Displaying publications 1 - 20 of 49 in total

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  1. Inhibition of lysosomal vacuolar proton pump down-regulates cellular acidification and enhances E. coli bactofection efficiency
    Akinsola RO, Lee CW, Sim EUH, Narayanan K
    Anal Biochem, 2021 03 01;616:114088.
    PMID: 33358938 DOI: 10.1016/j.ab.2020.114088
    Endosomal escape is considered a crucial barrier that needs to be overcome by integrin-mediated E. coli for gene delivery into mammalian cells. Bafilomycin, a potent inhibitor of the H+ proton pump commonly employed to lower endosomal pH, was evaluated as part of the E. coli protocol during delivery. We found an increase in green fluorescent protein expression up 6.9, 3.2, 5.0, 2.8, and 4.5 fold in HeLa, HEK-293, A549, HT1080, and MCF-7 respectively, compared to untreated cells. Our result showed for the first time that Inhibition of lysosomal V-ATPase enhances E. coli efficiency.
  2. RFP-based method for real-time tracking of invasive bacteria in a heterogeneous population of cells
    Akinsola RO, Adewoyin M, Lee CW, Sim EU, Narayanan K
    Anal Biochem, 2021 12 01;634:114432.
    PMID: 34695391 DOI: 10.1016/j.ab.2021.114432
    Quantification of bacterial invasion into eukaryotic cells is a prerequisite to unfold the molecular mechanisms of this vector's function to obtain insights for improving its efficiency. Invasion is traditionally quantified by antibiotic protection assays that require dilution plating and counting of colony-forming units rescued from infected cells. However, to differentiate between attached and internalized bacteria vector, this assay requires supplementation by a time-consuming and tedious immunofluorescence staining, making it laborious and reduces its reliability and reproducibility. Here we describe a new red fluorescent protein (RFP)-based high-throughput and inexpensive method for tracking bacterial adherence and internalization through flow cytometry to provide a convenient and real-time quantification of bacterial invasiveness in a heterogeneous population of cells. We invaded MCF-7, A549, and HEK-293 cells with the E. coli vector and measured RFP using imaging flow cytometry. We found high cellular infection of up to 70.47% in MCF-7 compared to 27.4% and 26.2% in A549 and HEK-293 cells, respectively. The quantitative evaluation of internalized E. coli is rapid and cell-dependent, and it distinctively differentiates between attached and cytosolic bacteria while showing the degree of cellular invasiveness. This imaging flow cytometry approach can be applied broadly to study host-bacteria interaction.
  3. Hericium erinaceus Promotes Anti-Inflammatory Effects and Regulation of Metabolites in an Animal Model of Cerebellar Ataxia
    Chau SC, Chong PS, Jin H, Tsui KC, Khairuddin S, Tse ACK, et al.
    Int J Mol Sci, 2023 Mar 23;24(7).
    PMID: 37047062 DOI: 10.3390/ijms24076089
    Cerebellar ataxia is a neurodegenerative disorder with no definitive treatment. Although previous study demonstrated the neuroprotective effects of Hericium erinaceus (H.E.), the mechanisms of H.E. treatment on the neuroinflammatory response, neurotransmission, and related metabolites remain largely unknown. We demonstrated that 3-AP rats treated with 25 mg/kg H.E. extracts had improved motor coordination and balance in the accelerated rotarod and rod tests. We showed that the H.E. treatment upregulated the expression of Tgfb1, Tgfb2, and Smad3 genes to levels comparable to those in the non-3-AP control group. Interestingly, we also observed a significant correlation between Tgfb2 gene expression and rod test performance in the 3-AP saline group, but not in the non-3-AP control or H.E.+3-AP groups, indicating a relationship between Tgfb2 gene expression and motor balance in the 3-AP rat model. Additionally, we also found that the H.E. treatment increased mitochondrial COX-IV protein expression and normalized dopamine-serotonin neurotransmission and metabolite levels in the cerebellum of the H.E.+3-AP group compared to the 3-AP saline group. In conclusion, our findings suggest that the H.E. treatment improved motor function in the 3-AP rat model, which was potentially mediated through neuroprotective mechanisms involving TGFB2-Smad3 signaling via normalization of neurotransmission and metabolic pathways.
  4. Induction of protein expression within Escherichia coli vector for entry into mammalian cells
    Chen Q, Lee CW, Sim EU, Narayanan K
    Hum Gene Ther Methods, 2014 Feb;25(1):40-7.
    PMID: 24134118 DOI: 10.1089/hgtb.2012.188
    Direct protein delivery into the cytosol of mammalian cells by invasive Escherichia coli (E. coli) bacterial vector will bypass the need to achieve nuclear entry and transcription of DNA, a major hurdle that is known to seriously limit gene transfer. The bacterial vector is induced to express the protein during its growth phase, before presentation for entry into mammalian cells and release of its content into the cellular environment. For this class of vector, crossing the plasma membrane becomes the primary step that determines the success of protein delivery. Yet, how the mechanics of protein expression within the vector affect its entry into the host is poorly understood. We found the vector's effectiveness to enter HeLa cells diminished together with its viability when phage N15 protelomerase (TelN) expression was induced continuously in the invasive E. coli despite producing an abundant amount of functional protein. By comparison, shorter induction, even as little as 3 hr, produced sufficient amounts of functional TelN and showed more effective invasion of HeLa cells, comparable to that of uninduced invasive E. coli. These results demonstrate that brief induction of protein expression during vector growth is essential for optimal entry into mammalian cells, an important step for achieving bacteria-mediated protein delivery.
  5. Fulltext Monitoring Consumption of Common Illicit Drugs in Kuala Lumpur, Malaysia, by Wastewater-Cased Epidemiology
    Du P, Liu X, Zhong G, Zhou Z, Thomes MW, Lee CW, et al.
    Int J Environ Res Public Health, 2020 01 31;17(3).
    PMID: 32023897 DOI: 10.3390/ijerph17030889
    Southeast Asian countries including Malaysia play a major role in global drug trade and abuse. Use of amphetamine-type stimulants has increased in the past decade in Malaysia. This study aimed to apply wastewater-based epidemiology for the first time in Kuala Lumpur, Malaysia, to estimate the consumption of common illicit drugs in urban population. Influent wastewater samples were collected from two wastewater treatment plants in Kuala Lumpur in the summer of 2017. Concentrations of twenty-four drug biomarkers were analyzed for estimating drug consumption. Fourteen drug residues were detected with concentrations of up to 1640 ng/L. Among the monitored illicit drugs, 3,4-methylenedioxy-methamphetamine (MDMA) or ecstasy had the highest estimated per capita consumptions. Consumption and dose of amphetamine-type stimulants (methamphetamine and MDMA) were both an order of magnitude higher than those of opioids (heroin and codeine, methadone and tramadol). Amphetamine-type stimulants were the most prevalent drugs, replacing opioids in the drug market. The prevalence trend measured by wastewater-based epidemiology data reflected the shift to amphetamine-type stimulants as reported by the Association of Southeast Asian Nations Narcotics Cooperation Center. Most of the undetected drug residues were new psychoactive substances (NPSs), suggesting a low prevalence of NPSs in the drug market.
  6. Synechococcus production and grazing loss rates in nearshore tropical waters
    Heng PL, Lim JH, Lee CW
    Environ Monit Assess, 2017 Mar;189(3):117.
    PMID: 28220442 DOI: 10.1007/s10661-017-5838-1
    Temporal variation of Synechococcus, its production (μ) and grazing loss (g) rates were studied for 2 years at nearshore stations, i.e. Port Dickson and Port Klang along the Straits of Malacca. Synechococcus abundance at Port Dickson (0.3-2.3 × 10(5) cell ml(-1)) was always higher than at Port Klang (0.3-7.1 × 10(4) cell ml(-1)) (p  0.25), but nutrient and light availability were important factors for their distribution. The relationship was modelled as log Synechococcus = 0.37Secchi - 0.01DIN + 4.52 where light availability (as Secchi disc depth) was a more important determinant. From a two-factorial experiment, nutrients were not significant for Synechococcus growth as in situ nutrient concentrations exceeded the threshold for saturated growth. However, light availability was important and elevated Synechococcus growth rates especially at Port Dickson (F = 5.94, p  0.30). In nearshore tropical waters, an estimated 69 % of Synechococcus production could be grazed.
  7. Fulltext Sphingosine 1-phosphate receptor 2 (S1P2) attenuates reactive oxygen species formation and inhibits cell death: implications for otoprotective therapy
    Herr DR, Reolo MJ, Peh YX, Wang W, Lee CW, Rivera R, et al.
    Sci Rep, 2016 Apr 15;6:24541.
    PMID: 27080739 DOI: 10.1038/srep24541
    Ototoxic drugs, such as platinum-based chemotherapeutics, often lead to permanent hearing loss through apoptosis of neuroepithelial hair cells and afferent neurons of the cochlea. There is no approved therapy for preventing or reversing this process. Our previous studies identified a G protein-coupled receptor (GPCR), S1P2, as a potential mediator of otoprotection. We therefore sought to identify a pharmacological approach to prevent cochlear degeneration via activation of S1P2. The cochleae of S1pr2(-/-) knockout mice were evaluated for accumulation of reactive oxygen species (ROS) with a nitro blue tetrazolium (NBT) assay. This showed that loss of S1P2 results in accumulation of ROS that precedes progressive cochlear degeneration as previously reported. These findings were supported by in vitro cell-based assays to evaluate cell viability, induction of apoptosis, and accumulation of ROS following activation of S1P2 in the presence of cisplatin. We show for the first time, that activation of S1P2 with a selective receptor agonist increases cell viability and reduces cisplatin-mediated cell death by reducing ROS. Cumulatively, these results suggest that S1P2 may serve as a therapeutic target for attenuating cisplatin-mediated ototoxicity.
  8. Storage and dynamics of soil organic carbon in allochthonous-dominated and nitrogen-limited natural and planted mangrove forests in southern Thailand
    Hu J, Pradit S, Loh PS, Chen Z, Guo C, Le TPQ, et al.
    Mar Pollut Bull, 2024 Jan 29;200:116064.
    PMID: 38290368 DOI: 10.1016/j.marpolbul.2024.116064
    Mangrove forests can help to mitigate climate change by storing a significant amount of carbon (C) in soils. Planted mangrove forests have been established to combat anthropogenic threats posed by climate change. However, the efficiency of planted forests in terms of soil organic carbon (SOC) storage and dynamics relative to that of natural forests is unclear. We assessed SOC and nutrient storage, SOC sources and drivers in a natural and a planted forest in southern Thailand. Although the planted forest stored more C and nutrients than the natural forest, the early-stage planted forest was not a strong sink relative to mudflat. Both forests were predominated by allochthonous organic C and nitrogen limited, with total nitrogen being a major driver of SOC in both cases. SOC showed a significant decline along land-to-sea and depth gradients as a result of soil texture, nutrient availability, and pH in the natural forest.
  9. Fulltext Sarcopenia is a useful risk stratification tool to prognosticate splenic abscess patients in the emergency department
    Hung SK, Kou HW, Hsu KH, Wu CT, Lee CW, Leonard Goh ZN, et al.
    J Formos Med Assoc, 2021 Mar;120(3):997-1004.
    PMID: 32917483 DOI: 10.1016/j.jfma.2020.08.039
    BACKGROUND/PURPOSE: Splenic abscess is a life-threatening surgical emergency which requires early diagnosis and intervention to maximize patient outcomes. This can be achieved through accurate risk stratification in the emergency department (ED). Sarcopenia refers to an age-related loss of skeletal muscle mass and strength that is accompanied by major physiologic and clinical ramifications, and often signifies decreased physiologic reserves. It is associated with poor clinical outcomes in sepsis, acute respiratory failure, oncological surgery, and liver transplantation. This study evaluates the utility of sarcopenia as a radiological stratification tool to predict in-hospital mortality of splenic abscess patients in the ED. This will assist emergency physicians, internists and surgeons in rapid risk stratification, assessing treatment options, and communicating with family members.

    METHODS: 99 adult patients at four training and research hospitals who had undergone an abdominal contrast computed tomography scan in the ED with the final diagnosis of splenic abscess from January 2004 to November 2017 were recruited. Evaluation for sarcopenia was performed via calculating the psoas cross-sectional area at the level of the third lumbar vertebra and normalising for height, before checking it against pre-defined values. Univariate analyses were used to evaluate the differences between survivors and non-survivors. Sensitivity, specificity, and predictive values of the presence of sarcopenia in predicting in-hospital mortality were calculated. Kaplan-Meier methods, log-rank test, and Cox proportional hazards model were also performed to examine survival between groups with sarcopenia versus non-sarcopenia.

    RESULTS: Splenic abscess patients with sarcopenia were 7.56 times more at risk of in-hospital mortality than those without sarcopenia (multivariate-adjusted HR: 7.56; 95% CI: 1.55-36.93). Presence of sarcopenia was found to have 84.62% sensitivity and 96.49% negative predictive value in predicting mortality.

    CONCLUSION: Sarcopenia is associated with poor prognoses of in-hospital mortality in patients with splenic abscess presenting to the ED. We recommend its use in the ED to rapidly risk stratify and predict outcome to guide treatment strategies.

  10. Fulltext Extraction of Roads Using the Archimedes Tuning Process with the Quantum Dilated Convolutional Neural Network
    Khan MJ, Singh PP, Pradhan B, Alamri A, Lee CW
    Sensors (Basel), 2023 Oct 28;23(21).
    PMID: 37960482 DOI: 10.3390/s23218783
    Road network extraction is a significant challenge in remote sensing (RS). Automated techniques for interpreting RS imagery offer a cost-effective solution for obtaining road network data quickly, surpassing traditional visual interpretation methods. However, the diverse characteristics of road networks, such as varying lengths, widths, materials, and geometries across different regions, pose a formidable obstacle for road extraction from RS imagery. The issue of road extraction can be defined as a task that involves capturing contextual and complex elements while also preserving boundary information and producing high-resolution road segmentation maps for RS data. The objective of the proposed Archimedes tuning process quantum dilated convolutional neural network for road Extraction (ATP-QDCNNRE) technology is to tackle the aforementioned issues by enhancing the efficacy of image segmentation outcomes that exploit remote sensing imagery, coupled with Archimedes optimization algorithm methods (AOA). The findings of this study demonstrate the enhanced road-extraction capabilities achieved by the ATP-QDCNNRE method when used with remote sensing imagery. The ATP-QDCNNRE method employs DL and a hyperparameter tuning process to generate high-resolution road segmentation maps. The basis of this approach lies in the QDCNN model, which incorporates quantum computing (QC) concepts and dilated convolutions to enhance the network's ability to capture both local and global contextual information. Dilated convolutions also enhance the receptive field while maintaining spatial resolution, allowing fine road features to be extracted. ATP-based hyperparameter modifications improve QDCNNRE road extraction. To evaluate the effectiveness of the ATP-QDCNNRE system, benchmark databases are used to assess its simulation results. The experimental results show that ATP-QDCNNRE performed with an intersection over union (IoU) of 75.28%, mean intersection over union (MIoU) of 95.19%, F1 of 90.85%, precision of 87.54%, and recall of 94.41% in the Massachusetts road dataset. These findings demonstrate the superior efficiency of this technique compared to more recent methods.
  11. Investigating the spatial distribution of phototrophic picoplankton in a tropical estuary
    Lee CW, Lim JH, Heng PL
    Environ Monit Assess, 2013 Dec;185(12):9697-704.
    PMID: 23748919 DOI: 10.1007/s10661-013-3283-3
    We sampled extensively (29 stations) at the Klang estuarine system over a 3-day scientific expedition. We measured physical and chemical variables (temperature, salinity, dissolved oxygen, total suspended solids, dissolved inorganic nutrients) and related them to the spatial distribution of phototrophic picoplankton (Ppico). Multivariate analysis of variance of the physicochemical variables showed the heterogeneity of the Klang estuarine system where the stations at each transect were significantly different (Rao's F₁₈, ₃₆ = 8.401, p < 0.001). Correlation analyses also showed that variables related to Ppico abundance and growth were mutually exclusive. Distribution of Ppico was best explained by the physical mixing between freshwater and seawater whereas Ppico growth was correlated with temperature.
  12. Influence of elevated river flow on hypoxia occurrence, nutrient concentration and microbial dynamics in a tropical estuary
    Lee CW, Lim JH, Heng PL, Marican NF, Narayanan K, Sim EUH, et al.
    Environ Monit Assess, 2020 Sep 25;192(10):660.
    PMID: 32975666 DOI: 10.1007/s10661-020-08625-3
    We sampled the Klang estuary during the inter-monsoon and northeast monsoon period (July-Nov 2011, Oct-Nov 2012), which coincided with higher rainfall and elevated Klang River flow. The increased freshwater inflow into the estuary resulted in water column stratification that was observed during both sampling periods. Dissolved oxygen (DO) dropped below 63 μM, and hypoxia was observed. Elevated river flow also transported dissolved inorganic nutrients, chlorophyll a and bacteria to the estuary. However, bacterial production did not correlate with DO concentration in this study. As hypoxia was probably not due to in situ heterotrophic processes, deoxygenated waters were probably from upstream. We surmised this as DO correlated with salinity (R2 = 0.664, df = 86, p  6.7 h), hypoxia could occur at the Klang estuary. Here, we presented a model that related riverine flow rate to the post-heavy rainfall hypoxia that explicated the episodic hypoxia at Klang estuary. As Klang estuary supports aquaculture and cockle culture, our results could help protect the aquaculture and cockle culture industry here.
  13. Investigating the decay rates of Escherichia coli relative to Vibrio parahemolyticus and Salmonella Typhi in tropical coastal waters
    Lee CW, Ng AY, Bong CW, Narayanan K, Sim EU, Ng CC
    Water Res, 2011 Feb;45(4):1561-70.
    PMID: 21146847 DOI: 10.1016/j.watres.2010.11.025
    Using the size fractionation method, we measured the decay rates of Escherichia coli, Salmonella Typhi and Vibrio parahaemolyticus in the coastal waters of Peninsular Malaysia. The size fractions were total or unfiltered, <250 μm, <20 μm, <2 μm, <0.7 μm, <0.2 μm and <0.02 μm. We also carried out abiotic (inorganic nutrients) and biotic (bacterial abundance, production and protistan bacterivory) measurements at Port Dickson, Klang and Kuantan. Klang had highest nutrient concentrations whereas both bacterial production and protistan bacterivory rates were highest at Kuantan. We observed signs of protist-bacteria coupling via the following correlations: Protistan bacterivory-Bacterial Production: r = 0.773, df = 11, p < 0.01; Protist-Bacteria: r = 0.586, df = 12, p < 0.05. However none of the bacterial decay rates were correlated with the biotic variables measured. E. coli and Salmonella decay rates were generally higher in the larger fraction (>0.7 μm) than in the smaller fraction (<0.7 μm) suggesting the more important role played by protists. E. coli and Salmonella also decreased in the <0.02 μm fraction and suggested that these non-halophilic bacteria did not survive well in seawater. In contrast, Vibrio grew well in seawater. There was usually an increase in Vibrio after one day incubation. Our results confirmed that decay or loss rates of E. coli did not match that of Vibrio, and also did not correlate with Salmonella decay rates. However E. coli showed persistence where its decay rates were generally lower than Salmonella.
  14. Fulltext An Overview of Maxillofacial Trauma in Oral and Maxillofacial Tertiary Trauma Centre, Queen Elizabeth Hospital, Kota Kinabalu, Sabah
    Lee CW, Foo QC, Wong LV, Leung YY
    Craniomaxillofac Trauma Reconstr, 2017 Mar;10(1):16-21.
    PMID: 28210403 DOI: 10.1055/s-0036-1584893
    The aims of this study were to provide an overview of maxillofacial trauma and its relationship to patient's demographic data and alcohol consumption within the state of Sabah. It was a retrospective study of maxillofacial trauma cases treated by Oral and Maxillofacial Surgery Department, Queen Elizabeth Hospital, Kota Kinabalu, Sabah, from January 1, 2009, until December 31, 2013. A total of 630 maxillofacial trauma cases were included. Details of the trauma were collected from patients' record, including patients' cause of injuries, injuries suffered, treatment indications, and treatment received. Patients' demographic data (age, gender), alcohol consumption in relation to causes, and type of maxillofacial injury were analyzed. There were 538 male (85.4%) and 92 female (14.6%) patients (ratio: 5.8:1), with mean age of 31.0 years. Most common causes of maxillofacial injury were motor vehicle accident (MVA; 66.3%), followed by fall (12.4%) and assault (11.6%). Motorcyclists made up more than half of the total cases (53.1%). Cases referred were primarily due to soft-tissue injury (458 cases). Other cases were dentoalveolar and maxillofacial bone fractures. Treatment provided for the fractures included open reduction and internal fixation (22.9%), closed reduction (28.7%), and conservative management (48.4%). Toilet and suturing were done for all patients with soft-tissue injury. Maxillofacial trauma is a major problem in Sabah. It affects mostly males in the age group of 21 to 30 years. Most of the MVA patients were motorcyclists. Mandibular fracture with parasymphysis involvement recorded the highest number. Most of the patients preferred conservative management, probably due to financial and logistic issue.
  15. Tunable metasurfaces for visible and SWIR applications
    Lee CW, Choi HJ, Jeong H
    Nano Converg, 2020 Jan 20;7(1):3.
    PMID: 31956942 DOI: 10.1186/s40580-019-0213-2
    Demand on optical or photonic applications in the visible or short-wavelength infrared (SWIR) spectra, such as vision, virtual or augmented displays, imaging, spectroscopy, remote sensing (LIDAR), chemical reaction sensing, microscopy, and photonic integrated circuits, has envisaged new type of subwavelength-featured materials and devices for controlling electromagnetic waves. The study on metasurfaces, of which the thickness is either comparable to or smaller than the wavelength of the considered incoming electromagnetic wave, has been grown rapidly to embrace the needs of developing sub 100-micron active photonic pixelated devices and their arrayed form. Meta-atoms in metasurfaces are now actively controlled under external stimuli to lead to a large phase shift upon the incident light, which has provided a huge potential for arrayed two-dimensional active optics. This short review summarizes actively tunable or reconfigurable metasurfaces for the visible or SWIR spectra, to account for the physical operating principles and the current issues to overcome.
  16. Fulltext Temporal variation of bacterial respiration and growth efficiency in tropical coastal waters
    Lee CW, Bong CW, Hii YS
    Appl Environ Microbiol, 2009 Dec;75(24):7594-601.
    PMID: 19820145 DOI: 10.1128/AEM.01227-09
    We investigated the temporal variation of bacterial production, respiration, and growth efficiency in the tropical coastal waters of Peninsular Malaysia. We selected five stations including two estuaries and three coastal water stations. The temperature was relatively stable (averaging around 29.5 degrees C), whereas salinity was more variable in the estuaries. We also measured dissolved organic carbon and nitrogen (DOC and DON, respectively) concentrations. DOC generally ranged from 100 to 900 microM, whereas DON ranged from 0 to 32 microM. Bacterial respiration ranged from 0.5 to 3.2 microM O2 h(-1), whereas bacterial production ranged from 0.05 to 0.51 microM C h(-1). Bacterial growth efficiency was calculated as bacterial production/(bacterial production + respiration), and ranged from 0.02 to 0.40. Multiple correlation analyses revealed that bacterial production was dependent upon primary production (r2 = 0.169, df = 31, and P < 0.02) whereas bacterial respiration was dependent upon both substrate quality (i.e., DOC/DON ratio) (r2 = 0.137, df = 32, and P = 0.03) and temperature (r2 = 0.113, df = 36, and P = 0.04). Substrate quality was the most important factor (r2 = 0.119, df = 33, and P = 0.04) for the regulation of bacterial growth efficiency. Using bacterial growth efficiency values, the average bacterial carbon demand calculated was from 5.30 to 11.28 microM C h(-1). When the bacterial carbon demand was compared with primary productivity, we found that net heterotrophy was established at only two stations. The ratio of bacterial carbon demand to net primary production correlated significantly with bacterial growth efficiency (r2 = 0.341, df = 35, and P < 0.001). From nonlinear regression analysis, we found that net heterotrophy was established when bacterial growth efficiency was <0.08. Our study showed the extent of net heterotrophy in these waters and illustrated the importance of heterotrophic microbial processes in coastal aquatic food webs.
  17. Fulltext Tumor Necrosis Factor-Alpha Exacerbates Viral Entry in SARS-CoV2-Infected iPSC-Derived Cardiomyocytes
    Lee CY, Huang CH, Rastegari E, Rengganaten V, Liu PC, Tsai PH, et al.
    Int J Mol Sci, 2021 Sep 13;22(18).
    PMID: 34576032 DOI: 10.3390/ijms22189869
    The coronavirus disease 2019 (COVID-19) pandemic with high infectivity and mortality has caused severe social and economic impacts worldwide. Growing reports of COVID-19 patients with multi-organ damage indicated that severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) may also disturb the cardiovascular system. Herein, we used human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (iCMs) as the in vitro platform to examine the consequence of SARS-CoV2 infection on iCMs. Differentiated iCMs expressed the primary SARS-CoV2 receptor angiotensin-converting enzyme-II (ACE2) and the transmembrane protease serine type 2 (TMPRSS2) receptor suggesting the susceptibility of iCMs to SARS-CoV2. Following the infection of iCMs with SARS-CoV2, the viral nucleocapsid (N) protein was detected in the host cells, demonstrating the successful infection. Bioinformatics analysis revealed that the SARS-CoV2 infection upregulates several inflammation-related genes, including the proinflammatory cytokine tumor necrosis factor-α (TNF-α). The pretreatment of iCMs with TNF-α for 24 h, significantly increased the expression of ACE2 and TMPRSS2, SASR-CoV2 entry receptors. The TNF-α pretreatment enhanced the entry of GFP-expressing SARS-CoV2 pseudovirus into iCMs, and the neutralization of TNF-α ameliorated the TNF-α-enhanced viral entry. Collectively, SARS-CoV2 elevated TNF-α expression, which in turn enhanced the SARS-CoV2 viral entry. Our findings suggest that, TNF-α may participate in the cytokine storm and aggravate the myocardial damage in COVID-19 patients.
  18. A Self-Replicating Linear DNA
    Liew PS, Tan TH, Wong YC, Sim EUH, Lee CW, Narayanan K
    ACS Synth Biol, 2020 04 17;9(4):804-813.
    PMID: 32196315 DOI: 10.1021/acssynbio.9b00478
    TelN and tos are a unique DNA linearization unit isolated from bacteriophage N15. While being transferable, the TelN cleaving-rejoining activities remained stable to function on tos in both bacterial and mammalian environments. However, TelN contribution in linear plasmid replication in mammalian cells remains unknown. Herein, we investigated the association of TelN in linear tos-containing DNA (tos-DNA) replication in mammalian cells. Additionally, the mammalian origin of replication (ori) that is well-known to initiate the replication event of plasmid vectors was also studied. In doing so, we identified that both TelN and mammalian initiation sites were essential for the replication of linear tos-DNA, determined by using methylation sensitive DpnI/MboI digestion and polymerase chain reaction (PCR) amplification approaches. Furthermore, we engineered the linear tos-DNA to be able to retain in mammalian cells using S/MAR technology. The resulting S/MAR containing tos-DNA was robust for at least 15 days, with (1) continuous tos-DNA replication, (2) correct splicing of gene transcripts, and (3) stable exogenous gene expression that was statistically comparable to the endogenous gene expression level. Understanding the activities of TelN and tos in mammalian cells can potentially provide insights for adapting this simple DNA linearization unit in developing novel genetic engineering tools, especially to the eukaryotic telomere/telomerase study.
  19. Phage N15 protelomerase resolves its tos recognition site into hairpin telomeres within mammalian cells
    Liew PS, Chen Q, Ng AWR, Chew YC, Ravin NV, Sim EUH, et al.
    Anal Biochem, 2019 10 15;583:113361.
    PMID: 31306622 DOI: 10.1016/j.ab.2019.113361
    Phage N15 protelomerase (TelN) cleaves double-stranded circular DNA containing a telomerase-occupancy-site (tos) and rejoins the resulting linear-ends to form closed-hairpin-telomeres in Escherichia coli (E. coli). Continued TelN expression is essential to support resolution of the linear structure. In mammalian cells, no enzyme with TelN-like activities has been found. In this work, we show that phage TelN, expressed transiently and stably in human and mouse cells, recapitulates its native activities in these exogenous environments. We found TelN to accurately resolve tos-DNA in vitro and in vivo within human and mouse cells into linear DNA-containing terminal telomeres that are resistant to RecBCD degradation, a hallmark of protelomerase processing. In stable cells, TelN activity was detectable for at least 60 days, which suggests the possibility of limited silencing of its expression. Correspondingly, linear plasmid containing a 100 kb human β-globin gene expressed for at least 120 h in non-β-globin-expressing mouse cells with TelN presence. Our results demonstrate TelN is able to cut and heal DNA as hairpin-telomeres within mammalian cells, providing a tool for creating novel structures by DNA resolution in these hosts. The TelN protelomerase may be useful for exploring novel technologies for genome interrogation and chromosome engineering.
  20. Temporal variation of phytoplankton growth and grazing loss in the west coast of Peninsular Malaysia
    Lim JH, Lee CW, Kudo I
    Environ Monit Assess, 2015 May;187(5):246.
    PMID: 25864082 DOI: 10.1007/s10661-015-4487-5
    Phytoplankton growth (μ) and grazing loss (g) rates were measured monthly by the Landry-Hassett dilution method over a 2-year period at both estuarine (Klang) and coastal water (Port Dickson) systems along the Straits of Malacca. Chlorophyll a (Chl a) concentration ranged from 0.20 to 4.47 μg L(-1) at Klang except on two occasions when Chl a spiked above 10 μg L(-1). In contrast, Chl a concentrations were relatively stable at Port Dickson (0.14 to 2.76 μg L(-1)). From the rate measurements, μ was higher (t = 2.01, df = 43, p  0.80). g ranged from 0.30 to 1.50 and 0.21 to 1.51 day(-1) at Klang and Port Dickson, respectively. In this study, grazing loss was coupled to phytoplankton growth, and the ratio of g/μ or grazing pressure which estimates the proportion of primary production grazed was 50% at Klang and lower than at Port Dickson (68%; t = 2.213, df = 36, p 
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