Role of biotin-containing membranes and nuclear distribution in differentiating human endometrial cells

Abstract

Human Ishikawa endometrial cells form domes when confluent monolayers are stimulated with fresh fetal bovine serum. Extensive structural and biochemical changes have been detected during the approximately 30 h differentiation period. The earliest detectable change involves the formation of multinucleated structures and the appearance of "granules" that stain for biotin within those structures. Nuclei become associated with each other and are ultimately enclosed within a biotin-containing membrane. Aggregated membrane-sheathed nuclei and the cells containing them begin to elevate from the dish as biotin staining becomes apparent in apical membranes. The elevated structures are called predomes and consist of one or more very large cells containing the sheathed nuclei. Apical membranes of these unusual cells extend far out into the medium in structures that resemble endometrial pinopods. A lumen under the elevated cells fills with transcytosed fluid. As differentiation proceeds, highly concentrated chromatin material that was flattened against apical and lateral membranes of the predome cells begins to disperse. Small mononuclear cells evolve from larger predome cells. Apical membranes of predome and dome cells continue to stain for biotin. Gel electrophoresis of SDS-solubilized biotin-containing membranes, followed by Western blot analysis using avidin-linked peroxidase, resulted in three stained bands with molecular weights similar to those of the mitochondrial carboxylases: propionyl carboxylase, methylmalonyl carboxylase, and pyruvate carboxylase.

Polyploid monolayer Ishikawa endometrial cells form unicellular hollow spheroids capable of migration

“…Walen (2002) The consensus seems to be that endocycling, DNA replication without cytokinesis, is the predominant way in which polyploid cells arise, with syncytial formation recognized as an alternative mechanism. It is syncytial formation that gives rise to polyploid predomes in Ishikawa endometrial cells as previously described (Fleming 1998 andFleming 2016).…”

“…In the final stages of detachment the biotin stain is found in the center of polyploid cells in structures that resemble the mitochondrial superstructures (mitonucleons) that envelop nuclear aggregates during dome formation (Fleming et al 1998;Fleming 2016;Fleming 2018). One function of these mitonucleons appears to be the retention of gases (Fleming 2016;Fleming, 2018), and in a single detached cell the retention of gases results in the formation of a unicellular, albeit polyploid, floating spheroid.…”

“…al., 1973) endometrial cells were stimulated to differentiate into fluid-filled hemispheres (Fleming et al, 1998;Fleming, 2016a,b,c). These hemispheres can ultimately extend into gland-like structures, a frequent epithelial differentiation that takes place in organs throughout the body and in pre-menopausal women every month in the endometrial lining of the uterus.…”

Mitonucleons formed during differentiation of Ishikawa endometrial cells generate vacuoles that elevate monolayer syncytia: Differentiation of Ishikawa domes, Part 1

“…Clones could be isolated that made more and larger domes including a clone that 68 routinely extended domes into everted gland-like structures (Fleming et al, 1998;69 Fleming, 1999;Fleming 2016c As the bubble-like vacuoles enlarge within and compress the heterochromatin, the 143 resulting structure ( fig.3) . 4a).…”

“…Started from a well-differentiated adenocarcinoma and found to contain estradiol and 48 progesterone receptors (Nishida et al 1985) the cells were shown capable of 49 functions characteristic of normal endometrial cells such as enhanced proliferation in 50 response to estradiol and tamoxifen (Holinka et al,1986a To characterize the process we needed to find conditions that predictably resulted in 57 dome formation (Fleming, 1995), determine what factors enhanced the process 58 (Fleming et al 1998), look for the synthesis of new proteins (Fleming et.al. 1995;59 Fleming 1999), and finally examine the structural changes underlying differentiation.…”