Stem Cells

Stem Cells 101
How these powerful entities operate and some recent developments in stem cell biology


REVIEW SUMMARY

1. Stem cells (SCs) are divided into two main types:


Embryonic stem cells derive from the blastocyst and can, in theory, give rise to all cell types in an organism.

Adult stem cells derive from mature tissue and usually have a restricted spectrum of possible differentiation (e.g., epidermal stem cells).


2. Stem cells have two cardinal features:


The ability to self-renew indefinitely through cell division

The ability to differentiate into varying cell types


3. Biologists have begun to unravel the genetic determinants that dictate "stemness."

4. Normal skin fibroblasts can be "reprogrammed" into a stem-cell–like phenotype (induced pluriopotent stem cells) through the introduction of a limited number of genes.

5. Stem cell therapy is theoretically ideal for degenerative human disorders; proof-of-principle has already been established in mice models of disease.

Stem cells have been a source of tremendous promise and controversy during the past decade. Some contention derives from conflicts between secular and religious definitions of life. Another source of controversy is the imprecise use of stem cell terminology and concepts. The purpose of this brief review is to clarify principles and showcase some recent advances in SC biology.

FUNCTIONAL ATTRIBUTES

SCs have traditionally been defined by their functional attributes. There are two cardinal features of all SCs. The first is self-renewal — the capacity to go through indefinite cycles of cell division while maintaining an undifferentiated state. To self-renew, SCs undergo either symmetric division or asymmetric division. In symmetric division, a stem cell gives rise to two identical daughter cells, both endowed with SC properties:

A A + A*
In asymmetric division, an SC gives rise to one stem cell and one progenitor cell with limited self-renewal potential:

A A + B*
Progenitor cells have the capacity to undergo several cycles of cell division,

B B + B*
but unlike stem cells, will eventually differentiate terminally.

B C + C*
(*A = adult stem cell; B = progenitor cell; C = terminal, differentiated cell)

The mechanisms that govern the activation of symmetric versus asymmetric division and that determine which daughter cells will be stem cells and which will be progenitor cells are still unclear.

The second cardinal feature of all SCs is potency — the ability to differentiate into specialized cell types. There are four major types of potency:


Totipotency is the ability of a single cell to expand in number, differentiate into embryonic and extraembryonic tissues, and develop into an organism.

Pluripotency is the ability to differentiate into any of the three germ layers: endoderm, mesoderm, or ectoderm.

Multipotency is the capacity to produce a related family of cells (e.g., hematopoietic SCs differentiate into various blood cell types).

Unipotency is the ability to differentiate into just one cell type (e.g., epidermal SCs generate only keratinocytes). Different sources and types of SCs may harbor different types of potency.


STEM CELL CLASSES
The two large classes of SCs are embryonic stem cells (ESCs), which are found in developing blastocysts, and adult stem cells (ASCs), which are found in mature tissues. ESCs derive from fertilized embryos; specifically, they are isolated from the epiblast tissue of the inner cell mass of a blastocyst or earlier morula. In humans, a blastocyst comprises about 50 to 150 cells and is approximately 4 to 5 days old. ESCs conform to the definition of pluripotency, in that they can develop as all three primary germ layers: ectoderm, endoderm, and mesoderm. ESCs do not contribute to the extraembryonic membranes or the placenta. Although there are phenotypic similarities between mouse and human ESCs, each requires very distinct experimental conditions for study.

ASCs can be found in the developed organism in both children and adults. ASCs are also known as somatic SCs and are often lineage-restricted (i.e., multipotent but not pluripotent). In the fully formed organism, ASCs are conventionally described according to the tissue of origin (e.g., epidermal SCs, adipose SCs, mesenchymal SCs). For instance, a certain type of ASC, the hematopoietic stem cell (HSC), has been used for decades in the treatment of certain bloodborne cancers. Bone marrow transplantation is successful because HSCs transfer and engraft. Michele De Luca and colleagues recently applied SC technology to treatment of junctional epidermolysis bullosa (JW Dermatol Jan 26 2007 and Nat Med 2006; 12:1397). From palmar skin, they isolated putative SCs, into which they introduced the defective laminin 5 (LAM5) subunit and used the engineered cells to prepare genetically corrected cultured epidermal grafts. The autologous grafts showed synthesis and proper assembly of normal levels of functional LAM5. Even after a year, the engineered epidermis remained stable and adherent without blisters, infections, inflammation, or immune response. Although longer-term follow-up with more patients is needed to validate this study, the use of ASCs for certain targeted tissues is appealing, as it does not involve embryos.

IDENTIFYING STEM CELLS AND STEM CELL UTILITY
The identification and isolation of SCs has been challenging. No single fixed criterion offers indisputable evidence of "stemness." The sine qua non of an SC is its ability to regenerate a certain tissue over the life of the organism; thus, SCs are particularly attractive therapeutic models for degenerative diseases, such as neuronal degeneration in Parkinson disease or pancreatic degeneration in diabetes. An extant example of SC utility is the reconstitution of a complete immune system from bone marrow, which, presumably, contains hematopoietic SCs. In vitro assays help us understand the cellular behavior of SCs. For example, in the well-established clonogenic assay, SCs are characterized by their capacity to form colonies in tissue culture plates when seeded at low density. By contrast, committed cells that have limited replicative or survival potential will not undergo enough rounds of cell division to form clonal colonies. More-recent work has attempted to identify SCs based on protein or RNA markers. For instance, ESCs have been isolated and found to express transcription factors that appear to maintain pluripotency (Cell 2005; 122:947). The ability to point to the master regulators of "stemness" has been leveraged to create a new generation of induced SCs from adult tissue.

STEM ALCHEMY: INDUCED PLURIPOTENT STEM CELLS
In the Middle Ages, the art of transforming common metals into gold became a source of great mystical enthusiasm (and ultimate disappointment). Given the controversial nature and rarity of ESCs, the transformation of somatic cells into ESCs has similarly engendered great excitement in the past few years. Unlike the alchemists, however, current researchers have had some successes in SC engineering that may eventually find a therapeutic outlet.

The theoretical framework for production of induced pluripotent stem cells (iPSCs) is relatively straightforward: Identify a set of genes that define the "stemness" of ESCs and then "reprogram" adult cells as ESC-like cells by introducing these genetic factors into the somatic cells. Induced PSCs were first generated in mice in 2006 (Cell 2006; 126:663). Initial observations showed that four key genes were essential for reprogramming differentiated cells into the pluripotent state: Oct-3/4, Sox2, c-Myc, and Klf4. Despite similarities with ESCs, early iPSC lines failed to produce viable chimeras when injected into mouse embryos; thus, they do not fulfill the in vivo criteria for ESCs. Chimeras are animals that develop from different genetic sources — in this case, animals derived from both the host and the iPSCs. Subsequent refinements in technique have led to the successful reprogramming of mouse fibroblasts into iPSCs that can, in fact, produce viable chimeras (Nature 2007; 448:313 and Nature 2007; 448:318), thereby fully substantiating their pluripotent nature.

Induced PSCs have been shown to be similar to ESCs in morphology, growth properties, and the expression of SC genes and SC markers. Most critically, iPSCs can be triggered to differentiate into cardiac and neural cells as well as form viable mouse chimeras. A major concern among iPSC investigators is the use of c-Myc — a known oncogene — in the reprogramming; in fact, a significant fraction of the derived mice later developed cancer. A more recent protocol may make it possible to dispense with c-Myc (Nat Biotechnol 2008; 26:101).

Figure 1: Skin fibroblasts as the source of pluripotent human stem cells of the future
Reprinted with permission from Macmillan Publishers Ltd: Nature Medicine copyright 2007.

Differentiated cells, such as skin fibroblasts, are isolated and genetically reprogrammed to become iPSCs via the introduction of certain genes such as POU5F1, MYC, KLF4, and SOX2. These genes in turn induce the expression of SC master regulators, POU5F1 and NANOG. The iPSCs can then be recovered and redifferentiated into therapeutically regenerative cells and tissue. For details, see Nat Med 2007; 13:783.

THERAPEUTIC IMPLICATIONS
So what are the potential therapeutic implications of iPSCs? No human trials of iPSCs are under way, but at MIT, Rudolf Jaenisch and colleagues have recently shown that fibroblasts can be reprogrammed into fully functional blood cells in a humanized mouse model of sickle cell anemia (Science 2007; 318:1917). The same group also showed that iPSCs can be efficiently differentiated into neural precursor cells; transplanted into the fetal mouse brain; and successfully form glia and neurons, including glutamatergic, GABAergic, and catecholaminergic subtypes. In addition, when transplanted into the adult brain in a rat model of Parkinson disease, these iPSCs were able to improve behavior (Proc Natl Acad Sci U S A 2008; 105:5856). Studies involving skin disease will undoubtedly be published over the course of the next few years.

The National Institutes of Health provide a good introductory discussion of stem cells and a useful glossary of related terms.

— Hensin Tsao, MD, PhD

Published in Journal Watch Dermatology June 13, 2008

UV Light and changes in nevi

This article looked at the changes in benign nevi after exposure to narrow band UVB and UVA. Both modalities caused significant dermatoscopic changes that should be taken into consideration if monitoring nevi in sun exposed areas.

J Am Acad Dermatol. 2008 May;58(5):763-8. Epub 2008 Jan 30.
Repeated equally effective suberythemogenic exposures to ultraviolet (UV)A1 or narrowband UVB induce similar changes of the dermoscopic pattern of acquired melanocytic nevi that can be prevented by high-protection UVA-UVB sunscreens.Manganoni AM, Tucci G, Venturini M, Farisoglio C, Calzavara-Pinton PG.
Department of Dermatology, University of Brescia, Brescia, Italy.

BACKGROUND: Sunlight modifies the size and the dermoscopic pattern of acquired melanocytic nevi (AMN). OBJECTIVE: We investigated whether repeated exposures to equally effective suberythemogenic doses of ultraviolet (UV)A or UVB can induce changes in the dermoscopic features of AMN. METHODS: Twenty volunteers received equally effective doses of narrowband UVB or UVA1. During exposures, an AMN was covered with an opaque tape, another was shielded with the sunscreen, and another was left unprotected.

RESULTS: Nevi exposed to either narrowband UVB and UVA1 showed statistically significant changes in their dermoscopic features: increased size, increase of pigment network, overall color darkening, formation of focal branched streaks, and increased number and size of brown dots and globules. LIMITATIONS: The study is a clinical cohort study on a small number of selected patients. CONCLUSION: AMN show similar changes in size and dermoscopic pattern after narrowband UVB and UVA1 exposures.

Blink versus Systematic examination in Dermoscopy

Melanoma mimicking seborrheic keratosis: an error of perception precluding correct dermoscopic diagnosis.Braga JC, Scope A, Klaz I, Mecca P, Spencer P, Marghoob AA.
Department of Dermatology, Memorial Sloan-Kettering Cancer Center, New York, New York 10022, USA.

Seborrheic keratosis is a common skin lesion that can usually be recognized either clinically or dermoscopically. However, melanomas mimicking seborrheic keratoses, as well as melanomas arising in association with seborrheic keratoses, have been described. We report the case of a patient with a lesion that initially revealed "classic" dermoscopic features of a seborrheic keratosis. However, during follow-up, changes in color developed within the center of the lesion that led the clinician to the correct diagnosis of melanoma. Upon retrospective evaluation of the baseline image of the lesion; the clinician was now able to "see" that which his brain could not appreciate on initial examination and to realize that the lesion had subtle features suspect for melanoma. This case represents a diagnostic pitfall due to errors in perception. Dermatologists should be cognizant of "errors in perception"; we suggest that a final dermoscopic judgment of a seborrheic keratosis be rendered by combining the gestalt diagnosis of the overall pattern, with deliberate dermoscopic analysis of all quadrants of the lesion.

PMID: 18328596 [PubMed - indexed for MEDLINE]
From Club Dermaweb
A dermatologist’s brain will always favour the overall impression of the right side of the brain over the detailed analysis of the left side of the brain, both with a dermoscope as well as with the naked eye. The first is intuitive, fast and often very beneficial and efficient. Detailed rational analysis, which involves looking at the tumour section by section and studying all the basic signs and dermoscopic patterns, is tedious and slow. It is therefore logical that the dermatologist’s first impression tends to influence the way he/she interprets the detailed analysis, or even convince them not to do it. In this example a pigmented lesion of the neck found during a routine examination was quickly identified as seborrheic keratosis because he/she found numerous pseudo-horny cysts. When the patient was seen a year later, the lesion was a lot darker and had a blue-white veil. Excision was performed and an SSM associated with seborrheic keratotic-like epidermal hyperplasia was found, even though it wasn’t possible to distinguish between a verrucous melanoma and a composite lesion. The dermatologist looked at the initial dermoscopic image again and noticed that some aspects which suggested MM were already present a year earlier in a limited area at the periphery of the lesion. Papillomatous epidermal hyperplasia with pseudo-horny cysts has already been found in a small number of cases of malignant melanomas and does not allow this diagnosis to be officially ruled out. The error in perception which makes dermatologists favour the overall picture in relation to detailed analysis should be kept in mind to avoid mistakes like this.

Sidedness and Melanoma

Left-Sided Excess in the Laterality of Cutaneous Melanoma
Jean-Luc Bulliard, PhD; Silvia Ess, MD; Andrea Bordoni, MD; Isabelle Konzelmann, MD; Fabio Levi, MD


Arch Dermatol. 2008;144(4):556-558.

An unequal distribution of cancer laterality, particularly in paired organs, has long been documented and generally thought to be related to asymmetries in organ size or behavioral factors such as handedness.1 Recently in a large series patients with cancers in the left testis, right lung, and left ovary were found to have a significantly better survival than those with contralateral cancers.2 Apart from anecdotal assertions and very sparse data that suggest asymmetrical differences in the frequency of cutaneous melanoma and photodamage,3-4 melanoma laterality has, to our knowledge, never been specifically studied. Investigation of laterality could thus contribute to a better understanding of cancer etiology and prognosis.

Methods

As part of a larger study,5 the laterality of 2143 first cutaneous melanomas was retrieved and clinically validated using a standardized body chart that allowed unequivocal marking of the location of the lesion.6 After excluding cases with unspecified laterality (n = 228 [11%]) or those on the midline (n = 254 [12%]), 1661 melanomas diagnosed between 1995 and 2002 in 5 Swiss population-based tumor registries (Neuchâtel, St Gallen/Appenzell, Vaud, Valais, and Ticino) were investigated. Results were expressed as left to right (L/R) ratios and stratified by cancer registration area, sex, age group, and subsite. Exact 2-sided 95% confidence intervals (95% CIs) were computed assuming that laterality was binomially distributed.


Results

This series included 890 left-sided and 771 right-sided melanomas, yielding an L/R ratio of 1.15 (95% CI: 1.05-1.27). The excess of left-sided lesions was consistently observed across all populations, sexes, age groups, body site, and categories of Breslow thicknesses (Table), although it only occasionally reached statistical significance. The upper limbs was the site with the greatest left-sided excess (27%). Left to right ratios higher than 1 were systematically found for clinical characteristics such as tumor behavior (invasive and in situ), skin type, and morphological type (data not shown).
Comment

This multicentric study suggests a moderate but consistently higher frequency of melanoma on the left side of about 15%. Four main potential explanations were identified and explored: chance finding, recording bias, differential sun exposure, and bilateral asymmetry in the number of melanocytes or tumor biological behavior.

Although chance finding cannot be excluded, we believe it is an unlikely explanation for our observation. The pattern was similar for every variable studied and, for instance, the probability of observing simultaneously an excess L/R ratio in all 5 populations was about 3% (1 in 32). The detailed site was thoroughly cross-validated from textual and pictorial support.6

Two nonmethodological explanations for a left-sided excess of melanoma can be speculated. Traveling in a motor vehicle is probably the only frequent human activity that results in side-specific UV exposure depending on the individual position in the car. Swiss drivers sit on the left side of the car and, until the recent availability of air conditioning, their left arm was more likely to be sun exposed through an open window, particularly in summertime. The largest left-sided excess observed for the upper limbs (an L/R ratio of 1.27, 95% CI: 1.05-1.54,Table) and the greater L/R ratio for men (an L/R ratio of 1.38, P = .02, data not shown) than women (an L/R ratio of 1.18, P = .22, data not shown) at this site supports this assumption and the known greater propensity for men to drive. Reports of a left-sided excess of facial photodamage lesions commensurate with time spent driving in the United States4 and the commoner occurrence of solar keratoses on the right upper limb among Australian men,3 where drivers sit on the right side of vehicles, concurred with our findings. This hypothesis, however, only partly explains our results, since it cannot account for the left-sided excess of melanomas observed at other body sites.

Several aspects in embryogenesis occur in asymmetric fashion. An asymmetry in the distribution of melanocytes favoring the left body side might occur when these cells migrate from the neural crest during embryonic development. This assumption could be challenged and eventually supported by investigating the laterality of nonmelanocytic skin cancers from the Vaud Cancer Registry database7 since L/R ratios computed for squamous and basal cell carcinomas registered over a 10-year period (1995-2004) were 1.03 (1286:1244) and 1.00 (2946:2939), respectively (F.L., oral communication, September 2007). An asymmetric development of the angiolymphatic system might lead to a higher progression of left-sided melanoma, which is compatible with our concomitant increase in L/R ratios and melanoma thickness.

This largest study to date to explore melanoma laterality suggests that an asymmetric, melanocytic distribution or, to a lesser extent, a differential sun exposure are plausible etiological explanations for the observed left-sided excess of melanomas but not of other types of skin cancers.


AUTHOR INFORMATION

Correspondence: Dr Bulliard, Unité d'épidémiologie du cancer, Institut universitaire de médecine sociale et préventive, rue du Bugnon 17, 1005 Lausanne, Switzerland (Jean-Luc.Bulliard@chuv.ch).