Fig. 5.

Treatments that prevent disulphide accumulation or pharmacological SLC7A11 inhibition rescued the cell death induced by TXNRD1 inhibitors in pre-OCs. NFATc1 knockdown in pre-OCs attenuated TXNRD1 inhibitor-induced cystine accumulation and decreased the sensitivity to TXNRD1 inhibitors. SLC7A11 knockout eliminated the difference in sensitivity of BMDMs and pre-OCs to TXNRD1 inhibitors, and overexpression of SLC7A11 or NFATc1 in BMDMs increased cell sensitivity to TXNRD1 inhibitors

a, the HPLC-MS/MS results of intracellular cystine concentrations in BMDMs, BMDMs treated with TXNRD1 inhibitors, pre-OCs, and pre-OCs treated with TXNRD1 inhibitors. n = 3 per group. b, the viability of pre-OCs was measured after 24 h of incubation with a medium containing 2 μM AF or 5 μM TRi-1 with or without 5 mM NAC. n = 3 per group. c, the HPLC-MS/MS results of intracellular cystine concentrations in pre-OCs treated with TXNRD1 inhibitors and pre-OCs treated with both TXNRD1 inhibitors and NAC. n = 3 per group. d, the viability of BMDMs and pre-OCs was measured after 24 h of incubation with a medium containing 2 μM AF or 5 μM TRi-1 with or without 500 μM l-Penicillamine (L-Pen). n = 3 per group. e, the HPLC-MS/MS results of intracellular cystine concentrations in pre-OCs treated with TXNRD1 inhibitors and pre-OCs treated with both TXNRD1 inhibitors and L-pen. n = 3 per group. f, the viability of BMDMs and pre-OCs was measured after 24 h of incubation with a medium containing 2 μM AF or 5 μM TRi-1 with or without 2 mM 2-Mercaptoethanol (2 ME). n = 3 per group. g, The HPLC-MS/MS results of intracellular cystine concentrations in pre-OCs treated with TXNRD1 inhibitors and pre-OCs treated with both TXNRD1 inhibitors and 2 ME. n = 3 per group. h, the viability of BMDMs and pre-OCs was measured after 24 h of incubation with medium containing 2 μM AF or 5 μM TRi-1 with or without Sulfasalazine (SSZ) (low dose 100 μM, high dose 400 μM). n = 3 per group. i, the HPLC-MS/MS results of intracellular cystine concentrations in pre-OCs treated with TXNRD1 inhibitors and pre-OCs treated with both TXNRD1 inhibitors and SSZ. n = 3 per group. (j and k), the viability of siCTL and siNFATc1 pre-OCs was measured after 24 h of incubation with AF (j) or TRi-1 (k). n = 3 per group. l, The HPLC-MS/MS results of intracellular cystine concentrations in siCTL and siNFATc1 pre-OCs treated with TXNRD1 inhibitors, n = 3 per group (m and n), the viability of SLC7A11 KO BMDMs and pre-OCs was measured after 24 h of incubation with AF (m) or TRi-1 (n). n = 3 per group (o and p), western blot analysis of SLC7A11 protein levels in EV and SLC7A11 overexpression BMDMs. n = 3 per group. q, RT-qPCR analysis of Slc7a11 in EV and SLC7A11 overexpression BMDMs. n = 3 per group. (r and s), the viability of EV, NFATc1 overexpression, and SLC7A11 overexpression BMDMs was measured after 24 h of incubation with AF (r) or TRi-1 (s). n = 3 per group. All data in this figure are represented as mean ± SD. ns, no significance, **P < 0.01, ***P < 0.001, ****P < 0.0001.