Copper sulfate cuso4

Gold (III) chloride trihydrate (HAuCl₄·3H₂O, Cat. No. 520918, Sigma-Aldrich Co., St. Louis, MO, USA), sodium borohydride (NaBH₄, Cat. No. 213462, Sigma-Aldrich Co., St. Louis, MO, USA), copper sulfate (CuSO₄, Cat. No. 451657, Sigma-Aldrich Co., St. Louis, MO, USA), l-ascorbic acid (C₆H₈O₆, Cat. No. A5960-25G, Sigma-Aldrich Co., St. Louis, MO, USA), cetyltrimethylammonium bromide (CTAB, Cat. No. 52370-500G, Sigma-Aldrich Co., St. Louis, MO, USA), palladium (II) chloride (PdCl₂, Cat. No. 323373-1G, Sigma-Aldrich Co., St. Louis, MO, USA), hydrochloric acid (HCl, Cat. No. 84415-500ML, Sigma-Aldrich Co., St. Louis, MO, USA), and sodium tetrachloropalladate (II) (Na₂PdCl₄, Cat. No. 205818-1G, Sigma-Aldrich Co., St. Louis, MO, USA) were used as received without further purification. All aqueous solutions were prepared using deionized water with a resistivity of 18.2 MΩ cm.

Spherical Ti64 powders with a size range of 15–53 µm, supplied by Oerlikon company (Switzerland), were used to fabricate Ti64 catalyst supports. Colloidal palladium activator, sodium chloride (NaCl, purity ≥99.5%), and hydrochloric acid (HCl, purity: 36–38 wt%) were manufactured by Jixin Chemical Technology Co. Ltd. (Guangzhou, China), Xilong Science Co., Ltd (Shantou, China), and Dongjiang Chemical Reagent Co., Ltd (Dongguan, China), respectively. Copper sulfate (CuSO₄, purity ≥99%) and edetate disodium (EDTA 2Na, purity ≥99.5%) were manufactured by Sigma Aldrich (America) and Dibai Biological Technology Co., Ltd (Shanghai, China), respectively. Sodium hydroxide (NaOH, purity: 99.9%), Potassium ferrocyanide trihydrate (K₄FeC₆N₆ 3H₂O, purity: 98%), Potassium sodium tartrate tetrahydrate (C₄H₄O₆KNa 4H₂O, purity: 99%), formaldehyde solution (HCHO, purity: 37 wt%), ammonium persulfate ((NH₄)₂S₂O₈, purity ≥98%), cobalt sulfate heptahydrate (CoSO₄ 7H₂O, purity ≥99%), nickel sulfate hexahydrate (NiSO₄ 6H₂O, purity: 99.9%), and CO(NH₂)₂ (purity ≥99.5%) were acquired from Aladdin Chemical Co., Ltd (Shanghai, China). Deionized (DI) water was acquired utilizing a water purification system (Mini‐Q Water). All chemical reagents were directly utilized without any purification.

The standard compounds, artemetin (1), casticin (2), hesperidin (3), luteolin (4), vitexin (5), and vanillic acid (6), were isolated from the extract of V. rotundifolia fruits by emeritus professor Sam Sik Kang from Seoul National University. The compounds were identified by NMR and their purities were assessed by NMR and TLC before use for this study. TrichloroAcetic acid (TCA), 1,1,3,3-tetramethoxypropane, copper sulfate (CuSO₄), bovine serum albumin (BSA), and dialysis tubing cellulose membrane were purchased from Sigma Aldrich (St. Louis, MO, USA). Coomassie brilliant blue R-250 and guanidine hydrochloride were obtained from Tokyo Chemical Industry (Tokyo, Japan). 2,4-Dinitrophenylhydrazine (DNPH) was purchased from Junsei Chemical Co., Ltd (Tokyo, Japan). Acetic acid and sodium chloride were purchased from Merck (Darmsstadt, Germany).

For actomyosin inhibition, blebbistatin (Selleck Chemical) and Y-27632 (Selleck Chemical) were used at 100 μM concentration and added 1h before stimulation. For aquaporin 3 (AQP3) inhibition, copper sulfate (CuSO4, Sigma Aldrich) was used at 12.5μM concentration and added 24h before stimulation. For Na+/K+ ATPase inhibition, ouabain octahydrate (Sigma Aldrich) was used at 100nM and added 24h before stimulation. For CFTR channel inhibition, CFTR(inh)-172 (Cayman) was used at 50μM and 100μM concentration and added 24h before stimulation for MDCK cysts and 48h before stimulation for ISC organoids. For NKCC inhibition, bumetanide was used at 100μM concentration and added 1h before stimulation. For PI3K inhibition, LY294002 (Medchemexpress) was used at 25 μM to stall electrotaxis at and 50 μM to reverse electrotaxis for both cysts and monolayers. The LY294002 treated samples were incubated at 37 °C for 60 min before being assembled into the bioreactor. Media with the same concentration of LY294002 was used for perfusion. Inhibitors were applied after the patterning stencils were removed to prevent the inhibitors from being absorbed into the PDMS. Identical concentration of inhibitors was supplemented in the perfusion media to prevent the inhibitors from washing out during stimulation.

Copper sulfate (CuSO₄) was supplied by Sigma Chemical Co., St. Louis, USA. Chrysin (5,7-dihydroxyflavone) was obtained from Alfa Aesar Thermo Fisher Scientific Co., Germany. All other chemicals used in the study were of analytical reagent grade.