Limits...
Differentiation of human mesenchymal stem cell spheroids under microgravity conditions.

Cerwinka WH, Sharp SM, Boyan BD, Zhau HE, Chung LW, Yates C - Cell Regen (Lond) (2012)

Bottom Line: Spheroids cultured in undifferentiated spheroids of 3 and 10 days retained expression of CD44, without expression of differentiation markers.Spheroids cultured in adipogenic and osteogenic differentiation media exhibited oil red O staining and von Kossa staining, respectively.The use of gelatin scaffolds holds promise to design implantable stem cell tissue of various sizes and shapes for future regenerative treatment.

View Article: PubMed Central - PubMed

Affiliation: Children's Healthcare of Atlanta, Emory University School of Medicine, 5445Meridian Mark Road, Suite 420, Atlanta, GA 30342 USA ; Georgia Pediatric Urology, 5445 Meridian Mark Rd, Suite 420, Atlanta, GA 30342 USA.

ABSTRACT
To develop and characterize a novel cell culture method for the generation of undifferentiated and differentiated human mesenchymal stem cell 3D structures, we utilized the RWV system with a gelatin-based scaffold. 3 × 10(6) cells generated homogeneous spheroids and maximum spheroid loading was accomplished after 3 days of culture. Spheroids cultured in undifferentiated spheroids of 3 and 10 days retained expression of CD44, without expression of differentiation markers. Spheroids cultured in adipogenic and osteogenic differentiation media exhibited oil red O staining and von Kossa staining, respectively. Further characterization of osteogenic lineage, showed that 10 day spheroids exhibited stronger calcification than any other experimental group corresponding with significant expression of vitamin D receptor, alkaline phosphatase, and ERp60 . In conclusion this study describes a novel RWV culture method that allowed efficacious engineering of undifferentiated human mesenchymal stem cell spheroids and rapid osteogenic differentiation. The use of gelatin scaffolds holds promise to design implantable stem cell tissue of various sizes and shapes for future regenerative treatment.

No MeSH data available.


Related in: MedlinePlus

Schematic of generation of multicellular spheroids. 2D cultured hMSC were suspended in growth media in the presence of scaffold. Cell suspensions containing scaffolds were transferred to RWV rotary cell culture system and cultured at 4 rounds per minute for designated time intervals. After designated time intervals multicellular spheroids were sectioned and examined for cellularity and differentiation status.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4230401&req=5

Fig1: Schematic of generation of multicellular spheroids. 2D cultured hMSC were suspended in growth media in the presence of scaffold. Cell suspensions containing scaffolds were transferred to RWV rotary cell culture system and cultured at 4 rounds per minute for designated time intervals. After designated time intervals multicellular spheroids were sectioned and examined for cellularity and differentiation status.

Mentions: Gelatin sponge is a scaffold, which has pores separated by thin (few μm in thickness) walls [15]. We employed a cell loading procedure utilizing empirically determined 1 × 1 × 1 mm sponge scaffold to maximize hMSC loading and future engrafting (Figure 1).Figure 1


Differentiation of human mesenchymal stem cell spheroids under microgravity conditions.

Cerwinka WH, Sharp SM, Boyan BD, Zhau HE, Chung LW, Yates C - Cell Regen (Lond) (2012)

Schematic of generation of multicellular spheroids. 2D cultured hMSC were suspended in growth media in the presence of scaffold. Cell suspensions containing scaffolds were transferred to RWV rotary cell culture system and cultured at 4 rounds per minute for designated time intervals. After designated time intervals multicellular spheroids were sectioned and examined for cellularity and differentiation status.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4230401&req=5

Fig1: Schematic of generation of multicellular spheroids. 2D cultured hMSC were suspended in growth media in the presence of scaffold. Cell suspensions containing scaffolds were transferred to RWV rotary cell culture system and cultured at 4 rounds per minute for designated time intervals. After designated time intervals multicellular spheroids were sectioned and examined for cellularity and differentiation status.
Mentions: Gelatin sponge is a scaffold, which has pores separated by thin (few μm in thickness) walls [15]. We employed a cell loading procedure utilizing empirically determined 1 × 1 × 1 mm sponge scaffold to maximize hMSC loading and future engrafting (Figure 1).Figure 1

Bottom Line: Spheroids cultured in undifferentiated spheroids of 3 and 10 days retained expression of CD44, without expression of differentiation markers.Spheroids cultured in adipogenic and osteogenic differentiation media exhibited oil red O staining and von Kossa staining, respectively.The use of gelatin scaffolds holds promise to design implantable stem cell tissue of various sizes and shapes for future regenerative treatment.

View Article: PubMed Central - PubMed

Affiliation: Children's Healthcare of Atlanta, Emory University School of Medicine, 5445Meridian Mark Road, Suite 420, Atlanta, GA 30342 USA ; Georgia Pediatric Urology, 5445 Meridian Mark Rd, Suite 420, Atlanta, GA 30342 USA.

ABSTRACT
To develop and characterize a novel cell culture method for the generation of undifferentiated and differentiated human mesenchymal stem cell 3D structures, we utilized the RWV system with a gelatin-based scaffold. 3 × 10(6) cells generated homogeneous spheroids and maximum spheroid loading was accomplished after 3 days of culture. Spheroids cultured in undifferentiated spheroids of 3 and 10 days retained expression of CD44, without expression of differentiation markers. Spheroids cultured in adipogenic and osteogenic differentiation media exhibited oil red O staining and von Kossa staining, respectively. Further characterization of osteogenic lineage, showed that 10 day spheroids exhibited stronger calcification than any other experimental group corresponding with significant expression of vitamin D receptor, alkaline phosphatase, and ERp60 . In conclusion this study describes a novel RWV culture method that allowed efficacious engineering of undifferentiated human mesenchymal stem cell spheroids and rapid osteogenic differentiation. The use of gelatin scaffolds holds promise to design implantable stem cell tissue of various sizes and shapes for future regenerative treatment.

No MeSH data available.


Related in: MedlinePlus