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Electrochemical biosensor for trypsin activity assay based on cleavage of immobilized tyrosine-containing peptide

Filippova TA, Masamrekh RA, Shumyantseva VV, Latsis IA, Farafonova TE, Ilina IY, et al.

Talanta, 2023 May 15;257:124341.
PMID: 36821964 DOI: 10.1016/j.talanta.2023.124341

In this work, we proposed a biosensor for trypsin proteolytic activity assay using immobilization of model peptides on screen-printed electrodes (SPE) modified with gold nanoparticles (AuNPs) prepared by electrosynthetic method. Sensing of proteolytic activity was based on electrochemical oxidation of tyrosine residues of peptides. We designed peptides containing N-terminal cysteine residue for immobilization on an SPE, modified with gold nanoparticles, trypsin-specific cleavage site and tyrosine residue as a redox label. The peptides were immobilized on SPE by formation of chemical bonds between mercapto groups of the N-terminal cysteine residues and AuNPs. After the incubation with trypsin, time-dependent cleavage of the immobilized peptides was observed by decline in tyrosine electrochemical oxidation signal. The kinetic parameters of trypsin, such as the catalytic constant (kcat), the Michaelis constant (KM) and the catalytic efficiency (kcat/KM), toward the CGGGRYR peptide were determined as 0.33 ± 0.01 min-1, 198 ± 24 nM and 0.0016 min-1 nM-1, respectively. Using the developed biosensor, we demonstrated the possibility of analysis of trypsin specificity toward the peptides with amino acid residues disrupting proteolysis. Further, we designed the peptides with proline or glutamic acid residues after the cleavage site (CGGRPYR and CGGREYR), and trypsin had reduced activity toward both of them according to the existing knowledge of the enzyme specificity. The developed biosensor system allows one to perform a comparative analysis of the protease steady-state kinetic parameters and specificity toward model peptides with different amino acid sequences.
Fulltext Saudi Nursing Students' Knowledge and Perception of Testicular Cancer Assessment and Management: A Cross-Sectional Study

Saleh ZT, Elshatarat RA, Almarwani AM, Alahmadi HA, Elneblawi NH, Al-Za'areer MS, et al.

Asian Pac J Cancer Prev, 2023 Apr 01;24(4):1289-1295.
PMID: 37116151 DOI: 10.31557/APJCP.2023.24.4.1289

INTRODUCTION: Testicular cancer (TC) incidence is increasing worldwide. This study aimed to investigate Saudi nursing students' knowledge and perception about TC.
METHOD: This cross-sectional study was done using convenience sampling method. In this study, 280 nursing students from different nursing schools in six cities of Saudi Arabia were recruited. A structured self-report questionnaire was used to collect data.
RESULT: About 49.2% of the participants received education about TC in their nursing schools. The findings showed lack of enough knowledge about TC among Saudi nursing students. Mostly, the participants reported that heredity factor and having family history of TC (48.9%) and age between 56 and 70 years (41.8%) were the most common risk factors of TC. According to the participants, physical examination was the most common diagnostic test usually used for early detection of TC (40.4%) and biopsy test was the most accurate test to confirm TC diagnosis (45.4%). Only one third of the participants (34.6%) knew that between 75% and 100% of TC cases can be cured in case of early detection. About half of the participants (51.8%) reported that surgical procedure was the most common treatment for TC. The nursing students who had high GPA (r=0.86, p<0.001), were unwilling to get more information on TC (r=0.24, p=0.04), had family history of TC (r= 0.53, p=0.02), medical problems with testicles (r= 0.69, p=0.01), received education about TC in their school of nursing (r=0.65, p=0.02), and were more self-confident in assessing and managing TC (r=0.38, p=0.03) had higher level knowledge about TC. Conclusion: Despite the importance of nurses' roles in assessing and managing TC, nursing students in Saudi Arabia still did not have enough knowledge about TC. Improving nursing programs' curricula and conducting health education programs are recommended.
Fulltext Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

Klionsky DJ, Abdelmohsen K, Abe A, Abedin MJ, Abeliovich H, Acevedo Arozena A, et al.

Autophagy, 2016;12(1):1-222.
PMID: 26799652 DOI: 10.1080/15548627.2015.1100356
Fulltext Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1

Klionsky DJ, Abdel-Aziz AK, Abdelfatah S, Abdellatif M, Abdoli A, Abel S, et al.

Autophagy, 2021 Jan;17(1):1-382.
PMID: 33634751 DOI: 10.1080/15548627.2020.1797280

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.