Automated Uniform Spheroid Generation Platform for High Throughput Drug Screening Process.

Three-dimensional (3D) spheroid models are crucial for cancer research, offering more accurate insights into tumour biology and drug responses than traditional 2D cell cultures. However, inconsistent and low-throughput spheroid production has hindered their application in drug screening. Here, we present an automated high-throughput platform for a spheroid selection, fabrication, and sorting system (SFSS) to produce uniform gelatine-encapsulated spheroids (GESs) with high efficiency. SFSS integrates advanced imaging, analysis, photo-triggered fabrication, and microfluidic sorting to precisely control spheroid size, shape, and viability. Our data demonstrate that our SFSS can produce over 50 GESs with consistent size and circularity in 30 min with over 97% sorting accuracy while maintaining cell viability and structural integrity. We demonstrated that the GESs can be used for drug screening and potentially for various assays. Thus, the SFSS could significantly enhance the efficiency of generating uniform spheroids, facilitating their application in drug development to investigate complex biological systems and drug responses in a more physiologically relevant context.

Biological monitoring of human exposure to environmental lead in Singapore.

An epidemiological survey was conducted in 1990-91 to determine the personal and environmental factors in Singapore which may affect blood lead levels. A total of 1,408 children and adults aged from 6 months to over 50 years were interviewed and their blood samples obtained for analysis using atomic absorption spectroscopy. Overall, the geometric mean blood lead level of the population surveyed was 76.6 micrograms/l. Multivariate analyses revealed that significant variations in blood lead concentrations were related, in decreasing order of importance, to: sex, age, active and passive smoking, exposure to traffic, and exposure to recent paintwork. The consumption of alcohol and traditional medicines were found not to be significant determinants. To reduce the environmental lead problem in Singapore, unleaded petrol has been introduced and control of lead additive in paints is being considered. The national anti-smoking programme should further contribute to an overall reduction in the blood lead level of the population.

Blood cadmium levels in non-occupationally exposed adult subjects in Singapore.

Cadmium concentrations in whole blood were measured in subjects with no occupational exposure to cadmium. The study covered 128 males and 150 females from the three main ethnic groups in Singapore (namely Chinese, Malays and Indians). The geometric means (GM) of blood cadmium (CdB) levels of non-smoking males and females were 0.21 microgram/l and 0.26 microgram/l, respectively. Smokers had higher GM CdB levels than non-smokers. Significant ethnic differences were observed in both sexes, with Indian males having the highest GM CdB level of 0.48 microgram/l. Among the females, the Chinese had the highest GM CdB level: 0.33 microgram/l. Differences in dietary habits may have contributed to the observed ethnic differences in blood cadmium levels.

Effect of metabolic inhibitors on growth and brefeldin A formation in Curvularia lunata.

Arsenite as opposed to azide exerted a stronger inhibitory effect on brefeldin A production than on mycelial growth of cultures of Curvularia lunata. However, azide was a more potent inhibitor of mycelial growth than arsenite at 0.5 mM and 1.0 mM. The inhibitory effects of iodoacetate on growth and brefeldin A formation were significantly less potent than those of iosoacetamide. Increases in the levels of fluoride elicited a variable inhibitory effect on brefeldin A production and a corresponding decrease in biomass.

Biochemical changes accompanying brefeldin A formation in Curvularia lunata.

Extracellular brefeldin A was detected in 4% glucose--peptone--mineral salts cultures of Curvularia lunata at the start of the exponential growth phase. Some fluctuations in brefeldin A levels occurred during the exponential growth phase followed by a significant reduction in level at the stationary growth phase. Broth glucose levels decreased according to a sigmoid relationship with time whereas broth pH remained fairly constant during the exponential growth phase followed by a gradual increase into the stationary growth phase. Mycelial brefeldin A levels were low throughout the various growth phases. The principal fatty acids present in decreasing order during the exponential growth phase were linoleic, oleic and palmitic acids. However, the content of linoleic acid was significantly reduced at the onset and during the stationary growth phase.

Effect of malonate and maleate on growth and brefeldin A formation in Curvularia lunata.

Malonate exerted a stronger inhibitory effect on brefeldin A production than on mycelia growth in cultures of Curvularia lunata especially at inhibitory levels of 100 to 200 mM. The extent of 200 mM malonate inhibition of growth and brefeldin A production was greater in cultures treated with malonate prior to inoculation than those treated following 5 days after inoculation. Maleate at levels of 40 to 220 mM activated brefeldin A formation in cultures though exerting variable effects on mycelia growth.

Effect of brefeldin A and actinomycin D on culture growth and brefeldin A yield in Curvularia lunata.

Cultures incorporated with increasing quantities of brefeldin A in the form of crude extracts of fungal metabolites prior to inoculation demonstrated reduced growth rate and no significant increase in brefeldin A content. On the other hand, cultures incubated with increasing levels of actinomycin D on the 8th day of cultivation showed slight stimulation of brefeldin A formation with insignificant effect on growth.

Effect of nutrient variations on growth and brefeldin A formation in Curvularia lunata.

The highest level of secreted brefeldin A was present in glucose-grown cultures, intermediate levels in glucose-fructose, and xylose cultures and low levels in fructose- and galactose-grown cultures of Curvularia lunata. The biomass decreased from glucose, fructose, xylose, glucose-fructose to galactose cultures. Brefeldin A levels and mycelial yields were low in citrate-, gluconate-, and succinate-grown cultures. Inorganic phosphate-limited cultures supported a high level of brefeldin A. Intermediate levels were present in trace elements-, and inorganic phosphate-trace elements-limited cultures.

Effect of substrates on the malonate inhibitory pattern and brefeldin A formation in Curvularia lunata.

The extracellular level of brefeldin A fluctuates with the length of malonate inhibition. Following treatment with malonate, mycelial multiplication as opposed to brefeldin A formation, was preferentially increased in the maleate, fumarate, succinate, citrate, methyl palmitate and glucose replacement cultures. Competitive maleate--malonate, fumarate--malonate, succinate--malonate and citrate--malonate-inhibited replacement cultures gave significantly higher mycelial and brefeldin A yields than the sole malonate-inhibited replacement cultures.

Fatal gassing due to methylene chloride - a case report.

Methylene chloride is a major component of paint and varnish strippers. Due to its high volatility, its use in unventilated or poorly ventilated confined spaces poses a serious health hazard as a result of accumulation of the solvent vapour. At high levels, methylene chloride can cause severe central nervous system depression and ultimately death, as illustrated in this case report. The importance of adequate forced ventilation, utilisation of proper personal protective equipment and enforcement of a permit-to-work system during work with solvents in confined spaces is emphasised.

Blood lead levels of a population group not occupationally exposed to lead in Singapore.

A survey was conducted between 1995 and 1997 to assess the impact of introduction of unleaded petrol and other public health measures on the blood lead level of the population. The geometric mean blood lead level of 269 government employees as determined by graphite furnace atomic absorption spectroscopy, was 66.0 microg/l, much lower than that recorded before introduction of lead-free petrol. Using multiple regression analysis, factors significantly associated with blood lead levels were: exposure to traffic, age (>50 years) and active smoking. Passive smoking, exposure to recent paint work, consumption of alcohol and traditional medicine were found not to be significantly associated with the blood lead level.

The determination of chlorofluorocarbons and extractives in the expanded and disposable polystyrene ware.

Residual chlorofluorocarbons (CFCs) and water-extractive residues in expanded polystyrene ware were determined because of the threat to environmental and public health arising from the ozone depleting potential of CFCs and the leaching of toxic monomers and oligomers into food and drink. Chlorofluorocarbon in expanded polystyrene ware was analysed using headspace gas chromatography in the flame ionization mode, and confirmed by gas chromatography-mass spectrometry. The amount of extractive residue leached from this type of polymeric material was determined using an aqueous medium simulating categories of food and beverages. A wide range of concentrations of CFC-12 (dichlorodifluorocarbon) was detected in the styrofoam ware. It fluctuated between a low of 32 ppm and a high of 35471 ppm in two different samples of styrofoam plates. Similarly, the highest and lowest levels of extractive residue detected in two different styrofoam plates were 54 ppm and 0.5 ppm respectively. CFC-22 or HCFC (chlorodifluorocarbon) was absent from these expanded polystyrene ware. The continued use of chlorofluorocarbons as a blowing agent will further exert a deleterious effect on the stratospheric ozone layer. Though the level of water extractive residue from these expanded polystyrene ware was relatively safe, public health could be further safe-guarded through improvmeents in raw material selection and mode of production.

Lead exposure in the lead-acid storage battery manufacturing and PVC compounding industries.

This study was conducted as part of the Human Exposure Assessment Location (HEAL) Project which comes under the United Nations Environment Programme/World Health Organisation (UNEP/WHO) Global environmental Monitoring System (GEMS). The objective of the study was to evaluate workers' exposure to lead in industries with the highest exposure. All subjects were interviewed about their occupational and smoking histories, the use of personal protective equipment and personal hygiene. The contribution of a dietary source of lead intake from specified foods known to contain lead locally and personal air sampling for lead were assessed. A total of 61 workers from two PVC compounding and 50 workers from two lead acid battery manufacturing plants were studied together with 111 matched controls. In the PVC compounding plants the mean lead-in-air level was 0.0357 mg/m3, with the highest levels occurring during the pouring and mixing operations. This was lower than the mean lead-in-air level of 0.0886 mg/m3 in the lead battery manufacturing plants where the highest exposure was in the loading of lead ingots into milling machines. Workers in lead battery manufacturing had significantly higher mean blood lead than the PVC workers (means, 32.51 and 23.91 mcg/100 ml respectively), but there was poor correlation with lead-in-air levels. Among the lead workers, the Malays had significantly higher blood lead levels than the Chinese (mean blood levels were 33.03 and 25.35 mcg/100 ml respectively) although there was no significant difference between the two ethnic groups in the control group. There were no significant differences between the exposed and control group in terms of dietary intake of specified local foods known to contain lead. However, Malays consumed significantly more fish than the Chinese did. There were no ethnic differences in the hours of overtime work, number of years of exposure, usage of gloves and respirators and smoking habits. Among the Malays, 94.3% eat with their hands compared with 9.2% of the Chinese. Workers who ate with bare hands at least once a week had higher blood lead levels after adjusting for lead-in-air levels (mean blood lead was 30.2 and 26.4 mcg/100 ml respectively). The study indicated that the higher blood lead levels observed in the Malay workers might have been due to their higher exposure and eating with bare hands.

Environmental and biological monitoring of occupational exposure to 1,1,1-trichloroethane.

Fifty workers involved in various degreasing and cleaning processes using 1,1,1-trichloroethane (1,1,1-TCE) were studied with respect to personal and static exposures. In addition, end-of-shift expired air and venous blood samples were taken for analysis of the parent compound. Urinary samples were also obtained at the same time for analysis of its metabolites-trichloroethanol (TCOH) and trichloracetic acid (TCA). The results show that open/manual degreasing processes generate the highest environmental solvent levels (mean = 819.9 mg/m3; SD = 781.9 mg/m3) followed by jet-spray cleaning (mean = 460.5 mg/m3; SD = 292.4 mg/m3), vapour degreasing (mean = 365.3 mg/m3; SD = 279.9 mg/m3) and ultrasonic degreasing (mean = 134.7 mg/m3; SD = 121.0 mg/m3). Personal exposure levels were well correlated with concentrations of 1,1,1-TCE in end-of-shift expired air (r = 0.81) and venous blood samples (r = 0.88) but only moderately correlated with concentrations of its metabolites in urine (r = 0.49 for TCOH; r = 0.58 for TCA). Static (area) samples were poorly correlated with the biological exposure indices studied.