| Резюме: | <p>D-alanine co-localization to corticotroph cells of the anterior pituitary inspired the research
in this thesis, as this co-localization may imply a relationship. Investigations into effects
that D-alanine have on the anterior pituitary, via AtT20 cells found that D-alanine does in
fact have an inhibitory relationship on ACTH secretion in AtT20 cells, and this was
robustly proven. Additionally, D-serine was investigated for comparison, and this D-amino
acid had similar effects. To ascertain inhibitory hormone effects by D-amino acids, an in
vivo study using CD1 mice found that animals treated with D-alanine or D-serine had
lower levels of corticosterone in their plasma. This supported the finding that these Damino acids have an inhibitory effect on HPA axis hormones.</p>
<p>In order to manipulate this regulation for potential therapeutic benefits the mechanism
behind this inhibition was sought, and I found that this may be via a combination of Asc-1
and ASCT2 amino acid transporters and the SGK-1-POMC system. NMDARs and GlyRs
were explored but the data did not support the direct involvement of these receptors in
inhibition of HPA axis hormones: ACTH and corticosterone. Asc-1 involvement was
determined by transfecting AtT20 cells with Asc-1 and observing a more pronounced
inhibition in cells overexpressed with this amino acid transporter and treated with Dalanine.
To support the in vitro data, 2 in vivo studies were performed – a model that increased Dalanine by feeding it to CD1 mice and a germ free model, which would lack D-alanine due
to the lack of gut microbiota, the source of mammalian D-alanine. However the ‘D-alanine
knockout model’ of germ-free mice did not have opposing effects on SGK1 and POMC in
the cortex – though this may be due to presence of D-serine in these animals, as D-serine
and D-alanine was found to exert similar effects on HPA axis hormones and SGK1.
Determining that D-amino acids may have a therapeutic benefit of HPA axis regulation,
the effects these molecules exert on the site of entry, the gut, was investigated in colorectal
Human carcinoma cells Caco-2, and it was found that D-serine had pro-inflammatory
effects on these gut cells, but not D-alanine, however the mechanism behind this
inflammation was not determined. However, this was an interesting find as it supports the
use of D-alanine over D-serine as a potential stress axis regulator, as this D-amino acid
does not promote gut inflammation.</p>
<p>The robust inhibition of ACTH in the anterior pituitary by D-alanine, and D-serine
(Chapter 2), is not directly linked to NMDA receptor or GlyRs activity, but may involve
the SGK1 system, which interacts with POMC, precursor to ACTH, and the amino acid
transporter Asc-1 (Chapter 3). However, ingestion of D-serine supplements may have a
pro-inflammatory effect in the gut, as observed in Caco-2 colorectal cells in Chapter 4.
The inhibition of HPA axis hormones (Corticosterone) by D-alanine and D-serine was also
observed in CD1 mice (Chapter 5).</p>
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