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Fulltext Optimisation of a PC12 cell-based in vitro stroke model for screening neuroprotective agents

Chua P, Lim WK

Sci Rep, 2021 04 14;11(1):8096.

PMID: 33854099 DOI: 10.1038/s41598-021-87431-4

Stroke causes death and disability globally but no neuroprotectant is approved for post-stroke neuronal injury. Neuroprotective compounds can be identified using oxygen glucose deprivation (OGD) of neuronal cells as an in vitro stroke model. Nerve growth factor (NGF)-differentiated PC12 pheochromocytoma cells are frequently used. However, investigators often find their clonal variant undifferentiable and are uncertain of optimal culture conditions. Hence we studied 3 commonly used PC12 variants: PC12 Adh, PC12 from Riken Cell Bank (PC12 Riken) and Neuroscreen-1 (NS-1) cells. We found DMEM the optimal media for PC12 Riken and NS-1 cells. Using a novel serum-free media approach, we identified collagen IV as the preferred adhesive substrate for both cell lines. We found PC12 Adh cells cannot attach without serum and is unable to differentiate using NGF. NS-1 cells differentiated to a maximal 72.7 ± 5.2% %, with substantial basal differentiation. We optimised differentiated NS-1 cells for an in vitro stroke model using 3 h of OGD resulting in ~ 70% viable cells. We screened 5 reported neuroprotectants and provide the first report that serotonin is antiapoptotic in a stroke model and the 5-HT1A agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) is neuroprotective in PC12 cells. Thus we demonstrate the optimisation and validation for a PC12 cell-based in vitro stroke model.

Fulltext The strategic uses of collagen in adherent cell cultures

Chua P, Lim WK

Cell Biol Int, 2023 Feb;47(2):367-373.

PMID: 36423248 DOI: 10.1002/cbin.11966

The culture of adherent mammalian cells involves adhesion to the tissue culture vessel. This requires attachment factors from serum and/or a suitable substrate on the vessel surface. Some cells require collagen or other substrates to promote neurite outgrowth, differentiation or growth. However, laboratories often lack guidance on the selection and/or optimisation of collagen. We model such selection/optimisation work in the PC12 neuronal cell line. PC12 (NS-1 variant) cells require a substrate for adherence. Comparing cell attachment against a series of substrates, we found collagen IV to be optimal. We show by comparison of morphology against a range of concentrations that 10 µg/ml is sufficient for supporting cell attachment, and also differentiation. PC12 cells from Riken Cell Bank do not require a substrate for routine culturing but only for differentiation. As all substrates supported attachment equally well, we used a novel serum-free approach and identified collagen IV as its preferred substrate. For these cells, Dulbecco's modified eagle's medium but not Roswell Park Memorial Institute (RPMI) media supports normal cell attachment. However, coating with collagen IV enabled the cells to grow equally well in RPMI. Hence the strategic use of collagen is essential in laboratories working with anchorage-dependent cell lines.

Fulltext Draft Genome Sequences of Two Cellulolytic Paenibacillus sp. Strains, MAEPY1 and MAEPY2, from Malaysian Landfill Leachate

Chua P, Yoo HS, Gan HM, Lee SM

Genome Announc, 2014;2(1).

PMID: 24526641 DOI: 10.1128/genomeA.00065-14

We report the draft genome sequences of two Paenibacillus species with cellulose-degrading abilities isolated from landfill leachate. An array of genes putatively involved in cellulose degradation have been identified in both genome sequences, which can benefit various biotechnological industries.

Fulltext Genome sequencing and annotation of Cellulomonas sp. HZM

Chua P, Har ZM, Austin CM, Yule CM, Dykes GA, Lee SM

Genom Data, 2015 Sep;5:40-1.

PMID: 26484221 DOI: 10.1016/j.gdata.2015.05.009

We report the draft genome sequence of Cellulomonas sp. HZM, isolated from a tropical peat swamp forest. The draft genome size is 3,559,280 bp with a G + C content of 73% and contains 3 rRNA sequences (single copies of 5S, 16S and 23S rRNA).

Fulltext Genome sequencing and annotation of Aeromonas sp. HZM

Chua P, Har ZM, Austin CM, Yule CM, Dykes GA, Lee SM

Genom Data, 2015 Sep;5:38-9.

PMID: 26484220 DOI: 10.1016/j.gdata.2015.05.008

We report the draft genome sequence of Aeromonas sp. strain HZM, isolated from tropical peat swamp forest soil. The draft genome size is 4,451,364 bp with a G + C content of 61.7% and contains 10 rRNA sequences (eight copies of 5S rRNA genes, single copy of 16S and 23S rRNA each). The genome sequence can be accessed at DDBJ/EMBL/GenBank under the accession no. JEMQ00000000.

Phytochemicals from Calophyllum canum Hook f. ex T. Anderson and their neuroprotective effects

Lizazman MA, Jong VYM, Chua P, Lim WK, Karunakaran T

Nat Prod Res, 2023 Jun;37(12):2043-2048.

PMID: 35997666 DOI: 10.1080/14786419.2022.2116021

Previous phytochemical investigations reported that Calophyllum spp have biosynthesized a wide range of bioactive phenolics such as xanthones and coumarins. The phytochemical study conducted on the stem bark of C. canum has led to the isolation of eight trioxygenated xanthones namely: 5-methoxytrapezifolixanthone (1), 5-methoxyananixanthone (2), caloxanthone C (3), 1,5-dihydroxy-3-methoxy-4-isoprenylxanthone (4), 6-deoxyisojacareubin (5), euxanthone (6), trapezifolixanthone (7), ananixanthone (8), together with three common triterpenoids, β-sitosterol (9), friedelin (10), and stigmasterol (11). Furthermore, xanthones 1 and 2 were isolated for the first time as naturally occurring xanthones from the plant extract. The structures of these compounds were identified and elucidated using advanced spectroscopic techniques such as 1 D & 2 D NMR, MS, and FTIR. The neuroprotective property of selected compounds was tested through in vitro stroke model. Among all tested compounds, 1 µm of compounds 8, 9, and 10 showed significant neuroprotective activity via reduction of apoptosis by ∼ 50%.

Lifestyle factors associated with cardiovascular risk among healthcare workers from the tertiary hospitals in Sarawak

Kuan PX, Chan WK, Chua PF, Yeo J, Sapri FE, Bujang MA, et al.

Malays Fam Physician, 2020;15(1):15-22.

PMID: 32284800

INTRODUCTION: A cross-sectional study is used to evaluate the lifestyle factors associated with cardiovascular disease (CVD) risk among healthcare workers in tertiary hospitals in Sarawak, Malaysia.

METHODS: A questionnaire-based survey using the Simple Lifestyle Indicator Questionnaire (SLIQ) was administered to, and anthropometric measurements were collected from, 494 healthcare workers.

RESULTS: The mean age of the subjects was 32.4±8.4, with a range of 19 to 59 years. The subjects were from the allied health (45.5%), management and professional (25.1%) and executive (29.4%) fields. Overall, 47.4% of the subjects were of normal weight, 30.2% were overweight, 17.2% were obese and 5.2% were underweight. The mean number of working hours per week for the subjects was 47.6±14.0 with the highest working hours found among the management and professional group, followed by the executive and allied health groups. Overall, 39.7% of the healthcare workers worked office hours, 36.6% worked within the shift system, 20.9% worked office hours and were on-call and the remaining 2.8% worked a mixture of office hours and shifts. Based on the SLIQ score, 58.1% were classified as at intermediate risk for CVD, 38.5% were in the healthy category and 3.4% were in the unhealthy category. Factors associated with a healthier lifestyle were being female (Odds Ratio [OR]= 12.1; CI=3.2-46.4), professional (mean score= 6.70), in the allied health group (mean score=7.33) and in the normal BMI group (OR= 9.3, CI= 1.8-47.0).

CONCLUSION: In our study, healthcare workers had an intermediate risk of developing CVD in the future. Thus, there is a need to intervene in the lifestyle factors contributing to CVD.

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