Artificial heart valve reinforced with silk woven fabric and poly (ethylene glycol) diacrylate hydrogels composite.

The present study describes the preparation of woven silk fabric (WSF) and poly(ethylene glycol) diacrylate (PEGDA) hydrogel composite reinforced artificial heart valve (SPAHV). Interestingly, the longitudinal and latitudinal elastic modulus of the SPAHV composite can achieve at 54.08 ± 3.29 MPa and 23.96 ± 2.18 MPa, respectively, while its volume/mass swelling ratio and water permeability was 1.9 %/2.8 % and 3 mL/(cm2∙min), respectively, revealing remarkable anisotropic mechanical properties, low water swelling property and water permeability. The in vitro & in vivo biocompatibility and anti-calcification ability of SPAHV were further examined using L929 mouse fibroblasts and Sprague Dawley (SD) male rat model under 8 weeks of subcutaneous implantation. The expression of pro-inflammatory cytokine TNF-α and anti-inflammatory cytokine IL-10 was determined by immunohistochemical staining, as well as the H&E staining and alizarin red staining were accessed. The results showed that the composites possess better biocompatibility, resistance to degradation and anti-calcification ability compared to the control group (p < 0.05). Thus, the SPAHV composite with robust mechanical properties and biocompatibility has potential application for artificial heart valves.

Supercritical CO2-assisted fabrication of CM-PDA/SF/nHA nanofibrous scaffolds for bone regeneration and chemo-photothermal therapy against osteosarcoma.

Concurrent treatment of tumor recurrence and bone defects after surgical resection of osteosarcoma remains a clinical challenge. Combination therapy based on local drug delivery systems shows great promise in the treatment of osteosarcoma. In this study, curcumin modified polydopamine nanoparticle loaded silk fibroin doped with nano-hydroxyapatite (CM-PDA/SF/nHA) nanofibrous scaffolds were developed to induce bone defect regeneration and chemo-photothermal synergistic effects against osteosarcoma. These scaffolds exhibited good photothermal conversion efficiency and photostability. Moreover, the results of ALP staining and alizarin red S (ARS) staining indicated that the CM-PDA/SF/1%nHA scaffolds had the most obvious promotion effect on early osteogenic differentiation. The results of in vitro and in vivo anti-osteosarcoma activity showed that the CM-PDA/SF/1%nHA scaffolds exhibited higher anti-osteosarcoma activity compared to the control and SF scaffolds. In addition, the CM-PDA/SF/1%nHA scaffolds could promote the proliferation and differentiation of bone marrow mesenchymal stem cells in vitro and new bone production in vivo. Thus, these results suggested that the CM-PDA/SF/1%nHA scaffolds could improve bone defect regeneration and achieve chemo-photothermal synergistic effects against osteosarcoma.

Biomedical applications of magnetosomes: State of the art and perspectives.

Magnetosomes, synthesized by magnetotactic bacteria (MTB), have been used in nano- and biotechnological applications, owing to their unique properties such as superparamagnetism, uniform size distribution, excellent bioavailability, and easily modifiable functional groups. In this review, we first discuss the mechanisms of magnetosome formation and describe various modification methods. Subsequently, we focus on presenting the biomedical advancements of bacterial magnetosomes in biomedical imaging, drug delivery, anticancer therapy, biosensor. Finally, we discuss future applications and challenges. This review summarizes the application of magnetosomes in the biomedical field, highlighting the latest advancements and exploring the future development of magnetosomes.

Sustainable Antibacterial and Anti-Inflammatory Silk Suture with Surface Modification of Combined-Therapy Drugs for Surgical Site Infection.

Silk sutures with antibacterial and anti-inflammatory functions were developed for sustained dual-drug delivery to prevent surgical site infections (SSIs). The silk sutures were prepared with core-shell structures braided from degummed silk filaments and then coated with a silk fibroin (SF) layer loaded with berberine (BB) and artemisinin (ART). Both the rapid release of drugs to prevent initial biofilm formation and the following sustained release to maintain effective concentrations for more than 42 days were demonstrated. In vitro assays using human fibroblasts (Hs 865.Sk) demonstrated cell proliferation on the materials, and hemolysis was 2.4 ± 0.8%, lower than that required by ISO 10993-4 standard. The sutures inhibited platelet adhesion and promoted collagen deposition and blood vessel formation. In vivo assessments using Sprague-Dawley (SD) rats indicated that the coating reduced the expression of pro-inflammatory cytokines interleukin-10 (IL-10) and tumor necrosis factor-α (TNF-α), shortening the inflammatory period and promoting angiogenesis. The results demonstrated that these new sutures exhibited stable structures, favorable biocompatibility, and sustainable antibacterial and anti-inflammatory functions with potential for surgical applications.

Identification and functional characterization of potential oncofetal targets in human hepatocellular carcinoma.

Here, we present a detailed protocol for the identification of potential oncofetal targets for hepatocellular carcinoma (HCC) patients through a hepatocyte differentiation model and a sorafenib refractory cell-line-derived xenograft model. We describe the procedures of tumor sphere formation, organoid generation, and subcutaneous tumor formation for functional studies. We then detail the procedures of immunohistochemistry and immunofluorescence for examination of changes in lineage-specific markers. Finally, we describe the development of antibody-based therapeutics targeting tumor lineage plasticity in HCC. For complete details on the use and execution of this protocol, please refer to Kong et al. (2021).1.

Targeting tumor lineage plasticity in hepatocellular carcinoma using an anti-CLDN6 antibody-drug conjugate.

Tumor lineage plasticity is emerging as a critical mechanism of therapeutic resistance and tumor relapse. Highly plastic tumor cells can undergo phenotypic switching to a drug-tolerant state to avoid drug toxicity. Here, we investigate the transmembrane tight junction protein Claudin6 (CLDN6) as a therapeutic target related to lineage plasticity for hepatocellular carcinoma (HCC). CLDN6 was highly expressed in embryonic stem cells but markedly decreased in normal tissues. Reactivation of CLDN6 was frequently observed in HCC tumor tissues as well as in premalignant lesions. Functional assays indicated that CLDN6 is not only a tumor-associated antigen but also conferred strong oncogenic effects in HCC. Overexpression of CLDN6 induced phenotypic shift of HCC cells from hepatic lineage to biliary lineage, which was more refractory to sorafenib treatment. The enhanced tumor lineage plasticity and cellular identity change were potentially induced by the CLDN6/TJP2 (tight junction protein 2)/YAP1 (Yes-associated protein 1) interacting axis and further activation of the Hippo signaling pathway. A de novo anti-CLDN6 monoclonal antibody conjugated with cytotoxic agent (Mertansine) DM1 (CLDN6-DM1) was developed. Preclinical data on both HCC cell lines and primary tumors showed the potent antitumor efficiency of CLDN6-DM1 as a single agent or in combination with sorafenib in HCC treatment.

Micronized curcumin fabricated by supercritical CO2 to improve antibacterial activity against Pseudomonas aeruginosa.

Curcumin (CM) is a natural polyphenolic compound with multiple biomedical functions. However, clinical applications face more challenges due to its low dissolution rate and poor bioavailability. Micronization is an effective strategy to overcome these drawbacks. Herein, CM nanoparticles (CM NPs, ∼300 nm) were fabricated using solution enhanced dispersion by supercritical CO2 (SEDS). The solubility of CM NPs was remarkably enhanced. Aim to study the effects of micronization on the biological functions of CM, we investigated the antibacterial activity of original CM and CM NPs upon Pseudomonas aeruginosa. In vitro, the minimal inhibitory concentrations (MIC) assay, solid-medium spot assay, growth kinetics assay and morphologic observation using atomic force microscopy (AFM) confirmed that the anti-P. aeruginosa activity of CM NPs was enhanced compared to original CM. Moreover, CM NPs also showed stronger inhibition for adhesion and biofilm formation of P. aeruginosa compared to original CM. Experiments on mice infected with P. aeruginosa showed that CM NPs have a better therapeutic effect than the original CM in vivo. In summary, CM NPs may be a novel and promising therapeutic candidate for bacterial infection.

Sequential delivery of dual drugs with nanostructured lipid carriers for improving synergistic tumor treatment effect.

To improve synergistic anticancer efficacy and minimize the adverse effects of chemotherapeutic drugs, temozolomide (TMZ) and curcumin (CUR) co-loaded nanostructured lipid carriers (NLCs) were prepared by microemulsion in this study. And the physicochemical properties, drug release behavior, intracellular uptake efficiency, in vitro and in vivo anticancer effects of TMZ/CUR-NLCs were evaluated. TMZ/CUR-NLCs showed enhanced inhibitory effects on glioma cells compared to single drug loaded NLCs, which may be owing to that the quickly released CUR can sensitize the cancer cells to TMZ. The inhibitory mechanism is a combination of S phase cell cycle arrest associated with induced apoptosis. Notably, TMZ/CUR-NLCs can accumulate at brain and tumor sites effectively and perform a significant synergistic anticancer effect in vivo. More importantly, the toxic effects of TMZ/CUR-NLCs on major organs and normal cells at the same therapeutic dosage were not observed. In conclusion, NLCs are promising nanocarriers for delivering dual chemotherapeutic drugs sequentially, showing potentials in the synergistic treatment of tumors while reducing adverse effects both in vitro and in vivo.

High-throughput single-cell analysis of exosome mediated dual drug delivery, in vivo fate and synergistic tumor therapy.

Exosomes could serve as delivery platforms, owing to their good biocompatibility, stability, and long blood circulation time. Tracking the biological fate of exosomes in vivo is essential for evaluating their functions, delivery efficacy, and biosafety, and it is invaluable for guiding exosome-based therapy. Here, we merged a single-cell technique, mass cytometry, with in vivo uptake analysis to comprehensively reveal the fate of exosomes at the single-cell level. In tandem with multivariate cellular phenotyping, in vivo uptake of exosomes labeled with heavy metal-containing tags was quantified in a high-throughput manner. Interestingly, an organ-dependent uptake landscape of exosomes by diverse cell types was distinctly demonstrated, which implied that cancer cells seemed to preferably take up more released drugs from the exosomes. Using these cellular insights, the administration method of drug-loaded exosomes was optimized to elevate their accumulation in tumor sites and minimize their spread into healthy organs. Dual drug-loaded exosomes were locally administered and superior synergistic tumor treatment effects were achieved in a solid tumor model. The disclosure of exosome cellular distribution, together with the successful engineering of exosomes with multiple anticancer capacities, provides a new level of insight into optimizing and enhancing exosome-based drug delivery and synergistic tumor therapy.

Loading of metal isotope-containing intercalators for mass cytometry-based high-throughput quantitation of exosome uptake at the single-cell level.

Nanometer-sized exosomes are being widely studied as cell-to-cell communicators and versatile drug vehicles. Characterizations of the biodistribution of these exosomes are essential for the evaluation of their biological functions and drug delivery efficacy. However, current technologies for exosome tracking rely on fluorescence and have the disadvantages of being low throughput due to the limited number of available channels and spectral spillover. Here, we reported the development of an engineering approach that involves loading of metal isotope-containing intercalators into exosomes to quantify exosome uptake at the single-cell level. We demonstrate that mass cytometry in conjunction with highly multivariate cellular phenotyping enables high-throughput identification of the in vivo fate of exosomes. Inspired by these insights into cellular distribution, we optimized the administration methods for exosome-based drug delivery, verifying the anticancer efficacy of these exosomes in a mouse model of breast cancer. The evaluation of exosome's fate in vivo at the single-cell level provides valuable insights into the functions of exosomes in vivo and facilitates the improvement of exosome-based therapy.

MRI reporter gene MagA suppresses transferrin receptor and maps Fe2+ dependent lung cancer.

As a magnetic resonance imaging (MRI) reporter gene, MagA has become a powerful tool to monitor dynamic gene expression and allowed concomitant high resolution anatomical and functional imaging of subcellular genetic information. Here we establish a stably expressed MagA method for lung cancer MRI. The results show that MagA can not only enhance both in vitro and in vivo MRI contrast by specifically alternating the transverse relaxation rate of water, but also inhibit the malignant growth of lung tumor. In addition, MagA can regulate magnetic nanoparticle production in grafted tissues and also suppress transferrin receptor expression by acting as an iron transporter, and meanwhile can permit iron biomineralization in the presence of mammalian iron homeostasis. This work provides experimental evidence for the safe preclinical applications of MagA as both a potential inhibitor and an MRI-based tracing tool for iron ion-dependent lung cancer.

Supercritical carbon dioxide-developed silk fibroin nanoplatform for smart colon cancer therapy.

PURPOSE: To deliver insoluble natural compounds into colon cancer cells in a controlled fashion. MATERIALS AND METHODS: Curcumin (CM)-silk fibroin (SF) nanoparticles (NPs) were prepared by solution-enhanced dispersion by supercritical CO2 (SEDS) (20 MPa pressure, 1:2 CM:SF ratio, 1% concentration), and their physicochemical properties, intracellular uptake efficiency, in vitro anticancer effect, toxicity, and mechanisms were evaluated and analyzed. RESULTS: CM-SF NPs (<100 nm) with controllable particle size were prepared by SEDS. CM-SF NPs had a time-dependent intracellular uptake ability, which led to an improved inhibition effect on colon cancer cells. Interestingly, the anticancer effect of CM-SF NPs was improved, while the side effect on normal human colon mucosal epithelial cells was reduced by a concentration of ~10 µg/mL. The anticancer mechanism involves cell-cycle arrest in the G0/G1 and G2/M phases in association with inducing apoptotic cells. CONCLUSION: The natural compound-loaded SF nanoplatform prepared by SEDS indicates promising colon cancer-therapy potential.

Composite Membranes of Recombinant Silkworm Antimicrobial Peptide and Poly (L-lactic Acid) (PLLA) for biomedical application.

Antimicrobial peptides, produced by innate immune system of hosts in response to invading pathogens, are capable of fighting against a spectrum of bacteria, viruses, fungi, parasites and cancer cells. Here, a recombinant silkworm AMP Bmattacin2 from heterologous expression is studied, indicating a broad spectrum of antibacterial activity and showing selective killing ability towards skin and colon cancer cells over their normal cell counterparts. For the purpose of biomedical application, the electrospinning fabrication technique is employed to load Bmattacin2 into PLLA nanofibrous membrane. In addition to a good compatibility with the normal cells, Bmattacin2 loaded nanofibrous membranes demonstrate instant antibacterial effects and sustained anticancer effects. The cancer cell and bacteria targeting dynamics of recombinant Bmattacin2 are investigated. With these characteristics, PLLA/Bmattacin2 composite membranes have a great potential for developing novel biomedical applications such as cancer therapies and wound healing treatments.

An implantable and controlled drug-release silk fibroin nanofibrous matrix to advance the treatment of solid tumour cancers.

The development of more effective cancer therapeutic strategies are still critically required. The maximization of the therapeutic effect in combination with avoiding the severe side effects on normal tissues when using chemotherapy drugs is still an urgent problem that requires improvements urgently. Here we provide implantable and controllable drug-release that utilises silk fibroin (SF) as a nanofibrous drug delivery system (DDS) for cancer treatment. A nanofibrous structure with controllable fibre diameter (<100 nm) was produced. The drug release rate of the SF DDS was controlled by applying a post-treatment process. In vitro anti-cancer (HCT116) results indicated that curcumin (CM)-SF nanofibrous matrix had a superior anti-cancer potential when the concentration was >5 µg/mL. The mechanism could be explained by the cell cycle being held in the S phase. The toxic effect on normal cells (NCM460) was minimized by using a treatment concentration range (5-20 µg/mL). Implantation of this DDS into the tumour site inhibited the growth of solid tumour; this offers an alternative approach for novel cancer therapy.

Nano-curcumin prepared via supercritical: Improved anti-bacterial, anti-oxidant and anti-cancer efficacy.

Curcumin (CM) possesses multiple biological activities. However, poor water solubility and low bioavailability limit its application in biomedical fields. CM nanoparticles (NPs) (230-240nm) were prepared by solution-enhanced dispersion via supercritical CO2 (SEDS) (22-22.5MPa pressure, 31-32.5°C temperature) and its biological functions were evaluated in this study. The Minimum inhibitory concentration of CM NPs against S. aureus (∼250µg/mL) was lower than CM-DMSO (∼500µg/mL). Meanwhile, CM NPs showed effective anti-oxidant ability at a concentration raging from 125 to 2000µg/mL. CM NPs showed time-dependent intracellular internalization ability, resulting in an enhanced anti-cancer effect on colorectal cancer cells (HCT116), and the mechanism could be explained by cell cycle arrest in G2/M phase associated with inducing apoptotic cells. Moreover, CM NPs exhibited reduced cytotoxicity on normal cells (NCM460) compared to CM-DMSO and 5-Fu. In conclusion, CM NPs prepared via SEDS showed potentials in biomedical applications.

Silk fibroin-based nanoparticles for drug delivery.

Silk fibroin (SF) is a protein-based biomacromolecule with excellent biocompatibility, biodegradability and low immunogenicity. The development of SF-based nanoparticles for drug delivery have received considerable attention due to high binding capacity for various drugs, controlled drug release properties and mild preparation conditions. By adjusting the particle size, the chemical structure and properties, the modified or recombinant SF-based nanoparticles can be designed to improve the therapeutic efficiency of drugs encapsulated into these nanoparticles. Therefore, they can be used to deliver small molecule drugs (e.g., anti-cancer drugs), protein and growth factor drugs, gene drugs, etc. This paper reviews recent progress on SF-based nanoparticles, including chemical structure, properties, and preparation methods. In addition, the applications of SF-based nanoparticles as carriers for therapeutic drugs are also reviewed.

Construction of a Novel Magnetic Targeting Anti-Tumor Drug Delivery System: Cytosine Arabinoside-Loaded Bacterial Magnetosome.

To ease the side effects triggered by cytosine arabinoside (Ara-C) for acute leukemia treatment, a novel magnetic targeting anti-tumor drug delivery system was constructed through bacterial magnetosomes (BMs) from Magnetospirillum magneticum AMB-1 combined with Ara-C by crosslinking of genipin (GP). The results showed that Ara-C could be bonded onto the membrane surface of BMs effectively through chemical crosslinking induced by dual hand reagents GP. The average diameters of BMs and Ara-C-coupled BMs (ABMs) were 42.0 ± 8.6 and 72.7 ± 6.0 nm respectively, and the zeta potentials (-38.1 ± 9.1) revealed that these systems were stable, confirming the stability of the system. The optimal encapsulation efficiency and drug loading were 89.05% ± 2.33% and 47.05% ± 0.64% respectively when crosslinking reaction lasted for 72 h. The system also presented long-term stability and release behaviors without initial burst release (Ara-C could be released 80% within three months). Our results indicate that BMs have great potential in biomedical and clinical fields as a novel anti-tumor drug carrier.