Thermoresponsive System Research Articles

Temperature-Triggered Colloidal Gelation through Well-Defined Grafted Polymeric Surfaces.

Sufficiently strong interparticle attractions can lead to aggregation of a colloidal suspension and, at high enough volume fractions, form a mechanically rigid percolating network known as a colloidal gel. We synthesize a model thermo-responsive colloidal system for systematically studying the effect of surface properties, grafting density and chain length, on the particle dynamics within colloidal gels. After inducing an attraction between particles by heating, aggregates undergo thermal fluctuation which we observe and analyze microscopically; the magnitude of the variance in bond angle is larger for lower grafting densities. Macroscopically, a clear increase of the linear mechanical behavior of the gels on both the grafting density and chain length arises, as measured by rheology, which is inversely proportional to the magnitude of local bond angle fluctuations. This colloidal system will allow for further elucidation of the microscopic origins to the complex macroscopic mechanical behavior of colloidal gels including bending modes within the network.

Defined and Scalable Differentiation of Human Oligodendrocyte Precursors from Pluripotent Stem Cells in a 3D Culture System

SummaryOligodendrocyte precursor cells (OPCs) offer considerable potential for the treatment of demyelinating diseases and injuries of the CNS. However, generating large quantities of high-quality OPCs remains a substantial challenge that impedes their therapeutic application. Here, we show that OPCs can be generated from human pluripotent stem cells (hPSCs) in a three-dimensional (3D), scalable, and fully defined thermoresponsive biomaterial system. We used CRISPR/Cas9 to create a NKX2.2-EGFP human embryonic stem cell reporter line that enabled fine-tuning of early OPC specification and identification of conditions that markedly increased the number of OLIG2+ and NKX2.2+ cells generated from hPSCs. Transplantation of 50-day-old OPCs into the brains of NOD/SCID mice revealed that progenitors generated in 3D without cell selection or purification subsequently engrafted, migrated, and matured into myelinating oligodendrocytes in vivo. These results demonstrate the potential of harnessing lineage reporter lines to develop 3D platforms for rapid and large-scale production of OPCs.

Development of a rapid in vitro tissue deadhesion system using the thermoresponsive sol-gel transition of hydroxybutyl chitosan

In regenerative medicine, it has become increasingly important to collect cultured tissues using non-invasive methods. Enzymatic deadhesion is normally used to collect cells, but this method cannot be used when trying to collect whole tissues in order to avoid damaging cell–cell interactions. In order to resolve this issue, a thermoresponsive culture dish with poly(N-isopropyl acrylamide) (PNIPAAm) has been employed. This system can change its hydrophobicity depending on temperature. Thus, tissues can attach above 37 °C and be detached below 20 °C. However, the PNIPAAm system has some issues related to cost and detachment time. In this study, we developed a novel thermoresponsive detachment system using a polysaccharide derivative. We chose hydroxy butyl chitosan (HBC) as the thermoresponsive polymer because of its high biocompatibility and rapid phase transition. We developed a novel method of HBC synthesis in conditions that were milder than previously reported. We used spin-coating to make a thin coating on two kinds of culture dishes with various concentrations of HBC solution. Seeded cells attached to the surface at less than 0.5 mg/ml HBC coating concentration, and they could be successfully detached by simply lowering the temperature of the suspension dishes without enzymatic treatment; the cells took only 5–20 min to detach. To evaluate this system, we measured three metrics related to cell culturing on culture dishes: initial attachment rate, detachment rate and tissue detachment time. The study revealed that tissues could be detached faster on the suspension dishes used in this study than on PNIPAAm grafted dishes when HBC was coated at 0.5 mg/ml. We successfully developed a novel tissue detachment system using HBC. These results suggest that smart polymers may be useful in regenerative medicine.

Development of an injectable pseudo-bone thermo-gel for application in small bone fractures

A pseudo-bone thermo-gel was synthesized and evaluated for its physicochemical, mechanical and rheological properties, with its application to treat small bone fractures. The pseudo-bone thermo-gel was proven to have thermo-responsive properties, behaving as a solution in temperatures below 25°C, and forming a gelling technology when maintained at physiological conditions. Poly propylene fumerate (PPF), Pluronic F127 and PEG-PCL-PEG were strategically blended, obtaining a thermo-responsive delivery system, to mimic the mechanical properties of bone with sufficient matrix hardness and resilience. A Biopharmaceutics Classification System (BCS) class II drug, simvastatin, was loaded in the pseudo-bone thermo-gel, selected for its bone healing properties. In vitro release analysis was undertaken on a series of experimental formulations, with the ideal formulations obtaining its maximum controlled drug release profile up to 14days. Ex vivo studies were undertaken on an induced 4mm diameter butterfly-fractured osteoporotic human clavicle bone samples. X-ray, ultrasound as well as textural analysis, undertaken on the fractured bones before and after treatment displayed significant bone filling, matrix hardening and matrix resilience properties. These characteristics of the pseudo-bone thermo-gel thus proved significant potential for application in small bone fractures.

Stiffness-Controlled Thermoresponsive Hydrogels for Cell Harvesting with Sustained Mechanical Memory.

Most mechanobiological investigations focused on in situ mechanical regulation of cells on stiffness-controlled substrates with few downstream applications, as it is still challenging to harvest and expand mechanically primed cells by enzymatic digestion (e.g., trypsin) without interrupting cellular mechanical memory between passages. This study develops thermoresponsive hydrogels with controllable stiffness to generate mechanically primed cells with intact mechanical memory for augmented wound healing. No significant cellular property alteration of the fibroblasts primed on thermoresponsive hydrogels with varied stiffness has been observed through thermoresponsive harvesting. When reseeding the harvested cells for further evaluation, softer hydrogels are proven to better sustain the mechanical priming effects compared to rigid tissue culture plate, which indicates that both the stiffness-controlled substrate and thermoresponsive harvesting are required to sustain cellular mechanical memory between passages. Moreover, epigenetics analysis reveals that thermoresponsive harvesting could reduce the rearrangement and loss of chromatin proteins compared to that of trypsinization. In vivo wound healing using mechanically primed fibroblasts shows featured epithelium and sebaceous glands, which indicates augmented skin recovery compared with trypsinized fibroblasts. Thus, the thermoresponsive hydrogel-based cell harvesting system offers a powerful tool to investigate mechanobiology between cell passages and produces abundant cells with tailored mechanical priming properties for cell-based applications.

On-command controlled drug release by diels-Alder reaction using Bi-magnetic core/shell nano-carriers

A novel bi-functional thermo-responsive system, consisting of core/shell bi-magnetic nanoparticles with furan surface functionality, is bonded with N-(2-Carboxyethyl)maleimide through Diels-Alder reaction. The chemotherapeutics doxorubicin is attached onto the surface, with a high loading efficiency of 92%. This system with high responsiveness to a high frequency external alternating magnetic field shows a very good therapeutic efficiency in hyperthermia and drug release at relatively low temperatures (50°C). Polyhedron-shaped bi-magnetic nanoparticles (Zn0.4Co0.6Fe2O4@Zn0.4Mn0.6Fe2O4) exhibit a significant increase of the specific energy absorption rate up to 455W/g compared with the core nanoparticles (200W/g). Real-time florescence spectroscopy studies demonstrate rapid release of doxorubicin up to 50% in 5min and up to 92% after 15min upon exposure to high frequency external alternating magnetic field. The stability is evaluated for 8 weeks in phosphate buffer saline with a doxorubicin payload of 85%. In vitro studies using standard MTT cell assays with HeLa and Hep G2 lines prove an excellent biocompatibility with about 90% of cell viability after 24h of treatment within the highest concentration of functionalized magnetic nanoparticles (200μg/mL). The results indicate a controlled drug release mediated by thermo-responsive switching under applied alternating magnetic field.

Smart Multifunctional Magnetic Nanoparticle-Based Drug Delivery System for Cancer Thermo-Chemotherapy and Intracellular Imaging.

In this research, a thermoresponsive drug release system was synthesized, which encapsulated the magnetic nanoparticles Fe3O4 and the drug model 5-fluorouracil with thermosensitive polymer poly(N-isopropylacrylamide) (PNIPAM). Mesoporous SiO2 was used as the channel of drug release, which could enhance the rate of drug loading and reduce drug loss. Chitosan (CHI) is a natural cationic linear polymer. The results showed successful coating of chitosan and rhodamine 6G (R6G) on the surface of the SiO2 sphere. The intermolecular interactions of the nanocomposites were confirmed by Fourier transform infrared spectroscopy. R6G is a typical fluorochrome which could be applied for cell imaging. Fluorescent imaging studies by confocal laser scanning microscopy indicated that the prepared nanocomposites Fe3O4/PNIPAM/5-Fu@mSiO2-CHI/R6G could specifically target tumor cells. Therefore, our work shows great potential in drug delivery and cancer therapy.

Human Immunodeficiency Virus (HIV) Separation and Enrichment via the Combination of Antiviral Lectin Recognition and a Thermoresponsive Reagent System.

In order to improve the detection limit of existing HIV diagnostic assays, we explored the use of a temperature-responsive magnetic nanoparticle reagent system in conjunction with cyanovirin-N for HIV recognition to rapidly and efficiently concentrate viral particles from larger sample volumes, ~ 1ml. Cyanovirin-N (CVN) mutant, Q62C, was expressed, biotinylated, and then complexed with a thermally responsive polymer-streptavidin conjugate. Confirmation of protein expression/activity was performed using matrix assisted laser desorption/ionization (MALDI) and a TZM-bl HIV inhibition assay. Biotinylated CVN mutant recognition with gp120 was characterized using surface plasmon resonance (SPR). Virus separation and enrichment using a thermoresponsive magnetic nanoparticle reagent system were measured using RT-PCR. Biotinylated Q62C exhibited a KD of 0.6nM to gp120. The temperature-responsive binary reagent system achieved a maximum viral capture of nearly 100% HIV, 1 × 10(5) virus copies in 100μl, using pNIPAAm-Q62C within 30minutes. Additionally, the same reagent system achieved nearly 9-fold enrichment by processing a 10-times larger sample of 1000μl (Fig.3). This work demonstrated a temperature-responsive reagent system that provides enrichment of HIV using antiviral lectin CVN for recognition, which is potentially amenable for use in point-of-care settings.

Polymerization and Rheological Behavior of the Thermoresponsive Gelcasting System Based on N‐isopropylacrylamide

Recently, we developed an efficient method for rapid drying and improving relative density of the gelcast green body. In this work, polymerization and rheological behavior of the thermoresponsive gelcasting system based on N‐isopropylacrylamide were investigated. To initiate reaction of the system with 4 mL/L of catalyst, 0.25–0.5 mg/mL of initiator is considered optimal in the optimized temperature range 15–20°C. Suspensions with 40–55 vol.% solid loading still show shear‐thinning behavior, which is favorable for mixing, degassing, and casting. The final storage moduli of consolidated suspensions are approximately 104 Pa, which is sufficiently high for gelcast green bodies.

<b>Histological evaluation of the rat dental pulp after indirect capping with sildenafil or L-NAME incorporated into a bioadhesive thermoresponsive system

We evaluated the histological dental pulp state in vivo after indirect pulp capping using sildenafil or L G -nitro-L-arginine (L-NAME), incorporated into a new bioadhesive thermoresponsive system (BTS). Male Wistar rats were subjected to an upper and lower first molar class I cavity preparation followed by indirect pulp capping with sildenafil or L-NAME. Calcium hydroxide (CaOH 2 ) was used as a control. The teeth and surrounding bone were properly dissected and processed for Nissl’s staining. Pulp state was evaluated considering the morphological aspects of the inflammatory response, type of inflammatory infiltrate, organization of the odontoblast layer, blood vessel condition, and presence of abscesses or necrosis. The results were expressed as average of observations. The most intense inflammatory response was observed 3 days after the cavity preparation. No identified changes were detected in the dental pulp response of the molars treated with L-NAME compared with those treated with CaOH 2 . A dual effect was observed in the teeth treated with sildenafil. While low sildenafil concentration (0.015% w w -1 ) promoted effects comparable to CaOH 2 , at a higher concentration (0.15% w w -1 ), sildenafil caused a severe inflammatory response and pulp necrosis. This pioneering suggest that NO pathway activity may be a determinant in the process of dental pulp healing.

A thermoresponsive injectable delivery system for therapeutic cells to the retina

A thermoresponsive injectable delivery system for therapeutic cells to the retina

Formulation and Evaluation of a Mucoadhesive Thermoresponsive System Containing Brazilian Green Propolis for the Treatment of Lesions Caused by Herpes Simplex Type I

The aim of the present work was to develop a topical delivery system that contains Brazilian green propolis extract (PE-8) to increase efficiency and convenience when applied to herpetic lesions. The cytotoxicity and antiherpetic activity was determined in vitro and in vivo. The PE-8 was added to a system that contained poloxamer 407 and carbopol 934P. The in vitro characterization of the system included rheological studies, texture profile analysis, and mucoadhesion analysis. The PE-8 inhibited the virus during the phase of viral infection, induced virion damage, and exhibited an ability to protect cells from viral infection. The system had advantageous mucoadhesive properties, including a suitable gelation temperature of approximately 25°C for topical delivery, a desirable textural profile, and pseudoplastic behavior. The in vitro release study showed a rapid initial release of the PE-8 in the first 3 h, and the rate of drug release remained constant for up to 24 h. The system appeared to be macroscopically and microscopically innocuous to skin tissue. Therefore, the mucoadhesive thermoresponsive system that contained the PE-8 appears to be promising for increasing bioavailability and achieving prolonged release of the PE-8 when applied to skin lesions caused by herpes simplex virus type 1.

Synthesis of Thermoresponsive Amphiphilic Polyurethane Gel as a New Cell Printing Material near Body Temperature.

Waterborne polyurethane (PU) based on poly(ε-caprolactone) (PCL) diol and a second oligodiol containing amphiphilic blocks was synthesized in this study. The microstructure was characterized by dynamic light scattering (DLS), small-angle X-ray scattering (SAXS), and rheological measurement of the PU dispersion. The surface hydrophilicity measurement, infrared spectroscopy, wide-angle X-ray diffraction, mechanical and thermal analyses were conducted in solid state. It was observed that the presence of a small amount of amphiphilic blocks in the soft segment resulted in significant changes in microstructure. When 90 mol % PCL diol and 10 mol % amphiphilic blocks of poly(l-lactide)-poly(ethylene oxide) (PLLA-PEO) diol were used as the soft segment, the synthesized PU had a water contact angle of ∼24° and degree of crystallinity of ∼14%. The dispersion had a low viscosity below room temperature. As the temperature was raised to body temperature (37 °C), the dispersion rapidly (∼170 s) underwent sol-gel transition with excellent gel modulus (G' ≈ 6.5 kPa) in 20 min. PU dispersions with a solid content of 25-30% could be easily mixed with cells in sol state, extruded by a 3D printer, and deposited layer by layer as a gel. Cells remained alive and proliferating in the printed hydrogel scaffold. We expect that the development of novel thermoresponsive PU system can be used as smart injectable hydrogel and applied as a new type of bio-3D printing ink.

Screening and In Vitro Evaluation of Mucoadhesive Thermoresponsive System Containing Methylene Blue for Local Photodynamic Therapy of Colorectal Cancer.

Photodynamic therapy (PDT) with methylene blue (MB) constitutes a potentially useful modality for colorectal cancer treatment. The limitations of the formulations containing MB are problems of administration and the inability to get the closeness contact at the site during the appropriate residence time. Present study aimed to develop and characterize mucoadhesive thermoresponsive system containing MB designed as platform for colorectal cancer therapy. Formulations composed of different amounts of poloxamer 407 (Polox), Carbopol 934P (Carb), and MB were developed and characterized as rheological, compressional, mucoadhesive and syringeability properties, toxicity, photodynamic action, in vitro MB release profile, and ex vivo MB intestinal permeation. The different compositions resulted in formulations with distinctive macroscopic characteristics and wide range of gelation temperatures. The compressional flow, mucoadhesive, syringeability, and rheological properties were significantly influenced by temperature and/or composition. The MB release from formulation was governed by anomalous transport. In addition, it was observed that MB permeated the intestinal membrane; the formulation possesses photodynamic activity and low toxicity. The data obtained from the system composed of 20% Polox, 0.15% Carb, and 0.25% MB indicated a potentially functional role in PDT of the colorectal cancer and suggest it is worthy of clinical evaluation.

Development and preclinical pharmacokinetics of a novel subcutaneous thermoresponsive system for prolonged delivery of heparin

Heparin is still widely used for treatment and prevention of thromboembolic diseases. Due to specific physicochemical properties, it requires frequent parenteral injections. In this study we present the development and in vitro evaluation of an advanced delivery system for prolonged subcutaneous release of heparin. The delivery system consisted of an in situ forming thermoresponsive poloxamer-based platform combined with pH-responsive polyelectrolyte heparin/chitosan nanocomplexes. Thermoresponsive hydrogels were tested for gelation temperature, gel dissolution and in vitro heparin release, whereas polyelectrolyte nanocomplexes were physico-chemically characterized, as well as tested for in vitro cytotoxicity and in vitro heparin release. Hydrogel combined of two poloxamers demonstrated the highest gelation temperature (28.6°C), while the addition of hydroxypropyl methylcellulose prolonged gel dissolution. On the other hand, nanocomplexes’ dispersions, prepared at 1:1 heparin/chitosan mass ratio and in the concentration range 0.375–1.875mg/mL, demonstrated mean diameter <400nm and zeta potential >34mV. Pharmacokinetics of selected formulations (thermoresponsive hydrogel, nanocomplexes and a dual system consisting of nanocomplexes incorporated into thermoresponsive hydrogel) were studied in rats. Heparin plasma concentration-time profiles revealed a double-peak phenomenon, probably due to heparin diffusion inside the polymer matrix and gel dissolution. Pharmacokinetic parameters were determined by a non-linear mixed effects modeling approach. It was demonstrated that thermoresponsive hydrogel with heparin/chitosan nanocomplexes enabled the lowest absorption rate of heparin into systemic circulation and provided heparin concentration above the prophylaxis threshold for 5 days. In situ gelling thermoresponsive matrix combined with chitosan nanocomplexes present a promising delivery system for heparin, requiring less frequent administration during long-term treatment.

Reversible Calcium(II)‐Ion Binding through an Apparent pKa Shift of Thermosensitive Block‐Copolymer Micelles

AbstractThere is an increasing need for smart materials capable of removing multivalent ions from aqueous streams without the inconvenience of brine regeneration as in ion‐exchange processes. Herein, we present a thermoresponsive micellar system consisting of polystyrene–poly(methoxy diethyleneglycol acrylate) block copolymer surfactants modified with carboxylic acid end groups (PS‐PMDEGA‐COOH) that can be used to switch between the adsorption and desorption of divalent calcium(II) cations by a mild temperature trigger, thus providing a new type of thermoregenerable ion‐adsorbing materials. The switch of calcium(II)‐binding capacity is demonstrated to result from a shift in the pKa value of the carboxylic acid groups by the collapse and redissolution of the PMDEGA block and the associated change in local polarity.

Reversible Calcium(II)-Ion Binding through an Apparent pKa Shift of Thermosensitive Block-Copolymer Micelles.

There is an increasing need for smart materials capable of removing multivalent ions from aqueous streams without the inconvenience of brine regeneration as in ion-exchange processes. Herein, we present a thermoresponsive micellar system consisting of polystyrene-poly(methoxy diethyleneglycol acrylate) block copolymer surfactants modified with carboxylic acid end groups (PS-PMDEGA-COOH) that can be used to switch between the adsorption and desorption of divalent calcium(II) cations by a mild temperature trigger, thus providing a new type of thermoregenerable ion-adsorbing materials. The switch of calcium(II)-binding capacity is demonstrated to result from a shift in the pKa value of the carboxylic acid groups by the collapse and redissolution of the PMDEGA block and the associated change in local polarity.

Rheological property and drying mechanism of thermoresponsive gelcasting of ZnO

To solve the problem of drying gelcast green body, the thermoresponsive gel system which contains macromonomer graft chains was used in gelcasting of ZnO. The effects of the amount and length of graft chains macromonomer PIPAAm, the total amount of organic matters, and the solid loading on the rheological properties of suspensions were investigated, and the drying mechanism of gelcast green body was analyzed. The results show that ZnO suspensions with the gel system still display shear-thinning rheological behavior, but its viscosity increases with increasing the addition amount and relative molecular mass of PIPAAm graft chain, and the total organic matter content. The PIPAAm graft chains inhibit or even eliminate the formation of the “dense layer” on the surface of gelcast ZnO green bodies, and accelerate the drying of green bodies. The introduction of PIPAAm graft chains facilitates the shrinkage of the gelcast ZnO green bodies, which is a feasible method to increase the relative density of green bodies.

Nanostructured Cubosomes in a Thermoresponsive Depot System: An Alternative Approach for the Controlled Delivery of Docetaxel.

The aim of the present study was to develop and evaluate a thermoresponsive depot system comprising of docetaxel-loaded cubosomes. The cubosomes were dispersed within a thermoreversible gelling system for controlled drug delivery. The cubosome dispersion was prepared by dilution method, followed by homogenization using glyceryl monooleate, ethanol and Pluronic® F127 in distilled water. The cubosome dispersion was then incorporated into a gelling system prepared with Pluronic® F127 and Pluronic® F68 in various ratios to formulate a thermoresponsive depot system. The thermoresponsive depot formulations undergo a thermoreversible gelation process i.e., they exists as free flowing liquids at room temperature, and transforms into gels at higher temperatures e.g., body temperature, to form a stable depot in aqueous environment. The mean particle size of the cubosomes in the dispersion prepared with Pluronic® F127, with and without the drug was found to be 170 and 280 nm, respectively. The prepared thermoresponsive depot system was evaluated by assessing various parameters like time for gelation, injectability, gel erosion, and in-vitro drug release. The drug-release studies of the cubosome dispersion before incorporation into the gelling system revealed that a majority (∼97%) of the drug was released within 12 h. This formulation also showed a short lag time (∼3 min). However, when incorporated into a thermoresponsive depot system, the formulation exhibited an initial burst release of ∼21%, and released only ∼39% drug over a period of 12 h, thus indicating its potential as a controlled drug delivery system.

Postcasting Contraction: Improving the Density of Gelcast Nanoparticle Green Bodies with Heated Liquid Desiccants

To achieve high‐density green bodies, a novel route without needing high solids loading suspensions, that is postcontraction process has been developed. In this process, the wet green body is immersed in heated liquid desiccants (polyethylene glycol [PEG] aqueous solution), which is prepared by gelcasting with thermoresponsive gel system. The density of green bodies is improved through releasing water quickly and shrinking rapidly. It is attributed to the cooperative action of the interior aggregation forces stemming from the shrinkage of network‐structured polymer and the exterior osmotic pressure from PEG aqueous solution. Alumina nanoparticle green bodies with different solids loadings were prepared by gelcasting with PNIPAM Poly(N‐isopropylacrylamide) gel system. The postcontraction process was almost completed in 30 min in liquid at 50°C, and meanwhile the bodies were almost dried. The relative densities of all green bodies and sintered parts are in accordance with each other, around 43% and 94%, respectively.