Dynamics of the Self-Assembly of Complex Cellular aggregates on Micromolded Nonadhesive Hydrogels
Dynamics of the Self-Assembly of Complex Cellular aggregates on Micromolded Nonadhesive Hydrogels
Napolitano, A., Chai, P., Dean, D., and Morgan, J.
http://www.liebertonline.com/doi/pdf/10.1089/ten.2006.0190
Cell aggregation to form three-dimensional spherical shapes combined with self-segregating properties that allow the formation of multilayered, tissue-esque, three-dimensional shapes are important to the development of living tissues. Spheroid tissue shapes serve to increase gene expression; they also generate appropriate cell-cell interactions. Since previous studies involving embryonic tissues and adult cells have shown such self-aggregating properties, Napolitano, et al. sought to create nonadhesive hydrogels to study size, shape, dynamics, and composition involved with the normal human fibroblast (NHF) and the human umbilical vein endothelial cell (HUVEC) self-assembly process. The micromolded hydrogels enabled the group to study the dynamics of cell assembly given various geometric forms, including a toroid shape.
The group designed arrays of 800micron tall pegs that served as molds for seeding wells. Using different diameter wells and different morphological shapes, the team evaluated recess shape in cell aggregation and found that aggregation was controlled by recesses in hemispherical, large, flat bottoms more so than by irregularly shaped recesses. Hemispherical wells generated better cell regularity than did flat-bottom wells. The researchers used time-lapse (brightfield) microscopy to assess parameters such as circumferential contraction, re-segregation potential*, spheroid viability, and aggregate geometry for the cells. They analyzed the ells via a live/dad and calcein staining. When HUVECs and NHFs were co-seeded, a multi-layered spherical structure formed wherein the HUVECs formed a multilayer surrounding a NHF core. They also noted that given HUVEC spheroids combined with a NHF cell suspension, the NHFs reorganized to form a core surrounded by a HUVEC layer. They thus showed that spheroids retain the ability o reassemble and that cells can assemble to form complex shapes.
I chose this paper since the group used various techniques used in class to study methods of enhancing cell morphology. Some techniques we’ve studied include: live-dead determination/calcein staining, brightfield microscopy, cell passaging, cell counting, etc. Overall, though, this paper is important since the group utilizes non-adhesive hydrogels that minimize cell-substrate interactions and maximize cell-cell interactions. Also, since three-dimensional spheroid shapes are important in increasing gene expression, this study is an intriguing look on ways to improve culture techniques.
* by this, I mean the ability for mature spheroids to re-segregate into multilayered elementary tissue structures
6 comments:
Yeah! This is the only paper that I can understand most of the terminology, since most of the lab techniques are similar to what we are doing in the class right now. I still remember our 3D Rex cell culture all sticked to the bottom of the well. I guess it would be interesting if we can do the experiment again with this nonadhesive hydrogel. May be we can suggest Terry about this, lol!
Haha, I'm glad you can understand it; I wanted it to be very readable.
But, I just noticed some errors that are irriating me. I'm sure you all are smart enough to understand it; even still, in the second paragraph, it should read:
“They analyzed the cells (not “ells”) via a live/dead (not “live/dad”) and calcein staining”
And “ spheroids retain the ability to (not “o”) reassemble and that cells can assemble to form complex shapes.”
And, yes, it would be a good idea to talk to Terry about using hydrogels.
~Angelee
Um, I didn't mean that Terry would say yes or anything; I just meant it would be a good idea to consult him if you are interested in this for your project.
~Angelee
The article talked about cellular aggregation. It is just applicable to NHF anf HUVEC in this case; however, it does remind me a lot of the sperical structure of Rex culture in 3D. It would be interesting to know if more cell types react in such a special way using this kind of experiment.
Did the researchers explain how the spheroid tissue shapes serve to increase gene expression? And also why the HUVEC surrounds NHF cells and what is that relevant to?
They mentioned that according to the self-assembly theory: the
differential adhesion hypothesis (DAH), cellular aggregates
assume spherical geometry to maximize intercellular
adhesion. The theory also implies that cell types self-segregate "due to differences in cell-cell adhesion or apparent surface tension, with those cells of highest cohesion(like to like adhesion) located on the inside of a spheroid and those cells with lower cohesion located on the outside."
HUVECs, then surround NHF cells due to the higher cohesion of the NHF cells. Even still, the spheroid structures are not static and retain fluidity.
The incorporation of both cell lines is important in tissue engineering applications, as fibroblasts play an important role in the production of extracellular matrix components in tissue. According to wikipedia, fibroblasts, like the NHF line, are important in wound healing and in structural support (by production of collagens, glycosaminoglycans, reticular and elastic fibers, and glycoproteins).
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