Evaluation of right atrial function by two-dimensional echocardiography and strain imaging in patients with RCA CTO recanalization.

OBJECTIVES: The right heart is mainly supplied with blood by the right coronary artery (RCA). The impact of RCA chronic total occlusion (CTO) on the function of the right heart [right atrium (RA) and ventricle (RV)] and whether successful recanalization of a RCA CTO improves the function of the right heart is not clearly understood yet. We aimed to evaluate right atrial function after recanalization of the RCA using transthoracic echocardiography with additional strain imaging. METHODS AND RESULTS: Fifty-five patients undergoing RCA CTO recanalization at the University Medical Center of Mainz were included in the study. Right atrial strain was assessed before and 6 months after successful CTO revascularization. The median age of the total collective was 66 (50-90) years. We did not find difference in our analysis of RA Volume (p 0.086), RA area (p 0.093), RA major dimension (p 0.32) and RA minor dimension (p 0.139) at baseline and follow-up. Mean RA reservoir strain at baseline was 30.9% (21.1-43.0) vs. 33.4% (20.7-47.7) at follow up (p < 0.001). Mean RA conduit strain was - 17.5% (- 10.7-(- 29.7)) at baseline vs. - 18.2% (- 9.6-(- 31.7)) at follow-up (p = 0.346). Mean RA contraction strain was - 12.9% (- 8.0- (- 21.3)) at baseline vs. - 15.5% (- 8.7-(- 26.6)) at follow-up (p < 0.001). CONCLUSION: Right atrial function was altered in patients with RCA CTO. Successful revascularisation of an RCA CTO improved RA function assessed by strain imaging at follow-up.

Ebony plays an important role in egg hatching and 30k protein expression of silkworm (Bombyx mori).

QiufengN is a silkworm strain. During the feeding process of QiufengN, a mutant of black pupal cuticle QiufengNBP was found. Some silkworm pupae of the mutant were unable to easily molt during pupation, and some silkworm eggs produced by developed normally but larvae were unable to break out of the eggshells. These phenomena had not been observed in other black pupa mutants. Genetic analysis showed that the melanization trait of QiufengNBP is controlled by a recessive gene located on the autosome and follows Mendelian inheritance. Results of positional cloning and qRT-PCR showed that the occurrence of black pupae was caused by the mutation of the ebony gene on chromosome 26. 2-DE analysis of the pupal cuticle of QiufengN and QiufengNBP found that the 30K protein, the main storage protein for the growth and development of silkworms and an important energy substance for embryonic development, has changed significantly. In addition, the expression level of Bombyx mori hatching enzyme (BmHEL), which can soften the eggshell during the hatching process of silkworm, was significantly higher in the eggs of black pupae before and after hatching than in normal eggs. The mutation of ebony makes hatching difficult for silkworms, and increases in BmHEL is needed to soften the eggshell. This study showed that ebony may have important effects on the formation of silkworm pigment and egg hatching, and its formation mechanism is complex and deserves further study.

Enhanced soluble expression of active recombinant human interleukin-29 using champion pET SUMO system.

Current research focuses on the soluble and high-level expression of biologically active recombinant human IL-29 protein in Escherichia coli. The codon-optimized IL-29 gene was cloned into the Champion™ pET SUMO expression system downstream of the SUMO tag under the influence of the T7 lac promoter. The expression of SUMO-fused IL-29 protein was compared in E. coli Rosetta 2(DE3), Rosetta 2(DE3) pLysS, and Rosetta-gami 2(DE3). The release of the SUMO fusion partner resulted in approximately 98 mg of native rhIL-29 protein with a purity of 99% from 1 l of fermentation culture. Purified rhIL-29 was found to be biologically active, as evaluated by its anti-proliferation assay. It was found that Champion™ pET SUMO expression system can be used to obtained high yield of biologically active soluble recombinant human protein compared to other expression vector.

Proteomics Reveals How the Tardigrade Damage Suppressor Protein Teaches Transfected Human Cells to Survive UV-C Stress.

The genome sequencing of the tardigrade Ramazzottius varieornatus revealed a unique nucleosome-binding protein named damage suppressor (Dsup), which was discovered to be crucial for the extraordinary abilities of tardigrades in surviving extreme stresses, such as UV. Evidence in Dsup-transfected human cells suggests that Dsup mediates an overall response in DNA damage signaling, DNA repair, and cell cycle regulation, resulting in an acquired resistance to stress. Given these promising outcomes, our study attempts to provide a wider comprehension of the molecular mechanisms modulated by Dsup in human cells and to explore the Dsup-activated molecular pathways under stress. We performed a differential proteomic analysis of Dsup-transfected and control human cells under basal conditions and at 24 h recovery after exposure to UV-C. We demonstrate via enrichment and network analyses, for the first time, that even in the absence of external stimuli, and more significantly, after stress, Dsup activates mechanisms involved with the unfolded protein response, the mRNA processing and stability, cytoplasmic stress granules, the DNA damage response, and the telomere maintenance. In conclusion, our results shed new light on Dsup-mediated protective mechanisms and increases our knowledge of the molecular machineries of extraordinary protection against UV-C stress.

Identification of major and cross-reactive allergens of local freshwater snail (Pila polita) and the impact of thermal and non-thermal food processing on allergen stability.

BACKGROUND: Snail allergy is rare but can be fatal. Pila polita, a freshwater snail, was considered as a popular exotic food, particularly in tropical countries, and consumed in processed forms. Thus, the purpose of this study was to identify the major and cross-reactive allergens of P. polita and to determine the impact of food processing on the allergen stability. RESULTS: Sodium dodecyl sulfate-polyacrylamide gel electrophoresis fractionated raw snail extract to approximately 24 protein bands, between 9 and 245 kDa. The prominent band at 33 kDa was detected in all raw and processed snail extracts. Immunoblotting tests of the raw extract demonstrated 19 immunoglobulin E (IgE)-binding proteins, and four of them, at 30, 35, 42 and 49 kDa, were revealed as the major IgE-binding proteins of P. polita. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identified the 49 and 42 kDa major allergens as actin, whereas the 30 and 35 kDa major allergens were identified as tropomyosin. Immunoblotting revealed that the raw snail had more allergenic proteins than the processed snail. The degree of allergenicity in decreasing order was raw > brine pickled> boiled > roasted > fried > vinegar pickled. The presence of cross-reactivity between P. polita and the shellfish tested was exhibited with either no, complete, or partial inhibitions. CONCLUSION: Actin and tropomyosin were identified as the major and cross-reactive allergens of P. polita among local patients with snail allergy. Those major allergens are highly stable to high temperatures, acidic pH, and high salt, which might played a crucial role in snail allergy in Malaysia. © 2023 Society of Chemical Industry.

Yeast Secretes High Amounts of Human Calreticulin without Cellular Stress.

The ER chaperone calreticulin (CALR) also has extracellular functions and can exit the mammalian cell in response to various factors, although the mechanism by which this takes place is unknown. The yeast Saccharomyces cerevisiae efficiently secretes human CALR, and the analysis of this process in yeast could help to clarify how it gets out of eukaryotic cells. We have achieved a secretion titer of about 140 mg/L CALR in our S. cerevisiae system. Here, we present a comparative quantitative whole proteome study in CALR-secreting yeast using non-equilibrium pH gradient electrophoresis (NEPHGE)-based two-dimensional gel electrophoresis (2DE) as well as liquid chromatography mass spectrometry in data-independent analysis mode (LC-MSE). A reconstructed carrier ampholyte (CA) composition of NEPHGE-based first-dimension separation for 2DE could be used instead of formerly commercially available gels. Using LC-MSE, we identified 1574 proteins, 20 of which exhibited differential expression. The largest group of differentially expressed proteins were structural ribosomal proteins involved in translation. Interestingly, we did not find any signs of cellular stress which is usually observed in recombinant protein-producing yeast, and we did not identify any secretory pathway proteins that exhibited changes in expression. Taken together, high-level secretion of human recombinant CALR protein in S. cerevisiae does not induce cellular stress and does not burden the cellular secretory machinery. There are only small changes in the cellular proteome of yeast secreting CALR at a high level.

The comparison between 2DE-MS and bottom-up LC-MS demands high-end techniques for both technologies.

In contrast to bottom-up LC-MS only 2DE-MS can separate and detect a huge number of human protein species. Kwiatkowski et al. (in this issue) established parameters to estimate the amount of protein speciation for each human protein. Proteins identified in 2DE-MS approaches showed more protein speciation than in bottom-up LC-MS. The authors state that protein speciation is likely to increase the chance of proteins to be determined in 2-DE/MS, though admitting that low-sensitivity 2DE-MS methods were used in this study. In agreement with Kwiatkowski et al., we are convinced that the difference between 2DE-MS and bottom-up LC-MS will disappear, if high-resolution 2DE is combined with identification by a high-sensitivity LC-Orbitrap-MS. Meta-analysis of proteomic data is surely a promising tool, though the technological progress in 2DE and MS has to reach a plateau to enable useful comparisons.

Design, synthesis, and anticancer evaluation of ammosamide B with pyrroloquinoline derivatives as novel BRD4 inhibitors.

Bromodomain-containing protein 4 (BRD4), which is a member of the bromodomain and extra-terminal domain (BET) family, plays an important role in the regulation of gene expression as the "reader" of epigenetic regulation. BRD4 has become a promising target to treat cancer, because the up-regulation of BRD4 expression is closely associated with the occurrence and development of various cancers. At present, several BRD4 inhibitors are in clinical trials for cancer therapy, but no BRD4 inhibitors are on the market. Here, we designed and synthesized a series of compounds bearing pyrrolo[4,3,2-de]quinolin-2(1H)-one scaffold through structural modification of natural products ammosamide B, which is a natural pyrroloquinoline derivative reported for its potential antitumor activity. All target compounds were evaluated for their BRD4 BD1 inhibition activities via the protein thermal shift assays or AlphaSceen assay. The representative compound 49 showed potent activity (IC50 = 120 nM). The co-crystal of compound 49 with BRD4 BD1 was solved to study the structure activity relationship, which showed that 49 could combine with the acetyl lysine binding site and formed a hydrogen bond with the conserved residue Asn140. The results demonstrate that compound 49 is worthy of further investigation as a promising BRD4 inhibitor.

Mechanisms of action of SGLT2 inhibitors and their beneficial effects on the cardiorenal axis.

Large clinical studies conducted with sodium-glucose co-transporter 2 inhibitors (SGLT2i) in patients with type 2 diabetes and heart failure with reduced ejection fraction have demonstrated their ability to achieve both cardiac and kidney benefits. Although there is huge evidence on SGLT2i-mediated clinical benefits both in diabetic and non-diabetic patients, the pathophysiological mechanisms underlying their efficacy are still poorly understood. Some favorable mechanisms are likely due to the prompt glycosuric action which is associated with natriuretic effects leading to hemodynamic benefits as well as a reduction in glomerular hyperfiltration and renin-angiotensin-aldosterone system activation. In addition to the renal mechanisms, SGLT2i may play a relevant role in cardiorenal axis protection by improving the cardiomyocyte metabolism, by exerting anti-fibrotic and anti-inflammatory actions, and by increasing cardioprotective adipokine expression. New studies will be needed to better understand the specific molecular mechanisms that mediate the SGLT2i favorable effects in patients suffering diabetes. Our aim is to first discuss about the molecular mechanisms underlying the cardiovascular benefits of SGLT2i in each of the main organs involved in the cardiorenal axis. Furthermore, we update on the most recent clinical trials evaluating the beneficial effects of SGLT2i in treatment of both diabetic and non-diabetic patients suffering heart failure.

Comparison of the outer membrane proteomes between clinical carbapenem-resistant and -susceptible Acinetobacter baumannii.

BACKGROUND AND AIM: Carbapenem resistance has become a major obstacle in combating Acinetobacter baumannii infections. Although enzymatic degradation by ß-lactamases is the pivotal mechanism of carbapenem resistance, porin deficiency has also been implicated in the mechanism. In this study, outer membrane proteins (OMPs) pattern of a clinical multidrug-resistant A. baumannii isolate were analysed in order to attain a deeper understanding of carbapenem-resistance strategies. METHODS: OMPs extracts, respectively, separated from carbapenem-resistant and -susceptible clinical A. baumannii isolates were compared using two-dimensional polyacrylamide gel electrophoresis. Differentially expressed proteins were identified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF). RESULTS: Twenty-three differently expressed proteins were identified between the resistant and susceptible isolates. Among them, six were annotated convincingly as OMPs in UniProt database. CarO was found absent from the resistant isolate and the expression levels of Omp33-36 and Omp25 were significantly lower than that in the susceptible counterpart. Strikingly, a LysM domain/BON superfamily protein, which has been linked to carbapenem resistance in Klebsiella pneumoniae, was found underexpressed by tenfold in the resistant isolate. CONCLUSION: Our study verified some porins which have been proven to play an important role in bacterial resistance against carbapenems. Underexpression of the LysM domain/BON superfamily protein may indicate its possible engagement in bacterial drug resistance, but its actual role requires more investigation.