Programmed release of vascular endothelial growth factor and exosome from injectable chitosan nanofibrous microsphere-based PLGA-PEG-PLGA hydrogel for enhanced bone regeneration.

The healing of large bone defects remains a significant challenge in clinical practice. Accelerating both angiogenesis and osteogenesis can promote effective bone healing. In the natural healing process, angiogenesis precedes osteogenesis, providing a blood supply that supports the subsequent progression of osteogenesis. Developing a biomimetic scaffold that mimics the in vivo environment and promotes the proper sequence of vascularization followed by ossification is crucial for successful bone regeneration. In this study, a novel injectable dual-drug programmed releasing chitosan nanofibrous microsphere-based poly(D, l-lactide-co-glycolide)-b-poly(ethylene glycol)-b-poly(D,l-lactide-co-glycolide) (PLGA-PEG-PLGA) hydrogel is fabricated by incorporating vascular endothelial growth factor (VEGF) and microspheres loaded with dental pulp stem cells-derived exosomes (DPSCs-Exo). Rapid release of VEGF promotes the swift initiation of angiogenesis, while DPSCs-Exo release ensures persistent osteogenesis. Our results demonstrate that chitosan microsphere-based PLGA-PEG-PLGA hydrogel significantly promotes angiogenesis in human umbilical vascular endothelial cells and enhances the osteogenic differentiation of pre-osteoblasts. Furthermore, in vivo transplantation of this injectable chitosan microsphere-based PLGA-PEG-PLGA hydrogel into calvarial bone defects markedly promotes bone formation. Overall, our study provides a promising approach for improving bone regeneration by temporally replicating the behavior of angiogenesis and osteogenesis.

Applications of Stimuli-Responsive Hydrogels in Bone and Cartilage Regeneration.

Bone and cartilage regeneration is an area of tremendous interest and need in health care. Tissue engineering is a potential strategy for repairing and regenerating bone and cartilage defects. Hydrogels are among the most attractive biomaterials in bone and cartilage tissue engineering, mainly due to their moderate biocompatibility, hydrophilicity, and 3D network structure. Stimuli-responsive hydrogels have been a hot topic in recent decades. They can respond to external or internal stimulation and are used in the controlled delivery of drugs and tissue engineering. This review summarizes current progress in the use of stimuli-responsive hydrogels in bone and cartilage regeneration. The challenges, disadvantages, and future applications of stimuli-responsive hydrogels are briefly described.

Mussel-inspired functionalization of electrospun scaffolds with polydopamine-assisted immobilization of mesenchymal stem cells-derived small extracellular vesicles for enhanced bone regeneration.

Mesenchymal stem cells-derived small extracellular vesicles (MSCs-sEV) have shown promising prospects as a cell-free strategy for bone tissue regeneration. Here, a bioactive MSCs-sEV-loaded electrospun silk fibroin/poly(ε-caprolactone) (SF/PCL) scaffold was synthesized via a mussel-inspired immobilization strategy assisted by polydopamine (pDA). This pDA modification endowed the as-prepared scaffold with high loading efficiency and sustained release profile of sEV. In addition, the fabricated composite scaffold exhibited good physiochemical, mechanical, and biocompatible properties. In vitro cellular experiments indicated that the MSCs-sEV-loaded composite scaffold promoted the adhesion and spreading of preosteoblast and endothelial cells, as well as enhanced osteogenic differentiation and angiogenic activity. In vivo experiments showed that the functionalized electrospun scaffolds promoted bone regeneration in a rat calvarial bone defect model. Results suggest that the developed MSCs-sEV-anchored pDA-modified SF/PCL electrospun scaffolds possess high application potential in bone tissue engineering owing to their powerful pro-angiogenic and -osteogenic capacities, cell-free bioactivity, and cost effectiveness.

Spike-specific circulating T follicular helper cell and cross-neutralizing antibody responses in COVID-19-convalescent individuals.

Coronavirus disease 2019 (COVID-19) is caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)1-3 and individuals with COVID-19 have symptoms that can be asymptomatic, mild, moderate or severe4,5. In the early phase of infection, T- and B-cell counts are substantially decreased6,7; however, IgM8-11 and IgG12-14 are detectable within 14 d after symptom onset. In COVID-19-convalescent individuals, spike-specific neutralizing antibodies are variable3,15,16. No specific drug or vaccine is available for COVID-19 at the time of writing; however, patients benefit from treatment with serum from COVID-19-convalescent individuals17,18. Nevertheless, antibody responses and cross-reactivity with other coronaviruses in COVID-19-convalescent individuals are largely unknown. Here, we show that the majority of COVID-19-convalescent individuals maintained SARS-CoV-2 spike S1- and S2-specific antibodies with neutralizing activity against the SARS-CoV-2 pseudotyped virus, and that some of the antibodies cross-neutralized SARS-CoV, Middle East respiratory syndrome coronavirus or both pseudotyped viruses. Convalescent individuals who experienced severe COVID-19 showed higher neutralizing antibody titres, a faster increase in lymphocyte counts and a higher frequency of CXCR3+ T follicular help (TFH) cells compared with COVID-19-convalescent individuals who experienced non-severe disease. Circulating TFH cells were spike specific and functional, and the frequencies of CXCR3+ TFH cells were positively associated with neutralizing antibody titres in COVID-19-convalescent individuals. No individuals had detectable autoantibodies. These findings provide insights into neutralizing antibody responses in COVID-19-convalescent individuals and facilitate the treatment and vaccine development for SARS-CoV-2 infection.

Seasonal variation of red clover (Trifolium pratense L., Fabaceae) isoflavones and estrogenic activity.

Red clover (Trifolium pratense L., Fabaceae) dietary supplements are currently used to treat menopausal symptoms because of their high content of the mildly estrogenic isoflavones daidzein, genistein, formononetin, and biochanin A. These compounds are estrogenic in vitro and in vivo, but little information exists on the best time to harvest red clover fields to maximize content of the isoflavones and thus make an optimal product. Samples of cultivated red clover above-ground parts and flower heads were collected in parallel over one growing season in northeastern Illinois. Generally, autohydrolytic extracts of above-ground parts contained more isoflavones and had more estrogenic activity in Ishikawa endometrial cells as compared with extracts of flower heads. Daidzein and genistein contents peaked around June to July, while formononetin and biochanin A contents peaked in early September. Flower head and total above-ground parts extracts exhibited differential estrogenic activity in an Ishikawa (endometrial) cell-based alkaline phosphatase induction assay, whereas nondifferential activity was observed for most extracts tested in an MCF-7 (breast) cell proliferation assay when tested at the same final concentrations. Ishikawa assay results could be mapped onto the extracts' content of individual isoflavones, but MCF-7 results did not show such a pattern. These results suggest that significant metabolism of isoflavones may occur in MCF-7 cells but not in Ishikawa cells; therefore, caution is advised in the choice of bioassay used for the biological standardization of botanical dietary supplements.

Comparison of the in vitro estrogenic activities of compounds from hops (Humulus lupulus) and red clover (Trifolium pratense).

Because the prevailing form of hormone replacement therapy is associated with the development of cancer in breast and endometrial tissues, alternatives are needed for the management of menopausal symptoms. Formulations of Trifolium pratense L. (red clover) are being used to alleviate menopause-associated hot flashes but have shown mixed results in clinical trials. The strobiles of Humulus lupulusL. (hops) have been reported to contain the prenylflavanone, 8-prenylnaringenin (8-PN), as the most estrogenic constituent, and this was confirmed using an estrogen receptor ligand screening assay utilizing ultrafiltration mass spectrometry. Extracts of hops and red clover and their individual constituents including 8-PN, 6-prenylnaringenin (6-PN), isoxanthohumol (IX), and xanthohumol (XN) from hops and daidzein, formononetin, biochanin A, and genistein from red clover were compared using a variety of in vitro estrogenic assays. The IC50 values for the estrogen receptor alpha and beta binding assays were 15 and 27 microg/mL, respectively, for hops and 18.0 and 2.0 microg/mL, respectively, for the red clover extract. Both of the extracts, genistein, and 8-PN activated the estrogen response element (ERE) in Ishikawa cells while the extracts, biochanin A, genistein, and 8-PN, significantly induced ERE-luciferase expression in MCF-7 cells. Hop and red clover extracts as well as 8-PN up-regulated progesterone receptor (PR) mRNA in the Ishikawa cell line. In the MCF-7 cell line, PR mRNA was significantly up-regulated by the extracts, biochanin A, genistein, 8-PN, and IX. The two extracts had EC50 values of 1.1 and 1.9 microg/mL, respectively, in the alkaline phosphatase induction assay. On the basis of these data, hops and red clover could be attractive for the development as herbal dietary supplements to alleviate menopause-associated symptoms.

Multiple myeloma regression mediated by bruceantin.

PURPOSE: Bruceantin has been shown to induce cell differentiation in a number of leukemia and lymphoma cell lines. It also down-regulated c-MYC, suggesting a correlation of down-regulation with induction of cell differentiation or cell death. In the present study, we focused on multiple myeloma, using the RPMI 8226 cell line as a model. EXPERIMENTAL DESIGN: The effects of bruceantin on c-MYC levels and apoptosis were examined by immunoblotting, 4',6-diamidino-2-phenylindole staining, evaluation of caspase-like activity, and 3,3'-dihexyloxacarbocyanine iodide staining. The potential of bruceantin to inhibit primary tumor growth was assessed with RPMI 8226 xenografts in SCID mice, and apoptosis in the tumors was evaluated by the terminal deoxynucleotidyl transferase-mediated nick end labeling assay. RESULTS: c-MYC was strongly down-regulated in cultured RPMI 8226 cells by treatment with bruceantin for 24 h. With U266 and H929 cells, bruceantin did not regulate c-MYC in this manner. Apoptosis was induced in the three cell lines. In RPMI 8226 cells, apoptosis occurred through proteolytic processing of procaspases and degradation of poly(ADP-ribose) polymerase. The mitochondrial pathway was also involved. Because RPMI 8226 cells were the most sensitive, they were used in a xenograft model. Bruceantin treatment (2.5-5 mg/kg) resulted in a significant regression of tumors without overt toxicity. Apoptosis was significantly elevated in tumors derived from animals treated with bruceantin (37%) as compared with the control tumors (14%). CONCLUSIONS: Bruceantin interferes with the growth of RPMI 8226 cells in cell culture and xenograft models. These results suggest that bruceantin should be reinvestigated for clinical efficacy against multiple myeloma and other hematological malignancies.