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Background/Aims: β-Dystroglycan (β-DG) is a transmembrane glycoprotein that links the intracellular cytoskeleton to the extracellular matrix and is crucial for the molecular pathway of lateral force transmission in muscle. We aimed to investigate the effect of decreasing sarcolemmal cholesterol on the distribution of β-DG, its interaction with dystrophin and the impact on the contraction efficiency of muscle. Methods: Isolated rat extensor digitorum longus muscles were incubated with methyl β-cyclodextrin (MβCD) to deplete cholesterol and with MβCD-cholesterol to restore cholesterol. Electric stimulation protocols were used to determine muscle force and fatigue. Detergent-resistant membranes (lipid rafts) were separated from isolated skeletal muscle sarcolemma. The distribution and interactions of β-DG, caveolin-3 and dystrophin were determined by an immunoreactivity analysis. Results: Cholesterol depletion in muscle results in a weakened force of contraction, which recovers after cholesterol restoration. The rate of fatigue is unaffected, but fatigue recovery is dependent upon cholesterol restoration. MβCD modifies the structures of lipid rafts obtained from MβCD-treated muscles by, displacing the membrane proteins β-DG and caveolin-3 f from the lipid raft, thus reducing the interaction of β-DG with dystrophin. Conclusion: Cholesterol depletion weakens the muscle contractile force by disturbing the sarcolemmal distribution of β-dystroglycan and its interaction with dystrophin, two key proteins in the alignment of lateral force transmission pathway.

Incubation of bovine adrenal chromaffin cells in high K ϩ (38 mM) during 24 -48 h enhanced 2.5 to five times the expression of SNAP-25 protein and mRNA, respectively. This increase was reduced 86% by furnidipine (an L-type Ca 2ϩ channel blocker) but was unaffected by either -conotoxin GVIA (an N-type Ca 2ϩ channel blocker) or -agatoxin IVA (a P/Q-type Ca 2ϩ channel blocker). Combined blockade of N and P/Q channels with -conotoxin MVIIC did, however, block by 76% the protein expression. The inhibitory effects of furnidipine were partially reversed when the external Ca 2ϩ concentration was raised from 1.6 to 5 mM. These findings, together with the fact that nicotinic receptor activation or Ca 2ϩ release from internal stores also enhanced SNAP-25 protein expression, suggest that an increment of cytosolic Ca 2ϩ concentration ([Ca 2ϩ ] i ), rather than its source or Ca 2ϩ entry pathway, is the critical signal to induce the protein expression. The greater coupling between L-type Ca 2ϩ channels and protein expression might be due to two facts: (a) L channels contributed 50% to the global [Ca 2ϩ ] i rise induced by 38 mM K ϩ in indo-1-loaded chromaffin cells and (b) L channels undergo less inactivation than N or P/Q channels on sustained stimulation of these cells.

Alzheimer's disease (AD) is the most prevalent neurodegenerative disease, presenting the most devastating consequences on human health and life quality. Coumarin‐quinoline hybrids were synthesized following a very efficient and versatile strategy. Small structural variations contributed to dual acetyl/butyrylcholinesterases (AChE/BuChE) activity or selectivity towards one of these enzymes. In addition, some of the studied compounds are interesting iron chelators, presenting a tendency to be neuroprotective. Moreover, the compounds are not cytotoxic for SH‐SY5Y neuroblastoma cells. Compound 9c proved to be the most interesting compound of the studied series. This compound is selective against AChE and proved to be an excellent iron chelating agent (iron chelation at 100 μM=72.87%). Molecular docking studies were performed to establish the nature of the interaction between the studied compounds and the binding pockets, leading to a rationalization of structure–activity relationships. Compound 9c forms a well‐defined π‐stacking interaction with Phe330 and interacts with Tyr121 residue via a hydrogen bond, while the inactive compounds cannot establish these interactions. Important preliminary results against different targets, as well as some structure–activity relationships, were concluded from the experimental results.

We established two immortalized cell lines from cerebral cortex of normal (CNh) and trisomy 16 (CTb) mouse fetuses, an animal model of human trisomy 21. Those cells loaded with the fluorescent Ca2+ dyes, Indo-1 and Fluo-3, exhibited increments of intracellular Ca2+ ([Ca2+]i) in response to external glutamate, NMDA, AMPA and kainate. CTb cells exhibited higher basal Ca2+ concentrations and had higher amplitude and slower time-dependent kinetics in the decay than CNh cells, suggesting an impaired Ca2+ buffering capacity in the trisomy 16-derived cell line. Nicotine also induced increments of [Ca2+]i. The CTb cell line could represent a model for studying cellular alterations related to Down syndrome.

BACKGROUND There is a considerable degree of subjectivity and, therefore, substantial interobserver and intraobserver disagreement in the diagnosis and grading of dysplastic lesions in Barrett's esophagus (BE). The aim of this study was to evaluate the usefulness of DNA flow cytometry and immunohistochemical staining for p53 protein as objective methods to complement the conventional histologic diagnosis of dysplasia in patients with this disease. The most common problems and the possible advantages of using these procedures are analyzed briefly in this article. METHODS Formalin fixed, paraffin embedded tissue from 55 patients diagnosed with BE were processed for flow cytometric measurements (ploidy and proliferation index) and p53 immunostaining. RESULTS Both the cytometric data and the positivity of staining for p53 revealed a statistically significant increase throughout the following sequence: no dysplasia → indefinite for dysplasia → low grade dysplasia → high grade dysplasia → adenocarcinoma. There was also a highly significant correlation between the results of the cytometric study and the positivity of staining for p53. CONCLUSIONS In the future, the use of this procedure could play an important role in the evaluation of patients with BE. Considering that staining for p53 is technically simple, economical, and quick, and the materials required are available to most pathology laboratories, this method appears to be a firm candidate for application as a biomarker in BE. The authors have shown that it is possible to obtain adequate results for cytometric analysis with small formalin fixed, paraffin embedded biopsies if a strict protocol for the acceptance of tissue samples and/or histograms is observed. Cancer 1998;83:641‐651. © 1998 American Cancer Society.

Sarcoglycans (SGs) and sarcospan (SSPN) are transmembrane proteins of the dystrophin-glycoprotein complex. Mutations in the genes encoding SGs cause many inherited forms of muscular dystrophy. In this study, using purified membranes of wild-type (WT) and δ-SG knockout (KO) mice, we found the specific localization of the SG-SSPN isoforms in transverse tubules (TT) and sarcoplasmic reticulum (SR) membranes. Immunoblotting revealed that the absence of δ-SG isoforms in TT and SR results in a secondary deficiency of γ-SG and µSPN. Our results showed augmented ATP hydrolytic activity, ATP-dependent calcium uptake and passive calcium efflux, probably through SERCA1 in KO compared to WT mice. Furthermore, we found a conformational change in SERCA1 isolated from KO muscle as demonstrated by calorimetric analysis. Following these alterations with mechanical properties, we found an increase in force in KO muscle with the same rate of fatigue but with a decreased fatigue recovery compared to WT. Together our observations suggest, for the first time, that the δ-SG isoforms may stabilize the expression of γ-SG and µSPN in the TT and SR membranes and that this possible complex may play a role in the maintenance of a stable level of resting cytosolic calcium concentration in skeletal muscle.

Objective Different genetic polymorphisms of human leukocyte antigen (HLA) have been associated with the risk and prognosis of autoimmune and infectious diseases. The objectives of this study were to determine whether there is an association between HLA genetic polymorphisms and the susceptibility to and mortality of coronavirus disease 2019 (COVID-19) patients. Design Observational and prospective study. Setting Eight Intensive Care Units (ICU) from 6 hospitals of Canary Islands (Spain). Patients COVID-19 patients admitted in ICU and healthy subjects. Interventions Determination of HLA genetic polymorphisms. Main variable of interest Mortality at 30 days. Results A total of 3886 healthy controls and 72 COVID-19 patients (10 non-survivors and 62 survivor patients at 30 days) were included. We found a trend to a higher rate of the alleles HLA-A*32 ( p = 0.004) in healthy controls than in COVID-19 patients, and of the alleles HLA-B*39 ( p = 0.02) and HLA-C*16 ( p = 0.02) in COVID-19 patients than in healthy controls; however, all these p-values were not significant after correction for multiple comparisons. Logistic regression analysis showed that the presence of certain alleles was associated with higher mortality, such as the allele HLA-A*11 after controlling for SOFA (OR = 7.693; 95% CI = 1.063–55.650; p = 0.04) or APACHE-II (OR = 11.858; 95% CI = 1.524–92.273; p = 0.02), the allele HLA-C*01 after controlling for SOFA (OR = 11.182; 95% CI = 1.053–118.700; p = 0.04) or APACHE-II (OR = 17.604; 95% CI = 1.629–190.211; p = 0.02), and the allele HLA-DQB1*04 after controlling for SOFA (OR = 9.963; 95% CI = 1.235–80.358; p = 0.03). Conclusions The new finding from our preliminary study of small sample size was that HLA genetic polymorphisms could be associated with COVID-19 mortality; however, studies with a larger sample size before definitive conclusions can be drawn.