Tailoring transfection for cardiomyocyte cell lines: balancing efficiency and toxicity in lipid versus polymer-based transfection methods in H9c2 and HL-1 cells.

Cardiovascular research relies heavily on the veracity of in vitro cardiomyocyte models, with H9c2 and HL-1 cell lines at the forefront due to their cardiomyocyte-like properties. However, the variability stemming from nonstandardized culturing and transfection methods poses a significant challenge to data uniformity and reliability. In this study, we introduce meticulously crafted protocols to enhance the culture and transfection of H9c2 and HL-1 cells, emphasizing the reduction of cytotoxic effects while improving transfection efficiency. Through the examination of polymer-based and lipid-based transfection methods, we offer a comparative analysis that underscores the heightened efficiency and reduced toxicity of these approaches. Our research provides an extensive array of step-by-step procedures designed to foster robust cell cultures and outlines troubleshooting practices to rectify issues of low transfection rates. We discuss the merits and drawbacks of both transfection techniques, equipping researchers with the knowledge to choose the most fitting method for their experimental goals. By offering a definitive guide to these cell lines' culturing and transfection, our work seeks to set a new standard in procedural consistency, ensuring that the cardiovascular research community can achieve more dependable and reproducible results, thereby pushing the boundaries of current methodologies toward impactful clinical applications.NEW & NOTEWORTHY We have developed standardized protocols that significantly reduce cytotoxicity and enhance transfection efficiency in H9c2 and HL-1 cardiomyocyte cell lines. Our detailed comparative analysis of polymer-based and lipid-based transfection methods has identified optimized approaches with superior performance. Accompanying these protocols are comprehensive troubleshooting strategies to address common issues related to low transfection rates. Implementing these protocols is expected to yield more consistent and reproducible results, driving the field of cardiovascular research toward impactful clinical breakthroughs.

TXB-001, a newly-developed polymer-conjugated anthracycline: Significantly lower adverse effects in animal models of alopecia and hand-foot syndrome.

Anthracycline anti-cancer drugs have been widely used in the treatment of several cancers; however, their use is limited by adverse effects (AEs). Alopecia is a common AE that is minimally invasive, but adversely affects mental health and reduces quality of life (QoL). Hand-foot syndrome (HFS) is a dose-limiting AE of DOXIL, a liposomal formulation of doxorubicin (DOX). Although it is not a life-threatening condition, HFS affects function and reduces QoL. TXB-001 is a new candidate polymer-conjugated anthracycline anti-cancer drug, and modified and optimized polymerized pirarubicin (THP), known as P-THP, is expected to have low toxicity and high efficacy. The anti-cancer effects of TXB-001 were examined using the 4T1 mouse model. An alopecia mouse model and HFS rat model were used to evaluate the alopecia- and HFS-inducing effects of TXB-001 and compare their severity with existing anthracycline anti-cancer drugs. A pharmacokinetic analysis of plasma as well as chest, palmar, and plantar skin samples after the single intravenous administration of DOXIL and TXB-001 to rats was also performed. The results obtained revealed that TXB-001 exerted similar anti-cancer effects to those of DOXIL in mice, weaker alopecia-inducing effects than DOX, DOXIL, and THP in mice, and no or markedly weaker HFS-like changes than DOXIL, which induced significant histopathological changes. The results of the pharmacokinetic analysis showed the accumulation of DOXIL, but not TXB-001, in skin, particularly palmar and plantar skin samples, and these differences were considered to contribute to their HFS-inducing effects.

Ultra-high-performance liquid chromatography with tandem mass spectrometry method for cellular toxicity and pharmacokinetic study of PEG1K polymers.

Polyethylene glycol (PEG) is one of the most commonly used polymers in drug delivery systems. The investigation of the pharmacokinetic behavior of PEG is important for revealing the toxicity and efficiency of PEG-related Nano-drug delivery systems. A high through-put and selective ultra-high-performance liquid chromatography with tandem mass spectrometry (UHPLC-MS/MS) method coupled with collision-induced dissociation (CID) in source technique was developed and validated to determine PEG1K polymers in cellular samples in this study. The countless precursor ions of PEG1K are dissociated in the source to generate numerous product ions which have different numbers of subunits. The transition of [M+H]+ precursor ions → product ions at m/z 177.1 (four subunits)→89.1 (two subunits) was selected to determine PEG1K due to its high sensitivity. The UHPLC-MS/MS method coupled with CID in the source showed good linearity over the range of 0.1-10 µg/mL. Intra-day and inter-day accuracies and precisions of the assay were all within ± 12.39%. The assay was successfully applied to a cellular pharmacokinetic study of PEG1K in human breast cancer cells. The cytotoxicity of PEG1K polymers was also studied and the results indicated that the cytotoxicity of PEG1K was not significant in the range of 5-1200 µg/mL.

Exploring structure/property relationships to health and environmental hazards of polymeric polyisocyanate prepolymer substances-3. Aquatic exposure and hazard of aliphatic diisocyanate-based prepolymers.

The water extractability and acute aquatic toxicity of seven aliphatic diisocyanate-based prepolymer substances were investigated to determine if lesser reactivity of the aliphatic isocyanate groups, as well as increased ionization potential of the expected (aliphatic amine-terminated) polymeric hydrolysis products, would influence their aquatic behavior compared to that of previously investigated aromatic diisocyanate-based prepolymers. At loading rates of 100 and 1,000 mg/L, only the substances having log Kow ≤9 exhibited more than 1% extractability in water, and a maximum of 66% water extractability was determined for a prepolymer having log Kow = 2.2. For the more hydrophobic prepolymer substances (log Kow values from 18-37), water extractability was negligible. High-resolution mass spectrometric analyses were performed on the water-accommodated fractions (WAF) of the prepolymers, which indicated the occurrence of primary aliphatic amine-terminated polymer species having backbones and functional group equivalent weights aligned to those of the parent prepolymers. Measurements of reduced surface tension and presence of suspended micelles in the WAFs further supported the occurrence of these surface-active cationic polymer species as hydrolysis products of the prepolymers. Despite these characteristics, the water-extractable hydrolysis products were practically non-toxic to Daphnia magna. All of the substances tested exhibited 48-h EL50 values of >1,000 mg/L, with one exception of EL50 = 157 mg/L. The results from this investigation support a grouping of the aliphatic diisocyanate-based prepolymers as a class of water-reactive polymer substances having predictable aquatic exposure and a uniformly low hazard potential, consistent with that previously demonstrated for the aromatic diisocyanate-based prepolymers.

Understanding ecotoxicological drivers and responses of freshwater green algae, Raphidocelis subcapitata, to cationic polyquaternium polymers.

Cationic polymer (CP) ecotoxicity is important to understand and investigate as they are widely used in industrial and consumer applications and have shown toxic effects in some aquatic organisms. CPs are identified as "polymers of concern" and are to be prioritized in upcoming regulatory reviews, (e.g., REACH). Algae have generally been found to be the most sensitive trophic level to CP. This study aimed at elucidating the magnitude of cationic polyquaternium toxicity towards algae and to understand key toxicological drivers. A suite of polyquaterniums with varying charge density (charged nitrogen moieties) and molecular weight were selected. Highly charged polyquaternium-6 and -16 were toxic towards the freshwater green microalgae Raphidocelis subcapitata with ErC50-values ranging between 0.12 and 0.41 mg/L. Lower charge density polyquaternium-10 materials had much lower toxicity with ErC50 > 200 mg/L, suggesting that charge density is an important driver of algal toxicity. These levels of toxicity were in line with historic CP data in literature. Algal agglomeration was observed in all tests but was not linked with impacts on algal growth rate. However, agglomeration can pose challenges in the technical conduct of tests and can impair interpretation of results. The toxicity mitigation potential of humic acid was also explored. The addition of 2-20 mg/L humic acid completely mitigated PQ6 and PQ16 toxicity at concentrations higher than clean water ErC50-values. CP toxicity mitigation has also been observed in fish and invertebrate tests, suggesting that CP mitigation should be accounted for in all trophic levels within an environmental safety framework.

Toxicity assessment of core-shell and superabsorbent polymers in cell-based systems.

The identification of health risks arising from occupational exposure to submicron/nanoscale materials is of particular interest and toxicological investigations designed to assess their hazardous properties can provide valuable insights. The core-shell polymers poly (methyl methacrylate)@poly (methacrylic acid-co-ethylene glycol dimethacrylate) [PMMA@P (MAA-co-EGDMA)] and poly (n-butyl methacrylate-co-ethylene glycol dimethacrylate)@poly (methyl methacrylate) [P (nBMA-co-EGDMA)@PMMA] could be utilized for the debonding of coatings and for the encapsulation and targeted delivery of various compounds. The hybrid superabsorbent core-shell polymers poly (methacrylic acid-co-ethylene glycol dimethacrylate)@silicon dioxide [P (MAA-co-EGDMA)@SiO2] could be utilized as internal curing agents in cementitious materials. Therefore, the characterization of their toxicological profile is essential to ensure their safety throughout manufacturing and the life cycle of the final products. Based on the above, the purpose of the present study was to assess the acute toxic effects of the above mentioned polymers on cell viability and on cellular redox state in EA. hy926 human endothelial cells and in RAW264.7 mouse macrophages. According to our results, the examined polymers did not cause any acute toxic effects on cell viability after any administration. However, the thorough evaluation of a panel of redox biomarkers revealed that they affected cellular redox state in a cell-specific manner. As regards EA. hy926 cells, the polymers disrupted redox homeostasis and promoted protein carbonylation. Concerning RAW264.7 cells, P (nBMA-co-EGDMA)@PMMA caused disturbances in redox equilibrium and special emphasis was placed on the triphasic dose-response effect detected in lipid peroxidation. Finally, P (MAA-co-EGDMA)@SiO2 activated cellular adaptive mechanisms in order to prevent from oxidative damage.

Plant extracts, polymers and new approach methods: Practical experience with skin sensitization assessment.

Over the last decade, research into methodologies to identify skin sensitization hazards has led to the adoption of several non-animal methods as OECD test guidelines. However, predictive accuracy beyond the chemical domains of the individual validation studies remains largely untested. In the present study, skin sensitization test results from in vitro and in chemico methods for 12 plant extracts and 15 polymeric materials are reported and compared to available in vivo skin sensitization data. Eight plant extracts were tested in the DPRA and h-CLAT, with the 2 out of 3 approach resulting in a balanced accuracy of 50%. The balanced accuracy for the 11 plant extracts assessed in the SENS-IS was 88%. Excluding 5 polymers inconclusive in vitro, the remainder, assessed using the 2 out of 3 approach, resulted in 63% balanced accuracy. The SENS-IS method, excluding one polymeric material due to technical inapplicability, showed 68% balanced accuracy. Although based on limited numbers, the results presented here indicate that some substance subgroups may not be in the applicability domains of the method used and careful analysis is required before positive or negative results can be accepted.

Safety Assessment of Polyfluorinated Polymers as Used in Cosmetics.

The Expert Panel for Cosmetic Ingredient Safety (Panel) reviewed the safety of 12 polyfluorinated polymers in cosmetic products; most of these ingredients have the reported function of film former in common. However, PTFE, the only ingredient that is reported as currently used in cosmetics, functions as a bulking agent and slip modifier, but not as a film former. The Panel reviewed data relevant to the safety of these ingredients under the intended conditions of use in cosmetic formulations, and concluded that PTFE and Hexafluoropropylene/Tetrafluoroethylene Copolymer are safe in cosmetics in the present practices of use and concentration described in the safety assessment; the data are insufficient to determine the safety of the 4 fluorinated-side-chain polymers and 6 fluorinated polyethers.

Sensitive visual detection of intracellular zinc ions based on signal-on polydopamine carbon dots.

The concentration of intracellular zinc ions is a significant clinical parameter for diagnosis. However, it is still a challenge for direct visual detection of zinc ions in cells at single-cell level. To address this issue, herein, water-soluble amino-rich polydopamine carbon quantum dots (PDA-CQDs) were successfully synthesized, with strong blue-green fluorescence as the probes for zinc ions detection in cells. The structure and properties of PDA-CQDs were confirmed by transmission electron microscopy (TEM), X-ray diffraction (XRD), Fourier transformed infrared (FT-IR), UV-visible spectrophotometry (UV-vis), and fluorescence spectroscopy. Importantly, by successfully linking salicylaldehyde (SA) to PDA-CQDs via nucleophilic reaction, the FL quenching and Zn ions induced FL-recovering system was built up, thus offering a signal-on platform for the detection of zinc ions. This PDA-CQDs-SA nanoprobe can be applied for the detection of Zn2+with a detection limit of 0.09µM, with good biocompatibility confirmed using cytotoxicity assay. Of significance, the results of fluorescence bioimaging showed that PDA-CQDs-SA is able to detect Zn2+in single-cell visually, with the detection limit of Zn ions in cells as low as 0.11µM per cell, which was confirmed using flow cytometry. Therefore, this work offers a potential probe for Zn2+detection in cells at single-cell level, towards the precise diagnosis of zinc ions related diseases.

Toxicity of polymer-modified CuS nanoclusters on zebrafish embryo development.

Despite the vast amount of research on the toxicity of copper-based nanoparticles, the toxicity of CuS nanoparticles is still largely unknown. Due to the application of CuS-based nanomaterials in biomedical engineering, it is necessary to study their potential toxicity and biological effects. In this study, we evaluated the toxicity of polymer-modified CuS nanoclusters (PATA3-C4@CuS) on embryo development through exposing zebrafish embryos to 1, 2.5, 5, 7.5, and 10 mg/L PATA3-C4@CuS at 0.75-h post-fertilization. The morphological results demonstrated that PATA3-C4@CuS at concentrations greater than 1 mg/L PATA3-C4@CuS induced abnormal phenotypes including smaller heads and eyes, pericardial edema, and epiboly retardation and it increased mortality, lowered the hatching rate, and inhibited swim bladder inflation. In situ hybridization and quantitative reverse transcription polymerase chain reaction showed that PATA3-C4@CuS could alter the expression patterns of tbxta, dlx3, and cstlb and increase the expression levels of wnt5 and wnt11, which suggested that PATA3-C4@CuS disrupts cell migration by increasing the levels of wnt5 and wnt11 during gastrulation. It was also discovered that PATA3-C4@CuS exposure caused a slow heart rate and smaller ventricles in zebrafish larvae. Immunofluorescence and behavioral analyses showed that PATA3-C4@CuS could damage the ventral projection of the primary motor neurons CaP, which was in accordance with the reduction in locomotion ability. Together, our data demonstrated that functional PATA3-C4@CuS could disrupt cell migration during gastrulation, affect cardiac development and function, and decrease locomotive activity.

Bulky Barbiturates as Non-Toxic Ionic Dye Insulators for Enhanced Emission in Polymeric Nanoparticles.

Bulky hydrophobic counterions (weakly coordinating anions) can insulate ionic dyes against aggregation-caused quenching (ACQ) and enable preparation of highly fluorescent dye-loaded nanoparticles (NPs) for bioimaging, biosensing and light harvesting. Here, we introduce a family of hydrophobic anions based on fluorinated C-acyl barbiturates with delocalized negative charge and bulky non-polar groups. Similarly to fluorinated tetraphenylborates, these barbiturates prevent ACQ of cationic dye alkyl rhodamine B inside polymer NPs made of biodegradable poly(lactic-co-glycolic acid) (PLGA). Their efficiency to prevent ACQ increases for analogues with higher acidity and bulkiness. Their structure controls dye-dye communication, yielding bright NPs with on/off switching or stable emission. They enhance dye encapsulation inside NPs, allowing intracellular imaging without dye leakage. Compared to fluorinated tetraphenylborates known as cytotoxic transmembrane ion transporters, the barbiturates display a significantly lower cytotoxicity. These chemically available and versatile barbiturate derivatives are promising counterion scaffolds for preparation of bright non-toxic fluorescent nanomaterials.

Cell viability and cytotoxicity of inkjet-printed flexible organic electrodes on parylene C.

This study reports on the fabrication of biocompatible organic devices by means of inkjet printing with a novel combination of materials. The devices were fabricated on Parylene C (PaC), a biocompatible and flexible polymer substrate. The contact tracks were inkjet-printed using a silver nanoparticle ink, while the active sites were inkjet-printed using a poly (3,4ethylenedioxythiophene)/polystyrene sulfonate (PEDOT:PSS) solution. To insulate the final device, a polyimide ink was used to print a thick film, leaving small open windows upon the active sites. Electrical characterization of the final device revealed conductivities in the order of 103 and 102 S.cm-1 for Ag and PEDOT based inks, respectively. Cell adhesion assays performed with PC-12 cells after 96 h of culture, and B16F10 cells after 24 h of culture, demonstrated that the cells adhered on top of the inks and cell differentiation occurred, which indicates Polyimide and PEDOT:PSS inks are non-toxic to these cells. The results indicate that PaC, along with its surface-treated variants, is a potentially useful material for fabricating cell-based microdevices.

Exploring the Toxicity, Lung Distribution, and Cellular Uptake of Rifampicin and Ascorbic Acid-Loaded Alginate Nanoparticles as Therapeutic Treatment of Lung Intracellular Infections.

Nanotechnology is a very promising technological tool to combat health problems associated with the loss of effectiveness of currently used antibiotics. Previously, we developed a formulation consisting of a chitosan and tween 80-decorated alginate nanocarrier that encapsulates rifampicin and the antioxidant ascorbic acid (RIF/ASC), intended for the treatment of respiratory intracellular infections. Here, we investigated the effects of RIF/ASC-loaded NPs on the respiratory mucus and the pulmonary surfactant. In addition, we evaluated their cytotoxicity for lung cells in vitro, and their biodistribution on rat lungs in vivo after their intratracheal administration. Findings herein demonstrated that RIF/ASC-loaded NPs display a favorable lung biocompatibility profile and a uniform distribution throughout lung lobules. RIF/ASC-loaded NPs were mainly uptaken by lung macrophages, their primary target. In summary, findings show that our novel designed RIF/ASC NPs could be a suitable system for antibiotic lung administration with promising perspectives for the treatment of pulmonary intracellular infections.

Novel Peptide-Polymer Conjugate with pH-Responsive Targeting/Disrupting Effects on Biomembranes.

Conjugating polymers to peptides has become a new strategy of designing functional antitumor agents for their improved stability and enhanced activity. In this paper, a novel peptide-polymer conjugate PEPc-PMAA with pH responsiveness was designed and synthesized. The isoelectric point of PEPc was studied by dynamic light scattering for the targeting effect. Also, the transmittances of PMAA at different pHs were measured using an ultraviolet-visible spectrophotometer for determining the triggering pH of the disrupting effect. The results showed that PEPc-PMAA was hydrophilic under neutral conditions and changed to be amphiphilic composed of positively charged PEPc and hydrophobic PMAA under acidic conditions. The interactions between PEPc-PMAA and mimic cells were investigated by the measurements of membrane fluidity and cargo leakage from 1,2-dipalmitoyl-sn-glycerol-3-phosphocholine and 1,2-dipalmitoyl-sn-glycerol-3-phospho-(1-rac-glycerol) (DPPG) liposomes. It proved that PEPc-PMAA caused a distinct membrane disturbance of the DPPG liposome at pH 5.5, resulting in more serious cargo leakage. Because of its targeting and disrupting effects on negatively charged biomembranes under acidic conditions, PEPc-PMAA showed its good potential as an antitumor agent.

Vasculature-on-chip for Assessment of Bioresorbable Scaffolds and Endothelial Barrier Integrity.

ABSTRACT: Endothelial cells adhere to one another through junctional structures formed by intercellular adhesion molecules. These intercellular proteins regulate barrier function in response to the hemodynamic shear rate and enable the selective passage of solutes and fluids across the endothelium. After endovascular device implantation, the endothelial barrier is compromised and becomes discontinuous, which increases permeability, allowing transmigration of leukocytes and lipoproteins and leading to the accumulation of lipid-laden foamy macrophages in the subendothelial space. Drug-coated bioresorbable vascular scaffold implants have been associated with unexpected thrombotic complications, which were not predicted in animals because of dissimilarities in endothelial regeneration and realignment between animals and humans. The development of a microengineered, microfluidics-based system of patterned channels lined with human endothelial and smooth muscle cells perfused with blood allows for the evaluation of endothelial function and barrier integrity. This review highlights the translational potential of vasculature-on-chip, which recreates the microphysiological milieu to evaluate the impact of drug-eluting bioresorbable vascular scaffolds on endothelial barrier integrity and to characterize polymer biodegradation behavior and drug release kinetic profiles over time.

Role of Structural Morphology of Commodity Polymers in Microplastics and Nanoplastics Formation: Fragmentation, Effects and Associated Toxicity in the Aquatic Environment.

With the continued growth in plastic production, its ubiquitous use and insufficient waste management and disposal, the increased levels of plastics in the environment have led to growing ecological concerns. The breakdown of these plastic macromolecules to smaller micro and nanosized particles and their detection in the aerial, aquatic, marine and terrestrial environments has been reviewed extensively, especially for thermoplastics. However, the formation of micro and nanoplastics has typically been explained as a physical abrasion process, largely overlooking the underlying chemical structure-morphology correlations to the degradation mechanisms of the plastics. This is particularly true for the common commodity thermosets. This review focuses on the degradation pathways for the most widely produced commodity thermoplastics and thermosets into microplastics (MP)s and nanoplastics (NP)s, as well as their behaviour and associated toxicity. Special emphasis is placed on NPs, which are associated with greater risks for toxicity compared to MPs, due to their higher surface area to volume ratios. This review also assesses the current state of standardized detection and quantification methods as well as comprehensive regulations for these fragments in the aquatic environment.

Ecotoxicity evaluation of polymeric nanoparticles loaded with ascorbic acid for fish nutrition in aquaculture.

BACKGROUND: Ascorbic acid (AA) is a micronutrient essential for the mechanisms of reproduction, growth, and defense in fish. However, the biosynthesis of this micronutrient does not occur in fish, so it must be supplied with food. A difficulty is that plain AA is unstable, due to the effects of light, high temperature, and oxygen, among others. The use of nanoencapsulation may provide protection and preserve the physicochemical characteristics of AA for extended periods of time, decreasing losses due to environmental factors. METHOD: This study evaluated the protective effect of nanoencapsulation in polymeric nanoparticles (chitosan and polycaprolactone) against AA degradation. Evaluation was made of the physicochemical stability of the nanoformulations over time, as well as the toxicological effects in zebrafish (Danio rerio), considering behavior, development, and enzymatic activity. For the statistical tests, ANOVA (two-way, significance of p < 0.05) was used. RESULTS: Both nanoparticle formulations showed high encapsulation efficiency and good physicochemical stability during 90 days. Chitosan (CS) and polycaprolactone (PCL) nanoparticles loaded with AA had mean diameters of 314 and 303 nm and polydispersity indexes of 0.36 and 0.28, respectively. Both nanosystems provided protection against degradation of AA exposed to an oxidizing agent, compared to plain AA. Total degradation of AA was observed after 7, 20, and 480 min for plain AA, the CS nanoparticle formulation, and the PCL nanoparticle formulation, respectively. For zebrafish larvae, the LC50 values were 330.7, 57.4, and 179.6 mg/L for plain AA, the CS nanoparticle formulation, and the PCL nanoparticle formulation, respectively. In toxicity assays using AA at a concentration of 50 mg/L, both types of nanoparticles loaded with AA showed lower toxicity towards the development of the zebrafish, compared to plain AA at the same concentration. Although decreased activity of the enzyme acetylcholinesterase (AChE) did not affect the swimming behavior of zebrafish larvae in the groups evaluated, it may have been associated with the observed morphometric changes, such as curvature of the tail. CONCLUSIONS: This study showed that the use of nanosystems is promising for fish nutritional supplementation in aquaculture. In particular, PCL nanoparticles loaded with AA seemed to be most promising, due to higher protection against AA degradation, as well as lower toxicity to zebrafish, compared to the chitosan nanoparticles. The use of nanotechnology opens new perspectives for aquaculture, enabling the reduction of feed nutrient losses, leading to faster fish growth and improved sustainability of this activity.

Hollow polydopamine nanoparticles loading with peptide RL-QN15: a new pro-regenerative therapeutic agent for skin wounds.

BACKGROUND: Although the treatments of skin wounds have greatly improved with the increase in therapeutic methods and agents, available interventions still cannot meet the current clinical needs. Therefore, the development of new pro-regenerative therapies remains urgent. Owing to their unique characteristics, both nanomaterials and peptides have provided novel clues for the development of pro-regenerative agents, however, more efforts were still be awaited and anticipated. RESULTS: In the current research, Hollow polydopamine (HPDA) nanoparticles were synthesized and HPDA nanoparticles loading with RL-QN15 (HPDAlR) that was an amphibian-derived peptide with obvious prohealing activities were prepared successfully. The characterization, biodistribution and clearance of both HPDA nanoparticles and HPDAlR were evaluated, the loading efficiency of HPDA against RL-QN15 and the slow-releasing rate of RL-QN15 from HPDAlR were also determined. Our results showed that both HPDA nanoparticles and HPDAlR exerted no obvious toxicity against keratinocyte, macrophage and mice, and HPDA nanoparticles showed no prohealing potency in vivo and in vitro. Interestingly, HPDAlR significantly enhanced the ability of RL-QN15 to accelerate the healing of scratch of keratinocytes and selectively modulate the release of healing-involved cytokines from macrophages. More importantly, in comparison with RL-QN15, by evaluating on animal models of full-thickness injured skin wounds in mice and oral ulcers in rats, HPDAlR showed significant increasing in the pro-regenerative potency of 50 and 10 times, respectively. Moreover, HPDAlR also enhanced the prohealing efficiency of peptide RL-QN15 against skin scald in mice and full-thickness injured wounds in swine. CONCLUSIONS: HPDA obviously enhanced the pro-regenerative potency of RL-QN15 in vitro and in vivo, hence HPDAlR exhibited great potential in the development of therapeutics for skin wound healing.

Customized 3D printed bioreactors for decellularization-High efficiency and quality on a budget.

Decellularization (DC) of biomaterials with bioreactors is widely used to produce scaffolds for tissue engineering. This study uses 3D printing to develop efficient but low-cost DC bioreactors. Two bioreactors were developed to decellularize pericardial patches and vascular grafts. Flow profiles and pressure distribution inside the bioreactors were optimized by steady-state computational fluid dynamics (CFD) analysis. Printing materials were evaluated by cytotoxicity assessment. Following evaluation, all parts of the bioreactors were 3D printed in a commercial fused deposition modeling printer. Samples of bovine pericardia and porcine aortae were decellularized using established protocols. An immersion and agitation setup was used as a control. With histological assessment, DNA quantification and biomechanical testing treatment effects were evaluated. CFD analysis of the pericardial bioreactor revealed even flow and pressure distribution in between all pericardia. The CFD analysis of the vessel bioreactor showed increased intraluminal flow rate and pressure compared to the vessel's outside. Cytotoxicity assessment of the used printing material revealed no adverse effect on the tissue. Complete DC was achieved for all samples using the 3D printed bioreactors while DAPI staining revealed residual cells in aortic vessels of the control group. Histological analysis showed no structural changes in the decellularized samples. Additionally, biomechanical properties exhibited no significant change compared to native samples. This study presents a novel approach to manufacturing highly efficient and low budget 3D printed bioreactors for the DC of biomaterials. When compared to standard protocols, the bioreactors offer a cost effective, fast, and reproducible approach, which vastly improves the DC results.

Alternative to Poly(2-isopropyl-2-oxazoline) with a Reduced Ability to Crystallize and Physiological LCST.

In this work, we sought to examine whether the presence of alkyl substituents randomly distributed within the main chain of a 2-isopropyl-2-oxazoline-based copolymer will decrease its ability to crystallize when compared to its homopolymer. At the same time, we aimed to ensure an appropriate hydrophilic/lipophilic balance in the copolymer and maintain the phase transition in the vicinity of the human body temperature. For this reason, copolymers of 2-ethyl-4-methyl-2-oxazoline and 2-isopropyl-2-oxazoline were synthesized. The thermoresponsive behavior of the copolymers in water, the influence of salt on the cloud point, the presence of hysteresis of the phase transition and the crystallization ability in a water solution under long-term heating conditions were studied by turbidimetry. The ability of the copolymers to crystallize in the solid state, and their thermal properties, were analyzed by differential scanning calorimetry and X-ray diffractometry. A cytotoxicity assay was used to estimate the viability of human fibroblasts in the presence of the obtained polymers. The results allowed us to demonstrate a nontoxic alternative to poly(2-isopropyl-2-oxazoline) (PiPrOx) with a physiological phase transition temperature (LCST) and a greatly reduced tendency to crystallize. The synthesis of 2-oxazoline polymers with such well-defined properties is important for future biomedical applications.