oil application and ultraviolet B irradiation on mouse skin: An ultrastructural study

Exposure to UV radiation can cause histopathological and ultrastructural changes in the skin. Origanum hypericifolium, an endemic Turkish plant,essential oil is mainly composed of monoterpenes. The effects of undiluted O. hypericifolium oil on the ultrastructural characteristics of the UVB-irradiated dorsal skin of mice were investigated using transmission electron microscopy. The BALB/c mice were shaved of dorsal hair and randomly housed into 4 groups: 1: control; 2: UVB-irradiated; 3: oil applied; and 4: oil applied and UVB-irradiated. The oil was applied topically to the dorsal skins of the mice on alternate days for 1 week prior to UVB exposure. The skins were irradiated for a total dose of 3.5 J/cm(2). The sections were stained with hematoxylin and eosin, semithin sections were stained with toluidine blue and ultrathin sections were contrasted with uranyl acetate/lead citrate. There were histopathological changes such as parakeratosis and squamous hyperplasia in the epidermal cell layers (Groups 3 and 4). There were also ultrastructural changes including lacunae formations throughout the stratum corneum layer (Groups 2, 3, and 4), enlargement of intercellular spaces (Groups 2 and 3), reduced desmosomes, narrow and elongated interdigitations, shortened, relatively indistinct and electron dense intermediate keratin filament bundles (Group 3). There were various sizes of cytoplasmic and perinucleolar vacuoles (Groups 3 and 4) and apoptotic bodies phagocytized by keratinocytes (Group 4). I conclude that undiluted oil has side-effects and the potential to inflict injury to the skin. The oil does not ameliorate the negative effects of UVB on epidermal skin cells.This work was supported by Pamukkale University Scientific Research Projects Coordination Unit through Grant No: 2014HZL01

A histochemical study of ultraviolet B irradiation and Origanum hypericifolium oil applied to the skin of mice

Ultraviolet (UV) rays cause skin damage. Chronic exposure to UV irradiation causes decreased collagen synthesis, degenerative changes in collagen bundles, accumulation of elastotic material and increased epidermal thickness. Origanum hypericifolium, an endemic Turkish plant, belongs to Lamiaceae family. The main constituents of its oil are monoterpenes including cymene, carvacrol, thymol and γ-terpinene. The effects of undiluted O. hypericifolium oil on UVB irradiated skin of mice were investigated histochemically. Four groups of female BALB/c mice, whose dorsal hair was shaved, were allocated as follows: non-UVB irradiated (Group 1), UVB-irradiated (Group 2), O. hypericifolium oil treated (Group 3), and O. hypericifolium oil treated and UVB irradiated (Group 4). Sections of dorsal skin samples were stained with Mallory's phosphotungstic acid hematoxylin for collagen fibers and Taenzer-Unna orcein for elastic fibers. Sections also were stained with hematoxylin and eosin to measure epidermal thickness. We observed intense staining of collagen and homogeneous, scattered thin elastic fibers in Group 1; scattered and weakly stained collagen and curled, amorphous, accumulate elastic fibers in Group 2; and intense staining of collagen in Groups 3 and 4. Accumulation of elastic fibers in the dermis was unremarkable in Groups 3 and 4. In Groups 3 and 4, O. hypericifolium oil treatment thickened the epidermis. Epidermal thickness was greatest in Group 4. We suggest that O. hypericifolium oil may block UVB induced alterations of collagen and elastic fibers, and increase epidermal thickness. © 2013 The Biological Stain Commission

Investigation of particular matters on the leaves of Pinus nigra Arn. subsp. pallasiana (Lamb.) Holmboe in Denizli (Turkey)

In urban environments, the main sources of emission of particular matters can be listed as follows: Emissions due to heavy traffic and industrial activities; emissions from power stations, local heating sources and road dust. Particular matters in variety of forms such as dust particles, globules, aggregates etc., accumulate on leaf surfaces of road side plants including higher plants. In this respect, investigation of leaf surfaces may indicate environmental pollution and other effects due to industrial and urban activities and thus, receive great deal of attention for environmental evaluations. Pinus nigra Arn. subsp. pallasiana (Lamb.) Holmboe are among the most cultivated plants along the roadsides of the municipal area of Denizli city, TURKEY. In this study, the accumulation and the elemental analysis of particular matters on leaf surfaces of Pinus nigra Arn. subsp. pallasiana (Lamb.) Holmboe were investigated using FESEM-EDS techniques during the years 2009 and 2010. Leaf samples were collected from roadsides of four different local areas of heavy traffic and densely urbanized, and from an area far away from heavy traffic. The chemical composition of particular matters on leaves showed distinct differences in between the two data. This finding indicates high level of increased urbanisation, growing industry and heavy traffic in some locations of Denizli city. As a result, leaves of higher plants may be used as bioindicators for the assessment of particular matters in urban areas