Seipin deficiency leads to increased endoplasmic reticulum stress and apoptosis in mammary gland alveolar epithelial cells during lactation†

Abstract

Seipin is an integral endoplasmic reticulum (ER) membrane protein encoded by Berardinelli-Seip congenital lipodystrophy type 2 (BSCL2/Bscl2) gene. Most litters (59%) from Bscl2-/- dams mated with wild type (WT) (Bscl2+/+) males did not survive postnatal day 5 (PND5) and pups (Bscl2+/-) lacked milk in their stomachs. The survived litters had reduced pup survival rate at PND21. It was hypothesized that seipin was critical for lactation. Bscl2 was upregulated and highly detected in the lactation day 1 (LD1) WT mammary gland alveolar epithelial cells. LD1 Bscl2-/- mammary glands lacked adipocytes and alveolar clusters and had varied alveolar morphology: from interconnected mammary gland alveoli with dilated lumen and sloughed epithelial cells to undifferentiated mammary gland alveoli with unexpanded lumen. Comparable levels of whey acidic protein (WAP, a major component in rodent milk) staining and Nile Red lipid droplet staining between WT and Bscl2-/- LD1 alveolar epithelial cells indicated normal milk protein synthesis and lipid syntheses in LD1 Bscl2-/- mammary glands. Significantly reduced percentage of larger lipid droplets was detected in LD1 Bscl2-/- alveoli with unexpanded lumen. There was no obviously impaired proliferation detected by PCNA staining but increased apoptosis detected by cleaved caspase-3 staining in LD1 Bscl2-/- alveolar epithelial cells. Increased expression of protein disulfide isomerase and binding immunoglobulin protein in the LD1 Bscl2-/- mammary gland alveolar epithelial cells indicated increased ER stress. This study demonstrates increased ER stress and apoptosis in LD1 Bscl2-/- mammary gland alveolar epithelial cells and reveals a novel in vivo function of seipin in lactation.

Exploring Seipin: From Biochemistry to Bioinformatics Predictions

“…In contrast, Liu et al observed ER stress markers positively induced in adipose-specific seipin-knockout mice [ 74 ]. Zowalaty et al also showed ER stress and increased apoptosis in mammary gland alveolar epithelial cells with BSCL2 gene deletion and revealed the importance of seipin for lactation [ 87 ]. BSCL2 knockout was also associated with intensified cerebral ER stress after a transient middle cerebral artery occlusion situation in another study [ 88 ].…”

Effects of Peptidoglycan, Lipoteichoic Acid and Lipopolysaccharide on Inflammation, Proliferation and Milk Fat Synthesis in Bovine Mammary Epithelial Cells

“…Triglyceride (TG) accounted for more than 98% of milk lipid [ 6 ]. Fatty acids can be rapidly taken up and converted into lipid droplets by the lactating mammary gland [ 7 ]. Non-esterified fatty acid (NEFA) is a source of fatty acids and can increase milk fat synthesis [ 8 ].…”

Dietary exposure to mycotoxin zearalenone (ZEA) during post-implantation adversely affects placental development in mice

“…The middle three sections with the largest areas were selected for area quantification. The total area of each placental section and the areas of labyrinth layer, junctional zone, and decidua, which were outlined manually and quantified using Image J (National Institutes of Health, Bethesda, MD, USA) [35][36][37][38]. The average of each area from three sections in the same placenta was counted as one data point for statistical analysis.…”

“…Cross sections (10 μm) of the middle part of frozen placentas from 0 ppm and 40 ppm ZEAtreated groups were collected on the same slides for immunofluorescence. The sections were fixed in 4% paraformaldehyde for 15 min, and blocked with 3% hydrogen peroxide in methanol for 10 min at 25°C, and 10% goat serum in 1x phosphate buffered saline (PBS) for 1 hour at 25°C, incubated with rabbit anti-proliferating cell nuclear antigen (PCNA (D3H8P)1:250, 13110XP, Cell Signaling Technology, Danvers, MA, USA) overnight [35,38,39]. On the second day, the slides were incubated with goat anti-rabbit secondary antibody (Alexa Fluor 288, 1:200, A-11034, ThermoFisher, Rockford, IL, USA).…”

“…One set of slides with all sections was counterstained with Hematoxylin (HHS16-550ML, Sigma Aldrich, St. Louis, MO, USA), washed in tap water for 10 min, and mounted with glycerol (BP229-1, Fisher Scientific, Pittsburgh, PA, USA). Another set of slides with serial sections were mounted with glycerol without further counterstaining for quantification of Oil red O staining using Image J [35][36][37][38]. Briefly, three 0.0359 mm 2 representative areas in the labyrinth layer of each section were selected.…”