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Abstract
Background Ulcerative colitis (UC) is a chronic, relapsing disease, characterised by inflammation in colon and rectum. Common symptoms include bloody diarrhoea, abdominal pain, and faecal abnormalities, collectively leading to an impaired quality of life for the patients. The aetiology is not yet completely elucidated but it is considered to include an interplay between genetic and environmental factors, oxidative stress conditions, unbalanced immune responses and the intestinal microbiota.
The number of affected peo ple increases worldwide making safe and efficient treatments highly requested. Even though several therapeutic strategies are currently available, these are linked to unresponsiveness and side effects for many patients. Hence, development of alternative strategies for alleviation of UC is needed. Among various tested approaches, it is shown that some probiotic bacteria and antioxidant candidates holds promise as alternative or add on treatments These include the probiotic mix VSL#3 and E. coli Nissle ( EcN), and the antioxidants curcumin, resveratrol, and vitamin C. It is hypothesised that a delivery strategy combining protection during gastrointestinal (GI) transit and local release at the site of disease, namely the colon, may further improve the beneficial effects of these agents. To obtain this, novel oral delivery systems may be useful. This includes micrometre size cylindrical carriers, called microcontainers, which are designed for unidirectional loading of a compound of interest. Microcontainers have previously been found to possess mucoadhesive properties and they protect its content until coating controlled release at a relevant intestinal site.
Aim The aim of the present PhD study was to evaluate the application of this oral microcontainer based system for delivery of probiotic bacteria and antioxidants as alternative strategies for alleviation of UC. Effects on disease manifestations and the intestinal microbial composition was investigated in relevant mouse models.
Probiotic delivery Initially, microcontainer based delivery of the probiotic, EcN was evaluated. To ensure storage stability and enable microcontainer loading, a freeze drying procedure was optimised (Chapter 3 ). It was found that freeze drying settings, storage and microcontainer preparation processes were important for probiotic survival. This work was followed by an in vivo competition experiment in streptomycin treated mice (Chapter 4, Manuscript I). Colonisation and competition ability of EcN against two UC associated E. coli strains (EcUC) was compared for different delivery strategies of EcN, including microcontainer delivery and classical oral delivery strategies . Beneficial effects of microcontainer supported protection and mucus targeted delivery was hypothesised. However, no advantages of microcontainer delivery were found and the probiotic EcN co colonised with the EcUC strains for all delivery methods. This outcome is likely linked to a long lag phase of the freeze dried EcN and a lacking need for protection during GI transit for this specific probiotic. For future studies, it is suggested that sensitive next generation probiotics may benefit from this protective delivery form, even though microcontainer preparation methods needs prior optimisation to support viability of sensitive bacteria .
Antioxidant delivery Microcontainer delivery of the antioxidant, vitamin C was tested in a colitis mouse model. It was hypothesised that microcontainer supported protection from uptake in the small intestine, combined with stabilisation of the antioxidant, and colon targeted delivery might improve alleviating effects on local oxidative stress conditions and disease associated microbial dysbiosis. Several in vitro and in vivo preparation studies were conducted to obtain initial knowledge (Chapter 5). These included pH measurements along the GI tract in healthy and UC diseased m ice followed by coating release experiments to ensure targeted delivery by the microcontainers. UC-alleviating effects of vitamin C was finally investigated in mice. Delivery of vitamin C in drinking water and in peanut butter with or without microcontainers was compared and a sick and a healthy control group was additionally included (Chapter 6 , Manuscript II). Despite similar presence of vitamin C in the colon content for the dosed groups, the observed effects of the treatment differed between delivery methods. Alleviating effects on disease markers were observed for vitamin C dosed in peanut butter with or without microcontainers. This may be related to stabilisation of the antioxidant. In contrast, vitamin C dissolved in drinking water did not confer any positive effects, potentially due to instability in water. The disease associated microbial dysbiosis was not affected by vitamin C delivery.
These results emphasises that the delivery method of vitamin C is very important and encourage to further investigations elucidating the relevance of these findings for human consumption of the antioxidant.
Conclusion Specific candidates of both probiotics and antioxidants are promising as alternative therapeutic strategies for UC. Microcontainers can be used for intestinal delivery of both agents. However, within the studies of the present PhD, no beneficial effects are assigned to this delivery method compared to simple oral strategies. This suggests optimisation of the microcontainer system or development of other delivery strategies to support efficient alleviating outcomes of alternative UC treatments.
The number of affected peo ple increases worldwide making safe and efficient treatments highly requested. Even though several therapeutic strategies are currently available, these are linked to unresponsiveness and side effects for many patients. Hence, development of alternative strategies for alleviation of UC is needed. Among various tested approaches, it is shown that some probiotic bacteria and antioxidant candidates holds promise as alternative or add on treatments These include the probiotic mix VSL#3 and E. coli Nissle ( EcN), and the antioxidants curcumin, resveratrol, and vitamin C. It is hypothesised that a delivery strategy combining protection during gastrointestinal (GI) transit and local release at the site of disease, namely the colon, may further improve the beneficial effects of these agents. To obtain this, novel oral delivery systems may be useful. This includes micrometre size cylindrical carriers, called microcontainers, which are designed for unidirectional loading of a compound of interest. Microcontainers have previously been found to possess mucoadhesive properties and they protect its content until coating controlled release at a relevant intestinal site.
Aim The aim of the present PhD study was to evaluate the application of this oral microcontainer based system for delivery of probiotic bacteria and antioxidants as alternative strategies for alleviation of UC. Effects on disease manifestations and the intestinal microbial composition was investigated in relevant mouse models.
Probiotic delivery Initially, microcontainer based delivery of the probiotic, EcN was evaluated. To ensure storage stability and enable microcontainer loading, a freeze drying procedure was optimised (Chapter 3 ). It was found that freeze drying settings, storage and microcontainer preparation processes were important for probiotic survival. This work was followed by an in vivo competition experiment in streptomycin treated mice (Chapter 4, Manuscript I). Colonisation and competition ability of EcN against two UC associated E. coli strains (EcUC) was compared for different delivery strategies of EcN, including microcontainer delivery and classical oral delivery strategies . Beneficial effects of microcontainer supported protection and mucus targeted delivery was hypothesised. However, no advantages of microcontainer delivery were found and the probiotic EcN co colonised with the EcUC strains for all delivery methods. This outcome is likely linked to a long lag phase of the freeze dried EcN and a lacking need for protection during GI transit for this specific probiotic. For future studies, it is suggested that sensitive next generation probiotics may benefit from this protective delivery form, even though microcontainer preparation methods needs prior optimisation to support viability of sensitive bacteria .
Antioxidant delivery Microcontainer delivery of the antioxidant, vitamin C was tested in a colitis mouse model. It was hypothesised that microcontainer supported protection from uptake in the small intestine, combined with stabilisation of the antioxidant, and colon targeted delivery might improve alleviating effects on local oxidative stress conditions and disease associated microbial dysbiosis. Several in vitro and in vivo preparation studies were conducted to obtain initial knowledge (Chapter 5). These included pH measurements along the GI tract in healthy and UC diseased m ice followed by coating release experiments to ensure targeted delivery by the microcontainers. UC-alleviating effects of vitamin C was finally investigated in mice. Delivery of vitamin C in drinking water and in peanut butter with or without microcontainers was compared and a sick and a healthy control group was additionally included (Chapter 6 , Manuscript II). Despite similar presence of vitamin C in the colon content for the dosed groups, the observed effects of the treatment differed between delivery methods. Alleviating effects on disease markers were observed for vitamin C dosed in peanut butter with or without microcontainers. This may be related to stabilisation of the antioxidant. In contrast, vitamin C dissolved in drinking water did not confer any positive effects, potentially due to instability in water. The disease associated microbial dysbiosis was not affected by vitamin C delivery.
These results emphasises that the delivery method of vitamin C is very important and encourage to further investigations elucidating the relevance of these findings for human consumption of the antioxidant.
Conclusion Specific candidates of both probiotics and antioxidants are promising as alternative therapeutic strategies for UC. Microcontainers can be used for intestinal delivery of both agents. However, within the studies of the present PhD, no beneficial effects are assigned to this delivery method compared to simple oral strategies. This suggests optimisation of the microcontainer system or development of other delivery strategies to support efficient alleviating outcomes of alternative UC treatments.
Original language | English |
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Place of Publication | Kgs. Lyngby |
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Publisher | Technical University of Denmark |
Number of pages | 107 |
Publication status | Published - 2023 |
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Alleviation of ulcerative colitis by application of new oral delivery systems
Bondegaard, P. W. (PhD Student), Licht, T. R. (Main Supervisor), Mortensen, M. S. (Supervisor), Kleerebezem, M. (Examiner) & Müllertz, A. (Examiner)
01/05/2018 → 10/06/2024
Project: PhD