Antimicrobial prescribing confidence and knowledge regarding drug resistance : perception of medical students in Malaysia and the implications

Background: Worldwide, microbes are becoming more challenging by acquiring virulent skills to adapt and develop antimicrobial resistance (AMR). This is a concern as AMR increases morbidity, mortality, and costs. Consequently, physicians need to be trained on appropriate antimicrobial prescribing, starting as medical students. Objective: To evaluate medical students’ confidence in antimicrobial prescribing and AMR. Methods: Cross-sectional study assessing medical students’ knowledge, perception, and confidence in prescribing antimicrobials and AMR in a Malaysian University. A universal sampling method was used. Results: Most responding students believed that educational input regarding overall prescribing was sufficient. Regarding the principle of appropriate and accurate prescriptions, female medical students had less knowledge (odds ratio (OR) = 0.51; 95% confidence interval (CI) 0.25–0.99; p = 0.050). Year-IV and year-V medical students had more excellent knowledge than year-III students regarding confidence in potential antibiotic prescribing once qualified. Year-V students also showed an appreciably higher confidence in the broad principles of prescribing, including antibiotics for infectious diseases, compared to those in other years. Conclusion: Overall, medical students gain more knowledge and confidence regarding the potential prescribing of antimicrobials as their academic careers progress. This is important given concerns with the current excessive use of antimicrobials in Malaysia

Regulation of expression of 5α-reductases in human prostate cells

The aim of this project was to study the regulation of expression of the 5α-reductase (5αR) I and II genes in human prostate cells. Both enzymes are expressed in prostate tissue and 5αRII is essential for normal prostate development. 5αR is a potential target for the treatment of benign prostatic hyperplasia and prostate cancer. To study the regulation of gene transcription, cells constitutively expressing the 5α-reductase enzymes were needed. However this was not possible for 5αRII, since none of the human prostate cell lines available express it. Therefore the first objective of the study was to grow primary cultures from primary and metastatic prostate cancers in bone, in serum-free medium and immortalise the cells with a temperature-sensitive conditionally immortalising T-antigen construct. Conditionally immortalised cell lines would be more likely to express differentiated characteristics such as 5αRII, PSA etc lacking in most available cell lines and therefore could be used to study the regulation of 5αRII gene and would serve as more representative in vitro models for prostatic research. However, the serum-free media (WAJC 404) used to maintain the epithelial cell cultures derived from the primary prostate cancers did not support their long-term growth. Attempts were also made to establish cell lines from prostate cancer metastases in the bone marrow of hormone-relapsed patients. However, most of the bone marrow aspirates did not contain prostate cancer cells and no cultures were established. On the basis of these studies, using a new serum free medium (PrEGM), an epithelial (Pre2.8) and stromal cell (S2.13, which expresses 5αRII) line from the same patient were established subsequently by others in the laboratory. To study the regulation of transcription, the 5' regulatory region of the 5αRI gene was isolated from a human genomic DNA library, cloned and sequenced. A series of deletion constructs ranging in size from 0.16-4.6kb upstream of the ATG start site were made in the promoterless luciferase reporter vector pGL2Basic, and transfected in prostate and non- prostate cells. A 0.6kb fragment upstream of the ATG start site produced the highest promoter activity in cell lines, including PC3 and DU145, derived from human prostate cancers. Primer extension analysis showed that the transcription start site was located at a putative TATA box, 56 bases upstream of the ATG codon. There are many potential transcription factor (TF) binding sites on the promoter, including AP2 and SP1, but transient transfection with expression vectors for these TFs did not influence transcription from the 5αRI promoter construct. The 5' regulatory region of the 5αRII gene was isolated from a human genomic DNA library, cloned and sequenced. A series of deletion constructs ranging in size from 0.15-0.7kb upstream of the ATG site were made in the promoterless luciferase reporter vector pGL2Basic, and transfected in cells as for the 5αRI constructs. A 0.4kb fragment produced the highest promoter activity in a prostate and non-prostate cell line. Attempts at cloning the 8kb sequence upstream of the 0.7kb, in the 0.7kb luciferase construct were unsuccessful. Because of the lack of human 5αRII expressing prostate cell lines, which could be used for testing therapeutic agents targeting this enzyme, a serum free derivative of the prostate cancer cell line DU145 called DUSF was stably transfected with a 5αRII expression vector. An overexpressing clone of DUSF was isolated and found to convert more testosterone to dihydrotestosterone in a crude metabolism assay. This is the first human cell line clone shown to express 5αRII. As a result of this work, novel information has been obtained concerning the regulation of transcription of the 5αR genes. In addition, new human prostate cell line clones have been developed which express 5αRII