The current study was conducted to evaluate the effect of different levels of milk thistle ( Silybum marianum ) meal on performance, egg quality, blood biochemistry parameters, ileal bacterial enumeration, intestinal histology, immunological responses, antioxidant indices of blood and liver, and fatty liver indices in laying hen fed diets with different levels of metabolizable energy. In this study, 160 hen layers were randomly distributed among 8 experimental of treatment with 4 replicates of 5 birds each. Dietary treatment consisted of four levels of milk thistle meal (0, 1.5, 3 and 6%) and two levels of metabolizable energy (2650 and 2950 kcal/kg) that fed during a 80 d feeding trial including 10 days for adaptation period and 70 days as main recording period. Performance parameters including feed intake, feed conversion ratio, egg production percentage, egg weight, egg mass and also egg quality were calculated on d 35, and 70 of experiment. To investigate of immunological response, sheep red blood cells (SRBC) were injected in two times with 10 d interval; thereby, hens were bled 6 and 12 d after the first and second injection, respectively. Moreover, Newcastle disease virus was injected and then sera were taken 6 d after injection. In addition, differential leukocyte counts were evaluated at the end of experiment. At the end of experiment, two birds of each cage were exsanguinated to evaluate the relative weight of liver, spleen as well as pancreas, and also to sample ileal content and jejunal morphology. The results showed that dietary inclusion of high level of metabolizable energy led to an improvement in feed conversion ratio (P 0.01), villus height (P 0.01), antibody titer against SRBC (P 0.05) and an increase in triglyceride, plasma (P 0.01) and liver (P=0.1) malondialdehyde concentrations, hepatic fat (P 0.05) percentage and enzyme activities of SGPT (P 0.05) and ALP (P=0.1). Supplementation of 1.5 % of milk thistle meal led to an improvement in systemic immunity responses such as an increase in primary and secondary antibody titer against SRBC (P 0.05), a decrease in heterophil to lymphocyte ratio (P 0.05) and an increase in Kupffer cells population (P 0.01). The lowest plasma cholesterol concentration (P=0.1), hepatic enzyme (P 0.05) activities and yolk triglyceride concentration (P 0.01) were observed in hens fed on the highest levels (6%) of milk thistle meal. Dietary supplementation of 3% milk thistle meal resulted in the best feed conversion ratio (P 0.01) via a decrease in ileal Escherichia coli (P 0.01) and an improvement in villus height to crypt depth ratio (P 0.05). Among of egg quality parameters, only eggshell hardness and thickness were affected; so, inclusion of 3 and 6% milk thistle meal had the best eggshell hardness and thickness at the end of experiment (P 0.05). The interaction between different levels of milk thistle meal and metabolizable energy on relative weight of liver, villus height and plasma malondialdehyde concentration were significant; so that, with the lowest levels of energy, dietary addition of 1.5% milk thistle meal caused a decrease in relative weight of liver (P 0.01), an increase in villus height (P 0.01) and a decrease in plasma malondialdehyde concentration (P 0.05). With highest levels of energy, administration of 3 and 6% levels of drug meal caused similar responses. In general, results indicated that while highest levels of metabolizable energy led to an improvement in performance and immunological responses, it also caused fatty liver. However, dietary inclusion of 3 and 6% milk thistle meal resulted in suppression in fatty liver indices and best performance responses. Keywords : Milk thistle meal, Metabolizable energy, Laying hens, Immunological responses, Fatty liver, Antioxidant activity