64 （228）

診療と新薬・第 53 巻

第 3 号（2016 年 3 月）

USEFULNESS OF THE SUPPLEMENT CONTAINING PROTEOGLYCANFOR JAPANESE HEALTHY PEOPLE FEELING KNEE S DISCOMFORT
Masatomo NAJIMA1), Mitsuhiko MUNEKATA2) and Yoshifumi SOEDA3)
1）JACTA (Japan Clinical Trial Association), 5-27-3 Sendagaya, Shibuyaku, Tokyo, Japan
2）OZ Clinic3）Smile-Japan co., ltd.

Abstract
Objectives: The objective of this study is to verify the knee joint pain relieving effect of ingesting foodcontaining proteoglycan.
Methods: A randomized placebo-controlled, double-blind study was conducted in 36 subjects with symptoms ofknee joint discomfort. The test food was a tablet containing proteoglycan. Subjects consumed three tablets
every day for 12 weeks. Visual analog scale (VAS) for knee pain, the knee range of motion measurements andthe modiﬁed J-KOOS (Japan-Knee injury and Osteoarthritis Outcome Score) was evaluated as primary outcomes.
Results: Out of the 36 subjects, 5 dropped due to personal reasons and the remaining 31 subjects were evaluated[the mean of the age is 55.0±7.6 (test food) and 52.1±11.1 (placebo)]. Pain Vas, signiﬁcant diﬀerences wereobserved in the intragroup comparison of the test food (in repose, in walking and in stairs going up and down)
and also in the intergroup comparison of changes from the baseline (in stairs going up and down). With respect
to the right and left knee extension and ﬂexion, there were signiﬁcant diﬀerences in the intragroup comparisonof the test food and in the intergroup comparison of changes from the baseline. Furthermore, several categories
on the questionnaire of modiﬁed J-KOOS showed the signiﬁcant diﬀerences of changes from the baseline in 10
items and the total score. In addition, no abnormal change had been triggered by the ingestion of the test food.
Conclusion: It is suggested that the ingestion of the supplement containing proteoglycan by Japanese healthypeople feeling knee s discomfort for 12 weeks contributed to improving the joint support.
Key words: proteoglycan, locomotive syndrome, knee joint, salmon

1. INTRODUCTION
Articulation consists of cartilage cells, cartilage matrix,
subchondral bone, tendons and ligaments around thearticulation and muscles. These components undergo
various changes with increasing age. The progression ofthese changes may trigger off the development of
osteoarthritis (OA), the symptom associated with chronic
pain1). Now in Japan, the number of patients who sufferfrom OA increases significantly due to the aging of
population2). Once developed, there is no definite cure/therapy to inhibit the progression of OA, therefore it is
crucially important to ﬁnd out the means, other than drugintervention, for avoiding the development of and/or
inhibiting the progression of OA.
Among the changes associated with the aging, thechange of cartilage matrix is a key factor for developing
OA. It is reported that the volume of articular cartilage ofthe knee joint, which mainly consists of cartilage matrix,
decreases with age 3). The cartilage matrix contains
various types of collagen and proteoglycan in addition to aplenty of moisture. The change of cartilage matrix withthe age is thought to be related to the fact that the
turnover period for the protein of these collagen andproteoglycan is relatively longer for the aged people andtherefore they are easily modiﬁed during the period4).

Proteoglycan, on the other hand, is regarded as an ediblein general, and is an ingredient which is extracted fromthe nasal cartilage of salmon. It is reported that the
supplement containing glucosamine and chondroitin
sulfate, which are components of salmon-derived
proteoglycan5) , possibly contributes to the pain relief of
and/or the inhibition of progression of OA6) 〜8). However,there are very few numbers of reports that illustrate therelationship between the ingestion of proteoglycan itself
by healthy aged person and the prevention ofdevelopment of OA.
In this study, we examined the effect of a foodcontaining proteoglycan to verify the joint support andthe safety of the food. The test targets were Japanese
healthy person who are feeling knee s discomfort, and thetest method was a randomized, placebo-controlled,
double-blind study.

2.METHOD
2.1. Trial Design
A randomized, placebo-controlled, double-blind study was
conducted with the aid of a fund from Smile-Japan co.,ltd.
(Fukushima) at two centers (OZ clinic, Tokyo and
JACTA, Tokyo).
The study period was 12 weeks, from June 27 th toSeptember 19th, 2015.

診療と新薬・第 53 巻

This study was conducted in accordance with the
ethical principles of the declaration of Helsinki. The study
protocol was approved by the Institutional Review Boardof LLP. Pharmaceutical Law Wisdoms (Tokyo). Written
informed consent was obtained from all subjects.
The allocation of the test product to the subjects was
carried out by the person in charge of allocation.
The allocation list was sealed and strictly controlled ina safe deposit box of JACTA until the end of the study.
The clinical trial was registered at UMIN clinical Trial
Registration (Trial ID: UMIN000017839).
2.2. Subject
Healthy subjects participated in the present study.
All of the subjects in this study were public volunteers
who had enrolled in the monitor bank of CROee Inc.
(Tokyo).
2.2.1. Inclusion Criteria(1) Healthy people aged between 40 and 69 years;(2) Modiﬁed J-KOOS (Japan-Knee injury and Osteoarthritis
Outcome Score)9) : Appendix.
2.2.2. Exclusion criteria(1) Individuals already undertaking medical treatment forknee pain or discomfort;(2) Individuals on taking medication, including herbal
medicines;(3) Individuals judged to be unsuitable to participate in
the trial by the doctor conducting present study.
2.2.3. Eﬃcacy eligibility
With respect to the analysis of efficacy, we set thefollowing criteria of exclusion:(1) Participants who consumed less than 80% of the
expected dose;(2) Participants without adequate records;(3) Participants who fell under the exclusion criteria after
enrolment;(4) Participants who had justiﬁable reason for exclusion.
2.3. Randomization
Subjects who fulfilled the eligibility criteria were 36
persons. The inclusion was judged by the principle
investigator. All subjects were sequentially assigned
Table 2Term
Item

● : Implementation↔ : Daily practice during the test period

65 （229）

based on a random number table to one of the masked
products and randomized to group P (placebo: 18) andgroup T (test sample: 18)
The allocation was pre-assigned on the basis ofrandomized numbers.
2.4. Description of test foods and blinding
The test food Locomo Ace ( LA ) is a tablet containing
salmon-derived proteoglycan. The amount of a daily
intake is 3 tablets (750 mg). In the 3 tablets, salmonderived proteoglycan is included over 10 mg, the placebodoes not include proteoglycan. Table 1. shows thenutritional content of the sample. Both tablets wereindistinguishable in shape, color or taste. Tablets weremanaged by the identiﬁcation symbol. All involved wereblinded.
2.5. Experimental procedures
2.5.1. Experimental protocol
Subjects consumed 3 tablets of the supplement with hotor cold water every day for 12 weeks. Subjects were
instructed as follows: to take the assigned foods as
indicated; to maintain their usual lifestyles and habits; toavoid excessive amounts of food, drink, or alcohol; tomaintain a daily record of lifestyle factors such as all thefood items they consumed during the test period; and tosend the diary to the study coordinator on every Friday
by mobile email.
2.5.2. Outcome
The objective of this study is to verify the knee joint pain
relieving eﬀect of ingesting food containing proteoglycan.

Table 1 Nutritional content of the sample per 100 g
Item
EnergyProtein
Lipid
Ash
Carbohydrates
Na

LA

Placebo

394 kcal
6.8 g
2.6 g
5.6 g
85.8 g
182 mg

215 kcal
0.06 g2.77 g1.48 g95.11 g0.706 mg

Schedule for the study

Screening

Modiﬁed J-KOOS
Informed consent
Selection and/or allocation
VAS, Knee extension and ﬂexion
Biochemical analysis of blood
Urine analysis
Ingestion of test foods
Log

第 3 号（2016 年 3 月）

Pre Trial
Test

●
●
●
●
●
●

Test period
4w

8w

12 w

●

●

●

●

●

●
●
●

66 （230）

診療と新薬・第 53 巻

第 3 号（2016 年 3 月）

Applicants (n = 140)
(M：63, F：77)
Exclusion (n = 104)
(M：45, F：59) <J-KOOS>

LA (n = 18)(M：8, F：10)

Placebo (n = 18)(M = 10, F = 8)

Withdrawn (n = 2)
(M：1, F：1)
<personal reasons>Withdrawn (n = 3)
(M：2, F：1)
<personal reasons>

Full analysis set
LA (n = 15)(M：11, F：9)

Placebo (n = 16)(M：10, F：7)

Figure 1 Flow diagram of subject disposition

Table 3 Subject demographics
Item
Subjects＊

Male : Female＊

Age
VAS (in repose)
VAS (in walking)
VAS (in going up and down stairs)
Modiﬁed J-KOOS

Unit

LA

Placebo

−
−
years
mm
mm
mm
point

15
9:6
55.0 ± 7.6
9.7 ± 11.2
19.4 ± 11.3
35.4 ± 18.1
52.9 ± 8.1

16
7:9
52.1 ± 11.1
18.0 ± 19.8
28.6 ± 17.3
40.4 ± 19.2
48.3 ± 11.7＊

Number of subjects
mean ± SD

To evaluate this objective, visual analog scale (VAS)10)
for knee pain was conducted as the primary outcomes.
The knee range of motion measurements and themodified J-KOOS was also observed as the primary
outcome. The questionnaire of the modified J-KOOS
covered: joint discomfort, pain, function and the qualityof life. Blood biochemical and urine parameters wererecorded to evaluate the safety of the test foods as thesecondary outcome.
To evaluate the safety of the test foods, adverse eventswere collected by means of a written questionnaireduring the study.
According to the schedule shown in Table 2, we
measured parameters on eﬃcacy and safety.
2.6. Data Analysis
All analyses were performed on the on-treatment
population in the study.
Data were expressed as mean ± SD. For pain VAS,
knee extension, flexion, and biochemical analyses ofblood and urine, changes from the baseline in the samegroup were assessed using the paired t-test. Student's
t-test was used for intergroup comparisons of changes
from the baseline. For the modified J-KOOS, changes
from baseline in the same group were assessed using the
Wilcoxon signed-rank test. The Mann‒Whitney U test
was used for intergroup comparisons of changes from thebaseline. Student s t-test was used to compare subject
backgrounds between groups. Statistical analyses were
performed using Statcel 3 (Yanai, 2011). The results
were considered signiﬁcant at the < 5% level in the twosided test.

3. RESULT
3.1. Participant Demographics
From all of 140 applicants, 104 were eliminatedaccording to the modiﬁed J-KOOS score. The 36 subjects
were randomly assigned to intervention groups and madea start with ingestion. 5 were withdrawn due to personal

診療と新薬・第 53 巻

Table 4Item
<unit>

Group

Baseline
Mean ± SD

第 3 号（2016 年 3 月）

Results of test analyses

4 weeks
Mean ± SD

67 （231）

P-value1)

8 weeks
P-value2)

Mean ± SD

P-value1)

12 weeks
P-value2)

Mean ± SD

P-value1)

VAS (in repose)
<mm>

LA
P

9.7 ± 11.2
18.0 ± 19.8

11.3 ± 15.1
8.0 ± 11.0

0.707
0.105

0.117

5.1 ± 7.3
18.1 ± 16.9

0.182
0.938

0.236

1.9 ± 3.3
15.5 ± 13.1

0.453

VAS (in walking)
<mm>

LA
P

19.4 ± 11.3
28.6 ± 17.3

19.9 ± 15.9
28.6 ± 17.4

0.913
0.984

0.938

18.3 ± 7.8
25.9 ± 15.3

0.728
0.298

0.702

11.1 ± 11.9
26.6 ± 17.4

0.040 ＊
0.549

0.109

VAS (in stairs going up
and down)
<mm>

LA
P

35.4 ± 18.1
40.4 ± 19.2

31.4 ± 14.5
37.5 ± 19.8

0.464
0.402

0.866

21.1 ± 10.2
36.5 ± 20.5

0.024 ＊
0.217

0.121

10.5 ± 5.3 ＜ 0.001 ＊＊31.4 ± 19.5
0.162

0.041 ＃

knee extension (right)<degree>LAP

23.0 ± 7.1
17.0 ± 10.4

16.6 ± 8.5
18.8 ± 10.7

0.033 ＊

0.013 ＃

16.4 ± 5.9
17.8 ± 9.3

0.003 ＊＊

12.2 ± 5.3
17.2 ± 5.5＜ 0.001 ＊＊0.510

0.005 ＃＃

knee extension (left)<degree>LAP

23.7 ± 7.5
20.4 ± 12.1

14.9 ± 10.3
21.2 ± 7.7

0.009 ＊＊
0.404

0.011 ＃

14.5 ± 4.4
16.1 ± 7.7＜ 0.001 ＊＊0.117

0.070‡

10.1 ± 3.4
17.5 ± 4.9＜ 0.001 ＊＊＜ 0.001 ＃＃0.299

knee ﬂexion (right)<degree>LAP

128.3 ± 8.9
129.0 ± 9.3

124.5 ± 8.1
125.7 ± 8.5

0.085†
0.104

0.721

129.3 ± 8.4
127.2 ± 8.2

0.726
0.336

0.417

131.8 ± 8.8
125.1 ± 8.9

0.192
0.013 ＊

0.020 ＃

knee ﬂexion (left)<degree>LAP

125.0 ± 6.0
125.3 ± 13.0

124.1 ± 8.9
126.9 ± 9.2

0.721
0.640

0.557

122.7 ± 11.2
126.5 ± 10.3

0.493
0.671

0.417

129.2 ± 6.3
124.4 ± 11.7

0.029 ＊
0.313

0.025 ＃

0.169

0.017

P-value2)

＊

0.843

0.066‡＜ 0.001 ＃＃n = 31
1) ＊ : P ＜ 0.05, ＊＊ : P ＜ 0.01 intragroup diﬃerences versus baseline
2) ＃ : P ＜ 0.05, ＃＃ : P ＜ 0.01 intergroup diﬀerences between changes from baseline

reasons (3; occupational conflict, 1; cold, 1;
tendovaginitis) and the remaining 31 subjects completed
the study. Thus, data obtained from the 31 subjects (testgroup; 15, placebo group; 16) were used for efficacy
analysis (Figure 1.). There were no significant
differences in the mean age, gender ratios, VAS or
modiﬁed J-KOOS observed between groups (Table 3.).
3.2. VAS
Pain VAS was conducted in three situations. All of whichshowed significant differences from the baseline of LAgroup after 12 weeks of ingestion. First and foremost,the significant Pain VAS recorded in repose showed achange of p = 0.017. Subsequently, the measurement ofchange on the Pain VAS of walking came to p = 0.040 and
the Pain VAS of going up and down the stairs recorded at
p < 0.001. Overall the intergroup comparison of changes
between LA group and placebo group came to p = 0.041
from going up and down the stairs (Table 4.).
3.3. Knee extension and ﬂexion
Table 4. summarizes the average knee extension andﬂexion changes over time for subjects supplemented witheither LA or placebo.
For the results of the right and left extension, the rightknee extension changed signiﬁcantly from the baseline of
LA group after 4,8 and 12 weeks of ingestion (p = 0.033,
p = 0.003, p < 0.001, respectively). Furthermore,significant differences were observed between thechanges from the baseline of two groups after 4, 8 and 12
weeks of ingestion (p = 0.013, p = 0.005, p < 0.001,
respectively). The left knee extension showed signiﬁcantdiﬀerences from the baseline of LA group after 4, 8, 12
weeks of ingestion (p = 0.009, p < 0.001, p < 0.001,
respectively). Moreover there were significant
differences between the change from baseline of twogroups after 4 and 12 weeks of ingestion (p = 0.011, p <0.001, respectively).
For the results of the right and left knee flexion, asignificant difference was observed in changes from thebaseline of the two groups after 12 weeks of ingestion.
The right knee showed p = 0.020, while the left knee
demonstrated p = 0.025.
3.4. Modiﬁed J-KOOS
Table 5. reveals the Modified J-KOOS. Stiffness, Pain,
Function (daily living) and Quality of Life were asked as
Modiﬁed J-KOOS.
The result based on Modiﬁed J-KOOS criteria, showedthat a change of the aggregate scores was significantly
improved at week 4 and 12 in the LA group compared tothe placebo group. In the intergroup difference ofchanges from baseline, Question S1. ( Stiffness ; Howsevere is your knee joint stiﬀness after ﬁrst wakening in
the morning?), S2. (How severe is your knee stiffness
after sitting, lying or resting later in the day?), P2.
( Pain ; Walking on flat surface), A1. ( Function, dailyliving ; Descending stairs), A3. (Rising from sitting), A5.
(Bending to ﬂoor/ picking up an object), A6 (Walking on
ﬂat surface), A12. (Sitting), Q1. ( Quality of life ; Dueto pain, do you restrict your hobbies or recreational
activities?) and Q3. (Is your overall health condition
affected by your knee condition?) illustrated significant
diﬀerences.
3.5. Blood and Urine test
Table 6. and 7. show the blood biochemical and urine
parameters.
The change of Total Protein, Neutral Fat (TG) and
Creatinine in LA group, revealed a signiﬁcant diﬀerence
after 12 weeks of ingestion. However the change of Total
Protein and Creatinine was within the standard value. 4
subjects with Neutral Fat changed over the standard
value after 12 weeks of ingestion, but the investigator68 （232）

診療と新薬・第 53 巻

第 3 号（2016 年 3 月）

Table 5 Results of questionnaire (Modiﬁed J-KOOS)
Item

Group

Baseline
Mean ± SD

4 weeks
Mean ± SD

P-value1)
＊＊

0.002

8 weeks
P-value2)

Mean ± SD

P-value1)

0.034 ＃

41.9 ± 10.9
43.4 ± 8.3＊＊

0.015 ＊

＜ 0.001 ＃＃1.6 ± 0.6
2.5 ± 0.5

0.001 ＊＊
0.310

12 weeks
P-value2)

Mean ± SD

P-value1)

0.093‡

38.9 ± 10.2
46.4 ± 8.6＊＊

P-value2)

0.235

0.236

1.7 ± 0.7
2.7 ± 0.7

0.001 ＊＊
0.123

＜ 0.001 ＃＃0.048 ＃

1.8 ± 0.9
2.3 ± 0.4

0.011 ＊
0.686

0.017 ＃

Total

LA
Placebo52.9 ± 8.1
48.3 ± 11.7

42.7 ± 10.7
44.6 ± 7.0

0.079†

S1.

LA
Placebo

2.9 ± 0.7
2.3 ± 0.6

1.7 ± 0.7
2.9 ± 0.6

0.006 ＊＊

S2.

LA
Placebo

2.7 ± 0.7
2.3 ± 0.5

2.2 ± 1.0
2.1 ± 0.3

0.173
0.225

0.363

2.1 ± 0.7
2.3 ± 0.5

1.000

P1.

LA
Placebo

3.9 ± 1.0
3.5 ± 0.8

3.1 ± 1.1
3.1 ± 0.6

0.114
0.018 ＊

0.514

3.2 ± 1.2
2.8 ± 0.7

0.131
0.008 ＊＊

0.984

2.7 ± 1.2
2.9 ± 0.9

0.025 ＊
0.059†

0.323

P2.

LA
Placebo

2.2 ± 0.4
1.9 ± 0.7

1.7 ± 0.7
2.1 ± 0.3

0.033 ＊

0.031 ＃

1.6 ± 0.6
1.5 ± 0.5

0.022 ＊
0.142

0.514

1.5 ± 0.6
2.1 ± 0.3

0.013 ＊
0.237

0.004 ＃＃

P3.

LA
Placebo

2.5 ± 0.7
2.4 ± 0.5

2.2 ± 0.7
2.3 ± 0.5

0.401
0.361

0.859

1.9 ± 0.6
2.1 ± 0.3

0.043 ＊

0.477

2.0 ± 0.5
2.3 ± 0.6

0.374

1.3 ± 0.5

0.317

1.2 ± 0.41.000

1.0 ± 0.0

0.109

1.0 ± 0.0

0.109

0.353

1.3 ± 0.6
1.2 ± 0.4

0.208
0.0499 ＊0.635

1.3 ± 0.5
1.4 ± 0.5

0.262
0.401

0.678

0.874

1.7 ± 0.6
2.2 ± 0.4

0.019 ＊
0.686

0.040 ＃

1.7 ± 0.6
2.3 ± 0.6

0.022 ＊
1.000

0.024 ＃

0.385

1.8 ± 0.8
2.1 ± 0.3

0.086†
0.225

0.385

1.7 ± 0.7
2.1 ± 0.3

0.033 ＊
0.225

0.185

0.001 ＊＊

0.401

LA

1.2 ± 0.4

1.1 ± 0.4

0.317

Placebo

1.2 ± 0.4

1.1 ± 0.3

0.593

P5.

LA
Placebo

1.5 ± 0.6
1.6 ± 0.9

1.4 ± 0.6
1.2 ± 0.4

0.529
0.0499 ＊A1.

LAPlacebo

2.3 ± 0.7
2.3 ± 0.6

1.9 ± 0.6
2.1 ± 0.3

0.273

A2.

LA
Placebo

2.3 ± 0.7
2.3 ± 0.6

1.8 ± 0.8
2.1 ± 0.3

0.063†
0.345

P4.

A3.

LA
Placebo

2.8 ± 1.0
2.7 ± 0.8

1.9 ± 0.6
2.5 ± 0.6

A4.

LA
Placebo

1.9 ± 0.8
1.9 ± 0.9

1.3 ± 0.6
1.3 ± 0.7

1.000

0.018 ＊

0.005

＊＊

0.374
0.033 ＊
0.012 ＊

A5.

LA
Placebo

2.7 ± 0.8
2.1 ± 0.9

1.9 ± 0.7
1.8 ± 0.8

0.005

A6.

LA
Placebo

2.0 ± 0.0
1.6 ± 0.5

1.3 ± 0.5
1.5 ± 0.5

0.180

A7.

LA
Placebo

2.1 ± 0.6
1.8 ± 0.6

1.7 ± 0.6
1.4 ± 0.5

A8.

LA
Placebo

1.6 ± 0.6
1.7 ± 0.8

A9.

LA
Placebo

A10.

0.003

0.033 ＊

0.074†

0.252＊＊

0.055‡

2.0 ± 0.8
2.4 ± 0.6

0.008
0.262

1.000

1.3 ± 0.5
1.3 ± 0.7

0.008 ＊＊＊＊

0.018 ＊

0.220

1.9 ± 0.7
1.8 ± 0.8

0.013
0.248

0.0101 ＃1.4 ± 0.5
1.6 ± 0.5

0.076†
0.068†

0.664

1.6 ± 0.6
1.5 ± 0.6

1.000
0.225

2.2 ± 0.4
2.2 ± 1.0

1.7 ± 0.5
1.8 ± 0.5

0.076†LA
Placebo

1.3 ± 0.6
1.3 ± 0.4

1.2 ± 0.6
1.1 ± 0.3

A11.

LA
Placebo

1.7 ± 0.9
1.5 ± 0.7

A12.

LA
Placebo

A13.

0.002

0.001 ＃＃

0.076†

0.406

＊＊

0.161

1.8 ± 0.7
2.8 ± 0.7

0.003
0.735

0.813

1.1 ± 0.4
1.3 ± 0.7

0.008 ＊＊＊

0.567

0.002 ＃＃

0.012 ＊

0.937

＊＊

0.236

1.8 ± 0.6
2.0 ± 0.5

0.005
0.575

0.008 ＊＊
0.593

0.018 ＃

1.3 ± 0.5
1.8 ± 0.4

0.005 ＊＊
0.361

0.002 ＃

1.7 ± 0.7
1.4 ± 0.5

0.110
0.068†

0.607

1.5 ± 0.5
1.7 ± 0.7

0.022 ＊
0.753

0.0504‡0.514

1.5 ± 0.5
1.6 ± 0.7

0.594
0.735

0.906

1.5 ± 0.5
1.6 ± 0.7

0.594
0.735

0.906

0.465

1.9 ± 0.6
1.9 ± 0.8

0.139
0.178

0.567

1.7 ± 0.6
2.1 ± 1.0

0.686

0.593
0.361

0.797

1.3 ± 0.5
1.1 ± 0.3

1.000
0.463

0.580

1.2 ± 0.4
1.1 ± 0.3

0.317
0.225

0.553

1.6 ± 0.6
1.3 ± 0.5

0.686
0.345

0.921

1.5 ± 0.5
1.3 ± 0.5

0.500
0.345

0.828

1.4 ± 0.5
1.6 ± 0.7

0.237
0.800

0.333

1.9 ± 0.7
1.7 ± 0.8

1.2 ± 0.4
1.6 ± 0.6

0.151
0.500

0.060‡

1.3 ± 0.5
1.4 ± 0.5

0.018 ＊
0.068†

0.236

LAPlacebo

1.3 ± 0.5
1.3 ± 0.5

1.2 ± 0.4
1.3 ± 0.4

0.686
0.317

0.953

1.5 ± 0.5
1.2 ± 0.4

0.034 ＊
0.180

A14.

LA
Placebo

2.1 ± 0.6
2.3 ± 1.1

2.0 ± 0.8
2.0 ± 0.5

1.000
0.333

0.937

2.1 ± 0.8
2.1 ± 0.9

A15.

LA
Placebo

2.0 ± 0.5
2.0 ± 1.0

2.2 ± 0.9
2.1 ± 1.0

0.401
0.594

0.813

Q1.

LA
Placebo

2.1 ± 0.3
1.4 ± 1.0

1.3 ± 0.6
1.3 ± 0.8

0.686

Q2.

LA
Placebo

1.3 ± 0.6
1.1 ± 0.3

1.2 ± 0.4
1.2 ± 0.4

0.554
0.361

Q3.

LA
Placebo

2.6 ± 0.7
2.1 ± 1.1

2.1 ± 1.0
1.9 ± 0.8

0.083†

0.110
0.005 ＊＊

0.012 ＊

0.0096 ＊＊0.529

0.025 ＊

0.033 ＃

0.053‡

1.3 ± 0.5

0.012 ＊

1.6 ± 0.7

0.593

0.089‡

1.3 ± 0.5
1.2 ± 0.4

1.000
0.180

1.000
0.374

0.514

1.7 ± 0.7
2.4 ± 0.7

0.310

2.1 ± 0.8
2.1 ± 0.8

0.735
0.767

0.363

1.7 ± 0.7
1.8 ± 0.8

0.237
0.310

0.874

0.020 ＃

1.3 ± 0.5
1.3 ± 0.5

0.002 ＊＊
1.000

0.016 ＃

1.2 ± 0.4
1.2 ± 0.4

0.002 ＊＊
0.500

0.005 ＃＃

0.453

1.1 ± 0.3
1.2 ± 0.4

0.178
0.361

0.179

1.0 ± 0.0
1.2 ± 0.4

0.361

0.027 ＃

1.8 ± 0.8
2.0 ± 0.8

0.021 ＊
0.859

0.138

1.7 ± 0.7
2.2 ± 0.8

0.012 ＊
0.575

n = 31
1) ＊＊ : P ＜ 0.01 intragroup diﬃerences versus baseline
2) ＃ : P ＜ 0.05, ＃＃ : P ＜ 0.01 intergroup diﬀerences between changes from baseline

0.076†

0.068†

0.034 ＃
0.580
0.0504‡0.101
0.027 ＃

診療と新薬・第 53 巻

Table 6

第 3 号（2016 年 3 月）

69 （233）

Changes in biochemical blood test
Baseline

12 weeks

Mean ± SD

Mean ± SD

LAPlacebo0.72 ± 0.40
0.81 ± 0.48

0.67 ± 0.35
0.84 ± 0.52

M/F

LA
Placebo

7.2 ± 0.4
7.4 ± 0.4

7.5 ± 0.4 ＊＊7.5 ± 0.4†

3.8‑5.3

M/F

LA
Placebo

4.4 ± 0.2
4.6 ± 0.2

4.4 ± 0.2
4.5 ± 0.2

U/L

8‑38

M/F

LA
Placebo20.1 ± 5.1
20.3 ± 4.4

22.1 ± 6.0†
21.4 ± 4.6†

ALT (GPT)

U/L

4‑43

M/F

LA
Placebo15.5 ± 5.6
17.1 ± 5.9

15.8 ± 6.2
17.6 ± 7.6

ALP

U/L

110‑354

M/F

LA
Placebo207.9 ± 59.2
224.9 ± 74.6

219.6 ± 70.2
227.0 ± 70.0

LD (LDH)U/L

121‑245

M/F

LA
Placebo190.6 ± 37.9
174.1 ± 27.4

202.6 ± 49.2
175.0 ± 22.8

86 andunder

M

LA
Placebo20.5 ± 7.7
42.6 ± 44.4

24.3 ± 7.8
49.9 ± 59.4†

48 andunder

F

LA
Placebo20.6 ± 12.8
68.9 ± 138.1

20.0 ± 9.4
58.0 ± 109.5

38‑196

M

LA
Placebo156.7 ± 71.8
114.8 ± 55.4

199.8 ± 63.8
126.8 ± 59.7

30‑172F

LA
Placebo110.8 ± 42.6
101.1 ± 26.9

140.0 ± 71.9
90.1 ± 29.3

LA
Placebo207.9 ± 38.2
218.1 ± 39.9

209.3 ± 38.5
219.3 ± 46.9

LA99.1 ± 46.7

Placebo

143.8 ± 63.8

128.1 ± 57.0 ＊141.3 ± 73.0

Item

Unit

Std. Value

Gender

Group

Total Bilirubin

mg/dL

0.2‑1.2

M/F

Total Protein

g/dL

6.5‑8.3

Albumeng/dl

AST (GOT)

γ-GT (γGTP)CK (CPK)

U/L

U/L

Total Cholesterolmg/dL

130‑219

M/F

Neutral Fat (TG)mg/dL

30‑149M/F

SodiummEq/L

135‑150

M/F

LA
Placebo

142.9 ± 2.2
142.9 ± 2.0

142.5 ± 2.0
142.6 ± 2.5

Chloride

mEq/L

98‑110

M/F

LAPlacebo105.7 ± 2.4
104.2 ± 2.1

105.2 ± 1.9
103.2 ± 1.9†

Potassium

mEq/L

3.5‑5.3

M/F

LA
Placebo

4.3 ± 0.3
4.1 ± 0.3

4.3 ± 0.2
4.1 ± 0.3

Calcium

mg/dL

8.4‑10.2

M/F

mg/dL

2.5‑4.5

M/F

mg/dL

8.0‑22.0

M/F

Inorganic
Phosphorus
Urea Nitrogen

Creatinine

mg/dL

9.6 ± 0.3

9.7 ± 0.3

9.6 ± 0.3

9.8 ± 0.4 ＊LA

3.5 ± 0.6

3.6 ± 0.4

Placebo

3.4 ± 0.4

3.4 ± 0.6
14.6 ± 3.9

LA14.1 ± 4.1

Placebo

14.4 ± 3.8

15.1 ± 4.1

0.81 ± 0.10
0.90 ± 0.19

0.76 ± 0.08 ＊0.88 ± 0.22

0.61‑1.04

M

LA
Placebo0.47‑0.79

F

LA
Placebo0.69 ± 0.09
0.66 ± 0.09

0.65 ± 0.09 ＊＃0.61 ± 0.09†

60‑109

M/F

LA
Placebo70.1 ± 6.1
73.8 ± 9.8

72.3 ± 7.0
73.8 ± 14.3

mg/dL

Blood Sugar(Serum)LA
Placebon = 31†: P ＜ 0.1,＃＊

: P < 0.05, ＊＊ : P ＜ 0.01 intragroup diﬀerences versus baseline: P ＜ 0.05 intergroup diﬀerences between changes from baseline70 （234）

診療と新薬・第 53 巻

第 3 号（2016 年 3 月）

Table 7 Transition of Urinalysis
Baseline

12 weeks

Mean ± SD

Mean ± SD

LAPlacebo1.016 ± 0.005
1.019 ± 0.006

1.017 ± 0.005
1.019 ± 0.006

LAPlacebo

5.8 ± 0.8
5.9 ± 0.8

5.8 ± 0.9
5.8 ± 0.8

Item

Unit

Std. Value

GenderGroup

Speciﬁc Gravity

mg/dL

1.010‑1.025

M/F

pH

g/dL

4.5‑8.0

M/F

n = 31judged it as within the range of physiological variation.
No signiﬁcant change was detected in the urine test.

4. DISCUSSION
We conducted a randomized, placebo-controlled, doubleblind study to verify the joint support effects of the
supplement containing proteoglycan. As the primary
outcome, the study showed signiﬁcant diﬀerences in thescores of VAS (Visual Analogue Scale), in the intragroupcomparison of LA group (in repose, in walking and instairs going up and down) and also in the intergroupcomparison of changes from the baseline (in stairs goingup and down). In addition, as for the right and left knee
extension and ﬂexion, there were signiﬁcant diﬀerencesin the intragroup comparison of LA group and in theintergroup comparison of changes from the baseline.
Furthermore, several categories on the questionnaire ofmodified J-KOOS ( Japanese Knee Injury and
Osteoarthritis Outcome Score) showed the significant
diﬀerences of changes from the baseline in 10 items andthe total score. At the same time, as the secondary
outcome the observation of clinical findings such as
medical interview, blood and urine test revealed noabnormal change had been triggered by the ingestion oftest products.
Evaluating the above findings comprehensively, it issuggested that the ingestion of the test productcontaining proteoglycan contributes to the improvementof the knee condition.
Proteoglycan contained in this test product is one of
the glycoproteins which are the complex of sugar andprotein, and it is a biogenic substance people have by
nature. It excels in moisture holding ability andcontributes to the maintenance of body tissue by
producing an extracellular matrix with collagen andhyaluronic acid. Especially in articulation (joints), it iscontained in the cartilage and helps the smooth
movement of joints by its buﬀering function.
Intravitally, proteoglycan is produced in the Golgi
body, by adding a number of sugar chains to the coreprotein (i.e. glycosylation) 11). On the other hand, theusage of proteoglycan for food and cosmetic product has
started after the technological establishment of extracting
it from the nasal cartilage of salmon. Although themechanism underlying proteoglycan absorption in theintestine is not clear, the recent study showed even by

the oral ingestion, it is directly absorbed in the intestineby the intermediary of clathrin-mediated endocytosis12).
Historically, for the maintenance of joint function,
glucosamine and chondroitin sulfate (both of which are
the components of proteoglycan) have been used as asupplement in expectation of their production of cartilage
matrix intravitally 7) 8). Glucosamine reportedly owns
functions such as protection of cartilage and antiinﬂammatory power for cartilage13), and an oral ingestionof chondroitin, on the other hand, has a functionality ofalleviating arthritic pain6) 14). Since proteoglycan is directly
absorbed in the intestine, it can be expected that thefunctionality from the mechanism totally different fromthe ingestion of its components should be achieved.
There is a study which reports proteoglycan owns
growth-stimulating eﬀect and diﬀerentiation- maintainingof cartilage cells 15) , and this study encourages the
expectation that proteoglycan can improve the conditionof knee-joint by regeneration and protection against
scuﬃng of cartilage. In addition, another study illustrates
the anti-inflammatory power of proteoglycan 16)17) ,therefore it suggests proteoglycan achieves a suppressionof inﬂammation, which is a cause of pain.
In this study, the improvement effect of knee-joints
was mainly seen in the situations when the subjects
experienced a low-impact on their knees, and this ﬁndingis suggested by the results such as VAS of in repose or
J-KOOS of walking on flat surface . It is thought that
this finding has some relations to the function ofproteoglycan as explained above. On the other hand, thesituations such as up/down stairs and sports when
the subjects experienced a relatively high-impact did notshow the improvements; this result indicates that the test
product may gradually improve the condition of kneejoints and therefore the longer ingestion period isrecommended.
Based upon the above discussion, it is suggested that inthis study, the ingestion of the test product containing
proteoglycan showed a trend toward improvement in thecondition of knee-joints.
During the test period five subjects discontinued thetest. Four of them stopped the test because of theirpersonal reasons such as impossibility of continuing thetest due to their business or diseases (i.e. cold). As forthe remaining one subject in the placebo group, on theother hand, the measurement after 12-week ingestion

診療と新薬・第 53 巻

showed an abnormal value of CK, but after the medical
interview the doctor determined the abnormity was dueto the tendovaginitis and ruled out any relationship withthe test itself. Therefore, based upon the medical
interview, blood test and urine test, we observed noharmful influence against biochemical and/or
physiological matters of the subjects which seem to havecausal relationship with LA.
These results indicated the safety of the ingestion of
the test product (LA) for the 12-week test period.
In Japan, as the society faces aging of population, it is
now a big issue to prevent the elderly people from theloss of self-support. The fourth cause for the elderly careis an articular disease. There is also a report thatprevalence rate of OA increases with advancing age18).
The progression of its symptom makes it more diﬃcult forthe patients to walk. However, a full-scale treatment for
OA has not yet been established and this fact strongly
supports the importance of the prevention of symptom.
Although it is inevitable for the articular cartilage to bechanged with aging, achieving the improvement of theirknee-joint condition by the ingestion of foods may
contribute to the self-support of elderly people and to theprevention of the degradation of QOL.
Until several years ago it was difficult to use
proteoglycan as a food because of its high price, but after
the establishment of extraction method of it from salmon
it became possible to use it as a food. Therefore, its
functionality is worth noting from now on.
However, the study of proteoglycan was mainly based
upon the in-vitro settings or the research using animals,
and the functional mechanism in the human body has not
yet been deeply speculated so far. There is also a reportthat concludes the ingestion of glucosamine andchondroitin sulfate (both of which are the components ofproteoglycan) is not related to the improvement of joint
pain12).
In addition, since articular cartilage does not have
many blood vessels, it is recognized that its capacity forrepair is limited19). It is not clear that the improvementeﬀect of knee-joints we have observed in this study waseither the result of inﬂammatory suppression or becauseof the repair of articular cartilage, and its functional
mechanism is a matter of speculation. These points
should be further scrutinized in the future.
In conclusion, we found out that the ingestion of the
supplement LA containing proteoglycan by Japanese
healthy people feeling knee s discomfort for 12 weeks

第 3 号（2016 年 3 月）

71 （235）

contributed to improving the joint support. In addition,no safety-related matter occurred during 12-week test
period.
REFERENCES
1）Loeser RF Jr. Aging cartilage and osteoarthritis--whatʼs the link? Sci
Aging Knowledge Environ. 2004; 2004(29): pe31.
2）Loeser RF Jr. Aging and the etiopathogenesis and treatment of
osteoarthritis. Rheum Dis Clin North Am. 2000; 26(3): 547-67.
3）Ding C, Cicuttini F, Blizzard L, et al. A longitudinal study of theeﬀect of sex and age on rate of change in knee cartilage volume inadults. Rheumatology (Oxford). 2007; 46(2): 273-9.
4）Verzijl N, DeGroot J, Thorpe SR, et al. Eﬀect of collagen turnover on
the accumulation of advanced glycation end products. J Biol Chem.
2000; 275(50): 39027-31.
5）Majima M, Takagaki K, Sudo S, et al. Effect of proteoglycan on
experimental colitis. International Congress Series. 2001; 1223: 2214.
6）Nakamura H. Arthralgia and drug, supplement. Juntendo Medical
Journal.2008; 54(3): 372-6.
7）McAlindon TE, LaValley MP, Gulin JP, et al. Glucosamine andchondroitin for treatment of osteoarthritis: a systematic quality
assessment and meta-analysis. JAMA. 2000; 283(11): 1469-75.
8）Gokan N, Suzuki N, Shiizuka K, et al. The Effect of the Dietary
Supplement Containing with Both Glucosamine and Chndoritin
Sulfate on Knee Joint Pain. J.New Rem. & Clin. 2011; 60(7): 147682.
9）Nakamura N, Takeuchi R, Sawaguchi T, et al. Cross-cultural
adaptation and validation of the Japanese Knee Injury and
Osteoarthritis Outcome Score (KOOS). J Orthop Sci. 2011; 16(5):516-23.
10）Aitken RC. Measurement of feelings using visual analogue scales.
Proc R Soc Med. 1969; 62(10): 989-93.
11）Watanabe H. Agrecan and its chondroitin sulfate in cartilage.
Biochem. 2008; 80(1): 28-31.
12）Tsuchiya Y, Tomita M, Tsuboi M, et al. Absorption of Proteoglycan
via Clathrin-Mediated Endocytosis in the Small Intestine of Rats.
Biosci Biotechnol Biochem. 2013; 77(3): 654-6.
13）Nagaoka I. Chondroprotective, anti-inflammatory and anti-platelet
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Habits.2008; 19(1): 3-8.
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Sulfate, and the Two in Combination for Painful Knee Osteoarthritis.
N Engl J Med. 2006; 354(8): 795-808.
15）Ohshika S, Kon A, Ishibashi Y, et al. Effect of cartilage-derived
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16）Merline R, Moreth K, Beckmann J, et al. Signaling by the matrix
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72 （236）

診療と新薬・第 53 巻

Appendix<Modiﬁed J-KOOS>
Stiﬀness
The following questions concern the amount of joint stiffness you haveexperienced during the last week in your knee. Stiﬀness is a sensation of
restriction or slowness in the ease with which you move your knee joint.
S1. How severe is your knee joint stiﬀness after ﬁrst wakening in
the morning?
None, Mild, Moderate, Severe, Extreme
S2. How severe is your knee stiﬀness after sitting, lying or restinglater in the day?
None, Mild, Moderate, Severe, Extreme
Pain
P1. How often do you experience knee pain?
None, Mild, Moderate, Severe, Extreme
What amount of knee pain have you experienced the last week during thefollowing activities?
P2. Walking on ﬂat surface
None, Mild, Moderate, Severe, Extreme
P3. Going up or down stairs
None, Mild, Moderate, Severe, Extreme
P4. At night while in bed
None, Mild, Moderate, Severe, Extreme
P5. Standing uprightNone, Mild, Moderate, Severe, Extreme
Function, daily living
The following questions concern your physical function. By this we mean
your ability to move around and to look after yourself. For each of thefollowing activities please indicate the degree of difficulty you haveexperienced in the last week due to your knee.
A1. Descending stairs
None, Mild, Moderate, Severe, Extreme
A2. Ascending stairs
None, Mild, Moderate, Severe, Extreme
For each of the following activities please indicate the degree of diﬃculty
you have experienced in the last week due to your knee.

第 3 号（2016 年 3 月）
A3. Rising from sitting
None, Mild, Moderate, Severe, Extreme
A4. Standing
None, Mild, Moderate, Severe, Extreme
A5. Bending to ﬂoor/ picking up an object
None, Mild, Moderate, Severe, Extreme
A6. Walking on ﬂat surface
None, Mild, Moderate, Severe, Extreme
A7. Getting in/ out of car
None, Mild, Moderate, Severe, Extreme
A8. Going shopping
None, Mild, Moderate, Severe, Extreme
A9. Putting on/ oﬀ socks/ stockings
None, Mild, Moderate, Severe, Extreme
A10. Lying in bed (turning over, maintaining knee position)
None, Mild, Moderate, Severe, Extreme
A11. Getting in/ out of bath
None, Mild, Moderate, Severe, Extreme
A12. Sitting
None, Mild, Moderate, Severe, Extreme
A13. Getting on/ oﬀ toilet
None, Mild, Moderate, Severe, Extreme
For each of the following activities please indicate the degree of diﬃculty
you have experienced in the last week due to your knee.
A14. Heavy domestic duties (moving heavy boxes, scrubbing ﬂoors,etc.)
None, Mild, Moderate, Severe, Extreme
A15. Activities such as sports, jogging, swimming, gateball, dancing,etc.
None, Mild, Moderate, Severe, Extreme
Quality of Life
Q1. Due to pain, do you restrict your hobbies or recreational
activities?
Not at all, Mildly, Moderately, Severely, TotallyQ2. Due to pain, do you stop going out?
Not at all, Mildly, Moderately, Severely, Totally
Q3. Is your overall health condition aﬀected by your knee condition?
Not at all, Mildly, Moderately, Severely, Totally