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Dendritic Cells Regulate Treg-Th17 Axis in Obstructive Phase of Bile Duct Injury in Murine Biliary Atresia.

Liu YJ, Li K, Yang L, Tang ST, Wang XX, Cao GQ, Li S, Lei HY, Zhang X - PLoS ONE (2015)

Bottom Line: By depleting the Th17 cells, the BA symptoms (onset of jaundice, acholic stools and retarded growth) were attenuated and the survival rate was improved.The level of IL-6, which has been shown to be abundantly secreted by activated dendritic cells (DCs), was remarkably elevated.In conclusion, the DC-regulated Treg-Th17 axis, probably in conjunction with other effector T cells, aggravates progressive inflammatory injury at the time of ductal obstruction.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatric Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.

ABSTRACT
Several cell types are considered to be effector cells in bile duct injury in rhesus rotavirus (RRV)-induced experimental biliary atresia (BA). Here, we identified an increased T helper 17 (Th17) cell population in a BA mode. By depleting the Th17 cells, the BA symptoms (onset of jaundice, acholic stools and retarded growth) were attenuated and the survival rate was improved. Furthermore, we found that in mice with BA, the percentage of CD4+CD25highFoxp3+ T regulatory (Treg) cells decreased along with the increased percentage of Th17 cells. However, the absolute numbers of Treg and Th17 cells were both increased in liver of RRV-injected mice compared to saline-injected mice. The proportion of Th17 cells at 7 days post-infection was decreased if Treg cells isolated from normal adult mice, but not Treg cells from the livers of mice with BA, were intraperitoneally transferred on day 5 of life. In vitro experiments also showed that Treg cells from mice with BA had a diminished suppressive effect on Th17 cell generation. To determine the mechanisms, we investigated the production of cytokines in the liver. The level of IL-6, which has been shown to be abundantly secreted by activated dendritic cells (DCs), was remarkably elevated. Importantly, in a Treg/Th17 cell suppression assay, IL-6 was demonstrated to paralyze the Treg cells' suppressive effect on Th17 cells and eventually the unrestrained increase of Th17 cells contributed to bile duct injury. In conclusion, the DC-regulated Treg-Th17 axis, probably in conjunction with other effector T cells, aggravates progressive inflammatory injury at the time of ductal obstruction.

No MeSH data available.


Related in: MedlinePlus

Activated DC produces abundant IL-6, hinders the Treg suppression function and promotes Th17 generation ex vivo.(A) DCs were isolated from mice liver on the 7th day after injection of RRV or saline and within 12 hrs of birth. DCs were added to the differentiation assay containing 3 ng/ml TGF-β, 20 ng/ml IL-6 and the other indicated cytokines, and a stimulus (vide supra). After 7 days, the percentage of Th17 was measured by FCM and the data is reported as the mean ± SEM (n = 12). * P<0.05, ** P<0.01. (B) Tregs were isolated from liver and spleen of mice on the 7th day after injection of RRV or saline within 12 hrs of birth. Different groups of Tregs were pretreated by co-culturing with DCs obtained from the liver of either RRV or saline injected mice, before adding them to the suppression assay as described above. In group F, Tregs were pretreated with supernatant from RRV-primed DC + IL-6 Ab. The production of IL-17A in the supernatant was measured by ELISA and is reported as the mean ± SEM (n = 11). * P<0.05, ** P<0.01.
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pone.0136214.g007: Activated DC produces abundant IL-6, hinders the Treg suppression function and promotes Th17 generation ex vivo.(A) DCs were isolated from mice liver on the 7th day after injection of RRV or saline and within 12 hrs of birth. DCs were added to the differentiation assay containing 3 ng/ml TGF-β, 20 ng/ml IL-6 and the other indicated cytokines, and a stimulus (vide supra). After 7 days, the percentage of Th17 was measured by FCM and the data is reported as the mean ± SEM (n = 12). * P<0.05, ** P<0.01. (B) Tregs were isolated from liver and spleen of mice on the 7th day after injection of RRV or saline within 12 hrs of birth. Different groups of Tregs were pretreated by co-culturing with DCs obtained from the liver of either RRV or saline injected mice, before adding them to the suppression assay as described above. In group F, Tregs were pretreated with supernatant from RRV-primed DC + IL-6 Ab. The production of IL-17A in the supernatant was measured by ELISA and is reported as the mean ± SEM (n = 11). * P<0.05, ** P<0.01.

Mentions: To address the physiological role of IL-6, we established a modified Th17 differentiation assay. In this assay, adding an excess of IL-6 (30 ng/ml) markedly increased the percentage of Th17 cells in the culture system containing naïve CD4+ T cells and TGF-β, together with the indicated cytokines and stimulus (P = 0.005, Fig 7A, column C vs B). When we added DCs (or their supernatant) from RRV-primed mice instead of IL-6, the percentage of Th17 cells was maintained (P = 0.408, Fig 7A, groups D and G compared with group C). In contrast, when DCs from saline-injected rather than RRV-injected mice were added, the Th17 cell percentage dropped to 1.1% (P = 0.012, Fig 7A, column E vs D). If RRV-primed DCs were added together with 20mg anti-IL-6 Ab (ab6672, Abcam, Cambridge, UK), Th17 cell production was also significantly decreased, comparable to what was observed for RRV-primed DCs alone (2.34 ± 0.94% vs 8.13 ± 2.56%, P<0.01, Fig 7A, column F vs D). These data indicate that the soluble cytokine IL-6, secreted by RRV-activated DCs rather than inactivated DCs, plays a vital role in Th17 differentiation.


Dendritic Cells Regulate Treg-Th17 Axis in Obstructive Phase of Bile Duct Injury in Murine Biliary Atresia.

Liu YJ, Li K, Yang L, Tang ST, Wang XX, Cao GQ, Li S, Lei HY, Zhang X - PLoS ONE (2015)

Activated DC produces abundant IL-6, hinders the Treg suppression function and promotes Th17 generation ex vivo.(A) DCs were isolated from mice liver on the 7th day after injection of RRV or saline and within 12 hrs of birth. DCs were added to the differentiation assay containing 3 ng/ml TGF-β, 20 ng/ml IL-6 and the other indicated cytokines, and a stimulus (vide supra). After 7 days, the percentage of Th17 was measured by FCM and the data is reported as the mean ± SEM (n = 12). * P<0.05, ** P<0.01. (B) Tregs were isolated from liver and spleen of mice on the 7th day after injection of RRV or saline within 12 hrs of birth. Different groups of Tregs were pretreated by co-culturing with DCs obtained from the liver of either RRV or saline injected mice, before adding them to the suppression assay as described above. In group F, Tregs were pretreated with supernatant from RRV-primed DC + IL-6 Ab. The production of IL-17A in the supernatant was measured by ELISA and is reported as the mean ± SEM (n = 11). * P<0.05, ** P<0.01.
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Related In: Results  -  Collection

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pone.0136214.g007: Activated DC produces abundant IL-6, hinders the Treg suppression function and promotes Th17 generation ex vivo.(A) DCs were isolated from mice liver on the 7th day after injection of RRV or saline and within 12 hrs of birth. DCs were added to the differentiation assay containing 3 ng/ml TGF-β, 20 ng/ml IL-6 and the other indicated cytokines, and a stimulus (vide supra). After 7 days, the percentage of Th17 was measured by FCM and the data is reported as the mean ± SEM (n = 12). * P<0.05, ** P<0.01. (B) Tregs were isolated from liver and spleen of mice on the 7th day after injection of RRV or saline within 12 hrs of birth. Different groups of Tregs were pretreated by co-culturing with DCs obtained from the liver of either RRV or saline injected mice, before adding them to the suppression assay as described above. In group F, Tregs were pretreated with supernatant from RRV-primed DC + IL-6 Ab. The production of IL-17A in the supernatant was measured by ELISA and is reported as the mean ± SEM (n = 11). * P<0.05, ** P<0.01.
Mentions: To address the physiological role of IL-6, we established a modified Th17 differentiation assay. In this assay, adding an excess of IL-6 (30 ng/ml) markedly increased the percentage of Th17 cells in the culture system containing naïve CD4+ T cells and TGF-β, together with the indicated cytokines and stimulus (P = 0.005, Fig 7A, column C vs B). When we added DCs (or their supernatant) from RRV-primed mice instead of IL-6, the percentage of Th17 cells was maintained (P = 0.408, Fig 7A, groups D and G compared with group C). In contrast, when DCs from saline-injected rather than RRV-injected mice were added, the Th17 cell percentage dropped to 1.1% (P = 0.012, Fig 7A, column E vs D). If RRV-primed DCs were added together with 20mg anti-IL-6 Ab (ab6672, Abcam, Cambridge, UK), Th17 cell production was also significantly decreased, comparable to what was observed for RRV-primed DCs alone (2.34 ± 0.94% vs 8.13 ± 2.56%, P<0.01, Fig 7A, column F vs D). These data indicate that the soluble cytokine IL-6, secreted by RRV-activated DCs rather than inactivated DCs, plays a vital role in Th17 differentiation.

Bottom Line: By depleting the Th17 cells, the BA symptoms (onset of jaundice, acholic stools and retarded growth) were attenuated and the survival rate was improved.The level of IL-6, which has been shown to be abundantly secreted by activated dendritic cells (DCs), was remarkably elevated.In conclusion, the DC-regulated Treg-Th17 axis, probably in conjunction with other effector T cells, aggravates progressive inflammatory injury at the time of ductal obstruction.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatric Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.

ABSTRACT
Several cell types are considered to be effector cells in bile duct injury in rhesus rotavirus (RRV)-induced experimental biliary atresia (BA). Here, we identified an increased T helper 17 (Th17) cell population in a BA mode. By depleting the Th17 cells, the BA symptoms (onset of jaundice, acholic stools and retarded growth) were attenuated and the survival rate was improved. Furthermore, we found that in mice with BA, the percentage of CD4+CD25highFoxp3+ T regulatory (Treg) cells decreased along with the increased percentage of Th17 cells. However, the absolute numbers of Treg and Th17 cells were both increased in liver of RRV-injected mice compared to saline-injected mice. The proportion of Th17 cells at 7 days post-infection was decreased if Treg cells isolated from normal adult mice, but not Treg cells from the livers of mice with BA, were intraperitoneally transferred on day 5 of life. In vitro experiments also showed that Treg cells from mice with BA had a diminished suppressive effect on Th17 cell generation. To determine the mechanisms, we investigated the production of cytokines in the liver. The level of IL-6, which has been shown to be abundantly secreted by activated dendritic cells (DCs), was remarkably elevated. Importantly, in a Treg/Th17 cell suppression assay, IL-6 was demonstrated to paralyze the Treg cells' suppressive effect on Th17 cells and eventually the unrestrained increase of Th17 cells contributed to bile duct injury. In conclusion, the DC-regulated Treg-Th17 axis, probably in conjunction with other effector T cells, aggravates progressive inflammatory injury at the time of ductal obstruction.

No MeSH data available.


Related in: MedlinePlus