A Microcavity Array-Based 4D Cell Culture Platform.

(1) Background: We describe a 4D cell culture platform with which we tried to detect and to characterize migration dynamics of single hematopoietic stem cells in polymer film microcavity arrays integrated into a microtiter plate. (2) Methods: The system was set up with CD34-expressing KG-1a cells as a surrogate for hematopoietic stem cells. We then evaluated the system as an artificial hematopoietic stem cell niche model comprised of a co-culture of human hematopoietic stem cells from cord blood (cord blood CD34+ cells, hHSCs) and human mesenchymal stromal cells (hMSCs) from bone marrow over a period of 21 days. We used a software-based cell detection method to count single hematopoietic stem cells (HSCs) in microcavities. (3) Results: It was possible to detect single HSCs and their migration behavior within single microcavities. The HSCs displayed a pronounced migration behavior with one population of CD34-expressing cells located at the bottom of the microcavities and one population located in the middle of the microcavities at day 14. However, at day 21 the two populations seemed to unite again so that no clear distinction between the two was possible anymore. (4) Conclusions: Single cell migration detection was possible but microscopy and flow cytometry delivered non-uniform data sets. Further optimization is currently being developed.

Coregulation of genetic programs by the transcription factors NFIB and STAT5.

Mammary-specific genetic programs are activated during pregnancy by the common transcription factor signal transducer and activator of transcription (STAT) 5. More than one third of these genes carry nuclear factor I/B (NFIB) binding motifs that coincide with STAT5 in vivo binding, suggesting functional synergy between these two transcription factors. The role of NFIB in this governance was investigated in mice from which Nfib had been inactivated in mammary stem cells or in differentiating alveolar epithelium. Although NFIB was not required for alveolar expansion, the combined absence of NFIB and STAT5 prevented the formation of functional alveoli. NFIB controlled the expression of mammary-specific and STAT5-regulated genes and chromatin immunoprecipitation-sequencing established STAT5 and NFIB binding at composite regulatory elements containing histone H3 lysine dimethylation enhancer marks and progesterone receptor binding. By integrating previously published chromatin immunoprecipitation-sequencing data sets, the presence of NFIB-STAT5 modules in other cell types was investigated. Notably, genomic sites bound by NFIB in hair follicle stem cells were also occupied by STAT5 in mammary epithelium and coincided with enhancer marks. Many of these genes were under NFIB control in both hair follicle stem cells and mammary alveolar epithelium. We propose that NFIB-STAT5 modules, possibly in conjunction with other transcription factors, control cell-specific genetic programs.