Research Paper Volume 13, Issue 15 pp 19460—19474

Pedunculoside protects against LPS-induced mastitis in mice by inhibiting inflammation and maintaining the integrity of blood-milk barrier

The effect of PE on tight junction proteins occludin and claudin-3. PE was given orally for 7 days. The fourth pair of milk ducts in mice were injected with LPS for 24 h. The mice were killed by dislocation and fixed on the operating platform after LPS injection 24 h. The midline of abdomen was cut to collect mammary gland. PE was added to the cell culture medium at a concentration of 10 μM. After 1 h, LPS was added to the culture medium at a concentration of 1 μg/mL. The co-stimulation time was 12 h. (NP40) was added to mice mammary gland and mMECs, and then Western blot samples were prepared to obtain protein bands., Immunofluorescence and Western blot methods were used to evaluate the expression changes of tight junction proteins occludin and claudin-3 in mammary gland and mMECs. Immunofluorescence of occludin (A) and claudin-3 (B) in frozen sections of mammary gland. Scale bar 100 μM. (C, E, F) Western blot assay of occludin and claudin-3 in mammary gland; (D, G, H) Western blot assay of occludin and claudin-3 in mMECs. Each immunoreactive band was digitized and expressed as a ratio of the β-actin level. Values are presented as means ± SEM, three independent repeated experiments were performed; #p**p  0.01 vs. LPS group.

Figure 7. The effect of PE on tight junction proteins occludin and claudin-3. PE was given orally for 7 days. The fourth pair of milk ducts in mice were injected with LPS for 24 h. The mice were killed by dislocation and fixed on the operating platform after LPS injection 24 h. The midline of abdomen was cut to collect mammary gland. PE was added to the cell culture medium at a concentration of 10 μM. After 1 h, LPS was added to the culture medium at a concentration of 1 μg/mL. The co-stimulation time was 12 h. (NP40) was added to mice mammary gland and mMECs, and then Western blot samples were prepared to obtain protein bands., Immunofluorescence and Western blot methods were used to evaluate the expression changes of tight junction proteins occludin and claudin-3 in mammary gland and mMECs. Immunofluorescence of occludin (A) and claudin-3 (B) in frozen sections of mammary gland. Scale bar 100 μM. (C, E, F) Western blot assay of occludin and claudin-3 in mammary gland; (D, G, H) Western blot assay of occludin and claudin-3 in mMECs. Each immunoreactive band was digitized and expressed as a ratio of the β-actin level. Values are presented as means ± SEM, three independent repeated experiments were performed; #p<0.01 vs. NT group; **p < 0.01 vs. LPS group.