Photo- and Schiff Base-Crosslinkable Chitosan/Oxidized Glucomannan Composite Hydrogel for 3D Bioprinting
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsIn this paper, A novel chitosan/oxyglucomannan composite hydrogel was prepared by photoinduced crosslinking of Schiff base and phenol. The freshly created product was examined using a variety of instruments to examine its characteristics. The subject of this study is interesting and the whole manuscript is clear and logical. I recommend this paper for acceptance by the Polysaccharides with some necessary revisions. The following are specific comments that I need to consider before final acceptance.
1. As mentioned in 2.6 “The samples were loaded into syringes and manually extruded through a tube of 1.5mm diameter to assess their extrudability.” Whether the results of manual extrusion will cause deviations between the results？
2. The peaks at 1716 cm-1 and 805 cm-1 are not distinct enough. It is recommended to conduct the test again and analyze it.
3. Significance was mentioned in 3.4, but no multiple experiments were conducted. Multiple parallel experiments need to be carried out for significance analysis.
4. Some relative works should be referred to in the introduction for improving the quality of this manuscript. e.g. https://doi.org/10.3389/fchem.2023.1327426, https://doi.org/10.1016/j.indcrop.2024.119517, https://doi.org/10.1002/adfm.202410698. These documents may contribute to the author's understanding of chitosan or hydrogels.
5. The explanation of the relationship between hydrophobicity and hydrogel contraction seems rather far-fetched. Please conduct a reanalysis.
6. The author needs to enhance the stability of the water gel with pH greater than 7 to determine the differences in the stability of the water gel under acidic or alkaline conditions.
Author Response
Comment1: As mentioned in 2.6 “The samples were loaded into syringes and manually extruded through a tube of 1.5mm diameter to assess their extrudability.” Whether the results of manual extrusion will cause deviations between the results？
Responce1: Thank you very much for the question. We think that the manual force applied to syringe or syringe diameter does not cause the difference. At least, the hydrogel was not decomposed after the extrusion. In addition, in the rheology measurement, high strain (1000 %) was applied to the hydrogel samples, and the sample retained hydrogel property after that. We think that the strain manually applied to the samples in the syringe was less than that applied by rheometer. Therefore, we believe that the results would not be changed by the force of manual extrusion. The purpose of this experiment was to whether the samples retain their shape, or hydrogel state after the loading to the syringe and the extrusion from it. In 3D bioprinting process, such operations are necessary to fill the syringe with a bioink. The ChPh-Ox glucomannan hydrogels retained their shape after the loading and extruding. We have added the explanation in page 7 LINE 261-262
Comment2: The peaks at 1716 cm-1 and 805 cm-1 are not distinct enough. It is recommended to conduct the test again and analyze it.
Responce2: We apologize for the insufficient explanation. The peak at 805 cm-1 disappeared in Ox-glucomannan sample, indicating the ring cleavage of glucomannan due to the oxidation. We apologize for the miswriting again. We have revised the manuscript in page6 LINE 240. And as you mentioned, the peak 1705 was too weak. So, we have conducted FTIR analysis again with higher resolution based on your comment (FigR1). However, we could not detect the significant peak at 1705 cm-1 probably due to the low resolution of FTIR or　overlapping of other peaks. We have added the explanation and a reanalyzed data in page6 LINE240-244, and the information regarding the FTIR condition in 2.4. Thank you very much for the comment.
FigR1. FTIR spectra of glucomannan and Ox-glucomannan. The analysis was reconducted at a resolution of 2 cm-1 .
Comment3: Significance was mentioned in 3.4, but no multiple experiments were conducted. Multiple parallel experiments need to be carried out for significance analysis.
Responce3: Thank you very much for the comment. We prepared three or four hydrogel disks on each sample for swelling test. And, the statistical analysis was conducted to compare the difference using Student’s p-test. We have added the section of statistical analysis(section2.12) and information in figure6.
Comment4: Some relative works should be referred to in the introduction for improving the quality of this manuscript. e.g. https://doi.org/10.3389/fchem.2023.1327426, https://doi.org/10.1016/j.indcrop.2024.119517, https://doi.org/10.1002/adfm.202410698. These documents may contribute to the author's understanding of chitosan or hydrogels.
Responce4: Thank you very much for the constructive comment. We have revised the introduction and added the references based on your comment (page2 LINE47-50 ref15, page2 LINE80 ref24.) We hope the revised introduction become more readable.
Comment5: The explanation of the relationship between hydrophobicity and hydrogel contraction seems rather far-fetched. Please conduct a reanalysis.
Responce5: Thank you very much for the comment. We reconsidered the reason. We think the main reason is due to the molecular weight of chitosan. We have revised the discussion and added limitations based on your comment (Page9 LINE325-337). We greatly appreciate the opportunity to reanalyze and revise this section. We are confident that these revisions have made the manuscript clearer and more comprehensible for readers. Thank you once again for your valuable input.
Comment6: The author needs to enhance the stability of the water gel with pH greater than 7 to determine the differences in the stability of the water gel under acidic or alkaline conditions.
Responce6: Thank you very much for the comment. We did the swelling test at 7 because the physiological environment is almost neutral condition. In addition, there are several reports that Chitosan has a wound healing property. We were expecting that the proposed hydrogel had a potential for wound dressing application. The pH environment of human skin is around 4-7, so we evaluated the stability under the acidic condition (pH4, ref1). We have added the explanation and purpose of the experiment in page9 LINE320-321, page10 LINE372-374. We believe that incorporating this additional explanation has made the manuscript more accessible and informative for readers. Thank you again for helping us improve the clarity of our work.
1)https://www.acne.org/what-is-the-ph-of-human-skin
2)https://optimalbreathing.com/blogs/marlenea/importance-of-your-body-ph
Author Response File: 
Author Response.docx
Reviewer 2 Report
Comments and Suggestions for AuthorsThe research article entitled “Photo- and Schiff Base-Crosslinkable Chitosan/Oxidized Glucomannan Composite Hydrogel for 3D Bioprinting” by Mitsuyuki Hidaka and Shinji Sakai describes the effectiveness of crosslinking of chitosan-based phenol and oxidized glucomannan modified hydrogel for 3D printing.
1. The main question addressed by the researchers in this research article is the change in properties and the efficiency of the hydrogels formed from chitosan derived polymers. These composite hydrogels have improved the stability compared to its untreated or single modified materials.
2. The topic is relevant to the field and research on this topic is needed because chitosan is an important biomaterial used as an antimicrobial. The application of these materials in the form of hydrogel facilitates its use over just chitosan-based material.
3. The authors have provided a detailed introduction and explained the importance of the current study. The authors talked about types of bioprinting techniques such as extrusion, inkjet, and vat-polymerization based bioprinting for biomedical applications. They also mentioned about the different materials used under these categories along with chitosan.
4. The authors have explored the crosslinking technique to increase the stability of the hydrogel formed from hybrid material. They used phenol modified materials for the photo induced crosslinking and oxidized glucomannan based materials for the Schiff base modification. The free amine of the chitosan was treated with 3-(4-Hydroxyphenyl) propionic acid in the presence of coupling reagent EDC.HCl to get the phenol modified chitosan derivative. In order to get the Schiff based modification, the authors first oxidized the glucomannan by using sodium periodate to get the dialdehyde derivative. This dialdehyde derivative was then coupled with free amine of the chitosan to get the Schiff base linked chitosan-glucomannan polymer.
5. The authors have carried out the characterization of these modified polymers by using FTIR technique.
6. After the hydrogel formation from the PBS solution, the gel was characterized using a rheometer and its swelling abilities. Also, the gel stability was checked by using different pH solutions. The hydrogel made up from the modification of phenol and Schiff base followed by the photo irradiation gave the highest stability. The only phenol modification or only the Schiff base modification did not provide good stability to the hydrogel.
7. The authors have shown the extrudabilities of the polymer composites and found that the higher glucomannan containing polymer composites were not injectable.
8. The authors have demonstrated the swelling behaviors of the chitosan-phenol-Schiff base composite polymers. They observed the shrinking of the hydrogel when treated with PBS solution.
9. They have carried out the cell viability study and found that these polymer composites were nontoxic.
10. The authors have finally studied the printability of these polymer composites and identified that small polymers are good to be used for printing. The higher analogs were clogged during printing.
11. Though the authors have investigated hydrogel made in PBS, this hydrogel shrank when tested for swelling. It was not clear whether phenol rings have any influence because the phenol rings are present before the swelling. The authors need to explain in detail about this.
12. The authors have not mentioned the frequency used for the rheology data presented in figure 5.
13. Figure 2 shows FTIR spectra of glucomannan and Ox-glucomannan. However, the authors have given different explanations on page 6. They have mentioned that “In addition, another peak appeared at 805 cm−1, indicating the ring cleavage of mannose.” The careful analysis of the spectra shows that there is no peak at 805 cm−1 for ox-glucomannan.
14. They have also mentioned that “an additional small peak attributable to the aldehyde group was observed at 1716 cm−1, indicating the oxidization of glucomannan”. However, the peak at 805 cm−1is also seen in both the spectra.
15. The explanation given by the authors does not match the spectra because they mentioned that the peaks for oxidized version are 1716 cm−1 and 805 cm−1. However, the spectra in which the peak at 1716 cm−1 is present does not have the peak 805 cm−1.
16. Apart from the core scientific comments, there are some issues that need to be addressed.
17. The introduction or any other section needs a schematic figure outlining the previous modifications to use the chitosan for 3D printing.
18. The authors have given the wrong chemical name in section 2.1, third line. They need to provide the correct name for the 3-(4-Hydroxyphenyl) propionic acid.
19. The authors have interchanged the caption in figure 1. They have given (b) Oxidation of glucomannan for the phenol crosslinked hydrogel shown in fig 1b. and (c) phenol crosslinking of ChPh, for the oxidation of the glucomannan. They need to correct this.
20. The authors need to show full spectra of FTIR because it is important to show the -OH stretching frequency.
21. In figure 3, the authors need to denote the photographs of upper right and lower left with appropriate notations.
22. There are many typos and grammatical errors that need to be fixed before publication.
a) section 3.5: para 2: line 4: the citation for “figure 8 (c)” can be replaced with “figure 7 (c)”
b) Throughout the reference list there is no consistency while writing the reference paper titles. In some places, the title contains all words starting with capital letters and in some places, only the first letter of the title starts with capital letters. The authors need to be consistent while writing the reference list throughout the reference list.
c) The authors need to use the standard abbreviations while writing the journal names.
d) In some of the cases there is no full-stop after the abbreviation of the journal name.
e) In the ref list, at some places the references are written with page numbers and at some places there are no page numbers. The authors need to be consistent throughout the references.
f) Please remove the double numbering for the reference list.
Author Response
Comment1: The main question addressed by the researchers in this research article is the change in properties and the efficiency of the hydrogels formed from chitosan derived polymers. These composite hydrogels have improved the stability compared to its untreated or single modified materials.
Responce1: Thank you very much for the comment. We believe this hydrogel has a great potential for tissue regeneration.
Comment2: The topic is relevant to the field and research on this topic is needed because chitosan is an important biomaterial used as an antimicrobial. The application of these materials in the form of hydrogel facilitates its use over just chitosan-based material.
Responce2: Thank you very much for the comment. We believe this hydrogel widen the application of chitosan.
Comment 3: The authors have provided a detailed introduction and explained the importance of the current study. The authors talked about types of bioprinting techniques such as extrusion, inkjet, and vat-polymerization based bioprinting for biomedical applications. They also mentioned about the different materials used under these categories along with chitosan.
Responce3: Thank you very much for the comment. We have added more explanation regarding the material in Page 1 LINE38-43, page 2 LINE47-50. We hope the introduction become more readable and understandable for the readers.
Comment4: The authors have explored the crosslinking technique to increase the stability of the hydrogel formed from hybrid material. They used phenol modified materials for the photo induced crosslinking and oxidized glucomannan based materials for the Schiff base modification. The free amine of the chitosan was treated with 3-(4-Hydroxyphenyl) propionic acid in the presence of coupling reagent EDC.HCl to get the phenol modified chitosan derivative. In order to get the Schiff based modification, the authors first oxidized the glucomannan by using sodium periodate to get the dialdehyde derivative. This dialdehyde derivative was then coupled with free amine of the chitosan to get the Schiff base linked chitosan-glucomannan polymer.
Responce4: Thank you very much for the comment. We believe the phenol was modified to chitosan, and glucomannan was oxidized.
Comment5: The authors have carried out the characterization of these modified polymers by using FTIR technique.
Response 5: Thank you very much for the comment. We have revised the section3.1.
Comment 6: After the hydrogel formation from the PBS solution, the gel was characterized using a rheometer and its swelling abilities. Also, the gel stability was checked by using different pH solutions. The hydrogel made up from the modification of phenol and Schiff base followed by the photo irradiation gave the highest stability. The only phenol modification or only the Schiff base modification did not provide good stability to the hydrogel.
Response 6: Thank you very much for the comment. That is what we wanted to show.
Cooment7: The authors have shown the extrudabilities of the polymer composites and found that the higher glucomannan containing polymer composites were not injectable.
Responce7: Thank you very much for the comment.
Comment8: The authors have demonstrated the swelling behaviors of the chitosan-phenol-Schiff base composite polymers. They observed the shrinking of the hydrogel when treated with PBS solution.
Responce8: Thank you very much for the comment.
Comment9: They have carried out the cell viability study and found that these polymer composites were nontoxic.
Responce9: Thank you very much for the comment. We have revised the section in Page 12 LINE 398-403.
Comment 10: The authors have finally studied the printability of these polymer composites and identified that small polymers are good to be used for printing. The higher analogs were clogged during printing.
Responce10: Thank you very much for the comment. We have added the limitation and future perspectives in Page 13 LINE442-447.
Comment 11: Though the authors have investigated hydrogel made in PBS, this hydrogel shrank when tested for swelling. It was not clear whether phenol rings have any influence because the phenol rings are present before the swelling. The authors need to explain in detail about this.
Responce11: Thank you very much for the comment. We have reconsidered the reason. We think the reason of shrinking is due to the low molecular weight of chitosan. We have revised the manuscript and added the limitation in Page9 LINE 324-334. We are deeply grateful for the opportunity to revisit and refine our interpretation. We believe that these revisions have undoubtedly improved the clarity and scientific rigor of our manuscript. Thank you once again for your insightful feedback.
Comment 12: The authors have not mentioned the frequency used for the rheology data presented in figure 5.
Responce12: Thank you very much for the comment. We have added the operation condition of rheometer in the figure legends of Fig.4 and5.
Comment 13: Figure 2 shows FTIR spectra of glucomannan and Ox-glucomannan. However, the authors have given different explanations on page 6. They have mentioned that “In addition, another peak appeared at 805 cm−1, indicating the ring cleavage of mannose.” The careful analysis of the spectra shows that there is no peak at 805 cm−1 for ox-glucomannan.
Responce13: We apologize for the insufficient explanation. The peak at 805 cm-1 disappeared in Ox-glucomannan sample, indicating the ring cleavage of glucomannan. We have revised the explanation in page6 LINE 240-243. We apologize for the miswriting again (FigR1). And, we have conducted FTIR analysis again with higher resolution. The condition has been added in 2.4., and the reanalyzed data has been added in Fig6.
FigR1. FTIR spectra of glucomannan and Ox-glucomannan. The analysis was reconducted at a resolution of 2 cm-1 .
Comment 14: They have also mentioned that “an additional small peak attributable to the aldehyde group was observed at 1716 cm−1, indicating the oxidization of glucomannan”. However, the peak at 805 cm−1is also seen in both the spectra.
Responce14: We apologize for the insufficient explanation. We have analyzed again and revised the FTIR data. However, the peak attributing to aldehyde could not be detected. probably due to the low resolution of FTIR or　overlapping of other peaks We have revised the explanation in page6 LINE 240-243.
Comment15: The explanation given by the authors does not match the spectra because they mentioned that the peaks for oxidized version are 1716 cm−1 and 805 cm−1. However, the spectra in which the peak at 1716 cm−1 is present does not have the peak 805 cm−1.
Responce15: We apologize for the insufficient explanation. We have analyzed and revised the FTIR data. We have revised the explanation in page6 LINE 240-243.
Comment16: Apart from the core scientific comments, there are some issues that need to be addressed.
Responce16: We apologize for the mistakes in the manuscript. We have revised it carefully.
Comment 17: The introduction or any other section needs a schematic figure outlining the previous modifications to use the chitosan for 3D printing.
Response 17: Thank you very much for the comment. We have added the figure printing structure using the ink (Figure10(a)).
Comment 18. The authors have given the wrong chemical name in section 2.1, third line. They need to provide the correct name for the 3-(4-Hydroxyphenyl) propionic acid.
Response 18: We apologize for the incorrect chemical name. We have revised it in this section. Thank you very much for the comment.
Comment19: The authors have interchanged the caption in figure 1. They have given (b) Oxidation of glucomannan for the phenol crosslinked hydrogel shown in fig 1b. and (c) phenol crosslinking of ChPh, for the oxidation of the glucomannan. They need to correct this.
Responce19: We apologize for the incorrect figure caption. We have revised the captions. Thank you very much for the comment.
Comment 20: The authors need to show full spectra of FTIR because it is important to show the -OH stretching frequency.
Response 20: Thank you very much for the constructive comment. We have added the data at 3000-4000 cm-1 and the explanation (Figure2).
Comment 21: In figure 3, the authors need to denote the photographs of upper right and lower left with appropriate notations.
Response 21: We appreciate for the constructive comment. We have added the information in Figure3. Thank you very much for the comment.
Comment 22: There are many typos and grammatical errors that need to be fixed before publication.
- a) section 3.5: para 2: line 4: the citation for “figure 8 (c)” can be replaced with “figure 7 (c)”
Response 22: We apologize for the incorrect figure numbers. After checking the manuscript carefully, we have revised the numbers (Section3.4., LINE 319, 323, Section 3.5. LINE352,　LINE360, Para2 LINE366).
- b) Throughout the reference list there is no consistency while writing the reference paper titles. In some places, the title contains all words starting with capital letters and in some places, only the first letter of the title starts with capital letters. The authors need to be consistent while writing the reference list throughout the reference list.
Response 22:
We apologize for the inconsistency. We have revised the reference styles.
- c) The authors need to use the standard abbreviations while writing the journal names.
Response 22:
We apologize for it. We have revised the abbreviation.
- d) In some of the cases there is no full-stop after the abbreviation of the journal name.
Response 22:
We apologize for it. We have revised the reference style.
- e) In the ref list, at some places the references are written with page numbers and at some places there are no page numbers. The authors need to be consistent throughout the references.
Response 22:
We apologize for the mistakes. We have added the page number. Thank you very much for the comment.
- f) Please remove the double numbering for the reference list.
Response 22:
Thank you very much for the comment. We have checked and revised the references carefully.
Author Response File: Author Response.docx
Reviewer 3 Report
Comments and Suggestions for AuthorsThis study presents a novel double-crosslinkable chitosan/oxidized glucomannan composite hydrogel and explores its potential application in 3D bioprinting. Overall, the paper is innovative in content, reasonable in experimental design, and encouraging in results. However, there are still some aspects that need improvement and further clarification. The following are my review comments:
1. Literature review: the review of existing studies in the introduction section is rather brief, and it is recommended to increase the discussion of related literature, especially the rationale for choosing chitosan as a matrix, and it is recommended to add a comparison with other known hydrogel materials (e.g., gelatin, poly(vinyl alcohol), etc.), and the current status and challenges of the application of chitosan and its derivatives in bioprinting, in order to enhance the depth of the research background.
2. Experimental Methods: In the Materials and Methods section, it is recommended that more detailed experimental steps be provided, including specific temperature, time, pH, and cross-linker concentration, etc., so that other researchers can repeat the experiments and verify the results.
3. Gradient selection: In the article, there is no experimental sample with the best performance, it is recommended to refine the gradient design to explore the optimal ratio and its effect on the performance mechanism, in order to enhance the value and significance of the study.
4. Cell activity evaluation: It is recommended to increase the time point and density data of cell activity evaluation, such as observing cell proliferation or survival after 1 or 2 weeks, in order to evaluate the cytocompatibility of hydrogel in long-term culture.
5. Antimicrobial experiment details: It is recommended that more mechanistic analysis be provided in the antimicrobial experiments, as the antimicrobial properties do not change decreasingly with Ox-glucomannan concentration, and it is recommended to point out the reasons for the different effects of Ox-glucomannan concentration on the antimicrobial properties.
6. 3D printing evaluation: the picture of the nasal structure printed by ChPh-Ox25 mentioned in the article did not show good printability, and it is suggested to improve it.
7. Graphical details: It is recommended that the graphs be optimized to ensure that the legends and labels are clear and readable, especially in the graphs of the rheology and antimicrobial experimental results, and it is recommended that the images and rulers be neatly aligned so that readers can quickly understand the data.
8. Discussion section: It is recommended that the discussion section be expanded to analyze in depth the potential applications and limitations of the developed hydrogels, as well as the direction of future research, in order to enhance the depth and breadth of the paper.
9. Clarity of conclusion: It is recommended that the main findings and contributions of the study be clearly summarized in the conclusion section, echoing the “double cross-links” in the abstract and keywords.
Author Response
Comment1: Literature review: the review of existing studies in the introduction section is rather brief, and it is recommended to increase the discussion of related literature, especially the rationale for choosing chitosan as a matrix, and it is recommended to add a comparison with other known hydrogel materials (e.g., gelatin, poly(vinyl alcohol), etc.), and the current status and challenges of the application of chitosan and its derivatives in bioprinting, in order to enhance the depth of the research background.
Responce1: Thank you very much for the constructive comment. We have added the explanation of other polymers including synthetic polymers and other polysaccharides in page 1 LINE 38-43. In addition, we have added to explanation to emphasize the difference of chitosan from other polysaccharide in page2 LINE47-50.
Comment2: Experimental Methods: In the Materials and Methods section, it is recommended that more detailed experimental steps be provided, including specific temperature, time, pH, and cross-linker concentration, etc., so that other researchers can repeat the experiments and verify the results.
Responce2: Thank you very much for the comment. We have added the concentration information or rheometer parameters in Fig. 3-10. In addition, we have added the temperature in section 2.2., 2.3, 2.5, 2.7. We hope these revisions make the contents more readable.
Comment3: Gradient selection: In the article, there is no experimental sample with the best performance, it is recommended to refine the gradient design to explore the optimal ratio and its effect on the performance mechanism, in order to enhance the value and significance of the study.
Resnponce3: Thank you very much for the comment. We have added the limitation and future perspectives in Page 13 LINE443-448, LINE461-464, and Page14 LINE478-480.
Comment4: Cell activity evaluation: It is recommended to increase the time point and density data of cell activity evaluation, such as observing cell proliferation or survival after 1 or 2 weeks, in order to evaluate the cytocompatibility of hydrogel in long-term culture.
Responce4: Thank you very much for the comment. It has been reported that chitosan is a cell compatible material [1] and long-term culture at least for 7 days are available on the hydrogel [2]. Our results showed that the cell survived and grew on the hydrogel. The purpose of this experiment was to check whether the hydrogel has a cell compatibility as well as the previously reported chitosan hydrogels. We believe that the long-term cultivation is possible by modifying the culture condition and other parameters. We have added the prospect and comparisons with other reports in Page 12 LINE 399-405.
[1] https://www.frontiersin.org/journals/chemistry/articles/10.3389/fchem.2023.1327426/full
[2] Enhancing cell penetration and proliferation in chitosan hydrogels for tissue engineering applications
Comment5: It is recommended that more mechanistic analysis be provided in the antimicrobial experiments, as the antimicrobial properties do not change decreasingly with Ox-glucomannan concentration, and it is recommended to point out the reasons for the different effects of Ox-glucomannan concentration on the antimicrobial properties.
Responce5: Thank you very much for the comment. We have added some explanations and discussions in Page12 LINE419-423.
Comment 6: 3D printing evaluation: the picture of the nasal structure printed by ChPh-Ox25 mentioned in the article did not show good printability, and it is suggested to improve it.
Responce6: Thank you very much for the comment. We have added the description regarding the low printability of ChPh-Ox25, and the reason as well. In addition, we have also added the future issues in page13 LINE442-447.
Comment 7: Graphical details: It is recommended that the graphs be optimized to ensure that the legends and labels are clear and readable, especially in the graphs of the rheology and antimicrobial experimental results, and it is recommended that the images and rulers be neatly aligned so that readers can quickly understand the data.
Response 7: We apologize for the insufficient figure. We have rearranged the figure, added the minor tick marks to the graphs in Figure4, 9. In addition, we have modified the rulers and their layout in Figure 6 and Figure7.
Comment 8: Discussion section: It is recommended that the discussion section be expanded to analyze in depth the potential applications and limitations of the developed hydrogels, as well as the direction of future research, in order to enhance the depth and breadth of the paper.
Response 8: Thank you very much for the comment. We have added the discussion regarding the swelling in page10 LINE367-369, the cell compatibility in page 12 LINE398-403, 3D printing in page13 LINE442-447, and LINE460-463.
Comment 9: Clarity of conclusion: It is recommended that the main findings and contributions of the study be clearly summarized in the conclusion section, echoing the “double cross-links” in the abstract and keywords.
Response 9: Thank you very much for the comment. We have added our contributions more in the conclusion.
Author Response File: Author Response.docx
Round 2
Reviewer 1 Report
Comments and Suggestions for AuthorsAfter carefully reading the revised manuscript and the corresponding response to the reviewer's comments, I think this manuscript has been abundantly improved and deserved to be published in this Journal at the present form.
Reviewer 2 Report
Comments and Suggestions for AuthorsThank you for the revision of the manuscript. The authors have addressed all the concerns and provided the explanation for the queries.
Reviewer 3 Report
Comments and Suggestions for AuthorsThe manuscript can be accepted.
Comments on the Quality of English LanguageThe English could be improved to more clearly express the research.
