A simple, disposable, and improved organ culture system for maintaining three-dimensional development of mouse embryonic molars

Abstract

Mandibular first molars from 17-d-old mouse embryos were cultured in vitro for 2 to 4 d by a simple, disposable, improved floatation method. This method consisted of using a 24-well multidish and a plastic culture chamber with a membrane filter. The improved floatation method, as well as our previous method, was capable of the three-dimensional development of tooth germs. Cytodifferentiation of odontoblasts and ameloblasts and formation of extracellular matrices were accelerated by the present culture system, in comparison with our previous method. All the molars cultivated by this method were very similar in morphology to in vivo. On Day 2 of culture the terminal cytodifferentiation of odontoblasts and the formation of predentin were ascertained in the bucco-lingual sections of the cultured molars. A thick layer of predentin was formed at the tip of the cusp and gradually decreased toward the cervical loop and the fissure between the buccal and lingual cusps. On Day 4 in vitro, secretory ameloblasts produced enamel matrix, and the mineralized enamel showed showed prismatic structure very similar to that in vivo. Dentin and predentin also were normal in ultrastructure. The extracellular matrices (enamel, dentine, and predentin) were formed in line with the pattern of the cusp and the formation of matrices normally started at the tip of the cusp. We conclude that the three-dimensional development of whole tooth germs in vitro may be very important for normal expression of the developmental program intrinsic to mouse embryonic molars.

In vitro three‐dimensional development of mouse molar tooth germs in a rotary cell culture system

“…In vitro methods of culturing tooth germ have been used widely to explore intracellular and extracellular mechanisms involved in odontogenesis. For more accurate results, it is necessary to employ an organ culture system to simulate conditions in vivo . This study found that the RCCS was able to supply efficient high nutrient transfer, low shear force, and microgravity suitable for the development of tooth germs with appropriate spatial morphology.…”

“…Alterations in the shape and spatial arrangement of the resultant tooth germs, however, were observed. Application of this method was also hindered by a complex manufacturing process, with which it was difficult to obtain a uniform agar chamber. In each of the aforementioned studies, tooth germ ingrowth stopped after several days of culture, which indicates that tooth germ development is limited in a relatively static culture environment.…”

Trauma induced tissue survival in vitro with a muscle-biomaterial based osteogenic organoid system: a proof of concept study

“…Hence, in vivo tissue based bone inductive studies remain to date the best models to study the effect of biomaterial behavior in vivo. As such, a tissue-based bioreactor platform [23,29,30] could be superior to that of a stem cell-based system as tissues possesses various biochemical building blocks and adult stem cell niches together with preestablished cell growth promoting environments that theoretically could provide a superior culturing milieu. However, the use of bone directly as a biomaterial growth environment in vitro is highly problematic, as culture medium cannot adequately diffuse across a hard tissue barrier [30].…”

“…As such, a tissue-based bioreactor platform [23,29,30] could be superior to that of a stem cell-based system as tissues possesses various biochemical building blocks and adult stem cell niches together with preestablished cell growth promoting environments that theoretically could provide a superior culturing milieu. However, the use of bone directly as a biomaterial growth environment in vitro is highly problematic, as culture medium cannot adequately diffuse across a hard tissue barrier [30]. Therefore, the present pilot study investigated the feasibility of two in vitro skeletal muscle-based biomaterial-culturing systems as this tissue, being considered among the promising candidate grafts for bone tissue engineering, allows for better nutrient flow [31][32][33].…”

Trauma induced tissue survival in vitro with a muscle-biomaterial based osteogenic organoid system: a proof of concept study

“…Hence, in vivo tissue based bone inductive studies remain to date the best models to study the effect of biomaterial behavior in vivo. As such, a tissue-based bioreactor platform [23,29,30] could be superior to that of a stem cell based system as tissues poses various biochemical building blocks and adult stem cell niches together with pre-established cell growth promoting environments that theoretically could provide a superior culturing milieu. However, the use of bone directly as a biomaterial growth environment in vitro is highly problematic, as culture medium cannot adequately diffuse across a hard tissue barrier [30].…”