Hexokinase cellular trafficking in ischemia–reperfusion and ischemic preconditioning is altered in type I diabetic heart

Diabetes mellitus (DM) has been reported to alter the cardiac response to ischemia–reperfusion (IR). In addition, cardioprotection induced by ischemic preconditioning (IPC) is often impaired in diabetes. We have previously shown that the subcellular localisation of the glycolytic enzyme hexokinase (HK) is causally related to IR injury and IPC protective potential. Especially the binding of HK to mitochondria and prevention of HK solubilisation (HK detachment from mitochondria) during ischemia confers cardioprotection. It is unknown whether diabetes affects HK localisation during IR and IPC as compared to non-diabetes. In this study we hypothesize that DM alters cellular trafficking of hexokinase in response to IR and IPC, possibly explaining the altered response to IR and IPC in diabetic heart. Control (CON) and type I diabetic (DM) rat hearts (65 mg/kg streptozotocin, 4 weeks) were isolated and perfused in Langendorff-mode and subjected to 35 min I and 30 min R with or without IPC (3 times 5 min I). Cytosolic and mitochondrial fractions were obtained at (1) baseline, i.e. after IPC but before I, (2) 35 min I, (3) 5 min R and (4) 30 min R. DM improved rate-pressure product recovery (RPP; 71 ± 10 % baseline (DM) versus 9 ± 1 % baseline (CON) and decreased contracture (end-diastolic pressure: 24 ± 8 mmHg (DM) vs 77 ± 4 mmHg (CON)) after IR as compared to control, and was associated with prevention of HK solubilisation at 35 min I. IPC improved cardiac function in CON but not in DM hearts. IPC in CON prevented HK solubilisation at 35 min I and at 5 min R, with a trend for increased mitochondrial HK. In contrast, the non-effective IPC in DM was associated with solubilisation of HK and decreased mitochondrial HK at early reperfusion and a reciprocal behaviour at late reperfusion. We conclude that type I DM significantly altered cellular HK translocation patterns in the heart in response to IR and IPC, possibly explaining altered response to IR and IPC in diabetes.     

Phylogenetic diversity of isolates belonging to genera Geobacillus and Aeribacillus isolated from different geothermal regions of Turkey

The phylogenetic diversity of 31 thermophilic bacilli belonging to genera Geobacillus and Aeribacillus were investigated which were isolated from various geothermal sites of Turkey. Twenty-seven of these isolates were found to be belonged within the genus Geobacillus, whereas 4 of them were identified as Aeribacillus pallidus. The comparative 16S rRNA gene sequence analyses revealed that the A. pallidus isolates displayed sequence similarity values from 98.0 to 99.6% to their closest relative. Furthermore, Geobacillus isolates showed sequence similarity values from 88.9 to 99.8% with the reference type strains. According to the phylogenetic analysis, isolates belonging to genus Geobacillus were diverged into nine clusters and among these isolates, 19 of them were identified as strains related to G. caldoproteolyticus, G. thermodenitrificans, G. stearothermophilus, G. thermoglucosidasius and G. toebii with the most abundant 13 isolates from G. caldoproteolyticus. Four of the Geobacillus isolates were named as unidentified mix group, as they found to be genetically very homogenous like their closely related type species: G. thermoleovorans, G. vulcani, G. lituanicus, G. kaustophilus, G. caldovelox, G. caldotenax, and G. uralicus. Moreover, the sequence comparisons of E173a, E265, C161ab and A142 isolates demonstrated that they represented novel species among genus Geobacillus as they shared lower than 96.7% sequence similarity to all the described type species. The AluI-, HaeIII- and TaqI-ARDRA results were in congruence with the 16S rRNA gene sequence analyses. By ARDRA results, the isolates were able to be differentiated and clustered, the discriminative restriction fragments of these isolates and type species were determined and the novelty of E173, E265, C161ab and A142 isolates could be displayed. Some differentiating phenotypic characters and the ability of amylase, glucosidase and protease production of these bacilli were also studied and biotechnologically valuable thermostable enzyme producing isolates were introduced in order to use in further studies.     

Acute effects of small-sided games combined with running drills on internal and external loads in young soccer players

The purpose of this study was to compare the effects of regular small-sided games (SSGreg) and SSGs combined with running drills (SSGcom) on players’ internal and external loads. Eighteen young male soccer players (average age: 18.2 ± 0.5 years) participated in 3 vs. 3 and 4 vs. 4 games, under both SSGreg and SSGcom conditions. SSGreg bouts were played for 4 minutes without additional running drills, while SSGcom bouts consisted of 3 min 30 s SSG and 15 s running before and after the bout, making the duration of each bout 4 minutes. During all SSGs, measurements of heart rate (HR) responses as well as distances covered in four different speed zones – walking (WLK), low-intensity (LIR), moderate-intensity (MIR) and high-intensity running (HIR) – were recorded. Technical characteristics were monitored during the SSGs, and the rating of perceived exertion (RPE) and blood lactate (La-) responses were determined at the end of each SSG condition. Compared to the SSGreg in both 3 vs. 3 and 4 vs. 4 formats, the SSGcom condition resulted in higher La- and RPE responses (p < 0.05), greater distance covered at MIR and HIR speeds and greater total distance (p < 0.05). The results of this study show that replacing 30 s within the 4-min bouts of SSGs (both 3- and 4-a-side) by 2 x 15 s of running drills is effective in increasing internal (La- and RPE) and external loads (MIR and HIR) without a significant decrease in total passes and successful passes in young soccer players.     

Fluctuation in secondary metabolite production and antioxidant defense enzymes in in vitro callus cultures of goat’s rue (Galega officinalis) under different abiotic stress treatments

Plant Cell, Tissue and Organ Culture (PCTOC) - Exogenous abiotic stress treatments are an effective way of accelerating stress tolerance by modulation of accumulation of various secondary...     

Bioaccumulation of Heavy Metals in Various Tissues of Some Fish Species and Green Tiger Shrimp (<Emphasis Type="Italic">Penaeus semisulcatus</Emphasis>) from İskenderun Bay,Turkey, and Risk Assessment for Human Health

In this study, concentrations of heavy metals were determined by ICP-MS in the edible tissues of common sole (Solea solea Linnaeus, 1758), whiting (Merlangius merlangus Linnaeus, 1758), silver sillago (Sillago sihama Forsskål, 1775) and muscle-exoskeleton of green tiger shrimp (Penaeus semisulcatus De Haan, 1844), from the seas of ?skenderun Bay, Eastern Mediterranean, Turkey, in January–February, 2016. The lowest and highest mean concentrations of Mn, Cr, Cd, Ni, Hg, As, Pb, and Co in fish and shrimp’ muscles were found, respectively, as follows: 0.166–0.382, 0.134–0.336, 0.005–0.008, 0.091–0.140, 0.026–0.228, 1.741–29.254, 0.087–0.110, and <0.0005–0.027 mg kg^?1; in the skin and exoskeleton, the values were found, respectively, as 0.103–15.819, 0.301–0.778, 0.007–0.026, 0.115–0.513, 0.021–0.243, 1.548–17.930, 0.148–0.295, and <0.0005–0.140 mg kg^?1. According to the results, mean concentrations of all metals in the muscles of fish, except for arsenic and chromium, were found to be below certain legal limit values, especially arsenic levels in shrimp that were found to be above all the legal limit values. Also, the hazard quotients (HQ) of individual heavy metals in fish and shrimp, except for As, revealed safe levels for human consumption. However, the HQ values of estimated inorganic As exceeded 1 in the muscles of shrimp (P. semisulcatus), which may constitute a risk to public health.     

High-yield expression and purification of soluble forms of the anti-apoptotic Bcl-x(L) and Bcl-2 as TolAIII-fusion proteins

A method is presented to produce large amounts of Bcl-2 and Bcl-x_L, two anti-apoptotic proteins of considerable biomedical interest. Expression constructs were prepared in which the Escherichia coli protein TolAIII, known to promote over expression of soluble product, was added to the N-terminus of Bcl-2 or Bcl-x_L proteins, which had their C-terminal hydrophobic anchors deleted. Here the expression of these TolAIII-fusion constructs, followed by a two-step metal-affinity based purification protocol is described. The method delivers at least 20 and 10 mg of more than 90% pure TolAIII-Bcl-x_LΔC and TolAIII-Bcl-2(2)ΔC proteins, respectively, per liter of E. coli cell culture. The proteins are released by proteolysis with thrombin providing >12 mg of Bcl-x_LΔC or >6 mg of Bcl-2(2)ΔC per liter of E. coli cell culture with a purity of more than 95%. Whereas Bcl-x_LΔC is soluble both before and after TolAIII removal, Triton X-100 can significantly increase the extraction of TolAIII- Bcl-2(2)ΔC from the bacterial cells and its subsequent solubility. Far-UV CD spectroscopy demonstrated that they both have an α-helical structure. Fluorescence spectroscopy was used to quantitatively analyze the binding of the respiratory inhibitor antimycin A to recombinant Bcl-2 and Bcl-x_L proteins as well as the displacement of this ligand from the hydrophobic pocket with BH3 Bad-derived peptide. Purified Bcl-x_LΔC and Bcl-2(2)ΔC both protect isolated mitochondria from Bax-induced release of cytochrome c. The ensemble of data shows that the expressed proteins are correctly folded and functional. Therefore, the TolAIII-fusion system provides a convenient tool for functional characterization and structural studies of anti-apoptotic proteins.     

Tubulopapillary mammary carcinoma in a brown bear (Ursus arctos)

A 28-yr-old, nulliparous female brown bear (Ursus arctos) at the Karacabey Ovakurusu Bear Sanctuary presented with an enlargement of the mammary gland. Three other nodules were also noted in the proximity of the mammary gland and over the vulva. Clinical, hematologic, ultrasonographic, and radiologic examinations were performed; the enlarged mammary gland was removed and the other masses were also excised. Histopathologic examination revealed tubulopapillary carcinoma of the mammary gland, and the other masses were diagnosed as epidermoid cysts. This is the first reported case of tubulopapillary mammary carcinoma accompanied by epidermoid cysts in a bear.