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Alleviation of skin inflammation after Lin(-) cell transplantation correlates with their differentiation into myeloid-derived suppressor cells.

Jeong Ryu S, Ju JM, Kim W, Bum Kim M, Hee Oh K, Sup Lee D, Lee H, Eun Oh J, Soo Park K, Young Choi E - Sci Rep (2015)

Bottom Line: Longitudinal in vivo imaging and flow cytometry analyses revealed that Lin(-) cells home directly to inflamed skin within 6 h, where they undergo extensive expansion with the peak on day 14 post-transplantation, and preferential differentiation into CD11b(+)Ly6G(int)Ly6C(+) cells by day 7.Cells with phenotypic profiles of neutrophils, macrophages, and DCs appeared in inflamed skin on day 14.Progenies of transplanted Lin(-) cells showed similar kinetics of expansion and myeloid differentiation in BM.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, Korea.

ABSTRACT
To understand the cellular mechanism underlying the therapeutic effects exerted by hematopoietic stem cell transplantation in the repair of tissue damage, we investigated the in vivo dynamics of bone marrow (BM) lineage-negative (Lin(-)) cells transplanted into mice with hyper sensitivity dermatitis. Longitudinal in vivo imaging and flow cytometry analyses revealed that Lin(-) cells home directly to inflamed skin within 6 h, where they undergo extensive expansion with the peak on day 14 post-transplantation, and preferential differentiation into CD11b(+)Ly6G(int)Ly6C(+) cells by day 7. Cells with phenotypic profiles of neutrophils, macrophages, and DCs appeared in inflamed skin on day 14. Progenies of transplanted Lin(-) cells showed similar kinetics of expansion and myeloid differentiation in BM. However, differentiation into CD11b(+)Ly6G(int)Ly6C(+) cells in the inflamed skin on day 7 was more skewed toward CD115(+) cells (≥60%) with immune suppressive function and higher expression levels of iNOS, arginase, and IL-10, compared with those in the BM. Transplantation of Lin(-) cells reduced the levels of Cd3 transcript and CD4(+)/CD8(+) cells in inflamed skin. These results demonstrate differentiation of transplanted Lin(-) cells into myeloid-derived suppressor cells in inflamed skin to be the basis of the alleviation of skin inflammation after Lin(-) cell transplantation.

No MeSH data available.


Related in: MedlinePlus

Early homing and expansion of Lin− cells at the site of inflammation and BM.(a–c) Flow cytometric analysis of CMFDA+ cells in inflamed skin and BM at 6 and 12 h after transplantation of CMFDA+CD45.1+Lin− cells. Dermatitis or vehicle-control CD45.2+ B6 mice were transplanted with CMFDA-labeled Lin− cells or Lin+ cells (1 × 106), and analyzed for the presence of CMFDA+ cells in the inflamed skin and BM at 6 and 12 h after the transplantation by flow cytometry; representative flow cytometric data are shown. The percentage of CMFDA+ cells in the leukocytes present in skin (a) or BM cells (b), and the numbers of CMFDA+ cells in the skin and BM per mouse are shown. (c) Flow cytometric analysis of lineage marker expression in CMFDA+ cells in inflamed skin and BM at 6 h post-transplantation of CMFDA-labelled CD45.1+Lin− or Lin+ cells. Representative data are shown. (d,e) Flow cytometric analysis of CD45.1+ cells in inflamed skin (d) and BM (e) 14 days post-transplantation of CD45.1+Lin− or Lin+ cells; representative data are shown. Data were processed as described in (a,b). Data shown (a-e) are representative of more than three independent experiments (n = 3 mice/group/experiment). Data (a,b and d,e) are presented as means ±SEM. P values were determined using two-tailed unpaired Student’s t-tests; ns, not significant, *P < 0.05, **P < 0.01.
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f3: Early homing and expansion of Lin− cells at the site of inflammation and BM.(a–c) Flow cytometric analysis of CMFDA+ cells in inflamed skin and BM at 6 and 12 h after transplantation of CMFDA+CD45.1+Lin− cells. Dermatitis or vehicle-control CD45.2+ B6 mice were transplanted with CMFDA-labeled Lin− cells or Lin+ cells (1 × 106), and analyzed for the presence of CMFDA+ cells in the inflamed skin and BM at 6 and 12 h after the transplantation by flow cytometry; representative flow cytometric data are shown. The percentage of CMFDA+ cells in the leukocytes present in skin (a) or BM cells (b), and the numbers of CMFDA+ cells in the skin and BM per mouse are shown. (c) Flow cytometric analysis of lineage marker expression in CMFDA+ cells in inflamed skin and BM at 6 h post-transplantation of CMFDA-labelled CD45.1+Lin− or Lin+ cells. Representative data are shown. (d,e) Flow cytometric analysis of CD45.1+ cells in inflamed skin (d) and BM (e) 14 days post-transplantation of CD45.1+Lin− or Lin+ cells; representative data are shown. Data were processed as described in (a,b). Data shown (a-e) are representative of more than three independent experiments (n = 3 mice/group/experiment). Data (a,b and d,e) are presented as means ±SEM. P values were determined using two-tailed unpaired Student’s t-tests; ns, not significant, *P < 0.05, **P < 0.01.

Mentions: The similar expansion kinetics of Lin− progenies in the inflamed skin and BM suggested that Lin− cells home to the skin and BM almost immediately after transplantation, although the signals from BM were rarely detected in the ventral BLI data at early time points (data not shown). To examine this on a single-cell basis, we performed flow cytometric analysis of cells infiltrating the inflamed skin and BM of CD45.2+ B6 dermatitis mice following transplantation of 5-chloromethylfluorescein diacetate (CMFDA)-labeled CD45.1+Lin− cells. CMFDA-positive (CMFDA+) cells were detected among the cells infiltrating the inflamed skin and in the BM of the upper tibia at 6 and 12 h post-transplantation (Fig. 3a,b), and even after the recipient mice were perfused with PBS prior to the analysis at 12 hr, to exclude the cells in blood (Fig. S3, Supplementary). The CMFDA+ cells in the inflamed skin were of lineage marker-negative status at 6 h (Fig. 3c). Taken together, these data indicate early or immediate homing of Lin− cells, following upon transplantation into inflamed sites as well as the BM.


Alleviation of skin inflammation after Lin(-) cell transplantation correlates with their differentiation into myeloid-derived suppressor cells.

Jeong Ryu S, Ju JM, Kim W, Bum Kim M, Hee Oh K, Sup Lee D, Lee H, Eun Oh J, Soo Park K, Young Choi E - Sci Rep (2015)

Early homing and expansion of Lin− cells at the site of inflammation and BM.(a–c) Flow cytometric analysis of CMFDA+ cells in inflamed skin and BM at 6 and 12 h after transplantation of CMFDA+CD45.1+Lin− cells. Dermatitis or vehicle-control CD45.2+ B6 mice were transplanted with CMFDA-labeled Lin− cells or Lin+ cells (1 × 106), and analyzed for the presence of CMFDA+ cells in the inflamed skin and BM at 6 and 12 h after the transplantation by flow cytometry; representative flow cytometric data are shown. The percentage of CMFDA+ cells in the leukocytes present in skin (a) or BM cells (b), and the numbers of CMFDA+ cells in the skin and BM per mouse are shown. (c) Flow cytometric analysis of lineage marker expression in CMFDA+ cells in inflamed skin and BM at 6 h post-transplantation of CMFDA-labelled CD45.1+Lin− or Lin+ cells. Representative data are shown. (d,e) Flow cytometric analysis of CD45.1+ cells in inflamed skin (d) and BM (e) 14 days post-transplantation of CD45.1+Lin− or Lin+ cells; representative data are shown. Data were processed as described in (a,b). Data shown (a-e) are representative of more than three independent experiments (n = 3 mice/group/experiment). Data (a,b and d,e) are presented as means ±SEM. P values were determined using two-tailed unpaired Student’s t-tests; ns, not significant, *P < 0.05, **P < 0.01.
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Related In: Results  -  Collection

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f3: Early homing and expansion of Lin− cells at the site of inflammation and BM.(a–c) Flow cytometric analysis of CMFDA+ cells in inflamed skin and BM at 6 and 12 h after transplantation of CMFDA+CD45.1+Lin− cells. Dermatitis or vehicle-control CD45.2+ B6 mice were transplanted with CMFDA-labeled Lin− cells or Lin+ cells (1 × 106), and analyzed for the presence of CMFDA+ cells in the inflamed skin and BM at 6 and 12 h after the transplantation by flow cytometry; representative flow cytometric data are shown. The percentage of CMFDA+ cells in the leukocytes present in skin (a) or BM cells (b), and the numbers of CMFDA+ cells in the skin and BM per mouse are shown. (c) Flow cytometric analysis of lineage marker expression in CMFDA+ cells in inflamed skin and BM at 6 h post-transplantation of CMFDA-labelled CD45.1+Lin− or Lin+ cells. Representative data are shown. (d,e) Flow cytometric analysis of CD45.1+ cells in inflamed skin (d) and BM (e) 14 days post-transplantation of CD45.1+Lin− or Lin+ cells; representative data are shown. Data were processed as described in (a,b). Data shown (a-e) are representative of more than three independent experiments (n = 3 mice/group/experiment). Data (a,b and d,e) are presented as means ±SEM. P values were determined using two-tailed unpaired Student’s t-tests; ns, not significant, *P < 0.05, **P < 0.01.
Mentions: The similar expansion kinetics of Lin− progenies in the inflamed skin and BM suggested that Lin− cells home to the skin and BM almost immediately after transplantation, although the signals from BM were rarely detected in the ventral BLI data at early time points (data not shown). To examine this on a single-cell basis, we performed flow cytometric analysis of cells infiltrating the inflamed skin and BM of CD45.2+ B6 dermatitis mice following transplantation of 5-chloromethylfluorescein diacetate (CMFDA)-labeled CD45.1+Lin− cells. CMFDA-positive (CMFDA+) cells were detected among the cells infiltrating the inflamed skin and in the BM of the upper tibia at 6 and 12 h post-transplantation (Fig. 3a,b), and even after the recipient mice were perfused with PBS prior to the analysis at 12 hr, to exclude the cells in blood (Fig. S3, Supplementary). The CMFDA+ cells in the inflamed skin were of lineage marker-negative status at 6 h (Fig. 3c). Taken together, these data indicate early or immediate homing of Lin− cells, following upon transplantation into inflamed sites as well as the BM.

Bottom Line: Longitudinal in vivo imaging and flow cytometry analyses revealed that Lin(-) cells home directly to inflamed skin within 6 h, where they undergo extensive expansion with the peak on day 14 post-transplantation, and preferential differentiation into CD11b(+)Ly6G(int)Ly6C(+) cells by day 7.Cells with phenotypic profiles of neutrophils, macrophages, and DCs appeared in inflamed skin on day 14.Progenies of transplanted Lin(-) cells showed similar kinetics of expansion and myeloid differentiation in BM.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, Korea.

ABSTRACT
To understand the cellular mechanism underlying the therapeutic effects exerted by hematopoietic stem cell transplantation in the repair of tissue damage, we investigated the in vivo dynamics of bone marrow (BM) lineage-negative (Lin(-)) cells transplanted into mice with hyper sensitivity dermatitis. Longitudinal in vivo imaging and flow cytometry analyses revealed that Lin(-) cells home directly to inflamed skin within 6 h, where they undergo extensive expansion with the peak on day 14 post-transplantation, and preferential differentiation into CD11b(+)Ly6G(int)Ly6C(+) cells by day 7. Cells with phenotypic profiles of neutrophils, macrophages, and DCs appeared in inflamed skin on day 14. Progenies of transplanted Lin(-) cells showed similar kinetics of expansion and myeloid differentiation in BM. However, differentiation into CD11b(+)Ly6G(int)Ly6C(+) cells in the inflamed skin on day 7 was more skewed toward CD115(+) cells (≥60%) with immune suppressive function and higher expression levels of iNOS, arginase, and IL-10, compared with those in the BM. Transplantation of Lin(-) cells reduced the levels of Cd3 transcript and CD4(+)/CD8(+) cells in inflamed skin. These results demonstrate differentiation of transplanted Lin(-) cells into myeloid-derived suppressor cells in inflamed skin to be the basis of the alleviation of skin inflammation after Lin(-) cell transplantation.

No MeSH data available.


Related in: MedlinePlus