Review ArticleMinimally Manipulated Mesenchymal Stem Cells for theTreatment of Knee Osteoarthritis: A Systematic Review ofClinical Evidence

B. Di Matteo ,1,2 F. Vandenbulcke ,1,2 N. D. Vitale,1,2 F. Iacono,1,2 K. Ashmore,1,2

M. Marcacci,1,2 and E. Kon 1,2,3

1Department of Biomedical Sciences, Humanitas University, Via Manzoni 113, 20089 Rozzano, Milan, Italy2Humanitas Clinical and Research Center, Via Manzoni 56, 20089 Rozzano, Milan, Italy3First Moscow State Medical University, Sechenov University, Moscow, Russia

Correspondence should be addressed to F. Vandenbulcke; filippovdb@gmail.com

Received 22 January 2019; Accepted 21 July 2019; Published 14 August 2019

Guest Editor: Brian Johnstone

Copyright © 2019 B. Di Matteo et al. This is an open access article distributed under the Creative Commons Attribution License,which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Background. The use of laboratory-expanded mesenchymal stem cells (MSCs) is subject to several restrictions, resulting in“minimal manipulation” methods becoming the current most popular strategy to increase the use of MSCs in an orthopaedicpractice. The aim of the present systematic review is to assess the clinical applications of “minimally” manipulated MSCs, eitheras bone marrow aspirate concentrate (BMAC) or as stromal vascular fraction (SVF), in the treatment of knee osteoarthritis(OA). Methods. A systematic review of three databases (PubMed, ScienceDirect, and Google Scholar) was performed using thefollowing keywords: “Knee Osteoarthritis” with “(Bone marrow aspirate) OR (bone marrow concentrate)” or with “(adipose-derived mesenchymal stem cells) OR (adipose derived stromal cells) OR (stromal vascular fraction) OR (SVF)” as eitherkeywords or MeSH terms. The reference lists of all retrieved articles were further reviewed for identification of potentiallyrelevant studies. Results. Twenty-three papers were included in the final analysis (10 on BMAC and 13 on SVF). Of these, only 4were randomized controlled trials (RCTs). Bias risk evaluation, performed using a modified Coleman score, revealed an overallpoor quality of the studies. In terms of clinical application, despite the apparent safety of minimally manipulated MSCs and theshort-term positive clinical outcomes associated with their use, clinicians reported different preparation and administrationmethods, ranging from single intra-articular injections to intraosseous applications to administration in combination with othersurgical procedures. Conclusions. The available literature is undermined by both the lack of high-quality studies and the variedclinical settings and different protocols reported in the few RCTs presently published. This prevents any recommendation onthe use of either product in a clinical practice. Nevertheless, the use of minimally manipulated MSCs (in the form of BMAC orSVF) has been shown to be safe and have some short-term beneficial effects.

1. Background

Among the large number of treatment possibilities for kneeosteoarthritis (OA), novel regenerative medicine strategiesare an area of growing interest [1]. This is especially the casein the challenging subset of younger OA patients, who havehigh functional demands yet limited indications for invasivesurgical treatments. Nowadays, the issue has even extendedto middle-aged active patients who increasingly expect tomaintain a high activity level and postpone or avoid metal

resurfacing. Despite the rising incidence of OA, no effectivetherapies have been shown to fully restore the original fea-tures and structure of the damaged articular surface. Further-more, no “conservative” technique is able to amelioratewidespread damage to all articular tissues, which arises dueto OA affecting the entire joint environment [2, 3].

Innovative therapies, ranging from platelet-derivedgrowth factors (GFs) to cell-based treatments (sometimescombined with various biomaterials) [4, 5], have been pro-posed as potential solutions for these patients. Mesenchymal

HindawiStem Cells InternationalVolume 2019, Article ID 1735242, 14 pageshttps://doi.org/10.1155/2019/1735242

stem cells (MSCs) have emerged as a possible therapeuticoption, thanks to their multilineage differentiation potential[6]. Mesenchymal cells are indeed able to differentiate intoseveral cell types such as osteoblasts, chondrocytes, or adipo-cytes and have high plasticity, limited self-renewal capabili-ties, and immune-suppressive and anti-inflammatoryactions [7]. Growth factors, cytokines, bioactive lipids, andmicrovesicles that are released from stem cells may also exertbeneficial effects including angiopoietic and antiapoptoticactions. Studies have also highlighted the presence of MSCsin numerous adult tissues including adipose and muscle tis-sues, the dermis, periosteum, synovial membrane, synovialfluid, and articular cartilage [8].

Choosing the correct stem cell approach is imperative inachieving optimal results in regenerative medicine. Amongthe available sources of MSCs, two have received the greatestscientific attention: adipose-derived mesenchymal stem cells(ASCs) and the more studied bone marrow mesenchymalstem cells (BMSCs) [9, 10].

The clinical application of MSCs is strictly regulatedespecially in regard to cell expansion. In this case, despitebeing an autologous product, MSC products are considered“drugs,” and therefore, their use is extremely limited in a rou-tine clinical setting, both in Europe and the USA [11]. Thisregulatory burden has incentivized clinicians to developalternative strategies for MSC use, resulting in the currentlycrucial concept of “minimal manipulation.” If cells are notexpanded but are rather manipulated within or “nearby”the operating room (OR), MSC application is easier and sim-pler, since the cells can be administered outside the bound-aries of clinical trials (approved by local authorities andNational Health Ministries). Following this reasoning, twomain treatment modalities have emerged: bone marrow aspi-rate concentrate (BMAC) and adipose-derived stromal vas-cular fraction (SVF). BMAC is obtained via bone marrowneedle aspiration (which can be performed on various sitesbut most commonly on the iliac crest) and subsequent con-centration via dedicated centrifuges which can be trans-ported and used directly within the OR [7, 12]. On theother hand, collection of adipose-derived SVF involves mul-tiple steps: first, a liposuction is performed to obtain adiposetissue. In order to isolate the ASCs from the extracellularmatrix (ECM), collagenase is added to the lipoaspirate [13].Subsequently, the collagenase in the mixture is removed viaa dilution method, which involves washing the lipid-enzyme mixture with normal saline solution, followed (insome cases) by sterile centrifugation. This results in the finalSVF product, ready to be administered to the patient.

The aim of the present systematic review is to assess theclinical applications of “minimally manipulated”MSCs, bothBMAC and SVF, in the treatment of knee OA.

2. Materials and Methods

2.1. Literature Search Strategy.We conducted this systematicreview according to the Preferred Reporting Items for Sys-tematic Reviews and Meta-Analyses (PRISMA) guidelines[14]. A systematic review of three medical electronic data-bases (PubMed, ScienceDirect, and Google Scholar) was per-

formed by two independent authors (N.D.V. and F.V.) from1998 to 20/10/2018. To achieve maximum search strategysensitivity, we combined the terms “Knee Osteoarthritis” or“Knee OA” with “(Bone marrow aspirate) OR (bone marrowconcentrate)” or with “(adipose-derived mesenchymal stemcells) OR (adipose derived stromal cells) OR (stromal vascu-lar fraction) OR (SVF)” as either keywords or MeSH terms.The reference lists of all retrieved articles were furtherreviewed for identification of potentially relevant studiesand assessed using the inclusion and exclusion criteria statedbelow.

Eligible studies for the present systematic review includedthose dealing with the intra-articular use of minimallymanipulated BMAC or SVF in knee OA. The initial titleand abstract screening was made using the following inclu-sion criteria: studies of any level of evidence, written inEnglish, and reporting clinical results following the intra-articular application of either BMAC or SVF as a treatmentapproach for OA. Conversely, articles dealing with expandedor otherwise manipulated mesenchymal stem cells wereexcluded. Papers where MSCs were applied for other clinicalindications (such as focal cartilage defects) were alsoexcluded. We further excluded all duplicate articles, articlesfrom non-peer-reviewed journals, or articles lacking accessto the full text. Conference presentations, narrative reviews,editorials, and expert opinions were also excluded. APRISMA [14] flowchart of the selection and screeningmethod is provided in Figure 1.

All data were extracted from article texts, tables, and fig-ures. Two investigators independently reviewed each article(F.V. and N.D.V.). Discrepancies between the two reviewerswere resolved by discussion and consensus. The final resultswere reviewed by senior investigators (B.D.M. and E.K.). Riskof bias assessment of the included articles was done followingthe Coleman methodology score modified by Kon et al. [15].The assessment was independently performed by 2 authors(F.V. and N.D.V.). Any discrepancy was discussed with andresolved by the senior investigators, who made the finaljudgement.

3. Results

According to the aforementioned inclusion and exclusioncriteria, 23 articles were ultimately included in the presentreview. Relevant data is detailed in Tables 1 and 2. Thirteenof the included articles involved the use of SVF; the otherten involved the use of BMAC.

3.1. BMAC

3.1.1. Application Methods and Quality Assessment of theAvailable Literature. The results of the studies’ quality assess-ment performed with the Coleman methodology score mod-ified by Kon et al. are detailed in Table 3. The average scorewas 37.4 out of 100, thus showing overall poor methodologyin the available literature.

The ten studies investigating BMAC involved 1710 kneesfrom 1386 patients affected by OA. The concentrate wasinjected intra-articularly in 8 studies [16–23] and within the

2 Stem Cells International

tibial and femoral subchondral bones in two other papers[24, 25]. The aspirated bone marrow was centrifuged withoutany other manipulation, expansion, or culture before admin-istering it to the patients.

Regarding the therapeutic protocol, five papers dealt withthe administration of BMAC alone: two of these involvedsubchondral BMAC injections [24, 25], whilst in the remain-ing three, BMAC was injected intra-articularly [16–18]. Ofthese last three papers, one study involved the administrationof 4 sequential injections within a 3-month timespan [17].Other authors administered combined BMAC and PRPwithin the same session [19, 21, 22] or administered PRPalone after a certain period as a booster injection [20].Finally, two authors injected BMAC in association withadipose tissue [22, 23].

In regard to the clinical outcome, the majority of authorsemployed reliable clinical scores (such as those derived fromWOMAC, IKDC, KOOS, or KSS), whereas only a fewauthors used less known questionnaires, improvement ratingscores, or patient satisfaction [17, 20–22]. All studies assessedpain through a visual or numerical score such as VAS orNPS, except for two authors [18, 25] who included WOMACand KSS questionnaires, which have sections on patient pain.Finally, three authors performed MRI prior to and followingthe procedure [19, 24, 25].

Concerning the study design of the 10 included articles,only 2 were randomized controlled trials (RCTs) [19, 25], 1was a prospective pilot trial [24], and the rest were retrospec-tive. Of the latter group, two were comparative studies: one

compared two groups of patients receiving BMAC at differ-ent cell concentrations [21], whilst the other compared agroup treated with BMAC+PRP to a group treated withBMAC+PRP+adipose tissue [22]. Although two RCTs wereidentified, in both cases, the control group was the contralat-eral knee, which introduces into data gathering and interpre-tation [19, 25].

3.1.2. Clinical Findings. The main finding of the reviewedpapers was a significant improvement in pain and functionin almost all cohorts. However, no superiority has been dem-onstrated over standard treatments such as viscosupplemen-tation or corticosteroids. Shapiro et al. [19] performed theonly placebo-controlled RCT available, and they found nostatistically significant differences between BMAC and salineinjections. Moreover, studies on MRI reported conflictingoutcomes. Whilst Shapiro et al. [19] found no evidence ofcartilage regeneration on MRI scans, in their pilot trial, Vadet al. [24] detected an increase in extracellular matrix thick-ness by an average of 14%, especially in younger patients.MRI outcomes were dramatically better in osteonecroticpatients where bone and cartilage repair was observed, witha reduction in bone marrow lesion size by 40% [25]. Never-theless, it seems that a higher BMAC cell concentration isassociated with a significantly better outcome [21]. Further-more, Shaw et al. suggested that multiple injections couldbe more effective than a single one since each subsequenttreatment provided additional benefit to the patients [17].On the other hand, the administration of adipose tissue in

Additional recordsidentified through Google

Scholar (n = 203)

Record identifiedthrough PubMed

searching (n = 224)

Records remaining afterremoval of duplicates

(n = 305)

Records screened (n = 305)

Studies included inqualitative synthesis (n = 23)

Full-text articles assessedfor eligibility (n = 24)

Records excluded byreading the title and abstract

(n = 281)

Additional records identified through Science

Direct (n = 176)Id

entif

iicat

ion

Scre

enin

gEl

igib

ility

Incl

uded

Records excluded due to theunavailability of full text

(n = 1)

Figure 1: PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analysis) flowchart of the systematic literature review.

3Stem Cells International

Table1:Clin

icalstud

iesregardingtheuseof

bone

marrowaspirateconcentrate(BMAC)in

thetreatm

entof

knee

osteoarthritis.

Pub

lication

Stud

ydesign

Disease

Therapeuticprotocol

Outcome

Patient

characteristic

F-up

Mainfind

ings

Hernigou

etal.[25]

RCT(TKAon

contralateral

knee)

BilateralO

Asecond

aryto

severe

ON

relatedto

corticosteroids

BMACgraftpercutaneously

injected

tothesubcho

ndrium

ofthefemur

andtibiavs.

TKAon

contralateralk

nee

MRI,radiograph

s,bone

marrow

lesion

volume,

Kneesocietyscore

30(30BMAC,

30TKA)

Age:18-41

Sex:M-F:12-18

K-L:IV

8-16

years

(mean:

12)

Decreasein

ON

size

by40%.

Cartilage

andbone

repairobserved.

Outcomewas

notstatistically

significantlydifferentbetween

BMACandTKA.T

hemajority

ofpatientspreferredBMAC.

Themistocleous

etal.[16]

Retrospective

OA

BMACinjectionalon

eNPSandOKS

121

Age:70(50-85)

Sex:M-F:36-85

K-L:III-IV

11mon

ths

(range

6-30)

Significant

improvem

entof

both

knee

pain

andfunction

.

Shaw

etal.[17]

Retrospective

OA

4sequ

entialBMAC

injections

in3mon

ths

Resting/active

NPS,overall

percentage

improvem

ent

andLE

FS

15(20kn

ees)

Age:67.7(7.9)

Sex:M-F:5-10

K-L:N

/A

24days

from

last

injection

Significant

improvem

entof

both

knee

pain

andfunction

.The

addition

albenefitwitheach

subsequent

treatm

entmay

suggest

that

multipleinjections

aremore

effective

than

asingleon

e.

Rod

riguez-Fon

tan

etal.[18]

Retrospective

OA

12mlB

MAC

injectionalon

eWOMAC

Satisfaction

rate

19:10kn

ees

and15

hips

Age:m

ean58

(30-80)

Sex:M-F: 3-16

K-L:I-II

6-24

mon

ths

Significantlyim

proved

WOMAC

score;no

significant

difference

betweensix-mon

thandlatest

follow-upscores.V

ariable

satisfaction

rate

(63.2%

yes,36.8%

no).

Shapiroetal.[19]

Single-blin

dRCT(placebo

oncontralateral

knee)

BilateralO

ABMAC+platelet-poor

plasma(PRP)vs.

salin

einjection

VAS,ICOAP,

andalgometer

25(25vs.

25kn

ees)

Age:m

edian

60(42-68)

Sex:M-F:7-18

K-L:I-III

12mon

ths

Significant

improvem

entin

pain

and

qualityof

life.Nosuperiorityto

salin

einjection.

Noevidence

for

cartilage

regeneration

onMRI(T2mapping).

Sampson

etal.[20]

Retrospective

OA

BMACinjectionfollowed

byaPRPboosterinjectionat

approxim

ately8weeks

VASandglobal

patientsatisfaction

73(100

knees)

Age:range

23-79

Sex:N/A

K-L:III-IV

5mon

ths

Significant

improvem

entof

knee

pain.H

ighlevelo

fpatientsatisfaction

.

Vad

etal.[24]

Pilo

ttrial

OA

Injectionof

tibialBMACto

thefemoralandtibial

chon

dral-bon

einterface

andintra-articularkn

eejointspaceviathePeC

aBoo

deliverysystem

MRI,WOMAC,

participant-repo

rted

numericpain

rating

scale

10Age:63.5(52-73)

Sex:M-L:4-6

K-L:III-IV

13-15mon

ths

(mean:

14)

Significant

improvem

entin

WOMAC

andNRSscores.M

RIdisplayedan

increase

inextracellularmatrix

thickn

essby

anaverageof

14%.

Improvem

entsweremore

substantialfor

patientsyoun

ger

than

63.5yearsold.

4 Stem Cells International

Table1:Con

tinu

ed.

Pub

lication

Stud

ydesign

Disease

Therapeuticprotocol

Outcome

Patient

characteristic

F-up

Mainfind

ings

Centeno

etal.[21]

Com

parative

retrospective

(registrydata)

Group

Avs.B

OA

(A)<4

×10

8cells

BMAC+PRP+PL

(B)>4

×10

8cells

BMAC+PRP+PL

NPS,LE

FS,IKDC,

improvem

ent

rating

score

373(424

knees):

(224

vs.185)

Age:54.5vs.50.2

Sex:M-F:143-81/

140-45

K-L:I-IV

3-15

mon

ths

Significant

improvem

entof

both

knee

pain

andfunction

.Significantly

higher

pain

redu

ctionin

patients

treatedwithBMACwithhigh

cellcontent.

Centeno

etal.[22]

Com

parative

retrospective

(registrydata)

Group

Avs.B

OA

(A)BMAC+PRPvs.

(B)BMAC+PRP+

adiposetissue

NPS,LE

FS,

improvem

ent

rating

score

681(840

knees):

(616

vs.224)

Age:54.3vs.59.9

Sex:M-F:

397:219/119

:105

K-L:I-IV

6-10

mon

ths

Significant

improvem

entof

both

knee

pain

andfunction

.Nodetectible

benefitwiththeaddition

ofan

adiposegraftto

theBMAC.

Kim

etal.[23]

Retrospective

OA

BMAC+adiposetissue

inj.+

arthroscop

icdebridem

ent(6),

microfractures(5),

andHTO

(1)

VAS,IKDC,SF-36,

KOOS,Lysholm

41(75kn

ees)

Age:60.7(53–80)

Sex:M-F:17-24

K-L:I-IV

8.7mon

ths

Significant

improvem

entof

both

knee

pain

andfunction

.Better

outcom

esin

earlyto

mod

erateph

ases.

5Stem Cells International

Table2:Clin

icalstud

iesregardingtheuseof

stromalvascular

fraction

(SVF)

inthetreatm

entof

knee

osteoarthritis.

Pub

lication

Stud

ydesign

Disease

Therapeuticprotocol

Outcome

Patient

characteristic

F-up

Mainfind

ings

Jonesetal.[28]

RCT

(NCT03242707)

OA

Com

parative

stud

y:ultrasou

nd-guided,

intra-articularinjection

ofautologous

adipose

tissue

vs.H

A

WOMAC,P

ROMIS

question

naire,

syno

vialfluid

analysis,sway

velocity

assessment

54(27vs.27)

Age:N

/ASex:M-F:

N/A

K-L:N

/A

6mon

ths

Ongoing.

Roato

etal.[29]

Prospective

OA

Followingdiagno

stic

arthroscop

y,35

ml

ofconcentrated

adiposetissue

was

injected

intra-articularly

WOMAC,V

AS,MRI,

immun

ohistochem

istry

of2kn

ees

20Age:59.6

Sex:M-F:

9-11

K-L:I-III

18mon

ths

Whilstboth

WOMACandVASscores

improved

significantly,W

OMAC

scores

show

edprogressivelybetter

outcom

es.M

RIOuterbridge

gradedid

notshow

significant

changes.

Immun

ohistochem

istrydisplayed

newtissue

grow

th.

Hon

getal.[33]

Dou

ble-blind

RCT(H

Ain

contralateral

knee)

Bilateral

OA

Com

parative

stud

y:arthroscop

icdebridem

ent

followed

byintra-articular

SVFinjectionvs.H

Ainjectionin

the

contralateralk

nee

VAS,WOMAC,R

OM,

who

le-organ

MRI

score,MRI

observation

ofcartilage

repairtissue

16(32kn

ees):

(16vs.16)

Age:18-70

years

Sex:M-F:

K-L:II-III

12mon

ths

VASandWOMACscores

and

knee

ROM

improved

significantly

forboth

grou

ps,but

these

improvem

entswereno

tlong

lastingin

thecontrolgroup

.MRI

analysisshow

edsignificantly

increasedcartilage

repairin

theSV

Fgrou

pcomparedto

thecontrol.

Hud

etzetal.[26]

Prospective

OA

Microfragmented

adiposetissue

injection

VAS,radiograph

s,dG

EMRIC

MRI,IgG

isolationfrom

plasma

andsyno

vialfluid

17(32kn

ees)

Age:40-85

Sex:M-F:

12-5

K-L:III-IV

12mon

ths

Significant

decrease

inVASscores.N

ochange

inIgGglycom

ecompo

sition

.dG

EMRIC

MRIanalysisdisplayed

increase

inproteoglycan

content

withintheECM.

Bansaletal.[30]

Prospective

(phase

I)NCT03089762

OA

SVF+

PRPinjection

WOMAC,6-m

inute

walking

distance,M

RI

10(13kn

ees)

Age:≥

50Sex:N/A

K-L:I-II

24mon

ths

Significant

improvem

entof

WOMAC

scores

and6-minutewalking

distance.

MRIshow

edincrease

incartilage

thickn

essin

allb

ut2patients.A

llpatientsaresatisfied

withtherapy.

Yokotaetal.[27]

Prospective

OA

Intra-articular

injectionof

SVF

VAS,WOMAC,

Japanese

Knee

Osteoarthritis

Measure

(JKOM)

13(26kn

ees)

Age:74.5

Sex:M-F:

2-11

K-L:III-IV

6mon

ths

AllVAS,WOMAC,and

JKOM

scores

improved

significantlyat

the6-mon

th(last)follow-up.

Nguyenetal.[37]

Com

parative

prospective

OA

Com

parative

stud

y:arthroscop

icmicrofracture

(AM)andSV

F+PRP

injectionvs.A

Malon

e

WOMAC,V

AS,and

Lysholm

scores,M

RI,

knee

jointfunction

30(15vs.15)

Age:58.60

vs.58.20

Sex:M-F:

3-12

vs.3-12

K-L:II-III

18mon

ths

WOMAC,L

ysho

lm,and

VASscores

improved

forboth

grou

psup

to12

mon

ths,bu

tat

18mon

ths,theSV

Fgrou

pwas

significantlybetter

than

the

controlgroup

.At12

mon

ths,theSV

Fgrou

pdisplayedsignificantlyless

bone

marrowedem

athan

thecontrolgroup

.

6 Stem Cells International

Table2:Con

tinu

ed.

Pub

lication

Stud

ydesign

Disease

Therapeuticprotocol

Outcome

Patient

characteristic

F-up

Mainfind

ings

Koh

etal.[36]

Prospective

OA

Followingarthroscop

iclavage

isintra-articular

injectionof

SVF+

PRP

tothemostsevere

cartilagino

usdefects

Lysholm,V

AS,and

KOOSscores,

radiograph

s,2n

d-look

arthroscop

y

30Age:≥

60Sex:M-F:

5-25

K-L:II-III

24mon

ths

Lysholm,V

AS,andKOOSscores

all

improved

significantly.Scores

increasedat

thesecond

year

compared

tothefirstyear

offollow-up.

Second

-look

arthroscop

ydeterm

ined

the

majorityof

kneesas

positive

orbetter.

Kim

etal.[34]

Prospective

comparative

OA

Com

parative

stud

y:followingarthroscop

icdebridem

ent,grou

p1

received

anintra-articular

injectionof

SVF;

grou

p2

received

anintra-articular

injectionof

SVF+

fibrin

glue

asascaffold

IKDC,T

egner,

2nd-look

arthroscop

icICRSgrading

54(56kn

ees):

37vs.17

Age:57.5

vs.57.7

Sex:M-F:

14-23

vs.8-9

K-L:I-II

29.2vs.

27.3mon

ths

IKDCandTegneractivity

scores

significantlyim

proved

inboth

grou

psbu

tshow

edno

significant

difference.

Statisticalsignificancebetweenthetwo

grou

pswas

observed

inICRSgrades,

withtheSV

Fgrou

pbeingmore

positive.A

higher

BMIresulted

inlesspo

sitive

outcom

es.

Koh

etal.[32]

Prospective

comparative

OA

Com

parative

stud

y:intra-articular

injectionof

SVF+

PRP

vs.onlyPRPpriorto

performingop

en-w

edge

high

tibialosteotom

y

Lysholm,K

OOS,

VAS,and

femorotibialangle.

Arthroscopic

evaluation

44(23vs.21)

Age:52.3

vs.54.2

Sex:M-F:

6-17

vs.5-16

K-L:I-III

24-25mon

ths

(mean:

24.4)

Lysholm,V

ASscore,andKOOS

improved

statistically

inboth

grou

ps.

KOOSim

provem

entswerestatistically

greaterin

theSV

Fgrou

p.Nodifference

inthepreoperative

andpo

stop

erative

femorotibialangles.SV

Fgrou

pdisplayedgreaterfibrocartilage

coverage.

Koh

etal.[35]

Retrospective

OA

Arthroscopic

debridem

ent+

administration

ofSV

Fto

articular

chon

drallesion

s

IKDCscoreand

Tegneractivity

scale,2n

d-look

arthroscop

icICRSgrading

35(37kn

ees)

Age:48-69

Sex:M-F:

14-21

K-L:I-II

24-34mon

ths

(mean:

26.5)

IKDCandTegneractivity

scores

significantlyim

proved.P

atients

repo

rted

high

satisfaction

scores.It

was

notedthat

ahigher

BMIresulted

inlesspo

sitive

outcom

es.

Bui

etal.[31]

Prospective

OA

Intra-articular

SVF+

PRPinjection

VASandLysholm

scores,M

RI

21Age:≤

18Sex:N/A

K-L:II-II

6mon

ths

Statistically

significant

improvem

ent

inVASandLysholm

scores.M

RI

analysisshow

edpartialregeneration

andthickening

ofarticularcartilage.

Koh

etal.[38]

Retrospective

OA

Infrapatellar

SVF+

PRP,

intra-articular

injection+

weekly

PRPinjection

for2weeks

Who

le-organ

MRI,

WOMAC,V

AS,and

Lysholm

scores

18Age:41-69

Sex:M-F:

6-12

K-L:III-IV

24-26mon

ths

(mean:

24.3)

Significant

decrease

ofWOMAC,V

AS,

andLysholm

scores.Significant

decrease

ofwho

le-organ

MRIscores.

Extentof

improvem

entwas

directly

correlated

withtheam

ount

ofMSC

sinjected.

NPS:nu

mericalpain

scale;OKS:OxfordKneeScore;LE

FS:Low

erExtremityFu

nction

alityScore;VAS:visualanalog

scale;OARSI:O

steoarthritisResearchSocietyInternational;ICOAP:Intermittent

andCon

stant

OsteoarthritisPain;

WOMAC:Western

Ontario

andMcM

asterUniversitiesArthritis

Index;

TKA:totalkn

eearthroplasty;IKDC:InternationalKneeDocum

entation

Com

mittee;

KOOS:

Kneeinjury

and

OsteoarthritisOutcomeScore;

MACI:Matrix-indu

cedAutologou

sCho

ndrocyte

Implantation

;ROM:rangeof

motion;

ICRS:

InternationalCartilage

Regeneration&

JointPreservationSociety;

PeC

aBoo:

percutaneous

cartilage-bon

einterfaceop

timizationsystem

;dGEMRIC:d

elayed

gado

linium-enh

ancedmagneticresonanceim

agingof

cartilage.

7Stem Cells International

Table3:BMACstud

yqu

alityassessmentwiththeColem

anmetho

dology

scoremod

ified

byKon

etal.

Stud

yTOT

Stud

ysize

Mean

f-up

Different

surg

proc

Typeof

stud

ySurg

proc

description

Postop

rehab

MRI

outcom

eHistological

outcom

eOutcome

criteria

Outcome

assessment

Selection

process

Bon

emarrowaspirate

concentrate

Hernigouetal.

[25]

707

1010

155

510

05

30

Themistocleous

etal.[16]

4410

010

05

50

05

36

Shaw

etal.[17]

230

010

05

00

05

30

Rod

riguez-Fon

tan

etal.[18]

300

210

05

20

05

33

Shapiroetal.[19]

517

24

155

010

02

33

Sampson

etal.

[20]

2610

04

05

20

02

03

Vad

etal.[24]

490

210

105

010

05

70

Centeno

etal.[21]

2610

04

05

20

02

30

Centeno

etal.[22]

2310

04

05

20

02

00

Kim

etal.[23]

3210

04

05

50

05

30

8 Stem Cells International

combination with BMAC showed no benefit in comparisonto BMAC without adipose tissue [22].

3.2. SVF

3.2.1. Application Methods and Quality Assessment of theAvailable Literature. The mean Coleman methodology score(modified by Kon et al.) of the 13 studies was 47 out of 100(Table 4), slightly better than that of BMAC studies but stillinsufficient to define the available evidence as “methodologi-cally” robust.

Several different therapeutic approaches were employedin the studies regarding SVF. Only 4 studies [26–29] includedin this review analysed the results of knee intra-articular SVFconcentrate injections with no other additional treatments.SVF was administered intra-articularly in another 3 studies[30–32], but this was injected in combination with platelet-rich plasma (PRP). The majority of the authors, however,injected SVF following an arthroscopic surgical procedure,with 3 of them administering SVF after arthroscopic debride-ment or lavage [33–35]. Conversely, Koh et al. [36] per-formed arthroscopic lavage prior to an injection of SVF andPRP combined. Nguyen et al. [37] injected SVF suspendedin PRP after arthroscopic microfracture of the osteoarthriticbone. Only one study [32] reported the use of arthrotomicsurgery in the therapeutic protocol, with SVF+PRP beinginjected after performing open-wedge high tibial osteotomy.Regarding clinical outcomes, only 1 [26] of the 13 papersconcerning SVF did not include at least one knee-specificpatient-reported score as calculated through validated ques-tionnaires, namely, WOMAC, IKDC, KOOS, Lysholm score,or Tegner activity scale. None of these scores were employedin the study performed by Hudetz et al. [26], which was con-versely the only study in which laboratory measurementswere taken into consideration. The authors examined theeffects of autologous microfragmented fat tissue injectionon the synthesis of proteoglycans within the synovial fluid.Immunohistochemical analysis was only reported in Roatoet al.’s study [29]. In almost all of the studies, the VAS scalewas used to quantify osteoarthritic pain relief followingSVF administration. Changes in MRI images following thetherapeutic procedure were taken into account in 7 studies[26, 29–31, 33, 37, 38]. Second-look arthroscopy of theaffected joint was instead performed in 4 different papers[34–36]. Our review of the literature revealed a paucity ofrandomized controlled trials concerning the clinical applica-tion of SVF in the treatment of OA, with only 2 trials out of13 being RCTs [28, 33]. Nine papers were prospective clinicalstudies, with 3 of them being comparative [34, 35, 37]. Therest were retrospective studies.

3.2.2. Clinical Findings. The review of the literaturehighlighted some important concepts about SVF and its clin-ical application. Firstly, regarding its safety in the treatmentof OA, no serious adverse events have been described inany of the 13 articles dealing with this topic, with only mildswelling and pain being reported in very few patients withinthe first few days following the therapeutic procedure. Inregard to donor site morbidity, liposuction was very well tol-

erated by patients mainly due to the limited amount of tissueharvested. Secondly, SVF administration has shown positiveclinical outcomes in the studies reviewed. A significantimprovement in terms of range of movement, pain, and artic-ular function during daily activities (measured via varied andcommonly used validated questionnaires such as WOMAC,IKDC, KOOS, Lysholm score, Tegner activity scale, andVAS pain scale) was reported in all 13 studies at the lastfollow-up. Thirdly, regarding MRI, most of the papers [26,30, 31, 33, 38] (including the RCT by Hong), described pos-itive changes in the imaging outcomes, reporting signs ofpartial regeneration of the articular cartilage. Similarly,Nguyen et al. [37] described significantly less bone marrowedema in patients receiving PRP+SVF treatment followingmicrofracture compared to the control group (microfractureonly). Nevertheless, Roato et al. [29] did not report a signifi-cant difference between MRI scores before SVF treatmentcompared to MRI scores at 18 months, as measured via theuse of Outerbridge scores, a quantitative parameter (rangingfrom 1 to 4) determining the OA grade of the affected knee.The same study also took into consideration the histologicaloutcome of two patients who left the study after 12 and 14months, respectively, to undergo knee arthroplasty. Kneebiopsies were performed and compared to those collectedfrom OA patients (matched for age, sex, and OA grading)who underwent arthroplasty alone. In both the joints previ-ously treated with SVF, the authors detected the presence ofnew tissue formation starting from the subchondral end ofthe osteochondral lesion, whereas this type of neotissue for-mation was undetectable in those that did not receive anSVF injection.

These encouraging results concerning nonexpanded adi-pose tissue injections are however weakened by the presenceof several biases in the studies’ therapeutic protocols. In fact,in only 4 out of 13 studies [26–29] was adipose tissue solelyadministered. In the remaining papers, concomitant surgeryor administration of other injective therapies (viscosupple-mentation, PRP) was performed, obviously confusing theobtained outcomes and making them difficult to compare.

4. Discussion

The main finding of the present systematic review is that theavailable literature concerning the use of BMAC and SVF forknee OA is characterized by a lack of sound methodologies,as represented by the Coleman methodology scores, with apaucity of RCTs being presently published, thus preventingus from making solid conclusions on the real therapeuticpotential of these novel methods compared with others.

Currently, there are several clear issues in the scientificliterature. Although clinical results seem positive, whenassessing the risk of bias, it becomes apparent that most ofthe studies present are of low quality and lack relevant meth-odologies, as detailed in Table 3. Average Coleman scores(modified by Kon et al.) were exceedingly poor mainly dueto the mean follow-up being short and half of the includedstudies being retrospective, with only four papers being RCTs[19, 25, 28, 33] and eight being prospective studies. Further-more, in many papers, the inadequate patient selection

9Stem Cells International

Table4:SV

Fstud

yqu

alityassessmentwiththeColem

anmetho

dology

scoremod

ified

byKon

etal.

Stud

yTOT

Stud

ysize

Mean

f-up

Different

surg

proc

Typeof

stud

ySurg

proc

description

Postop

rehab

MRI

outcom

eHistological

outcom

eOutcome

criteria

Assessm

entof

clinicalou

tcom

eSelection

process

Adipo

se-derived

stem

cells

Jonesetal.[28]

547

010

155

00

05

93

Roato

etal.[29]

670

210

105

510

105

73

Hon

getal.[33]

564

24

155

510

05

33

Hud

etzetal.

[26]

534

210

105

010

05

70

Bansaletal.

[30]

450

54

105

010

05

33

Yokotaetal.

[27]

340

010

105

20

02

50

Nguyenetal.

[37]

514

24

105

510

05

33

Koh

etal.[36]

364

54

105

00

05

30

Kim

etal.[34]

477

54

105

50

05

33

Koh

etal.[32]

457

54

105

00

05

36

Koh

etal.[35]

4010

54

05

50

05

33

Pham

2014

Biomed

Res

444

04

105

010

05

33

Koh

etal.[38]

404

54

05

210

05

23

10 Stem Cells International

process introduced another source of bias, since inclusionand exclusion criteria were seldom reported, the recruitmentrate was not stated, and a flowchart of the selection processwas often unavailable. Although all the authors adequatelydescribed the procedure, in most of the studies, manypatients underwent concomitant surgery, such as arthro-scopic debridement, microfracture, or high tibial osteotomy,thus preventing a clear understanding of the real contribu-tion and clinical potential of these stem cell-based products.Furthermore, whilst outcome criteria were always clearlydefined and mostly demonstrated good reliability and sensi-tivity, the procedures for assessing the clinical outcome werenot fully elucidated. In addition, MRI outcomes were onlyreported in 11 studies (10/23 43,5%).

The overall lack of sounding methodologies, along withthe inherent paucity of RCTs available, is not an uncommonobservation in the field of biologic therapies for OA. If welook at the recent past, in many cases, biotechnologies havebeen released onto the market without enough proof of evi-dence: the paradigmatic example is platelet-rich plasma(PRP), which “invaded” clinical practice on the basis of hugemedia exposure rather than solid scientific evidence [39].One reason for this lies within the possibility of taking regu-latory shortcuts such as the 510(k) exemption [40], based onthe fact that new medical devices “substantially equivalent”to others already present on the market can skip the standardFDA approval process. This led to a proliferation of commer-cially available kits for PRP production, and in a few years,the market was saturated by many different preparation sys-tems giving substantially different outputs in terms of biolog-ical features, thus inhibiting a “standardization” of PRPtherapy for the treatment of OA or other musculoskeletalconditions. The same risk is now impending on stem cell-based technologies, especially on the so-called “minimallymanipulated” products, which are not affected by the regula-tory burden currently imposed on products requiring cellexpansion in lab. Consequently, BMAC and SVF haveemerged as the easiest way to employ MSCs in the clinicalpractice, since these products can be rapidly harvesteddirectly within the OR and be immediately administered topatients in disparate ways, such as simple intra-articularinjections (sometimes in association with other carriers likePRP or hyaluronic acid), as intraosseous injection at thebone-cartilage interface, as augmentation for various scaf-folds, and as a topical application on focal defects [25, 34,38]. Whilst the wide range of potential applications is fasci-nating, it is also responsible for the current lack of clear indi-cations on the best method of therapeutic administration.The concurrent use of other biological agents or biomaterialsor the administration of these cellular products following“conventional” surgical procedures introduces additionalconfounding elements, thus preventing a fair comparison ofthe studies performed so far. Even the few RCTs publishedpresent some relevant biases, mainly due to the fact that inmost of them patients were treated bilaterally [19, 25, 33],which is not the ideal condition to assess the efficacy of atreatment considering that patients cannot evaluate one kneeindependently from the other. This results in a bias that isvery difficult to account for even when using dedicated statis-

tical tests [41]. Basic questions such as the number of cellsadministered, the optimal number of injections to achievethe best therapeutic effect, and the superiority of one prepa-ration method over another still remain unanswered. Despitethe attempts presently published, the literature is still lackingwell-designed trials to address these issues. Similarly, it hasbeen impossible to establish which, or whether, one of thetwo sources, either bone marrow or adipose tissue, providesbetter results. Whilst it has been shown that the immunophe-notypes of BMSCs and ASCs are more than 90% identical[42, 43], they still exhibit a number of distinct characteristics,for example, in their cell surface markers, differentiationpotentials, and in their distribution within the body. In vitroanalysis revealed that the great advantage of SVF is theirabundance: when compared to 100ml of bone marrow aspi-rate, up to 300-fold more stem cells can be obtained from100 g of adipose tissue [44, 45]. Regardless, since a dose-effect correlation has not been clearly demonstrated, thisadvantage still remains theoretical. Lastly, the issue of inter-human variability, which plays a central role in the successof these biological therapies (since a particular “patient’s pro-file” could respond better to a specific biologic stimulus com-pared to another), has not yet been faced. This exacerbatesthe need for more research, dedicated to understanding theunique features of a specific stem cell-based product in orderto target the unique features of the recipient joint of a partic-ular patient.

From a “surgical” point of view, bone marrow harvestingand lipoaspiration are both simple procedures with minimalside effects (with the latter being perhaps slightly more seri-ous due to the associated risks of hematoma and pain at thesite of lipoaspiration). Currently, the choice between bonemarrow and fat tissue is purely based on the surgeon’s pref-erence and experience and, keeping commercial issues inmind, on the different availability of preparation kits in dif-ferent countries. However, in recent years, more and moreindustries have been releasing their proprietary kits forBMAC and SVF preparation with new methods still beingdeveloped, such as the use of mechanical microfragmenta-tion of the adipose tissue to obtain an “adipose graft” con-taining intact stromal vascular niches with MSCs and othercells involved in the modulation of joint homeostasis [46].Whilst the increase in the number of preparation kits onthe market has contributed to a partial reduction of costsover time, it has also raised the risk of releasing products withdubious performances and without sufficient scientific datacertifying their capability of concentrating MSCs in a “mini-mal manipulation”-compliant approach. In addition, theoverexposure of biological products for OA has led to theirrapid growth (both within the market of “dedicated devices”and within clinical settings), without the support of robustevidence. Expanding the number of available treatmentoptions for patients affected by knee OA does not alwaysmean improving the standard of care, especially when thereis a lack of comparative trials assessing the effectiveness of anovel treatment compared to established ones.

Presently, stem cell-based products undoubtedly repre-sent an expensive technology, whose costs still cannot be sus-tained by National Health Systems, and (due to the lack of

11Stem Cells International

robust data on their effectiveness) cannot be endorsed as a“routine” treatment for knee cartilage degeneration. From aclinical point of view, despite the aforementioned methodo-logical limitations, the use of BMAC and SVF for the treat-ment of knee OA seems safe and able to provide positiveclinical outcomes, thus potentially offering a new minimallyinvasive therapeutic option for patients who are not eligiblefor more invasive approaches. Nevertheless, their promising,short-term results must be both confirmed in the long-termand compared to those of established treatments. Until then,the use of minimally manipulated MSCs, either BMAC orSVF, represents a therapeutic option that must be carefullyand thoroughly discussed between the physician and thepatient, especially when it is proposed as a first-line therapeu-tic approach to avoid more invasive solutions, rather than a“salvage procedure.”

5. Conclusion

The available literature is undermined by both the lack ofhigh-quality studies and the varied clinical settings and dif-ferent protocols reported in the small number of RCTs pub-lished. This prevents any recommendation on the use ofeither product in the clinical practice. Nevertheless, focusingon clinical results, BMAC and SVF have been shown to besafe and to have some short-term beneficial effect on thetreatment of knee cartilage degeneration. Currently, there isno evidence on the superiority of either bone marrow or adi-pose tissue as a source of minimally manipulated MSCs.

Conflicts of Interest

The authors declare that they have no conflicts of interest.

Acknowledgments

The present paper was funded by the 2015 PRIN (Progetti diRicerca di Rilevante Interesse Nazionale) project titled:“Amniotic epithelial stem cells (AECs) vs. adipose-derivedmesenchymal stem cells (ADSCs): translational potential asbiological injective treatment for osteoarthritis.”

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