CELL DEATH DIFFERENTIATION IN BLACK HEADED RAMS SPERMATOZOA, USING FLUORESCENT LABELED ANNEXIN V

Double staining kit of Annexin V Cy3.18/6-CFDA was used to investigate the changes in phospholipide asymmetry after treating sperm cells with dexamethasone. The % of spermatozoa with registered translocation of PS in treated with dexamethazone groups at the 10-th min and in control no treated varied from 2.74%±0.65 to 2.30%±0.89. After the 5 hour of incubation these % increased to 39.83±3.33 for the treated group and 23.44±1.12 for the control. It was concluded that Annexin V binding assay is more sensitive in the detection of deterioration in membrane function than other conventional methods such as motility analysis and supravital techniques

Surface structuring of β-TCP and transition to α-TCP induced by femtosecond laser processing

Tricalcium phosphate (Ca3(PO4)2, TCP), is one of the most studied and used as material for bioresorbable implants. The β phase has a slower dissolution dynamic and ensures mechanical support for a longer time in biological environment, while a faster release of ions characterize the α phase that trigger a stronger biological response. In this work a femtosecond laser system was used to process β-TCP pellets surface. The femtosecond laser processing results in surface morphology modification, by turning the flat mirror polished surface into a rough and opaque one. The morphological and phisycochemical characteristics of material surface were studied by means of SEM, AFM, Raman, XRD and contact angle measurement. The processed surface showed the formation of micro and nano roughness alongside, furthermore a partial phase transformation from β-TCP to α-TCP was detected. A significant improvement in surface wettability for three different liquids (i.e.water, ethylene glycol and diiodo-methane) is reported. This implies an increase in surface free energy as well. The combination of α and β phase, together with the increased roughness obtained by laser processing, could positively affect the cell adhesion and metabolic activity

Landscape-scale forest loss as a catalyst of population and biodiversity change

The BioTIME database was supported by ERC AdG BioTIME 250189 and ERC PoC BioCHANGE 727440. We thank the ERC projects BioTIME and BioCHANGE for supporting the initial data synthesis work that led to this study, and the Leverhulme Centre for Anthropocene Biodiversity for continued funding of the database. Also supported by a Carnegie-Caledonian PhD Scholarship and NERC doctoral training partnership grant NE/L002558/1 (G.N.D.), a Leverhulme Fellowship and the Leverhulme Centre for Anthropocene Biodiversity (M.D.), Leverhulme Project Grant RPG-2019-402 (A.E.M. and M.D.), and the German Centre of Integrative Biodiversity Research (iDiv) Halle-Jena-Leipzig (funded by the German Research Foundation; FZT 118, S.A.B.).Global biodiversity assessments have highlighted land-use change as a key driver of biodiversity change. However, there is little empirical evidence of how habitat transformations such as forest loss and gain are reshaping biodiversity over time. We quantified how change in forest cover has influenced temporal shifts in populations and ecological assemblages from 6090 globally distributed time series across six taxonomic groups. We found that local-scale increases and decreases in abundance, species richness, and temporal species replacement (turnover) were intensified by as much as 48% after forest loss. Temporal lags in population- and assemblage-level shifts after forest loss extended up to 50 years and increased with species’ generation time. Our findings that forest loss catalyzes population and biodiversity change emphasize the complex biotic consequences of land-use change.PostprintPeer reviewe

Engineering of N. benthamiana L. plants for production of N-acetylgalactosamine-glycosylated proteins - towards development of a plant-based platform for production of protein therapeutics with mucin type O-glycosylation

<p>Abstract</p> <p>Background</p> <p>Mucin type O-glycosylation is one of the most common types of post-translational modifications that impacts stability and biological functions of many mammalian proteins. A large family of UDP-GalNAc polypeptide:N-acetyl-α-galactosaminyltransferases (GalNAc-Ts) catalyzes the first step of mucin type O-glycosylation by transferring GalNAc to serine and/or threonine residues of acceptor polypeptides. Plants do not have the enzyme machinery to perform this process, thus restricting their use as bioreactors for production of recombinant therapeutic proteins.</p> <p>Results</p> <p>The present study demonstrates that an isoform of the human GalNAc-Ts family, GalNAc-T2, retains its localization and functionality upon expression in <it>N. benthamiana </it>L. plants. The recombinant enzyme resides in the Golgi as evidenced by the fluorescence distribution pattern of the GalNAc-T2:GFP fusion and alteration of the fluorescence signature upon treatment with Brefeldin A. A GalNAc-T2-specific acceptor peptide, the 113-136 aa fragment of chorionic gonadotropin β-subunit, is glycosylated <it>in vitro </it>by the plant-produced enzyme at the "native" GalNAc attachment sites, Ser-121 and Ser-127. Ectopic expression of GalNAc-T2 is sufficient to "arm" tobacco cells with the ability to perform GalNAc-glycosylation, as evidenced by the attachment of GalNAc to Thr-119 of the endogenous enzyme endochitinase. However, glycosylation of highly expressed recombinant glycoproteins, like magnICON-expressed <it>E. coli </it>enterotoxin B subunit:<it>H. sapiens </it>mucin 1 tandem repeat-derived peptide fusion protein (LTBMUC1), is limited by the low endogenous UDP-GalNAc substrate pool and the insufficient translocation of UDP-GalNAc to the Golgi lumen. Further genetic engineering of the GalNAc-T2 plants by co-expressing <it>Y. enterocolitica </it>UDP-GlcNAc 4-epimerase gene and <it>C. elegans </it>UDP-GlcNAc/UDP-GalNAc transporter gene overcomes these limitations as indicated by the expression of the model LTBMUC1 protein exclusively as a glycoform.</p> <p>Conclusion</p> <p>Plant bioreactors can be engineered that are capable of producing Tn antigen-containing recombinant therapeutics.</p

ОПРЕДЕЛЯНЕ А ЕКСПРЕСИЯТА НА ФОСФАТИДИЛСЕРИН ПРИ СПЕРМАТОЗОИДИ В РАННА АПОПТОЗА ЧРЕЗ ФЛУОРЕСЦЕНТНО БИЛЯЗАН АНЕКСИН V

Double staining kit of Annexin V Cy3.18/6-CFDA was used to investigate the changes in phospholipide asymmetry after treating sperm cells with dexamethasone. The % of spermatozoa with registered translocation of PS in treated with dexamethazone groups at the 10-th min and in control no treated varied from 2.74%±0.65 to 2.30%±0.89. After the 5 hour of incubation these % increased to 39.83±3.33 for the treated group and 23.44±1.12 for the control. It was concluded that Annexin V binding assay is more sensitive in the detection of deterioration in membrane function than other conventional methods such as motility analysis and supravital techniques.Приложен е кит за двойно оцветяване с Анексин V Cy3.18 и 6-CFDA, даващ възможност за диференциране на апоптични от мъртви клетки. Процентът сперматозоиди с регистрирана експресия на ФС при свежи нетретирани сперматозоиди – контролна проба и експериментални, третирани с дексаметазон на 10-тата минута, варира от 2.30%±0.89 до 2.74%±0.65. След съхранение, на 5-я час този процент нараства до 23.44±1.12 за контролата и до 39.83±3.33 за опитната група. С настоящето изследване се доказва, че Анексин V теста е по-чувствителен за определяне промените в ПМ, в сравнение с конвенционалните методи на изследване за подвижност и преживяемост на сперматозоидите

Ultra-fast laser modification of poly-lactic acid (PLA) -towards enhanced biocompatibility

peer reviewedIn this study, the interaction was investigated of femtosecond laser radiation (pulse duration τ = 130 fs, wavelength λ = 800 nm) with poly-lactic acid (PLA) 2D scaffolds. Two sets of laser fluences (F) and scanning speed (V) values were applied to PLA matrices -F = 1.66 J/cm2, V = 3.8 mm/s, and F = 0.83 J/cm2, V = 3.8 mm/s. The PLA samples thickness, roughness, and water contact angle (WCA) were characterized before and after the laser structuring. The fs-induced micro structures were investigated by SEM, EDX, and FTIR analyses. Preliminary cell fibroblast studies were performed. The results obtained clearly show that a precise laser surface structuring could orient the cells matrix ingrowth and thus make PLA bone tissue engineering interbody future application more successful and adaptable to the personal needs of the recipients

Optimization of titanium and titanium alloy surface properties by ultra-short laser processing for improved antibacterial characteristics

AbstractThe aim of the current study is to improve in a one step process the properties of Ti and Ti alloy surfaces by enhancing their bioactivity in order to provide better conditions for microbial rejection. We propose to alter the biomaterial characteristics by a method alternative to the chemical ones, namely, non-contact processing of the surface by ultra-short laser pulses. The laser-induced modification results in a surface with different topographic features and an increased presence of oxides. We performed hierarchical laser patterning of the surface inducing the formation of areas covered by nanostructures, or laser-induced periodic surface structures (LIPSS), alternating with areas covered by micropillars in their turn surmounted by LIPSS. The increased roughness achieved due to the presence of micropillars, together with a marked presence of oxides, has been proven by several studies to enhance the biocompatibility of the material by improving the surface wettability and, furthermore, promoting the cells adhesion and osseointegration, while reducing the adhesion of pathogens.The laser processed surface, consisting of a pattern of parallel lines, showed improved and anisotropic wettability. The water contact angle value decreased by ∼10° in a direction orthogonal to the lines of the pattern, and by ∼40° in a parallel direction