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Profiling of Proteins Regulated by Venlafaxine during Neural Differentiation of Human Cells.

Doh MS, Han DM, Oh DH, Kim SH, Choi MR, Chai YG - Psychiatry Investig (2015)

Bottom Line: To study the effects of venlafaxine during neural differentiation at the cellular level, we looked at its effect on protein expression and regulation mechanisms during neural differentiation.Treatment with venlafaxine decreased expression of prolyl 4-hydroxylase (P4HB), ubiquitin-conjugating enzyme E2K (HIP2) and plastin 3 (T-plastin), and up-regulated expression of growth factor beta-3 (TGF-β3), dihydropyrimidinase-like 3 (DPYSL3), and pyruvate kinase (PKM) after differentiation for 1 and 7 days.Our findings may contribute to improve understanding of molecular mechanism of venlafaxine.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Life Sciences, Hanyang University, Ansan, Republic of Korea.

ABSTRACT

Objective: Antidepressants are known to positively influence several factors in patients with depressive disorders, resulting in increased neurogenesis and subsequent relief of depressive disorders. To study the effects of venlafaxine during neural differentiation at the cellular level, we looked at its effect on protein expression and regulation mechanisms during neural differentiation.

Methods: After exposing NCCIT cell-derived EBs to venlafaxine during differentiation (1 day and 7 days), changes in protein expression were analyzed by 2-DE and MALDI-TOF MS analysis. Gene levels of proteins regulated by venlafaxine were analyzed by real-time RT-PCR.

Results: Treatment with venlafaxine decreased expression of prolyl 4-hydroxylase (P4HB), ubiquitin-conjugating enzyme E2K (HIP2) and plastin 3 (T-plastin), and up-regulated expression of growth factor beta-3 (TGF-β3), dihydropyrimidinase-like 3 (DPYSL3), and pyruvate kinase (PKM) after differentiation for 1 and 7 days. In cells exposed to venlafaxine, the mRNA expression patterns of HIP2 and PKM, which function as negative and positive regulators of differentiation and neuronal survival, respectively, were consistent with the observed changes in protein expression.

Conclusion: Our findings may contribute to improve understanding of molecular mechanism of venlafaxine.

No MeSH data available.


Related in: MedlinePlus

mRNA expression patterns of proteins regulated by venlafaxine during neural differentiation of NCCIT cells. EBs were treated with 10 µM retinoic acid (RA) in the presence or absence of 10 µM venlafaxine (VEN) for 1 or 7 days, and mRNA expression levels of targets differentially regulated by VEN were analyzed by real-time RT-PCR. A: Levels of mRNAs regulated by VEN after differentiation for 1 day. B: Spot intensity of proteins regulated by VEN after differentiation for 1 day. C: Levels of mRNAs regulated by VEN after differentiation for 7 days. D: Spot intensity of proteins regulated by VEN after differentiation for 7 days. Each bar in A and C represents the mean±SEM (N=3). *significantly different from CON by the two-sample t-test (p<0.05). ND: not detected by 2DE assay.
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Figure 3: mRNA expression patterns of proteins regulated by venlafaxine during neural differentiation of NCCIT cells. EBs were treated with 10 µM retinoic acid (RA) in the presence or absence of 10 µM venlafaxine (VEN) for 1 or 7 days, and mRNA expression levels of targets differentially regulated by VEN were analyzed by real-time RT-PCR. A: Levels of mRNAs regulated by VEN after differentiation for 1 day. B: Spot intensity of proteins regulated by VEN after differentiation for 1 day. C: Levels of mRNAs regulated by VEN after differentiation for 7 days. D: Spot intensity of proteins regulated by VEN after differentiation for 7 days. Each bar in A and C represents the mean±SEM (N=3). *significantly different from CON by the two-sample t-test (p<0.05). ND: not detected by 2DE assay.

Mentions: Although post-translational modification and protein turn over can affect the levels of protein expression, determining that an identified protein target is regulated at the mRNA level provides corresponding evidence of the reliability of the relative quantitative protein expression results. Thus, we measured transcript levels of selected proteins using real-time RT-PCR. Similar to the proteomic expression pattern, the expression levels of HIP2 and T-plastin mRNA were decreased while levels of PKM and TGF-β3 mRNA were increased in venlafaxine-treated cells compared to control cells after differentiation for 1 day (Figure 3A and B). After differentiation for 7 days, the levels of expression of P4HB and T-plastin mRNA were significantly decreased, while the expression of PKM and DPYSL3 mRNAs were increased in venlafaxine-treated cells compared to control cells. Together, these results corresponded with the observed patterns of protein expression (Figure 3C and D). On the other hand, although expression of DPYSL3 and HIP2 proteins could not be detected after 1 and 7 days of differentiation, respectively, changes in their expression at the mRNA level were confirmed. Based on these results, it appeared that, beginning almost immediately after initiation of differentiation and up to 7 days later, treatment with venlafaxine increased consistently levels of DPYSL3 and levels of HIP2 gradually decreased. Together, these data served to validate several proteins obtained from proteomic profiling, as well as provided information regarding estimated expression levels of proteins that were identified at only a single time point by proteomic analysis.


Profiling of Proteins Regulated by Venlafaxine during Neural Differentiation of Human Cells.

Doh MS, Han DM, Oh DH, Kim SH, Choi MR, Chai YG - Psychiatry Investig (2015)

mRNA expression patterns of proteins regulated by venlafaxine during neural differentiation of NCCIT cells. EBs were treated with 10 µM retinoic acid (RA) in the presence or absence of 10 µM venlafaxine (VEN) for 1 or 7 days, and mRNA expression levels of targets differentially regulated by VEN were analyzed by real-time RT-PCR. A: Levels of mRNAs regulated by VEN after differentiation for 1 day. B: Spot intensity of proteins regulated by VEN after differentiation for 1 day. C: Levels of mRNAs regulated by VEN after differentiation for 7 days. D: Spot intensity of proteins regulated by VEN after differentiation for 7 days. Each bar in A and C represents the mean±SEM (N=3). *significantly different from CON by the two-sample t-test (p<0.05). ND: not detected by 2DE assay.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4310925&req=5

Figure 3: mRNA expression patterns of proteins regulated by venlafaxine during neural differentiation of NCCIT cells. EBs were treated with 10 µM retinoic acid (RA) in the presence or absence of 10 µM venlafaxine (VEN) for 1 or 7 days, and mRNA expression levels of targets differentially regulated by VEN were analyzed by real-time RT-PCR. A: Levels of mRNAs regulated by VEN after differentiation for 1 day. B: Spot intensity of proteins regulated by VEN after differentiation for 1 day. C: Levels of mRNAs regulated by VEN after differentiation for 7 days. D: Spot intensity of proteins regulated by VEN after differentiation for 7 days. Each bar in A and C represents the mean±SEM (N=3). *significantly different from CON by the two-sample t-test (p<0.05). ND: not detected by 2DE assay.
Mentions: Although post-translational modification and protein turn over can affect the levels of protein expression, determining that an identified protein target is regulated at the mRNA level provides corresponding evidence of the reliability of the relative quantitative protein expression results. Thus, we measured transcript levels of selected proteins using real-time RT-PCR. Similar to the proteomic expression pattern, the expression levels of HIP2 and T-plastin mRNA were decreased while levels of PKM and TGF-β3 mRNA were increased in venlafaxine-treated cells compared to control cells after differentiation for 1 day (Figure 3A and B). After differentiation for 7 days, the levels of expression of P4HB and T-plastin mRNA were significantly decreased, while the expression of PKM and DPYSL3 mRNAs were increased in venlafaxine-treated cells compared to control cells. Together, these results corresponded with the observed patterns of protein expression (Figure 3C and D). On the other hand, although expression of DPYSL3 and HIP2 proteins could not be detected after 1 and 7 days of differentiation, respectively, changes in their expression at the mRNA level were confirmed. Based on these results, it appeared that, beginning almost immediately after initiation of differentiation and up to 7 days later, treatment with venlafaxine increased consistently levels of DPYSL3 and levels of HIP2 gradually decreased. Together, these data served to validate several proteins obtained from proteomic profiling, as well as provided information regarding estimated expression levels of proteins that were identified at only a single time point by proteomic analysis.

Bottom Line: To study the effects of venlafaxine during neural differentiation at the cellular level, we looked at its effect on protein expression and regulation mechanisms during neural differentiation.Treatment with venlafaxine decreased expression of prolyl 4-hydroxylase (P4HB), ubiquitin-conjugating enzyme E2K (HIP2) and plastin 3 (T-plastin), and up-regulated expression of growth factor beta-3 (TGF-β3), dihydropyrimidinase-like 3 (DPYSL3), and pyruvate kinase (PKM) after differentiation for 1 and 7 days.Our findings may contribute to improve understanding of molecular mechanism of venlafaxine.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Life Sciences, Hanyang University, Ansan, Republic of Korea.

ABSTRACT

Objective: Antidepressants are known to positively influence several factors in patients with depressive disorders, resulting in increased neurogenesis and subsequent relief of depressive disorders. To study the effects of venlafaxine during neural differentiation at the cellular level, we looked at its effect on protein expression and regulation mechanisms during neural differentiation.

Methods: After exposing NCCIT cell-derived EBs to venlafaxine during differentiation (1 day and 7 days), changes in protein expression were analyzed by 2-DE and MALDI-TOF MS analysis. Gene levels of proteins regulated by venlafaxine were analyzed by real-time RT-PCR.

Results: Treatment with venlafaxine decreased expression of prolyl 4-hydroxylase (P4HB), ubiquitin-conjugating enzyme E2K (HIP2) and plastin 3 (T-plastin), and up-regulated expression of growth factor beta-3 (TGF-β3), dihydropyrimidinase-like 3 (DPYSL3), and pyruvate kinase (PKM) after differentiation for 1 and 7 days. In cells exposed to venlafaxine, the mRNA expression patterns of HIP2 and PKM, which function as negative and positive regulators of differentiation and neuronal survival, respectively, were consistent with the observed changes in protein expression.

Conclusion: Our findings may contribute to improve understanding of molecular mechanism of venlafaxine.

No MeSH data available.


Related in: MedlinePlus