## 3 Results
### Effect of Doxorubicin-Mscari comosum Extract Treatment on the UPR Pathway
No significant changes in the expression of microtubule-associated protein 1A/1B-light chain 3 (LC3) were observed following treatment with doxorubicin alone or in combination with MET70 (Figure 7). However, while doxorubicin alone increased the expression of Beclin-1 (BECN1), the combination treatment attenuated this effect, suggesting that inhibition of autophagy may contribute to the enhanced cytotoxicity of doxorubicin. Modulation of ABC Transporters Expression by Combined Doxorubicin and Muscari comosum Extract (MET70) Treatment  
Multidrug resistance (MDR) remains a major obstacle in cancer therapy, often mediated by the efflux of chemotherapeutic agents through ATP-binding cassette (ABC) transporters. Among the most relevant transporters are ABCB1, ABCC1, and ABCG2. In this study,  
Figure 7: Effect of doxorubicin–MET70 combined treatment on the mRNA expression of genes involved in autophagy in HepG2 cells. Cells were treated with doxorubicin (0.5 uM) and MET70 (100 ug/mL), either alone or in combination, for 72 h. DMSO (0.02%) was used as the vehicle control. Relative mRNA expression levels of BECN1 and LC3 analyzed by qRT-PCR were calculated using the 2\({}^{-\Delta\text{CI}}\) method, with \(\beta\)-actin as the endogenous reference gene. Data are presented as mean \(\pm\) SEM from at least three independent experiments. Statistical significance was determined using one-way ANOVA followed by Holm–Sidak correction for multiple comparisons (\({}^{*}\)_p_ < 0.05; \({}^{***}\)_p_ < 0.001). we investigated the modulatory effects of the combined treatment with doxorubicin and _Muscari comosum_ extract (MET70) on the expression of these transporters. RT-qPCR and Western blot analyses demonstrated that doxorubicin treatment alone significantly reduced both mRNA and protein levels of ABCB1 and ABCG2 compared to control cells, while ABCC1 expression remained unchanged (Figure 8a,b). Interestingly, compared to doxorubicin alone, the combination with MET70 did not further alter ABCB1 expression, but significantly decreased ABCC1 levels and increased ABCG2 expression. Figure 8: Effect of doxorubicin-MET70 combined treatment on the mRNA expression (**a**) and protein levels (**b**) of key ABC transporters involved in multidrug resistance in HepG2 cells. Cells were treated with doxorubicin (0.5 \(\upmu\)M) and MET70 (100 \(\upmu\)g/mL), either alone or in combination, for 72 h. DMSO  
(0.02%) was used as the vehicle control. (**a**) Relative mRNA expression levels of ABCB1, ABCC1 and ABCG2, analyzed by qRT-PCR were calculated using the \(2^{-\Delta\text{Ct}}\) method, with \(\beta\)-actin as the endogenous reference gene. Data are presented as mean \(\pm\) SEM from at least three independent experiments. (**b**) Protein expression levels of ABC transporters assessed by Western blot analysis. Representative blots and densitometric quantifications are shown, with protein levels normalized to \(\beta\)-actin and expressed relative to control (set to 100%). Data are presented as mean \(\pm\) SEM from at least three independent experiments. Statistical significance was determined by one-way ANOVA followed by Holm-Sidak correction for multiple comparisons (* \(p<0.05\); ** \(p<0.01\); *** \(p<0.001\)). These findings suggest that MET70 may differentially modulate ABC transporter expression, potentially contributing to the sensitization of HepG2 cells to doxorubicin.