Kasper C., van Griensven M., P?rtner R. (Eds.) Bioreactor Systems for Tissue Engineering II: Strategies for the Expansion and Directed Differentiation of Stem Cells

Подождите немного. Документ загружается.

disease in dogs. Prior to implantation the cells were suspended in ﬁbrin glue. Implan-

tation was performed by transcutaneous injection, under CT or radiographic guide,

using a Jamshidi needle inserted through the femoral head and neck starting at the base

of the trochanter major.

In nine of the treated dogs the disappearance of pain was observed after about

3–4 weeks following cell administration. This also became obvious by a gradual

weightbearingontheaffectedlimbuptoacomplete remission of the symptomatology.

In the other two cases a femoral head and neck ostectomy was performed because

the recovery proceeded too slowly. Hist ological and immu nohistochemical studies

were performed on these samples and showed new formation of cartilage and

subchondral bone in the implantation area. Therefore cell therapy seems to be an

effective and minimal invasive therapeutic approach for the treatment of Legg–

Calve

–Perthe

s disease. The efﬁcacy is considered to be due to the osteogenetic as

well as anti-inﬂammatory capacity of the stromal cells which may ﬁrst lea d to pain

relieve and then to reparative activity within the bone causing a better sclerosis of

the femoral head [120].

Regardless of the clinical application, all mentioned studies share the common

observation of improved bone tissue formation upon local MSC application as an

essential part of the tissue engineering process. However, the application of MSCs

for bone repair in the veterinary implementation does not predominantly aim at the

clinical treatment of animal patients. More often, animals are used as appropriate

models to conduct preclinical studies before advancing to human clini cal trials.

Still, the principles tested in a species like the dog can directly be clinically

translated in the patient of the respective species.

4.4 Spinal Cord Injuries

Acute spinal cord injuries affect many dogs and cats. It has been reported that about

1–2% of all dogs admitted to animal hospitals suffer from injuries to the spinal cord

only due to intervertebral disc disease. Clearly there are many other conditions that

can lead to compression, concussion or laceration of the spinal cord [121].

Traumatic spinal cord injury causes loss of tissue, including myelinated ﬁbre

tracts responsible for carrying descending motor and asce nding sensory informa-

tion. Reduced myelination could be due to either loss of myelinated cells or reduced

oligodendrocyte myelin synthesis [122].

Although animals tend to recover a substantial amount of locomotor ability after

spinal cord injury, the natural CNS capacity to recover from injury is unfortunately

limited. Neuroanatomical differences between species may also be an important

factor that needs to be considered in the assessment of the recovery of spinal cord

injuries [121].

After spinal cord injury, massive oligodendrocyte death attributed to apoptosis

occurs. It seems that a complete restoration of the lost myelin in the injury zone

by endogenous oligodendrocytes is not possible. Therefore transplantation of cells

250 I. Ribitsch et al.

with the ability to differentiate into oligodendrocytes may be a feasible method for

myelin replacement. It was reported that stem cells implanted into spinal cord

lesions not only differentiate into astrocytes and oligodendrocytes but also integrate

into axonal pathways and thus regenerate injured axons [122].

At the moment lots of different sources of cells for neurotransplantation are

being evaluated, e.g. embryonic, bone marrow, adipose and UCB stem cells. The

cells obtained from these sources can migrate and differentiate into neural pheno-

types in the damaged brain and spinal cord [123].

Jeffery et al. [124] showed that recovery of locomotor activity of dogs with

spinal cord injuries following autologous olfactory glial cell transplantation

appeared to be considerably faster than reported in historical cases.

Adel and Gabr [125] reported signiﬁcant improvement in the motor power of six

dogs compared to the control group, based on intrathecal transplantation of autolo-

gous bone marrow derived MSCs 1 week after spinal cord injury.

It was also shown that allogeneic UCB deri ved MSC transplantation is feasible

to induce neuroregeneration using UCB MSCs derived from canine foetuses. UCB

contains more mesenchymal progenitor cells and is more pluripotent and geneti-

cally ﬂexible than bone marrow derived stem cells. Based on the fact that they are

less mature than other adult stem cells they may not elicit alloreactive responses

that modulate the immune system.

Dogs included in the study had more than 75% of their spinal canal occluded

over a 12-h period. This resulted in a manifest lesion with histologically severe

haemorrhage and vacuole formation. The dogs showed paraplegia and were not

expected to regain a normal gait. In the group with UCB MSC treatment the gait

improved from 2 weeks and the weight bearing of the pelvic limbs improved from

10% to 50% of the time. Therefore, the group with UBC MSC treatment appeared

to have improved spinal cord function after the experimentally induced spinal cord

injury. It is concluded that MSCs might improve the functional outcome by creating

new neuronal pathways in the ﬁbrous scar tissue. They have been observed to

integrate into the lesion in the central nervous tissue and a smaller percentage of

cavity formation was observed following UCB MSC injection. However, 8 weeks

after stem cell implantation magnetic resonance imaging and histology showed no

convincing evidence of spinal cord regeneration. Based on somatosensory evoked

potentials it was also demonstrated that the nerve conduction velocity was signiﬁ-

cantly improved. In addition a distinct structural consistency of the nerve cell

bodies was observed in lesions treated with MSCs [123].

After human UCB stem cell implantation fol lowing spinal cord injury in rats,

locomotor function was signiﬁcantly enhanced within 14 days after transplantation

as compared to the non-treated group. In contrast to the non-treated group, consis-

tent plantar stepping, forelimb-hindlimb coordination and no toe drag during

walking were observed. Findings demonstrated that hUCB stem cells differentiate

into oligodendrocytes and neurons in vivo and lead to improved locomotor function

[122]. Moreover, Dasari et al. [122] showed that the number of oligodendrocytes

as well as of myelinated axons was elevated in the treatment group compared to the

control group and that neuroptropins (NT3 and BDNF) secreted by these

Basic Science and Clinical Application of Stem Cells in Veterinary Medicine 251

oligodendrocytes in turn enhanced mye linogenesis as well as proliferation and

survival of oligodendrocyte precursors. Furthermore, hUCB stem cells producing

these neurotropins seem to promote neuritogenesis and axon myelination. Morpho-

logically normal appearing sheaths around the axons in the injured areas were found

as well, which was consistent with the observed rapid locomotor improvement.

These results are consistent with the hypothesised migration of stem cells to lesion

sites and their participation in healing of neurological defects caused by traumatic

injury. In the non-injured areas of the spin al cord no hUCB derived cells were

detectable [122].

First results from studies in primates (Rhesus monkeys) using bone marrow

derived MSCs were promising as well.

Corticosomatosensory evoked potential signals recovered signiﬁcantly 3 months

after MSC injection whereas in control animals the signals remained ﬂattened. The

same was observed for motorevoked potential. Healing and regeneration of the

spinal cords in animals transplanted with MSC derived cells was shown by H&E

staining. In contrast, the injured tissue of the control animals showed obvious

degeneration with the appearance of many holes and abundant dissolution of neural

tissue and cells. Re-establishment of the axon pathway across the contusive injury of

the spinal cord was evaluated by application of labelled cells that were later observed

in the rostral thoracic spinal cord, red nucleus and senso ry motor cortex [126].

It is not clear, however, whether the therapeutic potential of stem cells is based

on their attributed inherent ability to replace injured tissue s or if they repair

damaged tissue through the induction of neural protection and secretion of neuro-

trophic factors by various cell types within the graft. More precisely, stem cells

could either promote axonal regeneration by constituting a connection through a

lesion site which in turn supports axonal attachment or secrete certain growth

factors to attract injured axons. It also still needs to be determined if the enhanced

functional recovery is based on re-myelination of demyelinated axons or by trophic

support to prevent degeneration of the white matter [122].

MSCs have been shown to differentiate into neurons via ex vivo induction as

well as following in vivo transplantation. However, compared with native MSCs,

neural induced MSCs display a higher survival rate and support better functional

recovery after transplantation in rat models. As the microenvironment of acute

injury does not favour de novo neurogenesis, the brief induction of MSCs prior to

implantation might have a beneﬁcial effect on their in vivo differentiation [126].

4.5 Liver Disease

Also so-called liver progenitor cells (LPC) are hoped to be able to support liver

regeneration. LPCs, undifferentiated epithelial cells lying at the interface of the

hepatic cords and the biliary tree, offer a promising target for therapeutic interven-

tion in severe liver diseases [127]. They are bipotential cells who express hepato-

cytic, biliary and progenitor cell markers and can also be isolated from the smallest

252 I. Ribitsch et al.

and most peripheral branches of the biliary tree (Hering canals) [128, 129]. These

cells are deﬁned as side population. Side populations were identiﬁed in multiple

tissues and display an enriched population of authentic or potential tissue stem

cells. In vitro they show a greatly enriched haematopoietic stem cell potential

whereas in vivo they show haematopoietic reconstitution activity. In healthy livers,

LPCs remain in a quiescent stadium [129] and their presence is low, but they

proliferate and invade the liver parenchyma in several pathologic conditions

[128]. LPCs are only activated during liver regeneration when hepatocyte prolifer-

ation is insufﬁcient [127]. Activated LPCs can either differentiate into haemato-

poietic lineages [129] or mature hepatocytes as well as cholangiocytes in order to

regenerate the pathological changes in the liver [128].

In animal models it was shown that MSCs induced to adopt a hepatocytic

phenotype as well as BM mononuclear MSC subpopulations contribute to a histo-

logic decrease in hepatic ﬁbrosis and a rise in serum albumin level when infused

early enough after injury onset [130].

The results of the study performed by Arends et al. [129] provide a new option of

treatment approach in currently untreatable canine liver diseases. It is hypothesised

that liver reconstitution can be stimulated by injection of progenitor cells into

diseased livers or via stimulation of the endogenous progenitor cells. The potential

use of these cells for the treatment of naturally occurring liver dise ase in dogs is also

of interest for human medicine, as a high homology with human liver diseases at the

molecular as well as pathological level is described [128].

Another approach to achieve liver regeneration might be using bone marrow.

Bone marrow comp rises hepatic stellate cells and myoﬁbroblasts, which were

showntobeofMSCorigin[131]. Based on these ﬁndings it is hypothesised that

some hepatocyte regeneration may be achieved through bone marrow MSC

transplantation and might induce measurable improvements in hepatic function

after damage. Whether engraftment and origin restitution continues in the long

term has not been described yet. Another possible explanation for the reduced

ﬁbrosis is that hepatocyte proliferation and suppression of ﬁbrogenesis are

induced by critical growth f actors and cytokines supplied by migrating bone

marrow cells [130].

5 Future Prospects and Outlook

Based on all these reports it is obvious that regenerative medicine in the ﬁeld of

veterinary medicine is making great steps to become clinical reality but it is also

shown that several important questions still remain to be answered. One of the

fundamental questions is the adequate number of cells that would need to be

implanted in order to achieve optimal results. Proving that the effect of cell based

treatment regimes is in fact caused by the administered stem cells and not by any

other cells or biological factors applied simultaneously is still outstanding as well [11].

It also needs to be answered whether stem cells really functionally incorporate into

Basic Science and Clinical Application of Stem Cells in Veterinary Medicine 253

the tissue that requires regeneration or whether they excite a conducting role

recruiting and controlling resident cells to regenerate the respective tissue [11].

Maybe they rather synthesise and secrete growth factors which in turn promote

tissue function [5].

Considering all the aspects discussed above, it also seems that MSCs obtained

from different sources may have different properties and it will be necessary to

deﬁne the best source depending on the intended treatment.

6 Embryonic Stem Cells and Induced Pluripotent Stem Cells

Embryonic stem (ES) cells are pluripotent stem cells obtained from the inner cell

mass of the blastocyst – an early-stage embryo. In humans, for example, embryos

reach the blastocyst stage about 4–5 days post fertilisation. ES cells are capable of

self-renewal and thus have the inherent potential for exceptionally prolonged

culture (up to 1–2 years). So far ES cells have been recovered and maintained

from non-human primate, mouse [5 ] and horse blastocysts [132]. In addition,

bovine ES cells have been grown in primary culture and there are several reports

of ES cells derived from mink, rat, rabbit, chicken and pigs [5]. Advances in the

laboratory have led to development of feed er and animal-sera – free cells lines.

However, clinical application of ES cells remains faced with practical and ethical

concerns [133]. Their potential for uncontrolled proliferation and immune rejection

[133] as well as their tendency towards teratogenic degeneration in vivo remain

major obstacles. The potential to form teratoma consisting of tissues from all three

germ lines even serves as a deﬁnitive in vivo test for ES cells.

Recently veterinary scientists started to develop several equine ES cells lines so

that they can be genetically matched to patients to eliminate immune rejection

[133]. Horse ES cells were found to express ES cel l marker genes that differ from

both human and mouse ES cells, but that reﬂects the expression of these genes in the

inner cell mass of horse blastocysts. Therefore it may be concluded that species

differences exist even at this early stage of development and that horse ES cells may

provide a better tool to study early horse development than extrapolating data from

other species. Equine ES cel ls are able to generate derivatives of all three germ

layers upon differentiation in vitro. Interestingly, they seem not to generate terato-

mas upon implantation into severe combined immune deﬁcient (SCID) mice. This,

combined with a lack of expressio n of MHC class II antigens, may make horse ES

cells more suitable for use in cell transplantation therapies [134] than ES cells from

other species. Based on that, at least two companies are currently developing equine

ES cells and pilot studies are being performed to determine the efﬁcacy of equine

ES cells for tendon regeneration [133]. In addition, ES cells certainly remain an

important model system for studying cellular differentiation in relationship to

development and oncogenesis [133].

A major breakthrough in the ﬁeld of stem cell research was achieved in 2006,

when it was shown that induced pluripotent stem (IPS) cells could be obtained from

254 I. Ribitsch et al.

adult somatic cells through expression of a set of transcription factors such as Oct4,

Sox2, Klf4, c-Myc, NANOG and Lin28 [133]. Also IPS cells are capable of

differentiating into all three embryonic germ layers and, because of this, they

have enormous potential for biomedical research and regenerative therapy. These

ES-like cells have been generated from rodent, human and porcine somatic cells by

forcing the ectopic expression of four transcription factors, Oct4, Sox2, Klf4 and

cMyc. These IPS cells may have a great potential in medicine because they can be

produced in a patient-speciﬁc manner [135]. Generation of IPS cel ls allows for

development of patient-speciﬁc cell populations without the ethical controversy of

ES cells. Prior to clinical application, an important next step will be to identify ways

of assessing which IPS cell lines are sufﬁciently reprogrammed and safe for

therapeutic applications [133]. In addition it will again be necessary to overcome

their potential to form teratoma in vivo.

A speci es, other than humans, that is likely to beneﬁt from this potential is the

horse, particularly in regard to the treatment of musculoskeletal injuries. First

studies attempting to derive IPS cells from equine somatic cells have begun.

Putative equine IPS colonies were identiﬁed that tested positive for ALP and

Nanog. Clonal populations have continued to expand while maintaining their ES-

like morphol ogy over several passages. Current efforts are focused on deﬁnitely

establishing the pluripotency of these cell lines, including their potential for

differentiation into cells of all three embryonic germ layers. Initial results are

very encouraging for the eventual generation of IPS cell lines that may have great

potential for equine regenerative medicine [135]. However, it was shown that age,

origin and cell type have a deep impact on the reprogramming efﬁciency [136] and

most likely also the quality of the obtained IPS cells. IPS cells obtained from

somatic cells of adult patients are IPS cells with the biological age of the donor.

Therefore, recently a study on the gener ation of IPS cells from human cord blood

was carried out in order to obtain IPS cells from young cells which can be expected

to carry minimal somatic mutations and the immunological immaturity of newborn

cells [136]. This might offer major advantages for the future use of IPS cells.

In summary, it can clearly be said that in the future a lot of effort still needs to be

put into all ﬁelds of stem cell rese arch and veterinary medicine will also play an

important role because animals may serve as models for human medicine.

7 Animal Models

Cell therapy with adult pluripotent MSCs may revolutionise the treatment of a large

variety of diseases in veterinary as well as in human medicine in the future [7].

Veterinary medicine in the form of animal trials plays a major role in preclinical

and ﬁrst clinical phases of human medical trials. In animal studies MSCs seem to

provide disease-ameliorating effects in conditions like Alzheimer’s disease [137],

Huntington’s disease [138], amyotrophic lateral sclerosis [139], spinal cord injury

[122, 140–142] and myocardial infarction [143, 144].

Basic Science and Clinical Application of Stem Cells in Veterinary Medicine 255

Also, the effect of MSCs in orthopaedic disorders like OA and tendon injuries is

being studied using animal models.

In general, it is important to deﬁne the questions and goals of a preclinical

animal study before the required species is chosen. However, successful laboratory

studies provide valuable proof of principal demonstrating statistical differences in

outcome between small groups of treated and control animals with highly un iform

injuries, but translational stud ies aiming to determine whether MSC transplantation

provides a medically useful effect in large patient populations that have some

variability in the degree of injury severity still need to be carried out [124].

References

1. Agung M, Ochi M, Yanada S, Adachi N, Izuta Y, Yamasaki T, Toda K (2006) Mobilization

of bone marrow-derived mesenchymal stem cells into the injured tissues after intraarticular

injection and their contribution to tissue regeneration. Knee Surg Sports Traumatol Arthrosc

14:1307–1314

2. Barry FP, Murphy JM (2004) Mesenchymal stem cells: clinical applications and biological

characterization. Int J Biochem Cell Biol 36:568–584

3. Fortier LA, Nixon AJ, Williams J, Cable CS (1998) Isolation and chondrocytic differentia-

tion of equine bone marrow-derived mesenchymal stem cells. Am J Vet Res 59:1182–1187

4. Worster AA, Nixon AJ, Brower-Toland BD, Williams J (2000) Effect of transforming

growth factor beta1 on chondrogenic differentiation of cultured equine mesenchymal stem

cells. Am J Vet Res 61:1003–1010

5. Fortier LA (2005) Stem cells: classiﬁcations, controversies, and clinical applications. Vet

Surg 34:415–423

6. Dahlgren LA (2009) Fat-derived mesenchymal stem cells for equine tendon repair. In: World

Conference on Regenerative Medicine. Regen Med Suppl, Vol.4, No.6 (Suppl. 2), Nov 2009.

Ref Type: Conference Proceeding

7. Prockop DJ, Gregory CA, Spees JL (2003) One strategy for cell and gene therapy: harnessing

the power of adult stem cells to repair tissues. Proc Natl Acad Sci USA 100(Suppl 1):11917–

11923

8. Ryan JM, Barry FP, Murphy JM, Mahon BP (2005) Mesenchymal stem cells avoid alloge-

neic rejection. J Inﬂamm (Lond) 2:8

9. Csaki C, Matis U, Mobasheri A, Ye H, Shakibaei M (2007) Chondrogenesis, osteogenesis

and adipogenesis of canine mesenchymal stem cells: a biochemical, morphological and

ultrastructural study. Histochem Cell Biol 128:507–520

10. Hoynowski SM, Fry MM, Gardner BM, Leming MT, Tucker JR, Black L, Sand T, Mitchell

KE (2007) Characterization and differentiation of equine umbilical cord-derived matrix

cells. Biochem Biophys Res Commun 362:347–353

11. Koch TG, Berg LC, Betts DH (2008) Concepts for the clinical use of stem cells in equine

medicine. Can Vet J 49:1009–1017

12. Fan J, Varshney RR, Ren L, Cai D, Wang DA (2009) Synovium-derived mesenchymal stem

cells: a new cell source for musculoskeletal regeneration. Tissue Eng Part B Rev 15:75–86

13. Koch TG, Heerkens T, Thomsen PD, Betts DH (2007) Isolation of mesenchymal stem cells

from equine umbilical cord blood. BMC Biotechnol 7:26

14. Yoshimura H, Muneta T, Nimura A, Yokoyama A, Koga H, Sekiya I (2007) Comparison of

rat mesenchymal stem cells derived from bone marrow, synovium, periosteum, adipose

tissue, and muscle. Cell Tissue Res 327:449–462

256 I. Ribitsch et al.

15. Koerner J, Nesic D, Romero JD, Brehm W, Mainil-Varlet P, Grogan SP (2006) Equine

peripheral blood-derived progenitors in comparison to bone marrow-derived mesenchymal

stem cells. Stem Cells 24:1613–1619

16. Vidal MA, Kilroy GE, Johnson JR, Lopez MJ, Moore RM, Gimble JM (2006) Cell growth

characteristics and differentiation frequency of adherent equine bone marrow-derived mes-

enchymal stromal cells: adipogenic and osteogenic capacity. Vet Surg 35:601–610

17. Vidal MA, Kilroy GE, Lopez MJ, Johnson JR, Moore RM, Gimble JM (2007) Characteriza-

tion of equine adipose tissue-derived stromal cells: adipogenic and osteogenic capacity and

comparison with bone marrow-derived mesenchymal stromal cells. Vet Surg 36:613–622

18. Giovannini S, Brehm W, Mainil-Varlet P, Nesic D (2008) Multilineage differentiation

potential of equine blood-derived ﬁbroblast-like cells. Differentiation 76:118–129

19. Kadiyala S, Young RG, Thiede MA, Bruder SP (1997) Culture expanded canine mesenchy-

mal stem cells possess osteochondrogenic potential in vivo and in vitro. Cell Transplant

6:125–134

20. Martin DR, Cox NR, Hathcock TL, Niemeyer GP, Baker HJ (2002) Isolation and characteri-

zation of multipotential mesenchymal stem cells from feline bone marrow. Exp Hematol

30:879–886

21. Ringe J, Kaps C, Schmitt B, Buscher K, Bartel J, Smolian H, Schultz O, Burmester GR,

Haupl T, Sittinger M (2002) Porcine mesenchymal stem cells. Induction of distinct mesen-

chymal cell lineages. Cell Tissue Res 307:321–327

22. Bosnakovski D, Mizuno M, Kim G, Takagi S, Okumura M, Fujinaga T (2005) Isolation and

multilineage differentiation of bovine bone marrow mesenchymal stem cells. Cell Tissue

Res 319:243–253

23. Kulterer B, Friedl G, Jandrositz A, Sanchez-Cabo F, Prokesch A, Paar C, Scheideler M,

Windhager R, Preisegger KH, Trajanoski Z (2007) Gene expression proﬁling of human

mesenchymal stem cells derived from bone marrow during expansion and osteoblast differ-

entiation. BMC Genomics 8:70

24. Smith RK, Korda M, Blunn GW, Goodship AE (2003) Isolation and implantation of

autologous equine mesenchymal stem cells from bone marrow into the superﬁcial digital

ﬂexor tendon as a potential novel treatment. Equine Vet J 35:99–102

25. Huss R, Lange C, Weissinger EM, Kolb HJ, Thalmeier K (2000) Evidence of peripheral

blood-derived, plastic-adherent CD34(-low) hematopoietic stem cell clones with mesenchy-

mal stem cell characteristics. Stem Cells 18:252–260

26. Reed SA, Johnson SE (2008) Equine umbilical cord blood contains a population of stem cells

that express Oct4 and differentiate into mesodermal and endodermal cell types. J Cell

Physiol 215:329–336

27. Sakaguchi Y, Sekiya I, Yagishita K, Muneta T (2005) Comparison of human stem cells

derived from various mesenchymal tissues: superiority of synovium as a cell source. Arthritis

Rheum 52:2521–2529

28. Reich CM, Raabe O, Wenisch S, Bridger PS, Kramer M, Arnhold S (2009) Comparison of

canine adipose and bone marrow-derived mesenchymal stem cells. In: World Conference on

Regenerative Medicine. Regen Med Suppl,Vol.4, No.6 (Suppl. 2), Nov 2009. Ref Type:

Conference Proceeding

29. Colleoni S, Bottani E, Tessaro I, Mari G, Merlo B, Romagnoli N, Spadari A, Galli C, Lazzari

G (2009) Isolation, growth and differentiation of equine mesenchymal stem cells: effect of

donor, source, amount of tissue and supplementation with basic ﬁbroblast growth factor. Vet

Res Commun 33:811–821

30. Conrad S, Nufer F, Mundle K, Ihring J, Seid K, Walliser U, Skutella T (2009) Mesenchymale

Stammzellen aus dem Fettgewebe des Pferdes – Isolation, Expansion und Charakterisierung.

18. In: Tagung u

ber Pferdekrankheiten im Rahmen der Equitana. 119. 2009, 20-3-2009. Ref

Type: Conference Proceeding

31. Mundle K, Conrad S, Skutella T, Walliser U (2009) Mesenchymale Stammzellen aus

Fettgewebe – Neue Anwendungsmo

glichkeiten in der Orthopa

die. 18. In: Tagung u

ber

Pferdekrankheiten im Rahmen der Equitana, pp. 120–121. Ref Type: Conference Proceeding

Basic Science and Clinical Application of Stem Cells in Veterinary Medicine 257

32. De Ugarte DA et al (2003) Comparison of multi-lineage cells from human adipose tissue and

bone marrow. Cells Tissues Organs 174:101–109

33. Kern S, Eichler H, Stoeve J, Kluter H, Bieback K (2006) Comparative analysis of mesen-

chymal stem cells from bone marrow, umbilical cord blood, or adipose tissue. Stem Cells

24:1294–1301

34. Koga H, Muneta T, Nagase T, Nimura A, Ju YJ, Mochizuki T, Sekiya I (2008) Comparison

of mesenchymal tissues-derived stem cells for in vivo chondrogenesis: suitable conditions

for cell therapy of cartilage defects in rabbit. Cell Tissue Res 333:207–215

35. Vidal MA, Robinson SO, Lopez MJ, Paulsen DB, Borkhsenious O, Johnson JR, Moore RM,

Gimble JM (2008) Comparison of chondrogenic potential in equine mesenchymal stromal

cells derived from adipose tissue and bone marrow. Vet Surg 37:713–724

36. Passeri S et al (2009) Isolation and expansion of equine umbilical cord-derived matrix cells

(EUCMCs). Cell Biol Int 33:100–105

37. Mitchell KE et al (2003) Matrix cells from Wharton’s jelly form neurons and glia. Stem Cells

21:50–60

38. Weiss ML, Mitchell KE, Hix JE, Medicetty S, El-Zarkouny SZ, Grieger D, Troyer DL

(2003) Transplantation of porcine umbilical cord matrix cells into the rat brain. Exp Neurol

182:288–299

39. De Bari C, Dell’Accio F, Tylzanowski P, Luyten FP (2001) Multipotent mesenchymal stem

cells from adult human synovial membrane. Arthritis Rheum 44:1928–1942

40. Staszyk C, Mensing N, Hambruch N, Ha

ger J-D, Pfarrer C, Gasse H (2009) Equine

periodontal ligament: a source of mesenchymal stem cells for regenerative therapies in the

horse? In: World Conference on Regenerative Medicine. Regen Med Suppl, Vol. 4, No. 6

(Suppl.2), Nov 2009. Ref Type: Conference Proceeding

41. Warhonowicz M, Staszyk C, Rohn K, Gasse H (2006) The equine periodontium as a

continuously remodeling system: morphometrical analysis of cell proliferation. Arch Oral

Biol 51:1141–1149

42. McCulloch CA (1985) Progenitor cell populations in the periodontal ligament of mice. Anat

Rec 211:258–262

43. Staszyk C, Gasse H (2007) Primary culture of ﬁbroblasts and cementoblasts of the equine

periodontium. Res Vet Sci 82:150–157

44. Gould TR (1983) Ultrastructural characteristics of progenitor cell populations in the peri-

odontal ligament. J Dent Res 62:873–876

45. Gronthos S, Mrozik K, Shi S, Bartold PM (2006) Ovine periodontal ligament stem cells:

isolation, characterization, and differentiation potential. Calcif Tissue Int 79:310–317

46. Shirai K, Ishisaki A, Kaku T, Tamura M, Furuichi Y (2009) Multipotency of clonal cells

derived from swine periodontal ligament and differential regulation by ﬁbroblast growth

factor and bone morphogenetic protein. J Periodontal Res 44:238–247

47. Fujii S, Maeda H, Wada N, Tomokiyo A, Saito M, Akamine A (2008) Investigating a clonal

human periodontal ligament progenitor/stem cell line in vitro and in vivo. J Cell Physiol

215:743–749

48. Toma JG, Akhavan M, Fernandes KJ, Barnabe-Heider F, Sadikot A, Kaplan DR, Miller FD

(2001) Isolation of multipotent adult stem cells from the dermis of mammalian skin. Nat Cell

Biol 3:778–784

49. Jiang Y, Vaessen B, Lenvik T, Blackstad M, Reyes M, Verfaillie CM (2002) Multipotent

progenitor cells can be isolated from postnatal murine bone marrow, muscle, and brain. Exp

Hematol 30:896–904

50. Stewart A, Chen YJ, Caporali EH, Stewart A (2009) Isolation and chondrogenic differentia-

tion of cells isolated from equine synovial ﬂuid. In: World Conference on Regenerative

Medicine. Regen Med Suppl Vol. 4, No. 6 (Suppl. 2), Nov 2009. Ref Type: Conference

Proceeding

51. Durgam SS, Stewart AA, Caporali EH, Karlin WM, Stewart MC (2009) Effect of tendon-

derived progenitor cells on a collagenase-induced model of tendinitis in horses. In: World

258 I. Ribitsch et al.

Conference on Regenerative Medicine. Regen Med Suppl, Vol.4, No.6 (Suppl.2), Nov 2009.

Ref Type: Conference Proceeding

52. da Silva ML, Chagastelles PC, Nardi NB (2006) Mesenchymal stem cells reside in virtually

all post-natal organs and tissues. J Cell Sci 119:2204–2213

53. Krampera M, Glennie S, Dyson J, Scott D, Laylor R, Simpson E, Dazzi F (2003) Bone

marrow mesenchymal stem cells inhibit the response of naive and memory antigen-speciﬁc T

cells to their cognate peptide. Blood 101:3722–3729

54. Tse WT, Pendleton JD, Beyer WM, Egalka MC, Guinan EC (2003) Suppression of alloge-

neic T-cell proliferation by human marrow stromal cells: implications in transplantation.

Transplantation 75:389–397

55. Di Nicola M, Carlo-Stella C, Magni M, Milanesi M, Longoni PD, Matteucci P, Grisanti S,

Gianni AM (2002) Human bone marrow stromal cells suppress T-lymphocyte proliferation

induced by cellular or nonspeciﬁc mitogenic stimuli. Blood 99:3838–3843

56. Guest DJ, Smith MR, Allen WR (2008) Monitoring the fate of autologous and allogeneic

mesenchymal progenitor cells injected into the superﬁcial digital ﬂexor tendon of horses:

preliminary study. Equine Vet J 40:178–181

57. Chong AK, Ang AD, Goh JC, Hui JH, Lim AY, Lee EH, Lim BH (2007) Bone marrow-

derived mesenchymal stem cells inﬂuence early tendon-healing in a rabbit Achilles tendon

model. J Bone Joint Surg Am 89:74–81

58. Brehm W (2006) Stammzellen, Stammzelltherapie – Begriffserkla

rung, Zusammenha

nge

und mo

gliche klinische Anwendungen. Pferdeheilkunde 22:259–267

59. Herthel DJ (2001) Enhanced suspensory ligament healing in 100 horses by stem cells and

other bone marrow components. Proc Am Ass equine Practnrs 47:319–321

60. Richardson LE, Dudhia J, Clegg PD, Smith R (2007) Stem cells in veterinary medicine–

attempts at regenerating equine tendon after injury. Trends Biotechnol 25:409–416

61. Smith RK (2008) Mesenchymal stem cell therapy for equine tendinopathy. Disabil Rehabil

30:1752–1758

62. Dowling BA, Dart AJ, Hodgson DR, Smith RK (2000) Superﬁcial digital ﬂexor tendonitis in

the horse. Equine Vet J 32:369–378

63. Taylor SE, Smith RK, Clegg PD (2007) Mesenchymal stem cell therapy in equine musculo-

skeletal disease: scientiﬁc fact or clinical ﬁction? Equine Vet J 39:172–180

64. Nixon AJ, Dahlgren LA, Haupt JL, Yeager AE, Ward DL (2008) Effect of adipose-derived

nucleated cell fractions on tendon repair in horses with collagenase-induced tendinitis. Am J

Vet Res 69:928–937

65. Leppa

nen M, Miettinen S, Ma

kinen S, Wilpola P, Katiskalahti T, Heikkila

P, Tulamo R-M

(2009) Management of equine tendon and ligament injuries with expanded autologous

adipose-derived mesenchymal stem cells: a clinical study. In: World Conference on Regen-

erative Medicine. Regen Med Suppl, Vol.4, No.6 (Suppl. 2), Nov 2009. Ref Type: Confer-

ence Proceeding

66. Pacini S, Spinabella S, Trombi L, Fazzi R, Galimberti S, Dini F, Carlucci F, Petrini M (2007)

Suspension of bone marrow-derived undifferentiated mesenchymal stromal cells for repair

of superﬁcial digital ﬂexor tendon in race horses. Tissue Eng 13:2949–2955

67. Schnabel LV, Lynch ME, van der Meulen MC, Yeager AE, Kornatowski MA, Nixon AJ

(2009) Mesenchymal stem cells and insulin-like growth factor-I gene-enhanced mesenchy-

mal stem cells improve structural aspects of healing in equine ﬂexor digitorum superﬁcialis

tendons. J Orthop Res 27:1392–1398

68. Smith RK, Webbon PM (2005) Harnessing the stem cell for the treatment of tendon injuries:

heralding a new dawn? Br J Sports Med 39:582–584

69. Crovace A, Lacitignola L, De SR, Rossi G, Francioso E (2007) Cell therapy for tendon repair

in horses: an experimental study. Vet Res Commun 31(Suppl. 1):281–283

70. Smith R, Young N, Dudhia J, Kasashima Y, Clegg PD, Goodship A (2009) Effectiveness of

bone-marrow-derived mesenchymal progenitor cells for naturally occurring tendinopathy in

the horse. In: World Conference on Regenerative Medicine. Regen Med Suppl, Vol. 4, No. 6

(Suppl. 2), Nov 2009. Ref Type: Conference Proceeding

Basic Science and Clinical Application of Stem Cells in Veterinary Medicine 259