Protective effects of Danshensu from the aqueous extract of Salvia miltiorrhiza (Danshen) against homocysteine-induced endothelial dysfunction

Homocysteine (Hcy) is a by-product of methionine metabolism. An imbalance of Hcy in the body may lead to hyperhomocysteinemia, a condition with elevated Hcy concentration in blood that may be one of the risk factors responsible for the development of several vascular diseases (thromboembolism, atherosclerosis, stroke, vascular diseases and dementia). Radix Salvia miltiorrhiza (Danshen), a well-known Chinese medicinal herb that can activate and improve blood microcirculation, is noticeable for its beneficial effect in treating cardiovascular diseases. The present study is to demonstrate the protective effect of Danshen extract against the homocysteine-induced adverse effect on human umbilical vein endothelial cell (HUVEC). Homocysteine (5 mM) not only decreased the cell viability but also caused the disruption of capillary-like structure formation in vitro. The protective effect of Danshen aqueous extract and its active compounds on endothelial cell function were demonstrated through an in vitro tube formation assay, which mimics the new blood vessel formation. To identify the active components in the aqueous extract of Danshen, the content was characterized by instrumental analysis using high performance liquid chromatography with diode array detector (DAD) and electrospray tandem mass spectrometry (ESI-MS/MS). Interestingly, Danshen extract and its pure compounds showed different effectiveness in protecting HUVEC against Hcy-induced injury according to the following descending order: Danshen aqueous extract, 3-(3,4-dihydroxy-phenyl)-2-hydroxy-propionic acid (Danshensu), protocatechuic acid, catechin and protocatechualdehyde. We believed that such findings might provide evidence in understanding the beneficial effects of Danshen on the cardiovascular system.     

Molecular cloning and characterization of FHL2, a novel LIM domain protein preferentially expressed in human heart

A full-length cDNA clone encoding a novel LIM-only protein was isolated and sequenced from a human fetal heart cDNA library. This full-length clone consists of 1416 base pairs and has a predicted open reading frame (ORF) encoding 279 amino acids. The ORF of this polypeptide codes for the human heart-specific

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our and a

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alf

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IM-only protein

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(FHL2). It possesses an extra zinc finger that is a half LIM domain and four repeats of LIM domain. When the human FHL2 cDNA probe was used to hybridize with poly-A RNA of various human tissues, a very strong signal could be seen in heart tissues, and only moderately low signals could be detected in placenta, skeletal muscle and ovary. Virtually no signal could be detected in brain, lung, liver, kidney, pancreas, spleen, thymus, prostate, testis, small intestine, colon or peripheral blood leukocyte. FHL2 was mapped to chromosome 2q12–q13 by fluorescent in-situ hybridization (FISH).     

Cell surface nucleolin is a novel ADAMTS5 receptor mediating endothelial cell apoptosis

A Disintegrin and Metalloproteinase with ThromboSpondin motif (ADAMTS) 5 functions as an anti-angiogenic and anti-cancer protein independent of its metalloproteinase activity. Both full-length ADAMTS5 and TS5-p45, the autocatalytically cleaved C-terminal 45 kDa truncate of ADAMTS5, inhibits angiogenesis, and induces endothelial cell (EC) apoptosis. However, how ADAMTS5 triggers EC apoptosis remains unclear. This work shows that caspase-8 (Cas-8) and caspase-9 (Cas-9) are involved in TS5-p45-induced EC apoptosis. We identify cell surface nucleolin (NCL) as a novel high-affinity receptor for TS5-p45 in ECs, mediating TS5-p45’s cell surface binding and pro-apoptotic function. We show that the central RNA-binding domain (RBD) of NCL is essential and sufficient for its binding to TS5-p45. Upon interacting with EC surface NCL, TS5-p45 is internalized through clathrin- and caveolin-dependent endocytosis and trafficked to the nucleus via late endosomes (LEs). We demonstrate that the nuclear trafficking of TS5-p45 is important for its pro-apoptotic activity as disruption of LE membrane integrity with an endosomolytic peptide suppressed both nuclear trafficking and pro-apoptotic activity of TS5-p45. Through cell surface biotinylation, we revealed that cell surface NCL shuttles extracellular TS5-p45 to the nucleus to mediate apoptosis. Furthermore, blocking the importin α1/ß1 receptor hindered the nuclear trafficking of TS5-p45, suggesting the involvement of the nuclear importing machinery for this nuclear translocation. RNA-seq identified many apoptosis-related genes that are differentially expressed at least two-fold in TS5-p45-treated ECs, with 10 of them qRT-PCR-validated and at least 5 of these genes potentially contributing to TS5-p45-NCL-induced apoptosis. Altogether, our work identifies NCL as a novel cell surface receptor for ADAMTS5 and demonstrates the critical role of NCL-mediated internalization and nuclear trafficking for ADAMTS5-induced EC apoptosis. These findings reveal novel mechanistic insights of the secreted metalloproteinase ADAMTS5 in angiogenesis inhibition.Subject terms: Tumour angiogenesis, Apoptosis, Protein translocation     

A ubiquitin-independent proteasome pathway controls activation of the CARD8 inflammasome

CARD8 is a pattern-recognition receptor that forms a caspase-1-activating inflammasome. CARD8 undergoes constitutive autoproteolysis, generating an N-terminal (NT) fragment with a disordered region and a ZU5 domain and a C-terminal (CT) fragment with UPA and CARD domains. Dipeptidyl peptidase 8 and dipeptidyl peptidase 9 inhibitors, including Val-boroPro, accelerate the degradation of the NT fragment via a poorly characterized proteasome-mediated pathway, thereby releasing the inflammatory CT fragment from autoinhibition. Here, we show that the core 20S proteasome, which degrades disordered and misfolded proteins independent of ubiquitin modification, controls activation of the CARD8 inflammasome. In unstressed cells, we discovered that the 20S proteasome degrades just the NT disordered region, leaving behind the folded ZU5, UPA, and CARD domains to act as an inhibitor of inflammasome assembly. However, in Val-boroPro–stressed cells, we show the 20S proteasome degrades the entire NT fragment, perhaps due to ZU5 domain unfolding, freeing the CT fragment from autoinhibition. Taken together, these results show that the susceptibility of the CARD8 NT domain to 20S proteasome-mediated degradation controls inflammasome activation.     

Adverse events of special interest and mortality following vaccination with mRNA (BNT162b2) and inactivated (CoronaVac) SARS-CoV-2 vaccines in Hong Kong: A retrospective study

BackgroundSafety monitoring of coronavirus disease 2019 (COVID-19) vaccines is crucial during mass vaccination rollout to inform the choice of vaccines and reduce vaccine hesitancy. Considering the scant evidence directly comparing the safety profiles of mRNA and inactivated SARS-CoV-2 vaccines, this territory-wide cohort study aims to compare the incidence of various adverse events of special interest (AESIs) and all-cause mortality between CoronaVac (inactivated vaccine) and BNT162b2 (mRNA-based vaccine). Our results can help vaccine recipients make an informed choice.Methods and findingsA retrospective, population-based cohort of individuals who had received at least 1 dose of BNT162b2 or CoronaVac from 23 February to 9 September 2021 in Hong Kong, and had data linkage to the electronic medical records of the Hong Kong Hospital Authority, were included. Those who had received mixed doses were excluded. Individuals were observed from the date of vaccination (first or second dose) until mortality, second dose vaccination (for first dose analysis), 21 days after vaccination, or 30 September 2021, whichever came first. Baseline characteristics of vaccinated individuals were balanced between groups using propensity score weighting. Outcome events were AESIs and all-cause mortality recorded during 21 days of post-vaccination follow-up after each dose, except anaphylaxis, for which the observation period was restricted to 2 days after each dose. Incidence rate ratios (IRRs) of AESIs and mortality comparing between CoronaVac and BNT162b2 recipients were estimated after each dose using Poisson regression models. Among 2,333,379 vaccinated individuals aged 18 years or above, the first dose analysis included 1,308,820 BNT162b2 and 955,859 CoronaVac recipients, while the second dose analysis included 1,116,677 and 821,560 individuals, respectively. The most frequently reported AESI among CoronaVac and BNT162b2 recipients was thromboembolism (first dose: 431 and 290 per 100,000 person-years; second dose: 385 and 266 per 100,000 person-years). After the first dose, incidence rates of overall AESIs (IRR = 0.98, 95% CI 0.89–1.08, p = 0.703) and mortality (IRR = 0.96, 95% CI 0.63–1.48, p = 0.868) associated with CoronaVac were generally comparable to those for BNT162b2, except for Bell palsy (IRR = 1.95, 95% CI 1.12–3.41, p = 0.018), anaphylaxis (IRR = 0.34, 95% CI 0.14–0.79, p = 0.012), and sleeping disturbance or disorder (IRR = 0.66, 95% CI 0.49–0.89, p = 0.006). After the second dose, incidence rates of overall AESIs (IRR = 0.97, 95% CI 0.87–1.08, p = 0.545) and mortality (IRR = 0.85, 95% CI 0.51–1.40, p = 0.516) were comparable between CoronaVac and BNT162b2 recipients, with no significant differences observed for specific AESIs. The main limitations of this study include residual confounding due to its observational nature, and the possibility of its being underpowered for some AESIs with very low observed incidences.ConclusionsIn this study, we observed that the incidences of AESIs (cumulative incidence rate of 0.06%–0.09%) and mortality following the first and second doses of CoronaVac and BNT162b2 vaccination were very low. The safety profiles of the vaccines were generally comparable, except for a significantly higher incidence rate of Bell palsy, but lower incidence rates of anaphylaxis and sleeping disturbance or disorder, following first dose CoronaVac versus BNT162b2 vaccination. Our results could help inform the choice of inactivated COVID-19 vaccines, mainly administered in low- and middle-income countries with large populations, in comparison to the safety of mRNA vaccines. Long-term surveillance on the safety profile of COVID-19 vaccines should continue.

In a retrospective study, Carlos King Ho Wong, Kristy Tsz Kwan Lau, and colleagues study adverse events reported following COVID-19 vaccination in Hong Kong.     

Microstructural analysis of anaerobic granules

Summary Photos of scanning and transmission electron microscopy showed that sludge granules treating sucrose wastewater had a 20–40 m surface layer with diverse morphology, consisting of cocci and bacilli, and a loosely packed interior, mainly consisting of Methanothrix. Light microscopy using epi-fluorescent excitation showed not only the similar microstructure, but also the distribution of various genuses of methanogens.     

Evidence for major gene inheritance of Alzheimer disease in families of patients with and without apolipoprotein E epsilon 4.

Apolipoprotein E (APOE) genotype is the single most important determinant to the common form of Alzheimer disease (AD) yet identified. Several studies show that family history of AD is not entirely accounted for by APOE genotype. Also, there is evidence for an interaction between APOE genotype and gender. We carried out a complex segregation analysis in 636 nuclear families of consecutively ascertained and rigorously diagnosed probands in the Multi-Institutional Research in Alzheimer Genetic Epidemiology study in order to derive models of disease transmission which account for the influences of APOE genotype of the proband and gender. In the total group of families, models postulating sporadic occurrence, no major gene effect, random environmental transmission, and Mendelian inheritance were rejected. Transmission of AD in families of probands with at least one epsilon 4 allele best fit a dominant model. Moreover, single gene inheritance best explained clustering of the disorder in families of probands lacking epsilon 4, but a more complex genetic model or multiple genetic models may ultimately account for risk in this group of families. Our results also suggest that susceptibility to AD differs between men and women regardless of the proband's APOE status. Assuming a dominant model, AD appears to be completely penetrant in women, whereas only 62%-65% of men with predisposing genotypes develop AD. However, parameter estimates from the arbitrary major gene model suggests that AD is expressed dominantly in women and additively in men. These observations, taken together with epidemiologic data, are consistent with the hypothesis of an interaction between genes and other biological factors affecting disease susceptibility.     

Habituation to acid in Escherichia coli: conditions for habituation and its effects on plasmid transfer.

Induction of acid resistance (habituation) in Escherichia coli at pH 5.0 took ca 5 min in broth at 37 degrees C and 30-60 min in minimal medium. Induction occurred at a range of pH values from 4.0 to 6.0; it was dependent on continuing protein and RNA synthesis but substantial acid resistance appeared in the presence of nalidixic acid. Acid resistance was long-lasting; organisms grown at pH 5.0 retained most of their resistance after 2 h growth at pH 7.0. Organisms grown at pH 5.0 showed increased synthesis of a number of cytoplasmic proteins compared with the level in cells grown at pH 7.0. DNA repair-deficient strains carrying recA, uvrA or polA1 mutations were more acid-sensitive than the repair-proficient parents but were able to habituate at pH 5.0. Organisms grown at pH 5.0 transferred the ColV plasmid much more effectively at acid pH than did those grown at pH 7.0 and habituated recipients appeared better able to repair incoming acid-damaged plasmid DNA than did those that were non-habituated. Induction of acid resistance at pH 5.0 may be significant for the survival of organisms exposed to periodic discharges of acid effluent in the aquatic environment and habituation may also allow plasmid transfer and repair of acid-damaged plasmid DNA during or after such exposure.     

5,2′-dihydroxy-6,7-methylenedioxyisoflavone from seed balls of sugar beet

5,2′-Dihydroxy-6,7-methylenedioxyisoflavone was identified from the seed balls of sugar beet. A neolignan, 6-oxo-2-(4-hydroxy-3,5-dimethoxyphenyl)-3,7-dioxabicyclo-[3.3.0]-octane, and indole-3-carboxylic acid were also isolated.