Potential role of p53 protein as a novelbiomarker of sperm quality, able to predict thesuccess of ART techniques. EcoFoodFertility Project

Introduction: Protein p53 is well known as “The guardian of genome”; it changes its concentration in human spermatozoa DNAin relation to the damage of the latter. It has been suggested thatthe role of the p53 ancestral gene was to ensure the integrity ofthe genomic germline and the fidelity of the evelopment process.The aim of this study is to evaluate if different concentrations of p53 protein in human spermatozoa could influence embryo quality and pregnancy rate and possibly representing a potential predictive marker of sperm quality for successful fertilization .Methods: From July 2013 to June 2017 we have examinatedretrospectively 79 couples with 2-5 years of infertility history.Males had an average age of 27 ± 7,5 years, sperm concentrationof 33,8 ± 6,2 mil/ml, progressive motility of 41,4 ± 8,3 and a typical morphology of 16,5 ± 3,5 according to Kruger’s method. We have divided the couples on the basis of p53 levels: Group A:0,35–1,65 ng/mil (21 males); Group B: 1,66–3,57 ng/mil (32 males);Group C: 3,58–14,53 ng/mil (26 males). We have evaluated thenumber of embryos at stage of 6–8 cells, btained at the third dayof embryo development, in these three different group. In order toevaluate the concentration of p53 protein, we first proceeded toa DNA extraction with forensic method and then to a quantification p53 protein with ELISA-immunoenzymatic assay, expressedin ng/million of spermatozoa.Results: We have observed different percentage of embryo development at stage of 6-8 cells in the third day and different pregnancy rate (PR):Group A: 101 embryos at 6-8 cells/ 147 total number of obtained embryos in this group (68,4%) and PR = 52,38%.Group B: 128/240 (53,5%); PR = 37,50%; Group C: 79/216 (36,1%);PR = 7,69%. These results support the hypothesis that an high con-centration of p53 in human sperm DNA is associated to a low percentage of embryos able to reach the stage of 6-8 cells in the third day of development and also to a lower pregnancy rate. So p53 levels can be considered as a predictive value to embryo development and pregnancy rate.Conclusions: Protein p53 is a sequence-specific transcriptionfactor that responds to a wide variety of stress signals (environ-mental insults and bad lifestyle) as we are investigating within theecofoodfertility project. Particularly quantitative research of p53could be considered as a novel biomarker of sperm quality, able topredict the success of ART echniques, and could open a new roadfor infertility diagnosis

Rubber band ligation (RBL) for haemorrhoids. The experience of three surgical units

Haemorrhoids are the most common proctological disorder with a high incidence per year and a prevalence up to 39% in the general population 1 . This condition often leads to disruption in an individual’s personal and working life. Management has considerable cost implications, and therefore, economic consequences 2 . Due to the fear of SARS-COVID infection the most of patients actually regret hospitalization for surgery and choose to delay the time of treatment. RBL can be proposed as successful procedure to patients with II-III grade with a short stay in the hospital. Treatment consists initially of conservative measures such as lifestyle advice, diet and toilet behaviour. When conservative hemorrhoid therapy is ineffective, many physicians may choose other non-surgical modalities: rubber band ligation, injection sclerotherapy, cryotherapy, manual dilation of the anus infrared photocoagulation, bipolar diathermy 3 , direct current electrocoagulation 4 . Rubber band ligation (RBL) was established as one of the most important, cost-effective and commonly used treatments for first- to third-degree internal hemorrhoids. It is a very effective non-surgical treatment for internal hemorrhoids. causing fibrosis, retraction, and fixation of the hemorrhoidal cushions. Rubber band ligation is also more effective than sclerotherapy and infra-red coagulation, but more painful. Overall complications occur in less than 10%. A retrospective study of 186 patients outpatients who underwent RBL with a minimum follow-up of 12 months is reported. Results confirmed it is effective until 1 year with a low rate of complications and could be offered as conservative treatment for I to III grade hemorrhoids

Measuring the Radius of Gyration and Intrinsic Flexibility of Viral Proteins in Buffer Solution Using Small-Angle X-ray Scattering

Measuring structural features of proteins dispersed in buffer solution, in contrast to crystal form, is indispensable in understanding morphological characteristics of the biomolecule in a native environment. We report on the structure and apparent viscosity of unfolded α and β variants of SARS-CoV-2 spike proteins dispersed in buffer solutions. The radius of gyration of the β variant is found to be larger than that of the α variant, while the ab initio computation of one of the possible particle-like bodies is consistent with the small-angle X-ray scattering (SAXS) profiles resembling a conformation similar to the three-dimensional structure of the folded state of the corresponding α and β spike variant. However, a smaller radius of gyration with respect to the predicted folded state of 2.4 and 2.7 is observed for both α and β variants, respectively. Our work complements the structural characterization of spike proteins using cryo-electron microscopy techniques. The measurement/analysis discussed here might be useful for quick and cost-effective evaluation of several protein structures, let alone mutated viral proteins, which is useful for drug discovery/development applications

Peptide hydrogels from twisted ribbon aggregates

We have studied the rheology of an aqueous solution phase formed in the model peptide system A10K (A=alanine, K=lysine), where the short hydrophobic peptides self-assemble into twisted ribbon structures consisting of laminated beta-sheets. The ribbons are crystalline in 2 dimensions, therefore rigid, and they are weakly charged. The average ribbon lengths, \u3cL \u3e≈ 60 nm, corresponding to an aspect ratio, L/d≈10. With increasing concentration a transition from a viscous liquid into a gel-like solid occurs around a volume fraction f ≈ 0.02, that we identify as the overlap concentration f*. Coinciding with the overlap concentration is also a phase transition, from the low concentration isotropic liquid phase to a nematic phase. This concentration is significantly lower than what is predicted for hard rods by Onsager theory for this given aspect ratio. We attribute this to the ribbon charge and long range electrostatic interactions, stabilizing the nematic phase. In this nematic phase, the storage modulus G’ increases strongly with increasing Please click Additional Files below to see the full abstract

An HLD framework for cationic ammonium surfactants

The Hydrophilic-Lipophilic Difference (HLD) model can be described by additive contributions accounting for the effect of the oil and surfactant nature, temperature, ionic strength, and so on. The first step to build an HLD framework for a surfactant class is to have Winsor III phase equilibria in a restricted range of formulation variables. In this respect, anionic and nonionic surfactants are well suited for an HLD study. On the contrary, it is difficult achieve for pure cationic surfactant Winsor III phase equilibria without the addition of alcohols and this has precluded the extension of the HLD to cationic surfactants. In the present contribution, we first propose a system based on a blend of single-tailed and double-tailed cationic surfactant to study the oil contribution, and then we afforded the determination of the surfactant contribution trough an experimental approach (the “HLD-titration”) that is especially tailored for systems displaying a wide range of existence of Winsor III phase equilibria. HLD-titration results confirmed the ionic strength contribution to HLD as a logarithmic function of salinity for cationic-based microemulsions similarly to anionic ones. However, the oil carbon number contribution is almost four-fold larger (k=0.70.1) with respect to anionic surfactants. A clearing point was observed in correspondence of the Winsor III phase equilibria under stirring. This approach allows us the determination of the so-called characteristic curvature (Cc), i.e. the term describing the surfactant nature contribution to the film curvature, of the cationic surfactant. Finally, the method was adopted to determine Cc values of 7 quaternary ammonium surfactants differing in the polar heads nature and further three amine oxide surfactant at pH=1 where they are protonated

Arrested dynamics in a model peptide hydrogel system

We report here on a peptide hydrogel system, which in contrast to most other such systems, is made up of relatively short fibrillar aggregates, discussing resemblance with colloidal rods

Chemical composition and colloidal properties of dissolved organic matter in Norway spruce forest stands of different ages

Dissolved organic matter (DOM) is the most mobile and actively cycling fraction of soil carbon and acts as a carrier of nutrients and contaminants. It is consumed by microbes, photodegraded, or adsorbed in soils and sediments on its way to the ocean. Despite intensive research in the last two decades, the formation and fate of DOM in soils and its response to changes in land use and climate are poorly understood [1-3]. The changes in temperature and chemical composition of soils affect substantially the rates of microbial decomposition. It has previously been observed that afforestation had a positive effect on carbon stocks approximately 3 decades after land-use change [4]. The aim of this study was to identify the role of afforestation on the chemical composition and colloidal nature of DOM. We compared water extractable DOM from an organic horizon in three differently aged (35-, 61-, 90-years-old) Norway spruce stands growing in the same Tönnersjöheden area located at Simlångsdalen, south-west Sweden . Arable fields that were adjacent to each of these three forests served as control DOM samples and represented the soil material before afforestation. Chemical composition of DOM was inferred from 13C solid-state nuclear magnetic resonance (NMR), high-resolution 1H NMR, infrared spectroscopy (DRIFT) and elemental analysis measurements. Colloidal properties of DOM were investigated using small-angle X-ray and dynamic light scattering methods together with electrophoretic mobility measurements. The dialysis experiment was additionally performed in order to investigate the high molecular fraction of DOM. Elemental analysis revealed an increase in the ratio between total organic C and total N with forest age and no differences between three field DOM extracts. 1H and 13C NMR results showed that both field and forest DOM extracts were dominated by carbohydrates and also contained carboxylic and aliphatic compounds. The aromatic structures were not detected using NMR. However, some features of aromatics and phenolics were detected in IR spectra, especially in forest cold DOM. Scattering data showed that field and forest DOM contained locally stable colloidal aggregates of ca. 100 nm in radius. The structures of these aggregates are consistent with a combination of globular and cluster-like colloids. Field DOM contained slightly higher fraction of clusters than forest DOM. According to the dialysis experiment the half of DOM was presented in high molecular weight fraction (> 12-14 kDa). Overall, our data suggest that DOM extracted from forest and field organic soils had similar chemical and colloidal properties. The relative composition was dictated more by temperature at which DOM was extracted

Presence of p53 Protein on Spermatozoa DNA: A Novel Environmental Bio-Marker and Implications for Male Fertility

Many studies suggest a direct relationship between toxic effects and an increase in the p53 protein on cellular DNA. For our studies, we used sperm DNA as an indicator of environmental toxic effects, dosing p53 quantitatively. To assess possible variations, we used semen samples from two homogeneous male groups living permanently in areas with different environmental impact. The toxic effects of the selected high environmental impact area are caused by both soil and air pollution, while the selected low environmental impact area is a nature reserve where there are no landfills, but only rural factories. As we work with reproductive cells, our interest was inevitably focused on sperm DNA damage and whether this damage could affect their fertilizing capacity. The length of telomeres and the quantification of protamines are being studied to better define the possible damage

Self-assembly of the peptide A10K – Intermediate state in aggregate formation

We have studied the synthetic surfactant-like peptide A10K in solution. Upon mixing the freeze dried, essentially amorphous peptide powder in heavy water, the peptides self-assemble into long ribbon-like aggregates with a fixed cross section of circa 3x8 nm. It is still unclear whether this self-assembly is equilibrium like a surfactant micelle formation or whether it corresponds to a precipitation of a solid phase. Through light scattering measurements on dilution series, the solubility of the ribbons has been determined to 4.7 μM. However, quantitative NMR spectroscopy shows a monomer concentration of 3 mM, corresponding to a roughly 60 times supersaturation, and independent of the total concentration. Samples prepared directly at, or below this specific concentration shows no, or only minor signs of aggregation. In combination with a broad peak in high resolution 1H NMR spectroscopy we conclude that the formation of the A10K aggregates occurs through an intermediate state in equilibrium with the peptide monomers. Please click Additional Files below to see the full abstract