Self-crosslinkable hyaluronate-based hydrogels as a soft tissue filler.

With increasing interest in aging and skin care, the use of fillers to increase the volume of soft tissue volume is increasing globally. However, the side effects caused by the residual chemical crosslinking agents present in these fillers limit the effective application of commercialized filler products. Therefore, the development of a novel crosslinking system with a non-toxic chemical crosslinking agent is required to overcome the limitations of commercial hyaluronate (HA)-based fillers. In this paper, a new injectable hydrogel with enhanced mechanical properties, tissue adhesion, injectability, and biocompatibility is reported. The HA derivatives modified with catechol groups (HA-DA) were crosslinked by self-oxidation under in vivo physiological conditions (pH 7.4) without chemical crosslinkers to form hydrogels, which can be further accelerated by the dissolved oxygen in the body. The fabricated HA-DA filler showed excellent mechanical properties and could be easily injected with a low injection force. Further, the HA-DA filler stably attached to the injection site due to the tissue adhesion properties of the catechol groups, thus leading to an improved displacement stability. In addition, the HA-DA filler showed excellent cell viability, cell proliferation, and biocompatibility. Therefore, the HA-DA hydrogel is a novel soft tissue filler with great potential to overcome the limitations of commercial soft tissue fillers.

Bio-plotted hydrogel scaffold with core and sheath strand-enhancing mechanical and biological properties for tissue regeneration.

Three-dimensional bio-plotted scaffolds constructed from encapsulated biomaterials or so-called "bio-inks" have received much attention for tissue regeneration applications, as advances in this technology have enabled more precise control over the scaffold structure. As a base material of bio-ink, sodium alginate (SA) has been used extensively because it provides suitable biocompatibility and printability in terms of creating a biomimetic environment for cell growth, even though it has limited cell-binding moiety and relatively weak mechanical properties. To improve the mechanical and biological properties of SA, herein, we introduce a strategy using hydroxyapatite (HA) nanoparticles and a core/sheath plotting (CSP) process. By characterizing the rheological and chemical properties and printability of SA and SA/HA-blended inks, we successfully fabricated bio-scaffolds using CSP. In particular, the mechanical properties of the scaffold were enhanced with increasing concentrations of HA particles and SA hydrogel. Specifically, HA particles blended with the SA hydrogel of core strands enhanced the biological properties of the scaffold by supporting the sheath part of the strand encapsulating osteoblast-like cells. Based on these results, the proposed scaffold design shows great promise for bone-tissue regeneration and engineering applications.

3D Printing of Bone-Mimetic Scaffold Composed of Gelatin/ß-Tri-Calcium Phosphate for Bone Tissue Engineering.

3D printed scaffolds composed of gelatin and ß-tri-calcium phosphate (ß-TCP) as a biomimetic bone material are fabricated, thereby providing an environment appropriate for bone regeneration. The Ca2+ in ß-TCP and COO- in gelatin form a stable electrostatic interaction, and the composite scaffold shows suitable rheological properties for bioprinting. The gelatin/ß-TCP scaffold is crosslinked with glutaraldehyde vapor and unreacted aldehyde groups which can cause toxicity to cells is removed by a glycine washing. The stable binding of the hydrogel is revealed as a result of FTIR and degradation rate. It is confirmed that the composite scaffold has compressive strength similar to that of cancellous bone and 60 wt% ß-TCP groups containing 40 wt% gelatin have good cellular activity with preosteoblasts. Also, in the animal experiments, the gelatin/ß-TCP scaffold confirms to induce bone formation without any inflammatory responses. This study suggests that these fabricated scaffolds can serve as a potential bone substitute for bone regeneration.

Dual-crosslinked methylcellulose hydrogels for 3D bioprinting applications.

Thermo-sensitive methylcellulose (MC) hydrogel has been widely used as a scaffold material for biomedical applications. However, due to its poor mechanical properties, the MC-based hydrogel has rarely been employed in 3D bioprinting for tissue engineering scaffolds. In this study, the dual crosslinkable tyramine-modified MC (MC-Tyr) conjugate was prepared via a two-step synthesis, and its hydrogel showed excellent mechanical properties and printability for 3D bioprinting applications. The MC-Tyr conjugate formed a dual-crosslinked hydrogel by modulating the temperature and/or visible light. A combination of reversible physical crosslinking (thermal crosslinking) and irreversible chemical crosslinking (photocrosslinking) was used in this dual crosslinked hydrogel. Also, the photocrosslinking of MC-Tyr solution was facilitated by visible light exposure in the presence of biocompatible photoinitiators (riboflavin, RF and riboflavin 5'-monophophate, RFp). The RF and RFp were used to compare the cytotoxicity and salting-out effect of MC-Tyr hydrogel, as well as the initiation ability, based on the difference in chemical structure. Also, the influence of the printing parameters on the printed MC hydrogel was investigated. Finally, the cell-laden MC-Tyr bioink was successfully extruded into stable 3D hydrogel constructs with high resolution via a dual crosslinking strategy. Furthermore, the MC-Tyr scaffolds showed excellent cell viability and proliferation.

Fabrication of 3D plotted scaffold with microporous strands for bone tissue engineering.

Scaffold porosity has played a key role in bone tissue engineering aimed at effective tissue regeneration, by promoting cell attachment, proliferation, and osteogenic differentiation for new bone formation. Three-dimensional plotting systems (3DPSs) have been widely used to introduce porosity to the scaffold; however, introducing certain features in the scaffold strands that improve bone tissue regeneration remains a challenge. In this work, we fabricated bone tissue scaffolds with macro- and microporous structural features using a 3DPS and non-solvent-induced phase separation method. This approach allowed both macro- and micropores to be created in the scaffold strands. The surface morphology and mechanical and degradation properties of the perforated scaffolds were characterized carefully. Human marrow stromal cells were cultured on the scaffolds and then analyzed in vitro to assess scaffold bio-function. The highly porous scaffold exhibited mechanical properties similar to those of cancellous bone. Cell attachment, proliferation, and differentiation were significantly higher in porous scaffold compared to its nonporous counterpart. These results suggest that highly porous scaffolds have tremendous potential as a bone tissue regeneration platform.

Visible-light-induced hyaluronate hydrogel for soft tissue fillers.

Hyaluronic acid (HA) is widely used as a filler owing to its excellent biocompatibility and biodegradability. However, commercial HA-based filler products have some limitations and can cause side effects due to the presence of residual chemical crosslinking agents. In this study, tyramine (Tyr) was introduced into HA to impart photocrosslinking ability to HA, and a photocrosslinked hydrogel was formed using a less toxic vitamin B2 derivative as a photoinitiator. For injection, an injectable filler was prepared by converting the photocrosslinked hydrogel to a microgel form. The crosslinking of the tyramine-modified HA (HA-Tyr) hydrogel, which can be applied as a soft tissue filler, increased with an increase in the irradiation time, and the crosslinked hydrogel showed excellent mechanical strength, elastic recovery rate, and injectability. It also showed non-cytotoxicity and cell proliferation behavior in fibroblasts. Therefore, injectable HA hydrogels have great potential as an alternative to conventional commercial dermal fillers.

Algicidal effect of hybrid peptides as potential inhibitors of harmful algal blooms.

OBJECTIVES: To biochemically characterize synthetic peptides to control harmful algal blooms (HABs) that cause red tides in marine water ecosystems. RESULTS: We present an analysis of several short synthetic peptides and their efficacy as algicidal agents. By altering the amino acid composition of the peptides we addressed the mode of algicidal action and determine the optimal balance of cationic and hydrophobic content for killing. In a controlled setting, these synthetic peptides disrupted both plasma and chloroplast membranes of several species known to result in HABs. This disruption was a direct result of the hydrophobic and cationic content of the peptide. Furthermore, by using an anti-HAB bioassay in scallops, we determined that these peptides were algicidal without being cytotoxic to other marine organisms. CONCLUSIONS: These synthetic peptides may prove promising for general marine ecosystem remediation where HABs have become widespread and resulted in serious economic loss.

Cyclo(phenylalanine-proline) induces DNA damage in mammalian cells via reactive oxygen species.

Cyclo(phenylalanine-proline) is produced by various organisms such as animals, plants, bacteria and fungi. It has diverse biological functions including anti-fungal activity, anti-bacterial activity and molecular signalling. However, a few studies have demonstrated the effect of cyclo(phenylalanine-proline) on the mammalian cellular processes, such as cell growth and apoptosis. In this study, we investigated whether cyclo(phenylalanine-proline) affects cellular responses associated with DNA damage in mammalian cells. We found that treatment of 1 mM cyclo(phenylalanine-proline) induces phosphorylation of H2AX (S139) through ATM-CHK2 activation as well as DNA double strand breaks. Gene expression analysis revealed that a subset of genes related to regulation of reactive oxygen species (ROS) scavenging and production is suppressed by the cyclo(phenylalanine-proline) treatment. We also found that cyclo(phenylalanine-proline) treatment induces perturbation of the mitochondrial membrane, resulting in increased ROS, especially superoxide, production. Collectively, our study suggests that cyclo(phenylalanine-proline) treatment induces DNA damage via elevation of ROS in mammalian cells. Our findings may help explain the mechanism underlying the bacterial infection-induced activation of DNA damage response in host mammalian cells.

N-terminal truncated UCH-L1 prevents Parkinson's disease associated damage.

Ubiquitin C-terminal hydrolase-L1 (UCH-L1) has been proposed as one of the Parkinson's disease (PD) related genes, but the possible molecular connection between UCH-L1 and PD is not well understood. In this study, we discovered an N-terminal 11 amino acid truncated variant UCH-L1 that we called NT-UCH-L1, in mouse brain tissue as well as in NCI-H157 lung cancer and SH-SY5Y neuroblastoma cell lines. In vivo experiments and hydrogen-deuterium exchange (HDX) with tandem mass spectrometry (MS) studies showed that NT-UCH-L1 is readily aggregated and degraded, and has more flexible structure than UCH-L1. Post-translational modifications including monoubiquitination and disulfide crosslinking regulate the stability and cellular localization of NT-UCH-L1, as confirmed by mutational and proteomic studies. Stable expression of NT-UCH-L1 decreases cellular ROS levels and protects cells from H2O2, rotenone and CCCP-induced cell death. NT-UCH-L1-expressing transgenic mice are less susceptible to degeneration of nigrostriatal dopaminergic neurons seen in the MPTP mouse model of PD, in comparison to control animals. These results suggest that NT-UCH-L1 may have the potential to prevent neural damage in diseases like PD.

Disruption of microtubules sensitizes the DNA damage-induced apoptosis through inhibiting nuclear factor κB (NF-κB) DNA-binding activity.

The massive reorganization of microtubule network involves in transcriptional regulation of several genes by controlling transcriptional factor, nuclear factor-kappa B (NF-κB) activity. The exact molecular mechanism by which microtubule rearrangement leads to NF-κB activation largely remains to be identified. However microtubule disrupting agents may possibly act in synergy or antagonism against apoptotic cell death in response to conventional chemotherapy targeting DNA damage such as adriamycin or comptothecin in cancer cells. Interestingly pretreatment of microtubule disrupting agents (colchicine, vinblastine and nocodazole) was observed to lead to paradoxical suppression of DNA damage-induced NF-κB binding activity, even though these could enhance NF-κB signaling in the absence of other stimuli. Moreover this suppressed NF-κB binding activity subsequently resulted in synergic apoptotic response, as evident by the combination with Adr and low doses of microtubule disrupting agents was able to potentiate the cytotoxic action through caspase-dependent pathway. Taken together, these results suggested that inhibition of microtubule network chemosensitizes the cancer cells to die by apoptosis through suppressing NF-κB DNA binding activity. Therefore, our study provided a possible anti-cancer mechanism of microtubule disrupting agent to overcome resistance against to chemotherapy such as DNA damaging agent.

Regulation of polar flagellum genes is mediated by quorum sensing and FlhDC in Burkholderia glumae.

The bacterium Burkholderia glumae causes rice grain rot by producing toxoflavin, whose expression is regulated by quorum sensing (QS). We report a major deviation from the current paradigm for the regulation of bacterial polar flagellum genes. The N-octanoyl homoserine lactone (C8-HSL)-deficient mutant of B. glumae is aflagellate and has lost the ability to swim and swarm at 37 degrees C. Mutagenesis of the bacterium with the mini-Tn5rescue identified an IclR-type transcriptional regulator, called QsmR, which is important for flagellum formation. TofR, which is a cognate C8-HSL receptor, activated qsmR expression by binding directly to the qsmR promoter region. From the flagellum gene cluster, we identified flhDC homologues that are directly activated by QsmR. FlhDC in turn activates the expression of genes involved in flagellum biosynthesis, motor functions and chemotaxis in B. glumae. Non-motile qsmR, fliA and flhDC mutants produced toxoflavin, but lost pathogenicity for rice. The unexpected discovery of FlhDC in a polarly flagellate bacterium suggests that exceptions to the typical regulatory mechanisms of flagellum genes exist in Gram-negative bacteria. The finding that functional flagella play critical roles in the pathogenicity of B. glumae suggests that either QS or flagellum formation constitutes a good target for the control of rice grain rot.