PMC:13917 / 26154-28834
Annnotations
{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/13917","sourcedb":"PMC","sourceid":"13917","source_url":"http://www.ncbi.nlm.nih.gov/pmc/13917","text":"Expression of differentiation markers\nMilk protein synthesis in vivo occurs within clusters of differentiated mammary epithelial cells in response to lactogenic hormones, ECM interactions and cell-cell interactions [1]. In cultures of KIM-2 cells grown at the semipermissive temperature of 37°C or the nonpermissive temperature of 39°C, addition of lactogenic hormones induced partial differentiation as assessed by the synthesis of high levels of β-casein.\nFigure 9a shows a western blot of induced and uninduced late passage cells (P31) grown on tissue culture plastic. For induction, the cells are grown to confluence, EGF removed for 2 days, and the lactogenic hormones dexamethasone, insulin and prolactin added for the time period indicated. There appear to be at least two different forms of cytoplasmic β-casein induced. One has an apparent molecular mass of 29 kDa and there is also a lower molecular weight band, compared with 32 kDa for the secreted protein found in milk. The discrepancy in size is probably due to differences in post-translational modifications between intracellular β-casein and the secreted form, and was also observed in HC11 mammary cells (Fig. 9a). The level of β-casein induction is not affected by passage number or growth on collagen (data not shown). KIM-2 cells do secrete β-casein but at very low levels.\nOther milk proteins that are expressed later in pregnancy, such as WAP, appear to require other factors in addition to lactogenic hormones for their expression. We were interested in investigating the possibility of WAP expression in KIM-2 cultures, because this is a more appropriate measure of differentiation status. Figure 9b shows a time course of WAP mRNA induction with lactogenic hormones in KIM-2 cells grown on tissue culture plastic. Whereas β-casein expression was induced after 2 days, WAP expression was delayed until 4-8 days after hormone stimulation. These kinetics reflect the pattern of milk protein gene expression in the mammary gland, in which β-casein is expressed from day 10 of pregnancy but WAP is not expressed until approximately 4 days later. Recently a STAT5a knockout has been generated that exhibits an impairment of lobuloaveolar development during pregnancy and an inability to lactate [31]. Although these mice show normal β-casein expression, the levels of WAP expression were reduced considerably. Clearly the induction of WAP expression has more complex requirements that may include contacts with the ECM, expression of specific receptors and/or expression of differentiation-related genes. It is possible that prolonged stimulation of KIM-2 cells with lactogenic hormones induces these changes.","divisions":[{"label":"Title","span":{"begin":0,"end":37}}],"tracks":[{"project":"Colil","denotations":[{"id":"T29","span":{"begin":2267,"end":2269},"obj":"9009201"},{"id":"T33","span":{"begin":216,"end":217},"obj":"7001510"}],"namespaces":[{"prefix":"_base","uri":"http://pubannotation.org/docs/sourcedb/PubMed/sourceid/"}],"attributes":[{"subj":"T29","pred":"source","obj":"Colil"},{"subj":"T33","pred":"source","obj":"Colil"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"Colil","color":"#93b4ec","default":true}]}]}}