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Adipose Mesenchymal Stem Cell Secretome Modulated in Hypoxia for Remodeling of Radiation-Induced Salivary Gland Damage.

An HY, Shin HS, Choi JS, Kim HJ, Lim JY, Kim YM - PLoS ONE (2015)

Bottom Line: Repeated systemic infusion with the hAdMSC secretome resulted in improved salivation capacity and increased levels of salivary proteins, including amylase and EGF, relative to the PBS group.The hAdMSC secretome strongly induced proliferation of HPEC and led to a significant decrease in cell death in vivo and in vitro.Moreover, the anti-apoptotic effects of the hAdMSC secretome were found to be promoted after hypoxia-preconditioning relative to normoxia-cultured hAdMSC secretome.

View Article: PubMed Central - PubMed

Affiliation: Department of Otorhinolaryngology-Head and Neck Surgery, Inha University College of Medicine, Incheon, Republic of Korea; Translational Research Center, Inha University College of Medicine, Incheon, Republic of Korea.

ABSTRACT

Background and purpose: This study was conducted to determine whether a secretome from mesenchymal stem cells (MSC) modulated by hypoxic conditions to contain therapeutic factors contributes to salivary gland (SG) tissue remodeling and has the potential to improve irradiation (IR)-induced salivary hypofunction in a mouse model.

Materials and methods: Human adipose mesenchymal stem cells (hAdMSC) were isolated, expanded, and exposed to hypoxic conditions (O2 < 5%). The hypoxia-conditioned medium was then filtered to a high molecular weight fraction and prepared as a hAdMSC secretome. The hAdMSC secretome was subsequently infused into the tail vein of C3H mice immediately after local IR once a day for seven consecutive days. The control group received equal volume (500 μL) of vehicle (PBS) only. SG function and structural tissue remodeling by the hAdMSC secretome were investigated. Human parotid epithelial cells (HPEC) were obtained, expanded in vitro, and then irradiated and treated with either the hypoxia-conditioned medium or a normoxic control medium. Cell proliferation and IR-induced cell death were examined to determine the mechanism by which the hAdMSC secretome exerted its effects.

Results: The conditioned hAdMSC secretome contained high levels of GM-CSF, VEGF, IL-6, and IGF-1. Repeated systemic infusion with the hAdMSC secretome resulted in improved salivation capacity and increased levels of salivary proteins, including amylase and EGF, relative to the PBS group. The microscopic structural integrity of SG was maintained and salivary epithelial (AQP-5), endothelial (CD31), myoepithelial (α-SMA) and SG progenitor cells (c-Kit) were successfully protected from radiation damage and remodeled. The hAdMSC secretome strongly induced proliferation of HPEC and led to a significant decrease in cell death in vivo and in vitro. Moreover, the anti-apoptotic effects of the hAdMSC secretome were found to be promoted after hypoxia-preconditioning relative to normoxia-cultured hAdMSC secretome.

Conclusion: These results show that the hAdMSC secretome from hypoxic-conditioned medium may provide radioprotection and tissue remodeling via release of paracrine mediators.

No MeSH data available.


Related in: MedlinePlus

hAdMSC secretome exerts cytoprotection via anti-apoptotic effects on salivary parenchymal cells in vivo.(A) Representative images of an in vivo TUNEL assay from three experimental groups at 1, 2, and 4 weeks post-IR. Scale bars represent 50 μm. (B) The number of TUNEL-positive apoptotic cells was determined by a blinded researcher. Data are presented as the mean number of apoptotic cells per field ± SEM. Two-way ANOVA, Bonferroni post hoc test. *, compared to CON; #, compared to IR + PBS; †, compared to IR + NMX. *P <0.01, ***P <0.001, ##P < 0.01, ###P < 0.001, ††P<0.01, †††P<0.001. CON, normal control group (n = 21 sections); IR+ PBS, PBS-treated group (n = 21 sections); IR + SEC (NMX), IR + normoxic secretome-treated group (n = 15 sections); IR + SEC (HPX), hypoxia-conditioned secretome-treated group (n = 18 sections).
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pone.0141862.g005: hAdMSC secretome exerts cytoprotection via anti-apoptotic effects on salivary parenchymal cells in vivo.(A) Representative images of an in vivo TUNEL assay from three experimental groups at 1, 2, and 4 weeks post-IR. Scale bars represent 50 μm. (B) The number of TUNEL-positive apoptotic cells was determined by a blinded researcher. Data are presented as the mean number of apoptotic cells per field ± SEM. Two-way ANOVA, Bonferroni post hoc test. *, compared to CON; #, compared to IR + PBS; †, compared to IR + NMX. *P <0.01, ***P <0.001, ##P < 0.01, ###P < 0.001, ††P<0.01, †††P<0.001. CON, normal control group (n = 21 sections); IR+ PBS, PBS-treated group (n = 21 sections); IR + SEC (NMX), IR + normoxic secretome-treated group (n = 15 sections); IR + SEC (HPX), hypoxia-conditioned secretome-treated group (n = 18 sections).

Mentions: We next explored whether these cytoprotective effects of the hAdMSC secretome were related to inhibition of cell death by MSC-secreted paracrine factors. The number of TUNEL-positive apoptotic cells after infusion of PBS, normoxia-cultured hAdMSC secretome, or hypoxia-preconditioned hAdMSC secretome was determined at 1, 2, and 4 weeks post-IR (Fig 5A). A significant increase in apoptosis was observed after IR during the observation period (Fig 5B, P < 0.001), but this decreased significantly following hAdMSC secretome treatment relative to the PBS group (P < 0.01 at 1 week in the normoxia-secretome group and P < 0.001 at 2 and 4 weeks in the normoxia-secretome and all time points in the hypoxia-secretome). Treatment with the hypoxia-conditioned hAdMSC secretome resulted in enhanced anti-apoptotic effects relative to the normoxia-secretome (P < 0.01 at 1 and 2 weeks and P < 0.001 at 4 weeks after treatment).


Adipose Mesenchymal Stem Cell Secretome Modulated in Hypoxia for Remodeling of Radiation-Induced Salivary Gland Damage.

An HY, Shin HS, Choi JS, Kim HJ, Lim JY, Kim YM - PLoS ONE (2015)

hAdMSC secretome exerts cytoprotection via anti-apoptotic effects on salivary parenchymal cells in vivo.(A) Representative images of an in vivo TUNEL assay from three experimental groups at 1, 2, and 4 weeks post-IR. Scale bars represent 50 μm. (B) The number of TUNEL-positive apoptotic cells was determined by a blinded researcher. Data are presented as the mean number of apoptotic cells per field ± SEM. Two-way ANOVA, Bonferroni post hoc test. *, compared to CON; #, compared to IR + PBS; †, compared to IR + NMX. *P <0.01, ***P <0.001, ##P < 0.01, ###P < 0.001, ††P<0.01, †††P<0.001. CON, normal control group (n = 21 sections); IR+ PBS, PBS-treated group (n = 21 sections); IR + SEC (NMX), IR + normoxic secretome-treated group (n = 15 sections); IR + SEC (HPX), hypoxia-conditioned secretome-treated group (n = 18 sections).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4631328&req=5

pone.0141862.g005: hAdMSC secretome exerts cytoprotection via anti-apoptotic effects on salivary parenchymal cells in vivo.(A) Representative images of an in vivo TUNEL assay from three experimental groups at 1, 2, and 4 weeks post-IR. Scale bars represent 50 μm. (B) The number of TUNEL-positive apoptotic cells was determined by a blinded researcher. Data are presented as the mean number of apoptotic cells per field ± SEM. Two-way ANOVA, Bonferroni post hoc test. *, compared to CON; #, compared to IR + PBS; †, compared to IR + NMX. *P <0.01, ***P <0.001, ##P < 0.01, ###P < 0.001, ††P<0.01, †††P<0.001. CON, normal control group (n = 21 sections); IR+ PBS, PBS-treated group (n = 21 sections); IR + SEC (NMX), IR + normoxic secretome-treated group (n = 15 sections); IR + SEC (HPX), hypoxia-conditioned secretome-treated group (n = 18 sections).
Mentions: We next explored whether these cytoprotective effects of the hAdMSC secretome were related to inhibition of cell death by MSC-secreted paracrine factors. The number of TUNEL-positive apoptotic cells after infusion of PBS, normoxia-cultured hAdMSC secretome, or hypoxia-preconditioned hAdMSC secretome was determined at 1, 2, and 4 weeks post-IR (Fig 5A). A significant increase in apoptosis was observed after IR during the observation period (Fig 5B, P < 0.001), but this decreased significantly following hAdMSC secretome treatment relative to the PBS group (P < 0.01 at 1 week in the normoxia-secretome group and P < 0.001 at 2 and 4 weeks in the normoxia-secretome and all time points in the hypoxia-secretome). Treatment with the hypoxia-conditioned hAdMSC secretome resulted in enhanced anti-apoptotic effects relative to the normoxia-secretome (P < 0.01 at 1 and 2 weeks and P < 0.001 at 4 weeks after treatment).

Bottom Line: Repeated systemic infusion with the hAdMSC secretome resulted in improved salivation capacity and increased levels of salivary proteins, including amylase and EGF, relative to the PBS group.The hAdMSC secretome strongly induced proliferation of HPEC and led to a significant decrease in cell death in vivo and in vitro.Moreover, the anti-apoptotic effects of the hAdMSC secretome were found to be promoted after hypoxia-preconditioning relative to normoxia-cultured hAdMSC secretome.

View Article: PubMed Central - PubMed

Affiliation: Department of Otorhinolaryngology-Head and Neck Surgery, Inha University College of Medicine, Incheon, Republic of Korea; Translational Research Center, Inha University College of Medicine, Incheon, Republic of Korea.

ABSTRACT

Background and purpose: This study was conducted to determine whether a secretome from mesenchymal stem cells (MSC) modulated by hypoxic conditions to contain therapeutic factors contributes to salivary gland (SG) tissue remodeling and has the potential to improve irradiation (IR)-induced salivary hypofunction in a mouse model.

Materials and methods: Human adipose mesenchymal stem cells (hAdMSC) were isolated, expanded, and exposed to hypoxic conditions (O2 < 5%). The hypoxia-conditioned medium was then filtered to a high molecular weight fraction and prepared as a hAdMSC secretome. The hAdMSC secretome was subsequently infused into the tail vein of C3H mice immediately after local IR once a day for seven consecutive days. The control group received equal volume (500 μL) of vehicle (PBS) only. SG function and structural tissue remodeling by the hAdMSC secretome were investigated. Human parotid epithelial cells (HPEC) were obtained, expanded in vitro, and then irradiated and treated with either the hypoxia-conditioned medium or a normoxic control medium. Cell proliferation and IR-induced cell death were examined to determine the mechanism by which the hAdMSC secretome exerted its effects.

Results: The conditioned hAdMSC secretome contained high levels of GM-CSF, VEGF, IL-6, and IGF-1. Repeated systemic infusion with the hAdMSC secretome resulted in improved salivation capacity and increased levels of salivary proteins, including amylase and EGF, relative to the PBS group. The microscopic structural integrity of SG was maintained and salivary epithelial (AQP-5), endothelial (CD31), myoepithelial (α-SMA) and SG progenitor cells (c-Kit) were successfully protected from radiation damage and remodeled. The hAdMSC secretome strongly induced proliferation of HPEC and led to a significant decrease in cell death in vivo and in vitro. Moreover, the anti-apoptotic effects of the hAdMSC secretome were found to be promoted after hypoxia-preconditioning relative to normoxia-cultured hAdMSC secretome.

Conclusion: These results show that the hAdMSC secretome from hypoxic-conditioned medium may provide radioprotection and tissue remodeling via release of paracrine mediators.

No MeSH data available.


Related in: MedlinePlus