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Hyperosmotic stress activates the expression of members of the miR-15/107 family and induces downregulation of anti-apoptotic genes in rat liver.

Santosa D, Castoldi M, Paluschinski M, Sommerfeld A, Häussinger D - Sci Rep (2015)

Bottom Line: It was also identified that hyperosmolarity significantly reduces the expression of anti-apoptotic genes including Bcl2, Ccnd1, Mcl1, Faim, Aatf, Bfar and Ikbkb, which are either validated or predicted targets of these microRNAs.Moreover, through the application of NOX and JNK inhibitors as well as benzylamine it is shown that the observed response is mediated by reactive oxygen species (ROS), suggesting that miR-15a, miR-15b and miR-16 are novel redoximiRs.It is concluded that the response of these three microRNAs to osmotic stress is ROS-mediated and that it might contribute to the development of a proapoptotic phenotype.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, Hepatology and Infectious Diseases, Heinrich-Heine-University, Moorenstrasse 5, 40225 Düsseldorf, Germany.

ABSTRACT
microRNAs are an abundant class of small non-coding RNAs that negatively regulate gene expression. Importantly, microRNA activity has been linked to the control of cellular stress response. In the present study, we investigated whether the expression of hepatic microRNAs is affected by changes in ambient osmolarity. It is shown that hyperosmotic exposure of perfused rat liver induces a rapid upregulation of miR-15a, miR-15b and miR-16, which are members of the miR-15/107 microRNAs superfamily. It was also identified that hyperosmolarity significantly reduces the expression of anti-apoptotic genes including Bcl2, Ccnd1, Mcl1, Faim, Aatf, Bfar and Ikbkb, which are either validated or predicted targets of these microRNAs. Moreover, through the application of NOX and JNK inhibitors as well as benzylamine it is shown that the observed response is mediated by reactive oxygen species (ROS), suggesting that miR-15a, miR-15b and miR-16 are novel redoximiRs. It is concluded that the response of these three microRNAs to osmotic stress is ROS-mediated and that it might contribute to the development of a proapoptotic phenotype.

No MeSH data available.


Related in: MedlinePlus

Bcl2 is not significantly changed at the mRNA level in hyperosmotically perfused rat livers after addition of apocynin.(a) Bcl2 is not significantly changed by hyperosmolarity after addition of apocynin. (b) Downregulation of Ccnd1 under hyperosmotic exposure is attenuated by administration of apocynin. (c) Mcl1 is not significantly changed in both normoosmotic and hyperosmotic conditions, when apocynin is added. (d) Faim is significantly downregulated after 180 minutes of hyperosmotic stimulation and apocynin, while it is not significantly changed after 120 minutes in the same experiment. qPCR runs were normalized according to the ΔΔCt method using β-Tubulin as reference gene. Statistical analysis was carried out by unpaired student’s t-test. Data are shown as average ± S.E.M. of 3 independent experiments. The values of unstimulated controls (T0) were set arbitrarily to 100.
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f4: Bcl2 is not significantly changed at the mRNA level in hyperosmotically perfused rat livers after addition of apocynin.(a) Bcl2 is not significantly changed by hyperosmolarity after addition of apocynin. (b) Downregulation of Ccnd1 under hyperosmotic exposure is attenuated by administration of apocynin. (c) Mcl1 is not significantly changed in both normoosmotic and hyperosmotic conditions, when apocynin is added. (d) Faim is significantly downregulated after 180 minutes of hyperosmotic stimulation and apocynin, while it is not significantly changed after 120 minutes in the same experiment. qPCR runs were normalized according to the ΔΔCt method using β-Tubulin as reference gene. Statistical analysis was carried out by unpaired student’s t-test. Data are shown as average ± S.E.M. of 3 independent experiments. The values of unstimulated controls (T0) were set arbitrarily to 100.

Mentions: In order to evaluate whether miRNA expression was regulated at the transcriptional level, expression of miR-15a precursor (pre-miR-15a) was measured by using miQPCR. Our analysis identified that pre-miR-15a was significantly upregulated under hyperosmotic conditions (Supplementary Figure 3a), while this effect was blocked by administration of apocynin (Fig. 3d). These data point to a transcriptional regulation of miR-15a expression. Correspondingly, the finding that apocynin was also able to largely prevent the hyperosmolarity-induced downregulation of Bcl2, Ccnd1, Mcl1 and Faim (Fig. 4) confirms the inverse correlation between the levels of these miRNAs and the expression of anti-apoptotic genes. These findings suggest that NOX activity is required to mediate both the upregulation of miR-15a, -15b and -16 and the downregulation of Bcl2, Ccnd1, Mcl1 and Faim under hyperosmotic conditions.


Hyperosmotic stress activates the expression of members of the miR-15/107 family and induces downregulation of anti-apoptotic genes in rat liver.

Santosa D, Castoldi M, Paluschinski M, Sommerfeld A, Häussinger D - Sci Rep (2015)

Bcl2 is not significantly changed at the mRNA level in hyperosmotically perfused rat livers after addition of apocynin.(a) Bcl2 is not significantly changed by hyperosmolarity after addition of apocynin. (b) Downregulation of Ccnd1 under hyperosmotic exposure is attenuated by administration of apocynin. (c) Mcl1 is not significantly changed in both normoosmotic and hyperosmotic conditions, when apocynin is added. (d) Faim is significantly downregulated after 180 minutes of hyperosmotic stimulation and apocynin, while it is not significantly changed after 120 minutes in the same experiment. qPCR runs were normalized according to the ΔΔCt method using β-Tubulin as reference gene. Statistical analysis was carried out by unpaired student’s t-test. Data are shown as average ± S.E.M. of 3 independent experiments. The values of unstimulated controls (T0) were set arbitrarily to 100.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4508667&req=5

f4: Bcl2 is not significantly changed at the mRNA level in hyperosmotically perfused rat livers after addition of apocynin.(a) Bcl2 is not significantly changed by hyperosmolarity after addition of apocynin. (b) Downregulation of Ccnd1 under hyperosmotic exposure is attenuated by administration of apocynin. (c) Mcl1 is not significantly changed in both normoosmotic and hyperosmotic conditions, when apocynin is added. (d) Faim is significantly downregulated after 180 minutes of hyperosmotic stimulation and apocynin, while it is not significantly changed after 120 minutes in the same experiment. qPCR runs were normalized according to the ΔΔCt method using β-Tubulin as reference gene. Statistical analysis was carried out by unpaired student’s t-test. Data are shown as average ± S.E.M. of 3 independent experiments. The values of unstimulated controls (T0) were set arbitrarily to 100.
Mentions: In order to evaluate whether miRNA expression was regulated at the transcriptional level, expression of miR-15a precursor (pre-miR-15a) was measured by using miQPCR. Our analysis identified that pre-miR-15a was significantly upregulated under hyperosmotic conditions (Supplementary Figure 3a), while this effect was blocked by administration of apocynin (Fig. 3d). These data point to a transcriptional regulation of miR-15a expression. Correspondingly, the finding that apocynin was also able to largely prevent the hyperosmolarity-induced downregulation of Bcl2, Ccnd1, Mcl1 and Faim (Fig. 4) confirms the inverse correlation between the levels of these miRNAs and the expression of anti-apoptotic genes. These findings suggest that NOX activity is required to mediate both the upregulation of miR-15a, -15b and -16 and the downregulation of Bcl2, Ccnd1, Mcl1 and Faim under hyperosmotic conditions.

Bottom Line: It was also identified that hyperosmolarity significantly reduces the expression of anti-apoptotic genes including Bcl2, Ccnd1, Mcl1, Faim, Aatf, Bfar and Ikbkb, which are either validated or predicted targets of these microRNAs.Moreover, through the application of NOX and JNK inhibitors as well as benzylamine it is shown that the observed response is mediated by reactive oxygen species (ROS), suggesting that miR-15a, miR-15b and miR-16 are novel redoximiRs.It is concluded that the response of these three microRNAs to osmotic stress is ROS-mediated and that it might contribute to the development of a proapoptotic phenotype.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, Hepatology and Infectious Diseases, Heinrich-Heine-University, Moorenstrasse 5, 40225 Düsseldorf, Germany.

ABSTRACT
microRNAs are an abundant class of small non-coding RNAs that negatively regulate gene expression. Importantly, microRNA activity has been linked to the control of cellular stress response. In the present study, we investigated whether the expression of hepatic microRNAs is affected by changes in ambient osmolarity. It is shown that hyperosmotic exposure of perfused rat liver induces a rapid upregulation of miR-15a, miR-15b and miR-16, which are members of the miR-15/107 microRNAs superfamily. It was also identified that hyperosmolarity significantly reduces the expression of anti-apoptotic genes including Bcl2, Ccnd1, Mcl1, Faim, Aatf, Bfar and Ikbkb, which are either validated or predicted targets of these microRNAs. Moreover, through the application of NOX and JNK inhibitors as well as benzylamine it is shown that the observed response is mediated by reactive oxygen species (ROS), suggesting that miR-15a, miR-15b and miR-16 are novel redoximiRs. It is concluded that the response of these three microRNAs to osmotic stress is ROS-mediated and that it might contribute to the development of a proapoptotic phenotype.

No MeSH data available.


Related in: MedlinePlus