EXPERIMENTAL
Influences of Preservation at Various
Temperatures on Liposuction Aspirates
Daisuke Matsumoto, M.D.
Tomokuni Shigeura, M.S.
Katsujiro Sato, M.D.
Keita Inoue, M.D.
Hirotaka Suga, M.D.
Harunosuke Kato, M.D.
Noriyuki Aoi, M.D.
Syoko Murase, Ph.D.
Koichi Gonda, M.D.
Kotaro Yoshimura, M.D.
Tokyo and Kanagawa, Japan

Background: Aspirated fat is not only a filler material but also an abundantsource of adipose-derived stem cells. The aim of this study was to assess degeneration of aspirated fat during preservation and optimize the preservation
method for lipoaspirates.
Methods: Aspirated fat was preserved at room temperature for 1, 2, 4, and 24
hours (n ⫽ 10 each); at 4°C for 1, 2, and 3 days (n ⫽ 14 each); or at – 80°C for
1 month (n ⫽ 3). Morphologic changes were assessed with scanning electron
microscopy. Adipose-derived stem cell yield was measured after 1 week of culture. For aspirated fat preserved at room temperature, damaged adipocytes wereassessed by measuring the oil volume ratio after centrifugation (n ⫽ 6) andglycerol-3-phosphate-dehydrogenase activity in washing solution (n ⫽ 4). Cellsurface marker expression was examined by flow cytometry (n ⫽ 3).
Results: Although the scanning electron microscopic assay indicated no remarkable anatomical changes based on preservation methods, oil volume significantly increased in fat preserved at room temperature for 4 hours. Adiposederived stem cell yield was significantly reduced by preservation at room
temperature for 24 hours and by preservation at 4°C for 2 or 3 days. Flowcytometric analysis suggested that the biological properties of adipose-derived stem cells did not significantly change at 4°C up to 3 days. The cells
were isolated from cryopreserved fat, but the yield was much less than thatfrom fresh aspirated fat.
Conclusions: Aspirated fat should be transplanted as quickly as possible if
it is preserved at room temperature. For adipose-derived stem cell isolation,
aspirated fat can be stored or transported overnight if it is preserved at 4°C
without adipose-derived stem cell yield loss or changes in biologicalproperties. (Plast. Reconstr. Surg. 120: 1510, 2007.)

Although fat tissue has been used as a filler
material for more than 100 years,1,2 there
are several problems to be resolved, including unpredictability and fat necrosis resulting in infection and calcification.2,3 Autologous
fat transfer, however, is almost the only methodof soft-tissue augmentation that can be performed without detectable scarring on either adonor or a recipient site and without complications associated with foreign materials. Because
fat tissue is more easily damaged by ischemia
compared with other tissues such as skin andbone, transferring fat tissue as quickly as possible
From the Department of Plastic Surgery, University of Tokyo
School of Medicine, and the Division of Research and Development, Biomaster, Inc.
Received for publication August 15, 2006; accepted January
4, 2007.
Copyright ©2007 by the American Society of Plastic Surgeons
DOI: 10.1097/01.prs.0000288015.70922.e4

1510

after harvesting is recommended. Thus, it is ofgreat interest to investigate how adipose tissue
viability after aspiration changes over time depending on preservation methods. Clinically, aspirated fat tissue is usually preserved at room
temperature in a suction bottle, but how aspirated fat in the suction bottle changes during the
postoperative hours is unknown.
It was recently revealed that adipose tissue is aremarkable source of multipotent stem cells,4,5
which can differentiate into adipogenic, chondrogenic, osteogenic, myogenic, neurogenic, endothelial, and other lineages.6 Adipose-derived
stem cells have already been used in some clinical trials, including treatments for bone defects7;
rectovaginal fistula8; and soft-tissue augmentation, including breast enhancement, breast reconstruction, and facial rejuvenation.9 Adiposederived stem cells may be clinically used orbanked also for other therapeutic purposes in

www.PRSJournal.com

Volume 120, Number 6 • Preservation of Liposuction Aspirates
the near future. In practice, there is some timelag between liposuction and cell isolation; ittakes a few to several hours for liposuction surgery, depending on the volume and sites to suction, and a few hours to even a day or two fortransportation from an operating room to a cellprocessing unit. To maximize the potential of
adipose aspirates as a stem cell source, it is very
important to optimize protocols for their preservation.
Thus, aspirated fat is now valuable as an autologous filler material and as an abundant stem
cell source. We sought to comprehensively evaluate the influences of preservation at different
temperatures on the viability of aspirated fat andadipose-derived stem cells.

PATIENTS AND METHODS
Human Tissue Sampling and Preservation
We obtained liposuction aspirates from 21
healthy female donors undergoing liposuction of
the abdomen or thighs after informed consent
using an institutional review board–approved protocol. The adipose portion of the liposuction aspirates was preserved at room temperature for 1,
2, 4, or 24 hours (n ⫽ 10); at 4°C for 0, 1, 2, and3 days (n ⫽ 14); or at – 80°C for 1 month (n ⫽ 3)
and subjected to assays as described below. Excised fat obtained from a cosmetic surgery patient
was also used for comparison.
Cell Processing and Culture
Stromal vascular fractions were isolated fromthe fatty portion of liposuction aspirates as previously described.10 Briefly, the aspirated fat waswashed with phosphate-buffered saline and digested on a shaker at 37°C in phosphate-bufferedsaline containing 0.075% collagenase for 30 minutes. Mature adipocytes and connective tissues
were separated from pellets by centrifugation (800 g
for 10 minutes). The cell pellets were resuspended, filtered with a 100-␮m mesh (Millipore,Bedford, Mass.), plated (30,000 cells/cm2) onto
gelatin-coated dishes, and cultured at 37°C in an
atmosphere of 5% carbon dioxide in humid air.
The culture medium was M-199 containing 10%
fetal bovine serum, 100 IU of penicillin, 100
mg/ml streptomycin, 5 ␮g/ml heparin, and 2
ng/ml acidic fibroblast growth factor. Medium
was replaced every third day. After 7 days, adherent cells were trypsinized and counted witha cell counter (NucleoCounter; ChemoMetec,
Allerod, Denmark).

Flow Cytometric Analysis
Adherent adipose-derived stem cells were examined for surface marker expression using flow
cytometry after 1 week of culture (n ⫽ 3). The
following monoclonal antibodies were used:
CD29-PE; CD31-PE; CD34-PE; CD45-PE; CD90PE; CD133-PE; CD144-PE; HLA-A, B, and C-PE;
Tie-2-PE (BD Biosciences, San Diego, Calif.);
CD105-PE (Serotec, Oxford, United Kingdom);and Flk-1-PE (Techne, Princeton, N.J.). Cells wereincubated with the directly conjugated monoclonal antibodies in phosphate-buffered saline containing 0.5% bovine serum albumin for 30 minutes at 4°C, washed with phosphate-bufferedsaline containing 0.2% bovine serum albumin,and diluted in phosphate-buffered saline containing 0.1% bovine serum albumin. Flow cytometricanalyses were performed using an LSR2 (Becton
Dickinson, San Jose, Calif.).
Quantitative Analysis of Damaged Adipocytes in
Aspirated Fat
To assess damaged adipocytes in aspirated fat,
we measured the ratios of oil and fat volumes after
centrifugation (n ⫽ 6), and an extracellular activity of an enzyme, glycerol-3-phosphate-dehydrogenase, with a GPDH Activity Measurement Kit(Cell Garage, Tokyo, Japan) (n ⫽ 4). The enzyme
is normally strictly intracellular, and the enzyme
activity in a washing solution is proportional to theamount of adipocyte destruction.
For measurement of the oil ratio, the fatty
portion of liposuction aspirates was divided into
20 tubes (15-ml conical tubes) and preserved atroom temperature. After preservation for 1, 2, 4,
or 24 hours, five of the tubes were centrifuged
at 2330 g for 5 minutes to separate the oil, fat,and fluid into distinct layers from top to bottom(Fig. 1). The oil ratio was calculated as follows:
oil ratio ⫽ (oil volume)/[(oil volume) ⫹ (fatvolume)]. Data were collected from lipoaspirates
obtained from six patients.
For extracellular glycerol-3-phosphate-dehydrogenase assessment, after 1 g of aspirated fat
lobules was washed in a tube with 4 ml of phosphate-buffered saline, 1 ml of the phosphate-buffered saline solution was transferred to anothertube and centrifuged at 15,000 g for 5 minutes.
After the supernatant was appropriately (two to 20
times) diluted with the enzyme extracting reagent,
100 ␮l of each diluted sample was transferred onto
a 96-well plate. The enzyme reaction was thenstarted by the addition of 200 ␮l of substrate reagent containing dihydroxyacetone phosphate

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Plastic and Reconstructive Surgery • November 2007
dylate buffer for 1 week at room temperature andthen fixed in 1% osmium tetroxide. After dehydration, samples were dried with a supercriticalpoint carbon dioxide dryer (HCP-2; Hitachi, Tokyo, Japan), sputter-coated with platinum/palladium, and examined with a scanning electron microscope (S3500N; Hitachi).
Statistical Analysis
Results were expressed as mean ⫾ SE. Pairedor unpaired t tests were performed to evaluate
differences between groups, and no correction
was made for multiple comparisons.

RESULTS

Fig. 1. Analysis of adipocyte damage in lipoaspirates by centrifugation. After centrifugation, aspirated fat tissue was separated
into distinct layers from top to bottom: the oil, fat, and fluid layers.
Adipocyte damage by preservation was quantified by calculating the oil ratio in the volume as follows: oil ratio ⫽ (oil volume)/[(oil volume) ⫹ (fat volume)].

and nicotinamide adenine dinucleotide (NADH).
The assay measures the reduction of NADH to
NAD⫹ in the presence of glycerol-3-phosphatedehydrogenase. The absorbance at 340 nm wasrecorded every 30 seconds for 10 minutes by using
a spectrometer (DTX880; Beckman Coulter, Fullerton, Calif.) to produce an extinction curve forcalculating the maximum decrease, indicating theglycerol-3-phosphate-dehydrogenase activity.
Procedure for Cryopreserving and Thawing
Aspirated Fat
Fresh adipose tissue was mixed with an equalamount of freezing medium (Cell Banker BLC-1;
ZENOAQ, Fukushima, Japan) and cooled to– 80°C in a programmable freezing system (TNP87S; Nihon Freezer, Tokyo, Japan) at a freezing
rate of 1°C every 15 minutes. After 1 month, cryopreserved adipose tissue was thawed by placementof the cryotube into a water bath at 37°C.
Scanning Electron Microscopic Study
After preservation at room temperature or
4°C, aspirated fat was fixed with 2% paraformaldehyde and 2.5% glutaraldehyde in 0.2 M caco-

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Morphology of Adipocytes in Aspirated Fat
Aspirated fat was preserved at 4°C and fixed
for scanning electron microscopic study at different time points. Adipocytes from aspirated fat almost retained their round shape and showed nosignificant morphologic differences comparedwith those of excised fat (Fig. 2, above, left). No
remarkable change in adipocyte morphology was
found in aspirated fat tissues on days 0, 1, and 3(Fig. 2, above, second from left, second from right, andright).
Aspirated fat was preserved for 1, 2, 4, or 24hours at room temperature and evaluated by scanning electron microscopy also (Fig. 2, below). No
remarkable difference in adipocyte morphology
was identified.
Degeneration of Adipocytes with Preservation
Time
Because we clinically observe a gradual increase of oil volume in lipoaspirates, the oil ratio(oil volume/[oil volume ⫹ fat volume]) was used
as one of indices of degeneration of adipocytes inaspirated fat. The oil ratio increased over time
during preservation at room temperature (Fig. 3,
above, left). Oil ratios at 4 and 24 hours were significantly greater than those at 1 hour, and the oil
ratio per-hour increase slowed remarkably after 4hours (Fig. 3, above, right). Glycerol-3-phosphatedehydrogenase activity in a washing solution ofpreserved fat increased slightly up to 4 hours andincreased markedly after 24 hours of preservation(Fig. 3, below, left). Unlike the results with the oil
ratio, glycerol-3-phosphate-dehydrogenase activity
increase per hour did not change up to 4 hours
but significantly increased after that time point.

Volume 120, Number 6 • Preservation of Liposuction Aspirates

Fig. 2. Comparison with scanning electron microscopy of human aspirated fat tissues after preservation at 4°C or room temperature.
(Above, left) Excised adipose tissue was fixed immediately after the operation. Aspirated fat tissues preserved at 4°C were fixed on day
0 (above, second from left), day 1 (above, second from right), or day 3 (above, right), whereas those preserved at room temperature were
fixed at 1 hour (below, left), 2 hours (below, second from left), 4 hours (below, second from right), or 24 hours (below, right) after theoperation. Each sample was treated for evaluation with scanning electron microscopy, and representative photographs are shown.
No significant morphologic changes over time were found by scanning electron microscopy in aspirated fat, even in samples stored
at 4°C for 3 days or at room temperature for 24 hours. Scale bar ⫽ 250 ␮m.

Adipose-Derived Stem Cell Yield from
Aspirated Fat Preserved at Room or Cool
Temperature
When preserved at room temperature, adipose-derived stem cell yield was maintained up to
4 hours and decreased remarkably at 24 hours(Fig. 4). In contrast, after preservation at 4°C,
almost the same number of adipose-derived stem
cells were isolated from aspirated adipose tissue
on days 0 and 1 (Fig. 5). The number of isolated
adipose-derived stem cells was extensively decreased in some cases on day 2 and in all cases on
day 3. Statistical significance was seen between
days 0 and 3 (p ⬍ 0.001) and between days 1 and
3 (p ⬍ 0.001).

ervation time at a cool temperature, surface
marker analysis was performed on adipose-derived
stem cells isolated from aspirated fat tissues preserved for 0, 1, 2, and 3 days at 4°C (Table 1).

Surface Marker Expression of Adipose-Derived
Stem Cells Isolated from Aspirated Fat
Preserved at a Cool Temperature
To examine changes in the biological properties of adipose-derived stem cells based on pres-DISCUSSION

Adipose-Derived Stem Cell Yield from Cryopreserved Aspirated Fat
We also evaluated the possibility of isolating
adipose-derived stem cells from aspirated fat cryopreserved for 30 days (n ⫽ 3). Adipose-derived
stem cells were harvested from the cryopreserved
aspirated fat, but the adipose-derived stem cell
yield was significantly less than that obtained from
fresh aspirated fat (Fig. 6).

The scanning electron microscopic assay
showed that no significant morphologic change inadipocytes was found among aspirated fat tissues
preserved either at 4°C for up to 3 days or at1513

Plastic and Reconstructive Surgery • November 2007

Fig. 3. Assessment of damaged adipocytes after preservation at room temperature. Damaged adipocytes were evaluated by measurement of the oil ratio after centrifugation (above) or glycerol-3-phosphate-dehydrogenase activity of preserved fat in washingsolution (below). Statistical analysis was performed using paired t tests between groups. Values are mean ⫾ SE; *p ⬍ 0.05. (Above, left)
Oil ratios in aspirated fat preserved at room temperature for 1, 2, 4, or 24 hours are shown. The oil volume ratio gradually increased
with storage time, likely because of the breakdown of adipocytes. (Above, right) The oil ratio per-hour increase slowed remarkably
with storage time. (Below, left) Glycerol-3-phosphate-dehydrogenase activity of preserved fat in washing solution for 1, 2, 4, or 24hours was measured for assessment of damaged adipocytes. Glycerol-3-phosphate-dehydrogenase activity had slightly increased
up to 4 hours and was markedly increased at 24 hours. (Below, right) Glycerol-3-phosphate-dehydrogenase activity increase per hour
did not change significantly up to 4 hours, but did increase after 4 hours.

room temperature for up to 24 hours. However,quantitative analysis of damaged adipocytes bymeasuring the oil ratio and glycerol-3-phosphatedehydrogenase activity revealed that preserved
adipocytes were partly degenerated and ruptured
over preservation time when stored at room temperature. Thus, preservation at room temperature
resulted in damage to some adipocytes that may
have been located superficially; however, the remaining adipocytes retained almost-intact morphology.
In this study, damaged adipocytes were evaluated by measurement of the oil ratio after centrifugation and glycerol-3-phosphate-dehydrogenase activities in washing solution of preserved fat.
Boschert et al.11 reported that centrifugation at
greater than 100 g caused adipose cell destruction;
however, we recently found that centrifugation at

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400 g increased the oil portion in lipoaspirates butthat further centrifugation did not significantly
damage adipocytes or increase oil volume.12 In
addition, our histologic examinations of centrifuged aspirated fat with light and scanning electron microscopy showed that adipocytes appeared
to be intact even after centrifugation at 4300 g.12
Therefore, we considered that the increased oilvolume after centrifugation (for 5 minutes at 2330
g) in the present study was attributable to adipocyte damage from preservation at room temperature. The measurement of glycerol-3-phosphatedehydrogenase activity produced results that weresimilar to those of the oil ratio up to 4 hours butthat differed somewhat at 24 hours; glycerol-3phosphate-dehydrogenase activity had increased
remarkably after 24 hours of preservation. Thisdifference might be attributable to incomplete

Volume 120, Number 6 • Preservation of Liposuction Aspirates
Table 1. Surface Marker Expression of AdiposeDerived Stem Cells Isolated from Aspirated Fat
Tissues Preserved at 4ºC*

Fig. 4. Adipose-derived stem cell yield after preservation atroom temperature. We preserved aspirated adipose tissues atroom temperature for 1, 2, 4, or 24 hours and processed them forisolation of adipose-derived stem cells, which were then cultured
for 1 week. Ratios of adipose-derived stem cell yield to control(1-hour preservation) were calculated; data were obtained fromthree patients, and statistical analysis was performed using
paired t tests between 1-hour and other groups. Adipose-derived
stem cell yield seemed to be maintained for up to 4 hours of preservation and decreased remarkably when preserved for 24 hours
at room temperature. Values are mean ⫾ SE; *p ⬍ 0.05.

Fig. 5. Adipose-derived stem cell yields after preservation at 4°C.
We preserved aspirated fat tissues at 4°C for 0, 1, 2, and 3 days andprocessed them for isolation of adipose-derived stem cells,
which were then cultured for 1 week. Ratios of adipose-derived
stem cell yield to control (day 0, no preservation) were calculated;data were obtained from 14 patients (data for day 2 came fromfour of the 14 patients), and statistical analysis was performed
using unpaired t tests between day 0 and other groups. A statistical difference in adipose-derived stem cell yield was not foundbetween days 0 and 1, whereas adipose-derived stem cell yielddecreased significantly on days 2 and 3. Values are mean ⫾ SE;*p ⬍ 0.05.

release of leaked intracellular oil from damaged
adipocytes; the oil might be trapped in the surrounding matrices, proving difficult to measureusing the oil ratio assay. Thus, the present result

CD29
CD31
CD34
CD45
CD90
CD105
CD133
CD144
HLA-ABC
Flk-1
Tie-2

Day 0

Day 1

Day 2

Day 3

98.7 ⫾ 0.8
1.7 ⫾ 0.3
48.1 ⫾ 6.5
1.0 ⫾ 0.3
99.2 ⫾ 0.5
97.8 ⫾ 1.3
3.1 ⫾ 1.5
0.5 ⫾ 0.2
98.3 ⫾ 0.8
56.1 ⫾ 17.2
2.2 ⫾ 17.2

98.4 ⫾ 0.8
2.0 ⫾ 0.4
46.3 ⫾ 5.5
1.2 ⫾ 0.4
98.1 ⫾ 0.9
97.0 ⫾ 2.2
2.2 ⫾ 1.6
0.5 ⫾ 0.1
97.4 ⫾ 1.6
54.4 ⫾ 26.5
2.3 ⫾ 0.9

99.4 ⫾ 0.1
1.4 ⫾ 0.8
35.8 ⫾ 5.2
0.7 ⫾ 0.2
99.2 ⫾ 0.1
97.8 ⫾ 0.8
0.9 ⫾ 0.6
0.2 ⫾ 0.1
96.5 ⫾ 1.6
81.7 ⫾ 4.4
1.7 ⫾ 0.6

97.3 ⫾ 1.8
0.7 ⫾ 0.1
31.2 ⫾ 3.4
1.1 ⫾ 0.6
97.1 ⫾ 2.1
96.2 ⫾ 2.1
1.0 ⫾ 0.4
0.2 ⫾ 0.1
93.6 ⫾ 3.0
71.0 ⫾ 5.5
1.8 ⫾ 0.4

*Adipose-derived stem cells isolated from aspirated fat preserved at
4ºC for 0, 1, 2, and 3 days and flow cytometric analyses were performed on adipose-derived stem cells after culture for 1 week. Few
differences in expression profile of principal surface markers wereobserved among groups, suggesting that biological properties ofadipose-derived stem cells do not change by preservation at 4ºC for
up to 3 days. Vales are mean ⫾ SE.

Fig. 6. Adipose-derived stem cell yields from cryopreserved lipoaspirates. Fresh aspirated adipose tissue was mixed with anequal amount of freezing medium, cooled to – 80°C in a programmable freezing system, and stored at – 80°C for 1 month.
The cryopreserved adipose tissue was thawed and processed
to isolate adipose-derived stem cells, which were then cultured for 1 week. The ratio of adipose-derived stem cell yield
to control (fresh lipoaspirates) was calculated. Data were obtained from three patients, and statistical analysis was performed using paired t tests between groups. Adipose-derived
stem cells were isolated from cryopreserved aspirated fat(6.7 ⫾ 4.7 ⫻ 104 cells/ml after 1 week of culture), but the yield
was much less than that of the fresh fat. Values are mean ⫾ SE;*p ⬍ 0.05.

indicated that preservation at room temperaturefor 4 hours significantly damaged adipocytes inaspirated fat; thus, lipotransfer should be performed as quickly as possible after aspiration, especially when a large volume of aspirated fat is tobe transplanted.

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Plastic and Reconstructive Surgery • November 2007
Since the reports showing that adipose tissue
contains multipotent stem cells,4,5 aspirated adipose tissue has been regarded as not only a filler
material but also as an abundant source of stem
cells. Adipose-derived stem cells reside in adipose
tissue as progenitors of adipocytes, but it has
been suggested that adipose-derived stem cells
can differentiate into vascular endothelial
cells,13,14 can release angiogenetic factors underhypoxic conditions,15 and can contribute to ahigher graft take of transplanted fat.14,16 In thecurrent study, adipose-derived stem cell yield wasmaintained up to 4 hours at room temperature,
and an adipose-derived stem cell yield similar tothat of fresh aspirated fat was obtained from that
preserved at 4°C for 24 hours. This finding indicates that a 1-day delay in isolation of adiposederived stem cells from aspirated fat can be appropriate when the tissue is stored in arefrigerator. Therefore, overnight cooling transportation of aspirated fat to a specialized cell-processing center for isolation and banking of adipose-derived stem cells can be regarded aspractical, although adipose-derived stem cell yield
after 2 or 3 days would be uncertain, even withpreservation at 4°C.
Isolated adipose-derived stem cells can be frozen, thawed, and cultured again as well as almost
any other cell type. However, whether aspirated fat
tissue can be frozen as an effective filler material
or a source of adipose-derived stem cells has not
yet been established. In this study, we tried toisolate adipose-derived stem cells from aspirated
fat cryopreserved for 30 days and from fresh aspirated fat. The adipose-derived stem cell yieldfrom cryopreserved fat was much lower than thatof fresh aspirated fat. We tried several kinds offreezing conditions (rapid or slow freezing) andother freezing media (Dulbecco’s Modified Eagle
Medium or M-199 containing 10% dimethyl sulfoxide with 1% methylcellulose or 1% trehalose or
1 to 20% gelatin, or their mixture), but the adipose-derived stem cell yield was not improved(data not shown). Although there were a numberof red blood cells contaminating cell fractions isolated from fresh aspirated fat, almost no red blood
cells contaminated those from cryopreserved aspirated fat. Our result with adipose-derived stem
cell yield from cryopreserved fat contradicts a recent report17 showing that the adipose-derived
stem cell yield from cryopreserved lipoaspirates
was approximately 90 percent of that from fresh
lipoaspirates. The reported adipose-derived stem
cell yield from cryopreserved adipose is 3.7 ⫾1.4 ⫻ 105 cells/ml after 2 weeks of culture,17 which1516

is comparable to our result (6.7 ⫾ 4.7 ⫻ 104
cells/ml after 1 week of culture), because adiposederived stem cells proliferate 10 to 100 times in a
week, depending on culture conditions. However,
the reported adipose-derived stem cell yield from
fresh adipose (4.1 ⫾ 1.4 ⫻ 105 cells/ml after 2
weeks of culture17) was much less than that in thisstudy (7.9 ⫾ 1.5 ⫻ 105 cells/ml after 1 week ofculture). It is unknown why the reported adiposederived stem cell yield from fresh adipose differs
between the two studies, but it may be the resultof different methods of cell isolation.

CONCLUSIONS
We have demonstrated how the adipose-derived stem cell yield from aspirated fat changes
depending on preservation conditions and time
periods. Preservation for 4 hours at room temperature significantly damaged adipocytes but did not
significantly alter adipose-derived stem cell yield.
Adipose-derived stem cell yield significantly decreased with preservation for 24 hours at room
temperature but not with preservation at 4°C.
Thus, aspirated fat can be transported to a cellprocessing center for cell isolation on the day after
harvesting and for subsequent banking if it is kept
at 4°C. Adipose-derived stem cell yield from cryopreserved aspirated fat was minimal, and a further
optimization of methodology of freezing and preservation is needed for practical use of cryopreservation of aspirated fat intended as an adiposederived stem cell source.
Kotaro Yoshimura, M.D.
Department of Plastic Surgery
University of Tokyo School of Medicine
7-3-1, Hongo, Bunkyo-Ku
Tokyo 113-8655, Japan
yoshimura-pla@h.u-tokyo.ac.jp

ACKNOWLEDGMENTS

This study was supported by Japanese Ministry of
Education, Culture, Sports, Science, and Technology
grant-in-aid B2-16390507. The authors thank Dr. Satoru Fukuda for assistance in the histologic assay withscanning electron microscopy.
DISCLOSURE

None of the authors has any commercial associations
or financial interests that might pose or create a conflictof interest with information presented in this article.
REFERENCES
1. Neuber, K. Fettgewebstransplantation. Verh. Dtsch. Ges. Chir.
1: 66, 1893.
2. Shiffman, M. A., and Mirrafati, S. Fat transfer techniques:
The effect of harvest and transfer methods on adipocyte

Volume 120, Number 6 • Preservation of Liposuction Aspirates
viability and review of the literature. Dermatol. Surg. 27: 819,
2001.
3. Ersek, R. A., Chang, P., and Salisbury, M. A. Lipo layering ofautologous fat: An improved technique with promising results. Plast. Reconstr. Surg. 101: 820, 1998.
4. Zuk, P. A., Zhu, M., Mizuno, H., et al. Multilineage cells fromhuman adipose tissue: Implications for cell-based therapies.
Tissue Eng. 7: 211, 2001.
5. Zuk, P. A., Zhu, M., Ashjian, P., et al. Human adipose tissue
is a source of multipotent stem cells. Mol. Biol. Cell 13: 4279,
2002.
6. Tholpady, S. S., Llull, R., Ogle, R. C., et al. Adipose tissue:
Stem cells and beyond. Clin. Plast. Surg. 33: 55, 2006.
7. Lendeckel, S., Jodicke, A., Christophis, P., et al. Autologous
stem cells (adipose) and fibrin glue used to treat widespreadtraumatic calvarial defects: Case report. J. Craniomaxillofac.
Surg. 32: 370, 2004.
8. Garcia-Olmo, D., Garcia-Arranz, M., Herreros, D., et al. A
phase I clinical trial of the treatment of Crohn’s fistula byadipose mesenchymal stem cell transplantation. Dis. Colon
Rectum 48: 1416, 2005.
9. Yoshimura, K., Sato, K., Aoi, N., et al. Cell-assisted lipotransfer (CAL) for cosmetic breast augmentation: Supportive useof adipose derived stem/stromal cells [epub September 1,
2007.] Aesthetic Plast. Surg. DOI:10.1007/s00266-007-9019-4.
10. Yoshimura, K., Shigeura, T., Matsumoto, D., et al. Characterization of freshly isolated and cultured cells derived fromthe fatty and fluid portions of liposuction aspirates. J. Cell.
Physiol. 208: 64, 2006.
11. Boschert, M. T., Beckert, B. W., Puckett, C. L., et al. Analysis
of lipocyte viability after liposuction. Plast. Reconstr. Surg. 109:
761, 2002.
12. Kurita, M., Matsumoto, D., Shigeura, T., et al. Influences ofcentrifugation on cells and tissues in liposuction aspirates:
Optimized centrifugation for lipotransfer and cell isolation.
Plast. Reconstr. Surg. (in press).
13. Miranville, A., Heeschen, C., Sengenes, C., et al. Improvement of postnatal neovascularization by human adipose tissue-derived stem cells. Circulation 110: 349, 2004.
14. Matsumoto, D., Sato, K., Gonda, K., et al. Cell-assisted lipotransfer (CAL): Supportive use of human adipose-derived
cells for soft tissue augmentation with lipoinjection. TissueEng. 12: 3375, 2006.
15. Rehman, J., Traktuev, D., Li, J., et al. Secretion of angiogenic
and antiapoptotic factors by human adipose stromal cells.
Circulation 109: 1292, 2004.
16. Masuda, T., Furue, M., and Matsuda, T. Novel strategy forsoft tissue augmentation based on transplantation of fragmented omentum and preadipocytes. Tissue Eng. 10: 1672,
2002.
17. Pu, L. L., Cui, X., Fink, B. F., et al. Adipose aspirates as asource for human processed lipoaspirate cells after optimal
cryopreservation. Plast. Reconstr. Surg. 117: 1845, 2006.

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