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Hypergravity Provokes a Temporary Reduction in CD4+CD8+ Thymocyte Number and a Persistent Decrease in Medullary Thymic Epithelial Cell Frequency in Mice.

Tateishi R, Akiyama N, Miyauchi M, Yoshinaga R, Sasanuma H, Kudo T, Shimbo M, Shinohara M, Obata K, Inoue J, Shirakawa M, Shiba D, Asahara H, Yoshida N, Takahashi S, Morita H, Akiyama T - PLoS ONE (2015)

Bottom Line: We investigated the effects of hypergravity (2G) on murine thymic cells.Interestingly, a surgical lesion of the inner ear's vestibular apparatus inhibited these hypergravity effects.Overall, data suggest that 2G causes a temporary reduction of DP and a persistent reduction of mature mTECs in a vestibular system-dependent manner, and also dysregulates mTEC gene expression without involving the vestibular system.

View Article: PubMed Central - PubMed

Affiliation: Division of Cellular and Molecular Biology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan; Mouse epigenetics project, ISS/Kibo experiment, Japan Aerospace Exploration Agency, JAXA, Tsukuba, Japan.

ABSTRACT
Gravity change affects many immunological systems. We investigated the effects of hypergravity (2G) on murine thymic cells. Exposure of mice to 2G for three days reduced the frequency of CD4+CD8+ thymocytes (DP) and mature medullary thymic epithelial cells (mTECs), accompanied by an increment of keratin-5 and keratin-8 double-positive (K5+K8+) TECs that reportedly contain TEC progenitors. Whereas the reduction of DP was recovered by a 14-day exposure to 2G, the reduction of mature mTECs and the increment of K5+K8+ TEC persisted. Interestingly, a surgical lesion of the inner ear's vestibular apparatus inhibited these hypergravity effects. Quantitative PCR analysis revealed that the gene expression of Aire and RANK that are critical for mTEC function and development were up-regulated by the 3-day exposure and subsequently down-regulated by the 14-day exposure to 2G. Unexpectedly, this dynamic change in mTEC gene expression was independent of the vestibular apparatus. Overall, data suggest that 2G causes a temporary reduction of DP and a persistent reduction of mature mTECs in a vestibular system-dependent manner, and also dysregulates mTEC gene expression without involving the vestibular system. These data might provide insight on the impact of gravity change on thymic functions during spaceflight and living.

No MeSH data available.


Related in: MedlinePlus

Expressions of mTEC-related molecules in the thymus and plasma of mice exposed to 2G.(A) Expressions of RANK, Aire, and RANKL in the whole thymus of mice exposed to 2G (2G) for 3 days. “C” indicates 1G control. Vestibular apparatus are surgically disrupted in some groups of mice (labeled as VL). Expression of RANK (Rank), Aire (Aire), and RANKL (Rankl) mRNAs was evaluated by qPCR analysis. N = 5 each C, 2G, C with VL, and 2G with VL groups. The asterisks indicate statistical significance at **P < 0.01 and *P < 0.05 (Student’s t-test). (B) Expressions of RANK and Aire in the whole thymus of mice exposed to 2G for 14 days. Expression of RANK and Aire mRNAs was evaluated by qPCR analysis. N = 6 each C, C with VL, and 2G with VL groups. N = 5 for 2G. The asterisks indicate statistical significance at *P < 0.05, *P < 0.01, and ***P < 0.001 (Student’s t-test). NS indicates that the difference is not significant (Student’s t-test). (C) RANKL and OPG protein levels in plasma of mice exposed to 2G for 3 days. Plasma concentration of RANKL (left) and OPG (middle) protein was determined by ELISA. Ratio of RANKL to OPG (RANKL/OPG) was exhibited in the right figure. N = 5 each C, C with VL, and 2G with VL groups. N = 4 for 2G. The asterisks indicate statistical significance at **P < 0.01 (Student’s t-test). (D) Corticosterone level in plasma of mice exposed to 2G for 3 days. Concentration of corticosterone was determined by ELISA. N = 6 each C, G, C with VL, and 2G with VL groups. NS indicates that the difference is not significant (Student’s t-test).
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pone.0141650.g007: Expressions of mTEC-related molecules in the thymus and plasma of mice exposed to 2G.(A) Expressions of RANK, Aire, and RANKL in the whole thymus of mice exposed to 2G (2G) for 3 days. “C” indicates 1G control. Vestibular apparatus are surgically disrupted in some groups of mice (labeled as VL). Expression of RANK (Rank), Aire (Aire), and RANKL (Rankl) mRNAs was evaluated by qPCR analysis. N = 5 each C, 2G, C with VL, and 2G with VL groups. The asterisks indicate statistical significance at **P < 0.01 and *P < 0.05 (Student’s t-test). (B) Expressions of RANK and Aire in the whole thymus of mice exposed to 2G for 14 days. Expression of RANK and Aire mRNAs was evaluated by qPCR analysis. N = 6 each C, C with VL, and 2G with VL groups. N = 5 for 2G. The asterisks indicate statistical significance at *P < 0.05, *P < 0.01, and ***P < 0.001 (Student’s t-test). NS indicates that the difference is not significant (Student’s t-test). (C) RANKL and OPG protein levels in plasma of mice exposed to 2G for 3 days. Plasma concentration of RANKL (left) and OPG (middle) protein was determined by ELISA. Ratio of RANKL to OPG (RANKL/OPG) was exhibited in the right figure. N = 5 each C, C with VL, and 2G with VL groups. N = 4 for 2G. The asterisks indicate statistical significance at **P < 0.01 (Student’s t-test). (D) Corticosterone level in plasma of mice exposed to 2G for 3 days. Concentration of corticosterone was determined by ELISA. N = 6 each C, G, C with VL, and 2G with VL groups. NS indicates that the difference is not significant (Student’s t-test).

Mentions: Previous studies revealed that the development and maintenance of mTECs depend on TNF receptor family signaling [37]. In particular, TNF receptor family RANK signaling plays a critical role in the development of Aire-expressing mTECs [38–40], thereby suppressing the onset of autoimmunity. We investigated the expression level of Aire and RANK in the whole thymus of mice exposed to hypergravity. Exposure to 2G for 3 days significantly increased RANK expression (Fig 7A). Unexpectedly, the up-regulation of RANK expression was observed in the thymus of VL mice. Aire expression was also up-regulated by the 2G exposure in VL mice (Fig 7A) although an increment in the 2G-treated mice did not reach statistical significance. Thus, the 2G-dependent up-regulation of RANK and Aire expression occurs without the vestibular apparatus sensing gravity and therefore can not explained simply by the relative increase in mTEC frequency due to the reduction in DP number, which is dependent on the vestibular apparatus (Fig 1). Surprisingly, in contrast to the 3 day-exposure, 2G exposure for 14 days rather caused a reduction of Aire and RANK expression in the thymus (Fig 5B). Moreover, this 2G-mediated reduction in gene expression is independent of VL. Thus, the reduction of RANK and Aire expression caused by the 14-day 2G exposure also can not explained only by the reduction of mTEChi number, which is also dependent on the vestibular apparatus (Fig 5). Consequently, these data suggest that 2G exposure caused an early increment and subsequent reduction of mTEC gene expression without any involvement of the vestibular apparatus.


Hypergravity Provokes a Temporary Reduction in CD4+CD8+ Thymocyte Number and a Persistent Decrease in Medullary Thymic Epithelial Cell Frequency in Mice.

Tateishi R, Akiyama N, Miyauchi M, Yoshinaga R, Sasanuma H, Kudo T, Shimbo M, Shinohara M, Obata K, Inoue J, Shirakawa M, Shiba D, Asahara H, Yoshida N, Takahashi S, Morita H, Akiyama T - PLoS ONE (2015)

Expressions of mTEC-related molecules in the thymus and plasma of mice exposed to 2G.(A) Expressions of RANK, Aire, and RANKL in the whole thymus of mice exposed to 2G (2G) for 3 days. “C” indicates 1G control. Vestibular apparatus are surgically disrupted in some groups of mice (labeled as VL). Expression of RANK (Rank), Aire (Aire), and RANKL (Rankl) mRNAs was evaluated by qPCR analysis. N = 5 each C, 2G, C with VL, and 2G with VL groups. The asterisks indicate statistical significance at **P < 0.01 and *P < 0.05 (Student’s t-test). (B) Expressions of RANK and Aire in the whole thymus of mice exposed to 2G for 14 days. Expression of RANK and Aire mRNAs was evaluated by qPCR analysis. N = 6 each C, C with VL, and 2G with VL groups. N = 5 for 2G. The asterisks indicate statistical significance at *P < 0.05, *P < 0.01, and ***P < 0.001 (Student’s t-test). NS indicates that the difference is not significant (Student’s t-test). (C) RANKL and OPG protein levels in plasma of mice exposed to 2G for 3 days. Plasma concentration of RANKL (left) and OPG (middle) protein was determined by ELISA. Ratio of RANKL to OPG (RANKL/OPG) was exhibited in the right figure. N = 5 each C, C with VL, and 2G with VL groups. N = 4 for 2G. The asterisks indicate statistical significance at **P < 0.01 (Student’s t-test). (D) Corticosterone level in plasma of mice exposed to 2G for 3 days. Concentration of corticosterone was determined by ELISA. N = 6 each C, G, C with VL, and 2G with VL groups. NS indicates that the difference is not significant (Student’s t-test).
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pone.0141650.g007: Expressions of mTEC-related molecules in the thymus and plasma of mice exposed to 2G.(A) Expressions of RANK, Aire, and RANKL in the whole thymus of mice exposed to 2G (2G) for 3 days. “C” indicates 1G control. Vestibular apparatus are surgically disrupted in some groups of mice (labeled as VL). Expression of RANK (Rank), Aire (Aire), and RANKL (Rankl) mRNAs was evaluated by qPCR analysis. N = 5 each C, 2G, C with VL, and 2G with VL groups. The asterisks indicate statistical significance at **P < 0.01 and *P < 0.05 (Student’s t-test). (B) Expressions of RANK and Aire in the whole thymus of mice exposed to 2G for 14 days. Expression of RANK and Aire mRNAs was evaluated by qPCR analysis. N = 6 each C, C with VL, and 2G with VL groups. N = 5 for 2G. The asterisks indicate statistical significance at *P < 0.05, *P < 0.01, and ***P < 0.001 (Student’s t-test). NS indicates that the difference is not significant (Student’s t-test). (C) RANKL and OPG protein levels in plasma of mice exposed to 2G for 3 days. Plasma concentration of RANKL (left) and OPG (middle) protein was determined by ELISA. Ratio of RANKL to OPG (RANKL/OPG) was exhibited in the right figure. N = 5 each C, C with VL, and 2G with VL groups. N = 4 for 2G. The asterisks indicate statistical significance at **P < 0.01 (Student’s t-test). (D) Corticosterone level in plasma of mice exposed to 2G for 3 days. Concentration of corticosterone was determined by ELISA. N = 6 each C, G, C with VL, and 2G with VL groups. NS indicates that the difference is not significant (Student’s t-test).
Mentions: Previous studies revealed that the development and maintenance of mTECs depend on TNF receptor family signaling [37]. In particular, TNF receptor family RANK signaling plays a critical role in the development of Aire-expressing mTECs [38–40], thereby suppressing the onset of autoimmunity. We investigated the expression level of Aire and RANK in the whole thymus of mice exposed to hypergravity. Exposure to 2G for 3 days significantly increased RANK expression (Fig 7A). Unexpectedly, the up-regulation of RANK expression was observed in the thymus of VL mice. Aire expression was also up-regulated by the 2G exposure in VL mice (Fig 7A) although an increment in the 2G-treated mice did not reach statistical significance. Thus, the 2G-dependent up-regulation of RANK and Aire expression occurs without the vestibular apparatus sensing gravity and therefore can not explained simply by the relative increase in mTEC frequency due to the reduction in DP number, which is dependent on the vestibular apparatus (Fig 1). Surprisingly, in contrast to the 3 day-exposure, 2G exposure for 14 days rather caused a reduction of Aire and RANK expression in the thymus (Fig 5B). Moreover, this 2G-mediated reduction in gene expression is independent of VL. Thus, the reduction of RANK and Aire expression caused by the 14-day 2G exposure also can not explained only by the reduction of mTEChi number, which is also dependent on the vestibular apparatus (Fig 5). Consequently, these data suggest that 2G exposure caused an early increment and subsequent reduction of mTEC gene expression without any involvement of the vestibular apparatus.

Bottom Line: We investigated the effects of hypergravity (2G) on murine thymic cells.Interestingly, a surgical lesion of the inner ear's vestibular apparatus inhibited these hypergravity effects.Overall, data suggest that 2G causes a temporary reduction of DP and a persistent reduction of mature mTECs in a vestibular system-dependent manner, and also dysregulates mTEC gene expression without involving the vestibular system.

View Article: PubMed Central - PubMed

Affiliation: Division of Cellular and Molecular Biology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan; Mouse epigenetics project, ISS/Kibo experiment, Japan Aerospace Exploration Agency, JAXA, Tsukuba, Japan.

ABSTRACT
Gravity change affects many immunological systems. We investigated the effects of hypergravity (2G) on murine thymic cells. Exposure of mice to 2G for three days reduced the frequency of CD4+CD8+ thymocytes (DP) and mature medullary thymic epithelial cells (mTECs), accompanied by an increment of keratin-5 and keratin-8 double-positive (K5+K8+) TECs that reportedly contain TEC progenitors. Whereas the reduction of DP was recovered by a 14-day exposure to 2G, the reduction of mature mTECs and the increment of K5+K8+ TEC persisted. Interestingly, a surgical lesion of the inner ear's vestibular apparatus inhibited these hypergravity effects. Quantitative PCR analysis revealed that the gene expression of Aire and RANK that are critical for mTEC function and development were up-regulated by the 3-day exposure and subsequently down-regulated by the 14-day exposure to 2G. Unexpectedly, this dynamic change in mTEC gene expression was independent of the vestibular apparatus. Overall, data suggest that 2G causes a temporary reduction of DP and a persistent reduction of mature mTECs in a vestibular system-dependent manner, and also dysregulates mTEC gene expression without involving the vestibular system. These data might provide insight on the impact of gravity change on thymic functions during spaceflight and living.

No MeSH data available.


Related in: MedlinePlus