1. Mesenchymal Stem Cells Prevent Hypertrophic Scar Formation via Inflammatory Regulation when Undergoing Apoptosis 
Shiyu Liu, Lan Jiang, Haijian Li, Haigang Shi, Hailang Luo, Yongjie Zhang, Chunyan Yu, Yan Jin 
Journal of Investigative Dermatology 
Volume 134, Issue 10, Pages (October 2014)
DOI: /jid
Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Local injection of human mesenchymal stem cells (hMSCs) prevented hypertrophic scar formation in a rabbit scar model. (a) Images of wounds created on rabbit ear and the scar formation on postoperative day 28. Scale bar=5mm. (b) The analysis of the outstanding height of the scar on postoperative day 28 (n=12). (c) Hematoxylin and eosin (H&E) staining of wound area tissues on postoperative days 14 and 28. Bar=2mm. The double arrows indicate the granulation tissues on postoperative day 14. The dotted lines indicate the wound edges. (d) The analysis of scar elevation index (SEI) based on H&E staining on postoperative day 28 (n=6). (e) Masson's trichrome staining of wound area tissues followed by integrated optical density (IOD) analysis (f) on postoperative day 28 (n=12). Bar=200μm. (g h) Real-time PCR assay of collagen type I α1 (ColIal) and alpha-smooth muscle actin (α-SMA) expressions in wound area on postoperative day 7 (n=3). Values are means±SD. **P<0.01.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Human mesenchymal stem cells (hMSCs) regulated inflammation and prevented scar formation partially through TNF-alpha-stimulated gene/protein 6 (TSG-6). (a) Human genome-specific real-time PCR assay of TSG-6, inducible nitric oxide synthase (iNOS), indoleamine 2,3-dioxygenase (IDO), cyclooxygenase 2 (COX-2), and interleukin-10 (IL-10) levels in hMSCs at different time points post hMSC injection (n=3). (b) Local injection of hMSC prevented CD45-positive cell (red) and CD3-positive cell (green) infiltration on postoperative day 3 and day 5, respectively, which was attenuated by TSG-6 knockdown in hMSCs by small interfering RNA (siRNA). Bar=50μm. (c) The CD45- and CD3-positive cells were analyzed as the percentage of total detected cells (n=12). (d−h) Proinflammatory/profibrotic mediators in the wound area were analyzed by real-time PCR assay (d−f) (n=3) or ELISA (g h) (n=6) on postoperative day 3. (i) Images of wounds created on rabbit ear and the scar formation on postoperative day 28. Scale bar=5mm. (j) The analysis of the outstanding height of the scar on postoperative day 28 (n=12). Values are means±SD. **P<0.01.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Human mesenchymal stem cells (hMSCs) underwent extensive apoptosis and survived for a short time after injection. (a) DiO-labeled hMSCs (green) in the wound area at 24 and 72hours post injection. Bar=200μm. (b) Standard curve for real-time PCR assay of human-specific glyceraldehyde-3-phosphate dehydrogenase (hGAPDH). Values indicate ΔΔCt of primers for human/rabbit GAPDH (h/rGAPDH) genes and cDNA for hGAPDH on the same samples (n=3). (c) Percentage of surviving hMSCs in the wound area at different time points post injection based on the standard curve (n=3). (d) DiO-labeled hMSCs (green) and TUNEL-positive cells (red) at different time points post injection. The costained cells represented the apoptotic hMSCs, which were then analyzed as the percentage of total detected hMSCs (e) (n=12). Scale bar=50μm. (f) Real-time PCR assay of caspase-3, -7, and -9 expressions of hMSCs at different time points post injection (n=3). Values are means±SD. **P<0.01.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Apoptotic human mesenchymal stem cells (hMSCs) increased TNF-alpha-stimulated gene/protein 6 (TSG-6) secretion partially through caspase-3 activation. (a) Percentage of TUNEL-positive hMSCs after 6hours of treatment (n=12). The H2O2 treatment induced the apoptosis of hMSCs, which was attenuated by pretreatment of Z-DEVD-FMK, an inhibitor of caspase-3. (b) The caspase-3 activity of hMSCs after 6hours of treatment (n=6). (c) Real-time PCR assay of TSG-6 expression in hMSCs after 6hours of treatment (n=6). (d) ELISA of TSG-6 secreted by hMSCs after 12hours of treatment (n=6). Values are means±SD. **P<0.01.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
Apoptosis partially governed the therapeutic effects of human mesenchymal stem cells (hMSCs) on hypertrophic scar. (a) DiO-labeled hMSCs (green) and TUNEL-positive cells (red) at 24hours post injection. Untreated hMSCs, H2O2-treated hMSCs (apoptotic hMSCs), and Z-DEVD-FMK-treated hMSCs were applied to the rabbit scar model. The costained cells represented the apoptotic hMSCs, which were then analyzed as the percentage of total detected hMSCs (b) (n=12). Bar=50μm. (c) Percentage of surviving hMSCs in the wound area at different time points post injection. (d−g) Proinflammatory/profibrotic mediator levels in the wound area were analyzed by real-time PCR (d e) (n=3) or ELISA assay (f g) (n=6) on postoperative day 3. (h) Real-time PCR assay of human TNF-alpha-stimulated gene/protein 6 (TSG-6) expressions at different time points post injection (n=3). (i) Images of wounds created on rabbit ear and the scar formation on postoperative day 28. Scale bar=5mm. (j) The analysis of the outstanding height of the scar on postoperative day 28 (n=12). Values are means±SD. **P<0.01.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions