Engineered exosomes and composite biomaterials for tissue regeneration.

Exosomes, which are small vesicles enclosed by a lipid bilayer and released by many cell types, are widely dispersed and have garnered increased attention in the field of regenerative medicine due to their ability to serve as indicators of diseases and agents with therapeutic potential. Exosomes play a crucial role in mediating intercellular communication through the transfer of many biomolecules, including proteins, lipids, RNA, and other molecular constituents, between cells. The targeted transport of proteins and nucleic acids to specific cells has the potential to enhance or impair specific biological functions. Exosomes have many applications, and they can be used alone or in combination with other therapeutic approaches. The examination of the unique attributes and many functions of these factors has emerged as a prominent field of study in the realm of biomedical research. This manuscript summarizes the origins and properties of exosomes, including their structural, biological, physical, and chemical aspects. This paper offers a complete examination of recent progress in tissue repair and regenerative medicine, emphasizing the possible implications of these methods in forthcoming tissue regeneration attempts.

Double-layered microneedle patch loaded with bioinspired nano-vaccine for melanoma treatment and wound healing.

Malignant melanoma is a challenging problem worldwide, because the remaining tumor cells and extensive skin defects following surgical resection are difficult to treat. Biomaterial-mediated immunotherapy has emerged as a superior strategy for anti-tumor applications in recent years. Herein, a unique double-layer MNP was developed to address the problem of malignant melanoma. Hydroxyapatite (HAP) and short-chain peptides from tumor cells were self-assembled to prepare the bioinspired nano-vaccine, and then they were loaded onto the microneedle tips of methacrylated gelatin (GelMA)-based MNP. The products (dubbed HVMN) demonstrated relatively good biocompatibility and immune activity, inhibiting the proliferation and inducing apoptosis of malignant melanoma in a B16 cell-bearing model of C57BL/6 mice, and promoting skin tissue regeneration in a full thickness skin defect model of SD rats in 15 days. The putative molecular pathways were examined preliminarily. In conclusion, this research will develop a competitive microneedle patch with dual anti-tumor and pro-regenerative properties for the postoperative treatment of malignant melanoma.

Fabrication of a core-shell nanofibrous wound dressing with an antioxidant effect on skin injury.

Oxidative stress is one of the obstacles preventing wound regeneration, especially for chronic wounds. Herein, designing a wound dressing with an anti-oxidant function holds great appeal for enhancing wound regeneration. In this study, a biocompatible and degradable nanofiber with a core-shell structure was fabricated via coaxial electrospinning, in which polycaprolactone (PCL) was applied as the core structure, while the shell was composed of a mixture of silk fibroin (SF) and tocopherol acetate (TA). The electrospun PST nanofibers were proven to have a network structure with significantly enhanced mechanical properties. The PSTs exhibited a diameter distribution with an average of 321 ± 134 nm, and the water contact angle of their surface is 124 ± 2°. The PSTs also exhibited good tissue compatibility, which can promote the adhesion and proliferation of L929 cells. Besides, the dissolution of silk fibroin encourages the release of TA, which could play a synergistic effect and regulate the oxidative stress effect in the damaged area, for it promotes the adhesion and proliferation of skin fibroblasts (L929), reduces the cytotoxicity of hydrogen peroxide to cells, and lowers the level of reactive oxygen species. The animal experiment indicated that the PSTs would promote the reconstruction of skin. These nanofibers are expected to repair skin ulcers related to diabetes.

Local Delivery of Glabridin by Biomolecular Microneedle to Accelerate Infected Wound Healing.

Applying antibacterial polymers and pro-regenerative small molecules are two individual strategies for accelerating wound healing. However, integrating those two unique approaches into one therapeutic platform that meets clinical requirements is still a challenge. Herein, a series of antibacterial gelatin methacrylate (GelMA)/ε-polylysine (ε-PL) composite hydrogels (termed as GP-n HGs, n = 0, 10, 20, and 30, respectively) are innovatively fabricated by ultraviolet light (UV) crosslinking. The GP-n HGs are proved to be broad-spectrum antibacterial and biocompatible. Among those GP-n HGs, the GP-20 HG is selectively processed into microneedle following a mold-casting method. Then, the glabridin is loaded into those needles to produce composite microneedle termed GP-20@Gla MN. An S. aureus-infected full-thickness defect model in rats is created to evaluate the wound-healing effect of GP-20@Gla MN. Furthermore, an RNA sequencing assay is performed to explore the possible molecular mechanisms of glabridin in promoting tissue regeneration, and many positive routes are summarized. This work is of significant novelty in fulfilling complex clinical needs by simultaneously optimizing the advanced microneedles' chemical compositions and physical structures. This work will provide a promising therapeutic platform for treating infected and chronic wounds.

Cancer-associated fibroblasts in hematologic malignancies: elucidating roles and spotlighting therapeutic targets.

Hematologic malignancies comprise a diverse range of blood, bone marrow, and organ-related disorders that present significant challenges due to drug resistance, relapse, and treatment failure. Cancer-associated fibroblasts (CAFs) represent a critical component of the tumor microenvironment (TME) and have recently emerged as potential therapeutic targets. In this comprehensive review, we summarize the latest findings on the roles of CAFs in various hematologic malignancies, including acute leukemia, multiple myeloma, chronic lymphocytic leukemia, myeloproliferative neoplasms, and lymphoma. We also explore their involvement in tumor progression, drug resistance, and the various signaling pathways implicated in their activation and function. While the underlying mechanisms and the existence of multiple CAF subtypes pose challenges, targeting CAFs and their associated pathways offers a promising avenue for the development of innovative treatments to improve patient outcomes in hematologic malignancies.

Non-heme Iron Single-Atom Nanozymes as Peroxidase Mimics for Tumor Catalytic Therapy.

Single-atom nanozymes (SAzymes) open new possibilities for the development of artificial enzymes that have catalytic activity comparable to that of natural peroxidase (POD). So far, most efforts have focused on the structural modulation of the Fe-N4 moiety to mimic the metalloprotein heme center. However, non-heme-iron POD with much higher activity, for example, HppE, has not been mimicked successfully due to its structural complexity. Herein, carbon dots (CDs)-supported SAzymes with twisted, nonplanar Fe-O3N2 active sites, highly similar to the non-heme iron center of HppE, was synthesized by exploiting disordered and subnanoscale domains in CDs. The Fe-CDs exhibit an excellent POD activity of 750 units/mg, surpassing the values of conventional SAzymes with planar Fe-N4. We further fabricated an activatable Fe-CDs-based therapeutic agent with near-infrared enhanced POD activity, a photothermal effect, and tumor-targeting ability. Our results represent a big step in the design of high-performance SAzymes and provide guidance for future applications for synergistic tumor therapy.

Double-network cellulose-based hybrid hydrogels with favourable biocompatibility and antibacterial activity for wound healing.

Bacterial infections are among the leading causes of delayed wound healing. At present, a series of antibacterial materials, such as antibiotics, antimicrobial peptides (AMPs), metals and metal oxides (MMOs), have been used to fabricate antibacterial wound dressings. However, their translational potential is limited owing to their poor biocompatibility. ε-Polylysine (ε-PL) is a natural macromolecule with excellent biocompatibility and broad-spectrum antibacterial activity. Herein, ε-PL was incorporated into a cellulose/γ-polyglutamic acid (γ-PGA) composite hydrogel to form a novel double-network hydrogel termed as CGLH. The elastic modulus of CGLH increased from 0.097 ± 0.015 MPa to 0.441 ± 0.096 MPa, and the equilibrium swelling ratio increased from 382.7 ± 24.3 % to 611.2 ± 8.6 %. Several preclinical models were used to investigate the translational potential of this hydrogel. CGLH exhibited good biocompatibility and antibacterial activity, which promoted the healing of infected and critical-size wounds within 12 days. CGLH had positive effects on collagen synthesis, vascularization and cell proliferation. As a result, this study not only provided an effective alternative for wound healing but also proposed a double-network strategy for creating biocompatible and antibacterial biomaterials.

Lanosterol synthase loss of function decreases the malignant phenotypes of HepG2 cells by deactivating the Src/MAPK signaling pathway.

Cholesterol is critical for tumor cells to maintain their membrane components, cell morphology and activity functions. The inhibition of the cholesterol pathway may be an efficient strategy with which to limit tumor growth and the metastatic process. In the present study, lanosterol synthase (LSS) was knocked down by transfecting LSS short hairpin RNA into HepG2 cells, and cell growth, apoptosis and migratory potential were then detected by Cell Counting Kit-8 cell proliferation assay, flow cytometric analysis and wound healing assay, respectively. In addition, proteins associated with the regulation of the aforementioned cell biological behaviors were analyzed by western blot analysis. The activity of the Src/MAPK signaling pathway was measured by western blotting to elucidate the possible signal transduction mechanisms. LSS knockdown in the HepG2 liver cancer cell line inhibited cell proliferation, with cell cycle arrest at the S phase; it also decreased cell migratory ability and increased apoptosis. The expression proteins involved in the regulation of cell cycle, cell apoptosis and migration was altered by LSS knockdown in HepG2 cells. Furthermore, a decreased Src/MAPK activity was observed in the HepG2 cells subjected to LSS knockdown. LSS loss of function decreased the malignant phenotypes of HepG2 cells by deactivating the Src/MAPK signaling pathway and regulating expression of genes involved in cell cycle regulation, cell apoptosis and migration.

Monoporous Microsphere as a Dynamically Movable Drug Carrier for Osteoporotic Bone Remodeling.

To repair systematically osteoporotic bone defects, it is important to make an effort to both diminish osteoporosis and enhance bone regeneration. Herein, a specifically monoporous microsphere (MPM) carrier encapsulating dosage-sensitive and short half-time parathyroid hormone (PTH) is constructed to tackle the issue. Compared with conventional microsphere carriers involving compact, porous, and mesoporous microspheres, the MPM is desirable to achieve precisely in situ delivery and to minimize topical accumulation. The findings show that the PTH loaded inside MPMs can be gradually released from the single hole of MPMs to improve the initial drug concentration. Also, the MPMs can self-shift with the daily movement of experimental animals to effectively reduce the topical aggregation of released drugs in vitro. In vivo evaluation further confirms that the implant of MPMs-PTH plays a dual role in stimulating the regenerative repair of the cranial defect and relieving osteoporosis in the whole body. Consequently, the current work develops a dynamically movable drug delivery system to achieve precisely in situ delivery, minimize topical accumulation, and systematically repair osteoporotic bone defects.

Targeting pancreatic stellate cells in chronic pancreatitis: Focus on therapeutic drugs and natural compounds.

Chronic pancreatitis (CP) is a precancerous illness linked to pancreatic ductal adenocarcinoma (PDAC), although the evolutionary mechanism is uncertain. CP is distinguished by severe fibrosis caused by the activation of pancreatic stellate cells (PSCs). The current clinical therapeutic protocol for CP lacks specific therapeutic medicines for the prevention and suppression of inflammation and fibrosis aggravating in CP. More research on specifically targeting PSCs would help facilitate the development of novel therapies for pancreatic fibrosis. Notably, using natural compounds from medicinal plants as new antifibrotic agents has become a focus of recent research and is widely employed as an alternative and complementary approach. Our goal was to shed light on the role of PSCs in the development of CP and provide a focused update on the new potential therapeutic strategies against PSCs in CP models. Future studies can refer to these possible strategies for drug design, bioavailability, pharmacokinetics, and other issues to obtain better clinical outcomes for treating CP.

Robotic Intracellular Electrochemical Sensing for Adherent Cells.

Nanopipette-based observation of intracellular biochemical processes is an important approach to revealing the intrinsic characteristics and heterogeneity of cells for better investigation of disease progression or early disease diagnosis. However, the manual operation needs a skilled operator and faces problems such as low throughput and poor reproducibility. This paper proposes an automated nanopipette-based microoperation system for cell detection, three-dimensional nonovershoot positioning of the nanopipette tip in proximity to the cell of interest, cell approaching and proximity detection between nanopipette tip and cell surface, and cell penetration and detection of the intracellular reactive oxygen species (ROS). A robust focus algorithm based on the number of cell contours was proposed for adherent cells, which have sharp peaks while retaining unimodality. The automated detection of adherent cells was evaluated on human umbilical cord vein endothelial cells (HUVEC) and NIH/3T3 cells, which provided an average of 95.65% true-positive rate (TPR) and 7.59% false-positive rate (FPR) for in-plane cell detection. The three-dimensional nonovershoot tip positioning of the nanopipette was achieved by template matching and evaluated under the interference of cells. Ion current feedback was employed for the proximity detection between the nanopipette tip and cell surface. Finally, cell penetration and electrochemical detection of ROS were demonstrated on human breast cancer cells and zebrafish embryo cells. This work provides a systematic approach for automated intracellular sensing for adherent cells, laying a solid foundation for high-throughput detection, diagnosis, and classification of different forms of biochemical reactions within single cells.

FUS-induced circRHOBTB3 facilitates cell proliferation via miR-600/NACC1 mediated autophagy response in pancreatic ductal adenocarcinoma.

BACKGROUND: Circular RNAs (circRNAs) are becoming a unique member of non-coding RNAs (ncRNAs) with emerging evidence of their regulatory roles in various cancers. However, with regards to pancreatic ductal adenocarcinoma (PDAC), circRNAs biological functions remain largely unknown and worth investigation for potential therapeutic innovation. METHODS: In our previous study, next-generation sequencing was used to identify differentially expressed circRNAs in 3 pairs of PDAC and adjacent normal tissues. Further validation of circRHOBTB3 expression in PDAC tissues and cell lines and gain-and-loss function experiments verified the oncogenic role of circRHOBTB3. The mechanism of circRHOBTB3 regulatory role was validated by pull-down assays, RIP, luciferase reporter assays. The autophagy response of PANC-1 and MiaPaca-2 cells were detected by mCherry-GFP-LC3B labeling and confocal microscopy, transmission electron microscopy and protein levels of LC3B or p62 via Western blot. RESULTS: circRHOBTB3 is highly expressed in PDAC cell lines and tissues, which also promotes PDAC autophagy and then progression in vitro and in vivo. Mechanistically, circRHOBTB3 directly binds to miR-600 and subsequently acts as a miRNA-sponge to maintain the expression level of miR-600-targeted gene NACC1, which facilitates the autophagy response of PDAC cells for adaptation of proliferation via Akt/mTOR pathway. Moreover, the RNA-binding protein FUS (FUS) directly binds to pre-RHOBTB3 mRNA to mediate the biogenesis of circRHOBTB3. Clinically, circRHOBTB3, miR-600 and NACC1 expression levels are correlated with the prognosis of PDAC patients and serve as independent risk factors for PDAC patients. CONCLUSIONS: FUS-mediated circRHOBTB3 functions as a tumor activator to promote PDAC cell proliferation by modulating miR-600/NACC1/Akt/mTOR axis regulated autophagy.

CircNEIL3 regulatory loop promotes pancreatic ductal adenocarcinoma progression via miRNA sponging and A-to-I RNA-editing.

BACKGROUND: A growing number of studies have focused on investigating circRNAs as crucial regulators in the progression of multiple cancer types. Nevertheless, the biological effects and underlying mechanisms of circRNAs in pancreatic ductal adenocarcinoma (PDAC) remain unclear. METHODS: Differentially expressed circRNAs between cancerous tissue and adjacent normal tissues were identified by RNA sequencing in PDAC. Subsequently, in vitro and in vivo functional experiments were performed to investigate the functional roles of circNEIL3 in PDAC tumour growth and metastasis. Furthermore, RNA pull-down, dual-luciferase reporter assays, RNA immunoprecipitation (RIP) assays, fluorescent in situ hybridization (FISH) and Sanger sequencing assays were performed to examine the circular interaction among circNEIL3, miR-432-5p and adenosine deaminases acting on RNA 1 (ADAR1). RESULTS: CircNEIL3 was upregulated in PDAC and promoted the progression of PDAC cells both in vitro and in vivo. Mechanistically, circNEIL3 was shown to regulate the expression of ADAR1 by sponging miR-432-5p to induce RNA editing of glioma-associated oncogene 1 (GLI1), ultimately influencing cell cycle progression and promoting epithelial-to-mesenchymal transition (EMT) in PDAC cells. Moreover, we discovered that the circNEIL3/miR-432-5p/ADAR1 axis was correlated with the PDAC clinical stage and overall survival of PDAC patients, while ADAR1 may reduce the biogenesis of circNEIL3. CONCLUSIONS: Our findings reveal that circNEIL3 facilitates the proliferation and metastasis of PDAC through the circNEIL3/miR-432-5p/ADAR1/GLI1/cell cycle and EMT axis and that its expression is regulated by ADAR1 through a negative feedback loop. Therefore, circNEIL3 may serve as a prognostic marker and a therapeutic target for PDAC.

High Flexible and Broad Antibacterial Nanodressing Induces Complete Skin Repair with Angiogenic and Follicle Regeneration.

Complete skin reconstruction is a hierarchically physiological assembly involving epidermis, dermis, vasculature, innervation, hair follicles, and sweat glands. Despite various wound dressings having been developed for skin regeneration, few works refer to the complete skin regeneration, particularly lacking for vasculatures and hair follicles. Herein, an instructive wound dressing that integrates the antibacterial property of quaternized chitin and the mechanical strength and biological multifunction of silk fibroin through layer-by-layer electrostatic self-assembly is designed and reported. The resultant dressings exhibit a nanofibrous structure ranging from 471.5 ± 212.1 to 756.9 ± 241.8 nm, suitable flexibility with tensile strength up to 4.47 ± 0.29 MPa, and broad-spectrum antibacterial activity against Escherichia coli and Staphylococcus aureus. More interestingly, the current dressing system remarkably accelerates in vivo vascular reconstruction within 15 days, and the number of regenerated hair follicles reaches up to 22 ± 4 mm-2 , which is comparable to the normal tissue (27 ± 2 mm-2 ). Those crucial functions might originate from the combination between quaternized chitin and silk fibroin and the hierarchical structure of electrospun nanofiber. This work establishes an easy but effective pathway to design a multifunctional wound dressing for the complete skin regeneration.

Green Gas-Mediated Cross-Linking Generates Biomolecular Hydrogels with Enhanced Strength and Excellent Hemostasis for Wound Healing.

Forming biomolecular hydrogels with a combination of high strength and biocompatibility is still a challenge. Herein, we demonstrated a green gas (CO2)-mediated chemical cross-linking strategy that can produce a double-network cellulose/silk fibroin hydrogel (CSH) with significantly elevated mechanical strength while bypassing the toxicity of routine cross-linking agents. Specifically, cellulose and silk fibroin (SF) were first covalently cross-linked in NaOH/urea solution to create the primary network. Then, CO2 gas was introduced into the resultant CSH precursor gels to form carbonates to reduce the pH value of the intra-hydrogel environment from basic to neutral conditions. The pH reduction induced the ordered aggregation of cellulose chains and concomitant hydrogen bonding between these chains, leading to the formation of hydrogels with significantly improved mechanical strength. The CSHs could promote the adhesion and proliferation of the mouse fibroblast cell line (L929), and the CSHs proved to be of low hemolysis and could accelerate blood clotting and decrease blood loss. The CSHs with SF content of 1 wt % healed the wound in vivo within only 12 days through the acceleration of re-epithelialization and revascularization. Consequently, our current work not only reported a feasible alternative for wound dressings but also provided a new green gas-mediated cross-linking strategy for generating mechanically strong, hemostatic, and biocompatible hydrogels.

Advances in crosslinking strategies of biomedical hydrogels.

Biomedical hydrogels as sole repair matrices or combined with pre-seeded cells and bioactive growth factors are extensively applied in tissue engineering and regenerative medicine. Hydrogels normally provide three dimensional structures for cell adhesion and proliferation or the controlled release of the loading of drugs or proteins. Various physiochemical properties of hydrogels endow them with distinct applications. In this review, we present the commonly used crosslinking method for hydrogel synthesis involving physical and chemical crosslinks and summarize their current progress and future perspectives.

Poly(amino acid)-Based Carrier for Drug Delivery Systems.

Micro/nanoscale carrier is a crucial component of the drug delivery system that enables to be well designed for improving the pharmacological and therapeutic properties of drug administration. Herein we systemically review a special vector of natural poly(amino acid) (PAA) that has been extensively applied in the controlled delivery of drug, protein and gene due to its excellent biocompatibility, controlled degradability, and flexible physicochemical modification. Natural PAA is a group of poly(ionic) molecules that present various biological roles and putative technical functions. The intention of this review is to thoroughly summarize three classic types of PAA involving anionic, cationic and neutral PAA as well as their applications on drug delivery systems.

Poly(γ-glutamic acid) modulates the properties of poly(ethylene glycol) hydrogel for biomedical applications.

In this paper, a series of copolymer hydrogels were fabricated from methacrylated poly(γ-glutamic acid) (mPGA) and poly(ethylene glycol) diacrylate (PEGDA). The effect of ionic strength and pH on the swelling behavior and mechanical properties of these hydrogels were studied in detail. Release of Rhodamine B as a model drug from the hydrogel was evaluated under varied pH. In vitro photoencapsulation of bovine cartilage chondrocytes was performed to assess the cytotoxicity of this copolymer hydrogel. The results revealed that the copolymer hydrogel is ionic- and pH-sensitive, and does not exhibit acute cytotoxicity; this copolymer hydrogel may have promising application as matrix for controlled drug release and scaffolding material in tissue engineering.

Enzymatically cross-linked hyaluronic acid/graphene oxide nanocomposite hydrogel with pH-responsive release.

Hyaluronic acid (HA) is made up of repeating disaccharide units (ß-1,4-d-glucuronic acid and ß-1,3-N-acetyl-d-glucosamine) and is a major constituent of the extracellular matrix. HA and its derivatives which possess excellent biocompatibility and physiochemical properties have been studied in drug delivery and tissue engineering applications. Tyramine-based HA hydrogel with good compatibility to cell and tissue has been reported recently. However, inferior mechanical property may limit the biomedical application of the HA hydrogel. In this study, HA/graphene oxide (GO) nanocomposite (NC) hydrogel was prepared through a horseradish peroxidase catalyzed in situ cross-linking process. As compared with pure HA hydrogels, incorporation of GO to the HA matrix could significantly enhance the mechanical properties (storage moduli 1800 Pa) of the hydrogel and prolong the release of rhodamine B (RB) as the model drug from the hydrogel (33 h) as well. In addition, due to the multiple interactions between GO and RB, the NC hydrogels showed excellent pH-responsive release behavior. The release of RB from the NC hydrogel was prolonged at low pH (pH 4.0) in the presence of GO, which could be attributed to the enhanced interactions between GO and HA as well as with RB. In situ three-dimensional encapsulation of mouse embryonic fibroblasts (BALB 3T3 cells) in the NC hydrogels and cytotoxicity results indicated the cytocompatibility of both the enzymatic cross-linking process and HA/GO NC hydrogels (cell viability 90.6 ± 4.25%). The enzymatically catalyzed fabrication of NC hydrogels proved to be an easy and mild approach, and had great potential in the construction of both tissue engineering scaffolds and stimuli-responsive drug release matrices.