Studies reveal that alterations in membrane protein (MP) patterns are associated with underlying drug resistance to chemotherapy. Therefore, the tryptic-digested MPs from the bladder cancer cell line were subjected to global proteomics using LC-MS/MS to identify the highly expressed potential MPs in bladder cancer cells. Our findings revealed the identification of MP biomarkers, CD147, and caveolin-1. Immunocytochemistry analysis confirmed the presence of CD147 on the cell membrane, while caveolin-1 showed positive signals without apparent staining on the membrane, suggesting its existence in multiple locations. Western blot analysis confirmed the higher expression of CD147 in non-invasive (RT 112) and metastatic (UM-UC-13) bladder cancer cells compared to invasive bladder cancer cells (5637 and J82), suggesting its potential as an MP biomarker for both of the former subtypes. The identified MPs could be used as drug therapy targets aimed at improving drug sensitivity and enhancing treatment outcomes in bladder cancer patients. SIGNIFICANCE: Identification of the membrane proteins associated with bladder cancer recurrence is crucial to understanding the mechanisms underlying the drug resistance to chemotherapy.
Olive trees are one of the most widely cultivated fruit trees in the world. The chemical compositions and biological activities of olive tree fruit and leaves have been extensively researched for their nutritional and health-promoting properties. In contrast, limited data have been reported on olive flowers. The present study aimed to analyse bioactive compounds in olive flower extracts and the effect of fermentation-assisted extraction on phenolic content and antioxidant activity. High-performance thin-layer chromatography (HPTLC) hyphenated with the bioassay-guided detection and spectroscopic identification of bioactive compounds was used for the analysis. Enzymatic and bacterial in situ bioassays were used to detect COX-1 enzyme inhibition and antibacterial activity. Multiple zones of antibacterial activity and one zone of COX-1 inhibition were detected in both, non-fermented and fermented, extracts. A newly developed HPTLC-based experimental protocol was used to measure the high-maximal inhibitory concentrations (IC50) for the assessment of the relative potency of the extracts in inhibiting COX-1 enzyme and antibacterial activity. Strong antibacterial activities detected in zones 4 and 7 were significantly higher in comparison to ampicillin, as confirmed by low IC50 values (IC50 = 57-58 µg in zone 4 and IC50 = 157-167 µg in zone 7) compared to the ampicillin IC50 value (IC50 = 495 µg). The COX-1 inhibition by the extract (IC50 = 76-98 µg) was also strong compared to that of salicylic acid (IC50 = 557 µg). By comparing the locations of the bands to coeluted standards, compounds from detected bioactive bands were tentatively identified. The eluates from bioactive HPTLC zones were further analysed by FTIR NMR, and LC-MS spectroscopy. Multiple zones of antibacterial activity were associated with the presence of triterpenoid acids, while COX-1 inhibition was related to the presence of long-chain fatty acids.
This study compared the effectiveness of capillary dried blood spots (DBS) versus venous DBS in detecting metabolic changes related to drug-resistant epilepsy (DRE). DBS samples were collected from 142 epilepsy patients (58 drug-resistant, 84 drug-responsive) via venipuncture or fingerstick capillary sampling. Metabolomic analysis using gas chromatography-mass spectrometry compared DBS metabolite profiles between the two groups. While venous DBS profiles showed no distinct patterns, capillary DBS profiles revealed clustering patterns in principal components analysis, with the first two principal components explaining 14.5 %, and 13.5 % of the total variance, respectively. Orthogonal PLS-DA confirmed group discrimination (R2Y=0.989, Q2=0.742). Drug-resistant patients exhibited elevated capillary DBS levels of glutamine, pyruvic acid, and serine, and decreased palmitic acid compared to drug-responsive patients. Pathway analysis revealed disruptions in amino acid metabolism, neurotransmission, and cellular energy regulation. Elevated glutamine levels may contribute to an imbalance between excitatory glutamate and inhibitory GABA neurotransmission, key factors in epileptogenesis and drug resistance. Capillary DBS, likely enriched with arterial blood supply to the brain, appears to better capture central nervous system metabolic disturbances compared to venous DBS containing systemic contributions. This minimally invasive capillary DBS approach offers effective metabolic profiling of brain conditions like DRE, for monitoring disease progression and treatment response, enhancing personalized patient management in epilepsy.