Response to the Reviewers
Thank you for your comments and advices. We have revised the manuscript and responded to your
comments as listed below:
Response to Editor-in-Chief:
General comments:
The study addresses an important question and has the potential of enhancing our knowledge in the
field. The concerns raised and suggestions given by the reviewers seem important for improvement
of the manuscript.
Response: Thank you very much for your kindly comments. This study was to evaluate the
long-term effect of treadmill exercise on bone mass and articular cartilage. We hope that it can
address an important issue and the findings of our study can help enhance related knowledge in the
field.
Specific comments:
1. The authors need to clarify if the data were analyzed by blinded personnel. This is of particular
importance for the assessment of the histological preparations.
Response: We agree that the data were analyzed by blinded personnel is very important because the
individual bias will influence the results. In order to eliminate such bias, two authors involved in
this study and they have been blinded to group distribution in data analysis. This description was
shown in the Methods section (page 6 at the end of first paragraph) as “The histological evaluation
was performed by two of the coauthors (Chang and Chen) who were blinded to group distribution
when the analysis was made.” In addition, we also describe the crucial issue in the Discussion
section on page 14 as “Finally, the scoring of articular damage may be subjective and have an
individual bias; therefore, this analysis was carried out by two independent observers to ensure
repeatability and consistency.”
2. In the description of the statistical analysis, both the authors state that both t-tests and
non-parametric are used and for the same purpose. Data are given as mean and SD values,
suggesting that data are normally distributed. Please clarify your statistical approaches, and if
different methods are used, the method used in the different analyses should be specified.
Response: Thank you very much for your kindly mention. The Mann-Whitney U-test was chosen
for data analysis. The description of statistical method was modified as following. “The results of
this study were expressed as the mean values and standard deviations of the different continuous
variables. The Mann-Whitney U-test was used to analyze the results between different groups. A P
value of less than 0.05 was considered to be a significant difference.” The sentence showed on the
end of Method section.
3. A small detail: Correct the number of digits used for your SD values in the abstract to match the
numbers used for mean values.
Response: We have corrected the number of digits used for SD values in the abstract to match the
numbers used for mean values. The modification was appeared on page1, line19 as “….Mankin
score (7.7±1.4) compared to OVX-RUN group (4.8±1.0)…”.
Response to editorial:
1. Experimental research that is reported in the manuscript must have been performed with the
approval of an appropriate ethics committee. Research carried out on humans must be in
compliance with the Helsinki Declaration, and any experimental research on animals must follow
internationally recognized guidelines. A statement to this effect must appear in the Methods section
of the manuscript, including the name of the body which gave approval, with a reference number
where appropriate.
Response: This animal study has been approved by the Institutional Animal Care and Use
Committee (IACUC) of Mackay Memorial Hospital, Taipei, Taiwan. This claim has been added in
the Animals & Exercise Program subtitle of Method section and the approval is shown below.
2. Regarding the editorial comments, including manuscript format and competing interests
Response: The manuscript format has been rechecked with the suggestion of the esteemed journal
of BMC musculoskeletal disorders. In addition, we declare that we have no competing interests.
And this research grant was supported by the National Science Council, Taiwan, ROC. Above
statement was also shown on page 14-15.
My co-authors and I thank you again for your substantial effort to review and comment on this
manuscript. We are very pleased with the revised manuscript and hope that you will agree that your
reviews have helped us to refine our work into an important and interesting article.
Response to the Reviewers
Thank you for your comments and advices. We have revised the manuscript and responded to your
comments as listed below:
Reviewer #1:
General comments:
Chang et al. investigated the influence of long-term treadmill exercise on changes in bone mass and
articular cartilage in ovariectomized rats. They suggested that long-term exercise had a beneficial
effect on articular cartilage for the rats after ovariectomy, but appeared to have few improvements
on bone mass. This study seems to be interesting because the effect of long-term exercise on bone
and articular cartilage was examined.
Response: Thank you very much for your kindly comments. We believe the result of current study
would help address the important issue of the effect of long-term exercise on musculoskeletal
system. Reviewer’s other concerns were responded based on point-by-point rule as following.
Specific comments:
1. Although the authors obtained a positive result regarding articular cartilage in ovariectomized rats,
the assessment of BMD and bone mass may not be enough to conclude non significant effect of
exercise on bone tissue.
Response: Thank you for your advices. The purpose of this study was mainly focused on observing
the changes of articular cartilage and bone mass by giving a related long-term exercise program,
which has not been clearly addressed till now. In the current study, we observed moderate exercise
seems to provide a positive role of chondroprotective effect in ovariectomized rats. In addition to
BMD and bone mass, we also found that the BV/TV ratio in the OVX-RUN group is slightly
increased about 11.6% than that of the OVX group. But, no strongly effects were found on these
bony changes by determining current analyzed parameters. Further detailed studies would be
helpful to evaluate the long-term exercise on the effects of bone tissues. We have taken this point as
the limitation of our study. Description has been added on the end of page 13, as “……Thirdly,
current study found that moderate exercise seems to play a positive role on chondroprotective effect
in ovariectomized rats but seems no obviously changes on the bone tissue. Except for slightly
increase of the ratio of BV/TV (11.6%) was found in the OVX-RUN group when compared to the
OVX group. Further detailed assessments will be helpful to justify the realistic mechanism of
exercise on bone mass.…….”
2. Why didn’t the authors examine femoral or tibial BMD as well as lumbar spine BMD? There are
a lot of papers showing the positive effect of treadmill running exercise on femoral or tibial BMD
and bone mass.
Response: Both femoral and spine BMD were examined in the study. However, no significant
difference was found in the mean value between spinal BMD and femoral BMD. (Please see the
figure below). Furthermore, variation of bone density in femoral BMD was somewhat higher than
that of spinal BMD (Standard deviations in each group of femoral BMD are higher than those of
each group of spinal BMD). The possible reason resulting in a higher variation in the femoral BMD
would be owing to smaller dimension of femur. Therefore, we merely reported lumbar BMD here.
This was also considered as a limitation of this study and the description was added in the
discussion section as “We reported lumbar BMD instead of the distal femur or the proximal tibia
because higher variation of BMD was found on the femur than that of spine. The possible reason
resulting in higher variation for femoral BMD might be smaller dimension of femoral bone. Central
skeleton (Lumbar BMD) would eliminate the variation. In addition, the data of trabecular bone
mass further proved that relative osteoporosis was seen after ovariectomy, which also correlates
with the BMD result of spine.”
Higher variations are observed in each group of femoral BMD when comparing to those of spinal
group.
3. Please show initial and final body weight, and also show some parameters related to bone size
(growth), because the authors used young growing rats. Muscle weight and body composition are
also important parameters in this study.
Response: The initial body weight of experimental rats was 229 ± 9.0 g. Sixteen weeks later, the
mean body weight was 418.7 ± 58.8 g in the ovariectomized group (OVX, OVX-RUN), while in the
control group (CON, RUN) was 347.1 ± 30.9 g (**P<0.05). The final body weight in RUN and
OVX-RUN groups was 394.3 ± 55.8 g and 451.1 ± 74.0 g respectively, while the CON and OVX
groups was 430.0 ± 47.0 g and 568.3 ± 132.4 g in the end of the study. The figure was shown below.
We agree with the reviewer’s opinion that muscle weight and body composition are important
parameters in musculoskeletal study. Since the main purpose of our study is to evaluate the
influence of exercise on articular cartilage, we didn’t include muscle weight and body composition
in this study.
4. In figures, the order of bars should be CON, RUN, OVX, and OVX-RUN from the left side.
Please show the results of statistical analysis in the figure or figure legend.
Response: The order of bars was changed as the reviewer’s suggestion in Fig. 2, 3, and 5. The
results of statistical analysis also added in each figure and legend. Thank you again for your
recommendation.
5. The method of trabecular BV/TV measurement may not be adequately described. Please detail it.
Response: The histomophometric parameters (BV/TV) were measured according to Parfitt et al’s
method. (Parfitt AM et al.: Bone histomorphometry: standardization of nomenclature, symbols, and
units. Report of the ASBMR Histomorphometry Nomenclature Committee. J Bone Miner Res 1987,
2(6):595-610.). The definition of BV indicates as a two dimensional manner of bone volume
(referring to Parfitt et al); and definition of TV indicates tissue volume. Thank you for your kindly
suggestions. We then add this reference and give more description on page 6 as “.
Histomorphometric measurements of the cancellous bone of the proximal tibia were performed
semi-automatically with an Olympus BX 40 microscope and an Olympus DP-70 digital camera
(Olympus, Tokyo, Japan). Data were further analyzed by a computer with commercially available
software (Multi Gauge v2.1, Fuji film Co., Tokyo, Japan). The trabecular bone volume (BV/TV) in
the metaphysis of the proximal tibia was then measured according to Parfitt’s method [17]. The
definition of BV indicates bone volume (it was simplified to a two-dimensional manner [4, 17] );
and definition of TV indicates tissue volume.”
6. Please refer the intensity of exercise. Was it mild or moderate?
Response: The intensity of current exercise program can be considered as moderate level. Similar
treadmill exercise has also been applied for animal study and intensity is defined as “moderate”.
(Iwamoto J, Takeda T, Ichimura S: Effect of exercise on tibial and lumbar vertebral bone mass in
mature osteopenic rats: bone histomorphometry study. J Orthop Sci 1998, 3(5):257-263.). We have
added this reference and more description into the manuscript in the Method section on page 4.
Thank you for your question.
7. Bone formation and resorption parameters need to be shown, if they were measured.
Response: We didn’t measure these parameters in current study. We believe that related parameters
of bone formation and resorption are helpful to clarify the influence of exercise on bone tissue in
musculoskeletal disorders. We will plan to do more detailed measurement in future work.
8. In the abstract, results, and discussion, the following sentence needs to be revised: The results
showed that rats in groups without ovariectomy (CON and RUN) have significant higher BMD and
bone mass than in the groups with ovariectomy (OVX and OVX-RUN). This may be right: The
results showed that rats in groups with ovariectomy (OVX and OVX-RUN) had significant lower
BMD and bone mass than in the groups without ovariectomy (CON and RUN).
Response:. The modification was done and listed on Page 1. Thank you for your suggestion.
9. English needs to be corrected by native speakers.
Response: English writing throughout the manuscript has been corrected by a native speaker and
the final text has also been read and carefully rechecked by coauthors.
My co-authors and I thank you again for your substantial effort to review and comment on this
manuscript. We are very pleased with the revised manuscript and hope that you will agree that your
reviews have helped us to refine our work into an important and interesting article.
Response to the Reviewers
Thank you for your comments and advices. We have revised the manuscript and responded to your
comments as listed below:
Reviewer #2:
General comments:
The manuscript by Chang et al. aims at investigation of the influence of long-term treadmill
exercise on the changes of bone mass and articular cartilage in ovariectomized rats. By evaluating
changes in bone mass and articular cartilage with (1) trabecular bone volume and bone mineral
density and (2) histology analysis and a modified Mankin scoring method, respectively, in different
groups of rats with different treatments, the authors claimed that long-term exercise may exert
beneficial effects on articular cartilage for the rats with ovariectomy, but appears to have few
improvements on the bone mass in the rats after ovariectomy. This is an interesting study, and the
manuscript is well written. Since the loss of bone quality and change in articular cartilage are
commonly events after menopause, the present study has provided data for the understanding of the
role of running exercise in the changes in bone mass and articular cartilage in subjects after
menopause. The manuscript will be significantly improved by the additional data or more detailed
discussions on whether running exercise can influence changes in the concentrations of certain
hormones in animals by ovariectomy, which consequently leads to changes in articular cartilage
rather than bone mass. This will strengthen the mechanistic insight of the study.
Response: Thank you for your kindly comments. We believe the present study can help provide
interesting and important information about the role of running exercise on the changes of articular
cartilage in rats after menopause. We have added more explanation about the effect on articular
cartilage and hormone changes in animals after ovariectomy in Discussion section on the page 12,
second paragraph as “Cartilage degeneration after ovariectomy has been studied extensively before.
Sniekers et al. used a systemic approach to review studies regarding the osteoarthritic changes after
ovariectomy and the effect of estrogen treatment [23]. They found that 11 out of 14 studies showed
a detrimental effect of ovariectomy on articular cartilage in sexually mature animal. However, the
beneficial effect of estrogen therapy on cartilage was inconclusive in this review paper. They
suggested that OVX-related changes in hormones (estrogen, progesterone, follicle-stimulating
hormone, or luteinizing hormone) may play a role in the articular damage seen after ovariectomy,
and this may explain why estrogen treatment could not always reduce the OVX-induced changes. In
the current study, cartilage degenerative change was more severe in the OVX group than other
groups. The real mechanism of cartilage degeneration following an ovariectomy is complex and
multi-factorial, and further investigation is needed.”
My co-authors and I thank you again for your substantial effort to review and comment on this
manuscript. We are very pleased with the revised manuscript and hope that you will agree that your
reviews have helped us to refine our work into an important and interesting article.