Periodic Reporting for period 1 - RAMEN (RAMan spectroscopy for tracking Exosome based delivery of Nutraceuticals)
Okres sprawozdawczy: 2019-06-03 do 2021-06-02
The overall objective of the project is to design nanosized intestine targeted iron supplement delivery vehicles, based on milk derived exosomes to efficiently address anaemia, prevalent among 614 million women and 200 million children worldwide, a target of the 2nd United Nations Sustainable Development Goal (SDG) “zero hunger” under the 2030 agenda. Specifically, enterocytes, which are cells responsible for iron absorption and conversion to usable forms in the body, will be targeted, and the exosomes will be functionalised with cell penetrating peptides to circumvent endocytosis, facilitating direct delivery of the iron supplement to the cytosol. This uptake by enterocytes will be tracked using Raman microspectroscopy, a label-free live cell technique, observing the molecular signatures of exosomes and the changes in the signature of iron supplements, after delivery and conversion to usable forms in human cells and in vitro intestine models.
1.2 Work done towards achieving objectives set in the workpackage
From 17 June 2019 to 30 November 2019 , work was carried out towards realizing the objectives set in the first workpackage, Design and characterisation of functionalised nutrient loaded milk exosomes.
Task1.1 of the workpackage 1 was the focus of the work carried out in these 4 months. Low fat and high fat commercial milk were used to extract the exosomes and the preliminary studies showed that low fat milk handling was easier than full fat milk and the exosomes obtained did not block the filters during the centrifugation step.
Dynamic light scattering studies were used as described in the proposal to determine the size of the exosomes and their stability when stored at 4oC was followed over a few days.
Initial studies showed varying size distributions in the sample obtained and prefiltration steps were added to improve the size distribution of exosomes obtained.
Scanning electron microscopy was performed on the samples after dynamic light scattering to verify the presence of exosomes. Presence of protein aggregates was suspected from the scanning electron microscopy images and we were in the process of identifying a means to avoid protein aggregates and extract only exosomes.
This was when the fellow secured a permanent position with an industry in France and decided to move.
1.3 Impact
The fellowship has played a significant role in the fellow securing their current position. From the application process to my short stay, my mentor Prof. Hugh J Byrne had been very supportive and I learnt a lot of about different aspects of project management and proposal writing from him. This fellowship has given me the opportunity to expand my scientific network and would help my current work as well.
From 17 June 2019 to 30 November 2019 , work was carried out towards realizing the objectives set in the first workpackage, Design and characterisation of functionalised nutrient loaded milk exosomes.
Task1.1 of the workpackage 1 was the focus of the work carried out in these 4 months. Low fat and high fat commercial milk were used to extract the exosomes and the preliminary studies showed that low fat milk handling was easier than full fat milk and the exosomes obtained did not block the filters during the centrifugation step.
Dynamic light scattering studies were used as described in the proposal to determine the size of the exosomes and their stability when stored at 4oC was followed over a few days.
Initial studies showed varying size distributions in the sample obtained and prefiltration steps were added to improve the size distribution of exosomes obtained.
Scanning electron microscopy was performed on the samples after dynamic light scattering to verify the presence of exosomes. Presence of protein aggregates was suspected from the scanning electron microscopy images and we were in the process of identifying a means to avoid protein aggregates and extract only exosomes.
This was when the fellow secured a permanent position with an industry in France and decided to move.
1.3 Impact
The fellowship has played a significant role in the fellow securing their current position. From the application process to my short stay, my mentor Prof. Hugh J Byrne had been very supportive and I learnt a lot of about different aspects of project management and proposal writing from him. This fellowship has given me the opportunity to expand my scientific network and would help my current work as well.
In the recent years nanomaterials have been used for delivery of nutraceuticals especially Iron. Solid lipid nanoparticle based iron delivery is currently in clinical trial phase 2. There are other carriers that are being tested in vitro such as protein fibrils. These nanocarriers being tested are not modified for targeting or to enhance uptake. Hence, the efficiency of uptake may not be significantly higher than conventional supplements. In this study we intend to use nanoexosomes from milk that can survive stomach acids and have shown to be internalized by intestinal cells. Further, the nanoexosomes will be modified with a peptide to promote specific internalization pathwy to enhance release of iron in the cytosol of the cells. This we believe would increase iron delivered to the cells and the conversion to usable forms.
The impact would be to reduce side effects from conventional iron supplementation and a faster recovery of blood iron levels to acceptable levels.
The impact would be to reduce side effects from conventional iron supplementation and a faster recovery of blood iron levels to acceptable levels.