Rhein and hesperidin nanoparticles remodel tumor immune microenvironment by reducing CAFs and CCL2 secreted by CAAs for efficient triple-negative breast cancer therapy.

In triple-negative breast cancer (TNBC), the tumor immune microenvironment (TIME) is a highly heterogeneous ecosystem that exerts indispensable roles in tumorigenesis and tumor progression. Cancer-associated fibroblasts (CAFs) and cancer-associated adipocytes (CAAs) are the main matrix components in the TIME of TNBC. CAFs mediate the edesmoplastic response, which is a major driver of the immunosuppressive microenvironment to promote tumor growth. In addition, CAAs, a type of tumor-educated adipocyte, participate in crosstalk with breast cancer and are capable of secreting various cytokines, adipokines and chemokines, especially C-C Motif Chemokine Ligand 2 (CCL2), resulting in changes of cancer cell phenotype and function. Therefore, how to treat tumors by regulating the CAFs and the secretion of CCL2 by CAAs in TIME is investigated here. Our research group previously found that rhein (Rhe) has been identified as effective against CAFs, while hesperidin (Hes) could effectively diminish CCL2 secretion by CAAs. Inspired by the above, we developed unique PLGA-based nanoparticles loaded with Rhe and Hes (RH-NP) using the emulsion solvent diffusion method. The RH-NP particles have an average size of 114.1 ± 0.98 nm. RH-NP effectively reduces CAFs and inhibits CCL2 secretion by CAAs, promoting increased infiltration of cytotoxic T cells and reducing immunosuppressive cell presence within tumors. This innovative, safe, low-toxic, and highly effective anti-tumor strategy could be prospective in TNBC treatment.

Hesperidin PLGA nanoparticles potentiate the efficacy of aPD-1 in treating triple negative breast cancer by regulating CCL2 and ADPN expression in cancer-associated adipocytes.

Triple negative breast cancer (TNBC) represents a heterogeneous subtype of breast cancer characterized by an unfavorable prognosis due to its aggressive biology. Cancer-associated adipocytes (CAAs) play an active role in tumor development, invasion and metastasis, and response to treatment by secreting various cytokines. CAAs secrete CCL2 and ADPN which significantly affect the efficacy of aPD-1 in treating breast cancer. Our recent research has demonstrated that Hesperidin, a natural phenolic compound, significantly inhibits CCL2, elevates ADPN secreted by CAAs in vitro and in vivo, remodels the immune microenvironment, and potentiates the efficacy of aPD-1 in triple-negative breast cancer. We used Oil red staining, Bodipy 493/503 staining and quantitative real-time PCR to verify the formation of CAAs. ELISA was used to detect levels of CCL2, ADPN secreted by CAAs. Changes in the number of immune cells in mouse tumor tissues were detected using flow cytometry and immunofluorescence. Our data suggest that Hesperidin PLGA nanoparticles significantly reduced CCL2 and increased ADPN secreted by CAAs, which concurrently decreased the recruitment of M2 macrophages, Tregs and MDSCs while increased the infiltration of CD8+T cells, M1 macrophages and DCs into tumor, thus significantly potentiated the efficacy of aPD-1 in vivo. This study provides a new combined strategy for the clinical treatment of triple-negative breast cancer by interfering with CCL2, ADPN secreted by CAAs to enhance the efficacy of immunotherapy.

HOXA5 inhibits the proliferation and metastasis of cervical squamous cell carcinoma by suppressing the ß-catenin/Snail signaling.

HOXA5, as a transcription factor, plays an important role in a variety of malignant tumors. Nevertheless, its biological role in cervical squamous cell carcinoma (CSCC) is largely unknown. In our study, we aimed to explore the function of HOXA5 in CSCC and its molecular mechanism. Immunohistochemistry showed that HOXA5 expression was downregulated in human CSCC tissues and HOXA5 staining was negatively correlated with tumor size and histological grade of CSCC. Ectopic expression of HOXA5 inhibited proliferative and metastatic abilities of CSCC cells in vitro and in vivo. Furthermore, overexpression of HOXA5 inhibited the cell cycle by arresting the S/G2 phase by flow cytometry and that was related to the downregulation of Cyclin A. Further study showed that HOXA5 suppressed EMT by inhibiting the ß-catenin/Snail signaling resulting in reduced metastasis of CSCC cells. Altogether, our results suggested that HOXA5 inhibited the proliferation and metastasis via repression of the ß-catenin/Snail pathway, proposing the potential role of HOXA5 in the prevention and treatment of CSCC.

Riboflavin photo-cross-linking method for improving elastin stability and reducing calcification in bioprosthetic heart valves.

BACKGROUND: Glutaraldehyde cross-linked bioprosthetic heart valves might fail due to progressive degradation and calcification. METHODS: In this study, we developed a new BHVs preparation strategy named as "HPA/TRA/FMN" that utilized 3,4-hydroxyphenylpropionic acid (HPA)/tyramine (TRA) conjugated pericardium and riboflavin 5'-monophosphate (FMN) initiated photo-cross-linking method. HPA/TRA-pericardium conjugation would provide extra phenol groups for FMN initiated photo-cross-linking. RESULTS: The feeding ratio of riboflavin 5'-monophosphate was optimized. The collagenase and elastase enzymatic degradation in vitro, biomechanics, calcification, elastin stability in vivo, and macrophage marker CD68 were characterized. We demonstrated that riboflavin photo-cross-linked pericardiums had great collagen and elastin stability, improved mechanical properties, better resistance for calcification, and less CD68 positive macrophages in rat subdermal implantation study. CONCLUSIONS: This new riboflavin photo-cross-linking strategy would be a promising method to make BHVs which have better elastin stability, less calcification, and reduced inflammatory response.

Hydrogel hybrid porcine pericardium for the fabrication of a pre-mounted TAVI valve with improved biocompatibility.

Transcatheter aortic valve implantation (TAVI) has been developed years ago for patients who cannot undergo a surgical aortic valve replacement (SAVR). Although TAVI possesses the advantages of lower trauma and simpler manipulation compared to SAVR, the need for storage in glutaraldehyde (GLU) and a tedious intraoperative assembly process have caused great inconvenience for its further application. A pre-mounted TAVI valve assembled by mounting a dry valve frame to a delivery system is expected to address these problems. However, the currently used GLU treated leaflet cannot unfold normally after being crimped for a long-term and loses its function when the BHV is assembled to the catheter. Besides, its cytotoxicity and immune response after implantation are still problems to be solved. In the present study, a hydrogel hybrid porcine pericardium (HHPP) approach was developed to endow the BHVs with a favorable unfolding property and good biocompatibility. Three monomers with different charge characteristics (sodium acrylate, 2-methacryloyloxyethyl phosphorylcholine, and acryloyloxyethyltrimethyl ammonium chloride) were complexed with GLU treated PP (GLU-PP) to form three kinds of HHPPs (SAAH-PP, MPCH-PP, and DACH-PP). The results of the crimping simulation experiment showed that all HHPPs could quickly recover in PBS after being folded for 10 days, while the traditional BHVs (GLU-PP) could not recover under the same conditions. Bovine serum albumin adsorption and platelet adhesion test showed that SAAH-PP and MPCH-PP had good anti-adhesion abilities. A cell culture study indicated that all the three HHPPs promoted HUVEC growth and proliferation. In vivo biocompatibility studies showed that the immune response induced by MPCH-PP was reduced compared to that by GLU-PP. These studies demonstrated that the strategy of MPC hydrogel hybridization may be an effective approach to prepare a pre-mounted TAVI valve with improved biocompatibility.

Radical polymerization-crosslinking method for improving extracellular matrix stability in bioprosthetic heart valves with reduced potential for calcification and inflammatory response.

In recent years, the number of heart valve replacements has multiplied with valve diseases because of aging populations and the surge in rheumatic heart disease in young people. Among them, bioprosthetic heart valves (BHVs) have become increasingly popular. Transcatheter aortic valve implantation (TAVI) valve as an emerging BHV has been increasingly applied to patients. However, the current commercially used BHVs treated with glutaraldehyde (Glut) still face the problem of durability. BHVs derived from Glut-treated xenogenetic tissues would undergo structural degeneration and calcification sometimes even as short as less than 10 years. This issue has already become a big challenge considering more and more young patients at the age of 50-60 s are receiving the BHV replacement. In our study, an approach that is totally different from the previous techniques named by us as the radical polymerization-crosslinking (RPC) method was developed to improve extracellular matrix stability, prevent calcification, and reduce inflammatory response in BHVs. The porcine pericardium (PP) tissue was decellularized, functionalized with methacryloyl groups, and subsequently crosslinked by radical polymerization. We found that high-density RPC treatment remarkably improved the stability of collagen and elastin of PP, enhanced its endothelialization potential, and provided reliable biomechanical performance as compared to Glut treatment. The in vivo rat model also confirmed the increased componential stability and the reduced inflammatory response of RPC-treated PP. Moreover, the RPC-treated PP showed better in vivo anticalcification potential than Glut-treated PP. STATEMENT OF SIGNIFICANCE: Bioprosthetic heart valves (BHVs) manufactured from glutaraldehyde (Glut)-treated xenogeneic tissues have been used to treat valve-related diseases for several decades. However, the durability of BHVs remains unresolved and becomes more pronounced particularly in younger patients. Although a number of new alternative methods for Glut crosslinking have been proposed, their overall performance is still far from ready to use in humans. In this study, radical polymerization was investigated for crosslinking the porcine pericardium (PP). This treatment was found to have advantages compared to Glut-treated PP in terms of stability, biocompatibility, and anticalcification potential with the hope of addressing the needs of more robust biomaterials for the fabrication of BHVs.

Elastin Stabilization Through Polyphenol and Ferric Chloride Combined Treatment for the Enhancement of Bioprosthetic Heart Valve Anticalcification.

The lifetime of bioprosthetic heart valves (BHVs) is limited by the mechanical damage and calcification. The major components of BHVs are collagen and elastin. Collagen could be well protected by glutaraldehyde (GLUT) crosslinking, while elastin is not stabilized and has a high risk of degradation, which could lead to the calcification of BHVs. We aimed to develop methods for stabilizing elastin and decreasing calcification. We investigated the combined tannic acid (TA) or epigallocatechin gallate (EGCG) with ferric chloride to stabilize elastin and prevent calcification. We found that the amount of TA/EGCG bound to elastin was in a time-dependent pattern and this reaction showed better efficiency in acidic condition and ethanol-water mixed solvents. Moreover, Fe3+ could compete with Ca2+ to bind to polyphenol, which could reduce the calcium deposition on BHVs. Cytotoxicity test showed that all extracts from different treatments had similar cell viabilities (85-100%). Through the combined treatments of polyphenol and ferric chloride, the pericardium had a better resistance to elastase degradation and more excellent anticalcification performance.