* Unknown * | ACSJCA | JCA11.2.5208/W Library-x64 | manuscript.3f (R5.1.i3:5008 | 2.1) 2021/07/12 08:51:00 | PROD-WS-116 | rq_7422207 | 12/02/2021 14:15:13 | 12 | JCA-DEFAULT
www.acsami.org
Research Article
Design Strategy for a Hydroxide-Triggered pH-Responsive Hydrogel
2 as a Mucoadhesive Barrier to Prevent Metabolism Disorders
1
3
Rui-Chian Tang, Tzu-Chien Chen, and Feng-Huei Lin*
Cite This: https://doi.org/10.1021/acsami.1c17706
ACCESS
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
Metrics & More
Read Online
Article Recommendations
sı Supporting Information
*
ABSTRACT: Excess nutrient uptake is one of the main factors of
complications related to metabolism disorders. Therefore, efforts
have emerged to modulate nutrient transport in the intestine.
However, current approaches are mainly invasive interventions
with various side effects. Here, a pH-responsive hydrogel is
formulated by acidifying the hydroxide compounds within
sucralfate to allow electrostatic interactions between pectin and
aluminum ions. The pH responsiveness relies on the alternation of
cations and hydroxide species, providing reversible shifting from a
hydrogel to a complex coacervate system. It acts as a transient
physical barrier coating to inhibit intestinal absorption and changes
the viscosity and barrier function in different parts of the
gastrointestinal tract, showing enhanced mucoadhesive properties. The therapeutic hydrogel remarkably lowers the immediate
blood glucose response by modulating nutrient contact with bowel mucosa, suggesting potential in treating diabetes. In addition, it
significantly reduces weight gain, fat accumulation, and hepatic lipid deposition in rodent models. This study provides a novel
strategy for fabricating pH-responsive hydrogels, which may serve as a competent candidate for metabolism disorder management.
KEYWORDS: hydrogel, pH-responsive, hydroxide, intestine, metabolism disorders, pectin
reported.8 Consequently, a simultaneously safe and effective
alternative for bowel inhibition is needed. Thus, we aim to
develop an edible and biodegradable supplement that forms a
transient covering on the gastrointestinal tract as a temporary
barrier to prevent excessive absorption (Figure 1a). The
covering mimics the bypassing effect of bariatric surgeries and
duodenal sleeves, without requiring patients to undergo these
invasive interventions.
In this study, a mucoadhesive material would be considered
to serve as a covering on the gastrointestinal tract. Sucralfate is
an FDA-approved, orally administered polymer consisting of
sucrose octasulfate and polyaluminum complex. It inactivates
pepsin, adsorbs bile salts, and exhibits cytoprotective properties
to maintain epithelial integrity.9 Researchers have also shown
that treatment with HCl solution breaks the hydroxyl bonds of
the polyaluminum complex into shorter chains (Figure 1b).
These positively charged short chains bind to sucrose
octasulfate via electrostatic interactions and form a complex
coacervate system, which binds to the gastrointestinal tract as a
1. INTRODUCTION
Excess nutrition uptake can lead to obesity, impaired glucose
response, and dyslipidemia, which are generally termed under
the umbrella of metabolic syndrome.1 Numerous comorbidities such as type II diabetes and nonalcoholic fatty liver
disease (NAFLD) are closely related.2 However, no matter
how much the nutrient intake, metabolism disorders depend
highly on how much one absorbs. As the chief location for
digestion and absorption, the intestine plays a leading role in
the nutrient uptake.3 Therefore, regulating intestinal absorption could be one of the effective ways to treat metabolism
disorders. For example, the Roux-en-Y gastric bypass (RYGB)
operation shows more promising results than traditional
pharmaceuticals,4 providing adequate weight loss and glycemic
control. Nevertheless, invasive bariatric surgery irreversibly
alters the gastrointestinal tract, deterring more than 98% of the
eligible patients.5
Therefore, a less invasive procedure, EndoBarrier, was first
introduced to clinical studies in 2007.6 It is an endoscopically
implanted device anchored in the upper part of the proximal
duodenum. The duodenojejunal polymer sleeve would act as a
physical barrier to prevent contact between the ingested food
and the intestinal mucosa.7 Though helpful in achieving
improvements in metabolic parameters, EndoBarrier requires
annual removal, and numerous side effects, such as nausea,
gastrointestinal bleeding, and even device migration have been
© XXXX American Chemical Society
Received: September 14, 2021
Accepted: November 23, 2021
A
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
46
47
48
49
50 f1
51
52
53
54
55
56
57
58
59
60
61
62
63
64
ACS Applied Materials & Interfaces
www.acsami.org
Research Article
Figure 1. Schematic illustrations of the idea and materials used in this study. (a) Schematic illustrations of an orally administered polyelectrolyte
hydrogel as a transient barrier to inhibit excess nutrient uptake. (b) Illustrative schematic of the chemical structure of sucralfate and acid-treated
sucralfate. (c) Schematic illustration of the chemical structure of pectin. (d) Schematic representation of the pH-responsive hydrogel composed of
pectin and acid-treated sucralfate.
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
coating layer to reduce blood glucose response through its
barrier effect.10 However, the daily dosage of sucralfate to
achieve glycemic control would be costly. In addition, although
relatively rare, constipation, nausea, and urticaria may occur as
side effects under standard dosing.11 To address this issue, we
seek to reduce the amount of sucralfate required while
enhancing the barrier properties via creating complex
coacervation with charged polymers.
Natural mucoadhesive polysaccharides are of interest for
their beneficial effects on the gastrointestinal tract and low
cost. Pectin, a component of plant cell walls, was selected
because it promotes mucus secretion and maintains epithelial
integrity,12 prohibiting the infiltration of toxic lipopolysaccharides, which may induce mild inflammation of the intestine and
lead to a propensity for obesity and insulin resistance.13 Most
importantly, it consists of partially methoxy-esterified galacturonic acid units, which makes it an acidic polysaccharide with a
pKa of about 3.5 (Figure 1c).14 Given this property, pectin can
form a hydrogel by attracting multivalent cations through its
deprotonated carboxyl group at a specific pH, creating a 3D
network structure.15
Therefore, we predicted that pectin would form a hydrogel
with only traces of the acid-treated sucralfate via the attraction
between deprotonated carboxyl groups and aluminum ions,
forming a complex coacervate system with tight networking
barrier structures (Figure 1d). Since adjusting pH values alters
the ratio of aluminum cations and precipitated hydroxide
species as well as the number of deprotonated carboxyl groups,
we hypothesized that this unconventional hydrogel/complex
coacervate would possess pH-responsive properties in terms of
morphology and barrier function.
In this study, we synthesized the pectin sucralfate hydrogel
(PSH) with unique pH-responsive properties and enhanced
barrier function. Furthermore, we proposed a pH-responsive
gelation mechanism to serve as a design strategy to predict and
fabricate various hydrogels with similar behaviors. Finally, we
B
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
ACS Applied Materials & Interfaces
www.acsami.org
Research Article
Figure 2. Fabrication, physicochemical properties, and pH-dependent mechanism of PSH. (a) Representative pictures of PSH in different
gastrointestinal pH (gut, pH 1.2; duodenum, pH 3.5; intestine, pH 6.8). (b) Viscosity of pectin and PSH in different gastrointestinal pH (gut, pH
1.2; duodenum, pH 3.5; intestine, pH 6.8). One-way ANOVA with multiple comparisons (n = 3 per arm, **P < 0.002, ***P < 0.0001 compared to
pectin). (c) Change of rheological properties of PSH in different gastrointestinal pH (gut, pH 1.2; duodenum, pH 3.5; intestine, pH 6.8) (n = 3).
(d) Titration curve of 1% pectin. (e) Titration curve of 0.1% acid-treated sucralfate. (f) Titration curve of 0.1% sucralfate, 1% pectin, and 1% PSH.
(g) Proposed scenario for PSH in simulated gut fluid (pH 1.2), where the protonated pectin cannot attract the aluminum ions. (h) Proposed
scenario for PSH in simulated duodenal fluid (pH 3.5), where the deprotonated pectin forms a hydrogel with the aluminum ions. (i) Proposed
scenario for PSH in simulated intestinal fluid (pH 6.8), where the hydrogel structure decomposes due to the hydroxylation of aluminum ions,
leading to the formation of a complex coacervate system.
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
f2
116
evaluated the therapeutic effects of PSH on ameliorating oral
glucose tolerance response, weight gain under a high-fat diet,
body fat accumulation, hepatic lipid deposition, and other
metabolism disorders.
In contrast, neither the pectin itself nor the pectin added with
untreated sucralfate changed (Figure S1, Supporting Information). This suggests that pectin and acid-treated sucralfate can
indeed interact to form a hydrogel and that the pH-dependent
property is only involved in the formation of PSH. We found
that within a pH range of about 0.8−9, the change in viscosity
was reversible. However, when the environment became too
basic, PSH turned yellow and became much less viscous, and
the pH-responsive property was lost (Figure S2, Supporting
Information). This could be due to the β-elimination reaction
that cleaves the polymeric structure of pectin into smaller
fragments that are unable to form a tightly packed hydrogel.16
Further rheological analysis showed that PSH viscosity
increased slightly at pH 1.2 compared to pectin, became much
more viscous at pH 3.5, and reached an intermediate viscosity
at pH 6.8 (Figure 2b). This suggests that PSH is harder to be
washed away compared to pectin in the duodenum and
intestine, where most enzymatic reactions and nutrient
2. RESULTS AND DISCUSSION
2.1. Fabrication and Physicochemical Properties of
PSH. PSH was synthesized by combining acidified sucralfate in
HCl (0.2% w/v) and the same volume of citrus pectin solution
(2% w/v) to give a final concentration of 1% PSH (1% pectin
plus 0.1% acidified sucralfate). Specifically, pectin with over
74% of galacturonic acid was chosen to provide more carboxyl
groups for electrostatic interaction between the polyelectrolytes. PSH was adjusted to pH 1.2, 3.5, and 6.8 to simulate the
environments of the stomach, duodenum, and intestine,
respectively. In different environments, PSH changed its
viscosity, becoming more viscous and gel-like, resembling a
mixture of sticky paste and smashed jelly at pH 3.5 (Figure 2a).
C
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
ACS Applied Materials & Interfaces
www.acsami.org
Research Article
Figure 3. Barrier function, swelling, and adhesion assessments of PSH in vitro. (a) Schematic representation of the barrier function test. 1% of the
material was applied on a mucin-coated cellulose membrane and mounted onto a microfiltration laboratory apparatus, and the glucose permeation
was measured using a glucose meter. (b) Relative barrier function of pectin, acid-digested sucralfate, and PSH tested with glucose solution (500 mg
dL−1) in different gastrointestinal pH (gut, pH 1.2; duodenum, pH 3.5; intestine, pH 6.8). Two-way ANOVA with multiple comparisons (n = 3 per
arm, ***P < 0.0001 compared to pectin). (c) Relative barrier function of PSH formed with sucralfate digested in HCl solution (0.1−0.5 N) tested
with glucose solution (500 mg dL−1) in intestinal pH (n = 3). (d) Swelling of PSH in simulated duodenal fluid over time (n = 3). PSH was applied
evenly on a mucin-coated membrane and soaked in simulated duodenal fluid at 37 °C. Relative wet weights were measured and plotted with time as
the x-axis. (e) Zeta potential of pectin and PSH in different gastrointestinal pH (gut, pH 1.2; duodenum, pH 3.5; intestine, pH 6.8). Two-way
ANOVA with multiple comparisons (n = 3 per arm, ***P < 0.0001). (f) Schematic illustration of the in vitro adherence evaluation. FITC-albumin
encapsulated materials were loaded on a μ-slide seeded with IEC-6 cells and subjected to a constant medium flow. The retained fluorescence was
observed using a fluorescence microscope. (g) Mucus adhesion of pectin and PSH after being washed by medium for periods of time. FITCalbumin encapsulated materials are represented in green; the nucleus is represented in blue. Scale bar, 100 μm. (h) Remaining fluorescence over
time. Fluorescence at each time point was normalized to the initial fluorescence of pectin and PSH (100%).
135
136
137
138
139
the presence of solid behavior in a viscoelastic fluid in
oscillatory rheological analysis. Therefore, the result implies
that PSH exhibits more fluidlike rheological properties in a
simulated intestinal environment and may form a more
conformable coating on the intestinal epithelium. Also, the
absorption occur.17 We also found that the concentration of
HCl used to treat sucralfate did not affect the viscosity (Figure
S3, Supporting Information).
Moreover, the phase angle of PSH at pH 6.8 was higher than
that of PSH at pH 3.5 (Figure 2c). The phase angle measures
D
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
160
161
162
163
164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
180
181
182
183
184
185
186
187
188
189
190
191
192
193
194
f3
195
196
197
198
199
200
201
202
203
204
205
206
207
ACS Applied Materials & Interfaces
www.acsami.org
storage and loss modulii of PSH convey a similar concept
(Figure S4, Supporting Information).
2.2. pH-Responsive Mechanism of PSH. Typical
polysaccharide cation hydrogels, such as alginate hydrogel
cross-linked only by calcium ions, do not exhibit such drastic
morphological and rheological alternations as PSH at this
range of pH value.18 The unique pH-responsive property
might be attributed to the degree of pectin ionization and the
change in available aluminum cation species. For most pectins,
their pKa values are approximately 3.5,14 and over 99% of
carboxyl groups would be deprotonated at pH > 4.5 (Figure
2d). Furthermore, the pKa value of the first hydroxylation for
aluminum ions is 5.02.19 Thus, for acidified sucralfate,
aluminum ions are abundant at lower pH; at pH 4−5, most
of them convert into aluminum hydroxide species (mainly
Al(OH)2+),20 corresponding to the lower horizontal part of the
titration curve (Figure 2e).
We combine the titration curves and propose each scenario
at different pH values (Figure 2f). In the gastric environment,
despite the presence of aluminum ions, the pectin does not
have sufficient deprotonation of the carboxyl group to develop
into a hydrogel (Figure 2g). In the duodenal environment, the
negative ionized carboxyl groups bind to the positive
aluminum ions to convert into a hydrogel (Figure 2h). The
viscosity of PSH peaks at pH 4−5 and gradually decreases as
the aluminum ions are hydroxylated into aluminum monohydroxide, which may be structurally too large to be retained in
the hydrogel structure. Finally, the oppositely charged
polyelectrolytes transform into a complex coacervate system
in the intestinal environment (Figure 2i).
The observation of the titration curve could serve as a
template to predict new pH-responsive hydrogels consisting of
anionic gelling polymers and materials with multivalent
cationic hydroxide. Gelation occurs when the number of
cations and deprotonated anionic groups is sufficient, while the
hydrogel structure decomposes once the cations begin to
precipitate at a pH value equal to the first hydroxylation of
cations. That is, the formed hydrogels would transform into a
complex coacervate system.
Various existing hydroxides, whether with divalent or
trivalent ions, may serve as possible candidates. Furthermore,
manipulating the ratio of gelling polymers and hydroxides
changes the amount of alkali (or acid) needed to trigger
hydrogel transformation. This suggests the possibility of finetuning the reversible pH-responsive property in a targeted
environment.
2.3. Barrier Function. Improvements in the barrier
properties of PSH were investigated. The materials used in
the study spread on a mucin-coated porous cellulose
membrane were subjected to gravity loading, followed by a
permeability test with a D-glucose solution (Figure 3a).
Glucose was chosen as the main target for the effect on type
II diabetes. As shown in Figure 3b, acidified sucralfate (33%
blocked) exhibits a slightly better barrier effect than pectin
(26% blocked). Interestingly, the permeability changes notably
at different pH values for PSH. Although PSH exhibits lower
barrier properties in the simulated gut environment (21%
blocked), it shows remarkably enhanced barrier function in
simulated duodenal and intestinal environments (up to 63 and
85%, respectively). The result serves as a key implication that
PSH has the best barrier effect in environments where
digestion and nutrient absorption occur the most, which is
highly favorable in this study.
Intriguingly, the highest viscosity of PSH at pH 3.5 does not
provide the highest barrier function. Instead, PSH at pH 6.8
inhibits glucose permeation the most. From the proposed
mechanism, we deduced that it is because when pectin nearly
reaches total ionization, the attraction between cationic
aluminum hydroxide species and sucrose octasulfate along
intermolecular hydrogen bonds peaks to form a highly packed
complex coacervate system.
The influence of the acid concentration used to treat
sucralfate on the barrier function was investigated. For
digestion of sucralfate, 0.1−0.5 N HCl was used, and the
corresponding PSH was adjusted to pH 6.8 since our interest
lay in the formation of an intestinal barrier. The relative barrier
function was the best at 0.3 N (Figure 3c); therefore, sucralfate
was treated with this concentration to form PSH in subsequent
experiments.
2.4. Swelling and Zeta Potential. Given that PSH
becomes hydrogel-like at duodenal pH (pH 3.5), we were
curious if PSH would swell under the condition. Therefore,
PSH was evenly applied to a mucin-coated membrane and
soaked in simulated duodenal fluid at 37 °C. The hydrogel
layer on the mucus surface hardly swelled for at least 2 h in
simulated duodenal fluid (Figure 3d). This indicates that the
PSH layer coated on the intestinal tract retains its original
shape and the barrier properties change minimally during
meals.
Since the shift in electric charge plays an important role in
the pH-responsive property, we investigated the change in zeta
potential of pectin and PSH at different pH values. With
increasing pH, the zeta potential of pectin dropped from +3
mV at pH 1.2 to around −24 mV at pH 6.8, whereas the zeta
potential for PSH ranged only from −1 to −9 mV, which was
due to the neutralization of the aluminum cation species.
When the pH is gradually lowered, the cation species present
(Al3+, Al(OH)2+) is constantly attracted to the increasingly
deprotonated carboxyl group; therefore, the zeta potential of
PSH was closer to 0 mV compared to pectin (Figure 3e).
Considering that oligosaccharide chains confer a negative
charge to mucins through carboxyl and sulfate groups,21,22
many studies have shown that polymers or proteins with a
lower negative charge are better able to adhere to or penetrate
the mucus.21−24 Therefore, we reasoned that this lower
negativity might be another factor, in addition to higher
viscosity, that contributes to PSH adhering more firmly to the
luminal surface of the gastrointestinal tract.
2.5. Adhesion Assessment of PSH In Vitro. We
hypothesized that polyelectrolytes within the coacervate
system might bind more strongly to the negatively charged
mucin oligosaccharides, further increasing the adhesion of PSH
to the intestinal epithelium. Therefore, fluorescence-modified
materials were loaded onto a μ-slide seeded with IEC-6 cells,
and constant flow-induced shear was imposed on the system to
simulate stress under peristalsis (Figure 3f). As shown in
Figure 3g, the pectin and PSH showed similar fluorescence
intensity before constant shear (0 min). After 30 min, the
fluorescence of pectin decreased significantly, and 60 min later,
the fluorescence was barely visible. On the other hand, under
the same circumstances, a much larger proportion of PSH
remained on the μ-slide, indicating that PSH has better
adhesion to mucus than pectin.
2.6. Evaluation of Mucus Adherence In Vivo. C57BL/6
mice were gavaged with albumin FITC-loaded candidate
materials to test their ability to resist intestinal peristalsis over
E
Research Article
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
208
209
210
211
212
213
214
215
216
217
218
219
220
221
222
223
224
225
226
227
228
229
230
231
232
233
234
235
236
237
238
239
240
241
242
243
244
245
246
247
248
249
250
251
252
253
254
255
256
257
258
259
260
261
262
263
264
265
266
267
268
269
270
ACS Applied Materials & Interfaces
www.acsami.org
Research Article
Figure 4. Evaluation of mucus adherence in vivo and the reduced glucose response with PSH administration in mice. (a) Schematic illustration of
the in vivo mucus adhesion evaluation and the fluorescence image analysis. Mice were gavaged with materials (250 mg per kg mouse) encapsulated
with FITC-albumin, and the gastrointestinal tracts from the stomach to the colon were harvested in 1, 2, 4, and 24 h and imaged using an IVIS in
vivo imaging system. Mice gavaged with PBS were used as the control. (b) Schematic representation of the barrier effect of PSH in OGTT with oral
glucose administration. (c) OGTT curves of mice gavaged with pectin and PSH (250 mg per kg mouse). Mice gavaged with PBS were used as the
control; mice without glucose administration were used as shams. Two-way ANOVA with multiple comparisons (n = 4, ***P < 0.0001). (d) iAUC
of the OGTT curves. One-way ANOVA with multiple comparisons (n = 4 per arm, ***P < 0.0001 compared to control). (e) Schematic
representation of the no systemic effect of PSH in IPGTT where IP glucose administration bypassed the barrier coating. (f) IPGTT curves of mice
gavaged with PSH (250 mg per kg mouse). Mice gavaged with PBS were used as the control; mice without glucose administration were used as
shams. Two-way ANOVA with multiple comparisons (n = 4, ***P < 0.0001). (g) iAUC of the IPGTT curves. One-way ANOVA with multiple
comparisons (n = 4 per arm).
271
272
f4
273
274
275
276
277
compared to the small amount of pectin fluorescence retained.
The observation was also confirmed by quantifying the relative
fluorescence intensity. Thus, we conclude that PSH indeed
binds more strongly to the intestinal mucus than pectin itself,
in agreement with the results of the in vitro μ-slide test.
Moreover, there was no fluorescence signal after 24 h, which
indicated that PSH would only form a transient barrier on the
an extended period in vivo. The intestines were harvested at
different time points and visualized using the IVIS system
(Figure 4a). Fluorescent signals were detected mainly in the
duodenum and intestine within 1 h for both PSH and pectin.
The signal intensities gradually faded out with time, and a key
point to notice was at the fourth hour. At this time point, much
more PSH remained in the upper part of the intestine
F
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
278
279
280
281
282
283
284
ACS Applied Materials & Interfaces
www.acsami.org
Research Article
Figure 5. Long-term therapeutic effects of PSH on reducing weight gain, glucose response, and body fat deposition in a HFD-induced mice model.
(a) Illustrated schematic of the gastrointestinal barrier of PSH to inhibit HFD absorption in the mice model. (b) Weight gain (%) of mice fed with
HFD and HFD + PSH. Mice with rodent chow (regular food) were used as the control. Two-way ANOVA with multiple comparisons (n = 6, ***P
< 0.0001 for HFD + PSH vs HFD). (c) Food intake of mice fed with HFD and HFD + PSH. (d) Schematic illustration of a standard OGTT to
evaluate blood glucose response in mice. (e) OGTT curves of mice administered with glucose. Two-way ANOVA with multiple comparisons (n =
6, *P < 0.05, ***P < 0.0001 for HFD + PSH vs HFD). (f) iAUC of the OGTT curves. One-way ANOVA with multiple comparisons (n = 6 per
arm, ***P < 0.0001 compared to HFD). (g) 3D CT imaging of mice with a resolution of 35 μm voxel spacing. Skeletons are represented in white;
adipose tissue iss represented in red. (h) Representative pictures of eWAT. Scale bar, 1 cm. (i) Dry weight of the harvested eWAT. One-way
ANOVA with multiple comparisons (n = 6 per arm, ***P < 0.0001 compared to HFD).
285
286
287
288
289
290
291
292
293
294
295
296
297
298
299
300
301
302
Concerning metabolism disorders, dramatic fluctuations in
blood glucose are associated with various problems, including
coronary heart disease and aorta endothelial cell apoptosis.25,26
As opposed to the steep rise and fall in the control group,
where the highest blood glucose level was at around 350 mg
dL−1 and quickly dropped to 230 mg dL−1, the curve of the
PSH-fed group rose steadily to merely 230 mg dL−1 and fell
more slowly. This demonstrated the ability of PSH to keep a
steady blood glucose level after food intake, which is especially
vital for patients with diabetes.
An intraperitoneal glucose tolerance test (IPGTT) was
performed to examine whether PSH has a systemic effect on
blood glucose since glucose administration bypasses the
physical barrier of the PSH in the gastrointestinal tract (Figure
4e). In the IPGTT, the blood glucose level of both the control
and PSH groups rose to around 560 mg dL−1, and there was
no difference in the glucose response curves (Figure 4f), which
gastrointestinal tract, without continuing to accumulate in the
body.
2.7. Reduced Glucose Response with PSH Administration in Mice. We then evaluated the in vivo barrier effect
of PSH on nutrients, particularly glucose, for our interest in
metabolic syndrome-related diseases. An oral glucose tolerance
test (OGTT) was performed to determine whether the PSH
on the gastrointestinal mucosa could act as a physical barrier to
lower the blood glucose response (Figure 4b). The initial
blood glucose levels of both pectin and PSH were similar to
the control group, indicating that the materials were not
digested into absorbable fragments to influence blood glucose.
As shown in Figure 4c,d, PSH showed a remarkable reduction
in blood glucose response, with a 49.9% reduction in iAUC
compared to the saline control, which supports its feasibility as
a nutrient barrier. Moreover, pectin did not show the same
ability to inhibit glucose absorption, indicating the superior
performance of the engineered PSH.
G
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
303
304
305
306
307
308
309
310
311
312
313
314
315
316
317
318
319
ACS Applied Materials & Interfaces
www.acsami.org
Research Article
Figure 6. Therapeutic effects of PSH on attenuating fatty liver and LDL metabolism. (a) Representative pictures of livers. Scale bar, 1 cm. (b)
Biochemical markers for liver injury assessments. One-way ANOVA with multiple comparisons (n = 6 per arm, **P < 0.002 compared to HFD).
(c) Histological analysis of the liver section. Scale bar, 100 μm for 10× view; 25 μm for 40× view. (d) Biochemical markers for lipid metabolism.
One-way ANOVA with multiple comparisons (n = 6 per arm, *P < 0.05 compared to HFD).
320
321
322
323
324
325
326
327
328
329
indicates that the reduction in glucose response was due to the
localized barrier effect rather than a systemic effect.
The mucoadhesion of materials mostly depends on the
interfacial mucosa-material relationship. Given the electric
buffering properties that mucin possess, it will potentially affect
the physicochemical properties of materials, especially charged
colloidal systems such as PSH. Taking the complex and
dynamic environment into consideration, foods with various
pH values may also influence the structure of PSH. Moreover,
the interplay between mucus, PSH, and nutrients with different
forms (whether liquid, emulsion, or semisolid) can impact the
intestinal uptake.27 Therefore, more investigation into the
microscopic environment of PSH in actual gastrointestinal
tracts may allow future optimization of its function in vivo.
2.8. Long-Term Therapeutic Effects of PSH. C57BL/6
mice with 4−8 weeks of age were fed high-fat diet (HFD) to
establish an obesity model. PSH was administered orally (125
mg per kg) daily for 6 weeks to investigate its effects on
metabolic parameters compared with the HFD and control
groups (Figure 5a). As seen in Figure 5b, weight gain was
H
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
330
331
332
333
334
335
336
337
338
339 f5
340
341
342
343
344
345
346
347
348
349
350
351
352
353
354
355
356
357
358
359
360
361
362
363
364
365
366
367
368
369
370
371
372
373
374
375
376
377
378
379
380
381
382
383
384
385
386
387
388
389
390
391
392
393
394
395
396
397
398
399
400
401
402
ACS Applied Materials & Interfaces
www.acsami.org
significantly lower in the HFD + PSH group compared to the
HFD group from the second week onwards, and the difference
became greater as the experiment progressed, eventually
leading to a 28.4% decrease in weight gain. Compared to the
control group (normal diet), the HFD + PSH group had an
increased weight gain by only 9.1% even when given a HFD. In
terms of food intake, the HFD + PSH group was almost
identical to the HFD group (Figure 5c), suggesting that the
reduction in weight gain of the HFD + PSH group was not due
to an increased feeling of satiety for mice.
HFD is known to induce insulin resistance in mouse
models.28,29 Therefore, a standard OGTT (without PSH
gavage) was performed to evaluate the blood glucose response
after daily administration of the materials for 6 weeks (Figure
5d). In the control and HFD + PSH groups, the initial blood
glucose level was merely 130 and 115 mg dL−1, respectively,
while in the HFD group, it was 150 mg dL−1 (Figure 5e). This
implies that PSH could improve the diet-induced high fasting
blood glucose level. Moreover, the PSH-treated mice showed a
notable improvement in the blood glucose response. The peak
blood glucose level was only 260 mg dL−1 for the HFD + PSH
group and near 230 mg dL−1 for the control group, and the
curve patterns resemble. In contrast to the HFD group, which
had a 350 mg dL−1 peak value, the HFD + PSH group had a
30.3% reduction in iAUC (Figure 5f). The result suggests that
PSH can improve insulin sensitivity in the long term. We
deduce that PSH improves insulin sensitivity for several
reasons. First, PSH was synthesized mainly by pectin, which
promotes mucus secretion and maintains epithelial integrity.
Therefore, PSH may inhibit HFD-induced infiltration of toxic
lipopolysaccharides that contributes to susceptibility to insulin
resistance.13 Second, sucralfate present in PSH may also exert
cytoprotective properties to enhance epithelial repair by
stimulating prostaglandin synthesis.9
We further investigated fat accumulation to determine the
effect of HFD + PSH on body composition under HFD. A 3D
computed tomography (CT) imaging system was used to
visualize the total amount of adipose tissue in mice. As shown
in red, for the adipose tissue, HFD + PSH reduced its
deposition in the hip, abdominal, chest, and back regions of the
mice (Figures 5g and S11). The decreased visceral fat
formation was also confirmed by measuring the epididymal
white adipose tissue (eWAT) weight. The eWAT of PSH-fed
mice was only 53% the weight of the HFD group (Figure 5i).
These results suggest that PSH reduces the formation of total
and visceral fat. However, the total amount of adipose tissue
and the eWAT of the control group were significantly less
when compared to HFD + PSH. The result may indicate that
although PSH showed a promising effect in reducing weight
gain and fasting blood glucose level under a HFD, reversing
body fat accumulation to match the normal diet group might
be challenging.
Through histological analysis, hypertrophy of white
adipocytes was observed in the HFD group, while adipocyte
sizes were relatively smaller in the HFD + PSH group, in
agreement with the macroscopic observation mentioned above
(Figure S12).
2.9. Effects of PSH on NAFLD. NAFLD, a feature of
metabolic syndrome,30 is characterized by the accumulation of
triglycerides and cholesteryl esters in hepatocytes. It is due to a
constant imbalance between fatty acid influx and triglyceride
utilization.31 Given the barrier properties of PSH, we
speculated that it might inhibit excess fat uptake and thereby
reduce lipid deposition in the liver. Morphologically, the liver
of the HFD group was pale in color, while the HFD + PSH
group had a reddish shade similar to the control group (Figure
6a). Serum biochemical analysis showed that the HFD group
developed higher aspartate aminotransferase and alanine
aminotransferase, which are widely used as enzyme biomarkers
of liver injury.32 In contrast, the HFD + PSH group had a
profile similar to that of the control, suggesting that PSH may
attenuate liver injury correlated with excessive triglyceride
aggregation (Figure 6b).33 The histological analysis allows
direct observation of hepatic fat accumulation. The HFD
group had more and enlarged lipid droplets (white bubbles),
while the HFD + PSH group had smaller lipid droplets. The
result shows that PSH remarkably decreases hepatic fat
deposition despite HFD (Figure 6c).
Elevated plasma low-density lipoprotein (LDL) was found in
patients with NAFLD and insulin resistance.34 Moreover, LDL
is known to lead to cardiovascular diseases.35 Therefore, we
investigated the ability of PSH to lower serum LDL. As shown,
the HFD + PSH group had a remarkable decrease in
atherogenic LDL compared with the HFD group, although
the difference in HDL and other related biochemical markers
was not significant (Figure 6d). Therefore, we deduced that
PSH might reduce lipid accumulation in the liver and further
improve fat metabolism.
We showed that PSH forms a transient coating on the
gastrointestinal tract and essentially mimics the crucial part of
proximal bowel isolation of RYGB and EndoBarrier in a
noninvasive way. However, a majority of current gastric bypass
research is based on established obesity or type II diabetes
models.36,37 That is, the treatment comes after the disease. On
the contrary, our current research aims to prevent the
formation of metabolic complications in an earlier state.
Therefore, future studies may consider starting PSH treatment
after establishing obesity or diabetes models to give a more
comprehensive understanding of the efficacy of PSH compared
to existing solutions.
Concerning possible aluminum toxicity, no plaque was
observed in the histological analysis of brains in mice given
with PSH for 6 weeks (Figure S15, Supporting Information),
indicating that the minimal amount of aluminum ions present
would not lead to amyloid-β plaque accumulations in the brain.
Furthermore, there is no significant difference in the
histological analysis between HFD and HFD + PSH groups.
Hence, we concluded that the risk of aluminum toxicity is little
to none.
Research Article
3. CONCLUSIONS
In summary, acidifying the aluminum hydroxide component
within sucralfate attracts deprotonated pectin to form a pHresponsive hydrogel. The viscosity changes as the pH in the
gastrointestinal tract varies. By observing the titration curve of
the hydrogel, we proposed a possible mechanism for its unique
pH-responsive property. Both the barrier function and the
adhesion property of the hydrogel were remarkably better
when tested in vitro and in vivo as it formed a tightly packed
complex coacervate system. When administered daily, PSH
reduced weight gain, blood glucose response, fat accumulation,
and hepatic lipid storage in the HFD mouse model. Therefore,
we concluded that the hydrogel may serve as a competent
candidate for the treatment of metabolic syndrome-related
complications, such as type II diabetes, obesity, and NAFLD.
Due to its ease of use and low cost, this orally administered
I
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
403
404
405 f6
406 f6
407
408
409
410
411
412
413
414
415
416
417
418
419
420
421
422
423
424
425
426
427
428
429
430
431
432
433
434
435
436
437
438
439
440
441
442
443
444
445
446
447
448
449
450
451
452
453
454
455
456
457
458
459
460
461
462
463
464
465
466
467
468
ACS Applied Materials & Interfaces
www.acsami.org
polymer may appeal to a wider range of patients, even to those
with mild disease. Furthermore, the proposed mechanism may
give light to the synthesis of various pH-responsive hydrogels
containing gelling anionic polymers and multivalent hydroxides.
of candidate material containing 0.1 mg of FITC-albumin). The μslide was mounted on a programmable syringe pump (New Era Pump
Systems) to stimulate shear force during intestinal peristalsis. Cells
were under a constant flow of DMEM at a rate of 150 μL min−1,
which produced a shear stress of 1.8 μN cm−2. The materials were
observed with a fluorescence microscope at the time points of 0, 30,
and 60 min. The remaining fluorescence over time was quantified
using ImageJ and normalized to the initial fluorescence of pectin and
PSH (100%).
4.6. Fluorescence Imaging of the Gastrointestinal Tract
with PSH Gavage. To investigate the increased adherence of PSH in
the gastrointestinal tract, C57BL/6 mice were gavaged with PSH or
with pectin (dose, 250 mg per kg mouse) encapsulated with FITCalbumin (10% of effective dry weight), and the gastrointestinal tracts
were harvested after 1, 2, 4, and 24 h. Gastrointestinal tracts were then
imaged using an IVIS Lumina II in vivo imaging system
(PerkinElmer). Mice with phosphate-buffered saline (PBS) administration were used as controls, and all images were normalized using
the control.
4.7. OGTT and IPGTT. To evaluate the in vivo effect of PSH on
postprandial glucose uptake, C57BL/6 mice at 4−6 weeks of age were
administered PSH, followed by an OGTT. Based on the
recommendation of IACUC, the maximum gavage volume is 10 mL
per kg mouse. Given an average weight of 25 g for a mouse, the
maximum volume is 250 μL. Furthermore, PSH became too viscous
to be administered through the gavage needle when exceeding 2.5%.
Therefore, the dosage was 250 μL of 2.5% PSH for each mouse, which
was 250 mg per kg mouse (i.e., 6.25 mg dose per mouse), and the
PSH was adjusted to around pH 7. According to the human
equivalent dose calculation based on the body surface area
recommended by FDA, the human equivalent dosage of 250 mg
per kg mice is a 20.3 mg per kg human dose. If we consider a 60 kg
human, the PSH dosage is ∼1.2 g containing ∼1.1 g of pectin and 0.1
g of sucralfate. The dosage is much less than the clinical pectin usage
of 10−20 g daily and the maximum sucralfate dosing of 4−5 g daily.
The dosage may be further reduced through optimization. In the
OGTT experiments, C57BL/6 mice were fasted overnight (starting at
4 pm with access to water, fasting duration 18 h before treatment)
and gavaged with PSH, pectin, or PBS. Then, 1 h after material
administration, a 0.5 g per mL glucose solution was administered at a
dose of 3 g per kg mouse to measure the changes in glucose levels for
120 min (n = 4). Mice without glucose administration were used as
shams. Blood was collected from the tail vein to measure glucose
levels using a glucometer (Accu-Chek Instant, Roche). Each data
point was plotted with time as the x-axis, and iAUC was calculated
based on the plot and preadministration glucose level as the baseline.
Statistical difference was determined using two-way analysis of
variance (ANOVA). Results were considered significant when P ≤
0.05 (***P < 0.0001). In IPGTT, mice were treated the same as in
the OGTT, except that the glucose was injected directly into the
peritoneum.
4.8. Investigation of the Therapeutic Effects of PSH Long
Term. C57BL/6 mice at 4−8 weeks of age were fed a HFD consisting
of 60% kcal fat (Dyets) to establish the obesity model. Mice with
rodent chow only were used as the control group. Pectin and PSH
were administered via oral gavage at a dose of 125 mg per kg mouse
daily for 6 weeks, and the PSH was adjusted to around pH 7. DDW
was gavaged daily to the control and HFD groups to provide the same
stress conditions. Body weight and food intake were recorded once a
week. Mean values for each group were plotted with time as the x-axis.
In OGTT conducted after 6 weeks of daily gavaging, mice were
treated the same as previously mentioned, except that PSH was not
given before glucose administration.
4.9. Animals. All animal testing protocols were approved by the
National Taiwan University Institutional Animal Care and Use
Committee (20201042). Male C57BL/6 mice (LASCO) of similar
age were housed in groups (five per cage) on a 12 h light−dark cycle;
rodent chow and water were given ad libitum. Mice were acclimatized
for 1 week before performing the following experiments.
4. METHODS
469
470
471
472
473
474
475
476
477
478
479
480
481
482
483
484
485
486
487
488
489
490
491
492
493
494
495
496
497
498
499
500
501
502
503
504
505
506
507
508
509
510
511
512
513
514
515
516
517
518
519
520
521
522
523
524
525
526
527
528
529
530
4.1. Fabrication of PSH. To convert sucralfate into a complex
coacervate system capable of interacting with pectin, sucralfate was
treated with 0.3 N HCl (0.2% w/v) for 1 h. Subsequently, 2% w/v of
citrus pectin (>74% galacturonic acid, >6.7% methoxy groups, Mw =
195,000, Sigma-Aldrich) in distilled deionized water (DDW, pH 7)
was thoroughly mixed with the same volume of the treated sucralfate
for 1 min to obtain the PSH (with a final concentration of 1% pectin
and 0.1% treated sucralfate). The PSH was further adjusted to pH 1.2
to maintain its potency and stored at room temperature for use within
1 week.
4.2. Rheological Analysis. Rheological properties were measured
using a rheometer (TA Instruments, AR2000). A 40 mm plate was
chosen to measure the dynamic viscosity of PSH at different pH
values (shear rate, 0.1−10 s−1 in the logarithmic scale; a shear rate of
1 s−1 was chosen to compare the viscosity). The dynamic frequency
sweep was used to measure the dynamic phase angle (frequency
range, 0.1−10 Hz in logarithmic scale). All measurements were
performed at 37 °C to simulate the physiological environment.
4.3. Investigation of Barrier Properties Using Mucin-Coated
Membrane. Since the mucoadhesive property of the material mainly
depends on the interfacial interaction between the mucin and the
material, a mucin layer was coated on a porous membrane to mimic
the unstirred mucus layer. In addition, commercially available mucin
with a fixed concentration was used to obtain reproducible results,
given the inhomogeneity of mucin from different sources. A cellulose
membrane (pore size 6 μm, Advantec) was incubated in 3% w/v
porcine stomach mucin (Sigma-Aldrich) in DDW and gently shaken
for 1 h at room temperature. The excess mucin solution was removed
with 1 mL of DDW (pH 7). The mucin-coated membrane was used
within 1 h. Then, 1 mL of a 1% w/v candidate polymer (10 mg of
material) formulation in DDW (pH 7) was drawn with a pipette and
applied evenly to cover the whole mucin-coated membrane without
leaving any spare space to avoid leakage of glucose solution. The
membrane was further tilted to the vertical position for 1 min to
eliminate excess material that was not fully adhered to the membrane.
An exception to this procedure, sucralfate was dissolved in a 0.3 N
HCl. The polymer-coated membrane was mounted onto a microfiltration laboratory apparatus (Spectrum Chemical Mfg. Corp), 50
mL of glucose solution (500 mg dL−1) was added, and samples were
collected from the container of the apparatus after 10 min. Barrier
property tests were performed with three repeats for each material
individually. Glucose concentration was measured using a glucometer
(Accu-Chek Instant, Roche). Results were normalized to a mucincoated membrane without the application of a candidate polymer (0%
blocked).
4.4. Zeta Potential. To evaluate the change in electrical charges
for PSH at different pH values, the zeta potential was determined
using a 90 Plus/BI-MAS particle size analyzer (Brookhaven
Instruments). Measurements were performed under an electric field
of approximately 9 and 3 V cm−1 for pectin and PSH, respectively.
Samples were adjusted to 1% in DDW and heated to 37 °C before
analysis. Each sample was measured in triplicate runs in a polystyrene
cuvette (replicate analysis), each run consisting of 40 cycles.
4.5. In Vitro Adherence Evaluation. The μ-slide pumping
system was used to investigate the enhanced adhesion property of
PSH with mucin. 70,000 IEC-6 cells were seeded into the μ-Slide I 0.4
Luer (ibidi) and incubated for 24 h to generate sufficient mucin.
Then, the cells were stained with Hoechst for 30 min to visualize the
nucleus and washed three times with 100 μL of DMEM to remove
excess stain. Then, 100 μL of the 1% candidate material (adjusted to
pH 7) encapsulated with FITC-albumin (10% of material dry weight)
was applied and incubated with mucin for 30 min (equivalent to 1 mg
J
Research Article
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
531
532
533
534
535
536
537
538
539
540
541
542
543
544
545
546
547
548
549
550
551
552
553
554
555
556
557
558
559
560
561
562
563
564
565
566
567
568
569
570
571
572
573
574
575
576
577
578
579
580
581
582
583
584
585
586
587
588
589
590
591
592
593
594
595
596
597
598
599
ACS Applied Materials & Interfaces
600
601
602
603
■
ASSOCIATED CONTENT
sı Supporting Information
*
Morphology of pectin/pectin and sucralfate in different
gastrointestinal pH; deteriorated PSH when adjusted to
pH 11; viscosity of PSH formulated with sucralfate
digested in various HCl solutions (0.1−0.5 N);
frequency sweep measurements of PSH; change of
rheological properties of pectin; frequency sweep
measurements of pectin; SEM images of PSH; EDS
analysis result for PSH and pectin; WST-1 cell viability
assay for PSH and pectin; live/dead cytotoxicity assay
for pectin and PSH; IVIS imaging of gastrointestinal
tract from mouse gavaged with PBS; representative
pictures of mice fed with different diets; 3D CT imaging
of mice from various camera angles; histological analysis
of the adipose tissue section; and histological analysis of
the brain section (PDF)
605
606
607
608
609
610
611
612
613
614
615
616
617
618
Representative 3D CT imaging of mice from the control,
HFD, and HFD + PSH groups (ZIP)
619
621
622
623
624
625
626
627
628
629
630
631
632
■
AUTHOR INFORMATION
Corresponding Author
Feng-Huei Lin − Department of Biochemical Science and
Technology, College of Life Science, National Taiwan
University, Taipei 10617, Taiwan (ROC); Department of
Biomedical Engineering, College of Medicine and College of
Engineering, National Taiwan University, Taipei 10672,
Taiwan (ROC); Institute of Biomedical Engineering and
Nanomedicine, National Health Research Institutes, Miaoli
County 35053, Taiwan (ROC); orcid.org/0000-00022994-6671; Email: double@ntu.edu.tw
Authors
643
Rui-Chian Tang − Department of Biochemical Science and
Technology, College of Life Science, National Taiwan
University, Taipei 10617, Taiwan (ROC)
Tzu-Chien Chen − Department of Biochemical Science and
Technology, College of Life Science, National Taiwan
University, Taipei 10617, Taiwan (ROC); Department of
Biomedical Engineering, College of Medicine and College of
Engineering, National Taiwan University, Taipei 10672,
Taiwan (ROC)
Complete contact information is available at:
https://pubs.acs.org/10.1021/acsami.1c17706
644
Funding
645
646
R.-C.T., T.-C.C., and F.-H.L. all received funding from the
National Health Research Institutes grant BN-110-PP-01.
647
Notes
633
634
635
636
637
638
639
640
641
642
648
649
650
651
652
653
654
655
Research Article
REFERENCES
656
(1) Alberti, K. G. M.; Zimmet, P.; Shaw, J. The Metabolic
SyndromeA New Worldwide Definition. Lancet 2005, 366, 1059−
1062.
(2) Lakka, H.-M.; Laaksonen, D. E.; Lakka, T. A.; Niskanen, L. K.;
Kumpusalo, E.; Tuomilehto, J.; Salonen, J. T. The Metabolic
Syndrome and Total and Cardiovascular Disease Mortality in
Middle-Aged Men. JAMA 2002, 288, 2709−2716.
(3) Greenwood-Van Meerveld, B.; Johnson, A. C.; Grundy, D.
Gastrointestinal Physiology and Function. Handb. Exp. Pharmacol.
2017, 239, 1−16.
(4) Wolfe, B. M.; Kvach, E.; Eckel, R. H. Treatment of Obesity:
Weight Loss and Bariatric Surgery. Circ. Res. 2016, 118, 1844−1855.
(5) English, W. J.; DeMaria, E. J.; Brethauer, S. A.; Mattar, S. G.;
Rosenthal, R. J.; Morton, J. M. American Society for Metabolic and
Bariatric Surgery Estimation of Metabolic and Bariatric Procedures
Performed in the United States in 2016. Surg. Obes. Relat. Dis. 2018,
14, 259−263.
(6) Gersin, K. S.; Rothstein, R. I.; Rosenthal, R. J.; Stefanidis, D.;
Deal, S. E.; Kuwada, T. S.; Laycock, W.; Adrales, G.; Vassiliou, M.;
Szomstein, S.; Heller, S.; Joyce, A. M.; Heiss, F.; Nepomnayshy, D.
Open-Label, Sham-Controlled Trial of an Endoscopic Duodenojejunal Bypass Liner for Preoperative Weight Loss in Bariatric Surgery
Candidates. Gastrointest. Endosc. 2010, 71, 976−982.
(7) Patel, N.; Mohanaruban, A.; Ashrafian, H.; Le Roux, C.; Byrne,
J.; Mason, J.; Hopkins, J.; Kelly, J.; Teare, J. EndoBarrier(R): A Safe
and Effective Novel Treatment for Obesity and Type 2 Diabetes?
Obes. Surg. 2018, 28, 1980−1989.
(8) Ruban, A.; Ashrafian, H.; Teare, J. P. The EndoBarrier:
Duodenal-Jejunal Bypass Liner for Diabetes and Weight Loss.
Gastroenterol. Res. Pract. 2018, 2018, 7823182.
(9) German, A. J.; Maddison, J. E.; Guilford, G. Gastrointestinal
Drugs. In Small Animal Clinical Pharmacology, 2nd ed.; Maddison, J.
E., Page, S. W., Church, D. B., Eds.; W.B. Saunders: Edinburgh, 2008;
Chapter 19, pp 469−497.
(10) Lee, Y.; Deelman, T. E.; Chen, K.; Lin, D. S. Y.; Tavakkoli, A.;
Karp, J. M. Therapeutic Luminal Coating of the Intestine. Nat. Mater.
2018, 17, 834−842.
(11) McCarthy, D. M. Sucralfate. N. Engl. J. Med. 1991, 325, 1017−
1025.
(12) Beukema, M.; Faas, M. M.; De Vos, P. The Effects of Different
Dietary Fiber Pectin Structures on the Gastrointestinal Immune
Barrier: Impact via Gut Microbiota and Direct Effects on Immune
Cells. Exp. Mol. Med. 2020, 52, 1364−1376.
(13) Ding, S.; Lund, P. K. Role of Intestinal Inflammation as an
Early Event in Obesity and Insulin Resistance. Curr. Opin. Clin. Nutr.
Metab. Care 2011, 14, 328−333.
(14) Opanasopit, P.; Apirakaramwong, A.; Ngawhirunpat, T.;
Rojanarata, T.; Ruktanonchai, U. Development and Characterization
of Pectinate Micro/Nanoparticles for Gene Delivery. AAPS
PharmSciTech 2008, 9, 67−74.
(15) McKenna, B. A.; Nicholson, T. M.; Wehr, J. B.; Menzies, N. W.
Effects of Ca, Cu, Al and La on Pectin Gel Strength: Implications for
Plant Cell Walls. Carbohydr. Res. 2010, 345, 1174−1179.
(16) Diaz, J. V.; Anthon, G. E.; Barrett, D. M. Nonenzymatic
Degradation of Citrus Pectin and Pectate During Prolonged Heating:
Effects of pH, Temperature, and Degree of Methyl Esterification. J.
Agric. Food Chem. 2007, 55, 5131−5136.
(17) Johnson, I. T. Dietary Fiber|Physiological Effects and Effects on
Absorption. In Encyclopedia of Human Nutrition, 2nd ed.; Caballero,
B., Ed.; Elsevier: Oxford, 2005; pp 572−578.
(18) Chuang, J.-J.; Huang, Y.-Y.; Lo, S.-H.; Hsu, T.-F.; Huang, W.Y.; Huang, S.-L.; Lin, Y.-S. Effects of pH on the Shape of Alginate
Particles and Its Release Behavior. Int. J. Polym. Sci. 2017, 2017,
3902704.
(19) Holmes, L. P.; Cole, D. L.; Eyring, E. M. Kinetics of Aluminum
Ion Hydrolysis in Dilute Solutions. J. Phys. Chem. 1968, 72, 301−304.
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acsami.1c17706.
604
620
■
www.acsami.org
The authors declare no competing financial interest.
No unexpected or unusually high safety hazards were
encountered.
■
ACKNOWLEDGMENTS
This work was supported by the National Taiwan University
(NTU) and the National Health Research Institutes (BN-110PP-01) and made use of facilities at the Yonglin Biomedical
Engineering Hall.
K
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
657
658
659
660
661
662
663
664
665
666
667
668
669
670
671
672
673
674
675
676
677
678
679
680
681
682
683
684
685
686
687
688
689
690
691
692
693
694
695
696
697
698
699
700
701
702
703
704
705
706
707
708
709
710
711
712
713
714
715
716
717
718
719
720
721
722
723
724
725
726
727
728
729
730
731
732
733
734
735
736
737
738
739
740
741
742
743
744
745
746
747
748
749
750
751
752
753
754
755
756
757
758
759
760
761
762
763
764
765
766
767
768
769
770
771
772
773
774
775
776
777
778
779
780
781
782
783
784
785
786
787
788
789
790
791
ACS Applied Materials & Interfaces
www.acsami.org
(20) Kartikaningsih, D.; Shih, Y.-J.; Huang, Y.-H. Boron Removal
From Boric Acid Wastewater by Electrocoagulation Using Aluminum
as Sacrificial Anode. Sustain. Environ. Res. 2016, 26, 150.
(21) Li, L. D.; Crouzier, T.; Sarkar, A.; Dunphy, L.; Han, J.; Ribbeck,
K. Spatial Configuration and Composition of Charge Modulates
Transport Into a Mucin Hydrogel Barrier. Biophys. J. 2013, 105,
1357−1365.
(22) Marczynski, M.; Käsdorf, B. T.; Altaner, B.; Wenzler, A.;
Gerland, U.; Lieleg, O. Transient Binding Promotes Molecule
Penetration Into Mucin Hydrogels by Enhancing Molecular
Partitioning. Biomater. Sci. 2018, 6, 3373−3387.
(23) Ç elebioğlu, H. Y.; Lee, S.; Chronakis, I. S. Interactions of
Salivary Mucins and Saliva With Food Proteins: A Review. Crit. Rev.
Food Sci. Nutr. 2020, 60, 64−83.
(24) Takeuchi, H.; Thongborisute, J.; Matsui, Y.; Sugihara, H.;
Yamamoto, H.; Kawashima, Y. Novel Mucoadhesion Tests for
Polymers and Polymer-Coated Particles to Design Optimal
Mucoadhesive Drug Delivery Systems. Adv. Drug Delivery Rev.
2005, 57, 1583−1594.
(25) Zhang, X. G.; Zhang, Y. Q.; Zhao, D. K.; Wu, J. X.; Zhao, J.;
Jiao, X. M.; Chen, B.; Lv, X. F. Relationship Between Blood Glucose
Fluctuation and Macrovascular Endothelial Dysfunction in Type 2
Diabetic Patients With Coronary Heart Disease. Eur. Rev. Med.
Pharmacol. Sci. 2014, 18, 3593−3600.
(26) Wu, N.; Shen, H.; Liu, H.; Wang, Y.; Bai, Y.; Han, P. Acute
Blood Glucose Fluctuation Enhances Rat Aorta Endothelial Cell
Apoptosis, Oxidative Stress and Pro-Inflammatory Cytokine Expression In Vivo. Cardiovasc. Diabetol. 2016, 15, 109.
(27) Scheuble, N.; Schaffner, J.; Schumacher, M.; Windhab, E. J.;
Liu, D.; Parker, H.; Steingoetter, A.; Fischer, P. Tailoring Emulsions
for Controlled Lipid Release: Establishing In Vitro−In Vivo
Correlation for Digestion of Lipids. ACS Appl. Mater. Interfaces
2018, 10, 17571−17581.
(28) Winzell, M. S.; Ahrén, B. The high-fat diet-fed mouse: a model
for studying mechanisms and treatment of impaired glucose tolerance
and type 2 diabetes. Diabetes 2004, 53, S215.
(29) Park, S.-Y.; Cho, Y.-R.; Kim, H.-J.; Higashimori, T.; Danton, C.;
Lee, M.-K.; Dey, A.; Rothermel, B.; Kim, Y.-B.; Kalinowski, A.;
Russell, K. S.; Kim, J. K. Unraveling the Temporal Pattern of DietInduced Insulin Resistance in Individual Organs and Cardiac
Dysfunction in C57BL/6 Mice. Diabetes 2005, 54, 3530−3540.
(30) Marchesini, G.; Brizi, M.; Bianchi, G.; Tomassetti, S.;
Bugianesi, E.; Lenzi, M.; McCullough, A. J.; Natale, S.; Forlani, G.;
Melchionda, N. Nonalcoholic Fatty Liver Disease: A Feature of the
Metabolic Syndrome. Diabetes 2001, 50, 1844.
(31) Mensenkamp, A. R.; Havekes, L. M.; Romijn, J. A.; Kuipers, F.
Hepatic Steatosis and Very Low Density Lipoprotein Secretion: The
Involvement of Apolipoprotein E. J. Hepatol. 2001, 35, 816−822.
(32) Giannini, E. G.; Testa, R.; Savarino, V. Liver Enzyme
Alteration: A Guide for Clinicians. Can. Med. Assoc. J. 2005, 172,
367−379.
(33) Ipsen, D. H.; Lykkesfeldt, J.; Tveden-Nyborg, P. Molecular
Mechanisms of Hepatic Lipid Accumulation in Non-Alcoholic Fatty
Liver Disease. Cell. Mol. Life Sci. 2018, 75, 3313−3327.
(34) Tacer, K. F.; Rozman, D. Nonalcoholic Fatty Liver Disease:
Focus on Lipoprotein and Lipid Deregulation. J. Lipids 2011, 2011,
783976.
(35) Ference, B. A.; Ginsberg, H. N.; Graham, I.; Ray, K. K.;
Packard, C. J.; Bruckert, E.; Hegele, R. A.; Krauss, R. M.; Raal, F. J.;
Schunkert, H.; Watts, G. F.; Borén, J.; Fazio, S.; Horton, J. D.;
Masana, L.; Nicholls, S. J.; Nordestgaard, B. G.; van de Sluis, B.;
Taskinen, M.-R.; Tokgözoğlu, L.; Landmesser, U.; Laufs, U.; Wiklund,
O.; Stock, J. K.; Chapman, M. J.; Catapano, A. L. Low-Density
Lipoproteins Cause Atherosclerotic Cardiovascular Disease. 1.
Evidence From Genetic, Epidemiologic, and Clinical Studies. A
Consensus Statement From the European Atherosclerosis Society
Consensus Panel. Eur. Heart J. 2017, 38, 2459−2472.
(36) Aguilar-Olivos, N. E.; Almeda-Valdes, P.; Aguilar-Salinas, C. A.;
Uribe, M.; Méndez-Sánchez, N. The Role of Bariatric Surgery in the
Management of Nonalcoholic Fatty Liver Disease and Metabolic
Syndrome. Metabolism 2016, 65, 1196−1207.
(37) Vilarrasa, N.; De Gordejuela, A. G. R.; Casajoana, A.; Duran,
X.; Toro, S.; Espinet, E.; Galvao, M.; Vendrell, J.; López-Urdiales, R.;
Pérez, M.; Pujol, J. Endobarrier in Grade I Obese Patients With LongStanding Type 2 Diabetes: Role of Gastrointestinal Hormones in
Glucose Metabolism. Obes. Surg. 2017, 27, 569−577.
L
Research Article
https://doi.org/10.1021/acsami.1c17706
ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
792
793
794
795
796
797
798