immunoflu orescence analysis confirmed that ifn - γ treatment led to a marked accumulation of p - stat1 in the nucleus, reinforcing the notion that stat1 plays a crucial role in ifn - γ - mediated transcriptional regulation. these findings align with studies showing that sustained stat1 activation promotes an immunosupp ressive tumor microenvironment by inducing pd - l1 expression and other immune - regulatory genes. the upregulation of pd - l1 in response to ifn - γ was confirmed at both the mrna and protein levels, as demonstrated by qrt - pcr and western blotting. the increased surface expression of pd - l1 following ifn - γ treatment highlights the functional significance of this regulation, as surface pd - l1 interacts with pd - 1 on t cells to inhibit anti-tumor immune responses ( arak et al. 2021 ). these results are in accordance with previous reports showing that ifn - γ is one of the most potent inducers of pd - l1 in nsclc, facilitating immune escape and tumor progression ( pawelczyk et al. 2019 ). additionally, the use of fludarabine, a stat1 inhibitor, significantly attenuated ifn - γ - induced pd - l1 expression, confirming that stat1 is the primary mediator of this regulatory axis. this finding corroborates prior studies demonstrating that stat1 - deficient cells fail to upregulate pd - l1 in response to ifn - γ, emphasizing the centrality of stat1 in this pathway. one of the most significant findings of this study is the ability of curcumin to inhibit ifn - γ - induced stat1 activation and pd - l1 expression in a549 cells. western blot analysis revealed that curcumin suppressed stat1 phosphorylation in a dose - dependent manner, with the highest concentration ( 50 µm ) reducing phosphorylation by 68 %. this effect was not due to a decrease in total stat1 protein levels, indicating that curcumin selectively inhibits stat1 activation rather than its expression. previous studies have reported that curcumin can interfere with jak - stat signaling in other cancer types. curcumin directly inhibits the phosphorylation of stat3, a key component of the jak - stat signaling pathway in breast cancer ( golmohammadi et al. 2024 ) and prostate cancer ( li et al. 2024 ), as well as the downregulation of the stat1 in melanoma ( xu et al. 2018 ), but its specific effect on stat1 in ifn - γ - stimulated nsclc cells had not been previously explored. our findings extend these observations by demonstrating that curcumin effectively blocks stat1 activation in lung cancer cells, preventing the downstream induction of pd - l1. the suppression of pd - l1 expression by curcumin was observed at both the transcriptional and translational levels, as evidenced by qrt - pcr and western blotting. this finding is particularly relevant in the context of nsclc, where high pd - l1 expression correlates with poor prognosis and resistance to immunotherap y. previous studies have reported that curcumin downregulates pd - l1 in other cancer models, such as melanoma ( xu et al. 2018 ) and hepatocellular carcinoma ( guo et al. 2021 ), but the specific inhibition of ifn - γ - induced pd - l1 expression in nsclc had not been thoroughly investigated. our study provides the first evidence that curcumin can effectively suppress ifn - γ - mediated pd - l1 upregulation in lung cancer cells via stat1 pathway, highlighting its potential as an immune - modulatory agent. in addition to pd - l1, stat1 regulates the expression of multiple isgs involved in immune evasion, including socs1 ( ilangumaran et al. 2024 ) and isg15 ( desai 2015 ). our results demonstrated that ifn - γ significantly upregulated both socs1 and isg15, reinforcing the notion that ifn - γ signaling contributes to an immunosupp ressive tumor microenvironment. curcumin pretreatment, however, led to a significant reduction in both socs1 and isg15 expression, further supporting its ability to interfere with ifn - γ - driven stat1 signaling. socs1 is known to act as a feedback inhibitor of jak - stat signaling ( liau et al. 2018 ), but paradoxically, its overexpression in tumors has been associated with immune escape mechanisms. by suppressing socs1 expression, curcumin may enhance the responsiveness of tumor cells to immune-mediated clearance. similarly, isg15 has been implicated in tumor progression and resistance to therapy ( meng et al. 2024 ), suggesting that its downregulation by curcumin may have additional therapeutic benefits. finally, we observed that curcumin enhances the anti - proliferative effect of ifn - γ in a549 cells. while ifn - γ alone resulted in a modest reduction in cell viability ( 21 % ), the combination of ifn - γ and curcumin led to a significantly greater reduction ( 47 % ), suggesting a synergistic effect. these findings align with previous reports that curcumin enhances the anti-tumor activity of cytokines by modulating cell cycle regulators and apoptotic pathways ( hu et al. 2018 ). the precise mechanism by which curcumin sensitizes nsclc cells to ifn - γ - induced growth suppression remains to be elucidated, but it may involve inhibition of survival pathways downstream of stat1 activation. given that stat1 has been implicated in both pro - apoptotic and pro - survival signaling, the net effect of its inhibition may depend on the cellular context and additional regulatory factors. in conclusion, our study provides novel evidence that ifn - γ induces pd - l1 expression in nsclc cells via stat1 activation and that curcumin effectively inhibits this process by suppressing stat1 phosphorylation and nuclear translocation. furthermore, curcumin downregulates ifn - γ - induced isgs and enhances ifn - γ - mediated tumor cell growth suppression, highlighting its potential as a therapeutic adjuvant in nsclc. these findings suggest that curcumin could be used to improve the efficacy of immune checkpoint inhibitors by reducing tumor immune evasion. future studies should focus on elucidating the precise molecular mechanisms underlying curcumin ’ s effects on stat1 signaling and investigating its potential synergistic effects with existing immunotherap ies in preclinical and clinical settings. materials and methods statistical analysis all experiments were performed at least three times. a one - way anova followed by a tukey ' s post - hoc test was used to compare the rt - qpcr signals from the treated cells and the elisa results. for comparison of rt - qpcr signals from treated cells to the hypothetical value of 1 ( normalized to the signal of control cells ), a one - sample t - test was used, as in this type of statistical analysis, the variability of the values obtained from control cell signals is taken into consideration, although they appear without standard deviations ( sd = 0 ). p # x003c; 0. 05 was considered to indicate a statistically significant difference. all statistical analyses were performed in graphpad prism version 9 ( graphpad software, inc. ); means and standard deviations are provided as numbers or as scatter plots. results discussion confirming previously published data, it was shown that the human cell line mio - m1, an accepted model of human mueller cells, expresses agt, ace, ace2, angiotensin ii receptors at1 and at2, as well as the receptor of angiotensin ( 1 - 7 ) mas1 ( 17 - 19, 21 - 24, 28 ). the mrna expression levels of these were largely unchanged when cells were exposed to hyperglycem ic or hypoxic conditions or both. exposure of cells to these conditions seemed to not result in induction of cellular stress, which may adversely affect the outcome of the investigations, since expression of il - 6 mrna, a marker of cellular stress, remained stable. it could be suggested that an incubation time of 6 h is too short, but due to the short half - life of angiotensin ii, longer exposure times would likely not result in more relevant data.