Is the particle deposition in a cell exposure facility comparable to the lungs? A computer model approach

Figures & data

Figure 1. (a) Scheme of a VitroCell automated exposure station (air-liquid interface exposure system). The aerosol path from outside to the exposure site is kept at a temperature of 37 °C by the system containment heater and circulation. It is conditioned to a relative humidity of 85% in the humidifier. The particles are isokinetically sampled from the humidifier and transported to the exposure wells in horizontal sampling lines. Flow rate through the well is controlled for 100 cm³ min⁻¹. Six wells are grouped into an exposure module. The system holds three modules with identical properties. One exposure module is used for clean air reference. The section numbers indicate the calculation steps for particle loss estimation (Table 1c). (b) Scheme of the air-liquid interface (ALI) exposure setup. The aerosol is delivered to the ALI via a trumpet-shaped flow-guiding element. Particles deposit onto the cells by diffusion and sedimentation. The well keeps the insert in place. Cells grow on a membrane in the insert with the medium from the basolateral and the exposure aerosol from the apical side. R_i is the inlet radius, R_w the radius of the membrane of an insert in a well and h_t the distance of the trumpet from the cell membrane (see Table 1b).

Table 1. ALI characteristics. (a) Parameters for the ALI deposition Equation (1a(1a) $${DE}_{\mathit{ALI}}=\alpha \mathrm{\hspace{0.17em}}{\left(\frac{d_p}{d_0}\right)}^{\beta }+\frac{2\mathrm{ }{\left(\gamma \mathrm{ }{e}^{{\rho }_p\epsilon }+\mathrm{ }{m}_0\right)}^2\mathrm{\hspace{0.17em}}{d}_p^2\mathrm{ }}{R_i^2}$$(1a) ) are taken from Comouth et al. (2013), his Table 1. The value for m0 is modified here to fit the experimental data in his Figure 9. Model results are valid for the operational parameters in Table 1b. (b) Operational and geometric parameters of the ALI setup. (c) Parameters for the estimation of the particle transmission from the inlet of a VitroCell automated exposure station to the inlet of a well-trumpet (see Figure 1a).

Identifier	Value	Unit		Meaning
(a) ALI deposition model
α	8.81 × 10^−13	–		Constant
β	−1.33618	–		Constant
γ	−905.207	–		Constant
ε	9.71 × 10^-5	m^3 kg^−1		Constant
d0	1	m		Normalization constant
m0	805.16	–		constant m0 = 1015.43 in Comouth et al. (2013)
Ri	3 × 10^-3	m		Inlet radius
ρp	1000	kg m^-3		Particle density
(b) ALI Operational and geometric parameters
Q	100	cm³ min^−1		Well flow rate
	37	°C		Aerosol temperature
	85	%		Aerosol relative humidity
Rw	12.2	mm		Radius a well
ht	2	mm		Distance of trumpet from membrane surface (Comouth et al. 2013)
(c) ALI Particle Transmission Estimation
Q	1	m³ h^−1		ALI inlet flow rate (aspiration)
D; L	10; 250	mm; mm		Section 1: Vertical inlet tube after pre-impactor: diameter; length
D1; D2; L	10; 50; 50	mm; mm; mm		Section 2: Vertical transition cone to reactor: inlet diameter; outlet diameter; length
D; L	50; 400	mm; mm		Section 3: Vertical reactor: diameter; length
D; L	4; 50	mm; mm		Section 4: Isokinetic sampling tube: diameter; length
Q	100	cm³ min^−1		Flow rate through sampling tube to well-trumpet
φ; Rφ; D	90; 20; 4	°; mm; mm		Bend in sampling tube: bend angle; bend radius; tube diameter
ϴ; D; L	0; 4; 200	°; mm; mm		Section 5: Horizontal transfer line to the well, sedimentation: angle with gravity; diameter; length
D; L	4; 200	mm; mm		Transfer line to the well, diffusion: diameter; length
φ; Rφ; D	90; 50; 4	°; mm; mm		Bend in transfer hose: angle of bend; radius of bend; tube diameter
D; L	4; 50	mm; mm		Vertical transfer hose to trumpet: diameter; length

Figure 2. (a) Parameters of the lung structure model of Yeh and Schum (1980) corrected for a mean lung volume of 3 675 cm³ with Equation (2b(2b) $$f={\left(\frac{LV}{\mathit{LVM}}\right)}^{\raisebox{1ex}{$1$}\!\left/ \!\raisebox{-1ex}{$3$}\right.}$$(2b) ). The model is extended with a nose or mouth and an oropharynx (Ferron, Haider, and Kreyling 1988; Ferron, Kreyling, and Haider 1988). Data are presented as a function of the lung generation i. Nose or mouth is i = 1, the oropharynx i = 2, the trachea i = 3, the main bronchi i = 4, the last bronchi i = 11, the bronchioles i = 12 to 19 and the alveolar ducts i = 20 to 26 (ICRP 1994). Background colors indicate the extra-thoracic (ET, green), tracheo-bronchial (TB, blue) and alveolar (AL, red) lung region. (b) Parameters calculated from the lung structure model of Yeh and Schum (1980) (see part (a)). Surface area A_lung(i) of a lung generation i is calculated with Equation (4)(4a) $$A_{\mathit{lung}}\left(i\right)=\pi \mathrm{ }L\left(i\right)\mathrm{ }D\left(i\right)\mathrm{ }N\left(i\right)$$(4a) . (c) Lung deposition calculated with the HPLD model as a function of lung generation and particle size. Blue color marks nearly zero deposition, red color maximum deposition. Calculation is performed for spherical particles with a density of 1 g cm⁻³ and for a sitting male adult breathing by mouth with a tidal volume of 750 cm³, a constant respiration airflow, an equal in- and exhalation time (Table 3). (d) Same as (c), but for nose breathing.

Table 2. Surface area of the cell layer at the ALI and of the lung structure of Yeh and Schum (1980) corrected for a lung volume of 3675 cm³. The alveolar surface area is calculated with Equation (4c(4c) $$A_{\mathit{lung}}\left(AL\right)=A_{\mathit{lung}}\left(TL\right)-A_{\mathit{lung}}\left(ET\right)-A_{\mathit{lung}}\left(TB\right)$$(4c) ). Total lung surface area is taken from Reference Man (ICRP 1975).

Region	Surface area		Lung generation	Meaning
r	A(r)	unit	i
ALI	4.7	cm²	–	Cell area air-liquid interface
ET	89.5	cm²	1, 2	Mouth and nose, extrathoracic region
TB	0.33	m²	3–19	Tracheo-bronchial region
AL	74.7	m²	20–26	Alveolar region
TL	75	m²	–	Total lung (ICRP 1975)

Table 3. Parameters used for the HPLD model calculations (Ferron et al. 2013). The respiration conditions are for a quietly breathing male person (ICRP 1994).

Parameter	Value	Unit
Particle density (non-hygroscopic)	1.0	g/cm³
Breathing path	mouth and nose	–
Inhalation/exhalation time	2.5/2.5	s
Breath duration	5	s
Breathing frequency	12	min^−1
Breathing minute volume	9 × 10^3	cm³ min^−1
Hourly breath flow rate	0.54	m³ h^−1
In- and exhalation airflow	250	cm³ s^−1
Functional Residual Capacity (FRC)	3300	cm³
Lung volume (LV)	3675	cm³
Tidal Volume (VT)	750	cm³
Mouth & oropharynx	50	cm³
Nose & nasopharynx	50	cm³

Table 4. Properties of several human lung cells and ALI cell lines. The weighted average of type-I and type-II pneumocytes is used for the TL calculations, as type-I cells contribute 94% to the epithelial surface area and type-II cells 6%. For the ALI exposure, the A549 cell line is assumed. BEAS-2B and 16HBE cell lines are included for comparison.

Cell species	Number of cells per surface area [cells per cm²]	Area of one cell [µm²]	Remarks
Human type-I pneumocytes^a	14 413	6 938	Human alveolar lung cell
Human type-II pneumocytes^a	395 257	253	Human alveolar lung cell
Weighted average type-I and type-II^a	15 300	6 536	Average of total lung
A549^b	400 000	250	Alveolar cell line
BEAS-2B^c	300 000	333	Bronchial cell line
16HBE140^c	700 000	143	Human bronchial epithelial cell line

^a Stone et al. (1992).

^b Lenz et al. (2009).

^c ATCC (2018a, 2018b).

Table 5. Parameters for the lognormal particle size distribution used as airborne particle exposure scenario. It mimics the size distribution of a diesel emission with 100 nm modal diameter. The corresponding mass median diameter is derived by Hatch-Choate conversion (Hinds 1999). Number and mass distribution are adjusted for a total concentration of 10⁶ cm⁻³ and 1 mg m⁻³, respectively.

Particle emission distribution scenario
Parameter	Number	Mass	Unit	Meaning
median	100	194	nm	Count or mass median diameter
σg	1.6	1.6	–	Geometric standard deviation
C	10^6	7.07 × 10^5	cm^−3	Number concentration parameter
Total sum	10^6 cm^−3	1mg m^−3		Total concentration

Figure 3. (a) ALI and regional lung deposition. Deposition at the ALl is calculated with Equation (1a(1a) $${DE}_{\mathit{ALI}}=\alpha \mathrm{\hspace{0.17em}}{\left(\frac{d_p}{d_0}\right)}^{\beta }+\frac{2\mathrm{ }{\left(\gamma \mathrm{ }{e}^{{\rho }_p\epsilon }+\mathrm{ }{m}_0\right)}^2\mathrm{\hspace{0.17em}}{d}_p^2\mathrm{ }}{R_i^2}$$(1a) ) as a function of particle size. Total and regional lung deposition is calculated with the HPLD model for mouth respiration. Modeling conditions are listed in Tables 1a and 3 for ALI and lung, respectively. (b) Ratio DE_lung(r)/DE_ALI of the regional lung deposition and the ALI deposition (a). The horizontal bars show the half-width for the ET (green), TB (blue), AL (red) and TL (yellow) region (Table 6).

Table 6. Half-width ranges for the ratio DE_lung(r)/DE_ALI and DA_lung(r)/DA_ALI. All values between the maximum (100%) and half of the maximum (50%) of a size distribution are found within the HW range (Figures 3b and 4b). The value “mean” is at ¾ of the maximum. Ratio⁻¹ is the inverse DA-ratio meaning how many fold the dose at the ALI is higher than in the lung (“dose correction factor”).

Region	Half-width		Ratio DElung(r)/DEALI			Ratio DAlung(r)/DAALI
	Range [nm]		(Figure 3b)			(Figure 4b)			Ratio^−1
r	Lower	upper	100%	mean	50%	100%	mean	50%	100%	mean
ET	1.47	1075	1.88	1.41	0.938	9.85 × 10^−2	7.39 × 10^−2	4.93 × 10^−2	10.2	13.5
TB	39.8	468	41.4	31.1	20.7	5.89 × 10^−2	4.42 × 10^−2	2.95 × 10^−2	17.0	22.6
AL	42.8	429	135	101	67.3	8.47 × 10^−4	6.35 × 10^−4	4.23 × 10^−4	1180	1575
TL	41.8	438	177	133	88.6	1.11 × 10^−3	8.33 × 10^−4	5.55 × 10^−4	901	1201

Figure 4. (a) Deposition per surface area for the ALI and for the lung regions. Mean surface deposition in the ALl is calculated with Equation (5a(5a) $${DA}_{\mathit{ALI}}(d_p,{\rho }_p)=\frac{{DE}_{\mathit{ALI}}(d_p,{\rho }_p)}{A_{\mathit{ALI}}}$$(5a) ) as a function of particle size. Total and regional lung surface deposition is calculated with the HPLD model for mouth respiration using Equation (5b(5b) $${DA}_{\mathit{lung}}(r,d_p,{\rho }_p)=\frac{{DE}_{\mathit{lung}}(r,d_p,{\rho }_p)}{A_{\mathit{lung}}\left(r\right)}$$(5b) ) (see Tables 1–3). The ordinates on the right show the corresponding particle number and mass delivered per surface area at the ALI and in the lung regions. They are calculated with Equations (6b)(6b) $${TD}_{\mathit{ALI}}\left(d_p,{\rho }_p\right)=Q\mathrm{ }{t}_e\mathrm{ }\sum _{d_p}{C}_p\left(d_p,{\rho }_p\right)\mathrm{ }{DA}_{\mathit{ALI}}\left(d_p,{\rho }_p\right)\mathrm{ }$$(6b) and (6c)(6c) $${TD}_{\mathit{lung}}\left(r{,d}_p,{\rho }_p\right)=Q\mathrm{ }{t}_e\sum _r\sum _{d_p}{C}_p\left(d_p,{\rho }_p\right)\mathrm{ }{DA}_{\mathit{lung}}\left(r{,d}_p,{\rho }_p\right)\mathrm{ }$$(6c) , respectively. Ordinates represent simultaneously the TD for an exposure number concentration of 1 cm⁻³ and for an exposure mass concentration of 1 µg m⁻³. Thereby t_e is set to 1 h for both ALI and lung and Q to 100 cm³ min⁻¹ (Table 1b) for the ALI and to 0.54 m³ h⁻¹ for the lung, what results in a constant factor of 6 × 10³ and 540 × 10³, respectively. (b) Ratio DA_lung(r)/DA_ALI of the regional lung surface deposition and the ALI surface-deposition (Figure 4a). The horizontal bars show the half-width for the ET (green), TB (blue), AL (red) and TL (yellow) region (Table 6).

Figure 5. Particle transmission to the site of deposition as a function of particle size. ALI transmission is calculated (without any pre-impactor) from the ALI inlet to a well (see Table 1c and Figure 1a). Human inhalability follows the ICRP convention (Brown et al. 2013). Mouth and nose transmission is calculated with the HPLD model, showing the particle transfer to the beginning of the tracheo bronchial tract (TB, i > 2) and to the beginning of the alveolar space (AL, i > 19). The transmission efficiency for exhaled particles is added. The expression “mouth to TB” is used as a shortcut for “mouth breathing to enter the tracheo-bronchial region,” and “mouth to AL” as a shortcut for “mouth breathing to enter the alveolar region.”

Table 7. Particle transmission to the ALI and different lung regions for inhalation (see Figure 5). For comparison the HW of the exhaled particles is indicated.

Ranges for 50% Transmission Efficiency
Region	Range
r	lower	upper
ALI	1.10	7.373	nm
ICRP	–	>50.000	nm
mouth to TB	1.56	11.731	nm
nose to TB	2.47	3.557	nm
mouth to AL	9.44	7.443	nm
nose to AL	10.5	3.356	nm
mouth exhaled	42.7	2.894	nm
nose exhaled	44.0	2.280	nm

Figure 6. Surface-delivered particle mass (a) and cell-delivered particle number (b) at the ALI and in the lung regions. The airborne exposure concentration is 1 mg m⁻³ in (a) and 10⁶ cm⁻³ in (b). A lognormal emission distribution is applied (see Table 5). Part (b) compares the load for cells in different lung regions. The corresponding cell counts (cells per cm²) are indicated in the columns. For the ALI, the size of A549 cells is assumed, for ET and TB region the cell size is assumed to be identical with type-II pneumocytes. For the AL region the size of type-I pneumocytes and for TL the weighted average of type-I and type-II cells is assumed (Table 4).

Table 8. Deposition in the lung regions of an adult man calculated with the HPLD model (Table 2) compared to lung deposition data published by the ICRP, Annexe F (1994). Data are for a polydisperse aerosol with a mean particle diameter of 100 nm, a GSD of 1.5, a particle density of 3 g cm⁻³ and a shape factor of 1.5.

	Breathing path	Q [m³ h^−1]	ET [-]	TB [-]	AL [-]	TL [-]
Figure 3a, HPLD, dp= 100 nm	Nose	0.54	0.032	0.069	0.24	0.34	Annexe F page
ICRP, Annexe F, dp = 100 nm	Nose	0.54	0.052	0.077	0.22	0.35	422
	Nose	0.45	0.052	0.082	0.20	0.33	418
	Nose	1.2	0.064	0.055	0.21	0.33	416
	Mouth	1.2	0.036	0.066	0.21	0.30	416
	Nose	1.5	0.066	0.051	0.21	0.32	426
	Nose	1.7	0.061	0.048	0.20	0.31	417
	Mouth	1.7	0.033	0.049	0.21	0.29	417
	Mouth	3.0	0.044	0.040	0.20	0.28	430

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