Pusat Pengajian Sains Kesihatan - Tesis

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Detection of humoral immune response in mice immunised with purified protein of milk expressing selected multi-tuberculosis epitopes
(2025-02)
Ravi, Dharshiga
Despite numerous global efforts to reduce infections caused by Mycobacterium tuberculosis and the implementation of immunization programs, tuberculosis (TB) remains one of the leading causes of mortality worldwide. Researchers have sought alternative preventive strategies to decrease the incidence of TB and address the limitations of existing Bacillus Calmette-Guérin (BCG) vaccination methods. A significant drawback of BCG vaccinations is their inability to provide adequate mucosal immunity, highlighting the urgent need for the development of more effective mucosal vaccines that can prevent invasive infections, particularly as MTB primarily targets the respiratory system. Our research team has developed a mucosal vaccine utilizing specific TB epitopes, namely Ag85B, Acr, and RpfE. The vaccine was administered by transfecting a pregnant goat's mammary gland, and the resulting milk was collected daily for 21 days and subsequently purified. The objective of this study was to evaluate the efficacy of the purified milk protein in stimulating humoral immune responses in the serum and mucosal systems of Balb/C mice that received the vaccine candidate intranasally. The effectiveness of the purified protein was assessed by measuring the levels of antibodies generated on antigen-coated plates using the ELISA technique. After incubating the serum, saliva, and bronchoalveolar lavage (BAL) samples from the immunized mice, optical densities (OD) were read using an ELISA reader. The study's results indicated that both the purified protein alone and the combination of BCG with purified protein resulted in elevated levels of IgG antibodies in serum samples, as well as increased levels of IgA antibodies in saliva and BAL samples. In conclusion, the findings demonstrate that the purified protein derived from milk, which contains selected TB epitopes, can induce antibody production, whether administered alone or in conjunction with BCG. This suggests its potential as a viable vaccine candidate against TB
In vitro evaluation of the antioxidant and antimicrobial activities of tualang honey samples and purified 5-hydroxymethylfurfural
(2025-01)
Chandiran, Danesh
Aged tualang honey (ATH) remains a favoured choice due to its superior health benefits, despite many people being unaware that it contains elevated concentrations of 5-hydroxymethylfurfural (HMF). This study aimed to assess the therapeutic properties of TH samples of varying ages (FTH, ATH-24 and ATH-48) containing natural HMF in comparison with purified HMF (HMF low, medium and high doses). The antioxidant property was evaluated through a DPPH radical scavenging assay to calculate IC50. Further, the antimicrobial activity of the TH samples was assessed based on the zone of inhibition by the agar well diffusion method. According to the findings, all TH samples demonstrated higher free radical scavenging activities (ATH-48 exhibited the highest) compared to purified HMF groups. The calculated IC50 values of TH samples were lower than the IC50 values of purified HMF, indicating their superior antioxidant potential. Apart from that, TH samples exhibited antimicrobial activity against all tested bacteria. The greatest antimicrobial activity, indicated by the largest inhibition zone of each TH sample, was observed against S. aureus. ATH-24 has the highest antimicrobial efficacy with better zones of inhibition for both Gram-positive and Gram-negative bacteria compared to FTH and ATH-48. Purified HMF groups demonstrated dose-dependent antioxidant effects without any antimicrobial activities. Taking all data together, ATH-24 with a medium level of HMF demonstrated good antioxidant and antimicrobial properties. As we uncover the advantages of TH samples containing natural HMF, these findings suggest that HMF may synergistically enhance the antioxidant effects in TH samples but not the antimicrobial effects. This study contributes to expanding new insights in apitherapy research
Morphological evaluation of M1 and M2 macrophages derived from thp-1 cell under hypoxic microenvironment
(2025-01)
Siang, Chin Wei
Macrophages play a crucial role in the immune system, with M1 Macrophages promoting proinflammatory responses and M2 Macrophages facilitating anti-inflammatory and tissue repair mechanisms. This study investigates the morphological changes of M1 and M2 macrophages derived from the THP-1 cell under hypoxic microenvironment. Hypoxia, a common feature in various pathological environments, significantly influences morphological behaviour Polarization of THP-1 cells into M1 and M2 phenotypes was achieved using specific stimuli, and their morphological adaptations were analysed through microscopic imaging. Western blot analysis was employed to validate the successful induction of hypoxic microenvironment, confirming the upregulation of hypoxia-inducible factor-1 alpha (HIF-1α), a key marker of hypoxia. The results reveal distinct changes in cell shape, and structure between M1 and M2 macrophages under hypoxic microenvironment, highlighting the impact of oxygen deprivation on macrophage polarization and morphology. Further analysis demonstrated that hypoxic microenvironment enhances the phenotypic shift in macrophage, with minimal morphological changes observed in M1 macrophages. In contrast, M2 macrophages exhibited a significant transition toward an M1-like structure, characterized by decreased cell elongation and cytoplasmic protrusions. These findings contribute to a better understanding of macrophage behaviour in hypoxic microenvironments, offering insights into their roles in disease progression, including cancer, ischemia, and chronic inflammation. This study underscores the importance of exploring macrophage behaviour to identify potential therapeutic targets for hypoxia-associated diseases and their implications in immunotherapy and regenerative medicine
The evaluation of etlingera elatior flower (bunga kantan) aqueous extract (EEAE) on fatty liver disease in hypercholesterolaemic rats
(2025-01)
Ting, Betty Wong Shi
Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease with increasing prevalence worldwide. Hypercholesterolaemia (HC) is the leading cause of NAFLD because excessive high-cholesterol diet (HCD) intake disrupts cholesterol homeostasis. The current medications for NAFLD treatment bring various side effects, and there are no drugs approved by the Food and Drug Administration (FDA). A medicinal plant, Etlingera elatior flower, exhibits antioxidants and hepatoprotective properties. This study aims to evaluate the effects of Etlingera elatior flower (bunga kantan) aqueous extract (EEAE) on the HCD-induced hypercholesterolaemic rat model. Eighteen male Sprague-Dawley (SD) rats were randomly divided into three groups (n = 6): control rat (Group 1), HC-untreated rat (Group 2), and HC-treated 1000 mg/kg EEAE rat (Group 3). The rats were fed HCD for six weeks to induce HC, followed by six weeks of oral EEAE treatment. EEAE significantly reduced body mass index (BMI) and blood cholesterol levels. The histopathological examination of livers for Haematoxylin and Eosin (H&E) staining and Masson’s Trichrome (MT) staining was evaluated. The HC-untreated group developed non-alcoholic steatohepatitis (NASH), characterised by macrovesicular steatosis, inflammation, ballooning degeneration, and fibrosis. In contrast, the liver tissue in the HC-treated EEAE group was reversible from NASH to non-alcoholic fatty liver (NAFL). Furthermore, E. elatior flower was evaluated for its nutritional composition, and HPLC analysis demonstrated that EEAE contains high levels of cyanidin-3-O-glucoside chloride. The effect of EEAE on the liver weight to body weight ratio required further evaluation. Liver function tests (LFT) and lipid profile tests were conducted to evaluate the effect of EEAE in hypercholesterolaemic rats. Overall, EEAE exhibits antioxidant, anti-inflammatory, and antihypercholesterolaemia effects in the liver
Construction of plasmid vector for expression of preliminary microrna-367 in mammalian cell
(2025-01)
Yee, Angelyn Lee Chui
This thesis focuses on the construction of plasmid vector for the expression of preliminary microRNA-367, with a particular emphasis on its connection to miRNA-367-3p and its potential role in regulating key biological processes. Pre-miRNAs serve as essential intermediates in the biogenesis of mature miRNAs, which regulate gene expression by binding to complementary mRNA targets, leading to their degradation or translational inhibition. MiRNA-367-3p, in particular, has been implicated in various cellular functions such as apoptosis, cell proliferation, and differentiation, suggesting its potential therapeutic applications in diseases linked to miRNA dysregulation. The primary aim of this study was to construct a plasmid capable of expressing pre-miRNA 367 in mammalian cells, thereby facilitating the investigation of its functional roles and therapeutic potential. The methodology employed involved designing a synthetic pre-miRNA 367 sequence with necessary restriction sites for cloning into the pCMV-MIR plasmid vector. Following digestion with XhoI and BamHI, the pre-miRNA insert was ligated into the vector and subsequently transformed into E. coli XL1-Blue cells. Despite optimisation efforts, including adjustments to the ligation conditions and insert-to-vector molar ratio, no colonies were obtained, suggesting that potential issues during the ligation or transformation process may have contributed to the lack of successful plasmid incorporation. The discussion provides a detailed analysis of the experimental results, highlighting the steps taken to enhance the ligation process and the challenges encountered. While the cloning was unsuccessful, this study provides valuable insights into the challenges associated with the construction of pre-miRNA expression vectors and offers recommendations for overcoming these challenges to achieve successful expression of pre-miRNA 367 in mammalian cells

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