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studies, a 10-minute insulin pretreatment significantly increased the cephalexin uptake
and PepT1 protein expression on the apical membrane. Again, no changes were
observed in the PepT1 mRNA expression levels. These findings suggest that insulin
binds to its receptor on the membranes, promoting PepT1 translocation from the
intracellular pool to the apical membrane surface. Collectively, these results indicate
that insulin signals an increased PepT1 translocation from a preformed cytoplasmic
pool to the apical membrane as opposed to de novo synthesis, which is similar to
events observed with the unfolded protein response.
143
,
146
These results are acute, as
opposed to the studies in diabetic rat models that are reflective of a chronic response,
further suggesting an alternative pathway by which insulin might increase PepT1
transcription and translation.
In addition to insulin, other hormonal PepT1 regulators have been identified and
studied, such as epidermal growth factor (EGF) and leptin. In contrast to insulin, EGF
was found to inhibit PepT1 by decreasing the number of transporter molecules on the
apical membrane.
147
Kinetic studies demonstrated a significant decrease in
V
max
on
the apical uptake of Gly-Sar, whereas
K
m
remained constant. In contrast, basolateral
Gly-Sar uptake demonstrated no changes in either
V
max
or
K
m
after EGF treatment.
147
Inhibition of transcellular Gly-Sar flux was observed after 5 days of EGF treatment
and reached a maximum after 15 days, indicating that a decrease in peptide transport
occurs after long-term EGF treatment. Interestingly, brief exposure of EGF has been
demonstrated to enhance the functional activity of PepT1.
147
A dose-dependent in-
crease in the apical Gly-Sar uptake with no change in
K
m
was observed in Caco-2 cells
after 5 minutes of EGF exposure. Furthermore, short-term EGF treatment did not alter
PepT1 mRNA expression or the intracellular pH. Unfortunately, the actual mechanism
by which short-term EGF treatment increases PepT1 functional expression remains
unclear.
147
Down-regulation of PepT2 mRNA was also observed after long-term
treatment with EGF in the rat proximal tubule cell line SKPT0193 cl.2 (SKPT),
148
implicating either a transcriptional regulatory mechanism through an EGF-responsive
element or regulation of PepT2 mRNA stability. However, further studies are required
to discern if the EGF regulatory mechanisms for PepT1 and PepT2 share common
features.
Buyse et al. explored hormonal PepT1 regulation further by demonstrating that
leptin secreted by the stomach may reach the small intestinal lumen to mediate trans-
porter expression.
149
The addition of leptin to intestinal perfusate increased the ab-
sorption of Gly-Sar across the rat jejunum in in vivo perfusion studies.
149
In addition,
uptake studies conducted utilizing Caco-2 cells demonstrated that leptin significantly
increased the
V
max
of Gly-Sar and cephalexin without changes in
K
m
values. Further,
leptin addition to the apical side of Caco-2 cells resulted in a 60% increase in mem-
brane PepT1 protein, with a corresponding decrease in intracellular PepT1. Moreover,
enhanced Gly-Sar uptake in response to leptin was abolished with colchicine pretreat-
ment but unaffected by brefeldin.
149
Taken together, these results suggest increased
trafficking of PepT1 from the intracellular pool to the apical membrane as a possible
mechanism to enhance the transcellular flux of substrates across Caco-2 cells.
Interestingly, gastrointestinal hormones and/or insulin elicit their regulatory effects
through specific binding to their respective surface receptors, while thyroid hormones
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