Identification of existing pharmaceuticals and herbal medicines as inhibitors of SARS-CoV-2 infection.

The outbreak of COVID-19 caused by SARS-CoV-2 has resulted in more than 50 million confirmed cases and over 1 million deaths worldwide as of November 2020. Currently, there are no effective antivirals approved by the Food and Drug Administration to contain this pandemic except the antiviral agent remdesivir. In addition, the trimeric spike protein on the viral surface is highly glycosylated and almost 200,000 variants with mutations at more than 1,000 positions in its 1,273 amino acid sequence were reported, posing a major challenge in the development of antibodies and vaccines. It is therefore urgently needed to have alternative and timely treatments for the disease. In this study, we used a cell-based infection assay to screen more than 3,000 agents used in humans and animals, including 2,855 small molecules and 190 traditional herbal medicines, and identified 15 active small molecules in concentrations ranging from 0.1 nM to 50 µM. Two enzymatic assays, along with molecular modeling, were then developed to confirm those targeting the virus 3CL protease and the RNA-dependent RNA polymerase. Several water extracts of herbal medicines were active in the cell-based assay and could be further developed as plant-derived anti-SARS-CoV-2 agents. Some of the active compounds identified in the screen were further tested in vivo, and it was found that mefloquine, nelfinavir, and extracts of Ganoderma lucidum (RF3), Perilla frutescens, and Mentha haplocalyx were effective in a challenge study using hamsters as disease model.

Dysregulated lipid metabolism in TMZ-resistant glioblastoma: pathways, proteins, metabolites and therapeutic opportunities.

Glioblastoma (GBM) is a highly aggressive and lethal brain tumor with limited treatment options, such as the chemotherapeutic agent, temozolomide (TMZ). However, many GBM tumors develop resistance to TMZ, which is a major obstacle to effective therapy. Recently, dysregulated lipid metabolism has emerged as an important factor contributing to TMZ resistance in GBM. The dysregulation of lipid metabolism is a hallmark of cancer and alterations in lipid metabolism have been linked to multiple aspects of tumor biology, including proliferation, migration, and resistance to therapy. In this review, we aimed to summarize current knowledge on lipid metabolism in TMZ-resistant GBM, including key metabolites and proteins involved in lipid synthesis, uptake, and utilization, and recent advances in the application of metabolomics to study lipid metabolism in GBM. We also discussed the potential of lipid metabolism as a target for novel therapeutic interventions. Finally, we highlighted the challenges and opportunities associated with developing these interventions for clinical use, and the need for further research to fully understand the role of lipid metabolism in TMZ resistance in GBM. Our review suggests that targeting dysregulated lipid metabolism may be a promising approach to overcome TMZ resistance and improve outcomes in patients with GBM.

Estradiol-mediated inhibition of Sp1 decreases miR-3194-5p expression to enhance CD44 expression during lung cancer progression.

BACKGROUND: Sp1, an important transcription factor, is involved in the progression of various cancers. Our previous studies have indicated that Sp1 levels are increased in the early stage of lung cancer progression but decrease during the late stage, leading to poor prognosis. In addition, estrogen has been shown to be involved in lung cancer progression. According to previous studies, Sp1 can interact with the estrogen receptor (ER) to coregulate gene expression. The role of interaction between Sp1 and ER in lung cancer progression is still unknown and will be clarified in this study. METHODS: The clinical relevance between Sp1 levels and survival rates in young women with lung cancer was studied by immunohistochemistry. We validated the sex dependence of lung cancer progression in EGFRL858R-induced lung cancer mice. Wound healing assays, chamber assays and sphere formation assays in A549 cells, Taxol-induced drug-resistant A549 (A549-T24) and estradiol (E2)-treated A549 (E2-A549) cells were performed to investigate the roles of Taxol and E2 in lung cancer progression. Luciferase reporter assays, immunoblot and q-PCR were performed to evaluate the interaction between Sp1, microRNAs and CD44. Tail vein-injected xenograft experiments were performed to study lung metastasis. Samples obtained from lung cancer patients were used to study the mRNA level of CD44 by q-PCR and the protein levels of Sp1 and CD44 by immunoblot and immunohistochemistry. RESULTS: In this study, we found that Sp1 expression was decreased in premenopausal women with late-stage lung cancer, resulting in a poor prognosis. Tumor formation was more substantial in female EGFRL858R mice than in male mice and ovariectomized female mice, indicating that E2 might be involved in the poor prognosis of lung cancer. We herein report that Sp1 negatively regulates metastasis and cancer stemness in E2-A549 and A549-T24 cells. Furthermore, E2 increases the mRNA and protein levels of RING finger protein 4 (RNF4), which is the E3-ligase of Sp1, and thereby decreases Sp1 levels by promoting Sp1 degradation. Sp1 can be recruited to the promoter of miR-3194-5p, and positively regulate its expression. Furthermore, there was a strong inverse correlation between Sp1 and CD44 levels in clinical lung cancer specimens. Sp1 inhibited CD44 expression by increasing the expression of miR-3194-5p, miR-218-5p, miR-193-5p, miR-182-5p and miR-135-5p, ultimately resulting in lung cancer malignancy. CONCLUSION: Premenopausal women with lung cancer and decreased Sp1 levels have a poor prognosis. E2 increases RNF4 expression to repress Sp1 levels in premenopausal women with lung cancer, thus decreasing the expression of several miRNAs that can target CD44 and ultimately leading to cancer malignancy.

Restraint of FAM60A has a cancer-inhibiting role in pancreatic carcinoma via the effects on the Akt/GSK-3ß/ß-catenin signaling pathway.

Family with sequence similarity 60A (FAM60A) has been reported as a new cancer-related protein that affects the malignant progression of some cancers. However, whether FAM60A plays a part in pancreatic carcinoma is undetermined. This work was designed to examine the impact of FAM60A in pancreatic carcinoma. Abundant expression of FAM60A was observed in the primary tumor tissue of pancreatic carcinoma. Moreover, a high FAM60A level was related to a poor overall survival in pancreatic carcinoma patients. Malignant behaviors of pancreatic carcinoma cells, such as proliferation and invasiveness, were markedly affected by FAM60A depletion. In addition, FAM60A depletion enhanced the drug sensitivity of pancreatic carcinoma cells to gemcitabine. Further study revealed that FAM60A depletion impaired the activities of Akt and ß-catenin. Inhibiting the activity of Akt abolished FAM60A-mediated ß-catenin activation. Re-expression of ß-catenin partially diminished the FAM60A-depletion-mediated cancer suppressive effect in pancreatic carcinoma cells. In vivo experiments demonstrated that FAM60A depletion prohibited the xenograft formation of pancreatic carcinoma cells, with concurrent reductions of Akt and ß-catenin activities. Collectively, our findings indicate that FAM60A exerts a cancer-promoting role in pancreatic carcinoma through affection of the Akt/ß-catenin pathway. This work indicates that FAM60A acts as a tumor promoter in pancreatic carcinoma and can be utilized as a potential target for anti-pancreatic carcinoma therapy development.

Molecular detection and subtype distribution of Blastocystis in farmed pigs in southern China.

Blastocystis is one of the most common causative agents of intestinal diseases, which can cause enteric diseases in animals and humans. However, limited data is available on the prevalence or subtypes of Blastocystis infections in farmed pigs in southern China. In this study, a total of 396 fecal samples were collected from farmed pigs in three provinces in southern China in 2016, and screened for Blastocystis by PCR amplification of the small subunit rRNA (SSU rRNA) gene fragment. One hundred and seventy (42.93%) of the examined fecal samples were detected Blastocystis-positive, and two known zoonotic subtypes ST1 and ST5 were identified, with ST5 being the predominate subtype. Moreover, gender, age and region were considered as risk factors that associated with Blastocystis infection in farmed pigs. The present study revealed the prevalence and subtypes of Blastocystis infections in farmed pigs in southern China, which provided essential data for the control of Blastocystis infections in pigs, other animals and humans in China.

Upregulation of LASP2 inhibits pancreatic cancer cell migration and invasion through suppressing TGF-ß-induced EMT.

LASP2 (LIM and SH3 protein 2), a member of the LIM-protein subfamily of the nebulin group, was first identified as a splice variant of the nebulin gene. In the past, investigators mainly focused on the impact of LASP2 on cardiac diseases because of its identification in the myocardium. Recently, several studies have reported that LASP2 is associated with the progression of various cancers. However, there have been no investigations on the expression and function of LASP2 in pancreatic cancer (PC). In this study, we performed the quantitative real-time polymerase chain reaction and Western blot analysis to detect the expression of LASP2 in PC tissues and cell lines. PC cells were transfected with LASP2 overexpression plasmid or the negative control in the presence or absence of tumor growth factor-ß (TGF-ß). The transwell assays were used to measure the effects of LASP2 on PC cell migration and invasion. The protein expression of epithelial-mesenchymal transition (EMT) markers was detected using Western blot assay. Our results demonstrated that LASP2 was downregulated in PC tissues and cell lines. In addition, upregulation of LASP2 inhibited the PC cell migration and invasion. We also found that LASP2 upregulation reversed TGF-ß-induced EMT in PC cells. Taken together, we provided novel evidence supporting the tumor-suppressor role of LASP2 in PC and suggested it as a potential therapeutic target in PC treatment.

Enrichment of superoxide dismutase 2 in glioblastoma confers to acquisition of temozolomide resistance that is associated with tumor-initiating cell subsets.

BACKGROUND: Intratumor subsets with tumor-initiating features in glioblastoma are likely to survive treatment. Our goal is to identify the key factor in the process by which cells develop temozolomide (TMZ) resistance. METHODS: Resistant cell lines derived from U87MG and A172 were established through long-term co-incubation of TMZ. Primary tumors obtained from patients were maintained as patient-derived xenograft for studies of tumor-initating cell (TIC) features. The cell manifestations were assessed in the gene modulated cells for relevance to drug resistance. RESULTS: Among the mitochondria-related genes in the gene expression databases, superoxide dismutase 2 (SOD2) was a significant factor in resistance and patient survival. SOD2 in the resistant cells functionally determined the cell fate by limiting TMZ-stimulated superoxide reaction and cleavage of caspase-3. Genetic inhibition of the protein led to retrieval of drug effect in mouse study. SOD2 was also associated with the TIC features, which enriched in the resistant cells. The CD133+ specific subsets in the resistant cells exhibited superior superoxide regulation and the SOD2-related caspase-3 reaction. Experiments applying SOD2 modulation showed a positive correlation between the TIC features and the protein expression. Finally, co-treatment with TMZ and the SOD inhibitor sodium diethyldithiocarbamate trihydrate in xenograft mouse models with the TMZ-resistant primary tumor resulted in lower tumor proliferation, longer survival, and less CD133, Bmi-1, and SOD2 expression. CONCLUSION: SOD2 plays crucial roles in the tumor-initiating features that are related to TMZ resistance. Inhibition of the protein is a potential therapeutic strategy that can be used to enhance the effects of chemotherapy.

Stage-specific embryonic antigen-3 (SSEA-3) and ß3GalT5 are cancer specific and significant markers for breast cancer stem cells.

The discovery of cancer stem cells (CSCs), which are responsible for self-renewal and tumor growth in heterogeneous cancer tissues, has stimulated interests in developing new cancer therapies and early diagnosis. However, the markers currently used for isolation of CSCs are often not selective enough to enrich CSCs for the study of this special cell population. Here we show that the breast CSCs isolated with CD44(+)CD24(-/lo)SSEA-3(+) or ESA(hi)PROCR(hi)SSEA-3(+) markers had higher tumorigenicity than those with conventional markers in vitro and in vivo. As few as 10 cells with CD44(+)CD24(-/lo)SSEA-3(+) formed tumor in mice, compared with more than 100 cells with CD44(+)CD24(-/lo). Suppression of SSEA-3 expression by knockdown of the gene encoding ß-1,3-galactosyltransferase 5 (ß3GalT5) in the globo-series pathway, led to apoptosis in cancer cells specifically but had no effect on normal cells. This finding is further supported by the analysis of SSEA-3 and the two related globo-series epitopes SSEA4 and globo-H in stem cells (embryonic stem cells and induced pluripotent stem cells) and various normal and cancer cells, and by the antibody approach to target the globo-series glycans and the late-stage clinical trials of a breast cancer vaccine.

ANGPTL4 Induces TMZ Resistance of Glioblastoma by Promoting Cancer Stemness Enrichment via the EGFR/AKT/4E-BP1 Cascade.

Glioblastoma (GBM) is the most aggressive type of brain tumor, with strong invasiveness and a high tolerance to chemotherapy. Despite the current standard treatment combining temozolomide (TMZ) and radiotherapy, glioblastoma can be incurable due to drug resistance. The existence of glioma stem-like cells (GSCs) is considered the major reason for drug resistance. However, the mechanism of GSC enrichment remains unclear. Herein, we found that the expression and secretion of angiopoietin-like 4 protein (ANGPTL4) were clearly increased in GSCs. The overexpression of ANGPTL4 induced GSC enrichment that was characterized by polycomb complex protein BMI-1 and SRY (sex determining region Y)-box 2 (SOX2) expression, resulting in TMZ resistance in GBM. Furthermore, epidermal growth factor receptor (EGFR) phosphorylation induced 4E-BP1 phosphorylation that was required for ANGPTL4-induced GSC enrichment. In particular, ANGPTL4 induced 4E-BP1 phosphorylation by activating phosphoinositide 3-kinase (PI3K)/AKT and extracellular signal-regulated kinase (ERK) cascades for inducing stemness. To elucidate the mechanism contributing to ANGPTL4 upregulation in GSCs, chromatin immunoprecipitation coupled with sequencing (ChIP-Seq) revealed that specificity protein 4 (Sp4) was associated with the promoter region, -979 to -606, and the luciferase reporter assay revealed that Sp4 positively regulated activity of the ANGPTL4 promoter. Moreover, both ANGPTL4 and Sp4 were highly expressed in GBM and resulted in a poor prognosis. Taken together, Sp4-mediated ANGPTL4 upregulation induces GSC enrichment through the EGFR/AKT/4E-BP1 cascade.

Live Attenuated Pru:Δcdpk2 Strain of Toxoplasma gondii Protects Against Acute, Chronic, and Congenital Toxoplasmosis.

Background: The threat of Toxoplasma gondii infection in immunocompromised individuals and pregnant women necessitates the development of a safe and effective vaccine. Here, we examined the immune protection conferred by a live attenuated strain of T. gondii. Methods: We tested the efficacy of intraperitoneal vaccination using 500 Ca2+-dependent protein kinase 2 (cdpk2)-deficient tachyzoites of T. gondii Pru strain against acute, chronic, and congenital toxoplasmosis in mice. The kinetics of antibody response, cytokines, and other quantifiable correlates of protection against T. gondii infection were determined. Results: Vaccination with Pru:Δcdpk2 induced a high level of anti-T. gondii immunoglobulin G titer, type 1 T-helper (Th1) response at 28 days postvaccination, and a mixed Th1/type 2 T-helper response at 70 days postvaccination. All vaccinated mice survived a heterologous challenge with 1000 tachyzoites of RH or ToxoDB#9 (PYS or TgC7) strains. Also, vaccination protected against homologous infection with 20 T. gondii Pru cysts, and improved pregnancy outcome by reducing parasite cyst load in the brain, maintaining litter size and body weight of pups born to vaccinated dams challenged with 10 Pru cysts compared to pups born to unvaccinated dams. Conclusions: The use of T. gondii Pru:Δcdpk2 mutant strain represents a promising approach to protection against acute, chronic, and congenital toxoplasmosis in mice.

Feasibility and effectiveness of percutaneous balloon mitral valvuloplasty under echocardiographic guidance only.

OBJECTIVE: Percutaneous balloon mitral valvuloplasty (PBMV) is the treatment of choice in patients with isolated mitral stenosis. This study aimed to assess the feasibility of PBMV under echocardiography guidance only of isolated mitral stenosis (MS). METHODS: From October 2016 to Dec 2017, 20 consecutive patients with severe MS underwent PBMV with echocardiography as the only imaging modality at a single center. Outpatient follow-up including chest radiography, electrocardiography, and transthoracic echocardiography was conducted at 1, 3,6, and 12 months after the procedure. RESULTS: All 20 patients successfully underwent PBMV under echocardiography guidance without radiation and contrast agent. Among them, 2 patients were pregnant, 5 had chronic renal failure, and 1 had history of allergy to contrast. Mitral transvalvular pressure gradient measured at catheterization dropped from 13.35 ± 2.85 mm Hg to 5.10 ± 1.17 mm Hg (P < .01). Mitral valve area increased from 0.82 ± 0.10 cm2 pre-PBMV to 1.88 ± 0.24 cm2 post-PBMV (P < .01). Mean balloon diameter was 26.63 ± 0.93 mm. Mild mitral regurgitation developed in 6 patients. Mean follow-up duration was 6.27 ± 3.12 months. At last follow-up, mitral valve area remained high (1.71 ± 0.14 cm2 ) and mean transmitral pressure gradient low (6.07 ± 1.03 mm Hg). No pericardial effusion or peripheral vascular complications occurred. CONCLUSION: In this small experience, PBMV could be successfully performed under only echocardiography guidance and appeared safe and effective while avoiding radiation and contrast agent use.

Transcriptomic analysis reveals Toxoplasma gondii strain-specific differences in host cell response to dense granule protein GRA15.

Growth and replication of the protozoan parasite Toxoplasma gondii within host cell entail the production of several effector proteins, which the parasite exploits for counteracting the host's immune response. Despite considerable research to define the host signaling pathways manipulated by T. gondii and their effectors, there has been limited progress into understanding how individual members of the dense granule proteins (GRAs) modulate gene expression within host cells. The aim of this study was to evaluate whether T. gondii GRA15 protein plays any role in regulating host gene expression. Baby hamster kidney cells (BHK-21) were transfected with plasmids encoding GRA15 genes of either type I GT1 strain (GRA15I) or type II PRU strain (GRA15II). Gene expression patterns of transfected and nontransfected BHK-21 cells were investigated using RNA-sequencing analysis. GRA15I and GRA15II induced both known and novel transcriptional changes in the transfected BHK-21 cells compared with nontransfected cells. Pathway analysis revealed that GRA15II was mainly involved in the regulation of tumor necrosis factor (TNF), NF-κB, HTLV-I infection, and NOD-like receptor signaling pathways. GRA15I preferentially influenced the synthesis of unsaturated fatty acids in host cells. Our findings support the hypothesis that certain functions of GRA15 protein are strain dependent and that GRA15 modulates the expression of signaling pathways and genes with important roles in T. gondii pathophysiology. A greater understanding of host signaling pathways influenced by T. gondii effectors would allow the development of more efficient anti-T. gondii therapeutic schemes, capitalizing on disrupting parasite virulence factors to advance the treatment of toxoplasmosis.

Identification of two novel host proteins interacting with Toxoplasma gondii 14-3-3 protein by yeast two-hybrid system.

Toxoplasma gondii deploys many effector proteins in order to hijack and manipulate host cell signaling pathways, allowing parasite colonization, subversion of immune responses, and disease progression. T. gondii effector protein 14-3-3 (Tg14-3-3) promotes parasite dissemination inside the body, by enhancing the migratory ability of infected microglia and dendritic cells. Understanding both the mechanism of action and the host targets of Tg14-3-3 effector is important because of their importance to the parasite's virulence. The aim of the present study was to explore the function of Tg14-3-3 by utilizing the yeast two-hybrid system (Y2HS) to identify novel Tg14-3-3 interactors/substrates in host cells. A human cDNA library was screened using Tg14-3-3 as the bait. Tg14-3-3 (RH strain, Type I) was cloned into the pGBKT7 vector and expressed in the Y2HGold yeast strain. The bait protein expression was validated by Western blotting analysis, auto-activation, and toxicity investigation compared with control (Y2HGold yeast strain transformed with empty pGBKT7 vector). Two positive Tg14-3-3 interactors identified by this screening, hCG1821272 and eIF5B (eukaryotic translation initiation factor 5B), were isolated and characterized. This approach made it possible to gain a better understanding of the function of Tg14-3-3 in regulating host proteins involved in key cellular processes, such as translational initiation and cell migration.

Body mass index is inversely associated with arterial stiffness in Chinese adults with primary hypertension: results from the China Stroke Primary Prevention Trial (CSPPT).

BACKGROUND: This study aimed to elucidate the relationship between body mass index (BMI) and the presence of arterial stiffness in rural-dwelling Chinese adults with primary hypertension. METHODS: Primary hypertension patients (n = 19,375) receiving an average of 4.5 years of antihypertension therapy were selected from the Chinese Stroke Primary Prevention Trial (mean age: 64.7 ± 7.4 years, male: 37.8%). Anthropometric, demographic, hemodynamic, and biochemical data were obtained. Arterial stiffness was assessed using brachial-ankle pulse wave velocity (baPWV). RESULTS: BMI was inversely associated with baPWV after adjusting for gender, age, smoking, alcohol consumption, center, pulse, SBP, DBP, FBG, TC, TG, HDL-C, BUN, Scr, UA, HCY, antidiabetes treatment, lipid-lowing treatment, and antihypertensive treatment (ß (SE): -10.72 (0.69), P < 0.0001). Quintile1 (Q1) was used as a reference; Q2, Q3, Q4, and Q5 groups were all inversely associated with baPWV. The ß increased with increasing BMI, ß (SE) was -20.29 (6.74), -30.66 (7.01), -51.82 (7.27), and -103.1 (7.62), respectively, for Q2 - Q5, P < 0.05. BMI remained inversely correlated with baPWV across differences in gender, center, blood pressure, lipid levels, and the presence or absence of diabetes subgroups. CONCLUSION: Increased BMI is a positive factor against the development of arterial stiffness in Chinese rural-dwelling adults with primary hypertension undergoing antihypertension treatments, after adjusting for confounding factors.

Human CLEC18 Gene Cluster Contains C-type Lectins with Differential Glycan-binding Specificity.

The human C-type lectin 18 (clec18) gene cluster, which contains three clec18a, clec18b, and clec18c loci, is located in human chromosome 16q22. Although the amino acid sequences of CLEC18A, CLEC18B, and CLEC18C are almost identical, several amino acid residues located in the C-type lectin-like domain (CTLD) and the sperm-coating protein/Tpx-1/Ag5/PR-1/Sc7 (SCP/TAPS) domain, also known as the cysteine-rich secretory proteins/antigen 5/pathogenesis-related 1 proteins (CAP) domain, are distinct from each other. Genotyping by real-time PCR and sequencing further shows the presence of multiple alleles in clec18a/b/c loci. Flow cytometry analysis demonstrates that CLEC18 (CLEC18A, -B, and -C) are expressed abundantly in human peripheral blood cells. Moreover, CLEC18 expression is further up-regulated when monocytes differentiate into macrophages and dendritic cells. Immunofluorescence staining reveals that CLEC18 are localized in the endoplasmic reticulum, Golgi apparatus, and endosome. Interestingly, CLEC18 are also detectable in human sera and culture supernatants from primary cells and 293T cells overexpressing CLEC18. Moreover, CLEC18 bind polysaccharide in Ca(2+)-independent manner, and amino acid residues Ser/Arg(339) and Asp/Asn(421) in CTLD domain contribute to their differential binding abilities to polysaccharides isolated from Ganoderma lucidum (GLPS-F3). The Ser(339) (CLEC18A) → Arg(339) (CLEC18A-1) mutation completely abolishes CLEC18A-1 binding to GLPS-F3, and a sugar competition assay shows that CLEC18 preferentially binds to fucoidan, ß-glucans, and galactans. Because proteins with the SCP/TAPS/CAP domain are able to bind sterol and acidic glycolipid, and are involved in sterol transport and ß-amyloid aggregation, it would be interesting to investigate whether CLEC18 modulates host immunity via binding to glycolipids, and are also involved in glycolipid transportation and protein aggregation in the future.

Novel synthetic benzimidazole-derived oligosaccharide, M3BIM, prevents ex vivo platelet aggregation and in vivo thromboembolism.

BACKGROUND: Thrombus formation, a phenomenon primarily related to increased platelet activation, plays a key role in cardiovascular and cerebrovascular diseases. Although the established antiplatelet agents, such as aspirin and clopidogrel, have been shown to be beneficial in treating thromboembolic diseases, they have considerable limitations. Hence, the development of more effective and safe antithrombotic agents is necessary to satisfy a substantial unmet clinical need. In recent years, the favorable properties of imidazole-related drugs have prompted medicinal chemists to synthesize numerous novel therapeutic agents. The chemical structure of the benzimidazole backbone has proven antiplatelet properties. Moreover, synthetic oligosaccharides have exhibited antiplatelet properties. Therefore, we developed a new aldo-benzimidazole-derived oligosaccharide compound, M3BIM, for achieving a stronger antiplatelet effect than the drugs which are being used in clinical aspects. We investigated the effects of M3BIM on platelet activation ex vivo and its antithrombotic activity in vivo. RESULTS: M3BIM (10-50 µM) exhibited a more potent activity in inhibiting platelet aggregation stimulated by collagen than it did in inhibiting that stimulated by thrombin in washed human platelets. The M3BIM treatment revealed no cytotoxicity in zebrafish embryos, even at the highest concentration of 100 µM. In addition, M3BIM inhibited the phosphorylation of phospholipase Cγ2, protein kinase C (PKC), and mitogen-activated protein kinases (MAPKs; extracellular signal-regulated kinase 2 and c-Jun N-terminal kinase 1), and markedly reduced the ATP-release reaction and intracellular calcium mobilization in collagen-activated platelets. By contrast, M3BIM showed no effects on either collagen-induced p38 MAPK and Akt phosphorylation or phorbol 12, 13-dibutyrate-induced PKC activation and platelet aggregation. Moreover, the M3BIM treatment substantially prolonged the closure time in human whole blood, and increased the occlusion time in mesenteric microvessels and attenuated cerebral infarction in mice. For the study of anticoagulant activities, M3BIM showed no significant effects in the prolongation of activated partial thromboplastin time and prothrombin time in mice. CONCLUSION: The findings of our study suggest that M3BIM is a potential therapeutic agent for preventing or treating thromboembolic disorders.

Immunization of fucose-containing polysaccharides from Reishi mushroom induces antibodies to tumor-associated Globo H-series epitopes.

Carbohydrate-based vaccines have shown therapeutic efficacy for infectious disease and cancer. The mushroom Ganoderma lucidum (Reishi) containing complex polysaccharides has been used as antitumor supplement, but the mechanism of immune response has rarely been studied. Here, we show that the mice immunized with a l-fucose (Fuc)-enriched Reishi polysaccharide fraction (designated as FMS) induce antibodies against murine Lewis lung carcinoma cells, with increased antibody-mediated cytotoxicity and reduced production of tumor-associated inflammatory mediators (in particular, monocyte chemoattractant protein-1). The mice showed a significant increase in the peritoneal B1 B-cell population, suggesting FMS-mediated anti-glycan IgM production. Furthermore, the glycan microarray analysis of FMS-induced antisera displayed a high specificity toward tumor-associated glycans, with the antigenic structure located in the nonreducing termini (i.e., Fucα1-2Galß1-3GalNAc-R, where Gal, GalNAc, and R represent, respectively, D-galactose, D-N-acetyl galactosamine, and reducing end), typically found in Globo H and related tumor antigens. The composition of FMS contains mainly the backbone of 1,4-mannan and 1,6-α-galactan and through the Fucα1-2Gal, Fucα1-3/4Man, Fucα1-4Xyl, and Fucα1-2Fuc linkages (where Man and Xyl represent d-mannose and d-xylose, respectively), underlying the molecular basis of the FMS-induced IgM antibodies against tumor-specific glycans.

Nucleolin enhances internal ribosomal entry site (IRES)-mediated translation of Sp1 in tumorigenesis.

Our previous study indicated that specificity protein-1 (Sp1) is accumulated during hypoxia in an internal ribosomal entry site (IRES)-dependent manner. Herein, we found that the Sp1 was induced strongly at the protein level, but not in the mRNA level, in lung tumor tissue, indicating that translational regulation might contribute to the Sp1 accumulation during tumorigenesis. A further study showed that the translation of Sp1 was dramatically induced through an IRES-dependent pathway. RNA immunoprecipitation analysis of proteins bound to the 5'-untranslated region (5'-UTR) of Sp1 identified interacting protein - nucleolin. Knockdown of nucleolin significantly inhibited IRES-mediated translation of Sp1, suggesting that nucleolin positively facilitates Sp1 IRES activation. Further analysis of the interaction between nucleolin and the 5'-UTR of Sp1 mRNA revealed that the GAR domain was important for IRES-mediated translation of Sp1. Moreover, gefitinib, and LY294002 and MK2206 compounds inhibited IRES-mediated Sp1 translation, implying that activation of the epithelial growth factor receptor (EGFR) pathway via Akt activation triggers the IRES pathway. In conclusion, EGFR activation-mediated nucleolin phosphorylated at Thr641 and Thr707 was recruited to the 5'-UTR of Sp1 as an IRES trans-acting factor to modulate Sp1 translation during lung cancer formation.

Percutaneous Closure of Atrial Septal Defects Under Transthoracic Echocardiography Guidance Without Fluoroscopy or Intubation in Children.

OBJECTIVE: Demonstrate the benefits of percutaneous atrial septal defect (ASD) closure under guidance of transthoracic echocardiography (TTE) without fluoroscopy. METHODS: From February 2013 to April 2014, 127 consecutive patients with an isolated type II ASD were recruited to undergo percutaneous closure under either TTE (n = 60, TTE group) or TEE (n = 67, TEE group) guidance. The TTE group received local anesthesia or sedation with propofol, and the TEE group received general anesthesia with endotracheal intubation. Follow-up examinations were performed for both groups at 1 month, 3 months, 6 months, and 1 year after discharge and annually thereafter. RESULTS: The TTE group had a significantly shorter procedure time and respirator ventilation duration than the TEE group. The dose of propofol required, the cost, and the pharyngeal complication rate were significantly lower in the TTE group than in the TEE group. The median follow-up of 11.6 months was uneventful in all patients. CONCLUSIONS: Percutaneous ASD closure with TTE guidance as the only imaging tool avoids fluoroscopy, endotracheal intubation, and probe insertion and is associated with a satisfactory procedural success rate and lower costs. This procedure is a safe and reliable treatment for ASD.

Enantioselective inclusion of alcohols by solvent-controlled assembled flexible metal-organic frameworks.

Through judicious choice of the ligands and patient regulation of the solvent conditions, three unique chiral coordination polymeric isomers have been synthesized. Their structures, gas adsorption, and structural interconversion have been studied. One of the isomers displays dynamic behavior, and its use in the enantioselective separation of chiral alcohols has also been reported.