Bone Destruction-Chemotactic Osteoprogenitor Cells Deliver Liposome Nanomedicines for the Treatment of Osteosarcoma and Osteoporosis.

Therapeutic efficacy of skeletal diseases is usually limited by unfavorable drug delivery due to incapable bone targeting and low bone affinity of conventional drug carriers, as well as relatively reduced vascularization and dense structure of bone tissues. Due to CXC chemokine receptor 4 (CXCR4)/CXC chemokine ligand 12 (CXCL12) signal axis-guided recruitment, osteoprogenitor cells (OPCs) can actively migrate to bone disease nidus. Here, drugs-loaded nanoliposomes are prepared and decorated onto OPCs by biotin-streptavidin linkage for precise bone disease targeting and effective drug delivery. In mouse models of tibia defect and orthotopic osteosarcoma, superior targeting property of OPCs-based drug delivery systems toward diseased bone niduses is verified. By encapsulating antitumor and antiosteoporosis drugs into nanoliposomes, OPCs-based drug delivery systems effectively inhibit disease development and restore bone destruction in mouse models of orthotopic osteosarcoma and ovariectomized osteoporosis. This study reveals a cell-based drug delivery system for precise bone disease targeting and highly effective drug delivery, which will find great potential as a universal drug delivery platform for targeting treatment of various bone diseases.

A multifunctional self-reinforced injectable hydrogel for enhancing repair of infected bone defects by simultaneously targeting macrophages, bacteria, and bone marrow stromal cells.

Injectable hydrogels (IHs) have demonstrated huge potential in promoting repair of infected bone defects (IBDs), but how to endow them with desired anti-bacterial, immunoregulatory, and osteo-inductive properties as well as avoid mechanical failure during their manipulation are challenging. In this regard, we developed a multifunctional AOHA-RA/Lap nanocomposite IH for IBDs repair, which was constructed mainly through two kinds of reversible cross-links: (i) the laponite (Lap) crystals mediated electrostatic interactions; (ii) the phenylboronic acid easter bonds between the 4-aminobenzeneboronic acid grafted oxidized hyaluronic acid (AOHA) and rosmarinic acid (RA). Due to the specific structural composition, the AOHA-RA/Lap IH demonstrated superior injectability, self-recoverability, spatial adaptation, and self-reinforced mechanical properties after being injected to the bone defect site. In addition, the RA molecules could be locally released from the hydrogel following a Weibull model for over 10 days. Systematic in vitro/vivo assays proved the strong anti-bacterial activity of the hydrogel against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli). Moreover, its capability of inducing M2 polarization of macrophages (Mφ) and osteogenic differentiation of bone marrow stromal cells (BMSCs) was verified either, and the mechanism of the former was identified to be related to the JAK1-STAT1 and PI3K-AKT signaling pathways and that of the latter was identified to be related to the calcium signaling pathway, extracellular matrix (ECM) receptor interaction and TGF-ß signaling pathway. After being implanted to a S. aureus infected rat skull defect model, the AOHA-RA/Lap IH significantly accelerated repair of IBDs without causing significant systemic toxicity. STATEMENT OF SIGNIFICANCE: Rosmarinic acid and laponite were utilized to develop an injectable hydrogel, promising for accelerating repair of infected bone defects in clinic. The gelation of the hydrogel was completely driven by two kinds of reversible cross-links, which endow the hydrogel superior spatial adaption, self-recoverability, and structural stability. The as-prepared hydrogel demonstrated superior anti-bacterial/anti-biofilm activity and could induce M2 polarization of macrophages and osteogenic differentiation of BMSCs. The mechanism behind macrophages polarization was identified to be related to the JAK1-STAT1 and PI3K-AKT signaling pathways. The mechanism behind osteogenic differentiation of BMSCs was identified to be related to the ECM receptor interaction and calcium signaling/TGF-ß signaling pathways.

Drug-Free "Triboelectric Immunotherapy" Activating Immunity for Osteomyelitis Treatment and Recurrence Prevention.

Treatment of osteomyelitis is clinically challenging with low therapeutic efficacy and high risk of recurrence owing to the immunosuppressive microenvironment. Existing therapies are limited by drug concentration and single regulatory effect on the immune network, and emphasize the role of anti-inflammatory effects in reducing osteoclast rather than the role of proinflammatory effects in accelerating infection clearance, which is not conducive to complete bacteria elimination and recurrence prevention. Herein, a direct-current triboelectric nanogenerator (DC-TENG) is established to perform antibacterial effects and modulate immunological properties of infectious microenvironments of osteomyelitis through electrical stimulation, namely triboelectric immunotherapy. Seeing from the results, the triboelectric immunotherapy successfully activates polarization to proinflammatory (M1) macrophages in vitro, accompanied by satisfying direct antibacterial effects. The antibacterial and osteogenic abilities of triboelectric immunotherapy are verified in rat cranial osteomyelitis models. The effects on the polarization and differentiation of immune-related cells in vivo are investigated by establishing in situ tibial osteomyelitis models and immunosurveillance models in C57 mice respectively, indicating the ability of activating immunity and producing immunological memory for in situ infection and secondary recurrence, thus accelerating healing and preventing relapse. This study provides an efficient, long-acting, multifunctional, and wearable triboelectric immunotherapy strategy for drug-free osteomyelitis treatment systems.

Estrogen Deficiency Exacerbates Traumatic Heterotopic Ossification in Mice.

Background: Traumatic heterotopic ossification (HO) is a devastating sequela of orthopedic surgeries and traumatic injuries; however, few studies have explored the effects of the estrogen-deficient state on HO formation. In the present study, we investigated the impact of estrogen deficiency on ectopic cartilage and bone formation in tendon after Achilles tenotomy in an ovariectomized mouse model. Methods: A total of 45 female C57BL/6 mice were randomly divided into three groups: sham-operated (control), estrogen depletion by ovariectomy (OVX) and OVX with 17ß-estradiol supplementation (OVX + E2), with 15 animals in each group. Three weeks after OVX, all mice were subjected to an Achilles tenotomy using a posterior midpoint approach to induce HO. At 1, 3 and 9 weeks after tenotomy, the left hind limbs were harvested for histology, immunohistochemistry and immunofluorescence evaluations. The volume of ectopic bone was assessed by micro-CT. Results: Mice in the OVX group formed more ectopic cartilage 3 weeks after tenotomy, as well as ectopic bone 9 weeks after tenotomy, compared to the control group. Estrogen deficiency resulted in more severe inflammatory infiltration at the injury sites 1 week after tenotomy, involving the recruitment of more macrophages and mast cells, as well as increasing the expressions of pro-inflammatory mediators, including IL-1ß, IL-6, and TNF-α. Moreover, the local TGF-ß/SMAD signaling pathway was dysregulated after OVX, which manifested as upregulated expressions of TGF-ß and pSMAD2/3. E2 supplementation protected against OVX-induced HO deterioration, inhibited inflammatory infiltration, and downregulated the TGF-ß/SMAD signaling pathway. Conclusion: Estrogen deficiency exacerbated HO formation in the Achilles tenotomy model. These findings might be attributable to the disturbance of the inflammatory response and the activation of TGF-ß/SMAD signaling at the injury sites during the early stages of HO development.

In Situ Sprayed Exosome-Cross-Linked Gel as Artificial Lymph Nodes for Postoperative Glioblastoma Immunotherapy.

One key challenge in postoperative glioblastoma immunotherapy is to guarantee a potent and durable T-cell response, which is restricted by the immunosuppressive microenvironment within the lymph nodes (LNs). Here, we develop an in situ sprayed exosome-cross-linked gel that acts as an artificial LN structure to directly activate the tumor-infiltrating T cells for prevention of glioma recurrence. Briefly, this gel is generated by a bio-orthogonal reaction between azide-modified chimeric exosomes and alkyne-modified alginate polymers. Particularly, these chimeric exosomes are generated from dendritic cell (DC)-tumor hybrid cells, allowing for direct and robust T-cell activation. The gel structure with chimeric exosomes as cross-linking points avoids the quick clearance by the immune system and thus prolongs the durability of antitumor T-cell immunity. Importantly, this exosome-containing immunotherapeutic gel provides chances for ameliorating functions of antigen-presenting cells (APCs) through accommodating different intracellular-acting adjuvants, such as stimulator of interferon genes (STING) agonists. This further enhances the antitumor T-cell response, resulting in the almost complete elimination of residual lesions after surgery. Our findings provide a promising strategy for postsurgical glioma immunotherapy that warrants further exploration in the clinical arena.

A double-network porous hydrogel based on high internal phase emulsions as a vehicle for potassium sucrose octasulfate delivery accelerates diabetic wound healing.

Diabetic wounds are a difficult medical challenge. Excessive secretion of matrix metalloproteinase-9 (MMP-9) in diabetic wounds further degrades the extracellular matrix and growth factors and causes severe vascular damage, which seriously hinders diabetic wound healing. To solve these issues, a double-network porous hydrogel composed of poly (methyl methacrylate-co-acrylamide) (p(MMA-co-AM)) and polyvinyl alcohol (PVA) was constructed by the high internal phase emulsion (HIPE) technique for the delivery of potassium sucrose octasulfate (PSO), a drug that can inhibit MMPs, increase angiogenesis and improve microcirculation. The hydrogel possessed a typical polyHIPE hierarchical microstructure with interconnected porous morphologies, high porosity, high specific surface area, excellent mechanical properties and suitable swelling properties. Meanwhile, the p(MMA-co-AM)/PVA@PSO hydrogel showed high drug-loading performance and effective PSO release. In addition, both in vitro and in vivo studies showed that the p(MMA-co-AM)/PVA@PSO hydrogel had good biocompatibility and significantly accelerated diabetic wound healing by inhibiting excessive MMP-9 in diabetic wounds, increasing growth factor secretion, improving vascularization, increasing collagen deposition and promoting re-epithelialization. Therefore, this study provided a reliable therapeutic strategy for diabetic wound healing, some theoretical basis and new insights for the rational design and preparation of wound hydrogel dressings with high porosity, high drug-loading performance and excellent mechanical properties.

Bioinspired wet adhesive carboxymethyl cellulose-based hydrogel with rapid shape adaptability and antioxidant activity for diabetic wound repair.

Currently, adhesive hydrogels have shown promising effect in chronic diabetic wound repair. However, there are issues and challenges in treating diabetic wounds due to inadequate wet adhesion, unable to fill irregular and deep wounds, and oxidative stress. Herein, a mussel-inspired naturally hydrogel dressing with rapid shape adaptability, wet adhesion and antioxidant abilities for irregular, deep and frequently movement diabetic wounds repair was constructed by comprising catechol modified carboxymethyl cellulose (CMC-DA) and tannic acid. Benefiting from the reversible hydrogen bonding, the resulting hydrogels exhibited injectability, remarkable self-healing ability, rapid shape adaptability and strong tissue adhesion (45.9 kPa), thereby contributing to self-adaptive irregular-shaped wounds or moving joint parts. Especially, the adhesion strength of the hydrogel on wet tissue still remained at 14.9 kPa. Besides, the hydrogels could be easily detached from the skin by ice-cooling that avoided secondary damage caused by dressing change. Remarkably, the hydrogels possessed excellent antioxidant, satisfactory biocompatibility, efficient hemostasis and antibacterial properties. The in vivo evaluation further demonstrated that the hydrogel possessed considerable wound-healing promotion effect by regulating diabetic microenvironment, attributed to that the hydrogel could significantly reduce inflammatory response, alleviate oxidative stress and regulate neovascularization. Overall, this biosafe adhesive hydrogel had great potentials for diabetic wound management.

Chitin nanofibrils assisted 3D printing all-chitin hydrogels for wound dressing.

The direct ink writing technique used in 3D printing technology is generally applied to designing biomedical hydrogels. Herein, we proposed a strategy for preparing all-chitin-based inks for wound dressing via direct ink writing technique. The ß-chitin nanofibers (MACNF) with a high aspect ratio were applied as a nanofiller to modulate the rheological properties of the alkaline dissolved chitin solution. The printing fidelity significantly depends on the MACNF introduction amount to the composite ink. 5-10 wt% MACNF ratio showed superior printing performance. The printed scaffold showed a uniform micron-sized pore structure and a woven network of nanofibers. Due to the good biocompatibility of chitin and the stereoscopic spatial skeleton, this scaffold showed excellent performance as a wound dressing, which can promote cell proliferation, collagen deposition and the angiogenesis of wounds, demonstrating its potential in biomedical applications. This approach successfully balanced the chitinous printability and biofunctions.

An Adaptable Drug Delivery System Facilitates Peripheral Nerve Repair by Remodeling the Microenvironment.

The multifaceted process of nerve regeneration following damage remains a significant clinical issue, due to the lack of a favorable regenerative microenvironment and insufficient endogenous biochemical signaling. However, the current nerve grafts have limitations in functionality, as they require a greater capacity to effectively regulate the intricate microenvironment associated with nerve regeneration. In this regard, we proposed the construction of a functional artificial scaffold based on a "two-pronged" approach. The whole system was developed by encapsulating Tazarotene within nanomicelles formed through self-assembly of reactive oxygen species (ROS)-responsive amphiphilic triblock copolymer, all of which were further loaded into a thermosensitive injectable hydrogel. Notably, the hydrogel exhibits obvious temperature sensitivity at a concentration of 6 wt %, and the nanoparticles possess concentration-dependent H2O2-response capability with a controlled release profile in 48 h. The combined strategy promoted the repair of injured peripheral nerves, attributed to the dual role of the materials, which mainly involved providing structural support, modulating the immune microenvironment, and enhancing angiogenesis. Overall, this study opens up intriguing prospects in tissue engineering.

Multi-crosslinked hydrogel built with hyaluronic acid-tyramine, thiolated glycol chitosan and copper-doped bioglass nanoparticles for expediting wound healing.

The migration of fibroblasts and endothelial cells is a critical determinant of wound-healing outcomes for skin injuries. Here, hyaluronic acid-tyramine (HAT) and thiolated glycol chitosan (TGC) conjugates were combined with copper-doped bioglass (ACuBG) nanoparticles to build a novel type of multi-crosslinked hydrogel for stimulating the migration of cells, and thus, expediting wound healing. The optimally devised HAT/TGC/ACuBG gels had markedly improved strength and stiffness compared to the gels built from either HAT or TGC while showing sufficient elasticity, which contributes to stimulating the migration of fibroblasts. The sustainable release of silicon and copper ions from the gels was found to jointly induce the migration of human umbilical vein endothelial cells. The results based on mouse full-thickness skin defects demonstrated that they were able to fully restore the skin defects with formation of complete appendages within two weeks, suggesting their promising potency for use in expediting wound healing.

Real-time visualization of skeletal muscle necrosis in mice and swine through NIR-II/I fluorescence imaging.

Avulsion often occurs in the limb due to heavy shearing forces which not only damage skeletal muscle but also main vessels, resulting in life-threatening muscle ischemia and necrosis. Defining muscle activity is vital for surgical repair. Currently, the color, capacity of blood, contractibility, and consistency (4C) are the primary principles for evaluating the activities of torn muscles. Based on clinical experiences, this standard turns out to be delayed diagnosis, which is not defined by specific parameters. Recently, near-infrared (NIR) fluorescence probes emitting within the second near-infrared window (NIR-II, 1000-1700 nm) have been widely used for non-invasive optical imaging because the tissue absorption and autofluorescence in the NIR-II region are negligible, thus allowing deeper penetration depths with micrometer-scale spatial resolution in vivo. As pathogenesis and development of muscle necrosis, necrosis-related protein may participate in this procedure. There is promising future for NIR-II to be used in evaluating muscle activity in avulsion. A new approach is developed based on experiments with mice and large animals (swine). Myoblasts were incubated with indocyanine green (ICG) to identify the necrosis muscles. The model of extremity damaged muscle was established for the real-time visualization and detection of developed necrosis muscle field under new equipment, both in balb/c mice (female) and long-haired swines. A visible NIR-II/I imaging system was first used in a large animal injured skeletal muscle-related model. Our NIR-II/I imaging system is suitable for evaluating the normal and injured skeletal muscle ICG cycle and pointing to the necrotic skeletal muscle tissue. NIR-II imaging is superior to NIR-I imaging in estimating skeletal muscle, best with 1100 nm filter. NIR-II fluorescence with 1100 nm filter is suitable for analyzing the progress of necrosis muscle tissue, leading to a new approach for intraoperative evaluation.

Monitoring angiogenesis in skin autografts using photoacoustic microscopy.

BACKGROUND: Skin autografts have been broadly used to manage the skin and soft tissue defects. It is important for surgeons to assess the vitality of skin autografts via observing the angiogenesis. However, there is lack of reliable approach for giving the quantitative angiogenesis information on the skin autografts. Recently, photoacoustic microscopy imaging has attracted much attention based on its good performance in angiography. METHODS: In this study, we aim to monitor angiogenesis in skin autografts via PAM, and further verify its clinical potential for the early prediction of skin autografts clinical outcome. RESULTS AND CONCLUSIONS: The results indicate that PAM is a feasible, precise, high-resolution, noninvasive technique for the early prediction of necrosis of skin autografts via monitoring the angiogenesis, providing a promising tool for surgeons to use this surgical technology.

Inducing in situ M2 macrophage polarization to promote the repair of bone defects via scaffold-mediated sustained delivery of luteolin.

Immunoregulation mediated bone tissue engineering (BTE) has demonstrated huge potential in promoting repair of critical-size bone defects (CSBDs). The trade-off between stable immunoregulation function and extended immunoregulation period has posed a great challenge to this strategy. Here, we reported a 3D porous biodegradable Poly(HEMA-co-3APBA)/LUT scaffold, in which reversible boronic acid ester bond was formed between the 3APBA moiety and the catechol moiety of luteolin (LUT). The boronic acid ester bond not only protected the bioactivity of LUT but also extended the release period of LUT. The rationale behind the phenomenon of sustained LUT release was explained using a classical transition state theory. In vitro/in vivo assays proved the immunoregulation function of the scaffold in inducing M2 polarization of both M0 and M1 Mφ. The crosstalk between the scaffold treated Raw 264.7 and BMSCs were also investigated through the in vitro co-culture assay. The results demonstrated that the scaffold could induce immunoregulation mediated osteogenic differentiation of BMSCs. In addition, CSBDs model of SD rats was also applied, and the corresponding data proved that the scaffold could accelerate new bone formation, therefore promoting repair of CSBDs. The as-prepared scaffold might be a promising candidate for repair of CSBDs in the future.

Chimeric Exosomes Functionalized with STING Activation for Personalized Glioblastoma Immunotherapy.

A critical challenge of existing cancer vaccines is to orchestrate the demands of antigen-enriched furnishment and optimal antigen-presentation functionality within antigen-presenting cells (APCs). Here, a complementary immunotherapeutic strategy is developed using dendritic cell (DC)-tumor hybrid cell-derived chimeric exosomes loaded with stimulator of interferon genes (STING) agonists (DT-Exo-STING) for maximized tumor-specific T-cell immunity. These chimeric carriers are furnished with broad-spectrum antigen complexes to elicit a robust T-cell-mediated inflammatory program through direct self-presentation and indirect DC-to-T immunostimulatory pathway. This chimeric exosome-assisted delivery strategy possesses the merits versus off-the-shelf cyclic dinucleotide (CDN) delivery techniques in both the brilliant tissue-homing capacity, even across the intractable blood-brain barrier (BBB), and the desired cytosolic entry for enhanced STING-activating signaling. The improved antigen-presentation performance with this nanovaccine-driven STING activation further enhances tumor-specific T-cell immunoresponse. Thus, DT-Exo-STING reverses immunosuppressive glioblastoma microenvironments to pro-inflammatory, tumoricidal states, leading to an almost obliteration of intracranial primary lesions. Significantly, an upscaling option that harnesses autologous tumor tissues for personalized DT-Exo-STING vaccines increases sensitivity to immune checkpoint blockade (ICB) therapy and exerts systemic immune memory against post-operative glioma recrudesce. These findings represent an emerging method for glioblastoma immunotherapy, warranting further exploratory development in the clinical realm.

An injectable and adaptable hydrogen sulfide delivery system for modulating neuroregenerative microenvironment.

Peripheral nerve regeneration is a complex physiological process. Single-function nerve scaffolds often struggle to quickly adapt to the imbalanced regenerative microenvironment, leading to slow nerve regeneration and limited functional recovery. In this study, we demonstrate a "pleiotropic gas transmitter" strategy based on endogenous reactive oxygen species (ROS), which trigger the on-demand H2S release at the defect area for transected peripheral nerve injury (PNI) repair through concurrent neuroregeneration and neuroprotection processing. This H2S delivery system consists of an H2S donor (peroxyTCM) encapsulated in a ROS-responsive polymer (mPEG-PMet) and loaded into a temperature-sensitive poly (amino acid) hydrogel (mPEG-PA-PP). This multi-effect combination strategy greatly promotes the regeneration of PNI, attributed to the physiological effects of H2S. These effects include the inhibition of inflammation and oxidative stress, protection of nerve cells, promotion of angiogenesis, and the restoration of normal mitochondrial function. The adaptive release of pleiotropic messengers to modulate the tissue regeneration microenvironment offers promising peripheral nerve repair and tissue engineering opportunities.

3-AP inhibits the growth of human osteosarcoma by decreasing the activity of the iron-dependent pathway.

3-aminopyridine-2-carboxaldehyde thiosemicarbazone (3-AP) has broad-spectrum antitumor activity. However, its role in osteosarcoma (OS) remains unclear. Therefore, this study explored the effects of 3-AP on OS in vitro and in vivo using three human OS cell lines (MG-63, U2-OS, and 143B) and a nude mice model generated by transplanting 143B cells. The cells and mice were treated with DMSO (control) or gradient concentrations of 3-AP. Then, various assays (e.g., cell counting kit-8, flow cytometry, immunohistochemistry, and western blotting) were performed to assess cell viability and apoptosis levels, as well as γH2A.X (DNA damage correlation), ribonucleotide reductase catalytic subunit M1 and M2 (RRM1 and RRM2, respectively) protein levels (iron-dependent correlation). 3-AP time- and dose-dependably suppressed growth and induced apoptosis in all three OS cell lines, and ferric ammonium citrate (FAC) blocked these effects. Moreover, 3-AP decreased RRM2 and total ribonucleotide reductase (RRM1 plus RRM2) protein expression but significantly increased γH2A.X expression; treatment did not affect RRM1 expression. Again, FAC treatment attenuated these effects. In vivo, the number of apoptotic cells in the tumor slices increased in the 3-AP-treated mice compared to the control mice. 3-AP treatment also decreased Ki-67 and p21 expression, suggesting inhibited OS growth. Furthermore, the expression of RRM1, RRM2, and transferrin receptor protein 1 (i.e., Tfr1) indicated that 3-AP inhibited OS growth via an iron-dependent pathway. In conclusion, 3-AP exhibits anticancer activity in OS by decreasing the activity of iron-dependent pathways, which could be a promising therapeutic strategy for OS.

NIR-triggered photodynamic therapy of traumatic heterotopic ossification with a type II collagen-targeted photosensitizer.

Traumatic heterotopic ossification (HO) represents an intractable sequela following trauma with no currently effective prophylaxis or treatment. Photodynamic therapy (PDT) is a non-invasive treatment for various proliferative diseases. However, the specific effects of PDT on HO development remain unclear. In this study, the therapeutic potential of a near-infrared (NIR) probe-WL-808, composed of type II collagen-binding peptide (WYRGRL) and a PDT photosensitizer (IR-808), was evaluated for the innovative HO-targeted PDT approach. In vitro studies indicated that WL-808 could induce chondrocyte apoptosis and inhibit cell viability through ROS generation under NIR excitation. In vivo, the efficacy of WL-808-mediated PDT was tested on the tenotomy HO model mice. WL-808 specifically targeted the type II collagen cartilaginous template of HO, promoting cell apoptosis and enhancing extracellular matrix (ECM) degradation under 808 nm NIR excitation, which inhibited the final ectopic bone formation. Moreover, no obvious toxicity or side effects were detected after treatment with WL-808. Taken together, WL-808-mediated PDT significantly diminished ectopic cartilage and subsequent bone formation, providing a new perspective for HO prophylaxis and treatment.

Erratum: Author correction to 'Bioengineered miR-124-3p prodrug selectively alters the proteome of human carcinoma cells to control multiple cellular components and lung metastasis in vivo' [Acta Pharmaceutica Sinica B 11 (2021) 3950-3965].

[This corrects the article DOI: 10.1016/j.apsb.2021.07.027.].

Sustained notch signaling inhibition with a gamma-secretase inhibitor prevents traumatic heterotopic ossification.

Background: Traumatic heterotopic ossification (THO) is a devastating sequela following traumatic injuries and orthopedic surgeries. To date, the exact molecular mechanism of THO formation is still unclear, which hinders the development of effective treatments. The process of THO formation is believed to recapitulate a series of spatiotemporal cellular and signaling events that occur during skeletal development. The Notch signaling pathway is a critical genetic regulator in embryological bone development and fracture healing. However, few data are available concerning whether Notch signaling regulates THO development and maturation. Methods: We firstly detected the expressions of Notch target genes in both mouse and human THO samples with quantitative RT-PCR and immunohistochemistry (IHC). Then, tissue-resident mesenchymal progenitor cells (TMPCs) were isolated, and the abilities of the proliferation and osteogenic and chondrogenic differentiation of TMPCs were examined under the intervention of the gamma-secretase inhibitor-DAPT at different time points. Finally, DAPT was also administrated in THO mice by burn and Achilles tenotomy injury, and ectopic cartilage and bone formation were monitored by histology and micro-CT. Results: Several Notch target genes were upregulated in both mouse and human THO tissues. Sustained Notch signaling inhibition by DAPT reduced proliferation, osteogenic and chondrogenic differentiation of TMPCs in a time-dependent manner. Moreover, DAPT administration within 3 weeks could inhibit ectopic cartilage and bone formation in a mouse THO model without affecting the total body bone mass. Conclusions: The Notch signaling serves as an important therapeutic target during THO formation. And sustained gamma-secretase inhibition by DAPT has great potential in repressing chondrogenic and osteogenic differentiation of TMPCs, as well as inhibited ectopic cartilage and bone formation in vivo. The translational potential of this article: Sustained Notch inhibition via systemic DAPT (or other similar gamma-secretase inhibitors) administration has promising clinical utility for inhibiting THO formation, providing new insight into THO prophylaxis and treatment.

Deep-penetration functionalized cuttlefish ink nanoparticles for combating wound infections with synergetic photothermal-immunologic therapy.

The challenge of wound infections post-surgery and open trauma caused by multidrug-resistant bacteria poses a constant threat to clinical treatment. As a promising antimicrobial treatment, photothermal therapy can effectively resolve the problem of drug resistance in conventional antibiotic antimicrobial therapy. Here, we report a deep-penetration functionalized cuttlefish ink nanoparticle (CINP) for photothermal and immunological therapy of wound infections. CINP is decorated with zwitterionic polymer (ZP, namely sulfobetaine methacrylate-methacrylate copolymer) to form CINP@ZP nanoparticles. Natural CINP is found to not only exhibit photothermal destruction of methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli), but also trigger macrophages-related innate immunity and enhance their antibacterial functions. The ZP coating on the surface of CINP enables nanoparticles to penetrate into deeply infected wound environment. In addition, CINP@ZP is further integrated into the thermosensitive Pluronic F127 gel (CINP@ZP-F127). After in situ spraying gel, CINP@ZP-F127 is also documented notable antibacterial effects in mice wound models infected with MRSA and E. coli. Collectively, this approach combining of photothermal therapy with immunotherapy can promote delivery efficiency of nanoparticles to the deep foci of infective wounds, and effectively eliminate wound infections.