Reviewer's report 
 Integrative roles of transforming growth factor-alpha in the cytoprotection mechanisms of Title: 
gastric mucosal injury 
 1  20 February 2006 Version: Date: 
 Peter Konturek Reviewer: 
Reviewer's report: 
Manuscript by Kosone T et al entitled " Integrative roles of transforming growth factor alpha in the 
cytoprotection mechanism of gastric mucosal injury" deals with the gastroprotective role of TGF , 
which was studied in transgenic mice overexpressing TGF and in gastric mucosa-derived cell line 
treated with TGF . 
The overexpression of TGF was associated with an increased resistance to ethanol injury and this 
effect was due partly to: increased gastric blood flow, increased expression of constitutive NO synthase, COX-2 expression and anti-apoptotic factors (Bcl-2; Hsp70), and reduction in proapoptotic 
caspase-3. In the in vitro experiments, gastric mucosa derived cells were treated with TGF 
(5-100ng/ml) in the presence of of three different MAP kinase inhibitors. Similary to the experiments in animals, TGF treatmeant caused an upregulation of HSP70, COX-2 and Bcl-2 expression. Both inhibitors, PD98059 for ERK1/2 and SB203580 for p38 inhibited the expression of HSP70 and 
COX-2. In contrast, SP600125 for JNK1/2 enhance the expression of COX-2, but inhibited HSP70 
expression. 
Major points of criticism: 
1. It is surprising for me that the transgenic animals overexpressing TGF do not show hypertrophy of 
the gastric mucosa like in the Menetriere disease? How can be this phenomenon explained ??? (see 
paper by Takagi JCI 1992). 
2. The role of TGF as a protective factor in the gastric mucosa is well established (see Gangarosa 
LM et al; Baillieres Clin Gastroenterol 1996; 10:49-63) and all the results obtained in the present study are mostly the confirmation of the previous studies. I am missing an originality of this study. The difference to the previous studies is the use of transgene animals overexpressing TGF instead 
of application of exogenous TGF as used before. 
3. Since NFkappaB plays a crucial role in the transcriptional regulation of pro-survival genes and there is an evidence for interaction between TGF and NFkappaB ( Kanai et al Gastroenterology 
2001; 121: 56-67). It would be of great importance to check this aspect in TGFalpha transgenic mice. 
Authors should compare the activation of NFkappaB in the gastric mucosa of wild animals and 
transgene mice overexpressing TGFa. This would be a new and original finding instead of repetition 
of previous results. 4. It is well known that TGF has antisecretory affects. However, there was no difference in 
intragastric pH between WT animals and TG animals. I strongly suggest the measurement of gastric 
acid secretion in mice equipped with special gastric fistula. 
5. In addition to analysis of COX-2 expression, the authors should measure prostaglandin generation 
in both animals groups (WT vs. TG). 
6. Since gastrin play a crucial role in the regulation of gastric secretion and also shows 
gastroprotective effects, it would be of importance to measure gastrin in both groups. 
7. Although the discussion and conclusions are well balanced and adequately supported by the data, 
the whole manuscript is too long, and the in vitro data should be omitted. Discussion should be 
shortened. 
 Unable to decide on acceptance or rejection until the authors have responded to the What next?: 
major compulsory revisions 
 An article whose findings are important to those with closely related research Level of interest: 
interests 
 Needs some language corrections before being published Quality of written English: 
 No Statistical review: 
Declaration of competing interests: 
I declare that I have no competing interests 
Reviewer's report 
 Integrative roles of transforming growth factor-alpha in the cytoprotection mechanisms of Title: 
gastric mucosal injury 
 1  20 February 2006 Version: Date: 
 Peter Konturek Reviewer: 
Reviewer's report: 
Manuscript by Kosone T et al entitled " Integrative roles of transforming growth factor alpha in the 
cytoprotection mechanism of gastric mucosal injury" deals with the gastroprotective role of TGF , 
which was studied in transgenic mice overexpressing TGF and in gastric mucosa-derived cell line 
treated with TGF . 
The overexpression of TGF was associated with an increased resistance to ethanol injury and this 
effect was due partly to: increased gastric blood flow, increased expression of constitutive NO synthase, COX-2 expression and anti-apoptotic factors (Bcl-2; Hsp70), and reduction in proapoptotic 
caspase-3. In the in vitro experiments, gastric mucosa derived cells were treated with TGF 
(5-100ng/ml) in the presence of of three different MAP kinase inhibitors. Similary to the experiments in animals, TGF treatmeant caused an upregulation of HSP70, COX-2 and Bcl-2 expression. Both inhibitors, PD98059 for ERK1/2 and SB203580 for p38 inhibited the expression of HSP70 and 
COX-2. In contrast, SP600125 for JNK1/2 enhance the expression of COX-2, but inhibited HSP70 
expression. 
Major points of criticism: 
1. It is surprising for me that the transgenic animals overexpressing TGF do not show hypertrophy of 
the gastric mucosa like in the Menetriere disease? How can be this phenomenon explained ??? (see 
paper by Takagi JCI 1992). 
2. The role of TGF as a protective factor in the gastric mucosa is well established (see Gangarosa 
LM et al; Baillieres Clin Gastroenterol 1996; 10:49-63) and all the results obtained in the present study are mostly the confirmation of the previous studies. I am missing an originality of this study. The difference to the previous studies is the use of transgene animals overexpressing TGF instead 
of application of exogenous TGF as used before. 
3. Since NFkappaB plays a crucial role in the transcriptional regulation of pro-survival genes and there is an evidence for interaction between TGF and NFkappaB ( Kanai et al Gastroenterology 
2001; 121: 56-67). It would be of great importance to check this aspect in TGFalpha transgenic mice. 
Authors should compare the activation of NFkappaB in the gastric mucosa of wild animals and 
transgene mice overexpressing TGFa. This would be a new and original finding instead of repetition 
of previous results. 4. It is well known that TGF has antisecretory affects. However, there was no difference in 
intragastric pH between WT animals and TG animals. I strongly suggest the measurement of gastric 
acid secretion in mice equipped with special gastric fistula. 
5. In addition to analysis of COX-2 expression, the authors should measure prostaglandin generation 
in both animals groups (WT vs. TG). 
6. Since gastrin play a crucial role in the regulation of gastric secretion and also shows 
gastroprotective effects, it would be of importance to measure gastrin in both groups. 
7. Although the discussion and conclusions are well balanced and adequately supported by the data, 
the whole manuscript is too long, and the in vitro data should be omitted. Discussion should be 
shortened. 
 Unable to decide on acceptance or rejection until the authors have responded to the What next?: 
major compulsory revisions 
 An article whose findings are important to those with closely related research Level of interest: 
interests 
 Needs some language corrections before being published Quality of written English: 
 No Statistical review: 
Declaration of competing interests: 
I declare that I have no competing interests 
Please review this paper.
user
Reviewer's report 
 Integrative roles of transforming growth factor-alpha in the cytoprotection mechanisms of Title: 
gastric mucosal injury 
 1  20 February 2006 Version: Date: 
 Peter Konturek Reviewer: 
Reviewer's report: 
Manuscript by Kosone T et al entitled " Integrative roles of transforming growth factor alpha in the 
cytoprotection mechanism of gastric mucosal injury" deals with the gastroprotective role of TGF , 
which was studied in transgenic mice overexpressing TGF and in gastric mucosa-derived cell line 
treated with TGF . 
The overexpression of TGF was associated with an increased resistance to ethanol injury and this 
effect was due partly to: increased gastric blood flow, increased expression of constitutive NO synthase, COX-2 expression and anti-apoptotic factors (Bcl-2; Hsp70), and reduction in proapoptotic 
caspase-3. In the in vitro experiments, gastric mucosa derived cells were treated with TGF 
(5-100ng/ml) in the presence of of three different MAP kinase inhibitors. Similary to the experiments in animals, TGF treatmeant caused an upregulation of HSP70, COX-2 and Bcl-2 expression. Both inhibitors, PD98059 for ERK1/2 and SB203580 for p38 inhibited the expression of HSP70 and 
COX-2. In contrast, SP600125 for JNK1/2 enhance the expression of COX-2, but inhibited HSP70 
expression. 
Major points of criticism: 
1. It is surprising for me that the transgenic animals overexpressing TGF do not show hypertrophy of 
the gastric mucosa like in the Menetriere disease? How can be this phenomenon explained ??? (see 
paper by Takagi JCI 1992). 
2. The role of TGF as a protective factor in the gastric mucosa is well established (see Gangarosa 
LM et al; Baillieres Clin Gastroenterol 1996; 10:49-63) and all the results obtained in the present study are mostly the confirmation of the previous studies. I am missing an originality of this study. The difference to the previous studies is the use of transgene animals overexpressing TGF instead 
of application of exogenous TGF as used before. 
3. Since NFkappaB plays a crucial role in the transcriptional regulation of pro-survival genes and there is an evidence for interaction between TGF and NFkappaB ( Kanai et al Gastroenterology 
2001; 121: 56-67). It would be of great importance to check this aspect in TGFalpha transgenic mice. 
Authors should compare the activation of NFkappaB in the gastric mucosa of wild animals and 
transgene mice overexpressing TGFa. This would be a new and original finding instead of repetition 
of previous results. 4. It is well known that TGF has antisecretory affects. However, there was no difference in 
intragastric pH between WT animals and TG animals. I strongly suggest the measurement of gastric 
acid secretion in mice equipped with special gastric fistula. 
5. In addition to analysis of COX-2 expression, the authors should measure prostaglandin generation 
in both animals groups (WT vs. TG). 
6. Since gastrin play a crucial role in the regulation of gastric secretion and also shows 
gastroprotective effects, it would be of importance to measure gastrin in both groups. 
7. Although the discussion and conclusions are well balanced and adequately supported by the data, 
the whole manuscript is too long, and the in vitro data should be omitted. Discussion should be 
shortened. 
 Unable to decide on acceptance or rejection until the authors have responded to the What next?: 
major compulsory revisions 
 An article whose findings are important to those with closely related research Level of interest: 
interests 
 Needs some language corrections before being published Quality of written English: 
 No Statistical review: 
Declaration of competing interests: 
I declare that I have no competing interests 
assistant