Smart Protein-Based Formulation of Dendritic Mesoporous Silica Nanoparticles: Toward Oral Delivery of Insulin

Abstract

Oral insulin administration still represents a paramount quest that almost a century of continuous research attempts did not suffice to fulfill. Before pre-clinical development, oral insulin products have first to be optimized in terms of encapsulation efficiency, protection against proteolysis, and intestinal permeation ability. With the use of dendritic mesoporous silica nanoparticles (DMSNs) as an insulin host and together with a protein-based excipient, succinylated β-lactoglobulin (BL), pH-responsive tablets permitted the shielding of insulin from early release/degradation in the stomach and mediated insulin permeation across the intestinal cellular membrane. Following an original in vitro cellular assay based on insulin starvation, direct cellular fluorescent visualization has evidenced how DMSNs could ensure the intestinal cellular transport of insulin.

Keywords: beta-lactoglobulin tablets; cellular insulin starvation; dendritic mesoporous silica nanoparticles; oral insulin dosage; transcellular epithelial transport.

Figure 1

(a–c) TEM images of the DMSNs synthesized with either hexane (DMSN‐Hex), cyclohexane (DMSN‐Cyclo) or toluene (DMSN‐Tol); (d, e) Isotherms and respective pore size distributions obtained from N₂ physisorption analyses (−196 °C); (f) Particle size distribution of the DMSNs obtained from dynamic light scattering (DLS) measurements.

Scheme 1

Scheme representing the loading of insulin inside DMSNs and the formulation of the BL tablets for the release tests (ZP stands for Zeta‐potential).

Figure 2

(a) Cumulative release of insulin from the different tablets immersed 2 h at pH 1.2 and subsequently at pH 7.4. Data shown as mean ± SE (n=3), Significant difference express **p<0.01, ****p<0.0001, One‐way ANOVA and Fisher Test; (b–d) MS characterization of the released insulin or the fragments of insulin at different time periods in correlation with their respective proportion.

Figure 3

Live cell fluorescence images of the uptake of FITC‐labeled DMSNs by HCEC cells after 2 h, 6 h or 20 h. FITC is represented in light blue and the plasma membrane in white.

Figure 4

Live cell fluorescence imaging of HCEC cells incubated with Ins‐FITC or DMSN‐Ins‐FITC for 2 h; (a) 2D images and (b) Zoom in the vertical cross‐section of the 3D cut stack. The plasma membrane is represented in white and, the fluorescence coming from Ins‐FITC in light blue.

Figure 5

In vitro assay based on the presence (no starvation) or absence (starvation) of insulin in the culture media of HCEC cells prior to the incubation of different treatments: commercial human insulin at the concentration 10 or 15 μg mL⁻¹, insulin confined in pure DMSNs or thiol‐functionalized DMSN‐SH at the concentration equivalent to 15 μg mL⁻¹ of insulin. One‐way ANOVA and Fisher Test expressed significant difference by *p<0.05, **p<0.01, ***p<0.001.