NEWS: Tracking a Serial Killer: Could Ebola Mutate to Become More Deadly?

Why we need to terminate Ebola 2014 before the virus learns too much about us.

The Ebola virus, seen here through a scanning electron microscope, is mutating as cases increase, raising the odds that the virus could become more transmissible.

Photograph by NIAID, Alamy

 

By David Quammen

National Geographic News

 

Published October 15, 2014

 

Forty years ago, Ebola was just the name of a river. It was a small waterway of no particularly sinister character that flowed through northern Zaire, not far from the village hospital where the first known outbreak of a new viral disease had been centered. That river gave its name to the new virus, and now "Ebola" is a global byword for ugly death, misery, and fear of contagion.

 

The 2014 epidemic of Ebola virus disease in West Africa is unprecedented in scope, and much attention has been focused, rightly, on how it has gotten so badly out of control.

 

Behind that question are three others, less obvious, more complicated, and crucial to seeing Ebola in a broader context: Where did the virus come from? Where is it going? What's next? We do well to consider these questions even as we react to the daily headlines, urge our leaders to take more deeply committed action, and support the organizations (such as Doctors Without Borders) that are fighting the epidemic so courageously in West Africa.

 

Where Did It Come From?

The outbreak began in early December, in a village called Meliandou, southeastern Guinea, not far from the borders with both Liberia and Sierra Leone. The first known case was a two-year-old child who died, after fever and vomiting and passing black stool, on December 6. The child's mother died a week later, then a sister and a grandmother, all with symptoms that included fever, vomiting, and diarrhea. Then, by way of caregiving visits or attendance at funerals, the outbreak spread to other villages.

See graphic:
Ebola: Tracking the Outbreak

It wasn't until March, three months later, that local officials alerted the Guinean Ministry of Health about these clusters of a strange, lethal disease in the countryside. By then, human-to-human transmission had started to multiply the case count. But tracing linked cases raises the question of ultimate origin. How did that first child get sick?

 

Ebola virus is a zoonosis, meaning an animal infection transmissible to humans. The animal in which a zoonosis lives its customary existence, discreetly, over the long term, and without causing symptoms, is called a reservoir host. The reservoir host of Ebola virus is still unknown—even after 38 years of efforts to identify it, since the original 1976 outbreak—although one or more kinds of fruit bat, including the hammer-headed bat, are suspects. There are hammer-headed bats in southeastern Guinea. It's possible that somebody killed one for food and brought it to Meliandou, where the child became infected either by direct contact with the bat or by virus passed on the hands of an adult.

 

Why are these facts and suppositions significant? Because they remind us that Ebola virus abides endemically in the forests of equatorial Africa. It will never be eradicated as long as those forests exist, unless the reservoir host itself is eradicated (not recommended) or cured of the viral infection (not likely possible). The virus may retire into its hiding place for years at a time, but eventually it will return, as a result of some disruptive contact by humans with the reservoir host. Then it will spill over into us again. All thinking and planning about how to defend against Ebola virus disease in the future needs to take account of that reality.

 

Another puzzling fact about origins is that the West Africa epidemic involves a species of ebolavirus (that's the label for the group, which includes five species) previously known only from outbreaks in the Democratic Republic of the Congo and its close neighbors.

 

A different species has emerged in Ivory Coast, another West African country, just east of Guinea and Liberia. According to a study published in Science in late August by Stephen K. Gire of Harvard and a long list of co-authors, the virus in West Africa seems to have diverged from its lineage in Central Africa just within the past decade. It somehow leapfrogged over or around the Ivory Coast ebolavirus in order to situate itself in southeastern Guinea. That suggests the unnerving prospect that the Central African ebolavirus (the only one strictly known as Ebola virus) is expanding its range, either by infecting new populations of reservoir hosts or by migrations of those host animals.

 

One way or another, it has been on the move.

 

Fruit bats are sold at an outdoor market in Brazzaville, capital of the Republic of the Congo. The reservoir host of Ebola virus is still unknown, but one or more kinds of fruit bat are suspects.

 

Photograph by Jiro Ose/Redux

 

Where Is It Going?

 

The virus has also traveled within living human bodies. We know that it went from Liberia to Dallas within the late Thomas Eric Duncan, from Liberia to Nigeria by way of the late Patrick Sawyer, and from Sierra Leone to Spain by way of two Spanish missionary priests, both also now deceased, who were evacuated for treatment.

 

And it has been carried to Omaha, Atlanta, London, Paris, Hamburg, Frankfurt, and Oslo within infected people, mostly health and aid workers brought home to be treated.

 

But just as worrisome as the virus's geographic spread is its journey across the evolutionary landscape. Is it mutating in ways that could make it more dangerous to humans? Is there any chance that it might become transmissible through the air, like the flu, the SARS virus, or a common cold?

Although Ebola becoming airborne is the ultimate disease nightmare, that seems to be almost vanishingly improbable, for reasons well put in a recent article in the Washington Post by Laurie Garrett, a senior fellow for global health at the Council on Foreign Relations. What is now a fluid-borne virus attaching itself to cells lining the circulatory system can't easily change into one that targets the tiny air sacs in the lungs.

 

"That's a genetic leap in the realm of science fiction," Garrett wrote.

 

The virus probably will not go airborne, but it could conceivably increase its Darwinian fitness in other ways, becoming more subtle and elusive.

 

The genetic study by Gire and his colleagues (five of whom were dead of Ebola by the time their study appeared) found 341 mutations as of late August, some of which are significant enough to change the bug's functional identity. The higher the case count in West Africa goes, the more chances for further mutations, and therefore the greater possibility that the virus might adapt somehow to become more transmissible-perhaps by becoming less pathogenic, sickening or killing its victims more slowly and thereby leaving them more time to infect others.

 

That's why, the Gire group wrote, we need to stop this thing everywhere as soon as possible. Future spillovers of Ebola are bound to occur, but those freshly emerged strains of the virus, direct from the reservoir host, won't contain any adaptive mutations that the West Africa strain is acquiring now.

We need to terminate Ebola 2014 before the virus learns too much about us.

 

Kumba Conde cries after her sister Marie, 14, died from Ebola in Koundony, Guinea, in July 2014. The current outbreak began in December 2013 in southeastern Guinea, not far from the borders with both Liberia and Sierra Leone.

 

Photograph by Samuel Aranda, The New York Times/Redux

 

What's Next?

 

No one knows, of course, how much worse the epidemic in West Africa will get. The U.S. Centers for Disease Control and Prevention issued a report, in late September, projecting that under the worst-case scenario there could be 1.4 million cases by early next year. The World Health Organization said Tuesday that new cases could rise to 10,000 per week by December, ten times the rate of the previous month. And the World Bank has warned that costs of the epidemic could reach $32.6 billion, which would be an economic catastrophe for the three West African countries that would compound their health catastrophes.

 

Will the epidemic spread more widely, igniting outbreaks in other parts of the world? We hope not. Will it turn up as additional cases, here and there, among people who have traveled from West Africa unaware, as Thomas Eric Duncan was reportedly unaware, that they were infected before boarding the airplane? Probably.

 

What's the best way to limit such occurrences? Rigorous screening at airports, quarantine for travelers who test positive, travel restrictions, or perhaps total bans on commercial flights arriving from Liberia, Guinea, and Sierra Leone-these measures should help. The most important and effective thing we can do, though, is to provide all possible assistance toward ending the outbreak where it began, in West Africa.

 

The world won't be free of Ebola 2014 until West Africa is free of it. Even severe restrictions, barring entry to anyone traveling from West Africa, would not make it impossible for the virus to get into America, or Europe, or wherever. To understand why, consider what I call the Nairobi Tabletop Scenario.

Imagine a doctor who departs from Monrovia, the capital of Liberia, feeling fine, on a flight to Nairobi, Kenya's capital, in East Africa. In transit he begins suffering a headache-nothing terrible yet, just discomfort, but it's the first hint of Ebola. At the Nairobi airport, in a café, the Liberian doctor coughs onto a table. Five minutes later, an American businessman touches that table. He rubs his eye. He departs to Singapore and spends three days there, in good health, discussing finance for his project in Kenya. Then he flies home to Los Angeles. To the screeners at LAX, he is an American businessman arriving from Singapore, with no history of recent travel in West Africa. But he's now infected with Ebola, carrying it into the United States.

 

How do you defend against the Nairobi Tabletop Scenario? By doing everything possible to end the epidemic in West Africa, and thereby to ensure that the Liberian doctor is healthy when he visits Nairobi.

 

Our safety against the menace of killer viruses can never be an absolute safety. There are too many of them, lurking within reservoir hosts amid distant forests or closer to home-viruses such as Nipah in Bangladesh, Marburg in Uganda, Lassa in West Africa, Sin Nombre virus in the American West, all the new influenzas coming out of southeastern Asia, plus many others that haven't yet been identified and named.

 

And there are too many of us humans, sharing the landscape with the reservoir hosts and with one another. We are too interconnected by air travel and transport. Viruses are simple organisms but well-adapted to the modern world. This year it's Ebola, devastating and scary. Next year it will be something else.

 

David Quammen is a contributing writer for National Geographic and author of Spillover and The Song of the Dodo, among other works. His new book Ebola: The Natural and Human History of a Deadly Virus will be published by W.W. Norton next week.

 

 

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Cardiac valve cells and their microenvironment—insights from in vitro studies

Nature Reviews Cardiology | Review

 

In conditions of prolonged stress valvular cells can contribute to valve disease progression. In this Review, Wang and colleagues describe how the biophysical and biochemical properties of the extracellular matrix... can regulate valve cell function in the context of calcific valvular diseases. The authors also describe how new cell culture approaches can be employed to better understand the pathophysiology of valve diseases.

 

Abstract

 

During every heartbeat, cardiac valves open and close coordinately to control the unidirectional flow of blood. In this dynamically challenging environment, resident valve cells actively maintain homeostasis, but the signalling between cells and their microenvironment is complex. When homeostasis is disrupted and the valve opening obstructed, haemodynamic profiles can be altered and lead to impaired cardiac function. Currently, late stages of cardiac valve diseases are treated surgically, because no drug therapies exist to reverse or halt disease progression. Consequently, investigators have sought to understand the molecular and cellular mechanisms of valvular diseases using in vitro cell culture systems and biomaterial scaffolds that can mimic the extracellular microenvironment. In this Review, we describe how signals in the extracellular matrix regulate valve cell function. We propose that the cellular context is a critical factor when studying the molecular basis of valvular diseases in vitro, and one should consider how the surrounding matrix might influence cell signalling and functional outcomes in the valve. Investigators need to build a systems-level understanding of the complex signalling network involved in valve regulation, to facilitate drug target identification and promote in situ or ex vivo heart valve regeneration.

 

Key points

• The interaction between valve cells and their microenvironment signals determine the functional output of cardiac valve tissues; cardiac valve diseases might be caused by a disruption of these interactions
• In vitro cell culture studies can help understand cardiac valve disease pathobiology, which is enhanced by culturing cells on biomaterials that mimic the native valve cell matrix environment
• Many biochemical signals regulate pathogenic phenotypes of cells in the valve; however, their effects are dependent on the matrix microenvironment
• Synthetic biomaterials can be used to understand valve cell regulation by biochemical, biophysical, and other matrix-associated signals, especially related to the development of valvular diseases
• Understanding the dynamic interplay between microenvironmental signals and valvular cell phenotypes will enable the identification of new therapies and the design of ex vivo tissue engineered valves

Introduction

 

As the heart evolved from a single-chamber to a multiple-chamber structure, cardiac valves arose to control the unidirectional flow of blood during cardiac cycles. For example, aortic valves open in response to higher blood pressure in the left ventricle than in the aorta, and close when the pressure equilibrates. These valves function in a similar manner to valves in water dams or car engines, but cardiac valves are living tissue with the ability to repair and remodel in response to damage. During an average human life span, heart valves open and close approximately 3 billion times,¹ withstanding various mechanical stresses, including fluid shear stresses and bending stretch.^{2, 3}

 



Figure 1: Valve cells and their matrix regulate tissue homeostasis and disease progression.

a | Cardiac valves have three extracellular matrix layers: the ventricularis, spongiosa, and fibrosa. In this image, a cross-section of a porcine aortic valve was visualized by second-harmonic generation confocal imaging. Elastin in green, collagen in red. Scale bar: 100 μm. b | Two main cell populations comprise the cardiac valve. VECs, which are CD31⁺ (green), line the boundaries of the leaflets. VICs reside throughout the valve leaflet. Nuclear staining (DAPI; blue), Scale bar: 100 μm. c | In healthy valves (left), very few VICs and VECs are activated to repair local tissue damage during normal valve function. Collagen fibres are circumferentially aligned, and activated VICs undergo continual turn over into deactivated fibroblasts and other cell fates (such as apoptosis). In diseased valves (right), VICs and VECs are recruited to remodel the tissue, leading to excessive collagen accumulation, a degraded and disarrayed matrix, which in some cases becomes calcified. Valve cells lose the homeostatic equilibrium and remain in an activated state. Abbreviations: VEC, valvular endothelial cell; VIC, valvular interstitial cell.





The material composition and structure of cardiac valves confer their robustness and durability. In humans, cardiac valves are made of thin (~500 μm) pliable cusps, and only mitral and tricuspid valves have supporting chordae tendineae and papillary muscles.⁴ A close examination of the tissue architecture of an aortic valve reveals three distinct layers of extracellular matrix (ECM), rich in collagens, proteoglycans, or elastin (Figure 1a).⁴ These ECM proteins impart unique macroscopic mechanical properties to valves, enabling them to withstand tension when closed and flexure when open. For example, the elastin fibres on the flow side of the valves (known as ventricularis) are radially aligned and elastic, which extend when the valves open and recoil when valves close.⁵ Proteoglycans in the middle layer, or spongiosa, function as a cushion for absorbing tension and friction between the top and the bottom layers.⁴ Finally, the fibrosa layer contains circumferentially oriented collagen fibres, which confer stiffness and strength to the valves.⁴



Figure 2: Dynamic cellular phenotypes in cardiac valve diseases.

a | Mineralized spherical particles (red) have been observed in human calcified valves, but not in normal valves. Scale bars 2 μm. b | Cells in the valve adopt diseased phenotypes (myofibroblast and osteoblast-like cells) in response to environmental signals. Activated myofibroblasts can revert back to quiescence, senescence, or undergo apoptosis. Osteoblast-like cells might also undergo apoptosis. (1) Myofibroblasts are characterized by ACTA2-positive stress fibres (green). Scale bar 50 μm. (2) VICs can be induced to mineralize, indicated by Alizarin Red S staining. Scale bar: 50 μm. Abbreviations: ACTA2, actin, aortic smooth muscle; VEC, valvular endothelial cell; VIC, valvular interstitial cell. Permission for panel a obtained from NPG © Bertazzo, S. et al. Nano-analytical electron microscopy reveals fundamental insights into human cardiovascular tissue calcification.

 

 







Figure 3: Matrix signals regulate valve cell phenotypes.

a | Plastic culture plates have high stiffness and biochemical signals must be introduced as soluble factors in the media. The stiffness of biomaterial-based cultures can mimic native tissues, and enables cells to grow in 3D space, with biochemical signals presented throughout the matrix. b | Adhesive signals (yellow in schematic) are tethered to an inert matrix to study specific cellular phenotype effects. For example, an elastin-derived peptide (VGVAPG, green bars) or a collagen-derived peptide (P15, blue bars) covalently linked to PEG hydrogels has distinct effects on ACTA2 levels. c | Elasticity can elicit intracellular cytoskeletal changes. In a modulus gradient of 7–32 kPa, >80% of PAVICs activated to myofibroblasts near the stiffer region and <10 1="" 3="" abbreviations:="" acta2="" actin="" after="" aortic="" became="" blue="" culture="" day="" days="" differences="" end="" i="" line="" muscle="" myofibroblasts="" near="" not="" observed="" of="" on="" red="" smooth="" softer="" the="" these="" were="">E

, Young's modulus; PAVIC, porcine aortic valve interstitial cell; PEG, poly(ethylene glycol). Permission for panel b obtained from Gould, S. T. et al. Small peptide functionalized thiol-ene hydrogels as culture substrates for understanding valvular interstitial cell activation and de novo tissue deposition. Acta Biomaterialia, 8, 3201–3209 (2012). Permission for panel c obtained from Kloxin, A. M. et al. In situ elasticity modulation with dynamic substrates to direct cell phenotype.





Figure 4: Valve cells and the ECM microenvironment are dynamic and mutually regulated.

Valvular interstitial cells interact with the ECM directly through integrins, with biochemical signals via receptors, and with neighbouring cells via cadherins. In healthy tissue (left), the cells and ECM are in a homeostatic state. When microenvironmental signals of valvular disease are present (right), cells can be activated, and biological outputs are altered. Secretion of ECM proteins, matrix-remodelling enzymes, and biochemical signals is increased; more contractile stress fibres and cell-matrix adhesions form; apoptosis increases where effective matrix interactions are lost. These cellular functions alter the microenvironment and valve tissue structure in diseased valves. Abbreviations: ACTA2, actin, aortic smooth muscle; ECM, extracellular matrix; MMP, matrix metalloproteinase.



                                                                                                                                                                                                                                                                                                                                                                                                                                         Huan Wang,^1, Leslie A. Leinwand^2, & Kristi S. Anseth^1,

 

 

Published online 14 October 2014

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Novel antiplatelet agents in acute coronary syndrome

Nature Reviews Cardiology | Review   

Dual antiplatelet therapy (DAPT) with aspirin and the P2Y purinoceptor 12 (P2Y12)-receptor inhibitor clopidogrel has been considered the gold standard of care in patients with acute coronary syndrome (ACS). The FDA approval of the novel P2Y12-receptor antagonists prasugrel and ticagrelor, which are faster-acting and more potent than clopidogrel, in the past 5 years have improved clinical outcomes in patients with ACS. Even with these newer compounds, many patients continue to experience adverse ischaemic events. This Review will provide an overview of the antiplatelet agents currently used to treat patients with ACS. Furthermore, emerging antiplatelet therapies, including intravenous P2Y12 antagonists, oral PAR-1 antagonists and thromboxane-receptor inhibitors will be discussed.

Abstract

For more than 10 years, dual antiplatelet therapy with aspirin and clopidogrel has remained the cornerstone of treatment for patients with acute coronary syndrome (ACS). The novel oral P2Y purinoceptor 12 (P2Y₁₂)-receptor inhibitors prasugrel and ticagrelor were approved by the FDA for clinical use in 2009 and 2011, respectively. These agents have a faster-acting, more-potent, and more-predictable antiplatelet effect than clopidogrel, which translates into improved clinical outcomes in patients with ACS, albeit at the expense of an increased risk of bleeding. However, some patients continue to experience adverse ischaemic events despite treatment with aspirin and a P2Y₁₂-receptor antagonist, because platelets can remain activated via pathways not inhibited by these agents, such as the protease-activated receptor (PAR)-1 platelet activation pathway stimulated by thrombin. Emerging antiplatelet therapies that might address these limitations include intravenous P2Y₁₂ antagonists, oral PAR-1 antagonists, and thromboxane-receptor inhibitors. In this Review, we provide an overview of these novel antiplatelet drugs, including newly approved agents and emerging compounds currently under clinical development, and also discuss evolving concepts and unmet needs related to antiplatelet therapy for the treatment of ACS.

At a glance

• Platelet adhesion, activation, and aggregation after plaque rupture or erosion are the major determinants of arterial thrombosis leading to acute coronary syndrome (ACS)
• Antiplatelet therapy, which targets pathways of platelet activation and aggregation, is pivotal in both the acute treatment and long-term secondary prevention of ischaemic events in patients with ACS
• Dual antiplatelet therapy with a combination of aspirin and either prasugrel or ticagrelor should be the treatment of choice in patients with ACS
• Drug selection should be based on contraindications and the individual patient's characteristics; clopidogrel should be used when both prasugrel and ticagrelor are contraindicated or not available
• Patients might continue to experience adverse events despite the use of prasugrel or ticagrelor
• New intravenous P2Y purinoceptor 12 inhibitors and agents that target other platelet-activation pathways have been developed

Figure 1: Mechanism of thrombus formation.

Plaque rupture exposes subendothelial components. Platelet adhesion during the rolling phase is mediated by interactions between vWF and GP Ib/V/IX receptor complexes located on the platelet surface, and between platelet collagen recep…

Figure 2: Platelet activation pathways targeted by current and novel antiplatelet agents.

Platelet adherence to injured vascular endothelium is mediated by binding of GP receptors to exposed extracellular matrix proteins (collagen and vWF). Complex intracellular signalling processes result in the local production and releas…

Figure 3: Mechanisms of action of P2Y₁₂ inhibitors.

Activation of P2Y₁₂ inhibits AC, causing a decrease in cAMP and VASP-P levels, and activation of P2Y₁ causes an increase in intracellular Ca²⁺ levels. These changes promote platelet aggregation by altering the ligand-binding properties…

Introduction

Arterial thrombosis after atherosclerotic plaque rupture or erosion is the major determinant of acute coronary syndrome (ACS).^{1, 2} Platelet adhesion, activation, and aggregation have important roles in the development of arterial thrombi; antiplatelet therapy is, therefore, pivotal in the treatment of ACS.^{2, 3, 4} Multiple platelet-signalling pathways are involved in thrombus formation and are potential targets for therapies.^{2, 3, 4} At present, three main classes of antiplatelet drugs are clinically approved for the treatment and secondary prevention of thrombotic complications in patients with ACS: oral cyclooxygenase-1 (COX-1; also known as prostaglandin G/H synthase 1) inhibitors, oral P2Y purinoceptor 12 (P2Y₁₂)-receptor inhibitors, and intravenous glycoprotein (GP) IIb/IIIa (also known as integrin α_IIbβ₃) inhibitors.^{5, 6, 7, 8}

The synergistic platelet-inhibitory effects exerted by dual antiplatelet therapy (DAPT) consisting of aspirin-induced blockade of COX-1 in conjunction with P2Y₁₂-receptor inhibition led to more than 10 years of clinical trials in patients with manifestations of ACS, including unstable angina, non-ST-segment elevation (NSTE) myocardial infarction (MI) and ST-segment elevation MI (STEMI).^{5, 6, 7, 8, 9, 10} Clopidogrel is the most broadly investigated P2Y₁₂-receptor inhibitor, owing to its favourable safety profile compared with ticlopidine.¹¹

Therefore, clopidogrel in combination with aspirin is still the cornerstone of treatment in patients with ACS.^{5, 6, 7, 8} However, despite the undisputable benefits of DAPT, a considerable number of patients continue to experience adverse thrombotic events.¹² These adverse events have been in part attributed to the nonuniform platelet inhibitory effects induced by clopidogrel, also known as interindividual clopidogrel response variability, where individuals who persist with high platelet reactivity despite clopidogrel therapy are at an increased risk of atherothrombotic recurrences.^{12, 13, 14, 15} Moreover, despite adequate COX-1 and P2Y₁₂-receptor blockade,^{3, 4} other platelet signalling pathways continue to be activated, and can contribute to thrombus formation. These observations have prompted the development of novel antithrombotic agents.

In this Review, we provide an overview of advances in antiplatelet therapy in the setting of ACS, focusing on novel drugs that have already been approved for clinical use, in addition to emerging agents at different phases of clinical development. However, a detailed description of the role of novel oral anticoagulants is beyond the scope of this Review, and the use of such agents is only briefly mentioned.

Mechanisms of atherothrombosis

Plaque rupture, fissure, or erosion exposes the subendothelial layer and leads to the recruitment and activation of platelets, as well as the generation of excessive levels of thrombin, which ultimately results in the formation of a fibrin-rich thrombus.^{1, 2, 3, 4} NSTE-ACS is frequently characterized by a nonocclusive or transiently occlusive thrombus, whereas a more stable and occlusive thrombus is typical in STEMI.¹ Platelet-activated thrombosis involves three principal steps: platelet adhesion, activation and recruitment of additional platelets, and platelet aggregation (Figure 1).^{3, 4, 16} Platelet adhesion during the initial rolling phase is mediated by interactions between the GP Ib/V/IX receptor complex, located on the surface of the platelet, and von Willebrand factor (vWF) bound to collagen exposed at the site of vascular injury.¹⁷ Rolling also brings the collagen receptors GP VI and GP Ia on platelets into contact with collagen.¹⁷ Binding of collagen to these receptors initiates platelet activation and triggers intracellular mechanisms that shifts platelet integrins to a high-affinity state, which induces the release of activating factors, in turn promoting aggregation, further recruitment, and further activation of circulating platelets.^{3, 4, 16, 17} These factors include ADP, epinephrine, serotonin, thrombin, and thromboxane A₂.^{3, 4} Platelet activation by these and other mediators (such as collagen) induces changes in platelet shape, expression of proinflammatory molecules (such as P-selectin and soluble CD40 ligand) and platelet procoagulant activity.^{3, 4, 16} The final step in platelet aggregation and thrombus formation involves the conversion of platelet GP IIb/IIIa (the main receptor mediating platelet aggregation) into its active form, which subsequently binds to the extracellular ligands fibrinogen and vWF, leading to platelet aggregation and thrombus formation mediated by platelet–platelet interaction.^{17, 18} In addition, vascular injury also exposes subendothelial tissue factor, which forms a complex with factor VIIa, activating the clotting cascade and leading to thrombin generation.¹⁹ However, only a modest amount of thrombin is produced as a result of the coagulation cascade; its main source within a platelet plug is the surface of activated platelets.^{16, 20} Thrombin converts fibrinogen to fibrin, generating a fibrin-rich clot, and further activates platelets by binding to protease-activated receptor (PAR)-1 and PAR-4.^{3, 4, 21, 22} Pathogenic thrombosis, therefore, involves a complex interplay between platelets and plasma components (coagulation factors) that interact in an auto-amplified process (Figure 1).

Francesco Franchi^1, & Dominick J. Angiolillo^1,

Nature Reviews Cardiology Year published: (2014) DOI: doi:10.1038/nrcardio.2014.156

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For diabetes, stem cell recipe offers new hope

Researchers coax stem cells into becoming long-sought insulin-producing β cells

news.sciencemag.org

 

This week, researchers reported a significant step towards finding a cure for diabetes. They've found a recipe that can turn human stem cells into functional pancreatic β cells—the same cells that are destroyed by the body’s own immune system in type 1 diabetes patients.

Latest News



 

 Douglas Melton

Two weeks after transplant into a diabetic mouse, human pancreatic β cells made in the lab produce enough insulin (green) to cure the animal.

Douglas Melton is as impatient as anyone for a cure for diabetes. His son developed the disease as an infant, and his daughter was diagnosed at age 14. For most of the past 2 decades, the developmental biologist at the Harvard Stem Cell Institute has focused his research on finding a cure. This week, he and his colleagues report a potentially significant step toward that goal: a recipe that can turn human stem cells into functional pancreatic β cells—the cells that are destroyed by the body’s own immune system in type 1 diabetes patients such as Melton’s son and daughter. The cells the researchers produced respond to glucose by producing insulin, just as normal β cells do. And when implanted into mice with a form of diabetes, the cells can cure the disorder.

 

“The diabetes research community has been waiting for ages for this type of breakthrough,” says Jorge Ferrer, who studies the genetics of β cells at Imperial College London. The lab-generated cells should be a valuable tool for studying diabetes and, Melton hopes, could eventually be used to treat patients.

 

Throughout the day, the pancreas regulates the body’s blood sugar levels, responding to an increase in glucose after a meal by secreting insulin, which helps cells take up the sugar. In type 1 diabetes, the body’s immune system mistakenly kills the β cells for still-unknown reasons, and the body is left without insulin. People control their diabetes by injecting carefully calibrated doses of insulin. But matching the precise insulin control achieved by the healthy pancreas is almost impossible, so researchers have hoped for decades to find a way to replace the missing cells.

 

When scientists isolated human embryonic stem (ES) cells in 1998, hopes soared. ES cells are pluripotent, which means that in theory they can turn into any of the body’s cell types—including β cells. Indeed, one of the first things researchers tried to make from ES cells was pancreatic β cells. Later they tried with so-called induced pluripotent stem (iPS) cells, made by reprogramming adult cells into an embryolike state. Either way, “it’s proved to be an extraordinarily complicated undertaking,” says Mark Magnuson of Vanderbilt University in Nashville, who studies pancreatic development.

 

Several teams have turned stem cells into precursors of β cells, which mature when placed into experimental animals. But the cells take 6 weeks to become fully functional β cells, and they can’t be studied easily outside the body. Nevertheless, a clinical trial started last month to test their therapeutic use in patients.

 

In Cell this week, Melton and his colleagues report a complex recipe that can transform either human ES cells or iPS cells directly into functional β cells. The breakthrough is based on more than a decade of tenacious work in Melton’s lab. He and his colleagues have painstakingly studied the signals that guide pancreas development, applying what they and others have found to develop a method that turns stem cells into mature β cells. “There’s no magic to this,” Melton says. “It’s not a discovery so much as applied developmental biology.”

 

The protocol “is reproducible, but it is tedious,” Melton adds. The stem cells are grown in flasks and require five different growth media and 11 molecular factors, from proteins to sugars, added in precise combinations over 35 days to turn them into β cells. On the bright side, Melton says, the technique can produce 200 million β cells in a single 500 ml flask—enough, in theory, to treat a patient. Melton says the protocol seems to work equally well with ES and iPS cell lines.

 

Before the cells can be used to treat type 1 diabetes, researchers need to find a way to protect them from immunologic rejection. The same autoimmune response that triggered the disease would likely attack new β cells derived from the patient’s own iPS cells, and a normal immune response would destroy ES-derived β cells, which would appear foreign. (That has been a challenge for efforts to treat type 1 diabetes with received transplants of β cells from deceased organ donors.) Melton and colleagues are now exploring how to physically encapsulate their stem cell–derived β cells, as well as ways to modify the β cells to enable them to ward off immune attack.

 

In the meantime, the cells should help the study of the autoimmune disorder. The technique “potentially provides ways to create model systems for studying the genetic basis of diabetes, or to discover novel therapies to enhance existing β cells,” Ferrer says. Melton says his lab has iPS cell lines from people with diabetes—both type 1 and type 2, in which the β cells are not destroyed—and healthy controls. They are generating β cells from those cell lines to look for differences that might explain how the different forms of the disease develop. They will also screen for chemicals that can stop or even reverse the damage diabetes does to β cells.  

 

Melton says his son and daughter—now 23 and 27 years old—were pleased but unsurprised by his group’s progress. Reversing the parent-child role, they gently nagged him to “get going and solve the [immune-rejection] problem.”

 

By Gretchen Vogel 9 October 2014 1:00 pm

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Posted in Biology, Health

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Dendritic cells and macrophages in the kidney: a spectrum of good and evil

Nature Reviews Nephrology | Review

 

Renal dendritic cells and macrophages are key factors in the initiation and propagation of renal disease and tissue regeneration. This Review discusses the common and distinct characteristics of dendritic cells and macrophages as well as current understanding of the renal-specific functions of these important phagocytic, antigen-presenting cell types in potentiating or mitigating intrinsic kidney disease.

 

Abstract | Renal dendritic cells (DCs) and macrophages represent a constitutive, extensive and contiguous network of innate immune cells that provide sentinel and immune-intelligence activity; they induce and regulate inflammatory responses to freely filtered antigenic material and protect the kidney from infection. Tissue-resident or infiltrating DCs and macrophages are key factors in the initiation and propagation of renal disease, as well as essential contributors to subsequent tissue regeneration, regardless of the aetiological and pathogenetic mechanisms. The identification, and functional and phenotypic distinction of these cell types is complex and incompletely understood, and the same is true of their interplay and relationships with effector and regulatory cells of the adaptive immune system. In this Review, we discuss the common and distinct characteristics of DCs and macrophages, as well as key advances that have identified the renal-specific functions of these important phagocytic, antigen-presenting cells, and their roles in potentiating or mitigating intrinsic kidney disease. We also identify remaining issues that are of priority for further investigation, and highlight the prospects for translational and therapeutic application of the knowledge acquired.

 

Rogers, N. M. et al. Nat. Rev. Nephrol. advance online publication 30 September 2014; doi:10.1038/nrneph.2014.170

Figure 1: The heterogeneous but overlapping phenotype and functions of renal DCs and macrophages.

DCs are traditionally described as mediators of immune surveillance and antigen presentation, and as the primary determinants of responses to antigens—through initiation of either immune effector-cell functions or the development of tolerance. Macrophages also function as innate immune cells, predominantly through phagocytosis and production of toxic metabolites. However, the classical paradigm of DC versus macrophage phenotypes and functions is increasingly indistinct within the kidney, as these cells exhibit overlapping surface markers, functional capabilities, and ontogenic pathways. This molecular and phenotypic overlap between cell types and subsets complicates their identification and evaluation. *Marker described only in humans. Abbreviations: B-ATF-3, basic leucine zipper transcription factor ATF-like 3; BDCA-1, blood dendritic cell antigen 1; CCR, CC chemokine receptor; CSF-1R, colony-stimulating factor 1 receptor; CX3CR1, CX3C chemokine receptor 1; DC, dendritic cell; DC-SIGN, dendritic-cell-specific ICAM-3-grabbing non-integrin; ECM, extracellular matrix; EMR1, EGF-like module-containing mucin-like hormone receptor-like 1; FcγR(II/III), low affinity IgG Fc region receptor (II/III); FLT3, fms-like tyrosine kinase 3; Gr-1, granulocyte-differentiation antigen-1; ICAM-1, intercellular adhesion molecule 1; ID-2, inhibitor of DNA binding 2; IL-4R, IL-4 receptor; IL-10R, IL-10 receptor; IRF, interferon regulatory factor; Ly6(C/G), lymphocyte antigen 6(C/G); SIRPα, signal-regulatory protein α (also known as tyrosine-protein phosphatase nonreceptor type substrate 1); STAT3, signal transducer and activator of transcription 3; ZBTB46, zinc finger and BTB domain containing protein 46.

Figure 2: The ontogeny of kidney-resident DCs and macrophages.

Bone-marrow-resident MDPs differentiate into monocytes that are released to the peripheral circulation under homeostatic and inflammatory conditions. MDPs also develop into CDPs, which subsequently differentiate to pre-DCs that can migrate from bone marrow to the renal interstitial compartment via the blood, with regular turnover. Under the influence of different chemokines and growth factors, the pre-DCs differentiate to form distinct, tissue-based DC subsets (broadly characterized as CX3CR1⁺ DCs or CD103⁺ DCs) that are capable of exodus to the draining lymph nodes where they can present antigens to B cells and T cells. Monocytes can also localize to the kidney under the influence of chemokines such as CCR2, and subsequently differentiate into DCs. Pre-DCs also give rise to pDC, although the presence of pDCs in kidneys of mice is disputed (dashed arrow). Abbreviations: B-ATF-3, basic leucine zipper transcription factor ATF-like 3; BDCA-(1/2), blood dendritic cell antigen (1/2); CCL(19/21), CC chemokine ligand (19/21); CCR(2/7), CC chemokine receptor (2/7); CDP, common dendritic cell precursor; CLEC4K, C-type lectin domain family 4 member K; CLEC9A, C-type lectin domain family 9 member A; CSF-1, colony-stimulating factor 1; CSF-1R, CSF-1 receptor; CX3CR1, CX3C chemokine receptor 1; DC, dendritic cell; DC-SIGN, dendritic-cell-specific ICAM-3-grabbing non-integrin; FLT3, fms-like tyrosine kinase 3; FLT3LG, FLT3 ligand; GM-CSF, granulocyte-macrophage colony-stimulating factor; Ly6(C/G), lymphocyte antigen 6(C/G); M-CSF, macrophage colony-stimulating factor; MDP, monocyte–DC precursor; pDC, plasmacytoid DC.

Figure 3: Renal DC function in health and disease.

DCs perform homeostatic functions, including induction of tolerance to peripheral antigens typically cleared by the glomerulus, such as albumin, and anti-infection immunosurveillance. Interaction of DCs with bacteria causes them to generate chemokines to attract effector cells, such as neutrophils. The kidney-resident DCs also operate to exacerbate (proinflammatory DCs) or mitigate (anti-inflammatory DCs) a wide range of parenchymal disease, and the role of these cells in disease might be determined by either tissue-resident cells or influxing cells and antigens. For example, the responsiveness and maturation state of DCs might be regulated by ongoing interactions with tubular epithelial cells. Abbreviations: AKI, acute kidney injury; DC, dendritic cell; IRI, ischaemia–reperfusion injury; LPS, lipopolysaccharide; NTN, nephrotoxic nephritis; SLE, systemic lupus erythematosus; T_H17, type 17 T-helper; UUO, unilateral ureteral obstruction.

Figure 4: Macrophages in renal disease.

Tissue-resident macrophages or infiltrating proinflammatory monocytes can become classically activated by exposure to danger-associated molecular patterns or proinflammatory cytokines to take on an M1 phenotype, associated with production of IL-12 and IL-23, engagement of T cells for antigen presentation, activation or exacerbation of profibrotic parenchymal changes, and direct and indirect tissue injury. M1 macrophages can be reprogrammed to become alternatively activated M2 macrophages by stimulation with anti-inflammatory cytokines, such as IL-10 or CSF-1, or ingestion of apoptotic cells. M2 macrophages might facilitate and coordinate restoration of tubular cell and, therefore, kidney tubule integrity following injury. M2 macrophages can also express anti-inflammatory mediators, such as HO-1 and IL-10, which act to limit tissue injury and promote resolution of inflammation, but might also drive pericyte and myofibroblast activation through production of TGF-β, galectin 3 and PDGF. Abbreviations: AKI, acute kidney injury; CSF-1, colony-stimulating factor 1; ECM, extracellular matrix; HO-1, haem oxygenase-1; ICAM-1, intercellular adhesion molecule 1; M-CSF, macrophage colony-stimulating factor; PDGF, platelet-derived growth factor; TGF-β, transforming growth factor β; Wnt7b, wingless-related MMTV integration site 7B.

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