June 2021

Medical Mass Spectrometry Vol. 5 No. 1

Research Paper

N-acetyl-seryl-aspartyl-lysyl-proline:
A new potential serum biomarker of rheumatoid arthritis
1,2
1,3
2
1
3
1
Yusuke Oguri , Yohei Kawaguchi *, Naoe Tatematsu , Yuji Joyo , Ken Mizuguchi , Hiroki Yonezu ,
1

1

1

4

1

3

Hideki Okamoto , Masahiro Nozaki , Makoto Kobayashi , Gen Kuroyanagi , Hisaki Aiba , Kiyofumi Asai ,Koichi Inoue , Hideki Murakami , Yuko Waguri-Nagaya
5

1

1,2

1

Department of Orthopaedic Surgery, Nagoya City University Graduate School of Medical Sciences,Mizuho-ku, Nagoya, Aichi 467‒8601, Japan2
Department of Orthopaedic Surgery, Nagoya City East Medical Center, Chikusa-ku, Nagoya, Aichi 464‒8547, Japan3
Department of Glial Cell Biology, Nagoya City University Graduate School of Medical Sciences,Mizuho-ku, Nagoya, Aichi 467‒8601, Japan
4
Department of Rehabilitation Medicine, Nagoya City University Graduate School of Medical Sciences,Mizuho-ku, Nagoya, Aichi 467‒8601, Japan
5
Laboratory of Clinical and Analytical Chemistry, College of Pharmaceutical Sciences, Ritsumeikan University,
Nojihigashi, Kusatsu, Shiga 525‒8577, Japan
Abstract

Background. Angiogenic factors play an important role in the pathogenesis of rheumatoid arthritis (RA)through microvessel proliferation. N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) is a tetrapeptide cleaved from the N-terminal of thymosin β4, known as a stimulator of angiogenesis. The objective of this study was to investigate the expressionof sera and synovial ﬂuid levels of AcSDKP relative to clinical manifestations in patients with RA.
Methods. The concentration of AcSDKP in sera and synovial ﬂuids was measured using a high throughput method basedon on-line solid-phase extraction liquid chromatography tandem mass spectrometry (SPE-LC-MS/MS). Serum or synovial
ﬂuid samples were collected from 52 patients with RA, 22 patients with osteoarthritis (OA), and 5 healthy controls. We
included serum samples from 22 patients with RA treated with disease modifying antirheumatic drugs (DMARDs) at the
beginning and after 12 weeks of treatment.
Results. Median serum levels of AcSDKP were 2.64 (ng/mL), 1.65 (ng/mL) and 0.69 (ng/mL) in patients with RA, patientswith OA and healthy controls, respectively. There was positive correlation between serum AcSDKP and serum CRP in patients with RA. However there were no correlations between serum levels of AcSDKP and serum matrix metalloproteinase(MMP)-3, and ESR. Serum levels of AcSDKP and DAS28 [ESR] reﬂected improvements in the disease activity following
treatment with DMARDs.
Conclusions. This is the ﬁrst report demonstrating that the serum levels of AcSDKP might serve as a useful clinical marker of RA.
Key words: N-acetyl-seryl-aspartyl-lysyl-proline, biomarker, synovial ﬂuid, rheumatoid arthritis, On-line solid-phase extraction
liquid chromatography tandem mass spectrometry* Corresponding author

Introduction

Yohei Kawaguchi
Department of Glial Cell Biology, Nagoya City University
Graduate School of Medical Sciences, Mizuho-ku, Nagoya,
Aichi 467‒8601, Japan
Tel: ＋81‒52‒853‒8200, Fax: ＋81‒52‒842‒3316
E-mail: yohei21diver@yahoo.co.jp
Received: October 29, 2020. Accepted: March 15, 2021.
Epub April 23, 2021.
DOI: 10.24508/mms.2021.06.004

Rheumatoid arthritis (RA) is a progressive, autoimmune
disease, characterized by chronic synovial inﬂammationand consistent degradation of articular cartilage and
1,2)

bone

. Angiogenesis is also known to play a key role in

RA, and thus the ability to induce and sustain angiogenesis
3)
seems to be acquired in prolonged synovitis . The modulation of angiogenesis is suggested to be reasonable strategy
28June 2021

Medical Mass Spectrometry Vol. 5 No. 1

Table 1.

in treatment of RA, because a lot of angiogenic factors such

Characteristics of 52 patients with RA

as vascular endothelial growth factor (VEGF), gliostatin/

Variablethymidine phosphorylase and ﬁbroblast growth factor are
4‒8)overexpression

Demographic variables.

Thymosin β4 (Tβ4) is the major actin-sequestering mole-

Gender (F/M)

42/10

Age (years)

59

cule in all eukaryotic cells, and a potent regulator of actin
9)polymerization in mammals . Recent studies have pro-

RA related variablesvided evidence that Tβ4 might have a role in the pathogene-

CRP (mg/dL)

Disease duration (years)

sis of RA. The level of Tβ4 has been shown to signiﬁcantly
increase in the synovial ﬂuid (SF) and serum of patients
10,11).
with RAN-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) is a tetra12)
peptide cleaved from the N-terminal of Tβ4 . This tetra-

peptide is involved angiogenesis beside the regulation of
the immune response and has been recognized to be an(25‒83)10.5 (0.5‒49)
0.7 (0.03‒13.6)

ESR (mm/h)

30.5 (2‒108)

MMP-3 (ng/mL)

84.2 (10‒1752)

RF or ACPA positive/negative

37/15

DAS 28 [ESR]

3.72 (1.11‒6.66)

Current MTX usage (number, %)44(84.6)

Current GC usage (number, %)12(23.0)

Current biologic therapy (number, %)11(21.1)

Values are median (range) or numbers (%).
CRP: C-reactive protein; ESR: erythrocyte sedimentation rate;
RF: rheumatoid factor; ACPA: anticitrullinated protein antibodies;
DAS: disease activity score; MTX: methotrexate; GC: glucocorticoid.

angiogenic activity. It inhibits the proliferation of humanhematopoietic progenitors, and it has been reported as an
13,14)

angiogenic activity

Results.

We hypothesized that AcSDKP may play an important,pathogenic role in the proinﬂammatory cascade during the

16)

and

course of RA. The aim of this study was to investigate thepatients was evaluated with the DAS28 [ESR] score

distribution of AcSDKP between the serum and SF inpatients with RA and osteoarthritis (OA), and the serum of

was calculated at the time of blood collection during theclinical appointment. Anti-rheumatic therapy and any other

healthy controls. Our present study identiﬁed that serum

concomitant treatments were recorded at the same time.

AcSDKP could serve as a useful biomarker of RA.

The characteristics of the 52 patients with RA are presented
in Table 1. Most patients were positive for RF and ACPA

Materials and Methods

and had an erosive form of RA. Remission of RA according

Patients and controls

to DAS28 (＜2.6) was observed in 10 patients (8 females

52 patients who met the 2010 American College of Rheu-

and 2 males). At the time of examination, all patients werematology/European League Against Rheumatism (ACR/
were recruitedtreated with at least one conventional synthetic disease
modifying anti-rheumatic drugs (csDMARDs): methotrex-

from Nagoya City University Hospital for this study. Theseate (MTX) (44 patients), sulfasalazine (4 patients), and tac-patients had not any evidence of infectious diseases in thelast month prior to data collection and any history of malig-rolimus (15 patients). Biological DMARDs (bDMARDs)
were used in 11 patients (21.1%) (adalimumab in 4, etaner-

nancy and secondary Sjögren s syndrome. No cases had
been taking angiotensin converting enzyme (ACE) inhibi-

cept in 3, and tocilizumab in 4 cases, respectively). Concomitantly, low-dose prednisone ( ≤ 10 mg/d) was used intors. Demographical (age and gender) and disease-related

12 patients (23.0%).

15)

EULAR) criteria for the diagnosis of RA

parameters of all patients were recorded at the time ofblood sampling. With respect to disease speciﬁc parame-

Consecutive pairs of sera were collected at the time ofchange DMARDs and after 12 weeks in 22 patients with
RA who did not respond to previous treatments. The clini-ters, disease duration, tender joint count, swollen joint
count, and a patient global assessment were evaluated. In
addition, we measured the serum levels of C-reactive pro-cal details of these patients are shown in Table 2. Serumtein (CRP) and matrix metalloproteinase-3 (MMP-3), thesamples were obtained from 22 patients with OA (18females and 4 males) who met the ACR criteria for the clas-

erythrocyte sedimentation rate (ESR), and the presence ofthe rheumatoid factor (RF) as well as anti-cyclic citrulli-siﬁcation of OA . The median age of the patients was 73.5
years, with an age range of 50‒82 years. Five normal serumnated peptide antibody (ACPA). Disease activity of RA

samples were obtained from healthy individuals who vis-

17)

29

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Table 2.

Medical Mass Spectrometry Vol. 5 No. 1

Characteristics of 22 patients with RA at thebaseline

Statistical analysis

Laboratory variable

lysed using GraphPad Prism 7 (GraphPad Software, Inc.,

No. of patients (F/M)

All data were entered into an electronic database and ana-

San Diego, CA, USA). All statistical tests were performed
at a signiﬁcance level of α＝0.05, that is, all p-values ≤ 0.05

22

(16/6)
58.5 (26‒81)
9.1 (0.1‒40.0)

Age (years)
Disease duration (years)

were considered signiﬁcant. All conﬁdence intervals were2.65 (0.03‒13.57)(2‒88)CRP (mg/dL)
ESR (mm/h)
MMP-3 (ng/mL)

computed using a conﬁdence level of 95%. Comparisons of42

patient groups were done with analysis of the variance(ANOVA) and Bonferroni test if the distribution was nor-

289 (18.9‒934.5)
4.22 (2.32‒6.66)

DAS28 [ESR]
Current MTX usage (number, %)14(63.6)

Current GC usage (number, %)6(27.2)

Current biologic therapy (number, %)14(63.6)

mal, if not with the Kruskal‒Wallis with Dunn s post-hoc
test was used to determine diﬀerences across groups for
AcSDKP. All data were presented as median and interquartile (IQR). Pearson correlation test was used to determine
the association between log-transformed AcSDKP and clini-

Values are median (range) or numbers (%).
CRP: C-reactive protein; ESR: erythrocyte sedimentation rate;
RF: rheumatoid factor; ACPA: anticitrullinated protein antibodies;
DAS: disease activity score; MTX: methotrexate; GC: glucocorticoid.

cal and laboratory variables. The statistical signiﬁcance ofthe diﬀerences in the levels of the indicated parameters
between baseline and the indicated treatment time points
was examined using the Wilcoxon signed rank test.

ited our hospital for routine health tests and were without
prior history of chronic inﬂammation or any form of arthri-

Results

Calibration curve and multiple reaction-monitoring(MRM) chromatogram of Ac-SDKP and internal
standard in human plasma for recovery test

tis, as evidenced by normal levels of CRP and ESR.
SF samples were obtained from 10 patients with RA (9females and 1 male) and OA (7 females and 3 males) priorStandard solutions of Ac-SDKP were prepared in pure13
water and added to a ﬁxed concentration of Ac-SDKP- C6,

to total knee arthroplasty, respectively.
This study was approved by the Nagoya City University

N2 (1μg/mL) for a calibration curve covering the concentration range from 0.1 to 75 ng/mL for human plasma sam-

15

Ethics Committee (# 671), and informed consents wereobtained from all patients.

ples (Fig. 1a). We proposed the method to evaluate endogenous Ac-SDKP levels in human plasma samples.

On-line solid-phase extraction liquid chromatography
tandem mass spectrometry (SPE-LC-MS/MS) analysis

Quantitative analysis was performed using multiple reaction-monitoring (MRM) mode in order to maximize sensi-of AcSDKP

tivity of quantitative ion, and ratio of sample/internal stan-

Synthetic AcSDKP was obtained from Peptide Institute
13
15
Inc. (Osaka, Japan). The AcSDKP- C6, N2 was synthe-

dard (Fig. 1b).

sized based on the previous report . AcSDKP was measured using an SPE-LC-MS/MS system as previously
19)
described . The stock solutions (0.1 mg/mL) for Ac-SDKP
18)

Serum levels of AcSDKP in healthy controls and inpatients with RA and OA

13
15
and Ac-SDKP- C6, N2 were prepared by dissolving the

We compared the levels of AcSDKP in the serum of

appropriate amount of standard in pure water. Calibration

patient groups and healthy controls. We found that instandards were prepared by diluting an aliquot of the stock

healthy controls, AcSDKP was present only at low levels (asolution in pure water.

median level of 0.69 ng/mL) in the circulation. Patients

The lower limit of quantiﬁcation (LLOQ) in humanwith OA had a median serum level of AcSDKP of 1.65 ng/

plasma sample was detected as the concentration of thelowest calibration by on-line SPE-LC-MS/MS. The LLOQ

mL, whereas patients with RA had a median level of

was 0.1 ng/mL based on signal per noise (plasma control

were signiﬁcantly increased in the RA patients group when

2.64 ng/mL, respectively. The Serum levels of AcSDKP

18)

sample) of 10 .

compared to either healthy control or OA group (p＜0.05;
Fig. 2). There was no statistically signiﬁcant diﬀerence in30

June 2021

Fig. 1.

Medical Mass Spectrometry Vol. 5 No. 1(a) Calibration standards were prepared by diluting an aliquot of the stock solution in pure water. Standard solutions of Ac-SDKP were13
15
prepared in pure water and added to a ﬁxed concentration of Ac-SDKP- C6, N2 for a calibration curve covering the concentration
range from 0.1 to 75 ng/mL for human plasma samples. (b) Representative multiple reaction-monitoring (MRM) chromatogram of
13
15
AcSDKP. Lower graph indicates 1 ng/mL of AcSDKP- C6, N2 and the upper graph shows serum sample from RA patients.

patients with moderate to high disease activity (DAS28[ESR] ≥ 3.2). The RA group with DAS28 [ESR] lower than
3.2 (n＝23) had a median serum level of AcSDKP of
3.42 ng/mL and the RA group with DAS28 [ESR] higher
than 3.2 (n＝29) had a median level of 2.34 ng/mL. There
was no statistically signiﬁcant diﬀerence between thesegroups (data was not shown).

Fig. 2.

Synovial ﬂuid levels of AcSDKP in patients with RA
and OA

Serum and synovial ﬂuid levels of AcSDKP in patients with RA, OA, and healthy controls.

AcSDKP was detected in the SF of patients with RA andOA. The median SF levels of AcSDKP in patients with OA

Box and whisker plots denote maximum and minimum,
IQR and median. *p＜0.05 was considered signiﬁcant.

were shown to be 0.54 ng/mL, whereas patients with RA
exhibited a median level of 3.07 ng/mL. AcSDKP levels in

AcSDKP levels between healthy controls and OA patients.

the RA SF were shown to be signiﬁcantly higher than in

These data suggest that AcSDKP levels are elevated in RA

OA SF (Fig. 2).

patients compared to OA and healthy individuals.
RA disease activity is categorized as being in remission(DAS 28 [ESR]＜2.6), low (DAS 28 [ESR]＜3.2), moder-

Correlation between serum levels of AcSDKP andclinical featuresate (DAS 28 [ESR] ≥ 3.2), or high (DAS 28 [ESR]＞5.1)
based on speciﬁc DAS 28 [ESR] cut-oﬀ scores. Because thePearson correlation analysis conﬁrmed the correlation
between serum levels of log AcSDKP in RA patients andvast majority of patients with RA in our study were under-

clinical features. Serum levels of log AcSDKP showed

going treatment at the time of study, and most patients wereslightly positive correlation with CRP (r＝0.23, p＝0.95),

diagnosed with low to moderate disease activity, we

however there was no statistically signiﬁcant diﬀerence.

divided patients with RA into two groups based on the
DAS28 [ESR] score to evaluate the ability of AcSDKP to

Serum levels of log AcSDKP had no correlation in DAS28[ESR], MMP-3, ESR. In addition, log AcSDKP exhibited amonitor disease severity and activity: patients who were in

negative trend that did not reach statistical signiﬁcance withremission or low disease activity (DAS28 [ESR]＜3.2) andRF (Fig. 3).
31

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Fig. 3.

Medical Mass Spectrometry Vol. 5 No. 1

Correlation between serum levels of log AcSDKP and indicated clinical measures in patients with RA.

The x-axis reﬂects the concentration of CRP in mg/dL, the DAS28 [ESR] score, the concentration of MMP-3 in ng/mL, the measurement of ESR in mm/h, and the concentration of RF in U/mL. The y-axis reﬂects the plasma concentration of log AcSDKP in
ng/mL. Scatter plot graphs showing the correlation between (a) CRP and AcSDKP, (b) DAS28 [ESR] and AcSDKP, (c) MMP-3 andAcSDKP, (d) ESR and AcSDKP, and (e) RF and AcSDKP.
The relationship between these variables was evaluated using the Pearson correlation test. Trend lines indicate linear correlation.

Discussion

Consecutive 22 pairs of serum AcSDKP at the time ofchanging DMARDs and after 12 weeks

Recent progress in the treatment of RA has been madeThe mean DAS 28 [ESR] score of 22 patients was 4.22

through introduction of MTX, bDMRADs and Janus kinaseand decreased to 2.79 after 12 weeks of treatment (p＜0.01). Clinical and laboratory parameters decreased signiﬁ-

inhibitors. A delay in initiating therapy against RA could
adversely aﬀect treatment outcomes, such as disease activ-

cantly after 12 weeks of treatment in all patients: AcSDKP

ity, remission, functional ability, and radiological progres-levels from 3.74 to 1.98 ng/mL (mean values), CRP levels
from 2.65 to 0.25 mg/dL (mean values), MMP-3 levels fromsion. Therefore, early diagnosis and eﬀective treatment is
20,21)

289.0 to 107.3 ng/mL (mean values), and ESR levels from. There is an unmet need for speciﬁc andvery importanteasy-to-measure biomarkers for the diagnosis of RA and

42 mm/h to 18 mm/h (mean values) at the time of changing

identiﬁcation of patients who are at increased risk of devel-

DMARDs and after 12 weeks, respectively (Fig. 4).

oping erosive, joint destructive disease. We focused on32June 2021

Fig. 4.

Medical Mass Spectrometry Vol. 5 No. 1

Serum samples were collected from 22 patients with RA at baseline and after 12 weeks of treatment with DMARDs.

DAS28 [ESR] (a), serum levels of AcSDKP (b), CRP (c), MMP-3 (d) and ESR (e) were signiﬁcantly down-regulated at 12 weeks
after treatment compared with baseline. Results are presented as the mean±SEM. The mean diﬀerences in the clinical measures between baseline and 12 weeks later were evaluated with the Wilcoxon signed rank test. *p＜0.05 was considered signiﬁcant.

AcSDKP and investigated whether this peptide was an RA

Importantly, the serum levels of AcSDKP were low orbiomarker.

undetectable in healthy controls, as well as in patients with

Osteoarthritis (OA) is a joint disease in which articular

OA. Similarly, we found that the levels of AcSDKP in the

cartilage degeneration and collapse, as well as osseous pro-synovial ﬂuid of patients with RA were signiﬁcantly higher

liferation, are progressively impaired in joint function. Painis also strong, leading to signiﬁcant deterioration in activi-

than in patients with OA.

ties of daily living and quality of life. Compared to RA, OA
has generally been described as a non-inﬂammatory joint

ACE and partially eliminated in urine. Therefore, patients

disease. In recent years, it has become clear that variousof AcSDKP in both their plasma and urine

inﬂammatory cytokines are involved in the development of
OA, however the expression levels are extremely low com-study, none of our tested subjects were prescribed ACE

pared to RA. In this study, sera from healthy subjects and

The expression of AcSDKP was originally considered a
reﬂection of angiogenesis. Indeed, AcSDKP and its precur-

It is known that AcSDKP is physiologically degraded byreceiving ACE inhibitor treatments displayed higher levels
22,23). In ourinhibitors.

OA patients were used as controls.
According to our results, the level of AcSDKP in theserum of patients of RA was 3-fold higher than in healthy

sor peptide, Tβ4, enhanced angiogenesis and exerted antiﬁ-

controls, and 1.5-fold higher than in patients with OA.

function

brotic eﬀects associated with the normalization of organ
24,25)

33. High levels of Tβ4 and AcSDKP have beenJune 2021

Medical Mass Spectrometry Vol. 5 No. 1

nancies, with the levels of AcSDKP being reported to be

3‒4 h in sera . It is suggested that biomarkers with a shorthalf-life will more accurately indicate the disease activity atmarkedly elevated in solid neoplasms. An association

the time of measurement.

associated with tumor progression in hematologic malig-

38)

between the levels of AcSDKP and tumor angiogenesis has
14,26,27)

Pearson correlation analysis conﬁrmed the correlation. In rats after traumatic brain injury,

between serum AcSDKP levels in RA patients and clinicaladministration of AcSDKP promoted neurovascular remod-

features. AcSDKP levels were not signiﬁcantly correlatedeling include angiogenesis and neurogenesis, as well aswith clinical features. Serum AcSDKP levels showed

been observed

28)

improves functional recovery . In healthy condition, the. In recent

slightly positive correlation with CRP, DAS28 [ESR],
MMP-3 and ESR, but not with RF, which exhibited a nega-

study, AcSDPK content was not completely absent in micetive trend that did not reach statistical signiﬁcance. Overall,

12,29)

major source of the AcSDPK have been Tβ4
25,30). Nevertheless ACE activity in SF fromthese ﬁndings show a statistically no signiﬁcant association

patients with RA was signiﬁcantly greater than that in SF

between AcSDKP and clinical features. However, this trend

lacking Tβ4

31,32), the concentrations of AcSDKP wereneeds to be further investigated and validated using larger

higher in RA-derived SF than in OA-derived SF. As the rea-patient cohorts that cover the entire spectrum of RA diseaseson, the production of Tβ4, a precursor of AcSDKP, isactivity.

from OA patients

10,11), and there may be a source of AcSDKP

abundant in RA

From simply diagnostic point of view, the serum levels

diﬀerent from healthy condition.

of Tβ4 were preferable to AcSDKP, because it was reported

Vascular proliferation is known to be one of the earliest

that serum levels of Tβ4 were approximately 10 fold higher

events in the development of RA and could be the causal

in RA patients than in healthy controls. Although, from theview point of responsiveness to anti-rheumatic treatment,

process triggering RA, which has been described as an
33,34). A number of angioangiogenesis-dependent diseaseno signiﬁcant diﬀerence was shown in the change in Tβ4
10)genic factors such as gliostatin and VEGF have been shown

concentration before and after therapy .

to be involved in the neovascularisation process in the RA
joint. Measurements of the serum levels of gliostatin andIn the present study, we investigated the eﬀects of
DMARDs on the serum levels of AcSDKP. Serial labora-

VEGF have been useful methods for monitoring the disease

tory measurements revealed a reduction in the levels of

6,8,35). Therefore, we investigated whether theserum levels of AcSDKP were decreased following treat-AcSDKP following administration of DMARDs. As well as
clinical features, such as DAS28 [ESR], CRP, MMP-3 andment with bDMARDs or MTX. AcSDKP may be instrumental as an easy-to-measure serum marker that can facili-

ESR, the levels of AcSDKP was also shown to have a hightate diagnosis of RA and discriminate RA from OA andthat AcSDKP may be involved in the pathophysiology ofpotentially other forms of arthritis with an inﬂammatorycomponent. In previous reports, AcSDKP had been mea-

RA and may have an eﬀect in the chronic arthritis during

activity of RA

response to treatment with DMARDs. These results suggest

the process of RA.

sured using enzyme-linked immunosorbent assay. In search

Our study has several limitations. First, since the studyof more accurate measurements, we have adopted themethod of SPE-LC-MS/MS analysis which is the quantita-

population was small, the result may not be valid for alltive determination of AcSDKP with relatively simple andsuch as concomitant use of corticosteroid, bDMARDs, andpatients with RA. Second, several baseline characteristics,

18,19)

highly sensitive. Moreover, this analytical method is

csDMARDs were diﬀerent among our study population.

suitable for measuring large numbers of clinical samples.

Third, in real world RA therapy using the National Health

The expression of AcSDKP was determined locally inInsurance medical examination and blood sampling every

the inﬂamed joints and also systemically in circulation of

day or every week are diﬃcult. Serial data pertaining topatients with RA was measured extracellularly. Whether
AcSDKP found extracellularly originates from synovial tis-AcSDKP levels were obtained from some patients treatedsues or is a subject of active secretion is presently
unknown. AcSDKP has a half-life of 4.5 min in the circula-to be responsive to therapy and serum AcSDKP, CRP,
MMP-3 and DAS 28 [ESR] levels gradually decreased with36)
tion, because it is hydrolyzed by ACE . The half-life of37)
CRP is 19 h , while the half-life of AcSDKP is as short ascourse of treatment (data not shown).

with DMARDs every 4 weeks. These patients were found34

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Medical Mass Spectrometry Vol. 5 No. 1

Conclusions

2) Smolen JS, Aletaha D, Mcinnes IB: Rheumatoid arthritis.
Lancet 388: 2023‒2038, 2016.

In conclusion, we found that the levels of AcSDKP wereincreased in the serum and synovial ﬂuid of patients with3) Elshabrawy HA, Chen Z, Volin MV, Ravella S, Virupan-

RA. Moreover, the serum levels of AcSDKP reﬂected

navar S, et al: The pathogenic role of angiogenesis in

improvement in the disease activity during the clinicalrheumatoid arthritis. Angiogenesis 18: 433‒448, 2015.

course of patients with RA. Our results validate AcSDKP4) Mcinnes IB, Schett G: The pathogenesis of rheumatoid arthritis. N Engl J Med 365: 2205‒2219, 2011.

for future studies focusing on its potential as a biomarker
for RA.

5) Takeuchi M, Otsuka T, Matsui N, Asai K, Hirano T, et al:
Aberrant production of gliostatin/platelet-derived endo-

Declarations

thelial cell growth factor in rheumatoid synovium. Arthritis Rheum 37: 662‒672, 1994.

Ethics approval and consent to participate
The study protocol was approved by the Nagoya City

6) Waguri Y, Otsuka T, Sugimura I, Matsui N, Asai K, et al:

University Ethics Committee and Informed consent was

Gliostatin/platelet-derived endothelial cell growth factor

obtained from all the patients upon their enrollment in theas a clinical marker of rheumatoid arthritis and its regula-study.

tion in ﬁbroblast like synoviocytes. Br J Rheumatol 36:
315‒321, 1997.

Consent for Publication

7) Muro H, Waguri-Nagaya Y, Mukofujiwara Y, Iwahashi T,
Otsuka T, et al: Autocrine induction of gliostatin/plate-

Written informed consents were obtained from all the

let-derived endothelial cell growth factor (GLS/PD-EC-

patients for publication of this report.

GF) and GLS-induced expression of matrix metallopro-

Acknowledgements

teinases in rheumatoid arthritis synoviocytes. Rheumatol
38: 1195‒1202, 1999.

This research was supported, in part, by MEXT/

8) Tanikawa T, Waguri-Nagaya Y, Kusabe T, Aoyama M,

JSPSKAKENHI Grant Number 18K09114, 19K18473 andAsai K, et al: Gliostatin/thymidine phosphorylase-regulat-

20K09465.

ed vascular endothelial growth-factor production in hu-

We would like to thank Editage (www.editage.com) for

man ﬁbroblast-like synoviocytes. Rheumatol Int 27: 553‒

English language editing.

559, 2007.

Conﬂict of Interest

9) Crockford D, Turjman N, Allan C, Angel J: Thymosin

The authors declare that they have no conﬂicts of inter-beta4: structure, function, and biological properties sup-est.

porting current and future clinical applications. Ann N Y
Acad Sci 1194: 179‒189, 2010.

Authors Contributions

10) Song R, Choi HM, Yang HI, Yoo MC, Park YB, et al: As-

YO, YK, KA, KI and YW-N designed the project, per-sociation between serum thymosin β4 levels of rheuma-

formed most of the experiments and analyzed the data. YO

toid arthritis patients and disease activity and response to

and YK also wrote the manuscript. HO, YJ, HY, KM, HA,

therapy. Clin Rheumatol 31: 1253‒1258, 2012.

GK and NT assisted with the interpretation of the data and11) Kim KS, Yang HI: Thymosin β4 in rheumatoid arthritis:
Friend or foe. Biomed Rep 7: 205‒208, 2017.

the preparation of the manuscript. MK and MN provided
the RA serums and participated in designing some of theexperiments. YW-N, HM designed the study, analyzed the12) Hannappel E: Thymosin beta4 and its posttranslationaldata, supervised the overall project and wrote the manu-

13) Lenfant M, Wdzieczak-Bakala J, Guittet E, Prome Jc, Sot-

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