Regulation of Butyrate-Induced Resistance through AMPK Signaling Pathway in Human Colon Cancer Cells.

Butyrates inhibit cell growth in colon cancer cells by inhibiting histone deacetylases. However, chronic exposure to butyrates induces butyrate resistance in colon cancer cells. The mechanism underlying the acquisition of resistance is not yet fully understood. Here, butyrate-resistant (BR) colon cancer cells were developed in HCT116, HT29, and SW480 human colon cancer cells and were confirmed by the increase in the inhibitory concentrations of cell growth by 50% (IC50) compared to their respective parental (PT) cells. Chronic exposure to butyrate induced autophagy via higher expression of Beclin-1 and LC3B-II. The AMP-activated protein kinase (AMPK) was downregulated along with the activation of Akt and mammalian target of rapamycin (mTOR) and decrease in acetyl-CoA carboxylase (ACC) in BR colon cancer cells compared to those in their respective PT cells. Activation of AMPK by AICAR treatment in BR colon cancer cells suppressed cell proliferation by inhibiting Akt and mTOR and activating ACC. Taken together, chronic exposure to butyrate increased butyrate resistance in human colon cancer by inducing protective autophagy through the downregulation of AMPK/ACC and activation of Akt/mTOR signaling. Activation of AMPK restored sensitivity to butyrate by the inhibition of Akt/mTOR, suggesting that AMPK could be a therapeutic target for BR colon cancers.

Effects of Diabetes Mellitus on the Disposition of Tofacitinib, a Janus Kinase Inhibitor, in Rats.

Tofacitinib, a Janus kinase inhibitor, was developed for the treatment of rheumatoid arthritis. Recently, it has been associated with an increased change in arthritis development in patients with diabetes. Herein, we evaluated the pharmacokinetics of tofacitinib after intravenous (10 mg/kg) and oral (20 mg/kg) administration to rats with streptozotocin-induced diabetes mellitus and control rats. Following intravenous administration of tofacitinib to rats with streptozotocin-induced diabetes mellitus, area under the plasma concentration-time curve from time zero to infinity of tofacitinib was significantly smaller (33.6%) than that of control rats. This might be due to the faster hepatic intrinsic clearance (112%) caused by an increase in the hepatic cytochrome P450 (CYP) 3A1(23) and the faster hepatic blood flow rate in rats with streptozotocin-induced diabetes mellitus than in control rats. Following oral administration, area under the plasma concentration-time curve from time zero to infinity of tofacitinib was also significantly smaller (55.5%) in rats with streptozotocin-induced diabetes mellitus than that in control rats. This might be due to decreased absorption caused by the higher expression of P-glycoprotein and the faster intestinal metabolism caused by the higher expression of intestinal CYP3A1(23), which resulted in the decreased bioavailability of tofacitinib (33.0%) in rats with streptozotocin-induced diabetes mellitus. In summary, our findings indicate that diabetes mellitus affects the absorption and metabolism of tofacitinib, causing faster metabolism and decreased intestinal absorption in rats with streptozotocin-induced diabetes mellitus.

Stabilizing Coacervate by Microfluidic Engulfment Induced by Controlled Interfacial Energy.

Low interfacial energy, an intrinsic property of complex coacervate, enables the complex coacervate to easily encapsulate desired cargo substances, making it widely used in encapsulation applications. Despite this advantage, the low interfacial energy of the complex coacervate makes it unstable against mechanical mixing, and changes in pH and salt concentration. Hence, a chemical cross-linker is usually added to enhance the stability of the complex coacervate at the expense of sacrificing all intrinsic properties of the coacervate, including phase transition of the coacervate from liquid to solid. In this study, we observed an abrupt increase in the interfacial energy of the coacervate phase in mineral oil. By controlling the interfacial energy of the coacervate phase using a microfluidic device, we successfully created double engulfed PEG-diacrylate (PEGDA) coacervate microparticles, named DEPOT, in which the coacervate is engulfed in a cross-linked PEGDA shell. The engulfed coacervate remained as a liquid phase, retained its original low interfacial energy property to encapsulate the desired cargo substances, and infiltrated into the target site by a simple solvent exchange from oil to water.

Short-Term Teriparatide and Recombinant Human Bone Morphogenetic Protein-2 for Regenerative Approach to Medication-Related Osteonecrosis of the Jaw: A Preliminary Study.

Our objective was to examine whether adjunct teriparatide administration and local application of recombinant human bone morphogenetic protein-2 (rhBMP-2) is beneficial for the regeneration of jaw bone in patients with medication-related osteonecrosis of the jaw (MRONJ). This study enrolled 17 patients diagnosed with MRONJ. All patients received sequestrectomy under general or local anesthesia with suspension of bisphosphonate. The bone regeneration ratio was compared on cone beam computed tomography (CBCT) scans, acquired immediately post-operation and after 6 months. The patients were divided into groups, based on their treatment regimens: teriparatide combined with rhBMP-2 (parathyroid hormone [PTH]+BMP), rhBMP-2 (BMP), and the control. Biochemical markers were also evaluated at the baseline (T0), 1 month (T1), and 3 months (T2) after surgery. Significant increase was observed in the values of the biochemical markers, serum osteocalcin, and serum C-terminal telopeptide cross-link of type I collagen, within 3 months of surgery in the PTH+BMP group, whereas the mean value in the BMP group did not show a significant change. In all groups, the MRONJ lesions were healed and new bone formation was detected in the CBCT images. The regeneration ratio was significantly greater in the group PTH+BMP than in the BMP and control groups. Significantly greater amount of bone formation was observed in the group PTH+BMP than in the BMP and control groups. Local application of rhBMP-2 alone also had a beneficial effect on bone regeneration but was not more significant than control. Based on these findings, administration of short-term teriparatide with rhBMP-2 in MRONJ patients may maximize the regeneration of bone after surgery. © 2017 American Society for Bone and Mineral Research.

Salt Triggers the Simple Coacervation of an Underwater Adhesive When Cations Meet Aromatic π Electrons in Seawater.

Adhesive systems in many marine organisms are postulated to form complex coacervates (liquid-liquid phase separation) through a process involving oppositely charged polyelectrolytes. Despite this ubiquitous speculation, most well-characterized mussel adhesive proteins are cationic and polyphenolic, and the pursuit of the negatively charged proteins required for bulk complex coacervation formation internally remains elusive. In this study, we provide a clue for unraveling this paradox by showing the bulky fluid/fluid separation of a single cationic recombinant mussel foot protein, rmfp-1, with no additional anionic proteins or artificial molecules, that is triggered by a strong cation-π interaction in natural seawater conditions. With the similar condition of salt concentration at seawater level (>0.7 M), the electrostatic repulsion between positively charged residues of mfp-1 is screened significantly, whereas the strong cation-π interaction remains unaffected, which leads to the macroscopic phase separation (i.e., bulky coacervate formation). The single polyelectrolyte coacervate shows interesting mechanical properties including low friction, which facilitates the secretion process of the mussel. Our findings reveal that the cation-π interaction modulated by salt is a key mechanism in the mussel adhesion process, providing new insights into the basic understanding of wet adhesion, self-assembly processes, and biological phenomena that are mediated by strong short-range attractive forces in water.

Molecular and structural basis of low interfacial energy of complex coacervates in water.

Complex coacervate refers to a phase-separated fluid, typically of two oppositely charged polyelectrolytes in solution, representing a complex fluid system that has been shown to be of essential interest to biological systems, as well as for soft materials processing owing to the expectation of superior underwater coating or adhesion properties. The significance and interest in complex coacervate fluids critically rely on its low interfacial tension with respect to water that, in turn, facilitates the wetting of macromolecular or material surfaces under aqueous conditions, provided there is attractive interaction between the polyelectrolyte constituents and the surface. However, the molecular and structural bases of these properties remain unclear. Recent studies propose that the formation of water-filled and bifluidic sponge-like nanostructured network, driven by the tuning of electrostatic interactions between the polyelectrolyte constituents or their complexes may be a common feature of complex coacervate fluids that display low fluid viscosity and low interfacial tension, but more studies are needed to verify the generality of these observations. In this review, we summarize representative studies of interfacial tension and ultrastructures of complex coacervate fluids. We highlight that a consensus property of the complex coacervate fluid is the observation of high or even bulk-like water dynamics within the dense complex coacervate phase that is consistent with a low cohesive energy fluid. Our own studies on this subject are enabled by the application of magnetic resonance relaxometry methods relying on spin labels tethered to polyelectrolyte constituents or added as spin labeled probe molecules that partition into the dense versus the equilibrium coacervate phase, permitting the extraction of information on local polymer dynamics, polymer packing and local water dynamics. We conclude with a snapshot of our current perspective on the molecular and structural bases of the low interfacial tension of complex coacervate fluids.

Recovery of inferior alveolar nerve injury after bilateral sagittal split ramus osteotomy (BSSRO): a retrospective study.

BACKGROUND: Bilateral sagittal split ramus osteotomy (BSSRO) is the most widely used mandibular surgical technique in orthognathic surgery and is easy to relocate the distal segments, accelerating bone repair by the large surface of bone contact. However, it can cause neurosensory dysfunction (NSD) or sensory loss by injury of the inferior alveolar nerve. The purpose of the present study was to evaluate NSD after BSSRO and modifiers at NSD recovery. METHODS: In this study, NSD characteristics after BSSRO from 2009 to 2014 at the Kyung Hee University Dental Hospital were evaluated. The pattern of sensory recovery over time was also evaluated based on factors such as field of sensory dysfunction, surgical procedure, presence of pre-operative facial asymmetry, and postoperative medications. RESULTS: Most of the patients had shown NSD immediately after orthognathic surgery. Among the 1192 sides of 596 patients, NSD was observed in 953 sides and 544 patients. Sexual predilection was shown in males (p value = 0.0062). In the asymmetric group of 132 patients, NSD was observed in 128 patients (96.97 %). In the symmetric group of 464 patients, NSD was observed in 416 patients (89.45 %); on the other hand, NSD was observed significantly higher in the asymmetric group (p = 0.025). NSD-associated factors were analyzed, and vitamin B12 may be beneficial for NSD recovery. CONCLUSIONS: There was a difference between the symmetric group and the asymmetric group in NSD recovery. Vitamin B12 can be regarded as an effective method to nerve recovery. However, a further prospective study is needed.

Sugary interfaces mitigate contact damage where stiff meets soft.

The byssal threads of the fan shell Atrina pectinata are non-living functional materials intimately associated with living tissue, which provide an intriguing paradigm of bionic interface for robust load-bearing device. An interfacial load-bearing protein (A. pectinata foot protein-1, apfp-1) with L-3,4-dihydroxyphenylalanine (DOPA)-containing and mannose-binding domains has been characterized from Atrina's foot. apfp-1 was localized at the interface between stiff byssus and the soft tissue by immunochemical staining and confocal Raman imaging, implying that apfp-1 is an interfacial linker between the byssus and soft tissue, that is, the DOPA-containing domain interacts with itself and other byssal proteins via Fe3(+)-DOPA complexes, and the mannose-binding domain interacts with the soft tissue and cell membranes. Both DOPA- and sugar-mediated bindings are reversible and robust under wet conditions. This work shows the combination of DOPA and sugar chemistry at asymmetric interfaces is unprecedented and highly relevant to bionic interface design for tissue engineering and bionic devices.

Bicontinuous Fluid Structure with Low Cohesive Energy: Molecular Basis for Exceptionally Low Interfacial Tension of Complex Coacervate Fluids.

An exceptionally low interfacial tension of a dense fluid of concentrated polyelectrolyte complexes, phase-separated from a biphasic fluid known as complex coacervates, represents a unique and highly sought-after materials property that inspires novel applications from superior coating to wet adhesion. Despite extensive studies and broad interest, the molecular and structural bases for the unique properties of complex coacervates are unclear. Here, a microphase-separated complex coacervate fluid generated by mixing a recombinant mussel foot protein-1 (mfp-1) as the polycation and hyaluronic acid (HA) as the polyanion at stoichiometric ratios was macroscopically phase-separated into a dense complex coacervate and a dilute supernatant phase to enable separate characterization of the two fluid phases. Surprisingly, despite up to 4 orders of magnitude differing density of the polyelectrolytes, the diffusivity of water in these two phases was found to be indistinguishable. The presence of unbound, bulk-like, water in the dense fluid can be reconciled with a water population that is only weakly perturbed by the polyelectrolyte interface and network. This hypothesis was experimentally validated by cryo-TEM of the macroscopically phase-separated dense complex coacervate phase that was found to be a bicontinuous and biphasic nanostructured network, in which one of the phases was confirmed by staining techniques to be water and the other polyelectrolyte complexes. We conclude that a weak cohesive energy between water-water and water-polyelectrolytes manifests itself in a bicontinuous network, and is responsible for the exceptionally low interfacial energy of this complex fluid phase with respect to virtually any surface within an aqueous medium.

Complexation and coacervation of like-charged polyelectrolytes inspired by mussels.

It is well known that polyelectrolyte complexes and coacervates can form on mixing oppositely charged polyelectrolytes in aqueous solutions, due to mainly electrostatic attraction between the oppositely charged polymers. Here, we report the first (to the best of our knowledge) complexation and coacervation of two positively charged polyelectrolytes, which provides a new paradigm for engineering strong, self-healing interactions between polyelectrolytes underwater and a new marine mussel-inspired underwater adhesion mechanism. Unlike the conventional complex coacervate, the like-charged coacervate is aggregated by strong short-range cation-π interactions by overcoming repulsive electrostatic interactions. The resultant phase of the like-charged coacervate comprises a thin and fragile polyelectrolyte framework and round and regular pores, implying a strong electrostatic correlation among the polyelectrolyte frameworks. The like-charged coacervate possesses a very low interfacial tension, which enables this highly positively charged coacervate to be applied to capture, carry, or encapsulate anionic biomolecules and particles with a broad range of applications.

Nanomechanical Contribution of Collagen and von Willebrand Factor A in Marine Underwater Adhesion and Its Implication for Collagen Manipulation.

Recent works on mussel adhesion have identified a load bearing matrix protein (PTMP1) containing von Willebrand factor (vWF) with collagen binding capability that contributes to the mussel holdfast by manipulating mussel collagens. Using a surface forces apparatus, we investigate for the first time, the nanomechanical properties of vWF-collagen interaction using homologous proteins of mussel byssus, PTMP1 and preCollagens (preCols), as collagen. Mimicking conditions similar to mussel byssus secretion (pH < 5.0) and seawater condition (pH 8.0), PTMP1 and preCol interact weakly in the "positioning" phase based on vWF-collagen binding and strengthen in "locked" phase due to the combined effects of electrostatic attraction, metal binding, and mechanical shearing. The progressive enhancement of binding between PTMP1 with porcine collagen under the aforementioned conditions is also observed. The binding mechanisms of PTMP1-preCols provide insights into the molecular interaction of the mammalian collagen system and the development of an artificial extracellular matrix based on collagens.

Recombinant mussel proximal thread matrix protein promotes osteoblast cell adhesion and proliferation.

BACKGROUND: von Willebrand factor (VWF) is a key load bearing domain for mamalian cell adhesion by binding various macromolecular ligands in extracellular matrix such as, collagens, elastin, and glycosaminoglycans. Interestingly, vWF like domains are also commonly found in load bearing systems of marine organisms such as in underwater adhesive of mussel and sea star, and nacre of marine abalone, and play a critical load bearing function. Recently, Proximal Thread Matrix Protein1 (PTMP1) in mussel composed of two vWF type A like domains has characterized and it is known to bind both mussel collagens and mammalian collagens. RESULTS: Here, we cloned and mass produced a recombinant PTMP1 from E. coli system after switching all the minor codons to the major codons of E. coli. Recombinant PTMP1 has an ability to enhance mouse osteoblast cell adhesion, spreading, and cell proliferation. In addition, PTMP1 showed vWF-like properties as promoting collagen expression as well as binding to collagen type I, subsequently enhanced cell viability. Consequently, we found that recombinant PTMP1 acts as a vWF domain by mediating cell adhesion, spreading, proliferation, and formation of actin cytoskeleton. CONCLUSIONS: This study suggests that both mammalian cell adhesion and marine underwater adhesion exploits a strong vWF-collagen interaction for successful wet adhesion. In addition, vWF like domains containing proteins including PTMP1 have a great potential for tissue engineering and the development of biomedical adhesives as a component for extra-cellular matrix.

Structural basis for the recognition of lysozyme by MliC, a periplasmic lysozyme inhibitor in Gram-negative bacteria.

Lysozymes are an important component of the innate immune system of animals that hydrolyze peptidoglycan, the major bacterial cell wall constituent. Many bacteria have contrived various means of dealing with this bactericidal enzyme, one of which is to produce lysozyme inhibitors. Recently, a novel family of bacterial lysozyme inhibitors was identified in various Gram-negative bacteria, named MliC (membrane bound lysozyme inhibitor of C-type lysozyme). Here, we report the crystal structure of Pseudomonas aeruginosa MliC in complex with chicken egg white lysozyme. Combined with mutational study, the complex structure demonstrates that the invariant loop of MliC plays a crucial role in the inhibition of the lysozyme by its insertion to the active site cleft of the lysozyme, where the loop forms hydrogen and ionic bonds with the catalytic residues. Since MliC family members have been implicated as putative colonization or virulence factors, the structures and mechanism of action of MliC will be of relevance to the control of bacterial growth in animal hosts.