Three-dimensional Sphere-forming Cells Are Unique Multipotent Cell Population in Dental Pulp Cells

Lee, Sung Hee; Inaba, Andrew; Mohindroo, NK; Ganesh, Deepika; Martin, Charlotte; Chugal, Nadia; Kim, Reuben H.; Kang, Mo K.; Park, No‐Hee; Shin, Kihyuk

doi:10.1016/j.joen.2017.03.016

Cited by 17 publications

(22 citation statements)

References 31 publications

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“…However, positive staining (Alizarin red) for mineral deposits was only observed in osteogenically induced GPC/BMSC spheroids herein. Indeed, previous studies have reported superior in vivo bone regeneration by osteogenically induced spheroids of human BMSCs (Suenaga et al, 2015), DPCs (Lee et al, 2017) and PDLCs (Moritani et al, 2018), vs. monolayers. Thus, it may be hypothesized that MCC-like assemblies induced by spheroid culture intrinsically 'prime' MSCs towards osteoblastic commitment, although extrinsic signals/supplements may be necessary for terminal differentiation and/or matrix mineralization (Kale et al, 2000;Facer et al, 2005).…”

Section: Discussionmentioning

confidence: 98%

“…While a majority of the literature is focused on BMSCs, 3D cultures have also been reported to enhance the survival, stemness, paracrine/immunomodulatory activity, and multilineage differentiation of oral tissue-derived MSCs (Zhang et al, 2012;Lee et al, 2017;Moritani et al, 2018;Subbarayan et al, 2018). However, few studies have characterized MSC spheroids in xeno-free cultures to facilitate clinical translation (Ylostalo et al, 2017;Dong et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

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Xeno-Free Spheroids of Human Gingiva-Derived Progenitor Cells for Bone Tissue Engineering

Shanbhag

Suliman

Bolstad

et al. 2020

Front. Bioeng. Biotechnol.

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Gingiva has been identified as a minimally invasive source of multipotent progenitor cells (GPCs) for use in bone tissue engineering (BTE). To facilitate clinical translation, it is important to characterize GPCs in xeno-free cultures. Recent evidence indicates several advantages of three-dimensional (3D) spheroid cultures of mesenchymal stromal cells (MSCs) over conventional 2D monolayers. The present study aimed to characterize human GPCs in xeno-free 2D cultures, and to test their osteogenic potential in 3D cultures, in comparison to bone marrow MSCs (BMSCs). Primary GPCs and BMSCs were expanded in human platelet lysate (HPL) or fetal bovine serum (FBS) and characterized based on in vitro proliferation, immunophenotype and multilineage differentiation. Next, 3D spheroids of GPCs and BMSCs were formed via self-assembly and cultured in HPL. Expression of stemness-(SOX2, OCT4, NANOG) and osteogenesis-related markers (BMP2, RUNX2, OPN, OCN) was assessed at gene and protein levels in 3D and 2D cultures. The cytokine profile of 3D and 2D GPCs and BMSCs was assessed via a multiplex immunoassay. Monolayer GPCs in both HPL and FBS demonstrated a characteristic MSC-like immunophenotype and multi-lineage differentiation; osteogenic differentiation of GPCs was enhanced in HPL vs. FBS. CD271 + GPCs in HPL spontaneously acquired a neuronal phenotype and strongly expressed neuronal/glial markers. 3D spheroids of GPCs and BMSCs with high cell viability were formed in HPL media. Expression of stemness-and osteogenesisrelated genes was significantly upregulated in 3D vs. 2D GPCs/BMSCs; the latter was independent of osteogenic induction. Synthesis of SOX2, BMP2 and OCN was confirmed via immunostaining, and in vitro mineralization via Alizarin red staining. Finally, secretion of several growth factors and chemokines was enhanced in GPC/BMSC spheroids, while that of pro-inflammatory cytokines was reduced, compared to monolayers. In summary, monolayer GPCs expanded in HPL demonstrate enhanced osteogenic differentiation potential, comparable to that of BMSCs. Xeno-free spheroid culture further enhances stemness-and osteogenesis-related gene expression, and cytokine secretion in GPCs, comparable to that of BMSCs.

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Section: Discussionmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

Xeno-Free Spheroids of Human Gingiva-Derived Progenitor Cells for Bone Tissue Engineering

Shanbhag

Suliman

Bolstad

et al. 2020

Front. Bioeng. Biotechnol.

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“…These secretions can migrate to inflamed tissues and exert anti-inflammatory and immunosuppressive effects on both the adaptive and innate immune systems. Paracrine effects of MSCs include immunomodulation, angiogenesis, antiapoptosis, sustenance to the growth of cells, and chemo-attraction properties [ 9 , 10 , 11 ]. Transplantation of the conditioned medium (CM) that contains these paracrine factors have been reported to enhance wound healing in animal models [ 8 , 12 , 13 ].…”

Section: Introductionmentioning

confidence: 99%

Effect of Ascorbic Acid on Differentiation, Secretome and Stemness of Stem Cells from Human Exfoliated Deciduous Tooth (SHEDs)

Bhandi

Alkahtani

Mashyakhy

et al. 2021

JPM

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Stem cells from human exfoliated deciduous teeth (SHEDs) are considered a type of mesenchymal stem cells (MSCs) because of their unique origin from the neural crest. SHEDs can self-renewal and multi-lineage differentiation with the ability to differentiate into odontoblasts, osteoblast, chondrocytes, neuronal cells, hepatocytes, adipocytes, etc. They are emerging as an ideal source of MSCs because of their easy availability and extraordinary cell number. Ascorbic acid, or vitamin C, has many cell-based applications, such as bone regeneration, osteoblastic differentiation, or extracellular matrix production. It also impacts stem cell plasticity and the ability to sustain pluripotent activity. In this study, we evaluate the effects of ascorbic acid on stemness, paracrine secretion, and differentiation into osteoblast, chondrocytes, and adipocytes. SHEDs displayed enhanced multifaceted activity, which may have applications in regenerative therapy.

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“…They found that ALB and G6P were detected earlier and higher levels of urea and AFP were produced in the 3D culture compared to those in the 2D culture. Lee et al [12] reported that DPSC spheres created by ultra-low attachment (ULA) culture plates possess a greater multilineage differentiation capacity compared to that in monolayer DPSCs, suggesting that a 3D culture probably better reflects the in vivo microenvironment of stem cells.…”

Section: Introductionmentioning

confidence: 99%

In Vitro Characterization of Dental Pulp Stem Cells Cultured in Two Microsphere-Forming Culture Plates

Lee

et al. 2020

JCM

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Various three-dimensional (3D) culture methods have been introduced to overcome the limitations of in vitro culture and mimic in vivo conditions. This study aimed to evaluate two microsphere-forming culture methods and a monolayer culture method. We evaluated cell morphology, viability, osteo-, adipo-, and chondrogenic differentiation potential of dental pulp stem cells (DPSCs) cultured in 3D culture plates: ultra-low attachment (ULA) and U-bottomed StemFit 3D (SF) plates, and a two-dimensional (2D) monolayer plate. RNA sequencing (RNA-seq) revealed differentially expressed gene (DEG) profiles of the DPSCs. In contrast to an increasing pattern in the 2D group, cell viability in 3D groups (ULA and SF) showed a decreasing pattern; however, high multilineage differentiation was observed in both the 3D groups. RNA-seq showed significantly overexpressed gene ontology categories including angiogenesis, cell migration, differentiation, and proliferation in the 3D groups. Hierarchical clustering analysis revealed a similar DEG regulation pattern between the 3D groups; however, a comparatively different DEG was observed between the 2D and 3D groups. Taken together, this study shows that DPSCs cultured in microsphere-forming plates present superior multilineage differentiation capacities and demonstrate higher DEG expression in regeneration-related gene categories compared to that in DPSCs cultured in a conventional monolayer plate.

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Three-dimensional Sphere-forming Cells Are Unique Multipotent Cell Population in Dental Pulp Cells

Cited by 17 publications

References 31 publications

Xeno-Free Spheroids of Human Gingiva-Derived Progenitor Cells for Bone Tissue Engineering

Xeno-Free Spheroids of Human Gingiva-Derived Progenitor Cells for Bone Tissue Engineering

Effect of Ascorbic Acid on Differentiation, Secretome and Stemness of Stem Cells from Human Exfoliated Deciduous Tooth (SHEDs)

In Vitro Characterization of Dental Pulp Stem Cells Cultured in Two Microsphere-Forming Culture Plates

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