Ene 2017 MiPschool Obergurgl

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Isolated mitochondria transfer restores schizophrenia related deficits in-vitro and in-vivo.

Link: MitoEAGLE

Ene HM, Robicsek O, Karry R, Ben-Yehuda R, Yitzhaki O, Wiener I, Ben-Shachar D (2017)

Event: MiPschool Obergurgl 2017

COST Action MitoEAGLE

Malfunction of mitochondria, key players in various essential cell processes, has been repeatedly reported in schizophrenia (SZ) [1]. Recent studies have reported functional improvement and cellular viability following mitochondrial transplantation in several diseases [2]. Here we aimed to study (A) the effect of transfer of isolated active normal mitochondria (IAN-MIT) into SZ-derived iPSCs on their impaired differentiation into glutamatergic neurons [3]; and (B) the effect of IAN-MIT transfer into the medial prefrontal cortex (mPFC) in adolescence on SZ-related behavior and mitochondrial function in adulthood, using the poly I:C rat model of SZ. In this model, maternal immune activation by poly I:C leads to SZ-relevant behavioral and morphological deficits in adult offspring [4]. Notably, a significant decrease in mitochondrial ATP production was observed in poly I:C-treated mice, mainly attributed to a lower complex I activity.

In the in-vitro study, IAN-MIT were transferred into differentiating SZ-derived iPSCs into glutamatergic neurons. At the end of the differentiation period, the efficiency of neuronal differentiation was assessed by the expression of neuronal and glutamatergic markers (β3-Tubulin, Tbr1, PSD-95 and Synapsin1) using immunofluorescence staining. In addition, glutamate (Glu) and glutamine (Gln) metabolism were evaluated by high-pressure liquid chromatography (HPLC). In the in-vivo study, pregnant dams were injected with poly I:C or saline. On postnatal days 34-47, their offspring were administered with a single bilateral stereotaxic injection of freshly prepared IAN-MIT or vehicle into the mPFC. In adulthood, attentional function was assessed by the latent inhibition (LI) test, and mitochondrial function was evaluated in fronto-cortical freshly isolated JC-1 stained neurons by measuring mitochondrial membrane potential (Δym) and mitochondrial network dynamics, using confocal microscopy.

In the in-vitro study, IAN-MIT transfer improved differentiation of SZ-derived iPSCs into neurons, by increased expression of neuronal and glutamatergic markers β3-Tubulin, synapsin1 and Tbr1 and by an activation of the Glu-Gln cycle. In the in-vivo study, medial prefrontal IAN-MIT transfer in adolescence had a prolonged effect on animal behavior and mitochondrial function. In adult poly I:C-exposed offspring, IAN-MIT prevented disrupted LI and increased the reduced mitochondrial Δym. In control offspring however, IAN-MIT injection disrupted LI and increased mitochondrial Δym. IAN-MIT transfer did not affect mitochondrial cell distribution and network connectivity.

These findings demonstrate a beneficial effect of IAN-MIT transfer in-vitro and in-vivo, suggesting its therapeutic potential in diseases with bioenergetic and neurodevelopmental abnormalities such as SZ. The results suggest that in-vivo IAN-MIT transfer is beneficial when mitochondrial function is impaired, while under normal conditions it interferes with the homeostasis of mitochondrial diverse functions. Moreover, this study provides evidence for a possible relationship between mitochondria and attention, as an opposite pattern was obtained in controls and poly-I:C offspring for both mitochondrial and attentional functions.


Bioblast editor: Kandolf G


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Organism: Rat  Tissue;cell: Nervous system 




Event: B2, Oral 


Affiliations

Ene HM(1), Robicsek O(1), Karry R(1), Ben-Yehuda R(2), Yitzhaki O(1), Wiener I(2) Ben-Shachar D(1)
  1. Dept Neurosci, Psychobiol Res Lab, Rambam Medical Center, Bruce Rappaport Fac Med, Technion IIT, Israel
  2. Dept Socsci, TAU Univ, Isreal. - hila.ene1@gmail.com

Figures

Ene Figure1 MiPschool Obergurgl 2017.jpg

Figure 1. Glutamatergic differentiation following early and late IAN-MIT transfer. A. Bright-field images of neurons. B. Immunofluorescence staining and D. quantification of β3-Tubulin and Tbr1, both decreased in SZ neurons. Early and late IAN-MIT transfer increased β3–Tubulin, while Tbr1 increased only following late transfer. GLM analysis showed a time-dependent increase in both (β3–Tubulin– F=20.16, P<0.001. Tbr1-F=116.8, P<0.001). C. Immunofluorescence staining and E. quantification of synapsin1 and PSD-95 and their co-localization, which were decreased in SZ-neurons. Early and Late IAN-MIT transfer had a minor effect on PSD-95 and co-localization, yet profoundly affected synapsin1, which following late transfer reached control levels. GLM showed a time-dependent increase in synapsin1 (Synapsin1-F=90.15, P<0.002). Values are means±SEM of 2 controls and 2 patients (one iPSCs clone for each) from 2 experiments in duplicates with 4-6 cells in each. Scale bars A,B-100Im, C-25Im. F. Late, but not early, IAN-MIT transfer almost normalized Gln release and ameliorated abnormal Glu consumption. Values are means±SEM of 2 controls and 2 patients (one iPSCs clone for each) from 2 experiments in triplicates. *P<0.01 SZ vs. Cont; +P<0.008 SZ-Mit vs. Cont; #P<0.03 SZ-Mit vs. SZ.







Ene Figure2 MiPschool Obergurgl 2017.jpg

Figure 2. mPFC IAN-MIT transfer prevents disrupted LI and dissipated Δψm in poly-I:C exposed offspring. A. LI is manifested as shorter log times to complete licking criteria after tone onset of PE compared with NPE groups. Values are means±SEM. *P<0.0001 PE vs. NPE in saline-vehicle exposed offspring and in poly-I:C+IAN-MIT exposed offspring. B. Representative images of JC-1 stained cortical neurons. JC-1 in cytosol (green), in active mitochondria (red). Scale bars-5Im. C. Quantification of Δψm, which is reduced in poly-I:C rats and increased by IAN-MIT injection in both poly-I:C.


References

  1. Ben-Shachar D (2016) Mitochondrial multifaceted dysfunction in schizophrenia; complex I as a possible pathological target. Schizophr Res. In press.
  2. Masuzawa A, Black KM, Pacak CA, Ericsson M, Barnett RJ, Drumm C, Seth P, Bloch DB, Levitsky S, Cowan DB, McCully JD (2013) Transplantation of autologously derived mitochondria protects the heart from ischemia-reperfusion injury. Am J Physiol Heart Circ Physiol 304: 966-82.
  3. Robicsek O, Karry R, Petit I, Salman-Kesner N, Müller FJ, Klein E, Aberdam D, Ben-Shachar D (2013) Abnormal neuronal differentiation and mitochondrial dysfunction in hair follicle-derived induced pluripotent stem cells of schizophrenia patients. Mol Psychiatry 18(10):1067-76.
  4. Piontkewitz Y, Arad M, Weiner I (2011) Risperidone administered during asymptomatic period of adolescence prevents the emergence of brain structural pathology and behavioral abnormalities in an animal model of schizophrenia. Schizophr Bul. 37:1257-69.


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