Human levator veli palatini muscle: a novel source of mesenchymal stromal cells for use in the rehabilitation of patients with congenital craniofacial malformations

Daniela Franco Bueno,Reza Jarrahy,Diogenes Laercio Rocha,Cassio Eduardo Raposo-Amaral,Daniela Y S Tanikawa,Maria Rita Passos-Bueno,Carla Cristina Gomes Pinheiro,Gerson Shigueru Kabayashi,Akishige Hokugo,Yoichiro Shibuya,José Ricardo Muniz Ferreira,Patricia A Zuk

doi:10.1186/s13287-020-02017-7

Abstract

BackgroundBone reconstruction in congenital craniofacial differences, which affect about 2–3% of newborns, has long been the focus of intensive research in the field of bone tissue engineering. The possibility of using mesenchymal stromal cells in regenerative medicine protocols has opened a new field of investigation aimed at finding optimal sources of multipotent cells that can be isolated via non-invasive procedures. In this study, we analyzed whether levator veli palatini muscle fragments, which can be readily obtained in non-invasive manner during palatoplasty in cleft palate patients, represent a novel source of MSCs with osteogenic potential.MethodsWe obtained levator veli palatini muscle fragments (3–5 mm3), during surgical repair of cleft palate in 5 unrelated patients. Mesenchymal stromal cells were isolated from the muscle using a pre-plating technique and other standard practices. The multipotent nature of the isolated stromal cells was demonstrated via flow cytometry analysis and by induction along osteogenic, adipogenic, and chondrogenic differentiation pathways. To demonstrate the osteogenic potential of these cells in vivo, they were used to reconstruct a critical-sized full-thickness calvarial defect model in immunocompetent rats.ResultsFlow cytometry analysis showed that the isolated stromal cells were positive for mesenchymal stem cell antigens (CD29, CD44, CD73, CD90, and CD105) and negative for hematopoietic (CD34 and CD45) or endothelial cell markers (CD31). The cells successfully underwent osteogenic, chondrogenic, and adipogenic cell differentiation under appropriate cell culture conditions. Calvarial defects treated with CellCeram™ scaffolds seeded with the isolated levator veli palatini muscle cells showed greater bone healing compared to defects treated with acellular scaffolds.ConclusionCells derived from levator veli palatini muscle have phenotypic characteristics similar to other mesenchymal stromal cells, both in vitro and in vivo. Our findings suggest that these cells may have clinical relevance in the surgical rehabilitation of patients with cleft palate and other craniofacial anomalies characterized by significant bone deficit.

Highlights

The therapeutic potential of mesenchymal stromal cells in bone tissue engineering is promising, as their use may allow for the reconstruction of complex bone defects without the need for associated donor site morbidity, which is a distinct limitation when autologous bone grafts are used
Cells derived from levator veli palatini muscle have phenotypic characteristics similar to other mesenchymal stromal cells, both in vitro and in vivo
Our findings suggest that these cells may have clinical relevance in the surgical rehabilitation of patients with cleft palate and other craniofacial anomalies characterized by significant bone deficit

Summary

Introduction

The therapeutic potential of mesenchymal stromal cells in bone tissue engineering is promising, as their use may allow for the reconstruction of complex bone defects without the need for associated donor site morbidity, which is a distinct limitation when autologous bone grafts are used. In approximately 50% of cases, CP occurs as an isolated entity, while the remainder of cases are associated with various syndromes in which other structures are affected [4] In these syndromic cases, patients may exhibit other facial bone malformations that require surgical correction, as in the case of Treacher-Collins syndrome [5] and Goldenhar syndrome [6]. The benefits of these surgical procedures may be offset by complications such as donor site morbidity, postsurgical reabsorption, and infection [7, 8] To circumvent these problems, researchers have focused on the development of bone tissue engineering strategies using various combinations of osteogenic materials, growth factors, and stem cells that may offer alternative methods with comparatively minimal or no donor site morbidity and lower overall complication profiles [9,10,11,12]. To demonstrate the osteogenic potential of these cells in vivo, they were used to reconstruct a critical-sized fullthickness calvarial defect model in immunocompetent rats

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