Fig. 4

Artemisinin derivatives induce ROS-dependent DNA damage and oxidative stress in Theileria-infected cells: A Graph shows the relative fluorescence intensity of intracellular ROS produced in T. annulata-infected cells after treatment with H₂0₂ and different concentrations of NALC compared to untreated cells (Control). The error bars reflect mean plus/minus standard deviations. B The graph depicts the relative fluorescence intensity of intracellular ROS generated in T. annulata infected cells following treatment with various concentrations of ARS + BPQ and DHART + BPQ compared to untreated cells (Control) with or without NALC. The error bars reflect mean plus/minus standard deviations. C Comet assay visualisation of DNA damage in T. annulata-infected cells after 120 min of treatment with ARS + BPQ and DHART + BPQ, as compared to untreated parasites with or without NALC. D Comet assay evaluation of the olive tail moment (OTM) after 120 min of treatment with ARS + BPQ and DHART + BPQ, compared to untreated cells (Control) with or without NALC. Ordinary 1way ANOVA was performed to check the statistical significance and Tukey’s multiple comparisons test was used for the analysis. The mean of each group was compared with the control as well as with each other and the graph represent standard error mean. In the graph ****represents significance of (P < 0.0001) while ns represents nonsignificant results with P value > 0.05 while *represents significance of (P ≤ 0.05). The error bars reflect mean plus/minus standard deviations. E Immunofluorescence staining of 8-OHdG after 24 h of treatment with DHART; untreated and BPQ treated cells were used as a positive control. DAPI was used for nuclear staining. F The graph depicts the levels of 8-OHdG in T. annulata infected cells after 24 h of DHART treatment as measured by fluorescence intensity using the appropriate ZEN 3.3 program. G Immunofluorescence staining of 8-OHdG after 24 h and 48 h treatment with ARS at different concentrations (1× and 5×); untreated and BPQ treated cells were used as a positive control. DAPI was used for nuclear staining. H The graph depicts the levels of 8-OHdG in T. annulata infected cells after 24 h and 48 h treatment with ARS as measured by fluorescence intensity using the appropriate ZEN 3.3 program. A minimum of forty cells were counted for each experiment and measurement point. The error bars reflect mean plus/minus standard deviations. Ordinary 1way ANOVA was performed to check the statistical significance, and Dunnett's multiple comparisons test was used for the analysis in F, H. The mean of each group was compared with the control and each other, and the graph represents the standard error mean. In the F graph, ***represents the significance of (P = 0.0009) while ns represents nonsignificant results with a P value > 0.05. In H, ***represents the significance of (P ≤ 0.001), and ****represents the significance of (P < 0.0001)