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Background: Wet age-related macular degeneration (wAMD) is characterized by the presence of choroidal neovascularization (CNV). Although there are some clinical drugs targeting vascular endothelial growth factor (VEGF) and inhibiting CNV, two major side effects limit their application, including the excessive activity of anti-VEGF and frequent intraocular injections. To explore better treatment strategies, researchers developed a hypoxic modulator retinal pigment epithelium (RPE)- specific adeno-associated virus (AAV) vector expressing endostatin to inhibit CNV. However, the mechanism of endostatin is complex. Instead, soluble fms-like tyrosine kinase-1 (sFlt-1) can inhibit VEGF-induced angiogenesis through two simple and clear mechanisms, giving rise to sequestration of VEGF and forming an inactive heterodimer with the membrane-spanning isoforms of the VEGF receptor Flt-1 and kinase insert domain-containing receptor. Objective: In this study, we chose sFlt-1 as a safer substitute to treat wAMD by inhibiting VEGFinduced angiogenesis. Methods: The AAV2/8-Y733F-REG-RPE-sFlt-1 vector was delivered by intravitreal injection to the eyes of mice. AAV2/8-Y733F vector is a mutant of the AAV2/8 vector, and the REG-RPE promoter is a hypoxia-regulated RPE-specific promoter. Two animal models were used to evaluate the function of the vector. Results: In the cobalt chloride-induced hypoxia model, the results demonstrated that the AAV2/8- Y733F-REG-RPE-sFlt-1 vector induced the expression of the sFlt-1 gene in RPE cells through hypoxia. In the laser-induced CNV model, the results demonstrated that the AAV2/8-Y733F-REG-RPE-sFlt- 1 vector reduced laser-induced CNV. Conclusions: Hypoxia regulated, RPE-specific AAV vector-mediated sFlt-1 gene is a hypoxiaregulated antiangiogenic vector for wAMD.

Emodin has been recognized to be an anti‑cancer agent against a number of types of human cancer. It was demonstrated that TNF receptor‑associated factor 6 (TRAF6) was correlated with cancer angiogenesis and metastasis. The present study confirmed the association between TRAF6 and the angiogenesis/metastasis of anaplastic thyroid cancer (ATC). The anti‑angiogenesis and metastatic effects of emodin, in addition to its molecular mechanisms in ATC, were investigated. A total of two ATC cell lines, namely 8505c and SW1736, were studied. ATC cells were implanted into nude mice to form xenografts or to establish lung metastasis models. Emodin was used to incubate ATC cells or to treat animals orally. An MTT assay was used to assess cell proliferation. A wound healing assay was employed to evaluate cell migration. ELISA analysis was used to detect the vascular endothelial growth factor (VEGF) content. Western blotting was used to determine the protein expression levels. In the in vivo study, cancer angiogenesis was assessed by micro vascular density measurement. The lung metastatic rate was the criterion for cancer metastasis. The results of the present study demonstrated that the proliferation of ATC was inhibited by emodin. The activation of the TRAF6/hypoxia inducible factor (HIF)‑1α/VEGF and TRAF6/basigin (CD147)/matrix metalloproteinase‑9 (MMP9) pathways were associated with the angiogenesis and metastasis of ATC. In a concentration‑dependent manner, emodin inhibited the TRAF6/HIF‑1α/VEGF and TRAF6/CD147/MMP9 signaling pathways to suppress angiogenesis and metastasis. In conclusion, emodin exerted anti‑angiogenic and anti‑metastatic activities in ATC by affecting TRAF6‑mediated pathways.

Dysiarenone (1), a dimeric C

Four unusual meroterpenoids, dysivillosins A–D (1–4), were isolated from an organic extract of the marine sponge Dysidea villosa collected from the South China Sea. Their planar structures were determined by 1D and 2D NMR and HRESIMS techniques, while the relative and absolute configurations were elucidated by NOESY experiments and comparison between the calculated and experimental ECD spectra. To the best of our knowledge, dysivillosins A–D are the first examples of terpene-polyketide-pyridine hybrid metabolites from the nature. Anti-allergic activity evaluation showed that compounds 1–4 potently inhibited the release of β-hexosaminidase, a marker of degranulation, in a dose-dependent manner with IC50 values of 8.2–19.9 μM. Additionally, the four meroterpenoids could downregulate the production of lipid mediator leukotrienes B4 (LTB4) and pro-inflammatory cytokine interleukin-4 (IL-4) in the antigen-stimulated RBL-2H3 mast cells. Further biological investigations revealed that dysivillosin A (1) could suppress the phosphorylation of Syk and PLCγ1 in IgE/FcɛRI/Syk signaling pathway, which resulted in the inhibition of degranulation and the downregulation of LTB4 and IL-4 production in mast cells.

(±)-Quassidines I (1) and J (2), two pairs of new bis-β-carboline alkaloid enantiomers, were isolated from the stems of Picrasma quassioides. Their structures were determined by the analysis of spectroscopic data, including HRESIMS and 2D NMR, and confirmed by single-crystal X-ray diffraction analysis. The racemic mixtures of 1 and 2 were resolved into two pairs of enantiomers, (+)-S-1a and (-)-R-1b and (+)-S-2a and (-)-R-2b, by HPLC using a chiral Daicel IB-3 column, respectively, which represents the first successful example to resolve bis-β-carboline racemic mixtures. The absolute configurations of the two pairs of enantiomers were determined by comparison between the calculated and experimental ECD spectra. The cytotoxicity evaluation revealed that (+)-S-1a and (+)-S-2a showed more potent cytotoxicity against human cervical HeLa and gastric MKN-28 cancer cell lines with IC50 values of 4.03-6.30 μM than their enantiomers with IC50 values of 9.64-12.3 μM; however, the two (+)-S-quassidines showed similar activities to their enantiomers against the mouse melanoma B-16 cancer cell line.

An unusual meroterpenoid, dysidinoid A (1), was isolated from the South China Sea sponge Dysidea sp. Its structure was elucidated by extensive spectroscopic methods including HRESIMS and 2D NMR, and its absolute configuration was determined by single-crystal X-ray diffraction analysis. Dysidinoid A (1) is the first meroterpenoid from Nature bearing a 9,4-friedodrime skeleton and a 2,5-dionepyrrole unit. Dysidinoid A (1) showed potent antibacterial activity against two strains of pathogenic bacteria methicillin-resistant Staphylococcus aureus (MRSA) with MIC90 values of 8.0 μg/mL against both.

Four new tetracyclic meroterpnes, dysiherbols A-C (1-3) and dysideanone E (4), were isolated from a Dysidea sp. marine sponge collected from the South China Sea. Their complete structures and absolute configurations were unambiguously determined by a combination of NMR spectroscopic data, ECD calculations, and single-crystal X-ray diffraction analysis. Within the sesquiterpene quinol structures, dysiherbols A-C possess an intriguing 6/6/5/6-fused tetracyclic carbon skeleton. The NF-κB inhibitory and cytotoxic activity evaluation disclosed that dysiherbol A (1) showed potent activity with respective IC50 values of 0.49 and 0.58 μM, which were about 10-fold and 20-fold more potent than those of dysiherbols B (2) and C (3), which feature hydroxy and ketone carbonyl groups at the C-3 position.

To observe the effect of isolating the chondrocytes from articular cartilage with the method of three-step enzymatic digestion, and the biological characteristics of the isolated chondrocytes during cultivation in vitro in order to evaluate their biological activity.The method of three-step enzymatic digestion was designed that the articular cartilage was digested one by one with the 1 g/L trypsin and 1 g/L EDTA, 1 g/L hyaluronidase and 2 g/L collagenase I in the culture medium to isolate chondrocytes. The harvesting and viability rate of the primary chondrocytes were detected. During the passage cultivation in vitro, the changes of the chondrocytes shape and growth were observed, the changes of the collagen type I and II and aggrecan in the extracellular matrix were investigated and detected.(1) The extracellular matrix of articular cartilage was completely dissolved by the three-step enzymatic digestion, and the chondrocytes were completely isolated from the solid matrix. The number of the harvested chondrocytes from every gram of wet cartilage was 50.3 x 10(6) on average, and their viability rate was 98.8% on average. (2) The primary and first passage chondrocytes had triangle or multi-angle shape, and became elliptic shape at the growing confluence with the positive immunohistochemical stain of collagen type II and the strong heterochromia to toluidine blue. The content of sulfate glycosaminoglycans (GAG) in the extracellular matrix of the primary passage cells was (92 +/- 10) microg/cm(2). The chondrocytes after the third passaging gradually became spindle shape with the negative stain of collagen type II and the weak heterochromia to toluidine blue. The content of sulfate GAG of the fourth passage cells was (48 +/- 12) microg/cm(2).(1) The method of three-step enzymatic digestion can make the extracellular matrix of articular cartilage completely degraded, and has advantages in the high efficiency of harvesting primary chondrocytes, high cellular viability rate and simple manipulation. (2) The primary and first passage chondrocytes have fine biological activity, and the chondrocytes after the third passaging have lost their special biological activity.