Functional Analysis of Differentially Expressed Acetylated Spermatozoal Proteins in Infertile Men with Unilateral and Bilateral Varicocele

Sperm proteins undergo post-translational modifications, such as phosphorylation, acetylation, and ubiquitination, which in turn play a key role in determining their fertilizing ability. In the current study, we examined the sperm proteome of men with unilateral and bilateral varicocele to identify the key proteins affected by acetylation to gain an insight into the difference in the severity of affected sperm function in the latter. An LTQ-Orbitrap Elite hybrid mass spectrometer system was used to profile the sperm proteome in pooled unilateral and bilateral varicocele patients. Bioinformatics database and tools, such as UniProtKB, Ingenuity Pathway Analysis Software (IPA) and Metacore, were used to identify the differentially expressed proteins (DEPs) involved in the acetylation process. A total of 135 DEPs in the spermatozoa of unilateral and bilateral varicocele patients were found to be affected by acetylation. The majority of these DEPs found were regulated by key transcription factors such as androgen receptor, p53, and NRF2. Furthermore, the DEPs predicted to be affected by the acetylation process were associated with fertilization, acrosome reaction, mitochondrial dysfunction and oxidative stress. Aberrant expression of proteins and their differential acetylation process may affect the normal physiological functions of spermatozoa. Protein–protein interactions identified dysregulation of the proteasome complex in the bilateral varicocele group. Damage to the proteasome complex may result in aggregation of the misfolded proteins, which in turn increase sperm DNA damage and apoptosis in patients with bilateral varicocele.

The phenolic profile of strawberry tree (Arbutus unedo L.) honey

Despite of the many beneficial health effects of strawberry tree (Arbutus unedo L.) honey, due to its strong antioxidant activity derived mostly from polyphenols, a detailed phenolic profile has not been previously studied. The aims of this study were to identify the phenolic compounds, determine the total phenolic content (TPC) and evaluate the radical scavenging activity (RSA) of strawberry tree honey from south Croatia. Fifty-two polyphenolics (twenty- -seven phenolic acids and twenty-five flavonoids) were identified using ultra- -high-performance liquid chromatograph coupled to a hybrid mass spectrometer (LTQ Orbitrap MS). Our overall results point to the higher TPC (1038 mg gallic acid equivalents per kg of honey) and the stronger RSA (3.32 mmol Trolox equivalents per kg of honey) compared to the other monofloral honeys. Due to the presence of large quantities of polyphenolic compounds, strawberry tree honey may have great potential as a health promoting food.

IDENTIFICATION OF BIOMARKERS OF RENAL PARENCHYMA DAMAGE IN THE URINE OF PATIENTS WITH CHRONIC PYELONEPHRITIS BY ELISA AND MASS SPECTROMETRY METHODS

We performed clinical observations and laboratory examination of 22 patients with chronic pyelonephritis (chronic renal failure, CRF) and 30 healthy individuals. The patients with CRF were examined twice. The first group (Group I) included patients with exacerbation of the disease. The comparison series (Group II) was represented by the same patients who were examined 1.5-3 months after completion of treatment, without clinical exacerbation of chronic pyelonephritis (CPN). Laboratory signs of acute renal damage were not detectable in all the patients examined. Concentrations of VEGF, MCP-1, IL-8 and IL-18 were determined in urine samples of all examined persons by ELISA technique. Protein spectrum of urine was assessed in six patients from Group I, and in six cases of Group II by means of mass spectrometry, using Agilent 1100 chromatographic device, and LTQ-FT Ultra hybrid mass spectrometer. The results of parallel determination of urine proteins by the two methods have shown that the evolving CPN exacerbation is associated with local secondary immune deficiency at the level of renal tubular urothelium. Determination of urine proteome by means of mass spectrometry in exacerbating disease allows identify the proteins associated with damage to epithelial lining of renal tubules and development of local immune response.

APPROACHES TO CHORIONIC GONADOTROPIN QUANTITATIVE DETERMINATION IN ANTI-DOPING CONTROL

The article presents the results of the first stage of development of a new quantitative method for human chorionic gonadotropin (hCG) determination by means of ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) to uncover doping abuse by athletes. The identified tryptic peptides correspond to the most abundant hCG isoforms: the α- and β-subunits, the nicked and β-core fragment of the hormone. Identification and sequencing of specific fragments were performed with the use of nanoLC-MS/MS. A high resolution / high accuracy hybrid mass-spectrometer was applied. Optimization of mass-spectrometric determination of selected specific peptides was accomplished by UPLC-MS/MS. Quantitative evaluation of hCG using specific fragments determination by UPLC-MS/MS allows to detect corresponding hCG isoforms. This significantly increases the method specificity and decreases the probability of false-positive results.