Degradable rosin-ester-caprolactone graft copolymers.

We have carried out the synthesis of side-chain rosin-ester-structured poly(ε-caprolactone) (PCL) through a combination of ring-opening polymerization and click chemistry. Rosin structures are shown to be effectively incorporated into each repeat unit of caprolactone. This simple and versatile methodology does not require sophisticated purification of raw renewable biomass from nature. The rosin properties have been successfully imparted to the PCL polymers. The bulky hydrophenanthrene group of rosin increases the glass-transition temperature of PCL by >100 °C, whereas the hydrocarbon nature of rosin structures provides PCL excellent hydrophobicity with contact angle very similar to polystyrene and very low water uptake. The rosin-containing PCL graft copolymers exhibit full degradability and good biocompatibility. This study illustrates a general strategy to prepare a new class of renewable hydrocarbon-rich degradable biopolymers.

The encapsulation and intracellular delivery of trehalose using a thermally responsive nanocapsule.

The thermally responsive wall permeability of an empty core-shell structured Pluronic nanocapsule (together with its temperature dependent size and surface charge) was successfully utilized for encapsulation, intracellular delivery, and controlled release of trehalose, a highly hydrophilic small (M(W) = 342 D) molecule (a disaccharide of glucose) that is exceptional for long-term stabilization of biologicals (particularly at ambient temperatures). It was found that trehalose can be physically encapsulated in the nanocapsule using a soaking-freeze-drying-heating procedure. The nanocapsule is capable of physically withholding trehalose with negligible release in hours for cellular uptake at 37 degrees C when its wall permeability is low. A quick release of the encapsulated sugar can be achieved by thermally cycling the nanocapsule between 37 and 22 degrees C (or lower). A significant amount of trehalose (up to 0.3 M) can be delivered into NIH 3T3 fibroblasts by incubating the cells with the trehalose-encapsulated nanocapsules at 37 degrees C for 40 min. Moreover, cytotoxicity of the nanocapsule for the purpose of intracellular delivery of trehalose was found to be negligible. Altogether, the thermally responsive nanocapsule is effective for intracellular delivery of trehalose, which is critical for the long-term stabilization of mammalian cells at ambient temperatures and the eventual success of modern cell-based medicine.

Stability improvement and characterization of bioprinted pectin-based scaffold.

PURPOSE:: Bioprinting is an alternative method for constructing tissues/organs for transplantation. This study investigated the cross-linker influence and post-printing modification using oligochitosan and chitosan for stability improvement. METHODS:: Oligochitosan was tested as a novel cross-linker to replace Ca2+ for pectin-based bio-ink. Oligochitosan (2 kD) and different molecular weight of chitosan were used to modify the bioprinted scaffold. Fourier transform infrared (FTIR) spectroscopy and scanning electron microscopy (SEM) were used to characterize the scaffolds. RESULTS:: Oligochitosan failed to serve as a viable cross-linker. Successful post-printing modification was confirmed by FTIR and SEM analyses. CONCLUSION:: Regarding post-modification, chitosan-treated scaffolds showed enhanced stability compared to untreated scaffolds. In particular, scaffolds modified with 150 kD chitosan exhibited the highest stability.

Novel bioprinting method using a pectin based bioink.

One major challenge of bioprinting is to develop a viable bioink to act as an extracellular matrix. This study investigated a novel method for bioprinting using a pectin based bioink. Besides pectin, Pluronic® F-127 was incorporated into the bioink to obtain the desired shape during the initial bioprinting process at 37∘C. Once an object was printed it was treated with Ca2+ (pectin cross-linker) to create the final tissue/organ structure. The results indicated that pectin/Pluronic® F-127 is a potential bioink. Moreover, this methodology provides a novel and fast approach for bioprinting.

Development of a microscale red blood cell-shaped pectin-oligochitosan hydrogel system using an electrospray-vibration method: preparation and characterization.

PURPOSE: To develop and characterize a microscale pectin-oligochitosan hydrogel microcapsule system that could be applied in such biological fields as drug delivery, cell immobilization/encapsulation, and tissue engineering. METHODS: Microscale pectin-oligochitosan hydrogel microcapsules were prepared by using the vibration/electrostatic spray method. The morphology and chemistry of the hydrogel microcapsules were characterized by using scanning electron microscope (SEM) and Fourier Transform Infrared Spectroscopy (FTIR), respectively. The designed hydrogel microcapsule system was then used to study the responsiveness of the microcapsules to different simulated human body fluids as well as cell encapsulation. RESULTS: The designed hydrogel microcapsule system exhibited a large surface area-to-volume ratio (red blood cell-shaped) and great pH/enzymatic responsiveness. In addition, this system showed the potential for controlled drug delivery and three-dimensional cell culture. CONCLUSION: This system showed a significant potential not only for bioactive-agent delivery, especially to the lower gastrointestinal (GI) tract, but also as a three-dimensional niche for cell culture. In particular, the hydrogel microcapsule system could be used to create artificial red-blood-cells as well as blood substitutes.

Thermally responsive nanoparticle-encapsulated curcumin and its combination with mild hyperthermia for enhanced cancer cell destruction.

In this study, thermally responsive polymeric nanoparticle-encapsulated curcumin (nCCM) was prepared and characterized. The nCCM is ≈ 22 and 300 nm in diameter at 37 and 22 °C, respectively. The smaller size of the nCCM at 37 °C was found to significantly facilitate its uptake in vitro by human prostate adenocarcinoma PC-3 cancer cells. However, the intracellular nCCM decreases rapidly (rather than plateaus) after reaching its peak at ≈ 1.5 h during a 3-day incubation of the PC-3 cells with nCCM. Moreover, a mild hyperthermia (with negligible cytotoxicity alone) at 43 °C applied between 1 and 1.5 h during the 3-day incubation not only increases the peak uptake but also alters intracellular distribution of nCCM (facilitating its delivery into cell nuclei), which helps to retain a significantly much higher level of intracellular curcumin. These effects of mild hyperthermia could be due in part to the thermal responsiveness of the nCCM: they are more positively charged at 43 °C and can be more easily attracted to the negatively charged nuclear membrane to enter nuclei as a result of electrostatic interaction. Ultimately, a combination of the thermally responsive nCCM and mild hyperthermia significantly enhances the anticancer capability of nCCM, resulting in a more than 7-fold decrease in its inhibitory concentration to reduce cell viability to 50% (IC50). Further mechanistic studies suggest injury pathways associated with heat shock proteins 27 and 70 should contribute to the enhanced cancer cell destruction by inducing cell apoptosis and necrosis. Overall, this study demonstrates the potential of combining mild hyperthermia and thermally responsive nanodrugs such as nCCM for augmented cancer therapy.

One-step microfluidic generation of pre-hatching embryo-like core-shell microcapsules for miniaturized 3D culture of pluripotent stem cells.

A novel core-shell microcapsule system is developed in this study to mimic the miniaturized 3D architecture of pre-hatching embryos with an aqueous liquid-like core of embryonic cells and a hydrogel-shell of zona pellucida. This is done by microfabricating a non-planar microfluidic flow-focusing device that enables one-step generation of microcapsules with an alginate hydrogel shell and an aqueous liquid core of cells from two aqueous fluids. Mouse embryonic stem (ES) cells encapsulated in the liquid core are found to survive well (>92%). Moreover, ~20 ES cells in the core can proliferate to form a single ES cell aggregate in each microcapsule within 7 days while at least a few hundred cells are usually needed by the commonly used hanging-drop method to form an embryoid body (EB) in each hanging drop. Quantitative RT-PCR analyses show significantly higher expression of pluripotency marker genes in the 3D aggregated ES cells compared to the cells under 2D culture. The aggregated ES cells can be efficiently differentiated into beating cardiomyocytes using a small molecule (cardiogenol C) without complex combination of multiple growth factors. Taken together, the novel 3D microfluidic and pre-hatching embryo-like microcapsule systems are of importance to facilitate in vitro culture of pluripotent stem cells for their ever-increasing use in modern cell-based medicine.

Robust antimicrobial compounds and polymers derived from natural resin acids.

We report novel robust resin acid-derived antimicrobial agents that exhibit excellent antimicrobial activities against a broad spectrum of bacteria (6 Gram-positive and 7 Gram-negative) with selective lysis of microbial membranes over mammalian membranes. Our results indicate that hydrophobicity and unique structures of resin acids can be determining factors in dictating the antimicrobial activity.

Synthesis and characterization of thermally responsive Pluronic F127-chitosan nanocapsules for controlled release and intracellular delivery of small molecules.

In this study, we synthesized empty core-shell structured nanocapsules of Pluronic F127 and chitosan and characterized the thermal responsiveness of the nanocapsules in size and wall-permeability. Moreover, we determined the feasibility of using the nanocapsules to encapsulate small molecules for temperature-controlled release and intracellular delivery. The nanocapsules are ∼37 nm at 37 °C and expand to ∼240 nm when cooled to 4 °C in aqueous solutions, exhibiting >200 times change in volume. Moreover, the permeability of the nanocapsule wall is high at 4 °C (when the nanocapsules are swollen), allowing free diffusion of small molecules (ethidium bromide, MW = 394.3 Da) across the wall, while at 37 °C (when the nanocapsules are swollen), the wall-permeability is so low that the small molecules can be effectively withheld in the nanocapsule for hours. As a result of their thermal responsiveness in size and wall-permeability, the nanocapsules are capable of encapsulating the small molecules for temperature-controlled release and intracellular delivery into the cytosol of both cancerous (MCF-7) and noncancerous (C3H10T1/2) mammalian cells. The cancerous cells were found to take up the nanocapsules much faster than the noncancerous cells during 45 min incubation at 37 °C. Moreover, toxicity of the nanocapsules as a delivery vehicle was found to be negligible. The Pluronic F127-chitosan nanocapsules should be very useful for encapsulating small therapeutic agents to treat diseases particularly when it is combined with cryotherapy where the process of cooling and heating between 37 °C and hypothermic temperatures is naturally done.