Fig. 1: Silicon nanopore membranes (SNM) prevent diffusion of cytokines in vitro.
From: Feasibility of an implantable bioreactor for renal cell therapy using silicon nanopore membranes
a In vitro setup where human renal epithelial cells (HRECs) were cultured in apical and basal compartments of a 6-well plate, separated by an SNM bonded to a Transwell® membrane insert (blue). TNF-alpha (TNF-a) (300 ng/mL) was added to the apical compartment only and incubated for 6 h. b TNF-a concentration in the apical and basal compartments after 6 h, with and without HREC. There was no statistically significant change in TNF-a concentration within the apical compartment after 6 h [290.81 ± 8.51 and 289.72 ± 15.01 (mean ± standard deviation)]. The TNF-a concentration on the basal side of the SNM was 12.38 ± 7.05 (mean ± standard deviation) after 6 h, suggesting little to no passage of TNF-a through the SNM. In the presence of cells, basal concentration after 6 h was 45.16 ± 2.57 (mean ± standard deviation) (n = 3 independent experiments). c LIVE/DEAD™ imaging of apical and basal HREC using calcein-AM (green) and ethidium homodimer-1 (red) demonstrated more dead cells in the apical compartment after 6 h, relative to the 0-h time point and relative to the basal compartment. Cells exposed to TNF-a in the apical compartment demonstrated 44.2 ± 4.3% viability (mean ± standard deviation) after 6 h. Cells on the basal side of the SNM demonstrated 88.6 ± 2.1% viability (mean ± standard deviation) after 6 h (n = 3 independent experiments). d Tight junction and cell monolayer integrity were evaluated by ZO-1 expression (green) and DAPI nuclear staining (blue). Cells on the basal compartment maintained ZO-1 expression after 6 h. Cells directly exposed to TNF-a in the apical compartment had fewer cells (indicated by DAPI) with less defined ZO-1 expression relative to cells at 0 h. Note: apical cells were cultured directly on SNM, which attenuated immunofluorescence imaging. Nonetheless, the difference in DAPI and ZO-1 expression between 0 and 6-h time points remains evident (n = 3 independent experiments). Source data are provided as a Source Data file.
