Activation of nuclear factor kappa B in mammary epithelium promotes milk loss during mammary development and infection

Abstract

We investigated whether Nuclear Factor-kappa B (NF-κB), which exhibits a regulated pattern of activity during murine mammary gland development, plays an important role during lactation and involution, when milk production ceases and the gland undergoes apoptosis and remodeling. We generated a doxycycline inducible transgenic mouse model to activate NF-κB specifically in the mammary epithelium through expression of a constitutively active form of IKK2, the upstream kinase in the classical NF-κB signaling cascade. We found that activation of NF-κB during involution resulted in a more rapid reduction in milk levels and increased cleavage of caspase-3, an indicator of apoptosis. We also found that activation of NF-κB during lactation with no additional involution signals had a similar effect. The observation that NF-κB is a key regulator of milk production led us to investigate the role of NF-κB during mastitis, an infection of the mammary gland in which milk loss is observed. Mammary gland injection of E. coli LPS resulted in activation of NF-κB and milk loss during lactation. This milk loss was decreased by selective inhibition of NF-κB in mammary epithelium. Together, our data reveal that activation of NF-κB leads to milk clearance in the lactating mammary gland. Therefore, targeting of NF-κB signaling may prove therapeutic during mastitis in humans and could be beneficial for the dairy industry, where such infections have a major economic impact.

Intraductal Injection of LPS as a Mouse Model of Mastitis: Signaling Visualized via an NF-κB Reporter Transgenic

“…Among many findings, these studies illustrate that the inflammatory response is modulated by interactions with CD14, and that it results in a neutrophil influx regulated by mammary alveolar macrophages. As suggested in both our previously published work 4 , as well as that of others 2 this model of mastitis may be further used to study mechanisms and therapeutics relevant in humans, where as many as 3-33% of breastfeeding women suffer from this painful infection (percentage varies based on study methodology) 15 . It may also prove a useful research tool for those in the dairy industry, where mastitis is a costly disease leading to significant losses in milk production each year 16 .…”

“…This method was successful and allowed us to study the inflammation associated with LPS injection without any additional effects overlaid by the process of injection. In addition, this method also utilized an NF-κB luciferase reporter transgenic mouse and bioluminescent imaging technology to visually and quantitatively show increased NF-κB signaling within the LPS-injected gland 4 .These methods are of interest to researchers of many disciplines who wish to model disease within the lactating mammary gland, as ultimately, the technique described here could be utilized for injection of a number of substances, and is not limited to only LPS. …”

“…This method was successful and allowed us to study the inflammation associated with LPS injection without any additional effects overlaid by the process of injection. In addition, this method also utilized an NF-κB luciferase reporter transgenic mouse and bioluminescent imaging technology to visually and quantitatively show increased NF-κB signaling within the LPS-injected gland 4 .…”

Nucling, a Novel Apoptosis-associated Protein, Controls Mammary Gland Involution by Regulating NF-κB and STAT3

“…The loss of NF-B signaling has been shown to result in a decrease in caspase-3 cleavage and delayed involution (11). In contrast, a previous study demonstrated that constitutively active IKK-␤ increased NF-B activation, leading to a reduction in milk protein levels and increases the level of apoptosis during involution (10).…”

“…However, the activation of STAT3 is insufficient to induce involution in the absence of NF-B signaling. During mammary gland development, the activation of NF-B increases during pregnancy, decreases during lactation, and then increases again after weaning (9,10). The loss of NF-B signaling has been shown to result in a decrease in caspase-3 cleavage and delayed involution (11).…”