Targeted modulation of cell differentiation in distinct regions of the gastrointestinal tract via oral administration of differently PEG-PEI functionalized mesoporous silica nanoparticles

Targeted delivery of drugs is required to efficiently treat intestinal diseases such as colon cancer and inflammation. Nanoparticles could overcome challenges in oral administration caused by drug degradation at low pH and poor permeability through mucus layers, and offer targeted delivery to diseased cells in order to avoid adverse effects. Here, we demonstrate that functionalization of mesoporous silica nanoparticles (MSNs) by polymeric surface grafts facilitates transport through the mucosal barrier and enhances cellular internalization. MSNs functionalized with poly(ethylene glycol) (PEG), poly(ethylene imine) (PEI), and the targeting ligand folic acid in different combinations are internalized by epithelial cells in vitro and in vivo after oral gavage. Functionalized MSNs loaded with γ-secretase inhibitors of the Notch pathway, a key regulator of intestinal progenitor cells, colon cancer, and inflammation, demonstrated enhanced intestinal goblet cell differentiation as compared to free drug. Drug-loaded MSNs thus remained intact in vivo, further confirmed by exposure to simulated gastric and intestinal fluids in vitro. Drug targeting and efficacy in different parts of the intestine could be tuned by MSN surface modifications, with PEI coating exhibiting higher affinity for the small intestine and PEI–PEG coating for the colon. The data highlight the potential of nanomedicines for targeted delivery to distinct regions of the tissue for strict therapeutic control

Insights into the mechanical properties of epithelial cells: the effects of shear stress on the assembly and remodeling of keratin intermediate filaments

The effects of shear stress on the keratin intermediate filament (KIF) cytoskeleton of cultured human alveolar epithelial (A549) cells have been investigated. Under normal culture conditions, immunofluorescence revealed a delicate network of fine tonofibrils containing KIFs, together with many nonfilamentous, keratin-containing “particles,” mostly containing either keratin 8 (K8) or 18 (K18), but not both. Triton X-100 extracted ∼10% of the cellular keratin, and this was accompanied by a loss of the particles but not the KIFs. Shear stress dramatically reduced the soluble keratin component and transformed the fine bundles of KIFs into thicker, “wavy” tonofibrils. Both effects were accompanied by the disappearance of most keratin particles and by increased phosphorylation of K8 and K18 on serine residues 73 and 33, respectively. The particles that remained after shearing were phosphorylated and were closely associated with KIFs. We suggest that keratin particles constitute a reservoir of protein that can be recruited into KIFs under flow, creating a more robust cytoskeleton able to withstand shear forces more effectively.—Flitney, E. W., Kuczmarski, E. R., Adam, S. A., Goldman, R. D. Insights into the mechanical properties of epithelial cells: the effects of shear stress on the assembly and remodeling of keratin intermediate filaments

Type II Keratins Are Phosphorylated on a Unique Motif during Stress and Mitosis in Tissues and Cultured Cells

Epithelial cell keratins make up the type I (K9–K20) and type II (K1–K8) intermediate filament proteins. In glandular epithelia, K8 becomes phosphorylated on S73 ((71)LLpSPL) in human cultured cells and tissues during stress, apoptosis, and mitosis. Of all known proteins, the context of the K8 S73 motif (LLS/TPL) is unique to type II keratins and is conserved in epidermal K5/K6, esophageal K4, and type II hair keratins, except that serine is replaced by threonine. Because knowledge regarding epidermal and esophageal keratin regulation is limited, we tested whether K4–K6 are phosphorylated on the LLTPL motif. K5 and K6 become phosphorylated in vitro on threonine by the stress-activated kinase p38. Site-specific anti-phosphokeratin antibodies to LLpTPL were generated, which demonstrated negligible basal K4–K6 phosphorylation. In contrast, treatment of primary keratinocytes and other cultured cells, and ex vivo skin and esophagus cultures, with serine/threonine phosphatase inhibitors causes a dramatic increase in K4–K6 LLpTPL phosphorylation. This phosphorylation is accompanied by keratin solubilization, filament reorganization, and collapse. K5/K6 LLTPL phosphorylation occurs in vivo during mitosis and apoptosis induced by UV light or anisomycin, and in human psoriatic skin and squamous cell carcinoma. In conclusion, type II keratins of proliferating epithelia undergo phosphorylation at a unique and conserved motif as part of physiological mitotic and stress-related signals