Structural insights into the distinctive RNA recognition and therapeutic potentials of RIG-I-like receptors.

RNA viruses, including the coronavirus, develop a unique strategy to evade the host immune response by interrupting the normal function of cytosolic retinoic acid-inducible gene-I (RIG-I)-like receptors (RLRs). RLRs rapidly detect atypical nucleic acids, thereby triggering the antiviral innate immune signaling cascade and subsequently activates the interferons transcription and induction of other proinflammatory cytokines and chemokines. Nonetheless, these receptors are manipulated by viral proteins to subvert the host immune system and sustain the infectivity and replication potential of the virus. RIG-I senses the single-stranded, double-stranded, and short double-stranded RNAs and recognizes the key signature, a 5'-triphosphate moiety, at the blunt end of the viral RNA. Meanwhile, the melanoma differentiation-associated gene 5 (MDA5) is triggered by longer double stranded RNAs, messenger RNAs lacking 2'-O-methylation in their 5'-cap, and RNA aggregates. Therefore, structural insights into the nucleic-acid-sensing and downstream signaling mechanisms of these receptors hold great promise for developing effective antiviral therapeutic interventions. This review highlights the critical roles played by RLRs in viral infections as well as their ligand recognition mechanisms. In addition, we highlight the crosstalk between the toll-like receptors and RLRs and provide a comprehensive overview of RLR-associated diseases as well as the therapeutic potential of RLRs for the development of antiviral-drugs. Moreover, we believe that these RLR-based antivirals will serve as a step toward countering the recent coronavirus disease 2019 pandemic.

Endogenous Stem Cell-Based In Situ Tissue Regeneration Using Electrostatically Interactive Hydrogel with a Newly Discovered Substance P Analog and VEGF-Mimicking Peptide.

The use of chemoattractants to promote endogenous stem cell-based in situ tissue regeneration has recently garnered much attention. This study is the first to assess the endogenous stem cell migration using a newly discovered substance P (SP) analog (SP1) by molecular dynamics simulations as an efficient chemoattractant. Further, a novel strategy based on electrostatic interaction using cationic chitosan (Ch) and anionic hyaluronic acid (HA) to prepare an SP1-loaded injectable C/H formulation without SP1 loss is developed. The formulation quickly forms an SP1-loaded C/H hydrogel in situ through in vivo injection. The newly discovered SP1 is found to possess human mesenchymal stromal cells (hMSCs) migration-inducing ability that is approximately two to three times higher than that of the existing SP. The designed VEGF-mimicking peptide (VP) chemically reacts with the hydrogel (C/H-VP) to sustain the release of VP, thus inducing vasculogenic differentiation of the hMSCs that migrate toward the C/H-VP hydrogel. Similarly, in animal experiments, SP1 attracts a large number of hMSCs toward the C/H-VP hydrogel, after which VP induces vasculogenic differentiation. Collectively, these findings indicate that SP1-loaded C/H-VP hydrogels are a promising strategy to facilitate endogenous stem cell-based in situ tissue regeneration.

Computational-Driven Epitope Verification and Affinity Maturation of TLR4-Targeting Antibodies.

Toll-like receptor (TLR) signaling plays a critical role in the induction and progression of autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematous, experimental autoimmune encephalitis, type 1 diabetes mellitus and neurodegenerative diseases. Deciphering antigen recognition by antibodies provides insights and defines the mechanism of action into the progression of immune responses. Multiple strategies, including phage display and hybridoma technologies, have been used to enhance the affinity of antibodies for their respective epitopes. Here, we investigate the TLR4 antibody-binding epitope by computational-driven approach. We demonstrate that three important residues, i.e., Y328, N329, and K349 of TLR4 antibody binding epitope identified upon in silico mutagenesis, affect not only the interaction and binding affinity of antibody but also influence the structural integrity of TLR4. Furthermore, we predict a novel epitope at the TLR4-MD2 interface which can be targeted and explored for therapeutic antibodies and small molecules. This technique provides an in-depth insight into antibody-antigen interactions at the resolution and will be beneficial for the development of new monoclonal antibodies. Computational techniques, if coupled with experimental methods, will shorten the duration of rational design and development of antibody therapeutics.

Exploring the Binding Mechanism of PF-07321332 SARS-CoV-2 Protease Inhibitor through Molecular Dynamics and Binding Free Energy Simulations.

The novel coronavirus disease, caused by severe acute respiratory coronavirus 2 (SARS-CoV-2), rapidly spreading around the world, poses a major threat to the global public health. Herein, we demonstrated the binding mechanism of PF-07321332, α-ketoamide, lopinavir, and ritonavir to the coronavirus 3-chymotrypsin-like-protease (3CLpro) by means of docking and molecular dynamic (MD) simulations. The analysis of MD trajectories of 3CLpro with PF-07321332, α-ketoamide, lopinavir, and ritonavir revealed that 3CLpro-PF-07321332 and 3CLpro-α-ketoamide complexes remained stable compared with 3CLpro-ritonavir and 3CLpro-lopinavir. Investigating the dynamic behavior of ligand-protein interaction, ligands PF-07321332 and α-ketoamide showed stronger bonding via making interactions with catalytic dyad residues His41-Cys145 of 3CLpro. Lopinavir and ritonavir were unable to disrupt the catalytic dyad, as illustrated by increased bond length during the MD simulation. To decipher the ligand binding mode and affinity, ligand interactions with SARS-CoV-2 proteases and binding energy were calculated. The binding energy of the bespoke antiviral PF-07321332 clinical candidate was two times higher than that of α-ketoamide and three times than that of lopinavir and ritonavir. Our study elucidated in detail the binding mechanism of the potent PF-07321332 to 3CLpro along with the low potency of lopinavir and ritonavir due to weak binding affinity demonstrated by the binding energy data. This study will be helpful for the development and optimization of more specific compounds to combat coronavirus disease.

Role of Thioredoxin-Interacting Protein in Diseases and Its Therapeutic Outlook.

Thioredoxin-interacting protein (TXNIP), widely known as thioredoxin-binding protein 2 (TBP2), is a major binding mediator in the thioredoxin (TXN) antioxidant system, which involves a reduction-oxidation (redox) signaling complex and is pivotal for the pathophysiology of some diseases. TXNIP increases reactive oxygen species production and oxidative stress and thereby contributes to apoptosis. Recent studies indicate an evolving role of TXNIP in the pathogenesis of complex diseases such as metabolic disorders, neurological disorders, and inflammatory illnesses. In addition, TXNIP has gained significant attention due to its wide range of functions in energy metabolism, insulin sensitivity, improved insulin secretion, and also in the regulation of glucose and tumor suppressor activities in various cancers. This review aims to highlight the roles of TXNIP in the field of diabetology, neurodegenerative diseases, and inflammation. TXNIP is found to be a promising novel therapeutic target in the current review, not only in the aforementioned diseases but also in prolonged microvascular and macrovascular diseases. Therefore, TXNIP inhibitors hold promise for preventing the growing incidence of complications in relevant diseases.

Biocompatibility and Therapeutic Effect of 3 Intra-Tympanic Drug Delivery Vehicles in Acute Acoustic Trauma.

INTRODUCTION: The aim of this study was to assess the biocompatibility of several intra-tympanic (IT) drug delivery vehicles and to compare hearing outcomes. MATERIALS AND METHODS: After acute acoustic trauma, rats were treated with IT 10 mg/mL dexamethasone phosphate (D) and divided into the following groups for drug delivery: saline + D (n = 15), hyaluronic acid (HA) + D (n = 17), and methoxy polyethylene glycol-b-polycaprolactone block copolymer (MP) + D (n = 24). RESULTS: No inflammation was found in the saline + D or HA + D groups. The duration of vehicle/drug persistence in the bulla was significantly longer for the MP + D (47.5 days) and HA + D groups (1.8 days) than for the saline + D group (<1 day). The tympanic membrane was significantly thicker in the MP + D group than in the saline + D and HA + D groups. The proportion of ears with good hearing outcome was significantly higher (63.6%) in the HA + D group than in the MP + D group. The number of hair cells in the hearing loss (HL) control group was significantly lower than in the MP + D group. DISCUSSION/CONCLUSION: HA shows great potential as a biocompatible vehicle for D delivery via the IT route, without an inflammatory reaction and with better hearing outcomes. Considering inflammation and hearing, MP may not be a good candidate for IT drug delivery.

Cross-linked cartilage acellular matrix film decreases postsurgical peritendinous adhesions.

Cartilage extracellular matrix contains antiadhesive and antiangiogenic molecules such as chondromodulin-1, thrombospondin-1, and endostatin. We have aimed to develop a cross-linked cartilage acellular matrix (CAM) barrier for peritendinous adhesion prevention. CAM film was fabricated using decellularized porcine cartilage tissue powder and chemical cross-linking. Biochemical analysis of the film showed retention of collagen and glycosaminoglycans after the fabrication process. Physical characterization of the film showed denser collagen microstructure, increased water contact angle, and higher tensile strength after cross-linking. The degradation time in vivo was 14 d after cross-linking. The film extract and film surface showed similar cell proliferation, while inhibiting cell migration and cell adhesion compared to standard media and culture plate, respectively. Application of the film after repair resulted in similar tendon healing and significantly less peritendinous adhesions in a rabbit Achilles tendon injury model compared to repair only group, demonstrated by histology, ultrasonography, and biomechanical testing. In conclusion, the current study developed a CAM film having biological properties of antiadhesion, together with biomechanical properties and degradation profile suitable for prevention of peritendinous adhesions.

Injectable In Situ-Forming Hydrogels for Protein and Peptide Delivery.

Injectable in situ-forming hydrogels have been used clinically in diverse biomedical applications. These hydrogels have distinct advantages such as easy management and minimal invasiveness. The hydrogels are aqueous formulations, and a simple injection at the target site replaces a traditional surgical procedure. Here, we review injectable in situ-forming hydrogels that are formulated by physical and chemical methods to deliver proteins and peptides. Prospects for using in situ-forming hydrogels for several specific applications are also discussed.

Biomimetic Scaffolds for Bone Tissue Engineering.

The use of biomimetic scaffolds for bone tissue engineering has been studied for a long time. Biomimetic scaffolds can assist and accelerate bone regeneration that is similar to that of authentic tissue, which represents the environment of cells in a living organism. Currently, numerous biomaterials have been reported for use as a biomimetic scaffold. This review focuses on the design of biomimetic scaffolds, kinds of biomaterials and methods used to fabricate biomimetic scaffolds, growth factors used with biomimetic scaffold for bone regeneration, mobilization of biological agents into biomimetic scaffolds, and studies on (pre)clinical bone regeneration from biomimetic scaffolds. Then, future prospects for biomimetic scaffolds are discussed.

Preparation of Biodegradable and Elastic Poly(ε-caprolactone-co-lactide) Copolymers and Evaluation as a Localized and Sustained Drug Delivery Carrier.

To develop a biodegradable polymer possessing elasticity and flexibility, we synthesized MPEG-b-(PCL-co-PLA) copolymers (PCxLyA), which display specific rates of flexibility and elasticity. We synthesize the PCxLyA copolymers by ring-opening polymerization of ε-caprolactone and l-lactide. PCxLyA copolymers of various compositions were synthesized with 500,000 molecular weight. The PCxLyA copolymers mechanical properties were dependent on the mole ratio of the ε-caprolactone and l-lactide components. Cyclic tensile tests were carried out to investigate the resistance to creep of PCxLyA specimens after up to 20 deformation cycles to 50% elongation. After in vivo implantation, the PCxLyA implants exhibited biocompatibility, and gradually biodegraded over an eight-week experimental period. Immunohistochemical characterization showed that the PCxLyA implants provoked in vivo inflammation, which gradually decreased over time. The copolymer was used as a drug carrier for locally implantable drugs, the hydrophobic drug dexamethasone (Dex), and the water-soluble drug dexamethasone 21-phosphate disodium salt (Dex(p)). We monitored drug-loaded PCxLyA films for in vitro and in vivo drug release over 40 days and observed real-time sustained release of near-infrared (NIR) fluorescence over an extended period from hydrophobic IR-780- and hydrophilic IR-783-loaded PCxLyA implanted in live animals. Finally, we confirmed that PCxLyA films are usable as biodegradable, elastic drug carriers.

Inappropriate continued empirical vancomycin use in a hospital with a high prevalence of methicillin-resistant Staphylococcus aureus.

Vancomycin is frequently inappropriately prescribed, especially as empirical treatment. The aim of this study was to evaluate (i) the amount of inappropriate continued empirical vancomycin use as a proportion of total vancomycin use and (ii) the risk factors associated with inappropriate continued empirical vancomycin use. We reviewed the medical records of adult patients who had been prescribed at least one dose of parenterally administered vancomycin between January and June 2012, in a single tertiary care hospital. When empirically prescribed vancomycin treatment was continued after 96 h without documentation of beta-lactam-resistant Gram-positive microorganisms in clinical specimens with significance, the continuation was considered inappropriate, and the amount used thereafter was considered inappropriately used. We identified risk factors associated with inappropriate continued empirical vancomycin use by multiple logistic regression. During the study period, the amount of parenterally administered vancomycin prescribed was 34.2 defined daily doses (DDDs)/1,000 patient-days (1,084 prescriptions for 971 patients). The amount of inappropriate continued empirical vancomycin use was 8.5 DDDs/1,000 patient-days, which represented 24.9% of the total parenterally administered vancomycin used (8.5/34.2 DDDs/1,000 patient-days). By multivariate analyses, inappropriate continued empirical vancomycin use was independently associated with the absence of any documented etiological organism (adjusted odds ratio [aOR], 1.60 [95% confidence interval {CI}, 1.06 to 2.41]) and suspected central nervous system (CNS) infections (aHR, 2.33 [95% CI, 1.20 to 4.50]). Higher Charlson's comorbidity index scores were inversely associated with inappropriate continued empirical vancomycin use (aHR, 0.90 [95% CI, 0.85 to 0.97]). Inappropriate continued empirical vancomycin use represented 24.9% of the total amount of vancomycin prescribed, which indicates room for improvement.

Development of hypoparathyroidism animal model and the feasibility of small intestinal submucosa application on the parathyroid autotransplantation.

The purpose of this study is to evaluate the feasibility of small intestinal submucosa (SIS) application on the parathyroid autotransplantation in a rat model of hypoparathyroidism. The rats were divided into four groups: NC (no procedure, n = 5), PTX (total parathyroidectomy, n = 6), PT (total parathyroidectomy and parathyroid autotransplantation, n = 10) and PT + SIS group (total parathyroidectomy and parathyroid autotransplantation with SIS, n = 10). The levels of parathyroid hormone (PTH), calcium, and phosphorous were measured on 0, 3, 7, 21, 56 and 84 days after surgery. PTH level was expressed as median (interquartile range) and histological and immunohistochemical examinations were performed. PTH levels were significantly decreased to "not detectable level" from day 3 in PTX group. PTH was not detected in both PT and PT + SIS groups on the 21st day. On the 56th day, PTH levels were increased in both groups: 3 out of 8 rats (37.5%) in the PT group, 6 out of 9 rats (66.7%) in the PT + SIS group. The PTH level was fully recovered to its preoperative range on the day 84 as 6 of 8 rats (75%) of the PT group and 7 of 9 rats (77.8%) of the PT + SIS group were recovered; the PTH levels were 117.84 and 178.36 pg/ml, respectively. The neo-vascularization was well observed around the parathyroid tissue, and the number of new vessels formed was higher in the PT + SIS group (15 vessels/high power field) as compared to the PT group (10 vessels/high power field). This study showed the feasibility and the treatment effect of SIS as the success rate of autotransplantation of parathyroid tissue was significantly increased without severe inflammatory response in hypothyroidism animal model.

Cellular behavior of human adipose-derived stem cells on wettable gradient polyethylene surfaces.

Appropriate surface wettability and roughness of biomaterials is an important factor in cell attachment and proliferation. In this study, we investigated the correlation between surface wettability and roughness, and biological response in human adipose-derived stem cells (hADSCs). We prepared wettable and rough gradient polyethylene (PE) surfaces by increasing the power of a radio frequency corona discharge apparatus with knife-type electrodes over a moving sample bed. The PE changed gradually from hydrophobic and smooth surfaces to hydrophilic (water contact angle, 90° to ~ 50°) and rough (80 to ~120 nm) surfaces as the power increased. We found that hADSCs adhered better to highly hydrophilic and rough surfaces and showed broadly stretched morphology compared with that on hydrophobic and smooth surfaces. The proliferation of hADSCs on hydrophilic and rough surfaces was also higher than that on hydrophobic and smooth surfaces. Furthermore, integrin beta 1 gene expression, an indicator of attachment, and heat shock protein 70 gene expression were high on hydrophobic and smooth surfaces. These results indicate that the cellular behavior of hADSCs on gradient surface depends on surface properties, wettability and roughness.

Intra-articular hydrogel formulation prolongs the in vivo stability of Toll-like receptor antagonistic peptides for rheumatoid arthritis treatment.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by systemic inflammation that primarily affects joint tissue and requires frequent medication. Recently, we developed cyclic phage-display-derived inhibitory peptides (CPs), which act as Toll-like Receptor 4 antagonists. These CPs exhibited therapeutic efficacy against joint diseases by alleviating inflammatory factors. Nonetheless, CP exhibits in vivo instability and a short half-life. Therefore, this study sought to improve the in vivo stability of CP, thereby reducing the frequency of CP administration through the development of an injectable hydrogel depot formulation. To improve in vivo stability, CP was chemically conjugated to hyaluronic acid (HA-CP) and subsequently mixed into a temperature-sensitive hydrogel [methoxy polyethylene glycol-b-poly(ε-caprolactone)-ran-poly(lactide) (PC)] as an injectable depot (PC+(HA-CP)). For comparison, CP was physically mixed with HA and PC (PC+(HA+CP)). Both PC+(HA-CP) and PC+(HA+CP) were found to rapidly form depots upon injection into the joint space. Cell viability assays confirmed the non-toxic nature of PC+(HA-CP) and PC+(HA+CP), whereas both formulations exhibited inhibition of inflammatory factors. Furthermore, PC+(HA-CP) retained CP for a longer duration compared to PC+(HA+CP) in the presence of hyaluronidase and within the RA joint space. Following intra-articular injection, both the PC+(HA-CP) and PC+(HA+CP) depots exhibited reductions in RA symptoms, cartilage regeneration, and suppression of pro-inflammatory cytokine levels. Specifically, by extending the in vivo retention of CP, PC+(HA-CP) demonstrated superior RA treatment efficacy compared to PC+(HA+CP). In conclusion, intra-articular injection of PC+(HA-CP) was validated as an effective strategy for treating RA, owing to its ability to prolong the in vivo retention of CP. This approach holds promise for improving RA management and patient outcomes.

3D digital light process bioprinting: Cutting-edge platforms for resolution of organ fabrication.

Research in the field of regenerative medicine, which replaces or restores the function of human damaged organs is advancing rapidly. These advances are fostering important innovations in the development of artificial organs. In recent years, three-dimensional (3D) bioprinting has emerged as a promising technology for regenerative medicine applications. Among various techniques, digital light process (DLP) 3D bioprinting stands out for its ability to precisely create high-resolution, structurally complex artificial organs. This review explores the types and usage trends of DLP printing equipment, bioinks, and photoinitiators. Building on this foundation, the applications of DLP bioprinting for creating precise microstructures of human organs and for regenerating tissue and organ models in regenerative medicine are examined. Finally, challenges and future perspectives regarding DLP-based bioprinting, particularly for precision printing applications in regenerative medicine, are discussed.

Recombinant human fibroblast growth factor 7 obtained from stable Chinese hamster ovary cells enhances wound healing.

Although fibroblast growth factor 7 (FGF7) is known to promote wound healing, its mass production poses several challenges and very few studies have assessed the feasibility of producing FGF7 in cell lines such as Chinese hamster ovary (CHO) cells. Therefore, this study sought to produce recombinant FGF7 in large quantities and evaluate its wound healing effect. To this end, the FGF7 gene was transfected into CHO cells and FGF7 production was optimized. The wound healing efficacy of N-glycosylated FGF7 was evaluated in animals on days 7 and 14 post-treatment using collagen patches (CPs), FGF7-only, and CP with FGF7 (CP+FGF7), whereas an untreated group was used as the control. Wound healing was most effective in the CP+FGF7 group. Particularly, on day 7 post-exposure, the CP+FGF7 and FGF7-only groups exhibited the highest expression of hydroxyproline, fibroblast growth factor, vascular endothelial growth factor, and transforming growth factor. Epidermalization in H&E staining showed the same order of healing as hydroxyproline content. Additionally, the CP+FGF7 and FGF7-only group exhibited more notable blood vessel formation on days 7 and 14. In conclusion, the prepared FGF7 was effective in promoting wound healing and CHO cells can be a reliable platform for the mass production of FGF7.

Amplifying endogenous stem cell migration for in situ bone tissue formation: Substance P analog and BMP mimetic peptide-loaded click-crosslinked hyaluronic acid hydrogel.

Endogenous stem cell-driven in situ bone tissue formation has recently garnered increasing attention. Therefore, our study sought to refine methods to enhance the migration and subsequent osteogenic differentiation of these cells. Our innovative approach involves using an injectable hydrogel that combines click cross-linking sites and a BMP-2 mimetic peptide (BP) with hyaluronic acid (HA). This injectable formulation, hereinafter referred to as SPa + Cx-HA-BP, incorporates a substance P analog peptide (SPa) with Cx-HA-BP, proving versatile for in vitro and in vivo applications without cytotoxicity. The controlled release of SPa creates a gradient that guides endogenous stem cells towards the Cx-HA scaffold from specific tissue niches. Both Cx-HA and SPa+Cx-HA induced minimal changes in the expression of genes associated with osteogenic differentiation. In contrast, these genes were robustly induced by both SPa + Cx-HA+BP and SPa + Cx-HA-BP, in which BP was respectively integrated via physical and chemical methods. Remarkably, chemically incorporating BP (Cx-HA-BP) resulted in 4-9 times higher osteogenic gene expression than physically mixed BP in Cx-HA+BP. This study validates the role of SPa role in guiding endogenous stem cells toward the hydrogel and underscores the substantial impact of sustained BP presence within the hydrogel. Collectively, our findings offer valuable insights for the development of innovative strategies to promote endogenous stem cell-based tissue regeneration. The developed hydrogel effectively guides stem cells from their natural locations and facilitates sustained osteogenic differentiation, thus holding great promise for applications in regenerative medicine.

Induction of neuronal differentiation of rat muscle-derived stem cells in vitro using basic fibroblast growth factor and ethosuximide.

Several studies have demonstrated that basic fibroblast growth factor (bFGF) can induce neural differentiation of mesenchymal stem cells. In this study, we investigated the neural differentiation of muscle-derived stem cells (MDSCs) following treatment with bFGF and ethosuximide, a small molecule used as an anticonvulsant in humans. Stem cells isolated from rat skeletal muscle (rMDSCs) were pre-induced by culturing with 25 ng/mL bFGF for 24 h and then were transferred to a medium supplemented with or without 4 mM ethosuximide. Neuronal differentiation was assessed by immunocytochemical and western blotting analyses of marker expression. Immunocytochemistry of rMDSCs treated with bFGF and ethosuximide identified abundant cells expressing neuronal markers (TuJ1, neuron-specific class III ß-tubulin; NeuN, neuronal nuclear antigen; and NF-MH; neurofilament M and H). Olig2 (oligodendrocyte transcription factor 2)-positive cells were also observed, indicating the presence of oligodendrocyte lineage cells. These findings were substantiated by western blotting analysis of marker proteins. In particular, the expression of NeuN and TuJ1 was significantly higher in rMDSCs treated with ethosuximide and bFGF than in cells stimulated with bFGF alone (NeuN, p < 0.05 and TuJ1, p < 0.001). Expression of the astrocyte marker GFAP (glial fibrillary acidic protein) was not detected in this study. Collectively, the results showed that treatment with bFGF and ethosuximide induced effective transdifferentiation of rMDSCs into cells with a neural-like phenotype. Notably, rMDSCs treated with a combination of bFGF plus ethosuximide showed enhanced differentiation compared with cells treated with bFGF alone, implying that ethosuximide may stimulate neuronal differentiation.

Preparation and evaluation of injectable microsphere formulation for longer sustained release of donepezil.

In this study, donepezil-loaded PLGA and PLA microspheres (Dp-PLGA-M/Dp-PLA-M) and Dp-PLA-M wrapped in a polyethylene glycol-b-polycaprolactone (PC) hydrogel (Dp-PLA-M/PC) were prepared to reduce the dosing frequency of injections to treat Alzheimer's disease patients. Dp-PLGA-M and Dp-PLA-M with a uniform particle size distribution were repeatably fabricated in nearly quantitative yield and with high encapsulated Dp yields using an ultrasonic atomizer. The injectability and in vitro and in vivo Dp release, biodegradation, and inflammatory response elicited by the Dp-PLGA-M, Dp-PLA-M, and Dp-PLA-M/PC formulations were then compared. All injectable formulations showed good injectability with ease of injection, even flow, and no clogging using a syringe needle under 21-G. The injections required a force of <1 N. According to the biodegradation rate of micro-CT, GPC and NMR analyses, the biodegradation of Dp-PLA-M was slower than that of Dp-PLGA-M, and the biodegradation rate of Dp-PLA-M/PC was also slower. In the Dp release experiment, Dp-PLA-M sustained Dp for longer compared with Dp-PLGA-M. Dp-PLA-M/PC exhibited a longer sustained release pattern of two months. In vivo bioavailability of Dp-PLA-M/PC was almost 1.4 times higher than that of Dp-PLA-M and 1.9 times higher than that of Dp-PLGA-M. The variations in the Dp release patterns of Dp-PLGA-M and Dp-PLA-M were explained by differences in the degradation rates of PLGA and PLA. The sustained release of Dp by Dp-PLA-M/PC was attributed to the fact that the PC hydrogel served as a wrapping matrix for Dp-PLA-M, which could slow down the biodegradation of PLA-M, thus delaying the release of Dp from Dp-PLA-M. Dp-PLGA-M induced a higher inflammatory response compared to Dp-PLA-M/PC, suggesting that the rapid degradation of PLGA triggered a strong inflammatory response. In conclusion, Dp-PLA-M/PC is a promising injectable Dp formulation that could be used to reduce the dosing frequency of Dp injections.

Development and In Vivo Assessment of an Injectable Cross-Linked Cartilage Acellular Matrix-PEG Hydrogel Scaffold Derived from Porcine Cartilage for Tissue Engineering.

The cartilage acellular matrix (CAM) derived from porcine cartilage, which does not induce significant inflammation and provides an environment conducive for cell growth and differentiation, is a promising biomaterial candidate for scaffold fabrication. However, the CAM has a short period in vivo, and the in vivo maintenance is not controlled. Therefore, this study is aimed at developing an injectable hydrogel scaffold using a CAM. The CAM is cross-linked with a biocompatible polyethylene glycol (PEG) cross-linker to replace typically used glutaraldehyde (GA) cross-linker. The cross-linking degree of cross-linked CAM by PEG cross-linker (Cx-CAM-PEG) according to the ratios of the CAM and PEG cross-linker is confirmed by contact angle and heat capacities measured by differential scanning calorimetry. The injectable Cx-CAM-PEG suspension exhibits controllable rheological properties and injectability. Additionally, injectable Cx-CAM-PEG suspensions with no free aldehyde group are formed in the in vivo hydrogel scaffold almost simultaneously with injection. In vivo maintenance of Cx-CAM-PEG is realized by the cross-linking ratio. The in vivo formed Cx-CAM-PEG hydrogel scaffold exhibits certain host-cell infiltration and negligible inflammation within and near the transplanted Cx-CAM-PEG hydrogel scaffold. These results suggest that injectable Cx-CAM-PEG suspensions, which are safe and biocompatible in vivo, represent potential candidates for (pre-)clinical scaffolds.