Reviewer's report
 TNF-a induced endothelial MAdCAM-1 expression is regulated by exogenous, not endogenous Title:
nitric oxide
Authors:
Tadayuki Oshima (toshim@lsumc.edu )
Paul Jordan (pjorda@lsumc.edu )
Matthew B Grisham (mgrish@lsumc.edu )
Jonathan S Alexander (jalexa@lsumc.edu )
Merilyn Jennings (mjenni@lsumc.edu )
Makotoh Sasaki (msasak@lsumc.edu )
 3  18 Jun 2001 Version: Date:
 Dr J Panes Reviewer:
 A paper whose findings are important to those with closely related research interests Level of interest:
 Accept after revision, which I do not need to see Advice on publication:
In the current study Oshima et al. show that upregulation of MAdCAM-1 in response to TNF stimulation 
is inhibited by NO-donors. This inhibition is associated to a reduction in a4b7-dependent leukocyte 
adhesion. This is an original, well performed study, that adds significant information to our current knowledge on the regulation of adhesion molecules expression. The conclusions drawn adequately supported by the data shown, and the manuscript adheres to the 
relevant standards for reporting and data deposition. 
I have the following points for the authors' consideration: 
1. In the aims section the authors state that they examine time and concentration-dependent induction 
of MAdCAM-1 in response to TNF-a. I could not find in the manuscript data on this point, it seems they always used 20 ng/ml TNF-a. The authors should either provide such data or remove this part of the aims statement. 
2. The authors state in the abstract that experiments measuring MAdCAM-1 protein expression 
following TNF-a stimulation were performed at 24 hours. This should be also specified in the methods/results section of the manuscript. 3. In the experiments with NO synthesis inhibitors, for how log were cells exposed to L-NAME? 
4. The different ability of Spermine-NO and DETA-NO to inhibit MAdCAM-1 upregulation is most 
probably related to differences in half life of these compounds (230 min vs. 56 hours). Therefore, in experiments performed at 24 hours a significant amount of NO was probably present only during the first few hours after Spermine-NO supplementation. This should be more clearly considered in the 
discussion. 
5. The possibility exists that reduction of TNF-a-induced leukocyte adhesion in response to NO donors is also related to modulation of expression of VCAM-1. This should be considered in the discussion. Direct exploration of this possibility would improve the manuscript. 
6. The authors suggest the possibility that MAdCAM-1 might be upregulated in human IBD. This has 
been already proved (Briskin et al. Am J Pathol 1997 Jul;151:97-110, Souza et al. Gut 1999 
Dec;45:856-63). 
7. In the discussion, the paragraph beginning with "In this model, the mechanism...." seems to lack 
some text lines towards the end. 8. In Figure 2, the paragraph relating part C should be corrected. 9. The manuscript contains numerous typos that need correction. 
Competing interests:
None declared.
Reviewer's report
 TNF-a induced endothelial MAdCAM-1 expression is regulated by exogenous, not endogenous Title:
nitric oxide
Authors:
Tadayuki Oshima (toshim@lsumc.edu )
Paul Jordan (pjorda@lsumc.edu )
Matthew B Grisham (mgrish@lsumc.edu )
Jonathan S Alexander (jalexa@lsumc.edu )
Merilyn Jennings (mjenni@lsumc.edu )
Makotoh Sasaki (msasak@lsumc.edu )
 3  18 Jun 2001 Version: Date:
 Dr J Panes Reviewer:
 A paper whose findings are important to those with closely related research interests Level of interest:
 Accept after revision, which I do not need to see Advice on publication:
In the current study Oshima et al. show that upregulation of MAdCAM-1 in response to TNF stimulation 
is inhibited by NO-donors. This inhibition is associated to a reduction in a4b7-dependent leukocyte 
adhesion. This is an original, well performed study, that adds significant information to our current knowledge on the regulation of adhesion molecules expression. The conclusions drawn adequately supported by the data shown, and the manuscript adheres to the 
relevant standards for reporting and data deposition. 
I have the following points for the authors' consideration: 
1. In the aims section the authors state that they examine time and concentration-dependent induction 
of MAdCAM-1 in response to TNF-a. I could not find in the manuscript data on this point, it seems they always used 20 ng/ml TNF-a. The authors should either provide such data or remove this part of the aims statement. 
2. The authors state in the abstract that experiments measuring MAdCAM-1 protein expression 
following TNF-a stimulation were performed at 24 hours. This should be also specified in the methods/results section of the manuscript. 3. In the experiments with NO synthesis inhibitors, for how log were cells exposed to L-NAME? 
4. The different ability of Spermine-NO and DETA-NO to inhibit MAdCAM-1 upregulation is most 
probably related to differences in half life of these compounds (230 min vs. 56 hours). Therefore, in experiments performed at 24 hours a significant amount of NO was probably present only during the first few hours after Spermine-NO supplementation. This should be more clearly considered in the 
discussion. 
5. The possibility exists that reduction of TNF-a-induced leukocyte adhesion in response to NO donors is also related to modulation of expression of VCAM-1. This should be considered in the discussion. Direct exploration of this possibility would improve the manuscript. 
6. The authors suggest the possibility that MAdCAM-1 might be upregulated in human IBD. This has 
been already proved (Briskin et al. Am J Pathol 1997 Jul;151:97-110, Souza et al. Gut 1999 
Dec;45:856-63). 
7. In the discussion, the paragraph beginning with "In this model, the mechanism...." seems to lack 
some text lines towards the end. 8. In Figure 2, the paragraph relating part C should be corrected. 9. The manuscript contains numerous typos that need correction. 
Competing interests:
None declared.
Please review this paper.
user
Reviewer's report
 TNF-a induced endothelial MAdCAM-1 expression is regulated by exogenous, not endogenous Title:
nitric oxide
Authors:
Tadayuki Oshima (toshim@lsumc.edu )
Paul Jordan (pjorda@lsumc.edu )
Matthew B Grisham (mgrish@lsumc.edu )
Jonathan S Alexander (jalexa@lsumc.edu )
Merilyn Jennings (mjenni@lsumc.edu )
Makotoh Sasaki (msasak@lsumc.edu )
 3  18 Jun 2001 Version: Date:
 Dr J Panes Reviewer:
 A paper whose findings are important to those with closely related research interests Level of interest:
 Accept after revision, which I do not need to see Advice on publication:
In the current study Oshima et al. show that upregulation of MAdCAM-1 in response to TNF stimulation 
is inhibited by NO-donors. This inhibition is associated to a reduction in a4b7-dependent leukocyte 
adhesion. This is an original, well performed study, that adds significant information to our current knowledge on the regulation of adhesion molecules expression. The conclusions drawn adequately supported by the data shown, and the manuscript adheres to the 
relevant standards for reporting and data deposition. 
I have the following points for the authors' consideration: 
1. In the aims section the authors state that they examine time and concentration-dependent induction 
of MAdCAM-1 in response to TNF-a. I could not find in the manuscript data on this point, it seems they always used 20 ng/ml TNF-a. The authors should either provide such data or remove this part of the aims statement. 
2. The authors state in the abstract that experiments measuring MAdCAM-1 protein expression 
following TNF-a stimulation were performed at 24 hours. This should be also specified in the methods/results section of the manuscript. 3. In the experiments with NO synthesis inhibitors, for how log were cells exposed to L-NAME? 
4. The different ability of Spermine-NO and DETA-NO to inhibit MAdCAM-1 upregulation is most 
probably related to differences in half life of these compounds (230 min vs. 56 hours). Therefore, in experiments performed at 24 hours a significant amount of NO was probably present only during the first few hours after Spermine-NO supplementation. This should be more clearly considered in the 
discussion. 
5. The possibility exists that reduction of TNF-a-induced leukocyte adhesion in response to NO donors is also related to modulation of expression of VCAM-1. This should be considered in the discussion. Direct exploration of this possibility would improve the manuscript. 
6. The authors suggest the possibility that MAdCAM-1 might be upregulated in human IBD. This has 
been already proved (Briskin et al. Am J Pathol 1997 Jul;151:97-110, Souza et al. Gut 1999 
Dec;45:856-63). 
7. In the discussion, the paragraph beginning with "In this model, the mechanism...." seems to lack 
some text lines towards the end. 8. In Figure 2, the paragraph relating part C should be corrected. 9. The manuscript contains numerous typos that need correction. 
Competing interests:
None declared.
assistant