Intra-articular injection of modified citrus pectin and hyaluronate gel induces synergistic effects in treating osteoarthritis.

We previously found that modified citrus pectin (MCP), an inhibitor of pro-inflammatory factor Galectin-3 (Gal-3), has significant anti-inflammatory and chondroprotective effects. In this study, a hyaluronate (HA) gel-based sustained release system of MCP (MCP-HA) was developed as an anti-inflammatory agent for chronic inflammation for osteoarthritis (OA) treatment. The MCP-HA gel was injected into the knee joint cavities of OA rabbit models induced by anterior cruciate ligament transection (ACLT) or modified Hulth method once a week for five weeks. We found that MCP-HA could improve the symptoms and signs of OA, protect articular cartilage from degeneration, suppress synovial inflammation, and therefore alleviate OA progression. Proteomic analysis of the synovial fluid obtained from the knee joints of OA rabbits revealed that MCP-HA synergistically regulated the levels of multiple inflammatory mediators and proteins involved in metabolic pathways. Taken together, our results demonstrate that the MCP-HA shows a synergistic effect of HA and MCP by modulating both inflammation and metabolic processes, thereby alleviating OA progression. The MCP-HA sustained release system has promising potential for long-term use in OA treatment.

The effects of stiffness on the specificity and avidity of antibody-coated microcapsules with target cells are strongly shape dependent.

Antibody modification is a common method for endowing drug carriers with the ability to target specific cells. Recent studies suggest that the efficacy of these antibody-modified drug carriers is closely related to their physicochemical properties, such as size, shape, stiffness, charge, and surface chemistry. In this study, we functionalized microcapsules with antibodies to investigate the combined effect of shape and stiffness on their targeting ability. We synthesized hollow microcapsules, both spherical and rod-shaped, with adjustable stiffness using calcium carbonate particles as templates and silk fibroin (SF) as the shell material. These microcapsules were then functionalized with trastuzumab (TTZ) to enhance targeting capabilities. Our analysis revealed that increasing stiffness significantly improved the specificity and avidity of TTZ-coated rod-shaped microcapsules, but not spherical ones, indicating a strong shape-dependent influence of stiffness on these properties. Additionally, we explored the mechanisms of endocytosis using various inhibitors and found that both macropinocytosis and clathrin played critical roles in the cellular uptake of microcapsules. Furthermore, we loaded microcapsules with doxorubicin (DOX) to evaluate their anti-tumor efficacy. The stiffest TTZ-coated, DOX-loaded rod-shaped microcapsules demonstrated the most potent anti-tumor effects on BT-474 cells and the highest uptake in BT-474 3D spheroids. This research contributes to the development of more effective microcapsule-based target delivery systems and the realization of the full potential of microcapsule drug delivery systems.

Gelatin/Hyaluronic Acid Photocrosslinked Double Network Hydrogel with Nano-Hydroxyapatite Composite for Potential Application in Bone Repair.

The application of hydrogels in bone repair is limited due to their low mechanical strength. Simulating bone extracellular matrix, methylacrylylated gelatin (GelMA)/methylacrylylated hyaluronic acid (HAMA)/nano-hydroxyapatite(nHap) composite hydrogels were prepared by combining the double network strategy and composite of nHap in this study. The precursor solutions of the composite hydrogels were injectable due to their shear thinning property. The compressive elastic modulus of the composite hydrogel was significantly enhanced, the fracture strength of the composite hydrogel nearly reached 1 MPa, and the composite hydrogel retained its high water content at above 88%. The composite hydrogels possess good compatibility with BMSCS and have the potential to be used as injectable hydrogels for bone defect treatment.

The effect of blending poly (l-lactic acid) on in vivo performance of 3D-printed poly(l-lactide-co-caprolactone)/PLLA scaffolds.

Blending poly (l-lactic acid, PLLA) with poly (l-lactide-co-caprolactone, PLCL) is an effective strategy for developing new PLCL/PLLA blend based biomaterials. However, the effect of PLLA on in vivo performance of PLCL/PLLA blends is unclear yet. To address this issue, in this study, the effect of PLLA on in vivo biodegradability and biocompatibility of 3D-printed scaffolds of PLCL/PLLA blend was investigated. Three kinds of different 3D-printed PLCL/PLLA scaffolds using different blends with different mass ratios of the polymers, were prepared and implanted subcutaneously. The shrinkage and tissue responses were monitored by ultrasonography after the implantation. 2 months post-operation, the in vivo performances of the scaffolds were investigated histologically. All scaffolds showed good biocompatibility and allowed fast tissues ingrowth, however PLCL50/PLLA50 scaffold with the highest PLLA ratio induced the thickest the fibrous capsule surrounding the scaffolds and highest inflammatory scores. Furthermore, it was found that the fine porous structures of all scaffolds were well maintained, indicating the 3D-printed scaffolds were degraded through a surface erosion but not bulk erosion way. However, different scaffolds showed different shrinkage and degradation ratios, and PLCL50/PLLA50 scaffold resulted in a significant shrinkage, while PLCL90/PLLA10 scaffold showed the better structural stability. Therefore, PLLA at blending different ratio had different effects on the in vivo performance of 3D-printed PLCL/PLLA scaffolds. Particularly, PLCL/PLLA scaffolds blending with low ratio of PLLA, such as PLCL90/PLLA10 scaffold showed better application potential in tissue engineering. Our findings provide a new insight on the rational design, constrcution and application of the 3D-printed PLCL/PLLA scaffolds.

Locally delivered modified citrus pectin - a galectin-3 inhibitor shows expected anti-inflammatory and unexpected regeneration-promoting effects on repair of articular cartilage defect.

Treating the concomitant inflammation in the process of injury and repair, and simultaneously promoting cartilage regeneration is very important for the repair of articular cartilage (AC) defects. Nevertheless, this remains a massive challenge. To address this issue, a collagen membrane-based modified citrus pectin (MCP) delivery system (MCP-C) was developed in this study by targeting galectin-3 (Gal-3), an upstream proinflammatory factor. As expected, MCP shows anti-inflammatory effects; it downregulates the expressions of IL-1ß, MMP13, Gal-3, and COL1A2, inhibits the degenerative effects of Gal-3 on chondrocytes in vitro, and protects chondrocytes from degeneration and death in vivo. Unexpectedly, MCP promotes the proliferation of chondrocytes, upregulates the expression of COL2A1 and SOX9 in the chondrocytes in vitro, and enhances the repair of AC defect in rabbit knee, especially MCP500-C with a complete release of the loading amount of approximately 500 µg/cm2 in a day. Mechanistically, MCP upregulates the expressions of multiple endogenous growth factors for chondrogenesis via the transcriptome sequencing of MCP-treated chondrocytes, and downregulates the expressions of various inflammatory factors. These findings demonstrate that locally delivered MCP can simultaneously modulate both regenerative and inflammatory responses, and can enhance the repair of AC defects.

Blending with Poly(l-lactic acid) Improves the Printability of Poly(l-lactide-co-caprolactone) and Enhances the Potential Application in Cartilage Tissue Engineering.

Poly(l-lactide-co-caprolactone) (PLCL, 50:50) has been used in cartilage tissue engineering because of its high elasticity. However, its mechanical properties, including its rigidity and viscoelasticity, must be improved for compatibility with native cartilage. In this study, a set of PLCL/poly(l-lactic acid) (PLLA) blends was prepared by blending with different mass ratios of PLLA that range from 10 to 50%, using thermoplastic techniques. After testing the properties of these PLCL/PLLA blends, they were used to fabricate scaffolds by the 3D printing technology. The structures and viscoelastic behavior of the PLCL/PLLA scaffolds were determined, and then, the potential application of the scaffolds in cartilage tissue engineering was evaluated by chondrocytes culture. All blends demonstrate good thermal stability for the 3D printing technology. All blends show good toughness, while the rigidity of PLCL is increased through PLLA blending, and Young's modulus of blends with 10-20% PLLA is similar to that of native cartilage. Furthermore, blending with PLLA improves the processability of PLCL for 3D printing, and the compression modulus and viscoelasticity of 3D-printed PLCL/PLLA scaffolds are different from that of PLCL. Additionally, the stress relaxation time (t 1/2) of the PLCL/PLLA scaffolds, which is important for chondrogenesis, is dramatically shortened compared with the pure PLCL scaffold at the same 3D-printing filling rate. Consistently, the PLCL90PLLA10 scaffold at a 70% filling rate with much shorter t 1/2 is more conducive to the proliferation and chondrogenesis of in vitro seeded chondrocytes accompanied by upregulated expression of SOX9 than the PLCL scaffold. Taken together, these results demonstrate that blending with PLLA improves the printability of PLCL and enhances its potential application, particularly PLCL/PLLA scaffolds with a low ratio of PLLA, in cartilage tissue engineering.

Chitosan-Stabilized Self-Assembled Fluorescent Gold Nanoclusters for Cell Imaging and Biodistribution in Vivo.

Biocompatible, near-infrared luminescent gold nanoclusters were synthesized in situ using as-prepared chitosan grafted with N-acetyl-l-cysteine (NAC-CS). The fluorescent gold nanoclusters coated with chitosan-N-acetyl-l-cysteine (AuNCs@NAC-CS) were aggregated by multiple ultrasmall gold nanoclusters closing with each other, with strong fluorescence emission at 680 nm upon excitation at 360 nm. AuNCs@NAC-CS did not display any appreciable cytotoxicity on cells even at a concentration of 1.0 mg mL-1. AuNCs@NAC-CS were more insensitive to H2O2 and trypsin compared with fluorescent gold nanoclusters coated with Albumin Bovine V (AuNCs@BSA), which make them have long time imaging in HeLa cells. Furthermore, the obvious fluorescence signal of AuNCs@NAC-CS appeared in the liver and kidney of the normal mice after 6 h injection. And the fluorescence intensity decreased after that because of the highly efficient clearance characteristics of ultrasmall nanoparticles. These findings demonstrated that AuNCs@NAC-CS possessed good fluorescence, low cytotoxicity, and low sensitivity to some content of cells, allowing imaging of the living cells.

[The gene expression patterns of bone-marrow mesenchymal stem cells under different osteogenic induction].

OBJECTIVE: To investigate the molecular mechanism of osteogenetic differentiation of bone-marrow mesenchymal stem cells (BMSCs) and the probability using the BMSCs to gene therapy for bone fractures. METHODS: By gradient centrifugation and adherence to the culture plastic, the MSCs were separated and purified from mouse bone marrow. The BMSCs then were cultured and sub-cultured in the osteogenetic medium (100 nmol/L Dexamethasone, 10 mmol/L beta-glycerophosphate and 50 mg/mL ascorbic acid, osteogenic supplements, OS-medium) or the recombinant human bone morphogenetic protein-2 (rhBMP-2, 500 ng/mL) for the mineralized inductions of osteogenesis, and stained by alizarin red in inducing week 1 and 2 for the identification of calcium nodule formed. The gene expressions of Runx2, Osx, OCN, and Col I were detected by RT-PCR on day 1, 2 and 3 after doing the osteogenetic inductions. RESULTS: The BMSCs induced by OS-medium and rhBMP-2 were both of positive Ca nodules with alizarin red. However, the Ca nodule induced by OS-medium formed in 1 inducing week, but the one done by rhBMP-2 occurred in 2 inducing weeks, which meant it was a late for one week. In the OS-group, the mRNA of Runx2 could not be detected on inducing day 1, 2 and 3, but the Osx mRNA appeared on inducing day 2 and 3, and also the mRNAs of OCN and Col I could be detected in all the three inducing days. In rhBMP-2 group, the Runx2 gene expressed on inducing day 2, the Osx gene expressed on inducing day 2 and 3, the OCN and Col I genes expressed on inducing day 1, 2 and 3. CONCLUSION: The BMSCs induced by OS-medium are more likely to form bone nodules than that of rhBMP-2, because of their simpler mechanisms to differentiate into osteoblasts.

Chitosan-coated gold nanorods for cancer therapy combining chemical and photothermal effects.

To develop a novel type of nanoparticle for cancer therapy, gold nanorods (GNRs) are coated with chitosan (CS) derivatives to combine chemical and photothermal effects. Thiol-modified chitosan derivatives chemically conjugated to doxorubicin (DOX) are successfully synthesized and their in vitro effect is evaluated. Functional nanocarriers (DOX-CS-GNR) with good biocompatibility and optical properties are prepared by conjugating chitosan derivatives to GNRs. Two types of structures with different molar ratios of chitosan derivatives and GNRs are successfully obtained. In in vitro studies, GNR-loaded nanoparticles show low cytotoxicity and high potential for anti-cancer effects. Under conditions of short exposure time and low light intensity, DOX-CS-GNR nanocarriers with a side-by-side structure exhibit cytoxicity against tumor cells based on a combination of chemical and photothermal therapeutic effects.