Assessing the bioactivity of crystalline silica in heated high-temperature insulation wools

Figures & data

Table 1. Physicochemical characteristics of the HTIW, including surface area, solubility, crystalline silica content, and number-weighted arithmetic mean fiber length and diameter, as well as aspect ratio and % fibers with a diameter less than 3 µm. d.µm refers to the hydrodynamic diameter, and PdI to the polydispersity index.

Material	Surface area (m^2/g)	Solubility (ppm)		Heating schedule (temp/time)	Crystalline silica content (w/w%)	Av. Length (μm)	Av. Diameter (μm)	Aspect ratio	fiber diameter <3 µm (%)
		pH 4.5	pH 7.4
CMS-f	1.06	225.84	171.30	–	–	17.4	2.9	6.0:1	59.9
CMS-f/h	0.54	152.00	128.00	1150 °C/48h	19.4	20.0	3.9	5.1:1	37.7
MS-f	1.58	139.61	106.02	–	–	15.4	2.8	5.5:1	58.2
MS-f/h	0.58	6.32	3.51	1100 °C/60h	33.1	18.3	3.1	5.9:1	51.4
CaS-f	0.73	139.78	119.50	–	–	24.2	3.7	6.5:1	38.9
CaS-f/h	0.48	38.31	105.60	1100 °C/24h	29.4	27.1	4.7	5.8:1	24.8
RCF-f	0.64	16.19	7.49	–	–	21.3	3.8	5.6:1	42.5
RCF-f/h	0.64	5.59	5.17	1225 °C/54h	21.7	22.4	4.1	5.5:1	36.0
						d.µm	PdI
CMS-g	2.56	239.04	186.57	–	–	1.5 ± 0.2	0.18
CMS-g/h	0.68	125.67	60.21	1150 °C/48h	19.4 (assumed)	1.5 ± 0.5	0.73

Further details of the solubility assay can be seen in Table S2.

Figure 1. Scanning electron microscopy images of the fibers and particles investigated, including CMS-f (unheated fibers), CMS-f/h (heated fibers), MS-f (unheated fibers), MS-f/h (heated fibers), CaS-f (unheated fibers), CaS-f/h (heated fibers), RCF-f (unheated fibers), RCF-f/h (heated fibers). The scale of each image is included at the bottom right of each image, and represents 50 μm on the HTIW images, and 5 μm on the images of DQ12 and TiO₂.

Figure 2. SEM images of untreated/control J774A.1 cells, and cell treated with CMS-f (unheated fibers), CMS-f/h (heated fibers), MS-f (unheated fibers), MS-f/h (heated fibers), CaS-f (unheated fibers), CaS-f/h (heated fibers), RCF-f (unheated fibers), RCF-f/h (heated fibers). Cells were exposed to 40 μg/cm² for 24 h prior to SEM preparation, and imaged using a FEI Quanta 3D FEG scanning electron microscope. A scale bar is shown at the bottom right corner of each image.

Figure 3. Cell viability, determined by mitochondrial activity (AlamarBlue assay), in response to HTIW. A-C) A549 cells and D-F) J774A.1 cells were treated for 24 h with 0-80 μg/cm² heated and unheated CMS-f, MS-f, CaS-f and RCF-f, as well as particle controls DQ12 and TiO₂. Results are expressed as % viability compared to vehicle (medium-only) control, and each data point represents the mean ± SD of at least three independent biological replicates. Statistical significance of particle exposures in comparison to medium only is indicated with * when p < 0.05.

Figure 4. GRO-α release from A549 cells treated for 24 h with 0-40 μg/cm² heated and unheated CMS-f, MS-f, CaS-f and RCF-f, as well as particle controls DQ12 and TiO₂. Each data point represents the mean ± SD of at least three independent biological replicates. Statistical significance of particle exposures in comparison to medium only is indicated when p < 0.05, identifying markers are shown on corresponding legends, and with ∞ when a heated sample is significantly different to an unheated sample.

Figure 5. TNF-α release from J774A.1 cells treated for 24 h with 0-40 μg/cm² heated and unheated CMS-f, MS-f, CaS-f and RCF-f, as well as particle controls DQ12 and TiO₂. Each data point represents the mean ± SD of at least three independent biological replicates. Statistical significance of particle exposures in comparison to medium only is indicated when p < 0.05, identifying markers are shown on corresponding legends.

Figure 6. RANTES release from J774A.1 cells treated for 24 h with 0-40 μg/cm² heated and unheated CMS-f, MS-f, CaS-f and RCF-f, as well as particle controls DQ12 and TiO₂. Each data point represents the mean ± SD of at least three independent biological replicates. Statistical significance of particle exposures in comparison to medium only is indicated when p < 0.05, identifying markers are shown on corresponding legends, and with ∞ when a heated sample is significantly different to an unheated sample.

Figure 7. MIP-2 release from J774A.1 cells treated for 24 h with 0-40 μg/cm² heated and unheated CMS-f, MS-f, CaS-f and RCF-f, as well as particle controls DQ12 and TiO₂. Each data point represents the mean ± SD of at least three independent biological replicates. Statistical significance of particle exposures in comparison to medium only is indicated when p < 0.05, identifying markers are shown on corresponding legends, and with ∞ when a heated sample is significantly different to an unheated sample.

Figure 8. Effect of aluminum lactate coating on the release of pro-inflammatory mediators, including GRO-α from A549 cells (A), and TNF-α (B) and MIP-2 (C) from J774A.1 cells. Cells were treated for 24 h with 0-40 μg/cm² heated, crystalline silica-containing CMS-f/h. The test material was either coated with AL (grey lines) or processed in the same fashion in PBS without AL (black lines). Each data point represents the mean ± SD of at least three independent biological replicates. Statistical significance of PBS processed particles compared to medium only is indicated with * when p < 0.05, and when AL coating is significantly different to without AL with ∞ when p < 0.05.

Figure 9. (A) Scanning electron microscopy images of calcium magnesium silicate wools investigated after the fibers had been ground to create a less fibrous morphology, including CMS-g (unheated) and CMS-g/h (heated). The scale of each image is included at the bottom right of each image, and represents 50 μm. B) GRO-α release from A549 cells and MIP-2, RANTES and TNF-α release from J774A.1 cells, all treatments were for 24 h with 0-40 μg/cm² heated and unheated CMS-g. Each data point represents the mean ± SD of at least three independent biological replicates. Statistical significance of particle exposures in comparison to medium only is indicated when p < 0.05, with * for CMS-g and + for CMS-g/h.