Presentation is loading. Please wait.

Presentation is loading. Please wait.

ACS nano. Subventricular Zone (SVZ) Subgranular Zone (SGZ) Rostral Migratory Stream (RMS) Neurogenic Niches - SVZ Neural Stem Cells (NSCs) display three.

Similar presentations


Presentation on theme: "ACS nano. Subventricular Zone (SVZ) Subgranular Zone (SGZ) Rostral Migratory Stream (RMS) Neurogenic Niches - SVZ Neural Stem Cells (NSCs) display three."— Presentation transcript:

1 ACS nano

2 Subventricular Zone (SVZ) Subgranular Zone (SGZ) Rostral Migratory Stream (RMS) Neurogenic Niches - SVZ Neural Stem Cells (NSCs) display three cardinal features: -proliferation -self renewal -multipotency Santos et al., 2012, Integr Biol In adult mammalian brain there are cells with stem cell properties, responsible for the formation of new neurons

3 Many factors regulate neurogenesis (Maden et al. Nature 2002) One developmental molecule of particular interest is retinoic acid (RA). Enhancing neurogenesis in the injured brain is a promissing therapy Retinoic Acid (RA)

4 Maden et al., 2007, Nat Rev Neurosci - Regulates proliferation and differentiation of stem cells in the developing and adult brain Santos et al., 2012, Integr Biol - Can improve age-related neuronal and cognitive loss Crandall et al Can influence LTP, LTD and neurite and axonal outgrowth Chiang et al & Corcoran and Maden et al 1999 A better understanding on how RA regulates postnatal neurogenesis may therefore offer regenerative strategies to treat brain injury or degeneration Retinoic Acid

5 However, RA: Is rapidly metabolized Has poor water solubility Requires a fine-tuning of concentration Aim: We propose an innovative approach to induce neuronal differentiation by using RA- loaded nanoparticles for the controlled release of RA. Retinoic Acid The use of nanoparticles (NPs) can be a powerful strategy to overcome these limitations

6 Neurosphere+EGF/FGF2 Doetsch et al 1999 SVZ cultures RA delivery system Since RA is rapidly metabolized by cells and has low solubility in aqueous solutions DS/PEI RA-releasing nanoparticles (RA + -NPs) were used as a vehicle for intracellular delivery dissociation Methods

7 RA-NPs achieve a proneurogenic effect with a RA concentration 2500-fold lower than the one with free RA RA-NPs induce functional neuronal differentiation via nuclear RAR activation Retinoic Acid-Loaded Nanoparticles Maia & Santos et al., 2011, ACSnano Control RA + -NPs + BMS493 Control RA + -NPs + BMS493

8 Sox2 -/- Sox2 -/+ Sox2 +/+ Nuclei Sox2Dlx2 Merge Stemness Dissociation Treatment 5days Sphere counting dissociation Secondary spheres counting Self-renewal assay 24hrs -/- pro-commitment +/+ pro-stemness -/+ neutral Sox-2, Dlx2 Cell pair assay

9 Stemness Sox2+/+Sox2+/-Sox2-/- Self-renewal assayCell pair assay RA-NPs induce the commitment of SVZ NSCs

10 Phospho-JNK 6hrs RA-NP treatment increased the total length and the number of P-JNK positive ramifications on Tau positive axons Signalling

11 Cross-linking Sonication Immunoprecipitation (H3K4me3) Reverse cross links Purify DNA Quantitative PCR for the promoter region of proneurogenic genes Epigenetic regulation by histone methylation is thought to be involved in long-term maintenance of certain regions of the genome Several genes are reported to become more expressed during this process Mash1 and Neurogenin1 qChIP (Abcam protocols)

12 RA facilitates the expression of pro-neurogenic genes Day 0 Plating of neurospheres In differentiation conditions (without EGF/FGF) Day 7 Day 9 Day 11 qChIP NP-RA exposure Day 10 qChIP

13 Day 0 Plating of neurospheres In differentiation conditions (without EGF/FGF) Day 7 Day 9 Day 12 RT-PCR NP-RA exposure RA-NPs promote a stronger expression of proneurogenic genes than solubilized RA Mash1 Ngn1 RA + -NPs - + RA + -NPs qRT-PCR

14 RA-NPs induce the expression of proneurogenic genes in the SVZ neurogenic niche in vivo In Vivo

15 LV 100µm LV SVZ RA-NPs intracerebroventricular injection 7 days SVZ laser microdissection LV: lateral ventricle

16 Our work presents a novel method to modulate the differentiation of SVZ cells involving the use of retinoic acid-releasing nanoparticles The use of RA-NPs may open new perspectives for brain repair Nanoparticles proved to deliver efficiently RA within the cells and consequently promote neuronal differentiation Treated cultures presented a higher expression of neuronal markers and functional neuronal activity. Retinoic acid promotes de trimethylation of H3K4 upregulating the expression of pro-neurogenic genes Retinoic Acid activates JNK pathway We demonstrated successfully that a NP formulation could be used in vivo to control the differentiation of NSCs RA-NPs were more robust in maintaining the signature of gene expression both in vitro and in vivo than solubilized RA Conclusions


Download ppt "ACS nano. Subventricular Zone (SVZ) Subgranular Zone (SGZ) Rostral Migratory Stream (RMS) Neurogenic Niches - SVZ Neural Stem Cells (NSCs) display three."

Similar presentations


Ads by Google