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Knowledge for Clinical Practice

Dental Stem Cells: A Guide for Dental Professionals

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A PEER-REVIEWED PUBLICATIONA PEER-REVIEWED PUBLICATION

DENTAL LEARNING

Jeffrey Gruneich, PhD, MS; and Fiona M. Collins, BDS, MBA, MA

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Written fordentists, hygienists

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ABSTRACT

EDUCATIONAL OBJECTIVES

The overall objective of this article is to provide the participant with information on stem cells, particularly dental stem cells. Upon completing this course, the participant will be able to:

1. Delineate key elements of the AAPD policy on stem cells and how this translates into daily practice.

2. List the types, sources, and basic properties of stem cells.3. List and describe the range of potential clinical uses for

dental stem cells and the current status of these in type 1 diabetes and spinal cord injuries.

4. Describe the key elements involved in discussing dental stem cell banking services with patients and in providing these services.

The science of stem cells and their clinical use in regenerative medicine and dentistry have developed significantly over the last decade. Stem cells may be derived from various sources within the body: teeth and dental pulp, bone marrow, cord blood, and adipose tissue. Most importantly for the dental professional, teeth can provide a plenti-ful and potent source of stem cells which can be preserved rather than discarded as medical waste. Dental stem cells are a convenient, noncontroversial, and affordable source of stem cells for families that wish to preserve their stem cells for future use. Dental professionals are in a unique position to build awareness about the option of stor-ing stem cells from teeth, as well as to assist patients with the collec-tion of candidate teeth for subsequent lab processing and cryopreser-vation by the dental stem cell bank. In 2008, the American Academy of Pediatric Dentistry published a policy on stem cells that provides a useful framework for dental stem cells dental stem cell banking. This couse reviews the science behind dental stem cells and, specifically for dental professionals, the processes involved with collection of dental stem cells, their processing and preservation.

ABOUT THE AUTHORS

Dr. Jeffrey Gruneich was a worldwide biobank-ing specialist for IBM Healthcare and Life Sciences, providing IT solutions and consult-ing for leading pharmaceutical, biotechnology, academic, government, hospital, and consumer advocacy organizations as they created new processes, products, and services in translational research and regenerative medicine. Prior to

IBM, Dr. Gruneich co-founded eTechtransfer.com, an online exchange for biotech intellectual property, and Infoceutics Inc., a bioinformatics company that identified key infectious genes in microbiology. Jeff also developed immunoassays and reagents for food and environmental monitoring for Charm Sciences in Malden, MA. Dr. Gruneich received his Ph.D. in Bioengineering from the University of Pennsylvania with a focus in cell and gene therapy, an M.S. in chemistry at UC Berkeley as a National Science Foundation graduate fellow, and B.S. degrees in chemistry and mathematics at Southern Methodist University. He lives in Boston with his wife and two sons. AUTHOR DISCLOSURE: Dr. Gruneich is vice president and co-founder of Provia Laboratories. He can be reached at [email protected].

Dr. Fiona M. Collins has authored and presented CE courses to dental professionals and students in the United States and internationally, and has been an active consultant in the dental industry for several years. She is a member of the Ameri-can Dental Association and the Organization for Asepsis and Safety Procedures, and has been a member of the British Dental Association, Dutch Dental Association, the International Assocation for Dental Research and the Academy of General

Dentistry Foundation Strategy Board. Fiona earned her dental degree from Glasgow University and holds an MBA and MA from Boston University. AUTHOR DISCLOSURE: Dr. Collins does not have a leadership position or a commercial interest with any products that are mentioned in this article, or with products and services discussed in this educational activity. Dr. Collins can be reached at [email protected]

SPONSOR/PROVIDER: This is a Dental Learning, LLC continuing education activity. COMMERCIAL SUPPORTER: This course has been made possible through an unrestricted educational grant from Store-A-Tooth. DESIGNA-TION STATEMENTS: Dental Learning, LLC is an ADA CERP recognized provider. ADA CERP is a service of the American Dental Association to assist dental professionals in identifying quality providers of continuing dental education. ADA CERP does not approve or endorse individual courses or instructors, nor does it imply acceptance of credit hours by boards of dentistry. Dental Learning, LLC designates this activity for 2 CE credits. Dental Learning, LLC is also designated as an Approved PACE Program Provider by the Academy of General Dentistry. The formal continuing education programs of this program provider are accepted by AGD for Fellowship, Mastership, and membership maintenance credit. Approval does not imply acceptance by a state or provincial board of dentistry or AGD endorsement. The current term of approval extends from 2/1/2016 - 1/31/2020. Provider ID: # 346890. EDUCATIONAL METHODS: This course is a self-instructional journal and web activity. Information shared in this course is based on current information and evidence. REGISTRATION: The cost of this CE course is $29.00 for 2 CE credits. ORIGINAL RELEASE DATE: February 2012. REVIEW DATE: January 2015. EXPIRATION DATE: December 2017. REQUIREMENTS FOR SUCCESSFUL COMPLETION: To obtain 2 CE credits for this educational activity, participants must pay the required fee, review the material, complete the course evaluation and obtain a score of at least 70%. AUTHENTICITY STATEMENT: The images in this course have not been altered. SCIENTIFIC INTEGRITY STATEMENT: Information shared in this continuing education activity is developed from clinical research and represents the most current information available from evidence-based dentistry. KNOWN BENEFITS AND LIMITATIONS: Information in this continuing education activity is derived from data and information obtained from the reference section. EDUCATIONAL DISCLAIMER: Completing a single continuing education course does not provide enough information to result in the participant being an expert in the field related to the course topic. It is a combination of many educational courses and clinical experi-ence that allows the participant to develop skills and expertise. PROVIDER DISCLOSURE: Dental Learning does not have a leadership position or a commercial interest in any products that are mentioned in this article. No manufacturer or third party has had any input into the development of course content. CE PLANNER DISCLOSURE: The planner of this course, Joe Riley, does not have a leadership or commercial interest in any products or services discussed in this educational activity. He can be reached at [email protected]. TARGET AUDIENCE: This course was written for dentists, dental hygienists, and assistants, from novice to skilled. CANCELLATION/REFUND POLICY: Any participant who is not 100% satisfied with this course can request a full refund by contacting Dental Learning, LLC, in writing. Please direct all questions pertaining to Dental Learning, LLC or the administration of this course to [email protected]. Go Green, Go Online to www.dentallearning.net take your course. © 2015

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Introduction

The term ”stem cell” dates back more than a cen-tury.1 Since then, clinicians and scientists have come to define stem cells as undifferentiated cells that

possess both the capacity for self-renewal and the ability to give rise to differentiated cells. Due to these properties, stem cells will have increasingly transformational uses in medicine and dentistry. The layperson often associates stem cells with “embryonic” stem cells that come from embryos. However, while the collection of embryonic stem cells is controversial, adult stem cells can be collected in noncon-troversial ways from various tissue niches within the body. These stem cells retain the properties that are important for clinical use. In fact, adult stem cells, such as those extracted from bone marrow transplants, have been used in medicine for over 50 years to treat hundreds of thousands of people, primarily patients with leukemia or genetic blood disor-ders. The use of bone marrow in medicine is an example of regenerative medicine.

The U.S. Department of Health and Human Services has identified regenerative medicine as an emerging field that will revolutionize health care treatment, potentially treat heretofore incurable diseases, combat rising healthcare costs, and serve as an important new area of U.S. competitive-ness.2 The interdisciplinary field of regenerative medicine is rapidly evolving, with a wide array of approaches to repairing, replacing, or enhancing biological function lost to congenital abnormalities, injury, disease, or aging.

Historically, identifying a potent, noncontroversial, accessible, relatively inexpensive, and convenient source of stem cells has been challenging—one solution is dental stem cells. Since scientists at the National Institutes of Health discovered these cells in 2000, dental profession-als and dental patients have become increasingly aware

of the emerging uses of both dental stem cells and dental stem cell banking services. Dental stem cells can be readily sourced from various dental tissues for potential autolo-gous use at a future date. Advances in stem cell research and interest in dental stem cells and banking services are increasing such that there are now policies relating to dental stem cells. In 2008, the AAPD Council on Clini-cal Affairs published a Policy on Stem Cells, which was revised in 2013.3 It provides a useful framework for dental professionals as the dental stem cell field emerges.

AAPD Policy on Stem CellsThe AAPD policy acknowledges that the public is ex-

pressing increasing interest in dental stem cell banking and “encourages dentists to follow evidence-based literature in order to educate patients about the collection, stor-age, viability, and use of dental stem cells with respect to autologous regenerative therapies.” The policy also high-lights that “while sources of dental stem cells are readily accessible, those cells must be secured and stored properly to maintain the potential to proliferate and differentiate.” Finally, the policy gives an overview of the properties of dental stem cells and highlights a number of conditions for which stem cell therapy in general is currently being clini-cally tested as a treatment for conditions including diabe-tes, Parkinson’s disease, and spinal cord injuries. Suggested regenerative dentistry applications mentioned include the repair of craniofacial defects as well as dental and peri-odontal tissues.

Stem Cell BasicsA simple way to conceptualize stem cells is to envision

the paths leading from embryogenesis to the development of each cell in a fully developed organism. Cells start with


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high potential in a fertilized egg and, in fact, are consid-ered at this point to be totipotent. This means that a single cell (for example, from a blastocyst) can give rise to a complete organism. In practice, embryonic stem cells are obtained at this point in development from embryos or blastocysts. As development proceeds, the cells divide and become increasingly specialized, developing into the four germ layers: ectoderm, mesoderm, endoderm, and germ layers (Figure 1). At this point the cells are multipotent, meaning that they still have the potential to differenti-ate into more limited arrays of cell types. Ultimately in a developed organism nearing or following birth, the vast majority of cells are oligopotent or unipotent—that is, they have terminally differentiated into the over 200 special-ized cell types making up the 50–75 trillion cells that do the work of the body (e.g., producing blood, bone, muscle, organs, and neurons).

Some stem cells remain in tissues after birth and into adulthood; they are by convention termed “adult” stem cells. These cells reside in specialized stem cell niches in various tissues. The niches preserve the capacity of stem cells to proliferate and differentiate into many cell types.4 In disease or injury, the body naturally relies in part on stem cell niches to help repair and regenerate tissues over one’s lifetime. Clinicians are increasingly using this knowl-

edge, for example by transplanting or mobilizing stem cells from their stem cell niche, to improve clinical outcomes. Practically speaking, the most accessible stem cell niches are bone marrow, the umbilical cord, adipose tissue, and dental pulp.5 Importantly for dental professionals, the dental stem cell niche is proving to be easy to access and contains a range of cell types that may be useful for regen-erative dentistry and medicine.

“Adult mesenchymal stem cells, of which dental stem cells are a subset, are highly proliferative and have the ability to pro-liferate and differentiate into many cell lines.” –AAPD policy

Sources of Stem CellsStem cells can be organized into two main sources:

embryonic stem cells sourced from embryos, and adult stem cells sourced from an adult stem cell niche (e.g., bone marrow, umbilical cord blood, dental pulp, adipose tissue). If it is necessary to have a discussion with a patient about stem cells before collecting them, it is important to realize that laypersons often oversimplify the field and associate all stem cells with embryonic stem cells. For completeness, this article addresses embryonic stem cells and then focuses on adult stem cells—particularly dental stem cells.

A. Embryonic Stem Cells and Induced Pluripotent Stem Cells

When human embryonic stem cells were first isolated in 1998,6 scientists and clinicians were very hopeful that this technology would provide a ready source of stem cells useful for the cure of most, if not all, diseases due to their pluripotent nature (the ability to form most if not all cell types). However, this has proven difficult in practice for many reasons. First, there are ethical, legal, moral, and social issues associated with embryonic stem cells—for ex-ample, determining the rights of the embryo, manifold is-sues around therapeutic cloning, and whether U.S. federal funding should be used to support this field. Second, there are significant practical clinical and engineering challenges to using pluripotent stem cells, including inadequate cell numbers, immunorejection (caused by a mismatch of the immune systems of the donor and recipient), and tumori-

Figure 1. Stem Cell Characteristics

Stem Cell Potency

Totipotent Pluripotent Multipotent Unipotent

Nails/Lens/EnamelCNS/PNS Spinal Cord/Brain

Skeleton Muscles/Heart Connective/Tissues Blood Tissues Red Cells Kidney Ovary/Uterus/Testes

Lung Liver Digestive System Pancreas


5January 2015

genesis (the pluripotent nature of embryonic stem cells that frequently leads to teratomas—tumors containing one or more of the three embryonic germ tissue layers).7 The first human clinical trials of embryonic stem cells in the United States were initiated in 2009 after the federal ban on embryonic stem cell therapy was lifted. Compared to adult stem cell approaches, which have been used safely and effectively for decades, the enormous challenges to using embryonic stem cells are clearly illustrated by the leading embryonic stem cell therapeutics company Geron. After investing millions of dollars in R&D for over 10 years, Geron not only terminated its embryonic stem cell clinical trial for spinal cord injury but also completely exited the business of embryonic stem cell therapeutic development.8

Partially addressing the ethical, legal, moral, and social issues associated with embryonic stem cells, genetic engi-neers in 2006 induced the first pluripotent states in unipo-tent cells by a technique known as induced pluripotent stem cell or iPS technology. Basically, genetic engineering could reset cells back to an embryonic-like state.9 While this technology avoids the ethical issues of having to extract embryonic stem cells and uses a patient’s own tissues (thus reducing immunologic incompatibility), iPS cells have also been shown to cause teratomas and may, in practice, be pro-hibitively expensive to generate on a personalized basis.10

B. Adult Stem Cells1. Bone Marrow Stem Cells

In contrast to embryonic stem cells or iPS cells, adult stem cells from bone marrow have been used clinically for over 50 years. In the late 1950s, bone marrow from one identical twin was used to treat the other twin’s leukemia.11 This work was inspired by research into the survival of irradiated dogs following nuclear radiation poisoning. The dogs survived because their spleens con-tained cells that repopulated their otherwise radiation-ablated blood systems. When applied to twins where one twin had leukemia and the other was healthy, initial bone marrow transplants restored the diseased twin’s health for a few months, but many of the first patients died from relapse of leukemia or graft-versus-host disease. Years

of research and clinical advancements led to successful transplants between non-twins in the late 1960s.12 In 1990, Dr. E. Donnall Thomas was awarded the Nobel Prize in Medicine for his pioneering work in bone mar-row transplantation.13

Today, clinical bone marrow stem cell treatments are commonplace; over 50,000 bone marrow transplants are performed each year,14 and more than 1,500 clinical trials related to bone marrow are planned, ongoing, or have been completed, treating conditions ranging from leukemia to spinal cord injury.15

2. Cord Blood Stem CellsIn the mid-1980s, a lack of matched donors available for

bone marrow transplants in children with leukemia moti-vated the alternative use of hematopoietic stem cells from umbilical cord blood. By 1988, the first cord blood stem cell transplant was used to treat a young boy with Fan-coni’s anemia.16 That patient remains alive and well to this date.17 Today dozens of diseases are treatable with fresh or cryopreserved umbilical cord blood. The properties of cord blood stem cells to treat leukemia or blood-related genetic diseases and to be readily collected and cryopreserved for future use formed the basis for the umbilical cord blood banking industry. To date, more than 9,300 patients have been treated with stem cells from umbilical cord blood, and over 200,000 units are publicly banked.18 In addition to public banks, parents of over 550,000 newborns have pri-vately banked their children’s umbilical cord blood at birth19 due to its proven use in treating leukemia and genetic blood disorders, as well as for its potential to treat other diseases such as myocardial infarction, diabetes mellitus, and neuro-logical disorders.20 The American College of Obstetricians and Gynecologists21 and the American Pediatric Associa-tion22 each have official policies or position statements on umbilical cord blood banking.

3. Adipose Tissue Stem CellsAdipose tissue obtained from liposuction aspirates

or abdominoplasty procedures can provide a source of mesenchymal stem cells. Stem cells from adipose tissue

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are being investigated for repairing tissue defects result-ing from traumatic injury, tumor resection, and congenital defects,23 as well as from calvarial defects following severe head injury,24 and in dentistry for maxillary and mandibu-lar repair.25

4. Dental Stem Cells“Pulpal tissue of primary teeth and surgically removed

third molars may serve as a source of mesenchymal stem cells.” –AAPD policy

In 2000, scientists at the National Institute of Dental and Craniofacial Research discovered that mesenchymal stem cells can be readily accessed from the dental pulp.26 Suspecting that a stem cell population was responsible for the similarities between dental pulp and bone mar-row, they identified mesenchymal stem cells in the pulp of extracted third molars. Termed dental pulp stem cells (DPSCs), these stem cells are more prolific than bone mar-row mesenchymal stem cells, and were shown to generate calcified nodules and neuron-like cells.27 Stem cells from

human exfoliated deciduous teeth (SHED),28 periodontal ligament stem cells (PDLSCs),29 stem cells from apical papilla (SCAPs),30 and dental follicle stem cells (DFSCs)31 were subsequently identified.

The Science of Dental Stem CellsHundreds of peer-reviewed scientific studies have

since examined dental stem cells in laboratory, animal, and human studies, showing that dental stems display typical mesenchymal stem cell characteristics.32-39 These include generating dentin-producing odontoblasts, adipocytes, osteoblasts, bone, cartilage, and smooth and skeletal muscle.27, 40, 41,42 Additionally, dental stem cells either contain or can switch lineage to form ectodermal tissues such as neurons43 and epithelial-like stem cells,44 as well as cells associated with the endodermal lineage, including endothelial cells,40 hepatocytes,45 and insulin-producing cells.46 In addition, populations of dental stem cells have been identified that express embryonic

Figure 2. Milestones in the Adult Stem Cell Field


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stem cell markers.47-49 In other words, stem cells found associated with teeth may contain many of the cell types required for regenerative medicine. The biology of odontogenesis provides some clues as to the existence of dental stem cells. Early in fetal development, teeth arise from the neural crest through a series of interactions between neural, mesenchymal, and epithelial tissues.50,51 The developed tooth can be thought of as a source of an encapsulated population of quiescent stem cells. It has also been shown that the pluripotency of dental stem cells52 may be a function of the age of the tooth or the age of its donor.53 In other words, primary teeth, molars, and wisdom teeth of young adults all contain potent sources of dental stem cells.

Current and Emerging Clinical Uses of Dental Stem Cells

In human clinical studies, dental stem cells have dem-onstrated proof of concept to regenerate alveolar bone54 and for periodontal disease.55 In animal studies, human dental stem cells have shown the potential to treat a wide variety of conditions including the ability to regenerate damaged pulp;56 reconstruct craniofacial defects;57 gener-ate lamellar bone;40 engineer new teeth;58 repair damaged corneas;59 treat liver disease;60 repair myocardial infarc-tion;61 and treat muscular dystrophy,62 stroke,63 and spinal cord injury.64 Dental stem cells have also proven capable of generating islet-like aggregates that secrete insulin in a

glucose-responsive manner65and of modulating immune responses,66 both of which may be important strategies for treating diabetes (Table 1).

A complete review of all the progress in this field is beyond the scope of this article.67 Therefore, this section focuses on two potentially groundbreaking uses of dental stem cells that illustrate the transformative role that dental professionals may play in this emerging field.

A. DiabetesToday, there is no cure for type 1 diabetes (insulin-

dependent or juvenile-onset diabetes). In the United States alone, nearly 15,000 children and teenagers are newly

Table 1. Potential Applications for Dental Stem Cells

Dental Applications Pulpal regenerationCraniofacial reconstructionEngineering of new teeth

Medical ApplicationsCorneal repairLamellar bone generationTreatment of liver diseaseCardiac repair following myocardial infarctionTreatment of muscular dystrophyTreatment following a strokeSpinal cord regenerationTreatment of diabetes

Figure 3. Dental Stem Cell Niches

Mixed Dentition Permanent Dentition

Dental Pulp Stem Cells

Stem Cells from Human

Exfoliating Deciduous Teeth

Dental Follicle Stem Cells

Periodontal Ligament Stem Cells

Stem Cells from Apical Papilla

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diagnosed each year.68 Standard treatment for type 1 diabetes includes lifestyle management, frequent blood glucose monitoring, and daily insulin injections or the use of insulin pumps. Although the discovery and use of insu-lin by Frederick Banting and Charles Best69 is one of the major breakthroughs in medical history, Banting himself recognized in his Nobel Lecture delivered in 1925 that: “Insulin is not a cure for diabetes; it is a treatment.” The expected total lifetime medical and indirect costs for those newly diagnosed is estimated at about $10 billion, and over $400 billion in costs would be avoided if the disease was eliminated.70

Stem cells are being aggressively investigated as part of a cure for type 1 diabetes. This research is focused on two fronts: inducing tolerance to pancreatic antigens,71 and developing islet-like cells that secrete insulin in a glucose-responsive manner for cell replacement therapy. Sourcing cells that will work on either or both fronts has historically remained a challenge. However, cells sourced from exfoliating or extracted teeth have been reported to possess potentially useful properties pertain-ing to both of these strategies: immunomodulation66,72,73 and islet generation.65 As a result, dental stem cells may represent an easily available source of stem cells for potentially both of these therapeutic approaches to type 1 diabetes. In the future, we may see patients’ own exfo-liating or extracted teeth (heretofore discarded) serving as the cellular starting materials for treating their own

diabetes.

B. Spinal Cord InjuryThe dental pulp originates from the neural crest,74 a

structure in the developing animal that gives rise to the autonomic nervous system including the spinal cord.75 It is therefore not a huge leap that neurons have been gener-ated from dental stem cells,76 including from PDLSCs,77 DPSCs,43 DFSCs,78 and SHEDs.79 More than 10 years ago, an animal model of spinal cord injury showed that dental stem cells increased the survival rates of motoneurons fol-lowing injury.64 When implanted into animals, dental pulp promotes proliferation, cell recruitment, and maturation of endogenous stem/progenitor cells by modulating the local microenvironment,80 and also secretes factors that induce the axon guidance of endogenous neurons.81 Continuing work since then has led to increased understanding of the techniques for inducing dental pulp into functional neural cells.82,83

Recently, dental pulp from human exfoliating decidu-ous teeth and from extracted third molars were studied in an animal model of a spinal cord injury and demonstrated multifaceted abilities to mediate the inflammatory process, direct axon growth, and even provide a source of neurons for cell therapy, resulting in the gain of significant hind leg motor function.84 Although significant hurdles remain before applying dental pulp becomes a routine clinical practice, dental stem cells may represent an easily available

Figure 4. A Typical Dental Stem Cell Banking Process


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source of stem cells for therapeutic treatments for spinal cord injuries. Compared to the earlier technology that uses embryonic stem cells, collecting dental stem cells is simpler, avoids the use of embryos, and is also less expensive.

The Basics of Stem Cell CryopreservationThe ready availability, simplicity, convenience, low cost,

and high potency of dental stem cells means that, as with stem cells derived from bone marrow, they may become just another logical and routine source for regenerative medicine.

Given that the clinical need for stem cells and the loss of teeth typically do not happen simultaneously, there is a need to store these tissues until they are needed. This can be ac-complished through cell cryopreservation, a well-established practice with, for example, bone marrow,85 cord blood,86 and fertilized embryos.87 Dental stem cells from the peri-odontal ligament,88 apical papilla,89 and dental pulp90,91 have been successfully cryopreserved while retaining their ability to differentiate into other tissue types. Dental stem cell banking works in the same manner as cord blood bank-ing, and these are complimentary. However, there is only one opportunity for an individual’s umbilical cord blood to be banked—at birth. The stem cells from cord blood are used primarily for hematopoietic uses, while stem cells from dental pulp are primarily mesenchymal stem cells and neuroprogenitors—thereby providing a complimentary cell

type for patients. The cryopreservation of dental stem cells simply represents the application of existing technology to a new source of stem cells.

After the dentist collects a tooth, it is transported to the laboratory in a sterile, isotonic solution. This package is shipped chilled to reduce the growth of contaminating microbes, and is delivered to the laboratory as quickly as possible. Although stem cells can successfully be recovered from teeth several days post-extraction, the yield declines significantly with time.92 The laboratory must have vali-dated processes with appropriate quality control metrics in place, verifying its ability to remove contaminating oral flora from the tooth and to recover viable stem cells. Ide-ally, the laboratory should have the ability to grow these cells in culture and to verify that the cultured cells exhibit the baseline set of cell surface markers expected for mesen-chymal stem cells.93

“While sources of dental stem cells are readily acces-sible, those cells must be secured and stored properly to maintain the potential to proliferate and differentiate.” –AAPD policy

Cryopreservation typically involves equilibrating the cells with a cryoprotectant solution, a solvent that protects the cells from the formation of ice crystals and helps pre-serve the integrity of cell membranes upon thawing. The temperature is typically slowly brought down to freezing using programmable controlled-rate freezers. The frozen

Figure 5. A Simple Guide to Help You Get Started with Dental Stem Cell Banking

Step 1. Deciden Decide if your practice

will offer dental stem cell banking services.

n Would your patients want to know?

n Will you do education or tooth collection or both?

Step 2. Select a VendorChecklist:n Professional supportn Patient education &

supportn Tooth collection &

transport processn Stem cell processing,

storage, reporting

Step 3. Get Startedn Delegate point personn Internal/external

practical plann Patient educationn Tooth collection

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cells are then transferred to vapor-phase liquid nitrogen freezers for long-term storage at ultra-low temperatures, typically at about –150°C. Freezing drastically slows down all biochemical processes, allowing the frozen cells to remain stable indefinitely.

The clinical use of cryopreserved tissues is regulated at both state and federal levels. Laboratories storing or expanding human cells for future clinical use must be registered with the FDA,94 licensed by the health department of the state where the laboratory operates, and should be accredited by the American Association of Blood Banks and/or the American Association of Tissue Banks.

Incorporation of Dental Stem Cell Services into Your Practice

“The American Academy of Pediatric Dentistry recog-nizes the emerging field of regenerative medicine and en-courages dentists to follow future evidence-based literature in order to educate parents about the collection, storage, viability, and use of dental stem cells with respect to au-tologous regenerative therapies.” –AAPD policy

Incorporating dental stem cell banking into any den-tal practice is surprisingly simple and boils down to two things: 1) informing patients that this is an option; and 2) helping patients with tooth collection. Since every prac-tice is unique, this section is intended to provide a general

guide to help you develop your own approach that works for your practice.

Step 1: Decide if you will incorporate dental stem cell banking into your practice.

Collectively, 20 to 30 million patients in the United States lose teeth every year—many of which are intact, vital teeth that can be suitable sources from which to obtain dental stem cells. This includes exfoliating or atraumatically extracted teeth (e.g., extracted third molars, teeth pulled in relation to orthodontic procedures). The enormous number of patients eligible for this new service means that dental professionals are in a unique position to be at the forefront of this field by building awareness about the option to store stem cells from teeth and collecting teeth for stem cell banking. All dental professionals can provide information to patients and the public about this important new field. General dentists and some specialists dealing with exfoliat-ing or extracted teeth will also be called on by their patients to assist with high-quality tooth collection.

Cost considerationsDifferent vendors provide different options for provid-

ers and patients. The costs to a dental practice may range from $0 to $90 or more per year. The cost to patients for privately banking their dental stem cells is typically around $600 for the first year plus an annual charge of roughly

Table 2. Dental Professionals Play a Number of Roles from Education to Providing High-quality Service

SpecialtyPatient/Parent

EducationTooth Collection

Exfoliating Ortho 3rd molars

Pediatric/General/Family Dentist ✔ ✔ ✔ ✔

Orthodontist ✔

Oral Maxillofacial Surgeon ✔ ✔ ✔

Hygienist ✔

Periodontist ✔

Endodontist ✔

Dental Assistant/ Office Staff ✔


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$100 for cryopreservation. Some companies provide a range of pricing and financing plans that provide families with more options. For reference, dental stem cell banking is substantially less expensive than private umbilical cord blood banking which is typically $1,000–2,000 for the first year and $100–150 per year after that.

Step 2: Considerations when selecting a vendorOnce you decide to offer patients the opportunity to

bank their dental stem cells, the next step is to select a ven-dor. The AAPD’s Policy on Stem Cells may help provide a useful framework during your vendor selection and evalu-ation process. Below are a few factors to consider:• Professional support. Look for the availability of dedi-

cated professional support to help your practice and to coordinate with your point person. This can make the difference between success and failure. Training, continuing education courses, webinars, and progress updates in the field are important for keeping current.

• Patient education. Look for the availability of simple and accurate tools for patient education from vendors such as informational brochures, videos, online Web tools, and newsletters. These can greatly simplify aware-ness building in your patient base. Consider the level of customer support you want from a vendor to assist those patients who want more information.

• Tooth collection and transport. The AAPD policy high-lights the need for appropriate informed consent and patient privacy. Ask about how the company transports your patient’s tooth to the lab while maintaining high cell viabilities.

• Stem cell harvesting and storage. Ultimately the cells collected may be used clinically. Ask how the company meets industry standards for human tissue banking for clinical use, and how they will provide you and or your patient with evidence that the stem cells were properly stored.

Step 3: Incorporate dental stem cell services into your everyday practice.

Now you are ready to incorporate dental stem cell ser-

vices into your everyday practice workflows and deal with patient education and tooth collection.

Patient education: Patients appreciate being informed about this option by their dental provider in time to make an informed decision about banking their own stem cells. Like any other service you provide, patients will expect the dental team to be current and provide them with informa-tion on dental stem cell banking. Accurate information becomes especially important if a patient’s family has the types of diseases discussed above, such as diabetes, where dental stem cells may play an important role in their future health. These patients and their families will be particu-larly grateful to your team for providing them with this information. Your vendor can provide you with support and materials to help you streamline patient education. In turn, providing this information to patients is not only potentially valuable for their health, it may also help retain patients and attract new ones.

The layperson already has a general idea of what stem cells are; therefore, patient education can be streamlined. For practices that see children, new patient orientation, newsletters, or hygiene visits prior to a tooth exfoliating can all be good opportunities for sharing information. When teeth need to be extracted as part of a treatment plan, the initial treatment planning visit and the extrac-tion consultation are both examples of good times when information fits naturally into the process. This gives patients time to consider their options between when they first learn they are eligible to when they need to bring their tooth collection kit to your office.

Tooth collection. Instead of being discarded, stem cells collected during the normal course of dental care can be readily cryopreserved for future clinical applica-tion.42,49,95,96,97 As a result, dental stem cells may present the simplest, least expensive way for families and medical professionals to collect potent stem cells from each person. When it comes to tooth collection, pulpal viability should be evaluated to decide if each tooth is a good candidate for banking. Different dental stem cell banking vendors approach this issue differently. Consult with your vendor about which teeth provide high-quality banked samples.

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Evidence suggests that dental stem cells may be more proliferative the younger the tooth/patient is, although patients with teeth extracted in their 30s and beyond can still be eligible. Typically third molars yield a higher total number of cells than primary teeth about to exfoliate, especially when multiple teeth are extracted as part of the treatment plan. Getting started.

As with other services in your practice, having a des-ignated point person in the practice who can work with patients, your team, your network of referring providers, and vendors can make it easier to get started. This person can be a dentist, hygienist, office manager, patient coordi-nator, or experienced dental assistant—anyone on the team who is enthusiastic about this service can be a good point person. Depending on your practice philosophy, this is also an opportunity to develop a participatory process where the dentists and hygienists may develop or co-develop a plan with the extended team for dental stem cell banking. Practical considerations include determining how you will let your patients, your referral network, and the outside world know that you offer this service.

“The public is increasingly aware of this emerging sci-ence, and more parents are expressing interest in harvest-ing/banking dental stem cells.” –AAPD policy

ConclusionsInterest in dental stem cells for regenerative dentistry

and medicine continues to increase as more research demonstrates their viability for the potential treatment of a range of diseases and conditions. The technology to collect and store tissues that contain these cells for future use is proven and available today. Millions of people will become eligible for tooth banking this year. Dental professionals are ideally positioned to help patients by providing both education and dental stem cell banking services, thereby giving patients the opportunity to store stem cells from vi-able teeth for future use.

TerminologyAllogeneic Use: This use requires a human leukocyte antigen (HLA) match; the donor and the recipient are two

different people. Autologous Use: The donor is the same person as the recipient (biological definition) or is a close blood rela-tive (FDA definition); the implantation, transplantation, infusion, or transfer of human cells or tissue back into the individual from whom the cells or tissue were recovered ensures an HLA match.Biomarker: A characteristic that is objectively measured and evaluated as an indicator of normal biologic processes, pathogenic processes, or pharmacologic responses to a therapeutic intervention. Frequently biomarkers are mea-surable proteins such as DNA or RNA. Cryopreservation: The process of using very low tempera-tures, typically around -300° F (-150° C), to keep cells preserved for future use. Differentiation: The process by which a less specialized cell becomes a more specialized cell. Flow Cytometry: A technique for measuring the presence of various biomarkers associated with cells. Hematopoietic: Blood forming.Homologous Use: The repair, reconstruction, replacement, or supplementation of a recipient’s cells or tissues with those that perform the same basic function(s) in the recipi-ent as in the donor. Human Leukocyte Antigen: The major histocompatibility complex in humans. HLAs on the surface of cells inform the immune system as to the contents of the cell (class I) and its environment (class II), and are therefore critical to the success of cell and organ therapies. Mesenchymal: The type of tissues arising from the me-soderm germ layer (e.g., myogenic or muscle forming, osteogenic or bone forming, adipogenic or fat forming, chondrogenic or cartilage forming). Multipotent: Capable of developing into many cell or tis-sue types. Pluripotent: A type of stem cell that has the potential to differentiate into endoderm, mesoderm, or ectoderm. Pluripotent stem cells can give rise to any fetal or adult cell type, but cannot develop into a fetal or adult animal because they lack the potential to contribute to extra-em-bryonic tissue, such as the placenta.


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Regenerative Medicine: The process of creating living, functional tissues to repair or replace tissue or organ func-tion lost due to age, disease, damage, or congenital defects. Stem Cell: An unspecialized cell that has the potential to develop into many different cell types and that, under certain conditions, can be induced to become a tissue- or organ-specif-ic cell.Totipotent: A type of stem cell that is capable of differenti-ating into all cell types. Totipotent stem cells can even form a complete organism. Unipotent: Capable of developing into only one type of cell or tissue. Vitrification Agent: This is added to the cryopreservation solution prior to freezing In order to prevent cell death due to the freezing process. It reduces the formation of ice crys-tals that can damage cells in the freeze-to-thaw process. This improves the post-thaw viability of cells.

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60. Ikeda E, Yagi K, Kojima M, et al. Multipotent cells from the human third molar: feasi-bility of cell-based therapy for liver disease. Differentiation. 2008 May;76(5):495-505.

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62. Kerkis I, Ambrosio CE, Kerkis A, et al. Early transplantation of human immature dental pulp stem cells from baby teeth to golden retriever muscular dystrophy (GRMD) dogs: local or systemic? J Transl Med. 2008 Jul 3;6:35.

63. Yang KL, Chen MF, Liao CH, Pang CY, Lin PY. A simple and efficient method for generating Nurr1-positive neuronal stem cells from human wisdom teeth (tNSC) and the potential of tNSC for stroke therapy. Cytotherapy. 2009;11(5):606-617.

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15January 2015

1. __________ is a source of stem cells.a. Bone marrowb. Dental pulpc. Adipose tissued. all of the above

2. Stem cells __________.a. are only unipotentb. differentiate into specific tissue typesc. can be banked for future used. b and c

3. Dental stem cells were first discovered by scientists at the Na-tional Institutes of Health in __________.a. 1990b. 1995c. 2000d. 2005

4. In 2008, the __________ published a Policy on Stem Cells. a. American Dental Association Council on Clinical Affairsb. American Academy of Periodontology Council on Clinical Affairsc. American Academy of Pediatric Dentistry Council on Clinical Af-fairsd. all of the above

5. The AAPD policy ‘encourages dentists to follow evidence-based literature in order to educate patients about the __________ of dental stem cells with respect to autologous regenerative therapies.’ a. collection b. bankingc. used. all of the above

6. Adult stem cells are those stem cells that remain in tissues __________.a. until birthb. after birthc. after birth and into adulthoodd. all of the above

7. In disease or injury, the body naturally relies in part on _________ to help repair and regenerate tissues over one’s lifetime. a. embryonic stem cellsb. stem cell nichesc. blood cellsd. a and b

8. Dental stem cells from the pulp __________.a. are more prolific than bone marrow mesenchymal stem cells b. can only be sourced when the patient is a young childc. are readily accessible compared to other sources of stem cellsd. a and c

9. Adult mesenchymal stem cells, of which dental stem cells are a subset, display __________.a. inherent plasticityb. the ability to proliferatec. the ability to differentiate into many cell linesd. all of the above

10. __________ is/are a significant practical challenge to the wide-spread use of embryonic stem cells. a. Immunorejectionb. Tumorigenesisc. Ethical and moral issuesd. all of the above

11. Clinical bone marrow stem cell treatments __________.a. have been used for over 50 yearsb. are commonplace with over 50,000 bone marrow transplants

per year c. are used to treat diseases such as leukemiad. all of the above

12. Cord blood stem cells __________.a. are collected from the umbilical cord at birth for bankingb. are clinically proven in the treatment of leukemia and genetic

blood disorders c. have potential to treat diseases such as myocardial infarction,

diabetes mellitus, and neurological disordersd. all of the above

13. __________ is/are a source of dental stem cells. a. Human exfoliating deciduous teethb. The periodontal ligamentc. The apical papillad. all of the above

14. Dental pulp has been shown to contain __________.a. mesenchymal stem cellsb. cells of endodermal lineagec. neuro-progenitorsd. all of the above

15. Dental stem cells have been found to __________.a. have only one possible lineageb. be able to switch lineagec. be difficult to sourced. be difficult to bank

16. Recently, dental pulp from human exfoliating deciduous teeth and from extracted third molars demonstrated the ability to __________ in animal studies.a. mediate the inflammatory process b. direct axon growth c. provide a source of neurons for cell therapyd. all of the above

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17. __________ teeth are __________ potent sources of stem cells than __________ teeth.a. Geriatric; more; youngerb. Geriatric; less; youngerc. Younger; less; embryonicd. none of the above

18. Clinical proof of concept in humans for using dental stem cells to regenerate tissues __________.a. does not existb. has been shown for alveolar bone regenerationc. has been shown for diabetesd. a and b

19. Shipping the sample _________ helps reduce the growth of contaminating microbes. a. frozenb. lukewarmc. chilledd. any of the above

20. The dental pulp originates from the neural crest, a structure in the developing animal that gives rise to the __________.a. autonomic nervous systemb. the spinal cordc. a and bd. none of the above

21. Cells sourced from __________ teeth have been reported to possess potentially useful properties for immunomodulation and islet generation. a. exfoliating or extractedb. degeneratedc. periodontally-involvedd. apically infected

22. Stem cells can be saved for future use through the process of __________. a. cyanopreservationb. cryodessicationc. cryopreservationd. all of the above

23. Incorporating dental stem cell banking into any dental practice boils down to two things: _________.a. informing patients this is an option and helping patients with

tooth collectionb. informing patients this is an option and referring them for

tooth collectionc. informing patients only if they are old and helping these patients

with tooth collectiond. a and b

24. Dental stem cell banking works in the same manner as _________. a. bone marrow bankingb. cord blood bankingc. blood banksd. all of the above

25. After a tooth is collected by the dentist for dental stem cell banking, the sample is transported to the laboratory in _________ solution. a. a sterile, isotonicb. an alkalinec. an acidicd. none of the above

26. Dental stem cells have also been shown to __________.a. be capable of generating islet-like aggregates that secrete

insulinb. modulate immune responsesc. elevate glucose levelsd. a and b

27. The laboratory where dental stem cells will be banked in place _________.a. must have validated processes with appropriate quality control

metricsb. should have the ability to grow these cells in culturec. should have the ability to verify that the cultured cells exhibit the

baseline set of cell surface markers expected for mesenchymal stem cells

d. all of the above

28. Cryopreservation typically involves equilibrating the cells with a cryoprotectant solution that _________.a. protects the cells from the formation of ice crystalsb. helps preserve the integrity of cell membranes upon thawingc. protects the solutiond. a and b

29. Laboratories storing or expanding human cells for future clinical use must be registered with the _________.a. ADAb. EPAc. FDAd. all of the above

30. Laboratories storing or expanding human cells for future clinical use should be accredited by the _________.a. American Association of Blood Banksb. American Association of Tissue Banksc. both a and bd. a and/or b

CE QUIZ

17January 2015


www.dentallearning.net/DSC-ceCE ANSWER FORM (E-mail address required for processing)

*Name: Title: Speciality

*Address: NPI No.

*City: *State: *Zip: AGD Identification No.

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*Telephone: License Renewal Date:

Please direct all questions pertaining to Dental Learning, LLC or the administration of this course to [email protected]. COURSE EVALUATION and PARTICIPANT FEEDBACK: We encourage participant feedback pertaining to all courses. Please be sure to complete the evaluation included with the course. INSTRUCTIONS: All questions have only one answer. Participants will receive confirmation of passing by receipt of a verification certificate. Verification certificates will be processed within two weeks after submitting a completed examination. EDUCATIONAL DISCLAIMER: The content in this course is derived from current information and research based evidence. Any opinions of efficacy or perceived value of any products mentioned in this course and expressed herein are those of the author(s) of the course and do not necessarily reflect those of Dental Learning. Completing a single continuing education course does not provide enough information to make the participant an expert in the field related to the course topic. It is a combination of many educational courses and clinical experience that allows the participant to develop skills and expertise. COURSE CREDITS/COST: All participants scoring at least 70% on the examination will receive a CE verification certificate. Dental Learning, LLC is an ADA CERP recognized provider. Dental Learning, LLC is also designated as an Approved PACE Program Provider by the Academy of General Dentistry. The formal continuing education programs of this program provider are accepted by AGD for Fellowship, Mastership, and membership maintenance credit. Please contact Dental Learning, LLC for current terms of acceptance. Participants are urged to contact their state dental boards for continuing education requirements. Dental Learning, LLC is a California Provider. The California Provider number is RP5062. The cost for courses ranges from $19.00 to $90.00. RECORD KEEPING: Dental Learning, LLC maintains records of your successful completion of any exam. Please contact our offices for a copy of your continuing education credits report. This report, which will list all credits earned to date, will be generated and mailed to you within five business days of request. Dental Learning, LLC maintains verification records for a minimum of seven years. CANCELLATION/REFUND POLICY: Any participant who is not 100% satisfied with this course can request a full refund by contacting Dental Learning, LLC in writing or by calling 1-888-724-5230. Go Green, Go Online to www.dentallearning.net to take this course. © 2015

PLEASE PHOTOCOPY ANSWER SHEET FOR ADDITIONAL PARTICIPANTS.

QUIZ ANSWERSFill in the circle of the appropriate answer that corresponds to the question on previous pages.

EDUCATIONAL OBJECTIVES1. Delineate key elements of the AAPD policy on stem cells and how this translates into daily practice.2. List the types, sources, and basic properties of stem cells.3. List and describe the range of potential clinical uses for dental stem cells and the current status of these

in type 1 diabetes and spinal cord injuries.4. Describe the key elements involved in discussing dental stem cell banking services with patients and in

providing these services.

If you have any questions, please email Dental Learning at [email protected] or call 888-724-5230.

COURSE SUBMISSION: 1. Read the entire course.2. Complete this entire answer sheet in

either pen or pencil.3. Mark only one answer for each question.4. Mail answer form or fax to 732-303-0555. For immediate results:1. Read the entire course.2. Go to www.dentallearning.net/DSC-ce.3. Log in to your account or register to create an

account.4. Complete course and submit for grading to

receive your CE verification certificate.

A score of 70% will earn your credits.

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Price: $29 CE Credits: 2Save time and the environment by taking this course online.

COURSE EVALUATIONPlease evaluate this course using a scale of 3 to 1, where 3 is excellent and 1 is poor.

1. Clarity of objectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 2 1

2. Usefulness of content . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 2 1

3. Benefit to your clinical practice . . . . . . . . . . . . . . . . . . . . 3 2 1

4. Usefulness of the references . . . . . . . . . . . . . . . . . . . . . . 3 2 1

5. Quality of written presentation . . . . . . . . . . . . . . . . . . . . 3 2 1

6. Quality of illustrations . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 2 1

7. Clarity of quiz questions . . . . . . . . . . . . . . . . . . . . . . . . . 3 2 1

8. Relevance of quiz questions . . . . . . . . . . . . . . . . . . . . . . 3 2 1

9. Rate your overall satisfaction with this course . . . . . . . . 3 2 1

10. Did this lesson achieve its educational objectives? Yes No

11. Are there any other topics you would like to see presented in the future? __________________________________________________________________________

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