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Fabry disease is a heritable lipid disorder caused by the low activity of α-galactosidase A and characterized by the systemic accumulation of globotriaosylceramide (Gb3). Recent studies have reported a structural heterogeneity of Gb3 in Fabry disease, including Gb3 isoforms with different fatty acids and Gb3 analogs with modifications on the sphingosine moiety. However, Gb3 assays are often performed only on the selected Gb3 isoforms. To precisely determine the total Gb3 concentration, here we established two methods for determining both Gb3 isoforms and analogs. One was the deacylation method, involving Gb3 treatment with sphingolipid ceramide N-deacylase, followed by an assay of the deacylated products, globotriaosylsphingosine (lyso-Gb3) and its analogs, by ultra-performance LC coupled to tandem MS (UPLC-MS/MS). The other method was a direct assay established in the present study for 37 Gb3 isoforms and analogs/isoforms by UPLC-MS/MS. Gb3s from the organs of symptomatic animals of a Fabry disease mouse model were mainly Gb3 isoforms and two Gb3 analogs, such as Gb3(+18) containing the lyso-Gb3(+18) moiety and Gb3(−2) containing the lyso-Gb3(−2) moiety. The total concentrations and Gb3 analog distributions determined by the two methods were comparable. Gb3(+18) levels were high in the kidneys (24% of total Gb3) and the liver (13%), and we observed Gb3(−2) in the heart (10%) and the kidneys (5%). These results indicate organ-specific expression of Gb3 analogs, insights that may lead to a deeper understanding of the pathophysiology of Fabry disease.

(Princeton University, Engineering School) New findings about critical cellular structures have upended common assumptions about their formation and composition and provided new insight how molecular machines are built in living cells.

(University of California - Irvine) Cyanobacteria living in rocks in Chile's Atacama Desert extract water from the minerals they colonize and, in doing so, change the phase of the material from gypsum to anhydrite. Researchers at the University of California, Irvine and Johns Hopkins University gained verification of this process through experiments, and the work points to possible strategies for humans to stay hydrated in harsh environments.

(American Geriatrics Society) For only the second time in its near 80-year history, the American Geriatrics Society (AGS) will award one of its highest honors typically reserved for individuals to West Health, a family of nonprofit organizations dedicated to lowering healthcare costs to enable older adults to successfully age in place.

Fabry disease is a heritable lipid disorder caused by the low activity of α-galactosidase A and characterized by the systemic accumulation of globotriaosylceramide (Gb3). Recent studies have reported a structural heterogeneity of Gb3 in Fabry disease, including Gb3 isoforms with different fatty acids and Gb3 analogs with modifications on the sphingosine moiety. However, Gb3 assays are often performed only on the selected Gb3 isoforms. To precisely determine the total Gb3 concentration, here we established two methods for determining both Gb3 isoforms and analogs. One was the deacylation method, involving Gb3 treatment with sphingolipid ceramide N-deacylase, followed by an assay of the deacylated products, globotriaosylsphingosine (lyso-Gb3) and its analogs, by ultra-performance LC coupled to tandem MS (UPLC-MS/MS). The other method was a direct assay established in the present study for 37 Gb3 isoforms and analogs/isoforms by UPLC-MS/MS. Gb3s from the organs of symptomatic animals of a Fabry disease mouse model were mainly Gb3 isoforms and two Gb3 analogs, such as Gb3(+18) containing the lyso-Gb3(+18) moiety and Gb3(−2) containing the lyso-Gb3(−2) moiety. The total concentrations and Gb3 analog distributions determined by the two methods were comparable. Gb3(+18) levels were high in the kidneys (24% of total Gb3) and the liver (13%), and we observed Gb3(−2) in the heart (10%) and the kidneys (5%). These results indicate organ-specific expression of Gb3 analogs, insights that may lead to a deeper understanding of the pathophysiology of Fabry disease.

Quentin Perraud
Apr 1, 2020; 19:589-607
Research

Georg H H Borner
Apr 28, 2020; 0:R120.001971v1-mcp.R120.001971
Review

Eelke P. Béguin
Apr 24, 2020; 0:RA120.001964v1-mcp.RA120.001964
Research

Karl A. T. Makepeace
Apr 1, 2020; 19:624-639
Research

Research Event

Event participants

Louise Mushikiwabo, Secretary General, Organisation Internationale de la Francophonie (OIF)
Chair: Bob Dewar CMG, Associate Fellow, Africa Programme, Chatham House

An experimental and computational approach for identification of protein-protein interactions by ex vivo chemical crosslinking and mass spectrometry (CLMS) has been developed that takes advantage of the specific characteristics of cyanurbiotindipropionylsuccinimide (CBDPS), an affinity-tagged isotopically coded mass spectrometry (MS)-cleavable crosslinking reagent. Utilizing this reagent in combination with a crosslinker-specific data-dependent acquisition strategy based on MS2 scans, and a software pipeline designed for integrating crosslinker-specific mass spectral information led to demonstrated improvements in the application of the CLMS technique, in terms of the detection, acquisition, and identification of crosslinker-modified peptides. This approach was evaluated on intact yeast mitochondria, and the results showed that hundreds of unique protein-protein interactions could be identified on an organelle proteome-wide scale. Both known and previously unknown protein-protein interactions were identified. These interactions were assessed based on their known sub-compartmental localizations. Additionally, the identified crosslinking distance constraints are in good agreement with existing structural models of protein complexes involved in the mitochondrial electron transport chain.

Bacteria secrete siderophores to access iron, a key nutrient poorly bioavailable and the source of strong competition between microorganisms in most biotopes. Many bacteria also use siderophores produced by other microorganisms (exosiderophores) in a piracy strategy. Pseudomonas aeruginosa, an opportunistic pathogen, produces two siderophores, pyoverdine and pyochelin, and is also able to use a panel of exosiderophores. We first investigated expression of the various iron-uptake pathways of P. aeruginosa in three different growth media using proteomic and RT-qPCR approaches and observed three different phenotypic patterns, indicating complex phenotypic plasticity in the expression of the various iron-uptake pathways. We then investigated the phenotypic plasticity of iron-uptake pathway expression in the presence of various exosiderophores (present individually or as a mixture) under planktonic growth conditions, as well as in an epithelial cell infection assay. In all growth conditions tested, catechol-type exosiderophores were clearly more efficient in inducing the expression of their corresponding transporters than the others, showing that bacteria opt for the use of catechol siderophores to access iron when they are present in the environment. In parallel, expression of the proteins of the pyochelin pathway was significantly repressed under most conditions tested, as well as that of proteins of the pyoverdine pathway, but to a lesser extent. There was no effect on the expression of the heme and ferrous uptake pathways. Overall, these data provide precise insights on how P. aeruginosa adjusts the expression of its various iron-uptake pathways (phenotypic plasticity and switching) to match varying levels of iron and competition.

The vessel wall is continuously exposed to hemodynamic forces generated by blood flow. Endothelial mechanosensors perceive and translate mechanical signals via cellular signaling pathways into biological processes that control endothelial development, phenotype and function. To assess the hemodynamic effects on the endothelium on a system-wide level, we applied a quantitative mass spectrometry approach combined with cell surface chemical footprinting. SILAC-labeled endothelial cells were subjected to flow-induced shear stress for 0, 24 or 48 hours, followed by chemical labeling of surface proteins using a non-membrane permeable biotin label, and analysis of the whole proteome and the cell surface proteome by LC-MS/MS analysis. These studies revealed that of the >5000 quantified proteins 104 were altered, which were highly enriched for extracellular matrix proteins and proteins involved in cell-matrix adhesion. Cell surface proteomics indicated that LAMA4 was proteolytically processed upon flow-exposure, which corresponded to the decreased LAMA4 mass observed on immunoblot. Immunofluorescence microscopy studies highlighted that the endothelial basement membrane was drastically remodeled upon flow exposure. We observed a network-like pattern of LAMA4 and LAMA5, which corresponded to the localization of laminin-adhesion molecules ITGA6 and ITGB4. Furthermore, the adaptation to flow-exposure did not affect the inflammatory response to tumor necrosis factor α, indicating that inflammation and flow trigger fundamentally distinct endothelial signaling pathways with limited reciprocity and synergy. Taken together, this study uncovers the blood flow-induced remodeling of the basement membrane and stresses the importance of the subendothelial basement membrane in vascular homeostasis.

Protein subcellular localization is an essential and highly regulated determinant of protein function. Major advances in mass spectrometry and imaging have allowed the development of powerful spatial proteomics approaches for determining protein localization at the whole cell scale. Here, a brief overview of current methods is presented, followed by a detailed discussion of organellar mapping through proteomic profiling. This relatively simple yet flexible approach is rapidly gaining popularity, due to its ability to capture the localizations of thousands of proteins in a single experiment. It can be used to generate high-resolution cell maps, and as a tool for monitoring protein localization dynamics. This review highlights the strengths and limitations of the approach, and provides guidance to designing and interpreting profiling experiments. 

Perlecan is a critical proteoglycan found in the extracellular matrix (ECM) of cartilage. In healthy cartilage, perlecan regulates cartilage biomechanics and we previously demonstrated perlecan deficiency leads to reduced cellular and ECM stiffness in vivo. This change in mechanics may lead to the early onset osteoarthritis seen in disorders resulting from perlecan knockdown such as Schwartz-Jampel syndrome (SJS). To identify how perlecan knockdown affects the material properties of developing cartilage, we used imaging and liquid chromatography–tandem mass spectrometry (LC-MS/MS) to study the ECM in a murine model of SJS, Hspg2C1532Y-Neo. Perlecan knockdown led to defective pericellular matrix formation, whereas the abundance of bulk ECM proteins, including many collagens, increased. Post-translational modifications and ultrastructure of collagens were not significantly different; however, LC-MS/MS analysis showed more protein was secreted by Hspg2C1532Y-Neo cartilage in vitro, suggesting that the incorporation of newly synthesized ECM was impaired. In addition, glycosaminoglycan deposition was atypical, which may explain the previously observed decrease in mechanics. Overall, these findings provide insight into the influence of perlecan on functional cartilage assembly and the progression of osteoarthritis in SJS.

A fundamental feature of the eukaryotic cell membrane is the asymmetric arrangement of lipids in its two leaflets. A cell invests significant energy to maintain this asymmetry and uses it to regulate important biological processes, such as apoptosis and vesiculation. The dynamic coupling of the inner or cytoplasmic and outer or exofacial leaflets is a challenging open question in membrane biology. Here, we combined fluorescence lifetime imaging microscopy (FLIM) with imaging total internal reflection fluorescence correlation spectroscopy (ITIR-FCS) to differentiate the dynamics and organization of the two leaflets of live mammalian cells. We characterized the biophysical properties of fluorescent analogs of phosphatidylcholine, sphingomyelin, and phosphatidylserine in the plasma membrane of two mammalian cell lines (CHO-K1 and RBL-2H3). Because of their specific transverse membrane distribution, these probes allowed leaflet-specific investigation of the plasma membrane. We compared the results of the two methods having different temporal and spatial resolution. Fluorescence lifetimes of fluorescent lipid analogs were in ranges characteristic for the liquid ordered phase in the outer leaflet and for the liquid disordered phase in the inner leaflet. The observation of a more fluid inner leaflet was supported by free diffusion in the inner leaflet, with high average diffusion coefficients. The liquid ordered phase in the outer leaflet was accompanied by slower diffusion and diffusion with intermittent transient trapping. Our results show that the combination of FLIM and ITIR-FCS with specific fluorescent lipid analogs is a powerful tool for investigating lateral and transbilayer characteristics of plasma membrane in live cell lines.

VOLUME 295 (2020) PAGES 4079–4092There was an error in the abstract. “The pyridinium cation hampers uptake of OPs into the central nervous system (CNS)” should read as “The pyridinium cation hampers uptake into the central nervous system (CNS).”

The Open Arms Development Centre, a homeless shelter in downtown Kingston, and Jamaica’s COVID-19 Response Fund Food Relief are among 25 charitable and community organisations across the island to benefit from the latest round of grants from US-...

As England’s presumed consent law for 2020 clears parliament, Veronica English, head of medical ethics and human rights at the BMA, say that evidence from Wales and other countries shows that it could increase transplantation rates. But Blair L Sadler, physician and senior adviser to California State University, consider such legal changes a...

Margaret Heffernan has thought a lot about whistleblowing, and why companies don't respond well to it. She wrote the "Book Wilful Blindness: Why We Ignore the Obvious at our Peril". In this podcast she talks about how culture, and groupthink, leads to a culture where whistleblowers are ignored, and why the NHS needs to change the way it treats...

Reports from Italy, and more recently from the U.S. show the strain the healthcare system is under during this pandemic. We know that staff will step up in an emergency, but this isn’t a fire or a bombing, this is going to last for months. So how can organisations be proactive in supporting staff, and how can leaders try to mitigate the...

In his first public address on his inaugural official visit to Washington, DC, IOM Director General António Vitorino discusses his vision for IOM; reforms in the UN system; coordination with other organizations in addressing humanitarian protection; and the operational steps IOM is taking to respond to forced migration and displacement.

In his first public address during his inaugural formal visit to Washington, DC. International Organization for Migration (IOM) Director General António Vitorino discussed his vision for the agency; reforms in the UN system designed to address migration matters better; the coordination of efforts between IOM, the UN High Commissioner for Refugees, and other international partners in addressing humanitarian protection; and the operational steps IOM is taking to respond to forced migration and displacement in hotspots around the world, including Venezuela, Bangladesh, and Libya.

Francesco Rubino
Jun 1, 2016; 39:861-877
Metabolic Surgery and the Changing Landscape for Diabetes Care

Francesco Rubino
Jun 1, 2016; 39:861-877
Metabolic Surgery and the Changing Landscape for Diabetes Care

The Organized Dentistry Coalition is applauding Congress for introducing legislation aimed at improving oral health literacy.

The ADA, along with a large group of other dental organizations, told the U.S. Department of Labor that they are concerned about provisions in HR 6201, the Families First Coronavirus Response Act as related to family and medical leave and paid sick leave.

The ADA and nearly 100 other organizations are asking Congress and the White House to help businesses survive the coronavirus crisis by reducing their costs and increasing their cash flow as much as possible in the coming weeks.

The ADA and other health care organizations signed a public statement March 27 saying they “strongly” support emergency efforts at the federal level to dramatically increase the production and distribution of personal protective equipment (PPE) and other necessary medical equipment and supplies during the COVID-19 pandemic.

The ADA, Organized Dentistry Coalition, and many state dental associations are asking the U.S. Department of Health and Human Services and Centers for Medicare & Medicaid Services to release funding from the Coronavirus Aid, Relief and Economic Security Act Provider Relief Fund to assist dental Medicaid providers.

The Organized Dentistry Coalition is asking Congress for flexibility in the Paycheck Protection Program to allow dentists to purchase personal protective equipment as states begin the reopening phase of the COVID-19 pandemic.

Neurons in the hippocampus categorize what we experience into abstract, discrete events, such as taking a walk versus having lunch

-- Read more on ScientificAmerican.com

Days before the Sept. 11, 2001, attacks, Osama bin Laden left his compound in Kandahar in Afghanistan and headed into the mountains. His driver, Salim Ahmed Hamdan, traveled with him. As U.S. and Northern Alliance forces stood poised to capture Kandahar a few months later, bin Laden told Hamdan t...

OBJECTIVE

To construct and internally validate prediction models to estimate the risk of long-term end-organ complications and mortality in patients with type 2 diabetes and obesity that can be used to inform treatment decisions for patients and practitioners who are considering metabolic surgery.

Two forthcoming pieces in History of the Human Sciences may be of interest to AHP readers. Full details below. “Cold War Pavlov: Homosexual aversion therapy in the 1960s,” by Kate Davison. Abstract: Homosexual aversion therapy enjoyed two brief but intense periods of clinical experimentation: between 1950 and 1962 in Czechoslovakia, and between 1962 and 1975 … Continue reading Forthcoming in HHS: Homosexual Aversion Therapy, Comte on Organism-Environment Relationships →

"Tom's Holiday is all about road safety for young children. Tom and his family park the car and head for the beach at a little beachside village. The beachside village is different from home in the city - there are no footpaths. Mum and Dad teach Tom that we must be careful because holiday towns don't have kerbs and gutters. The roads can be thin and there aren't white lines. It's very important that children hold their parent's hand" -- From the Internet.

Speculative fiction, Australian.

Leipzig : G. Thieme, 1890-1891.

Berlin : A. Hirschwald, 1890.

London : Macmillan, 1896.