PCI + Amphinex og Amylin

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PCIB 26.06.2018 kl 13:00 949

Takker osten1 for linken https://www.sciencedirect.com/science/article/pii/S1572100018302060.
Som SafeInvest skriver er nedlagsfelt for PCI-teknologien stort og i artikkelen (pub 21 juni då) omhandler det PCI Amphinex i insulin produserende beta-celler.
Artikkelen har ikke en oppsummerende konklusjon. så det overlater vi til kompente skribenter her på HO

Aims: (a) To optimize the concentration and exposure set up of the photosensitizing compound
meso-disulfonated tetraphenyl chlorin TPCS2a (AmphinexⓇ) for use in insulin producing beta
cells. (b) Further, to utilize the photosensitizing technique to probe for intracellular effects in
beta cells by amylin.


Results. Five concentrations of TPCS2a were tested at different light exposure periods. Toxicity
was evaluated by the MTT assay [7], (Figure 1). Using 4ng/ml, 18h about 70% of the cells were
unperturbed after 60s of blue light exposure. This concentration was deemed optimal for amylin
experiments. Dark toxicity effects of the photosensitizer alone (no light), showed marginal
toxicity (2%-4%) after incubation of 2-10ng/ml of the TPCS2a, whereas 200ng/ml resulted in
17% decrease by MTT.

Based on pilot experiments we chose a 10μM concentration of amylin, also used in other studies.
No toxic effect on viability was observed by exposure to human amylin per se during the
present experimental conditions (Figure 2). However, human amylin in combination with TPCS2a
reduced the MTT parameter of viability above the effect of TPCS2a and illumination alone (by 15
± 3.6%, n=6, p< 0.007). To document specificity, we also tested rat amylin, which does not form
oligomers [1]. Rat amylin failed to exert toxicity in combination with TPCS2a and illumination
(Figure 2).
There was no difference in levels of apoptosis or necrosis after incubation with amylin in
combination with TPCS2a (4ng/ml, 18h) and illumination (60s) vs. TPCS2a and illumination alone,
details given in Supplementary information. Total glucose-stimulated insulin secretion was
direction-wise but not significantly reduced by the combination of human amylin (10µM) with
TPCS2a (4ng/ml) and illumination (60s) more than by TPCS2a and illumination alone. Cellular
insulin contents were not affected, details in Supplementary Information.
Discussion. Prior to experiments with amylin, it was necessary to find a useful combination of
TPCS2a exposure and light so that relative differences between amylin exposed versus non
amylin exposed cells could be detected. Achieving this first aim of the study, we proceeded to
test for dissociation of effects of amylin exposure with or without treatment with TPCS2a.
Importantly, we observed no significant toxic effect on viability by exposure to amylin alone,
making effects in combination with TPCS2a compatible with an intracellular effect. Absence of
effects by rat amylin supports this notion, since rat amylin does not form oligomers [1].

It is of interest that functional effects of amylin exposure seemed to affect insulin secretion but
not insulin contents. Interference of oligomers of amylin primarily on the signal-secretion chain
of events leading to glucose-induced insulin secretion has indeed been reported [8].
Our data on insulin secretion are however insufficient to determine whether the functional effects of
exposure to amylin are exerted intra- or extracellularly.

The main limitation of our study is the inability to demonstrate an increase in amylin oligomers
inside the beta cells before or after photosensitization. This is due to lack of specific antibodies
that are commercially available. In addition, our study has been restricted to clonal beta cells,
which may differ in important respects from native beta cells. Nevertheless, our data are at least
compatible with the notion that intracellularly formed oligomers from amylin can exert toxicity
intracellularly in beta cells.
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