In Vitro Evidence of Differential Immunoregulatory Response between MDA-MB-231 and BT-474 Breast Cancer Cells Induced by Bone Marrow-Derived Mesenchymal Stromal Cells Conditioned Medium
, Víctor A. Cortés-Morales 3, José R. Navarro-Betancourt 1, Janneth Peralta-Ildefonso 1, Bulmaro Cisneros 2 and Salomón Hernández-Gutiérrez 1,*Round 1
Reviewer 1 Report
Arenas-Luna et al. performed in vitro study to show the effect of human bone marrow derived mesenchymal stem/stromal cells on immunoregulatory properties of two different cancer lines (MDA-MB 231, BT-474). They demonstrated that MDA-MB 231 cells, but not BT-474, in co-culture with mononuclear cells (MNCs) in the presence of hBM-MSCs conditioned medium inhibited MNCs proliferation and induced Treg phenotype. They showed that in such an experimental setting MDA-MB 231 cells secreted increased levels of IL-10 and TGFb and IDO. Authors conclude that those molecules may be involved in observed immunoregulatory response of MDA-MB 231 cancer cells. Overall, the study is of good quality, despite that potential mechanism of hBM-MSCs on cancer cells is missing.
There are only a few minor drawbacks that need to be corrected:
1) Please provide information on the number of healthy adult donors (n=…) participated in bone marrow aspiration procedure, how many donor specimens were used in the study (2. Material and methods 2.1. Cell cultures, lines 98 and 117)
2) Please provide information whether isolated BM-MSCs were pooled from several donors or each donor derived BM-MSCs was treated as separate cell line. It is important due to great heterogeneity in secreted molecules from donor to donor. The impact of BM-MSs-cm on immunoregulatory response of cancer cells may be dependent of various batches of BM-MSCs. Please refer to this aspect in Discussion section (cite doi: 10.1186/s13287-019-1331-9)
3) In all figures there is no indicated number of repetitions of a given experiment. Please complete this information under each figure.
4) Please describe Figure 6 in more detail either under the figure or in the text.
5) Why the authors did not perform analysis on molecules secreted by hBM-MSCs in Fig. 4?
6) I would recommend in all figures to switch the order of the displayed data, “A” as MDA-MB-231, “B” BT-474
Line:
162 – conditioned medium (…), which the media were replaced – please clarify conditioned medium composition: with or without FBS?
200 – then, the MNCs were incubated (…) – how long?
209 – the ratios are: 2:1 (…) 3:1 (…) – why are not the same?
422,23,24 – typo: CMNs, change to MNCs
Author Response
"Please see the attachment."
Author Response File: 
Author Response.pdf
Reviewer 2 Report
This manuscript by Víctor M Arenas-Luna and colleagues explores the regulatory effect of mesenchymal stromal cells on the immune microenvironment of breast cancer with the breast cancer cell line model MDA-MB-231 and BT-474. In an MDA-MB-231 cell line-specific way, mesenchymal stromal cells induced the compromised anti-tumor immune response through secreted soluble factors.
The topic of breast cancer immune microenvironment and mesenchymal stromal cells is interesting. There are some points in the paper that need to be further clarified or improved.
(1) In abstract, the results should be better summarized, and conclusions/significance of this study need to be mentioned.
(2) Western blot of MDA-MB-231 and BT-474 need to be done to make the convincing conclusion that IDO protein levels is increased under those conditions.
(3) The authors need to explain why TNF, IL-4, IL-10 and TGF-β were specifically chosen out of all cytokines.
(4) In figure 6B, IL-10 is shown to increase, which is not mentioned in the results context. The authors have to explain the inconsistency.
(5) The discussion section must be improved. At this stage, it is not succinct and concise enough and redundantly mentioned too much research background.
The manuscript is overall well-written except for some minor issues:
(1) Line 22, “Therefore, this in vitro work aimed to study, the effect of the conditioned medium of human bone” the comma needs to be deleted.
(2) Line 399 to 401, it is an incomplete sentence.
(3) Line 489 to 491, the sentence needs to be rephrased to make it easy to understand
(4) Line 579 to 580, there might be a typo here, so this sentence needs to be clarified.
Author Response
"Please see the attachment."
Author Response File: Author Response.pdf
Reviewer 3 Report
The MS entitled "In vitro Evidence of Differential Immunoregulatory Response 2 Between MDA-MB-231 and BT-474 Breast Cancer Cells Induced 3 by Bone Marrow-derived Mesenchymal Stromal Cells Condi- 4 tioned Medium” is well written.
Author Response
"Please see the attachment."
Author Response File: Author Response.pdf
