Effects of Taurine on Nitric Oxide and 3-Nitrotyrosine Levels in Spleen During Endotoxemia

Taurine (2-aminoethanesulfonic acid) is a free sulfur-containing β-amino acid which has antioxidant, antiinflammatory and detoxificant properties. In the present study, the role of endotoxemia on peroxynitrite formation via 3-nitrotyrosine (3-NT) detection, and the possible antioxidant effect of taurine in lipopolysaccharide (LPS)-treated guinea pigs were aimed. 40 adult male guinea pigs were divided into four groups; control, endotoxemia, taurine and taurine+endotoxemia. Animals were administered taurine (300 mg/kg), LPS (4 mg/kg) or taurine plus LPS intraperitoneally. After 6 h of incubation, when highest blood levels of taurine and endotoxin were attained, the animals were sacrificed and spleen samples were collected. The amounts of 3-nitrotyrosine and taurine were measured by HPLC, and reactive nitrogen oxide species (NOx) which are stable end products of nitric oxide was measured spectrophotometrically in spleen tissues. LPS administration significantly decreased the concentration of taurine whilst increased levels of 3-NT and NOx compared with control group. It was determined that taurine treatment decreased the levels of 3-nitrotyrosine and NOx in taurine+endotoxemia group. The group in which taurine was administered alone, contradiction to well-known antioxidant effect, taurine caused elevated concentration of 3-NT and NOx. This data suggest that taurine protects spleen against oxidative damage in endotoxemic conditions. However, the effect of taurine is different when it is administered alone. In conclusion, taurine may act as an antioxidant during endotoxemia, and as a prooxidant in healthy subjects at this dose.     

Cytokine gene polymorphisms as potential risk and protective factors in renal cell carcinoma

Serum tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 levels in the serum of 34 patients with head and neck cancer (HNC) undergoing locoregional radiotherapy (RT) were examined. The aim of the RT was definitive in 19 and postoperative adjuvant in 15 patients. Serum TNF-alpha and IL-6 levels were recorded before RT and after the completion of the fifth week of RT. The mean TNF-alpha levels before and after RT were 28.26 +/- 2.87 and 83.03 +/- 7.47, and the mean IL-6 levels were 61.56 +/- 14.32 and 122.45 +/- 30.66, respectively. The statistical analysis yielded a significant rise in TNF-alpha levels with RT in all patients (p < 0.0001) and also in IL-6 levels in patients treated with postoperative adjuvant RT (p = 0.001). Irradiation is likely to cause an acute phase response, and the cytokines studied may be used to monitor this clinically important response in further trials.     

Effect of glutamine and sugars after bull spermatozoa cryopreservation

The objective of this study was to evaluate the effects of the addition of different sugars (raffinose, sucrose, and trehalose) on bull spermatozoa cryopreserved in a commercial extender (Optidyl) supplemented with glutamine on semen parameters, fertilizing ability and superoxide dismutase (SOD) activity. Nine ejaculates for each bull were used in the study. Semen was frozen in five different extenders: raffinose 25 mM plus glutamine 3 mM (RGO), sucrose 25 mM plus glutamine 3 mM (SGO), trehalose 25 mM plus glutamine 3 mM (TGO), glutamine 3 mM (GO) and control (O). Insemination doses were processed so that each 0.25 mL straw contained 15 x 10⁶sperm. Groups of GO and RGO resulted in the higher rates of subjective (54.0 ± 1.7% and 64.0 ± 1.1%; P < 0.01) and CASA motilities (53.0 ± 2.7% and 61.0 ± 4.4%; P < 0.001), respectively compared to the other groups. The supplementation of additives did not provide an effect on the level of post-thaw sperm CASA progressive motilities, the sperm motion characteristics and pregnancy rates. GO and RGO provided the better protective effect for sperm acrosome (4.0 ± 0.5% and 12.0 ± 0.6%) and total abnormalities (5.0 ± 0.3% and 13.0 ± 0.7%; P < 0.001), respectively. At the HOST values, the additives did not give to result the protective effect in comparison to Optydil extender without additives (P > 0.05). For pregnancy rates, there were no significant differences among the groups. The supplementation of additives did not provide any significant difference on the level of SOD activity (P > 0.05). It can be also thought that these sugars might have worked with glutamine in a synergy. Thereby, sugars such as raffinose and sucrose with glutamine in freezing extender may be recommended to facilitate bull semen freezability.     

Using SCC8, SCF27 and VMC7f2 markers in grapevine breeding for seedlessness via marker assisted selection

We used molecular markers associated with seedlessness in grapes, namely SCC8, SCF27 and VMC7f2, to improve the efficiency of seedless grapevine breeding via marker assisted selection (MAS). DNA from 372 F(1) hybrid progeny from the cross between seeded "Alphonse Lavallée" and seedless "Sultani" was amplified by PCR using three markers. After digestion of SCC8 marker amplification products by restriction enzyme BgIII, 40 individuals showed homozygous SCC8+/SCC8+ alleles at the seed development inhibitor (SdI) locus. DNA from 80 of the progeny amplified with the SCF27 marker produced bands; 174 individuals had 198-bp alleles of the VMC7f2 marker associated with seedlessness. In the second year, based on MAS, 183 F(1) hybrids were designated as seedless grapevine candidates because they were positive for a minimum of one marker. Twenty individuals were selected as genetic resources for future studies on seedless grapevine breeding because they carried alleles for the three markers associated with seedlessness. The VMC7f2 SSR marker was identified as the marker most associated with seedlessness.     

The effects of different sugars on motility, morphology and DNA damage during the liquid storage of rat epididymal sperm at 4°C

This study evaluated the protective effects of supplementation with three different sugars on the motility, morphology and DNA integrity of rat epididymal sperm chilled and stored at 4°C Epididymides were obtained from each donor. Rat epididymal sperm was diluted in Ham's F10 plus raffinose, Ham's F10 plus trehalose, Ham's F10 plus fructose, and Ham's F10 medium for control purposes. Thereafter, the extended sperm were chilled and stored in liquid form at 4°C. Sperm motility, morphological abnormalities and DNA damage were determined at 0 and 12h after chilling. No significant difference was observed in any of the parameters evaluated at 0h, before storage (P>0.05). After 12h of storage, all sugar additives led to statistically higher motility, normal sperm morphology and DNA integrity in comparison to the control group. Raffinose gave the best motility percentages (32.86±1.84%) after 12h of storage at 4°C, compared to the other groups (P<0.001). In conclusion, Raffinose, trehalose and fructose provided a better protection of sperm functional parameters against chilling injury, in comparison to the control group.     

<Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis>-mediated genetic transformation of a recalcitrant grain legume,lentil (<Emphasis Type="Italic">Lens culinaris</Emphasis> Medik)

A simple and reproducible Agrobacterium-mediated transformation protocol for a recalcitrant legume plant, lentil (Lens culinaris M.) is reported. Application of wounding treatments and efficiencies of three Agrobacterium tumefaciens strains, EHA105, C58C1, and KYRT1 were compared for T-DNA delivery into lentil cotyledonary node tissues. KYRT1 was found to be on average 2.8-fold more efficient than both EHA105 and C58C1 for producing transient β-glucuronidase (GUS) gene (gus) expression on cotyledonary petioles. Wounding of the explants, use of an optimized transformation protocol with the application of acetosyringone and vacuum infiltration treatments in addition to the application of a gradually intensifying selection regime played significant roles in enhancing transformation frequency. Lentil explants were transformed by inoculation with Agrobacterium tumefaciens strain, KYRT1 harboring a binary vector pTJK136 that carried neomycin phosphotransferase gene (npt-II) and an intron containing gusA gene on its T-DNA region. GUS-positive shoots were micrografted on lentil rootstocks. Transgenic lentil plants were produced with an overall transformation frequency of 2.3%. The presence of the transgene in the lentil genome was confirmed by GUS assay, PCR, RT-PCR and Southern hybridization. The transgenic shoots grafted on rootstocks were successfully transferred to soil and grown to maturity in the greenhouse. GUS activity was detected in vegetative and reproductive organs of T₀, T₁, T₂ and T₃ plants. PCR assays of T₁, T₂ and T₃ progenies confirmed the stable transmission of the transgene to the next generations.     

Application of platelet derived growth factor-BB and diabetic wound healing: the relationship with oxidative events

The reasons that cause delay in wound healing in diabetes are a decrease in the level of growth factors secretion, an increase in the destruction of growth factors and in oxidative stress. Platelet derived growth factor (PDGF) is one of the important growth factors that play a role in all phases of wound healing. This study investigates time-dependent effects of topically PDGF-BB administration on oxidative events on the healing of dorsolateral-excisional wounds in diabetic rats. Forty-two female Wistar-albino rats with streptozotocin-induced diabetes were divided into four groups: control group, untreated group, chitosan-treated group, chitosan?+?PDGF-BB-treated group. Two identical full-thickness excisional skin wounds were made under anaesthesia in all rats except for the control group. In the PDGF-BB-treated and chitosan-treated groups, the wounds were treated topically PDGF-BB (7?ng/mL, single daily dose) and blank chitosan gel (equal amount) after wounding, respectively. After these administrations, on day 3 and day 7 of wound healing, rats were sacrificed. Thiobarbituric acid reactive substances, glutathione, nitric oxide, ascorbic acid levels, and superoxide dismutase activity in wound tissues were spectrophotometrically measured. PDGF-BB administration significantly increased TBARS levels and non-enzymatic antioxidant levels in early phase of diabetic wound healing. PDGF-BB dramatically reduced NO_x levels on day 3 and sharply increased NO_x levels on day 7 of wound healing. Consequently, PDGF-BB administration can be effective on oxidative balance in the early phase of diabetic wound healing.     

Changes in zinc levels and superoxide dismutase activities in the skin of acute, ultraviolet-B-irradiated mice after treatment with ginkgo biloba extract

Acute ultraviolet-B (UV-B) irradiation is known to act as an initiator in the formation of reactive oxygen species. These oxygen products are highly reactive and they are able to cause irreversible damage to cellular components. Oxygen free radicals are normally neutralized by very efficient systems in the body. These include antioxidant enzymes like superoxide dismutase (SOD). In a healthy subject, there is a balance between free radicals and the levels of antioxidants. In some pathological conditions such as oxidative stress, the level of antioxidants is significantly reduced. The skin contains relatively high levels of zinc (Zn), an essential element known to be a cofactor in some metabolic pathways. Zinc has also been reported to have antioxidant properties. In the present study, we investigated the effect of ginkgo biloba extract (Gbe), a potent free-radical scavenger, on UV-B-irradiated skin by measuring SOD activity and Zn levels in the skin, before and after treatment. The SOD activity was decreased after UV-B exposure, in comparison with the control group (p<0.05). After Gbe treatment, the SOD activity increased (p<0.05) as compared with the untreated UV-B irradiated group. The Zn levels changed in the same pattern as the SOD activity values.     

Effects of anti-oxidant additives on microscopic and oxidative parameters of Angora goat semen following the freeze–thawing process

The anti-oxidant system of reduced glutathione (GSH), glutathione peroxidase (GSH-PX), catalase (CAT), and superoxide dismutase (SOD) has been described as a defense functioning mechanism against lipid peroxidation (LPO) in semen, and is important in maintaining sperm motility and viability. This anti-oxidant capacity of sperm cells may be insufficient in preventing LPO during the freeze–thawing process. The aim of this study was thus to determine the influence of varying doses of anti-oxidant additives on standard semen parameters, lipid peroxidation and anti-oxidant activities after the freeze–thawing of goat semen. Ejaculate samples (artificial vagina) obtained from 4 mature Angora goats were evaluated and pooled at 37 °C. The semen samples diluted with a Tris-based extender, containing taurine (25, 50, 75 mM), trehalose (25, 50, 75 mM), and cysteine (5, 10, 15 mM), and an extender containing no anti-oxidant additives (control) were again evaluated. Diluted semen was cooled down to 5 °C and frozen in 0.25 ml French straws, prior to being stored in liquid nitrogen. Frozen straws were thawed in a water bath (37 °C) for 30 s for microscopic sperm evaluation. Upon evaluation of parameters for semen quality, the use of a Tris-based extender supplemented with anti-oxidant additives was found to cause no significant improvement in sperm mortality, when compared to the controls. Increasing doses of taurine and trehalose decreased (P < 0.05) the sperm motility following the freeze–thawing of the goat semen. In biochemical assays, the application of taurine (75 mM) produced the lowest level of malondialdehyde (MDA) (4.46 ± 0.31 nmol/ml), compared to the controls (P < 0.001). Lower GSH levels were higher in the groups in which cysteine was included at 10 and 15 mM (3.27 ± 0.11 and 3.45 ± 0.28 nmol/ml) – compared to the group which received 5 mM cysteine, as well as the controls (2.27 ± 0.08 and 2.50 ± 0.08 nmol/ml respectively, P < 0.001). Compared to the controls, taurine at a concentration of 25 and 75 mM, and increasing doses (50 and 75 mM) of trehalose, significantly increased the GSH-PX activity (P < 0.01). The maintenance of CAT activity was demonstrated to be higher with the addition of 10 and 15 mM cysteine, compared to the other groups (P < 0.001). Vitamin A (VitA) levels were significantly higher, compared to the controls (267.34 ± 9.68 mg/dl and 267.34 ± 9.68 mg/dl, respectively), when 25 mM taurine (329.61 ± 6.35 mg/dl) and 10 mM (318.64 ± 6.34 mg/dl) cysteine was added to the extender (P < 0.001). The results of this study provide a new approach to the cryopreservation of Angora goat semen and could contribute to the improvement of this technology in the goat industry.