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In Vitro Cell.Dev.Biol.

Animal (2017) 53:363370


DOI 10.1007/s11626-016-0121-2

Labeling of adipose-derived stem cells with quantum


dots provides stable and long-term fluorescent signal
for ex vivo cell tracking
Clautina R. M. Costa 1 & Matheus L. T. Feitosa 1 & Dayseanny O. Bezerra 1 &
Yulla K. P. Carvalho 1 & Rodrigo F. G. Olivindo 1 & Pablo B. Fernando 1 &
Gustavo C. Silva 1 & Mirna L. G. Silva 1 & Carlos E. Ambrsio 2 & Airton M. Conde Jnior 3 &
Napoleo M. Argolo Neto 1 & Las M. Costa Silva 1 & Maria A. M. Carvalho 1

Received: 13 October 2016 / Accepted: 7 December 2016 / Published online: 30 December 2016 / Editor: Tetsuji Okamoto
# The Society for In Vitro Biology 2016

Abstract Stem cells derived from adipose tissue (ADSC) nanocrystals (Qdots) enabling their in vitro and ex vivo
have been used in cell therapy as an alternative to treat chronic tracking.
and degenerative diseases. Using biomedical and image trials
to track the cells when infused in the target tissue is essential to Keywords Qdots . Adipose-derived stem cells . Cell
control cell migration and adhesion. The objective of the pres- tracking . Stem cells
ent study was to label and assess the adhesion of goat adipose
tissue-derived stem cells (g-ADSC) after cell infusion in ani-
mal models by tracking luminescent intracytoplasmatic
nanocrystals. The cells were labeled by using Qdots. The g- Introduction
ADSCs infused with nanocrystal were prepared either fresh or
fixed and further visualized under a fluorescence microscope. Adipose tissue is an abundant source of multipotent stem cells
The labeled cells were infused in the goat mammary glands (Zuk et al. 2002; Kershaw and Flier 2004). The use of stem
and mouse testicles and kidneys via tail vein injection. Thirty cells derived from adipose tissue (ADSC) has been widely
days after cell infusion, biopsy was carried out for analyses. investigated because of its potentiality in tissue healing and
The g-ADSC cultures were presented with high cellularity and differentiation into a great variety of cells, such as adipocytes,
fibroblast morphology, even after infusion of the nanocrystals. endothelial cells, smooth muscle cells, fibroblasts, immune
It was possible, by processing in paraffin and under fluores- system cells, and matrix connective tissue (Kim et al. 2008;
cence microscopy, demonstrating the success of the labeling Lin et al. 2008).
in the long term. Freezing mammary gland biopsies in liquid The greatest interest in using ADSCs in cell therapy is
NO 2 did not alter the quality of labeling with Qdots. focused on their capacity to renew and replace the cells of
Therefore, g-ADSCs can be labeled with intracytoplasmatic the resident tissue (Katz et al. 2005; Moseley et al. 2006;
Nakagami et al. 2006; Giordano et al. 2007), and they are an
alternative for treating several chronic and degenerative dis-
eases (Bunnell et al. 2008; Zimmerlin 2010; Ren et al. 2012).
* Maria A. M. Carvalho In cell therapy preclinical trials, protocols need to be adopted
mcelina@ufpi.edu.br
with non-invasive methods of in vitro labeling that can follow
1
the effective movement of the transplanted cells in animal
Integrated Nucleus of Morphology and Stem Cell Research
(NUPCelt), Federal University of Piau (UFPI), Teresina, Piau,
models in vivo (Noh et al. 2008).
Brazil According to Shaner et al. (2005) and Akins and Dubey
2
Veterinary Medicine Department, Faculty of Animal Science and
(2008), labeling methods should express efficiently, without
Food Engineering (FZEA), University of So Paulo (USP), So toxicity to the chosen niche, supply a reliable bright signal
Paulo, Brazil with the exact location of the cells of interest, and should
3
Morphology Department, Federal University of Piau, furthermore be light-stable throughout the experiment, permit-
Teresina, Piau, Brazil ting images over days, months, and even years. They should
364 COSTA ET AL.

also be insensitive to environmental effects that may cause Use of Laboratory Animals of the National Institutes of
wrong interpretations. Chudakov et al. (2005) further added Health. The protocol was approved by the Committee on the
that they should be compatible with life systems. Ethics of Animal Experiments of the Federal Univesity of
There are different types of labelers that have satis- Piau (Permit Number 037/2012).
factory characteristics for cell tracking, such as the
green fluorescent protein (GFP) gene so that their ex-
pression is not lost during cell replication and differen- Obtaining goat adipose tissue-derived stem cells The
tiation. Unlike other labelers such as DAPI and CM-Dil ADSCs were obtained from an Anglo-Nubian kid
(Ocarino et al. 2008; Weir et al. 2008), the use of fluo- (10 mo old) that was submitted to pre-anesthesia with
rescent GFP is based on the act of property as nonin- ketamine (Dopalen) at a dose of 8 mg/kg combined
vasive probes (Iafolla et al. 2008; Moreau et al. 2010; with midazolam (Dormonid) at a dose of 0.5 mg/kg
Dedecker et al. 2013); a beta-galactosyltransferase (lacZ) by intramuscular injection and infiltrative anesthesia
gene cleaves the colorless X-gal substrate, an analog of with subcutaneous lidocaine at the dose of 4 mg/kg in
the lactose in galactose; this divides spontaneously and the sternum region. An incision was made in the skin
is oxidized in a blue-colored soluble product, making over the manubrium region, and fat was obtained and
the enzyme-producing cell easy to identify (Pfeifer placed in 50 mL falcon tubes containing culture medi-
et al. 2001); labeling with bioluminescence resonance um and transported in an isothermal box to the Stem
energy transfer (BRET), in which excitation is observed Cell Culture Laboratory (LABCelt) of the Stem Cell
by means of a chemical reaction and thus the energy is Morphology and Research Integrated Nucleus
transferred from a donor to a chemiluminescent receptor (NUPCelt)-Federal University of Piau. The isolated
(Pfleger and Eidne 2006), and labeling with Quantum cells were cultured in DMEM/F12 medium (GIBCO
dots (Qdots), which release fluorescent nanocrystals in Dulbeccos modified Eagle medium/Nutrient Mixture
live cell cytoplasm and can be tracked through various F12, Invitrogen, Carlsbad, CA).
cell generations, were performed (Chan et al. 2002).
Qdots are bright light stable fluorophores that have a wide Protocols used for labeling and tracking g-ADSC with
excitation spectrum and permit efficient multicolor imaging of Qdots (Invitrogen, Life Technologies, EUA) Six passage
biological samples when submitted to narrow wavelengths cells were trypsinized and the pellet was re-suspended in
(Chan et al. 2002); in addition, the nanocrystals can be ob- 1 mL culture medium (alfa-MEM GIBCO Minimum
served by using any fluorescence microscope and remain la- Essential Medium Eagle, Invitrogen). The Qdots compo-
beled for many days. nentsemission (655 nm) and excitation (405615 nm)
Experiments using nanocrystals (Qdots) have been were homogenized according to the manufacturers in-
carried out to label and track stem cells in animal and structions. The g-ADSCs were added to the dot compo-
human models (Rosen et al. 2007; Oliveira et al. 2009; nents, incubated in a CO2 chamber at 37 C for 50 min,
Tognoli et al. 2009). According to Oliveira et al. and shaken in the vortex every 10 min. Next, 1 mL alfa-
(2014), the Qdots have the ability to schedule and track MEM medium was added and centrifuged twice, 462g
almost any type of material of biological interest as well for 5 min, and the pellet was re-suspended in 1 mL phys-
as being a definite contribution to know the regenerative iological solution. Protocols were adopted to visualize un-
potential and respond to the various issues related to der a fluorescence microscope the g-ADSC-nac labeling
stem cell transplants, or in the implant site or after of fresh and fixed cells in vitro and ex vivo, by biopsy,
migration, in vivo and ex vivo. According to Kaur after cell infusion in animal models (mouse, Mus
and Singhal (2012), the Qdots are considered suitable musculus, and goat, Capra hircus).
for the visualization of stem cells due also to its
photostability and longevity.
The objective of the present study was to assess the quality Tracking intracytoplasmatic Qdots in vitro g-ADSC Two
of intracytoplasmatic labeling with Qdots and g-ADSCs ad- samples were used to visualize the g-ADSC-nac, one
hesion to the surround tissue after cell infusion in animal fresh, on a slide covered with a side cover for assess-
models (mouse and goat). ment under a BX41 fluorescence microscope (Olympus,
Shinjuku, Tokyo, Japan). The other g-ADSC-nac sample
(1 105) was cultured on a slide in a six-well plate,
Material and methods removed after 3 d, fixed with 4% paraformaldehyde,
and mounted on a slide with Entellan (Merck,
Ethics statement This study was carried out in strict accor- Darmstadt, Hessen, Germany) for observation under a
dance with the recommendations in the Guide for the Care and BX41 fluorescent microscope (OLYMPUS).
LABELING OF ADIPOSE-DERIVED STEM CELLS WITH QUANTUM DOTS 365

Local infusion and tracking of g-ADSC-nac in goat mam- Previously labeled cells with the Qdots issued high cytoplas-
mary gland paraffin and frozen cuts The g-ADSCs after mic fluorescence with excitation filter N2.1 (red) and L5
sixth passage were trypsinized after 80% conference and (blue). The labeled and grown cells on coverslip were larger
centrifuged to 462g for 10 min. A total of 3.5 106 than fresh cells because by joining the surface, they are
g-ADSC was labeled with Qdots, re-suspended in 1 mL sprawling and assume characteristic fibloblastoide format.
of NaCl solution, and infused in the left mammary Cells when they are not in culture present themselves epi-
gland parenchyma of previously anesthetized goats thelioid. The Qdots issues can be displayed in blue and red
(0.05 mg/kg xylazine intramuscularly, after 15 min, filter in the fluorescent microscope because it has excitation
anesthetized with ketamine (Dopalen) at a dose of wavelengths ranging from 405 and 615 nm, so the color
8 mg/kg combined with midazolam (Dormonid) at a emitted by the quantum dots can be changed. In addition,
dose of 0.5 mg/kg intravenously). The right mammary the broad absorption spectrum allows the excitation of sev-
gland was infused with phosphate-buffered saline (PBS; eral populations of nanocrystals simultaneously.
GIBCO, Invitrogen) as a control. After 30 , biopsy In the histological assessment, to track the cells by fluores-
was made on the right and left mammary glands (lateral cence microscopy, the nanocrystal labeler was visualized in
and medial regions); the goats were anesthetized with paraffin-embedded tissue sections. The analyses made after 30,
the same protocol to block in paraffin and freeze in 60, and 90 d labeling showed that the nanocrystal particles
liquid nitrogen. Paraffin cuts without staining and cuts remained in the infused g-ADSC, locally, in the organs of the
stained with hematoxylin-eosin were analyzed histologi- animal models, goat mammary gland (Fig. 2AC) and mouse
cally at 30, 60, and 90 d after collection. Frozen cuts testicles (Fig. 2DF). In addition, nanocrystal particles were
stained with Evans blue and without staining were ana- found in the mouse kidneys, after infusion through the tail vein,
lyzed after 90 d freezing. The slides were mounted with confirming cell migration through the blood circulation system
VECTASHIELD Hardset with DAPI (4, 6-diamidino- (Fig. 2GI).
2-phenylindol) (Vector Laboratories, Burlingame, CA). The samples of goat mammary gland tissue, frozen and ana-
lyzed at 90 d, acquired a sufficiently rigid texture to obtain thin
Goat ADSC-nac infusion and tracking in mouse testicle cuts in the cryostat. The material was of high morphological
and kidney To infuse the g-ADSC-nac in mice, two 106 sam- quality, so the fluorescent labelers present in the cytoplasm of
ples were re-suspended in 1 mL physiological solution. As the infused g-ADSC could be identified (Fig. 2Jl).
anesthetic protocol for cell infusion, ketamine was adminis- g-ADSC-nac were identified in the right and left antimere
tered at a dose of 80 mg/kg combined with xylazine at a dose of the goat mammary tissues, although cell infusion had been
of 10 mg/kg intramuscularly. One sample was infused locally made only in the left antimere, demonstrating cell migration
in the testicles and the other systemically through the tail vein. into the contralateral mammary gland (Fig. 2MN). Mammary
After 30 d, the mouse was anesthetized and euthanized with tissue analyses before g-ADSC-nac infusion did not show any
an overdose of thiopental sodium 100 mg/kg IP. Harvesting of fluorescent structure (Fig. 2O.), since no infusion of
tissue for biopsy was made in the testicles (right, in the upper/ nanocrystals.
mid/lower region, and left, in the upper/mid/lower region) and Fluorescence microscopy analysis of the unstained tissues
in the kidneys (right upper/lower and left upper/lower region), (Fig. 3DF) showed better g-ADSC-nac tracking compared to
to track g-ADSCs-nac. The tissue from the biopsies was fixed the analysis of the stained slides (Fig. 3GI). The g-ADSC-nac
and blocked in paraffin to make slides (30, 60, and 90 d) in the kidney, testicle, or mammary gland tissues were not
without staining and stained with hematoxylin-eosin, to track tracked by an optical microscope (Fig. 3AC).
the cells under a BX41 fluorescence microscope The frozen cuts presented the advantages compared to the
(OLYMPUS). paraffin-blocked samples which presented similar morpholo-
gy (Fig. 3K), and the tissue counterstained with DAPI
(Fig. 3J), the filter for this labeling, was evidenced cell nuclei
Results and identified the organization breast tissue goats; for further
analysis, confocal microscopy could be utilized. The slides
The Qdots fluorescent nanocrystals were incorporated into stained with Evans blue showed satisfactory results for visu-
the g-ADSC cytoplasm, confirmed by cells fixed with 4% alizing the fluorescence.
paraformaldehyde (Fig. 1A) and fresh cells. The protocol
was shown to be adequate, and after 60 min, the labels could
be identified in the cytoplasm of the fresh cells by a fluo- Discussion
rescent microscope (Fig. 1B). The nanocrystals were
transported into the cells passively, and after that period, The nanocrystals were distributed throughout the cytoplasm
they were stabilized and presented themselves as labeled. and the g-ADSC-nac that were not fixed or modified their
366 COSTA ET AL.

Fig. 1 Photomicrography of goat


adipose stem cells labeled with
Qdots-655 fluorescent
nanocrystals. A Fluorescent
photomicrographs of cultured
in vitro g-ADSC-nac on slides in
six-well plates, after 3-d culture,
fixed with 4% paraformaldehyde,
10 20. B Fluorescent
photomicrographs of the fresh
g-ADSC-nac on the slide,
covered with a slide.

morphology, becoming rounded or epithelioid, because when in vivo tracking 1 wk after infusion in the myocardium.
they are not under in vitro culture, they do not remain adhered. Oliveira et al. (2009) stated that the Qtracker-655 nano-
The labeling protocol in vitro cell before proceeding infusion crystal is very efficient in cell labeling, but the viability
in animal model was efficient, both in fresh and after cultiva- time as labeler is relatively short. Similarly, Muccioli et al.
tion and fixation. The in vitro labeling was residence time (2011) used Qdots labeling in frozen rat spleen tissue but
need not be evaluated, only confirmation that the cells were reported good results for only 3 d after cell infusion.
labeled with Qdots, and may thus be infused and subsequently Likewise, Oliveira et al. (2014) demonstrated an efficient
screened into tissues after 30, 60, and 90 d. Efficiency was model, in a 7-d labeling period of equine bone marrow
found in in vitro labeling with Qdots even after fixed with mesenchymal stem cells infused in the equine superficial
paraformaldehyde and cell division. Chan et al. (2002) conju- digital flexor tendon with Qdots, to treat experimental
gated luminescent Qdots with transferrin and after endocytosis injury. The protocol developed in this study showed more
mediated by receptors; the individual Qdots were visible, satisfactory results, since the labeling time for cell track-
caught in the vessels inside live cancerous cells. The g- ing in infused tissue was higher than that reported in
ADSC-nac fixed with paraformaldehyde maintained fibro- literature.
blast morphology and adherent plastic and after The Qdots nanolabeler was adequate to visualize infused
multiplication, their fluorescence. These results were in line stem cells in live tissues due to its light stability and long
with those by Oliveira et al. (2014) who reported that lasting labeling, both in paraffin and frozen tissues.
nanocrystals can remain active for at least four cell genera- However, the technique of blocking tissue in paraffin with
tions, because they label specific cytoplasmic proteins inside labeled cells/Qdots transplanted in vivo is a simple procedure
the g-ADSC. for their identification, and in addition, the costs are lower
The labeling methods to express themselves efficiently, compared to the freezing technique.
in addition to other criteria, should be light-stable, Considering that the fluorescence of the Qdots nanolabeler
allowing images over days, months, and even years is not transferred from the labeled cells to the tissue in which
(Shaner et al. 2005; Akins and Dubey 2008). The present they are infused, the g-ADSC-nac were seen in a different
study demonstrated g-ADSC-nac tracking in paraffin cuts way, with intense brightness, confirming the hypothesis that
for up to 90 d of Qdots labeling maintained in goat mam- they are really the transplanted cells. Even though the
mary glands and mouse testicle and kidney, indicating the transplanted cells die, their labeling will not be incorporated
suitability of this labeler for tracking g-ADSC-nac. by the resident cells.
Tognoli et al. (2009) assessed the nanocrystals as efficient
labelers to identify mesenchymal stem cells delivered Fig. 2 Fluorescent photomicrographs after g-ADSCs-nac infusion in
subconjunctivally for treatment of corneal ulcer in dogs, mice and goat mammary gland. A, B, C Mice testis. D, E, F Mice kidneys.
but in the short term, unlike the results in the present G, H, I Analysis at 30, 60, and 90 d, blocking and processing in paraffin.
J, K, l Fluorescent photomicrographs of goat mammary tissue in frozen
study. In g-ADSC-nac identification in frozen goat mam- cuts after g-ADSC-nac infusion. Analysis at 50-d conservation in liquid
mary gland cuts, cellular labeling with Qdots was also nitrogen, 10 20. M, N Fluorescent photomicrographs of g-ADSC-nac in
visualized for up to 90 d. Similarly, Rosen et al. (2007) paraffin goat mammary tissue cuts 30 d after cell infusion, the left and
froze mouse heart tissue 8 wk after infusing human mes- right antimere, respectively, 10 20. O Photomicrographs of goat mam-
mary gland before g-ADSC-nac infusion, 10 20 not showing fluores-
enchymal stem cells and visualized many fluorescent cence. P Photomicrographs of mice kidneys before g-ADSC-nac infu-
signals. Oliveira et al. (2009) labeled mesenchymal cells sion, 10 20. Q Photomicrographs of mice testis before g-ADSC-nac
from human umbilical cord with Qdots that allowed infusion, 10 20.
LABELING OF ADIPOSE-DERIVED STEM CELLS WITH QUANTUM DOTS 367

The g-ADSC-nac infused in the left antimere of the the gland, indicating collateral cell migration. Noh et al.
goat mammary gland migrated to the right antimere of (2008) used Qdots-670 to track dendritic cell migration in
368 COSTA ET AL.

Fig. 3 Tracking of goat adipose stem cells. A, B, C Optical (mouse), and mammary gland (goat) 30 d after g-ADSC infusion, respec-
photomicrographs of the kidney, testicle (mouse), and mammary gland tively, HE, 10 20. J Photomicrographs of fluorescence of frozen goat
(goat) 30 d after g-ADSC infusion, respectively, HE, 10 40. D, E, Ff mammary gland cuts 30 d after g-ADSC-nac infusion stained with Evans
Photomicrographs of fluorescence of the kidney, testicle (mouse), and blue counterstained with DAPI. K Photomicrographs of fluorescence of
mammary gland (goat) 30 d after g-ADSC infusion, respectively, without frozen goat mammary gland cuts 30 d after g-ADSC-nac infusion, with-
staining, 10 20. G, H, I Photomicrographs of the kidney, testicle out staining, 10 20.

lymph nodes, highlighting the advantages of the Qdots, facilitate the traffic, adhesion, and infiltration of cells in-
especially for cell labeling and ex vivo tracking. These fused in injury sites.
authors stated that nanocrystals are semiconductors with Infusion locally gave good adherence, and furthermore,
stable, strong, and non-toxic fluorescence, unlike the fluo- where there was inflammatory process, a great number of
rescent organic stains. This potential for cell migration molecules were produced, including proteins, which have ad-
can occur in the in vivo applications, because they can hesive action among the cells preventing losses and ensuring
suppress local inflammatory processes or replace dam- that they perform their therapeutic functions.
aged cells. This way, they enable the creation of an It was possible to track the g-ADSC-nac infused systemat-
environment-favoring tissue repair. For example, ically, through the mouse tail vein and in the kidneys, but the
Barbash et al. (2003) and Karp and Leng Teo (2009) re- labeling was less evident than that of local infusion made in
ported that they are attracted to places with inflammation the goat mammary gland and mouse testicles. According to
because they express specific receptors or ligands that Karp and Leng Teo (2009), via systemic infusion, the cells can
LABELING OF ADIPOSE-DERIVED STEM CELLS WITH QUANTUM DOTS 369

reach other organs that are not those of interest and there may Iafolla MA, Mazumder M, Sardana V, Velauthapillai T, Pannu K,
McMillen DR (2008) Dark proteins: effect of inclusion body forma-
be a significant reduction in the number of cells at the injury
tion on quantification of protein expression. Proteins 72:12331242
site of interest. Karp JM, Leng Teo GS (2009) Mesenchymal stem cell homing: the devil
HE-stained tissues hindered visualization of the Qdots la- is in the details. Cell Stem Cell 4:206216
beling because they presented components that emitted little Katz AJ, Tholpady A, Tholpady SS, Shang H, Ogle RC (2005) Cell
surface and transcriptional characterization of human adipose-
fluorescence. However, tissues stained with Evans blue
derived adherent stromal (hADAS) cells. Stem Cells 23:412423
allowed the visualization of the labeling nanocrystals, but Kaur S, Singhal B (2012) When nano meets stem: the impact of nano-
the best g-ADSC-nac tracking was shown in the unstained technology in stem cell biology. J Biosci Bioeng 113:14
tissues bearing in mind there was no interference from the Kershaw EE, Flier JS (2004) Adipose tissue as an endocrine organ. J Clin
Endocrinol Metab 89:548556
stain in the fluorescence.
Kim WS, Park BS, Kim HK, Park JS, Kim KJ, Choi JS, Chung SJ, Kim
DD, Sung JH (2008) Evidence supporting antioxidant action of
adipose-derived stem cells: protection of human dermal fibroblasts
from oxidative stress. J Dermatol Sci 49:133142
Conclusions Lin G, Garcia M, Ning H, Banie L, Guo Y, Lue TF, Lin CS (2008)
Defining stem and progenitor cells within adipose tissue. Stem
The marking of adipose tissue-derived stem cells, with Cells Dev 17:10531064
intracytoplasmic nanocrystals (Qdots), was sufficient for Moreau MJ, Morin I, Schaeffer PM (2010) Quantitative determination of
protein stability and ligand binding using a green fluorescent protein
both paraffin-embedded and frozen sections, enabling the reporter system. Mol BioSyst 6:12851292
tracking of g-ADSCs, observed by fluorescence microscopy. Moseley TA, Zhu M, Hedrick MH (2006) Adipose-derived stem and pro-
It has fundamental importance for assessing compliance and genitor cells as fillers in plastic and reconstructive surgery. Plast
monitoring of migration of infused cells in different tissues, as Reconstr Surg 118:121128
Muccioli M, Pate M, Omosebi O, Benecia F (2011) Generation and
well as its relevance for understanding the biology of stem labeling of murine bone marrow-derived dendritic cells with Qdot
cells in vitro and in vivo, because it is photostable and bright, nanocrystals for tracking studies. J Vis Exp 52:15
and thus has applications for local and systemic route studies. Nakagami H, Morishita R, Maeda K, Kikuchi Y, Ogihara T, Kaneda Y
(2006) Adipose tissue-derived stromal cells as a novel option for
regenerative cell therapy. J Atheroscler Thromb 13:7781
Noh YW, Lim YT, Chung BH (2008) Noninvasive imaging of dendritic
Acknowledgements We are thankful to the National Scientific cell migration into lymph nodes using near-infrared fluorescent
and Technological Development Council-CNPq (Process: 552400/ semiconductor nanocrystals. FASEB J 22:39083918
11-4; 311 684/2012-2) for their financial support. This study was Ocarino NM, Bozzi A, Pereira RD, Breyner NM, Silva VL, Castanheira
carried out in strict accordance with the recommendations in the P, Goes AM, Serakides R (2008) Behavior of mesenchymal stem
Guide for the Care and Use of Laboratory Animals of the cells stained with 4, 6-diamidino-2-phenylindole dihydrochloride
National Institutes of Health. The protocol was approved by the (DAPI) in osteogenic and non osteogenic cultures. Biocell 32:
Committee on the Ethics of Animal Experiments of the Federal 175183
Univesity of Piau (Permit Number 037/2012). Oliveira DM, Almeida BO, Marti LC, Sibov TT, Pavon LF, Malheiros
DMAC, Campos AH (2009) Labeling of human mesenchymal stem
cells with QDs allows tracking of transplanted cells engrafted in
infarcted pig hearts. Einstein 7:284289
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