This document discusses dental and orofacial mesenchymal stem cells and their potential applications in craniofacial regeneration from the perspective of prosthodontists. It provides an overview of stem cell sources in dentistry, including dental tissue-derived stem cells like dental pulp stem cells, stem cells from exfoliated deciduous teeth, periodontal ligament stem cells, and gingival mesenchymal stem cells. It also discusses the use of hydrogels, particularly alginate hydrogel, as scaffolds for tissue engineering and encapsulating dental stem cells. Recent studies encapsulating dental stem cells in alginate hydrogels show promise for regenerating tissues like cartilage, tendon, and muscle.

1. DENTAL AND OROFACIAL MESENCHYMAL
STEM CELLS IN CRANIOFACIAL
REGENERATION:
THE PROSTHODONTIST’S POINT OF VIEW
Sahar Ansari, Tara Aghaloo, Jackson T. Seagroves, Chider Chen,
Benjamin M. Wu, Kumar Shah, Sompop Bencharit and Alireza Moshaverinia
THE JOURNAL OF PROSTHETIC DENTISTRY
ARTICLE IN PRESS
AAMIR ZAHID GODIL
SECOND YEAR P.G.
DEPARTMENT OF PROSTHODONTICS
M.A.R.D.C.

2. CONTENTS
• INTRODUCTION
• TERMINOLOGIES
• STEM CELLS AND TISSUE ENGINEERING: DENTAL ASPECTS
• SOURCES OF STEM CELLS IN DENTISTRY
• DENTAL TISSUE DERIVED STEM CELLS
• GINGIVAL MESENCHYMAL STEM CELLS (GMSCs)
• STEM CELLS FROM PERIODONTAL LIGAMENT (PDLSCs)
• HYDROGELS
• ALGINATE HYDROGEL AS SCAFFOLD FOR TISSUE ENGINEERING
• RECENT UPDATES
• CONCLUSION
• CRITIQUE

3. INTRODUCTION
• The reconstructive procedures for craniofacial tissue
regeneration are usually complex because the craniofacial
region is a complex construct, consisting of bone, cartilage, soft
tissue and neurovascular bundles.
AUTOLOGOUS
BONE
GRAFTS
(DISADVANTAG
ES)
RISK OF HEMATOMAS
DONOR SITE MORBIDITY
COST
INFECTION
INFLAMMATION
Monaco E, Bionaz M, Hollister SJ, Wheeler MB. Strategies for regeneration of the bone using porcine adult adipose-derived mesenchymal stem cells.
Theriogenology 2011;75:1381-99. 68
Sachlos E, Czernuszka JT. Making tissue engineering scaffolds work. Review on the application of solid freeform fabrication technology to the production of
tissue engineering scaffold. Eur Cell Mater 2003;5:29-39
Chen FM, Zhang J, Zhang M, An Y, Chen F, Wu ZF. A review on endogenous regenerative technology in periodontal regenerative medicine. Biomaterials

4. TERMINOLOGIES
• “Stem cell” refers to clonogenic, undifferentiated cell that is capable of self renewal
(the ability to go through numerous cell division cycles while maintaining the
undifferentiated state) and multilineage differentiation (potency or plasticity, which is
the capacity to differrentiate into specialized cell types)
Taylor CJ, Bolton EM, Bradley JA. Immunological considerations for embryonic and induced pluripotent stem
cell banking. Phil Trans R Soc Lond B Biol Sci 2011;366:2312-22.
They can recreate the entire organismTOTIPOTENT
They can create tissues of all 3 germ layers (ectoderm, mesoderm, and endoderm)PLURIPOTENT
• Embryonic stem cells (derived from inner cell mass of preimplantation embryo) have been categorized as predominately pluripotent
They can produce cell types from more than one (but not all) lineageMULTIPOTENT
• Adult stem cells (derived from many ectodermal and mesodermal organs in adults are mainly multipotent

5. STEM CELLS AND TISSUE
ENGINEERING
DENTAL ASPECTS
• The severe bone resorption in edentulous areas makes it difficult
to restore the missing teeth with dental implants or denture treatment
• Therefore, stem cell and tissue engineering therapies are
expected to provide a novel capability to regenerate large defects in
periodontal tissues and alveolar bone and to ultimately replace the
lost tooth itself.
• The tissues and organs targeted for such regenerative medicine
strategies in dentistry include the salivary gland, tongue and
craniofacial skeletal muscles, as well as the condylar cartilage of
the temporomandibular joint.

6. Adult stem
cells
Bone marrow-derived MSCs (BMSCs)- From iliac crest and orofacial bones
Dental tissue-derived stem cells
Periosteum-derived stem/progenitor cells
Salivary gland-derived stem cells
Adipose tissue-derived stem cells (ASCs)
Pluripotent
stem cells
Embryonic stem cells (ES)
Induced pluripotent stem cells (iPS)
SOURCES OF STEM CELLS IN
DENTISTRY

7. DENTAL TISSUE DERIVED STEM
CELLS
HUMAN PULP TISSUE
(POSTNATAL DENTAL PULP STEM
CELLS)Gronthos S, Brahim J, Li W, Fisher LW, Cherman N, Boyde A,
et al. Stem cell properties of human dental pulp stem cells. J
Dent Res 2002;81:531-5.
STEM CELLS FROM
EXFOLIATED DECIDUOUS TEETH
Miura M, Gronthos S, Zhao M, Lu B, Fisher LW, Robey PG, et
al. SHED: stem cells from human exfoliated deciduous teeth.
Proc Natl Acad Sci U S 2003;100:5807-12.
STEM CELLS FROM
PERIODONTAL LIGAMENT (PDLSCs)
Seo BM, Miura M, Gronthos S, Bartold PM, Batouli S, Brahim J,
et al. Investigation of multipotent postnatal stem cells from
human periodontal ligament. Lancet 2004;364:149-55.
STEM CELLS FROM APICAL PAPILLA
Sonoyama W, Liu Y, Fang D, Yamaza T, Seo BM, Zhang C, et al.
Mesenchymal stem cell-mediated functional tooth regeneration in swine.
PLoS One 2006;1:70-9.
Sonoyama W, Liu Y, Yamaza T, Tuan RS, Wang S, Shi S, et al.
Characterization of the apical papilla and its residing stem cells from
human immature permanent teeth: a pilot study. J Endod 2008;34:166-71.
DENTAL FOLLICLE PRECURSOR CELLS
Morsczeck C, Götz W, Schierholz J, Zeilhofer F, Kühn U, Möhl C, et
al. Isolation of precursor cells (PCs) from human dental follicle of
wisdom teeth. Matrix Biol 2005;24:155-65.
GINGIVAL MESENCHYMAL STEM
CELLS (GMSCs)
Tomar GB, Srivastava RK, Gupta N, Barhanpurkar AP, Pote ST, Jhaveri
HM, et al. Human gingiva-derived mesenchymal stem cells are superior to
bone marrow-derived mesenchymal stem cells for cell therapy in
regenerative medicine. Biochem Biophys Res Commun 2010;393:377-83.

9. After a tooth
was cut
horizontally, the
pulp tissue
(arrow) in the
pulp chamber
was extirpated.
This pulp
provides dental
pulp stem cells
(DPSCs)
Extracted
impacted third
molar (10-year
old female)
containing the
dental follicle
(dotted line) that
provides dental
follicle stem
cells (DFSCs)
Extracted
impacted third
molar (18-year
old male)
containing root
apical papillae
(asterisks) that
are a source of
stem cells from
apical papilla
(SCAP)

10. • A subpopulation of stem cells derived from human dental pulp
has been identified with osteogenic differentiation capacity that
can form bonelike tissues in vivo.
• In contrast with hBMMSCs, dental-derived MSCs are more
committed to be differentiated to odontogenic tissues rather
than osteogenic tissues.
• In addition to their osteogenic potential, several studies have
reported that subpopulations of dental-derived MSCs have
adipogenic and neurogenic differentiation potential as they
exhibit adipocyte- and neuronal-like cell morphologies and high
expression levels for related gene markers.
• More recent studies have confirmed that dental-derived MSCs
have better growth properties than hBMMSCs

11. • Among dental-derived stem cells, GMSCs and PDLSCs are of special
interest as they are easily accessible and can be found in the oral
environment.
GINGIVAL MESENCHYMAL STEM CELLS (GMSCs)
ADVANTAGESOF
GMSCs
EASE OF ISOLATION
HOMOGENOUS
FASTER PROLIFERATION THAN BMMSCs
FASTER HEALING OF DONOR SITE
STABLE MORPHOLOGY IN LONG TERM CULTURES
IMMUNOMODULATORY CHARACTERISTICS SIMILAR TO BMMSCs
MULTILINEAGE DIFFERENTIATION CAPACITY

12. • Periodontal ligament (PDL) contains cell populations that can
differentiate into cementoblasts or osteoblasts. MSCs derived from
PDL tissues can regenerate periodontal tissue.
• Human PDLSCs have been successfully isolated from extracted
human teeth and ex vivo-expanded PDLSCs are capable of
regenerating a typical cementum/PDL-like structure when
transplanted into immunocompromised mice using hydroxyapatite/
tricalcium phosphate as a carrier.
STEM CELLS FROM PERIODONTAL LIGAMENT
(PDLSCs)
Iwata T, Yamato M, Zhang Z, Mukobata S, Washio K, Ando T, et al. Vali dation of human periodontal ligament-
derived cells as a reliable source for cytotherapeutic use. J Clin Periodontol 2010;37:1088-99.

14. • The cell delivery vehicle has an important role in the in vivo
performance of stem cells and could dictate the success of the
regenerative therapy.
• Microscopically, a hydrogel is a network of cross-linked polymer
chains interspersed with water molecules. These polymer chains can
have a low modulus of elasticity and exhibit tissue like flexibility
because of their high water content.
• Macroscopically, a hydrogel is a porous material. The pore size can
selectively allow only certain sizes of soluble molecules or cells to
go through and promote angiogenesis.
Ahmed EM. Hydrogel: preparation, characterization, and applications: a review. J Adv Res 2015;6:105-21.
Annabi N, Nichol JW, Zhong X, Ji C, Koshy S, Khademhosseini A, et al. Controlling the porosity and microarchitecture of
hydrogels for tissue engineering. Tissue Eng Part B Rev 2010;16:371-83.
HYDROGELS

15. • Hydrogels are capable of forming 3-dimensional matrices for
the encapsulation of cells or sensitive bioactive molecules.
• Many natural and synthetic hydrogel biomaterials have
been used for craniofacial tissue regeneration and repair.
ALGINATE GELATIN FIBRIN COLLAGEN
POLYESTERS
PLA AND
PLGA
Ahmed EM. Hydrogel: preparation, characterization, and applications: a review. J Adv Res 2015;6:105-21.
Annabi N, Nichol JW, Zhong X, Ji C, Koshy S, Khademhosseini A, et al. Controlling the porosity and microarchitecture of
hydrogels for tissue engineering. Tissue Eng Part B Rev 2010;16:371-83.

16. ALGINATE HYDROGEL AS SCAFFOLD
FOR
TISSUE ENGINEERING• Among the available hydrogel biomaterials for cell encapsulation,
alginate-based hydrogels have shown promising results in
biomedical applications.
• Alginate scaffolds encapsulating MSCs can provide a suitable
microenvironment for cell viability and differentiation for tissue
regeneration applications
• Because of its inherent biocompatibility, modifiable
immunosuppression and degradation properties alginate
hydrogel has been recognized as a promising biomaterial for
biomedical applications.Weir MD, Xu HH. Human bone marrow stem cell-encapsulating calcium phosphate scaffolds for bone repair. Acta Biomater
2010;6:4118-26.
Orive G, Santos E, Poncelet D, Hernández RM, Pedraz JL, Wahlberg LU, et al. Cell encapsulation: technical and clinical advances.

17. • Several types of cells have been reported to maintain their
morphology in alginate hydrogel scaffolds
Chondrocytes have been successfully encapsulated in
alginate scaffolds and have been differentiated toward
chondrogenic tissues.Perng CK, Kao CL, Yang YP, Lin HT, Lin WB, Chu YR, et al. Culturing adult human
bone marrow stem cells on gelatin scaffold with pNIPAAm as transplanted grafts for
skin regeneration. J Biomed Mater Res A 2008;84: 622-30.
Hepatocytes have been encapsulated in alginate hydrogel as a 3-
dimensional scaffold and encapsulated cells were found to grow in
alginate scaffoldsNunamaker EA, Purcell EK, Kipke DR. In vivo stability and
biocompatibility of implanted calcium alginate disks. J Biomed Mater
Res A 2007;83:1128-37
Other studies have reported the encapsulation of different types of
cells such as cardiomyocytes, rat marrow cells, and Schwann cells in
hydrogel scaffolds.Lee KY, Rowley JA, Eiselt P, Moy EM, Bouhadir KH, Mooney DJ. Controlling mechanical and swelling properties of alginate hydrogels independently by cross-linker
type and cross-linking density. Macromolecules 2000;33:4291-4.
Strand B, Gåserød O, Kulseng B, Espevik T, Skjåk-Bræk G. Alginate-polylysine-alginate microcapsules: effect of size reduction on capsule properties. J Microencapsul
2002;19:615-30.

18. • Microencapsulation systems are one of the most popular
strategies in tissue engineering.
• Hydrogels and specifically alginate are quite promising
because of the ease of infusion of nutrients and oxygen
between encapsulated stem cells within the alginate
hydrogels and the microenvironment while the
encapsulated stem cells stay within the 3-dimensional
matrix.
• The alginate microspheres can be injected at the time
of transplantation and can be used as drug delivery
devices, making them the materials of choice for many

19. • Alginate hydrogel scaffold can be considered as a promising
encapsulating biomaterial for dental- and orofacial derived MSCs,
including GMSCs and PDLSCs
• Alginate microspheres can fill the gaps of irregularly shaped
defects, enabling minimally invasive surgical procedures.
• Because of the biodegradability of alginate hydrogel, an additional
clinical visit to remove the scaffold can be eliminated.
• Alginate hydrogel scaffold is capable of enhancing hard and soft
tissue regeneration to accomplish minimally invasive dental and
orthopedic surgeries.

21. http://www.reelabs.com/stem-cell-banking/dental-stem-cell-
banking.html
DENTAL STEM CELLS
COLLECTION AND
MAINTENANCE
(REELABS, MUMBAI- INDIA)

22. Moshaverinia et al
• Moshaverinia et al developed a novel co-delivery system
based on transforming growth factor (TGF)-b1-loaded alginate
microspheres encapsulating PDLSCs and GMSCs for cartilage
regeneration applications.
• Moreover, Moshaverinia et al confirmed that PDLSCs and
GMSCs encapsulated in biodegradable and injectable alginate
hydrogel scaffold containing appropriate inductive signals (TGF-
b3) can be used as an alternative treatment in tendon tissue
engineering. MSC and alginate constructs were able to
effectively differentiate and organize their extracellular matrix
into tendon tissue.
Moshaverinia A, Xu X, Chen C, Akiyama K, Snead ML, Shi S. Dental mesenchymal stem cells encapsulated in an alginate hydrogel co-delivery
microencapsulation system for cartilage regeneration. Acta Biomater 2013;9: 9343-50.
Moshaverinia A, Xu X, Chen C, Ansari S, Zadeh HH, Snead ML, et al. Application of stem cells derived from the periodontal ligament or gingival
tissue sources for tendon tissue regeneration. Biomaterials 2014;35: 2642-50.

23. Ansari et al
• More recently, Ansari et al confirmed that GMSCs, when
encapsulated in an alginate hydrogel microencapsulation system with
multiple growth factor delivery capacity (loaded with myogenic growth
factors), can be used in the repair and regeneration of muscular
tissues.
• They also reported that the GMSC/alginate hydrogel construct could
be considered a promising candidate for vascularized tissue
engineering.
• Based on the results of this study, GMSCs appear be an
advantageous alternative therapeutic approach for tongue muscle
repair and regeneration.
Ansari S, Chen C, Xu X, Annabi N, Zadeh HH, Wu BM, et al. Muscle tissue engineering using
gingival mesenchymal stem cells encapsulated in alginate hydrogels containing multiple growth
factors. Ann Biomed Eng 2016;44: 1908-20.

24. CONCLUSION

25. CRITIQUE
• Very little information about signaling molecules and
their clinical implications
• Feasibility in day-to-day practice is questionable
• Inadequate evidence based clinical data mentioned

26. Thank You