Sulfated Galactofucan from Sargassum Thunbergii Attenuates Atherosclerosis by Suppressing Inflammation Via the TLR4/MyD88/NF-κB Signaling Pathway.

PURPOSE: Sulfated galactofucan (SWZ-4), which was extracted from Sargassum thunbergii, has recently been reported to show anti-inflammatory and anticancer properties. The present study aimed to evaluate whether SWZ-4 attenuates atherosclerosis in apolipoprotein E-knockout (ApoE-KO) mice by suppressing the inflammatory response through the TLR4/MyD88/NF-κB signaling pathway. METHODS: Male ApoE-KO mice were fed with a high-fat diet for 16 weeks and intraperitoneally injected with SWZ-4. RAW246.7 cells were treated with lipopolysaccharide (LPS) and SWZ-4. Atherosclerotic lesions were measured by Sudan IV and oil red O staining. Serum lipid profiles, inflammatory cytokines, and mRNA and protein expression levels were evaluated. RESULTS: SWZ-4 decreased serum TNF-α, IL-6 and IL-1 levels, but did not reduce blood lipid profiles. SWZ-4 downregulated the mRNA and protein expression of TLR4 and MyD88, reduced the phosphorylation of p65, and attenuated atherosclerosis in the ApoE-KO mice (p < 0.01). In LPS-stimulated RAW 264.7 cells, SWZ-4 inhibited proinflammatory cytokine production and the mRNA expression of TLR4, MyD88, and p65 and reduced the protein expression of TLR4 and MyD88 and the phosphorylation of p65 (p < 0.01). CONCLUSION: These results suggest that SWZ-4 may exert an anti-inflammatory effect on ApoE-KO atherosclerotic mice by inhibiting the TLR4/MyD88/NF-κB signaling pathway in macrophages and therefore may be a treatment for atherosclerosis.

EGCG binds intrinsically disordered N-terminal domain of p53 and disrupts p53-MDM2 interaction.

Epigallocatechin gallate (EGCG) from green tea can induce apoptosis in cancerous cells, but the underlying molecular mechanisms remain poorly understood. Using SPR and NMR, here we report a direct, µM interaction between EGCG and the tumor suppressor p53 (KD = 1.6 ± 1.4 µM), with the disordered N-terminal domain (NTD) identified as the major binding site (KD = 4 ± 2 µM). Large scale atomistic simulations (>100 µs), SAXS and AUC demonstrate that EGCG-NTD interaction is dynamic and EGCG causes the emergence of a subpopulation of compact bound conformations. The EGCG-p53 interaction disrupts p53 interaction with its regulatory E3 ligase MDM2 and inhibits ubiquitination of p53 by MDM2 in an in vitro ubiquitination assay, likely stabilizing p53 for anti-tumor activity. Our work provides insights into the mechanisms for EGCG's anticancer activity and identifies p53 NTD as a target for cancer drug discovery through dynamic interactions with small molecules.

Effective Inhibition of SARS-CoV-2 Entry by Heparin and Enoxaparin Derivatives.

Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has caused a pandemic of historic proportions and continues to spread globally, with enormous consequences to human health. Currently there is no vaccine, effective therapeutic, or prophylactic. As with other betacoronaviruses, attachment and entry of SARS-CoV-2 are mediated by the spike glycoprotein (SGP). In addition to its well-documented interaction with its receptor, human angiotensin-converting enzyme 2 (hACE2), SGP has been found to bind to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we pseudotyped SARS-CoV-2 SGP on a third-generation lentiviral (pLV) vector and tested the impact of various sulfated polysaccharides on transduction efficiency in mammalian cells. The pLV vector pseudotyped SGP efficiently and produced high titers on HEK293T cells. Various sulfated polysaccharides potently neutralized pLV-S pseudotyped virus with clear structure-based differences in antiviral activity and affinity to SGP. Concentration-response curves showed that pLV-S particles were efficiently neutralized by a range of concentrations of unfractionated heparin (UFH), enoxaparin, 6-O-desulfated UFH, and 6-O-desulfated enoxaparin with 50% inhibitory concentrations (IC50s) of 5.99 µg/liter, 1.08 mg/liter, 1.77 µg/liter, and 5.86 mg/liter, respectively. In summary, several sulfated polysaccharides show potent anti-SARS-CoV-2 activity and can be developed for prophylactic as well as therapeutic purposes.IMPORTANCE The emergence of severe acute respiratory syndrome coronavirus (SARS-CoV-2) in Wuhan, China, in late 2019 and its subsequent spread to the rest of the world has created a pandemic situation unprecedented in modern history. While ACE2 has been identified as the viral receptor, cellular polysaccharides have also been implicated in virus entry. The SARS-CoV-2 spike glycoprotein (SGP) binds to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we report structure-based differences in antiviral activity and affinity to SGP for several sulfated polysaccharides, including both well-characterized FDA-approved drugs and novel marine sulfated polysaccharides, which can be developed for prophylactic as well as therapeutic purposes.

Sulfated galactofucan from Sargassum thunbergii induces senescence in human lung cancer A549 cells.

Isolated compounds from Sargassum thunbergii (S. thunbergii) have shown to exhibit diverse biological activities, including anti-cancer activity. In this study, we examined the effect of sulfated galactofucan (SWZ-4-H), which was successfully isolated from S. thunbergii, and its underlying mechanism on human lung cancer (LC) A549 cell growth in vitro and in vivo. In vitro experiment indicated that SWZ-4-H decreased cell growth and number in a dose-dependent manner (P < 0.05 vs. control). Besides, cells treated with SWZ-4-H had irregular morphology, including increased cell volumes, and large nuclei, which suggested senescence-like changes. Moreover, SWZ-4-H increased senescence-related ß-galactosidase (SA-ß-Gal) staining in a dose-dependent manner; however, while lower (1 mg mL-1) concentration induced mainly senescence without causing cell death, higher dosage (3 mg mL-1) induced both senescence and cell death. The effect of SWZ-4-H was further confirmed by analyzing the expression of p53, p21, p16, and Rb (p-RB); SWZ-4-H significantly increased the expression of p53, p21, and p16 and decreased phosphorylated Rb (p-RB) in a dose-dependent manner. Moreover, in vivo experiment showed that SWZ-4-H significantly reduced the tumor volume without affecting the body weight. To sum up, our data indicated that SWZ-4-H could induce lung cancer senescence by regulating p53, p21, p16, and p-Rb, thus providing a novel perspective on anti-cancer mechanisms of SWZ-4-H in human lung cancer A549 cells.

Structure Analysis and Anti-Tumor and Anti-Angiogenic Activities of Sulfated Galactofucan Extracted from Sargassum thunbergii.

Sulfated galactofucan (ST-2) was obtained from Sargassum thunbergii. It was then desulfated to obtain ST-2-DS, and autohydrolyzed and precipitated by ethanol to obtain the supernatant (ST-2-S) and precipitate (ST-2-C). ST-2-C was further fractionated by gel chromatography into two fractions, ST-2-H (high molecular weight) and ST-2-L (low molecular weight). Mass spectrometry (MS) of ST-2-DS was performed to elucidate the backbone of ST-2. It was shown that ST-2-DS contained a backbone of alternating galactopyranose residues (Gal)n (n ≤ 3) and fucopyranose residues (Fuc)n. In addition, ST-2-S was also determined by MS to elucidate the branches of ST-2. It was suggested that sulfated fuco-oligomers might be the branches of ST-2. Compared to the NMR spectra of ST-2-H, the spectra of ST-2-L was more recognizable. It was shown that ST-2-L contain a backbone of (Gal)n and (Fuc)n, sulfated mainly at C4 of Fuc, and interspersed with galactose (the linkages were likely to be 1→2 and 1→6). Therefore, ST-2 might contain a backbone of (Gal)n (n ≤ 3) and (Fuc)n. The sulfation pattern was mainly at C4 of fucopyranose and partially at C4 of galactopyranose, and the branches were mainly sulfated fuco-oligomers. Finally, the anti-tumor and anti-angiogenic activities of ST-2 and its derivates were determined. It was shown that the low molecular-weight sulfated galactofucan, with higher fucose content, had better anti-angiogenic and anti-tumor activities.

A study of neuroprotective and antioxidant activities of heteropolysaccharides from six Sargassum species.

Heteropolysaccharides were extracted from Sargassum integerrimum (S.I), Sargassum maclurei (S.M), Sargassum naozhouense (S.N), Spiraea thunbergii (S.T), Sargassum hemiphyllum (S.H) and Sargassum fusiforme (S.F), and their neuroprotective effects and antioxidant activities were investigated. It showed that S.I, S.N, S.T and S.F exhibited neuroprotective activities, whereas S.H and S.M did not. For this reason, they were separated by anion-exchange chromatography. It was apparent that the fraction 2 represented the principal difference between the active and non-active compounds. However, it did not correlate with neuroprotective effect. In addition, the results on antioxidant activities showed that the hydroxyl-radical scavenging effect contribute to the neuroprotective effect of S.T and S.N, and the DPPH-radical scavenging effect and reducing power contribute to S.T, S.F and S.I. However, the superoxide-radical scavenging effect did not correlate with neuroprotective activity. The conclusion was that the neuroprotective activity of the family of compounds investigated depended on a variety of factors.

Hypoglycemic property of acidic polysaccharide extracted from Saccharina japonica and its potential mechanism.

In the present study, a sulfated polysaccharide fucoidan extracted from Saccharina japonica was administered to normal and alloxan-diabetic rats/mice, and its effects on glycemia, insulin and serum lipid levels were evaluated. Fucoidan administered at 200 or 1200 mg/kg body weight/day could significantly reduce the blood glucose level by 22% and 34%, respectively, in alloxan-induced diabetic rats. Serum insulin levels in diabetic mice were increased by the administration of fucoidan (P<0.05). The results of an oral glucose tolerance test (OGTT) revealed that fucoidan treatment had some effect on glucose disposal after 15 days of treatment. Furthermore, fucoidan altered plasma lipid levels by lowering cholesterol, triglyceride and plasma low-density lipoprotein concentrations, while elevating plasma high-density lipoprotein cholesterol at 100 or 300 mg/kg body weight/day. The results suggested that fucoidan exhibited a considerable hypoglycemic effect, possibly by stimulating pancreatic release of insulin and/or by reducing insulin metabolism. Our results indicated that fucoidan could be developed as a potential oral hypoglycemic agents or functional food for the management of diabetes.

The neuroprotective activities and antioxidant activities of the polysaccharides from Saccharina japonica.

The crude polysaccharide (W) was extracted by water from Saccharina japonica and five fractions were separated by anion-exchange chromatography. And their chemical constituents, neuroprotective activities and antioxidant activities were studied. It showed that W had the neuroprotective activity while its fractions did not. In addition, the fractions displayed higher activities on hydroxyl-radical scavenging effects and reducing power than these of W. Moreover, it was speculated that the neuroprotective activities of samples were related to the hydroxyl-radical scavenging effect and reducing power while did not relate to superoxide-radical scavenging effect. Finally, it was concluded that some fractions could be good candidate antioxidants in food chemistry owing to the high antioxidant activities and their non-toxic characteristics.