Assessment of Pathogenicity of Community-Acquired MRSA Isolates in Mice-Induced Peritonitis

@#Introduction: Methicillin-Resistance Staphylococcus aureus (MRSA) is known as a major nosocomial pathogen in healthcare. However, it has now spread in the community known as community-acquired MRSA (CA-MRSA). Thus, the survival and pathogenicity of CA-MRSA isolates were assessed using in vivo peritonitis model with comparison to ATCC-MRSA. Two CA-MRSA isolates; CA-MRSA1 and CA-MRSA2 that were isolated from healthy population, were studied and compared. Methods: Mice were assigned into 4 groups and injected intraperitoneally with ATCC-MRSA, CA-MRSA1 or CA-MRSA2, respectively. Sterile Dulbecco’s Phosphate-Buffered Saline (DPBS) represents negative control. Mice were observed twice daily, 0-72 hours of post-infection. Any signs of distress were recorded for severity score and survival analyses. Mice were euthanised at 72 hours post-inoculation or by referring to the Peritonitis Severity Scoring (PSS) system. Organs of interest, peritoneal lavage and abscess were processed for bacterial counts. Tissue samples were analysed for histopathological scores. Results: All mice inoculated with MRSA showed clear signs of illness with peritonitis symptoms of p<0.001 and comparable PSS scores were recorded in all infected mice groups. Intraperitoneal injection of lethal dose of MRSA resulted in significant death of ATCC-MRSA (p<0.05) and CA-MRSA-infected mice (p<0.01), compared to the un-infected. Bacterial burden was significantly high in all samples harvested from mice challenged with CA-MRSA2 compared to ATCC-MRSA except in abscess and lung. Significant liver necrosis and spleen inflammation were observed in CA-MRSA1, and lung inflammation in ATCC-MRSA-infected mice. Conclusion: Nasal carriage CA-MRSA isolates from a healthy population has the potential to cause peritonitis with comparable severity as ATCC-MRSA.

Review on the In Vitro Cytotoxicity Assessment in Accordance to the International Organization for Standardization (ISO)

@#Cytotoxicity is a predominant biological evaluation applied to search for a suitable and non-toxic bioactive compound and to determine the biocompatibility of medical devices-related human body. The broad usage of cytotoxicity tests leads to a robust establishment of cytotoxicity assays with high sensitivity and prompt results. In vitro assays are always prioritized over in vivo due to the reproducible data, reduce numbers of animal used and easily accessible material. Compounds concentration that execute 50% of cell population is determined by calculating the IC50. According to ISO10993, cytotoxicity tests must be performed to determine the biocompatibility of medical devices that has contact with human body. This is crucial to ensure the safety of research and its clinical use. Under the recommendation of ISO10995-Part 5, three categories of tests have been documented; extract elution, direct contact and indirect contact test. Each category plays significant role depending on the nature of experiment and sample used.

Cytotoxic Effect of 2,6-bis(4-Hydroxy-3-Methoxybenzylidene) cyclohexanone (BHMC) and Curcumin on Human Liver Cancer Cells, HepG2

@#Introduction: Curcumin is an active constituent derived from turmeric with a variety of pharmacological activities. It suppressed cell proliferation and induced apoptosis in several cancer cell lines. However, due to its poor bioavailability, derivative analogue of curcumin has been synthesized to enhance the drug-like effects. BHMC was synthesized by removing β-diketone moiety from curcumin structure and modify it into conjugated double bonds. It has been proved to exhibit stronger anticancer effects with improved bioavailability compared to curcumin. Objective: This study aims to investigate the toxicity effect of BHMC and curcumin on human liver cancer, HepG2 and non-cancer mouse fibroblast, 3T3. Methods: Both cell lines were purchased from ATCC and cultured in supplemented DMEM. Cell viability was determined via MTT assay and confirmed with trypan blue assay. Morphology hallmarks of apoptosis of both treated cells were analyzed using inverted microscope at 40X magnifications. Results: BHMC and curcumin were very potent towards HepG2 and normal 3T3. These data were further confirmed with trypan blue assay which showed that both compounds significantly reduced the percentage of HepG2 and 3T3 cells viability. Both treated cells also displayed all the morphology hallmarks of apoptosis upon treatment. Conclusion: BHMC has a greater cytotoxicity effect on HepG2 compared to curcumin despite its non-selective cytotoxicity effect on non-cancer 3T3.