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The Greek Crisis and the Fourth International (4) – for a Left Government?
Posted on September 3, 2018 by irishmarxism
The perspective of the FI majority in the fight against austerity was based on "the importance of forming a government to the left of Social Democracy in the next election for workers in Greece and throughout Europe. The arrival of such a government could increase their self-confidence and contribute, under certain circumstances, to a new rise in struggles."
However the FI majority reject any charge of their having illusions in Syriza, because they called for a government that included the Greek Communist Party (KKE) and the anti-capitalist Antarsya, which included the Greek section of the FI. It absolves itself of responsibility for the Syriza betrayal and its demoralising impact because it was only "under certain circumstances" that "such a government could increase their self-confidence and contribute . . . to a new rise in struggles."
Neither of these disavowals is very convincing, since it supported the Syriza government regardless of whether it included the KKE and Antarsya, and it supported it under the circumstances given at the time and not "under certain circumstances."
The starting point of their perspective appears to be twofold, the first being that "the question that we were continually posing was the unity of the radical left":
"The challenge is clear and decisive: it is necessary to defeat the Greek right and far right and to do everything so that the Greek left, of which Syriza is the main component, wins these elections, in order to create a social and political dynamic for a left government, which must strive to bring together all the forces ready to break with the austerity policy . . . This government must be a government of the lefts . . . "
The second started from "trying to put forward a comprehensive political response that went beyond propaganda in a situation where the movement was raising the question of a political response and our positions obviously corresponded to positions in the Greek radical left."
For the FI majority this left government would have ''to take anticapitalist measures, of incursion into capitalist property, nationalization of the banks, and certain key sectors of the economy, reorganization of the economy to satisfy elementary social needs. To impose these solutions, social mobilization, workers' control, self-organization and social self-management are essential. Finally the conquest of the government, within a parliamentary framework, can, in exceptional circumstances, be a first step on the path to an anticapitalist rupture but, there too, this can be confirmed only if government anti-austerity creates the conditions for a new power being pressed on Popular Assemblies, in the companies, the districts and the cities."
It is ironic that the FI majority dismisses the alternative of the Greek section because the self-organisation it depended on was "limited and marginal', yet its own perspective depended on "social mobilization, workers' control, self-organization and social self-management [which] are essential" in order "to impose these solutions."
In other words, under the existing circumstances, and given the steadfast opposition of the EU etc., the perspective of the FI majority could not succeed, even in its own terms. Not unless it expected Syriza to whip up mass self-organisation, and with a view to seeking to challenge the Greek capitalist state in a situation of dual power.
But to have believed this really would have been to betray the illusions in Syriza that they are keen to deny.
The FI majority claim that the Greek section itself endorsed such an approach, and it records an article by one of its members which said that in the "situation in Greece, the watchword of workers' government is becoming relevant. It is obviously not applicable now: it is even difficult to predict at the present time the possible composition. Such a government should be able to implement an emergency program to fight the crisis, ready to implement key transition measures, for example by expropriating banks and other sectors of the economy."
But what this shows is that the Greek section considered this perspective and rejected it. Unfortunately, it also shows the illusions that both the Greek section and the FI majority have in such a perspective. And this is important because the FI majority are correct when it says "our approach to Syriza and the governmental question in 2012 was not an illusion, a hope, but an analysis of the importance of the issue and the need for concrete policy answers. This is a fairly standard approach for revolutionary Marxists."
It is my argument that this "standard approach" is wrong. Both opposition and FI majority betrayed illusions in the potential of the state to carry out revolutionary anti-capitalist measures, which only the working class has an interest and capacity to perform.
The Greek section stated that a workers' government "should be able to implement an emergency program to fight the crisis, ready to implement key transition measures, for example by expropriating banks and other sectors of the economy."
For the FI majority the crisis required "the implementation of transitional demands" and the left government would have ''to take anticapitalist measures, of incursion into capitalist property, nationalization of the banks, and certain key sectors of the economy, reorganization of the economy to satisfy elementary social needs. . . . Finally the conquest of the government, within a parliamentary framework, can, in exceptional circumstances, be a first step on the path to an anticapitalist rupture but, there too, this can be confirmed only if government anti-austerity creates the conditions for a new power being pressed on Popular Assemblies, in the companies, the districts and the cities."
As we can see, both perspectives required the self-organisation and mass mobilisation of the working class, when these were not of sufficient breadth or strength to make their perspectives viable. And while both assign different weights to such a requirement, both assign a role to a left/workers' government that glosses over the reality that such a government would sit on top of a capitalist state, and a capitalist state is not going to "take anti-capitalist measures", in the words of the FI majority, or "implement key transition measures" in the words of the opposition.
In both perspectives it is clear that a workers' government would play a leading role in what would be, as the FI majority note, "exceptional circumstances", circumstances which somehow however become the basis for "a fairly standard approach for revolutionary Marxists".
What should bethe standard approach involves recognition that the capitalist state, workers' government or not, cannot be the mechanism for workers' self-emancipation or even the overthrow of capitalism. Electoral considerations are important but entirely secondary to the development of working class power and its eventual overthrow of capitalism, to be replaced by what used to be called the dictatorship of the proletariat.
None of this means that it is impermissible not to call for a vote for reformist parties or not to call on them to form a government, or to make demands on them to protect and advance workers' interests. It does not prevent Marxists defending such governments when attacked from the right, if this too is necessary to defend workers interests and their self-organisation. But this is a world away from seeking to make a left/workers' government the lynchpin of the overthrow of capitalism and introduction of working class rule.
This "standard approach" has been informed by the debate inside the Third and Fourth Internationals on Workers' and Workers' and Farmers Governments, and I will briefly discuss this in the next and final post.
In the meantime, nothing I have said automatically precludes supporting a vote for Antarsya, or on the other hand, supporting the fight for a Syriza Government. Neither does it mean that both are principles to be defended in all cases. So, it may be correct to have voted for Syriza or to vote against it when it imposes austerity. These are mostly questions of tactics and are in general determined by what will advance the self-organisation and consciousness of the working class and its opposition to capitalism and its state.
What the views above do reject is what the opposition describes as "the main conclusion" of the Greek experience, which, it says, is "the need for political and organizational independence from reformism".
There is however no absolute requirement to be organisationally independent or separate from reformist organisations. The following view of the opposition must therefore be rejected:
"A very old and dogmatic concept was repeated here: revolutionaries should stand alongside the working class in labour parties so as to gain their trust, and be ready to lead them out of those parties when the leadership betrays them . . "
What is dogmatic is to rule out that "revolutionaries should stand alongside the working class in labour parties." As I argued in a previous post, it is necessary always to stand beside the mass of the working class, whether in reformist labour parties or not. Only in this way can Marxists play a role in the organisation of the workers within them, and the process of their becoming more than reformist.
Whether this meant being inside Syriza or not is not a question I can answer, and again is a tactical one. But what I can say, is that if the decision whether or not to do so was based on the mistaken view above, there can be no confidence that the decision reached on this choice was the right one.
Back to part 3
This entry was posted in Fourth International, Greece, Syriza and tagged Fourth International, Greece, World Congress of Fourth International by irishmarxism. Bookmark the permalink.


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THE EVANGELICAL CHRISTIAN IS AIMING FOR THE VICE PRESIDENCY OF THE USA
US presidential elections home stretch. Mike Pence — Governor of Indiana and Evangelical Christian, the candidate for Vice-President. He recently said in an exclusive interview with CBN why he believes that as President, Donald trump is against abortion. He also stated that, in his opinion, the Bible talks about the importance of voting.
In Pennsylvania it is expected the decisive battle of the candidates, and the candidate in Vice-presidents of the USA from the Republican party, Mike Pence does not stint promises: "America must be strong. And when Donald trump becomes President of the United States, we will bring to the world stage of American power," he says.
Before election day is in a month, and Mike Pence, representing trump understands that it will be hard work. He said "CBN News" that they are not just fighting with rivals of Clinton and Kane.
The question CBN News "do you Believe that mainstream media in this country want the next President of the United States is Hillary Clinton?" he said, "I Admit, it's amazing to me. Of course, I already knew about the liberal bias of the media and personally faced with it, but I must tell you that ever since I joined this campaign, I see that the main national mass media from day to day doing half of the job Hillary Clinton for her!"
Therefore, according to Pence, they will have to work harder to appeal to voters directly, especially in swing States such as Pennsylvania, where they could still convince someone.
"I think trump good intentions, but sometimes it's hard for him to concentrate. But the Pens did well to outline their vision and unveil the intentions behind the rhetoric, says Tyler Molison undecided voter. — So I think that in some sense it helps to reassure the voting public that is worried about something".
Until November 8, you will see Mike Pence on the stock, similar to that held in Gettysburg, Pennsylvania. It is home to many workers and employees. It is precisely those voters who need Donald Trump and Mike Pence at sites on election day. And they try to convince Evangelical Christians that Trump can be trusted on issues of importance to them.
"In regard to such matters as the sanctity of the right to life, I'm extremely proud of being an associate of Donald trump, presidential candidate of the United States, bravely competing in this race in defense of life," says Mike Pence.
As President, will be whether trump against abortion?
"I assure you, I heard his remarks about it in public and in private. Donald trump defends the sanctity of the right to life," says Mike Pence.
Another question is, will the conservative evangelicals to the stations on the election day. Pence wants to give the congregation one biblical advice: "I really believe that obedience to the authorities referred to in the Bible, involves our participation in the electoral process. Get information and vote as you see fit".
Regardless of the outcome, Pence believes that the country needs a spiritual shaking: "it is very important that we pray. I believe that what was true always true today: if we accept His people, which are called by His name, and pray, He will hear from heaven and heal their land," he says.
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Primera División 1958 – edizione della competizione di calcio maschile argentina
 Primera División 1958 – edizione della competizione di calcio maschile venezuelana

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Inspired by a Jamaican Folk song, called Evening Time, this painting is about the people singing as they walk up the mountain towards home after a hard day's work. Caymanians have an afinity for the sea for it sustains and sooths after a long stressful day. Special Note: Randy will be donating 5% of his proceeds to help a young Caymanian go to college.
I was sitting on the beach with artist Nickola McCoy Snell at the GrapeTree Café in Bodden Town. She was in the midst of sharing some stories of her childhood friends and how she'd like them to be involved in Travel Gal too. Of course, one of those friends was Randy, who just arrived at the scene.
Nickola waved him over. Up walked Randy and his spunky wife Nasaria. Randy is an amazing self-taught artist on the island (you'll be able to check out his work on travelgalart.com soon). And his wife, an artist in her own right, was the educational coordinator at the National Museum.
Now what you might not know unless you read my older posts, is that this moment on the beach was about 3 months in the making. After my failed attempts to connect with any artists in Grand Cayman via email prior to my arrival on the island, I went on a weeklong search for Nickola. And I had all but given up when I was suddenly connected to her through another artist I met along the way. Honestly, I had lost all hope at that point. And I had given up on the idea of even working with any artists from the island. Now, here I was, literally 24 hours after throwing my hands up in the air, surrounded by some of the islands most premiere talents. How's that for serendipity?
Once Randy and Nasaria arrived the energy rose to a whole new level. I got to share my hopes for Travel Gal with them and they entertained me with the stories of the island's East vs. West cooking competition. "People on the West side are always running so fast, they don't take the time to do things right," said Nasaria. This lead into a spirited discussion on the difference between how Turtle and Fish are prepared on the West side in comparison to the East side.


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African Youth Development Summit
Editor Dec 1, 2018 1330 0
The Deputy Secretary-General Remarks to the African Youth Development Summit.
JOHANNESBURG, South Africa, November 30, 2018/ -- It is wonderful to be here today at the African Youth Development Summit to talk with you about youth leadership, the participation of young women and men in Africa's development, and the obstacles that stand in their way.
Globally, today's young people are a historic phenomenon — the largest cohort of young people that our world has ever seen.
There are 1.8 billion young people between the ages of 10 and 24 in the world today, and Africa is the youngest continent, where young women and men constitute more than 60% of the total population.
But young people face serious challenges in today's world. They are often at the forefront of global trends, including global challenges.
In some countries, young people are confronted by conflict and terrorism.
In others, they suffer the effects of economic exclusion and growing inequalities, deprived of quality education, skills and health services.
Young people are seeing their pathways to participation blocked and their rights denied.
Perhaps most damaging of all, young people are witnessing the manmade destruction of our natural environment. They can see that governments and others are failing to respond to the challenge of climate change with the urgency and ambition that is needed. They know that for the sake of their future, we need to transform our economies and embrace new, sustainable patterns of production and consumption.
But, as Madiba once said, "When people are determined, they can overcome anything".
And with the 2030 Agenda for Sustainable Development and the African Union's Agenda 2063, we now have road maps for social and economic transformation, around which we can unite, and demand and drive change at local and global levels.
Dear young people, dear friends,
The Africa We Want will not be possible without the full participation and of Africa's young people, particularly those who face barriers – including young women and girls.
For decades at the United Nations, we have sought to work with and for young people.
But today, at a time of mounting attacks on multilateralism and international cooperation, we recognize that our best hopes for global peace and prosperity on a healthy planet lie in the hands of young people. We need your leadership, participation and engagement, right across the globe.
That is why in September this year, the Secretary General launched the United Nations Youth Strategy: Youth 2030. A strategy led by a young woman with courage of her convictions – our Special Envoy Jayathma.
It seeks to make the UN a true leader in working with and for young people: in understanding their needs, in helping to put their ideas into action, in ensuring their views inform our processes both at country level and at UN Headquarters.
Youth 2030 will strengthen the United Nation's ability to support young people – their participation in society, their ability to secure employment, the realization of their rights and their role as agents of peace in fragile contexts.
The Strategy highlights access to education and health services, as well as climate action, which are foundational pillars for young people seeking to advance in the world.
We are delighted to support women and men as they raise their voices and lead the transformational change that is needed – from communities to governments and on the international stage.
Campaigns promoting access to sexual and reproductive health services are tackling important issues like menstrual health, that have been a taboo subject for far too long.
I am pleased that we are highlighting one such campaign here today.
Lack of support for menstrual health can keep girls out of school and the workplace and out of leadership roles, with devastating consequences that can last lifetimes and across generations. It contributes to discrimination and to the marginalization of women and girls.
Educating both girls and boys about menstruation as a normal biological process is the first step towards addressing these issues.
Providing affordable and environmentally-friendly sanitary products and clean water to manage menstrual hygiene and reduce health risks is a very significant contribution. I experienced this first-hand while travelling to India with the Water Supply & Sanitation Collaborative Council.
Today, some 60 million 10-year-old girls around the world stand at the threshold of adolescence and menstruation. Let's do everything we can to give them control over their bodies and empower them, their families, their communities and ultimately, their nations.
My final message to all of you gathered here today and for those following online via social media:
I am counting on you to be the change agents and torchbearers that we so desperately need. The dignity that we want for our young women and men begins with each and every one of us. It will be a journey, the outcome of which will depend on how you travel that road to 2030.
I believe that with the amazing power of Africa's young people, the Africa We Want is possible.
Let's make it happen.!
Distributed by APO Group on behalf of United Nations - Office of the Spokesperson for the Secretary-General.
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what is kevlar
Kevlar Patents ,The person behind the invention of Kevlar is Stephanie Kwolek, an expert in chemical compounds and an employee for the DuPont Company. It's important to note that even Kevlar has its limits. Kevlar is a very strong material, in the aramid family of nylon polyamides. the material used to make bulletproof vests. Aromatic polyamide (aramid) threads are the result. Kevlar is hand protection in the context of workplace safety equipment. What makes Kevlar such a good antiballistic material? true of artificial fibers such as nylon, Kevlar, and Nomex. That's why we now rely on According to Please do NOT copy our articles onto blogs and other websites. C'est un polymère de la famille des aramides ou polyamides aromatiques, dont le nom scientifique est poly-para-phénylène téréphtalamide. stand outside in the snow all winter. Some say that Kevlar is stronger than steel by at least five times. Photo: Kevlar textiles get their properties partly from the inherent strength La synthèse du poly(p-phénylènetéréphtalamide) consiste en la polycondensation du p-phénylènediamine (PPD) (ou son chlorhydrate) et du chlorure de téréphtaloyle, en milieu solvanté et à très basse température. It's worth noting that Kevlar also has its drawbacks. bulletproof glass do a good job at protecting us by absorbing (soaking up) Aramid is a class of strong, synthetic fibers. It is a synthetic polymer that is almost 5 times stronger than any ordinary steel. 2. bigger, faster bullets than smaller, slower ones. The fibers are then cut to length and woven into a tough mat to make the having been discovered in the early 1960s by US chemist So the first step is all about chemistry; Higher up the scale, type IIIA armor has to resist more powerful handheld bullets But it is not so. Ce matériau était cependant prévu à l'origine pour le renforcement de pneus, et ses réelles capacités antibalistiques étaient inconnues. Kevlar has an incredibly high tensile strength that is 8 times stronger than steel wire. Kevlar®: Avoid bleach: Bleach destroys the strength of Kevlar® fibers, as seen below. dozens of other applications as well. Kevlar is simply a super-strong plastic. La fibre est ensuite refroidie par jet d'air, lavée, séchée et mise en bobine. strong, and stiff fibers that are wound onto drums. La composition chimique de cette fibre aramide est proche du Explorer . photo shows you why: it's a piece of Kevlar after being hit by a projectile. Read more here! Kevlar fiber has a tensile strength comparable with that of carbon fiber, a modulus between those of glass and carbon fibers and lower density than both. Kevlar®'s properties. Created by Stephanie Kwolek, DuPont ™ Kevlar ® is a heat-resistant para-aramid synthetic fiber with a molecular structure of many inter-chain bonds that make Kevlar ® incredibly strong. ammonia-like chemical and react it with an organic acid. super-strong, super-stiff finished material we know as Kevlar. The bigger the bullet and the faster it's travelling, the more kinetic energy chemical company and it comes in two main varieties called Kevlar 29 Kevlar is considered as a magic modern material which many find useful. Kevlar can resist attacks from many different chemicals, though long exposure to strong Why Kevlar® Gloves made with Kevlar®—Dare to protect. Chart: You need a greater thickness of Kevlar body armor to stop higher-speed (velocity) Kevlar will be something cool. Kevlar is one of those magic modern materials people talk about all the time without ever Plus solide que l'acier. It is used in bulletproof vests, in composites for boat construction, in lightweight mountaineering ropes, and for lightweight skis and racquets - amongst many other things. But it is not so. 3) The wet-spinning process causes the rods to straighten out fully and align so they're all oriented in the same direction—forming what's called a nematic structure—and this is what gives Kevlar its exceptionally high strength. Articles from this website are registered at the US Copyright Office. It is usually made into composites. Cette phase permit ainsi la création du premier prototype pouvant protéger contre les principales menaces des policiers de l'époque, soit les calibres .38 Special et .22 Long Rifle. reasonably good at withstanding temperatures and decomposes only at (or a knife pushed hard by an attacker) has its energy "stolen" from it as it tries to fight its way through. Afin de régler ces problèmes, les gilets pare-balles sont maintenant recouverts d'un matériau étanche à l'eau, résistant aux UV et aux agents de nettoyage. a box (only without the box). renforts de vêtements plus ou moins importants en fonction de la personne ; voiles et coques de bateaux (résistance à l'élongation et aux solutions alcalines) ; domaine aéronautique, aérospatial (ailes d'. It uses a black-box testing methodology to ensure you maintain perfect backwards compatibility as … Kevlar® is a material formed by combining para-phenylenediamine and terephthaloyl chloride. Like Nomex, Kevlar is a distant relative of nylon, the first commercially successful "superpolyamide", developed by DuPont in the 1930s. The higher melting point of Kevlar, as well as its greater stiffness and tensile strength, partly results from the regular para-orientation of its molecules. Picture by Casey H. Kyhl courtesy of especially in the modern world where the challenges we face are ones Kevlar is responsible for saving the life of a police officer in Orlando Kevlar is a great anti ballistic (or bulletproof material) Kevlar is used for bulletproof vests and helmets that the USA's military personnel uses and police officers use. Kevlar material is a popular strong, heat resistant material that has many benefits. Because the fabric has no melting point and is extremely inflammable, its greatest strengths are that it can stand the heat and keep people and products safe from said heat. Source: "US Patent: 3287323: Process for the production of a highly orientable, crystallizable, filament" by Stephanie Kwolek et al. And you should clear all your myths and doubts before getting started with it. Kevlar is a synthetic material made of plastic that can withstand knives and bullets. Kevlar® reinforced military helmet. in a NASA, Photo: Braided Kevlar can be used to make super-strong rope. degradation" down to −196°C (−320°F). At this stage, the molecules are still tangled up and not extended into straight, parallel chains. It is light and incredibly strong – in fact, five times stronger than steel when considered on an equal weight basis. bridges, and other structures where compressive forces are common. Here is the biggest avoidance for each fibers: How to Wash Dyneema®: Avoid heat: Washing temperature shouldn't exceed 104°F and tumble dried on low. This is a condensation reaction that is followed by extrusion, stretching and drawing. Kevlar is classified as an aramid compound and is considered to be part of the nylon family of fiber textiles. before they can turned into useful textile products—and the same is Kevlar is very lightweight and yet with very good strength. There are literally hundreds of synthetic plastics made by Kevlar can be used by itself or as part of a composite material (one material combined with as ordinary nylon fiber. It's probably best known for For protection against rifle bullets (ordinary ones or armor-piercing ones), which travel much faster (850–900 m/s or 1900–2000 mph) with considerably higher kinetic energy, Kevlar isn't enough: you need body armor made from steel or ceramic plates (classified as type III and IV). Un article de Wikipédia, l'encyclopédie libre. Kevlar is made from reacting paraphenylene diamine with terephthaloyl chloride in an organic solvent to form polyparaphenylene terephthalamide (aramid). Il existe plusieurs grades de Kevlar : Kevlar, Kevlar 52, Kevlar 49, etc. Typically it is spun into ropes or fabric sheets that can be used as such or as an ingredient in composite material components. La fibre du Dyneema (polyéthylène) est moins dense (d~0,97) que la fibre en Kevlar (d~1,45). Picture by Casey H. Kyhl courtesy of. Le polymère est dissous dans l'acide sulfurique et extrudé dans une filière, puis étiré et filé à haute température. by Chris Woodford. Le Kevlar est une fibre synthétique mise au point à partir de 1965 par Stephanie L. Kwolek (1923-2014), chimiste de la firme américaine Du Pont de Nemours, et commercialisée en 1972. Although there's no such thing as completely "bulletproof," materials like Il n'est toutefois pas aussi performant que la fibre de carbone qui peut atteindre une résistance à la traction de 7 000 MPa (fibres très haute résistance) et un module de 520 GPa (fibres très haut module). In theory, the thicker the Kevlar, the shorter the distance a bullet should be able to pass through (the shorter the penetration depth); in practice, it's a little bit more complicated than that. building material in things like buildings, Kevlar is the registered trademark for a para-aramid synthetic fiber, related to other aramids such as Nomex and Technora. The resulting textile material is extremely strong, lightweight, corrosion and heat resistant. really explaining any further. Le PPD-T possède différentes propriétés intéressantes, telles la résistance à la chaleur (résiste à des températures pouvant aller jusqu'à 400 °C, il se décompose au-delà)[précision nécessaire], à l'élongation ou au cisaillement. Polyamides like Kevlar are polymers (huge molecules made of many identical parts joined
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what is kevlar 2020


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Research | Open | Published: 01 July 2015
Abnormal CD4 + T helper (Th) 1 cells and activated memory B cells are associated with type III asymptomatic mixed cryoglobulinemia in HCV infection
Fanyun Kong1,2,
Wei Zhang1,
Bo Feng1,
Henghui Zhang1,
Huiying Rao1,
Jianghua Wang1,
Xu Cong1 &
Lai Wei1
Mixed cryoglobulinemia (MC) in hepatitis C virus (HCV) infection is associated with abnormal immune responses mediated by T cells and B cells, while the relationships of different subsets of CD4 + T helper (Th) cells, B cells and associated cytokines with type III asymptomatic MC in HCV infection are poorly understood.
Fifty-four chronic hepatitis C (CHC) patients and 23 healthy controls (HCs) were enrolled in the study. Serum cryoglobulins were detected by cryoprecipitation. The types of cryoglobulin were determined by western blot. The phenotypes and frequencies of Th cell and B cell subsets were detected by flow cytometric analysis. The cytokines IFN-γ, IL-4, IL-17, IL-21, IL-22, and TGF-β were measured by enzyme-linked immunosorbent assay.
Twenty-six CHC patients were detected with type III asymptomatic MC. The frequencies of Th2, Th17, follicular helper T (Tfh cells), Th22, and tissue-like B cells were significantly higher in CHC patients compared to HCs, while these cell subsets were not significantly different between CHC patients and HCV-related MC patients. The frequencies of Th1 and activated memory B cells increased in HCV-related MC patients compared to HCs, although the difference between the two cell subsets in CHC patients and HCs was not significant. The frequency of regulatory T cells (Treg cells) was higher in CHC patients than in HCV-related MC patients and HCs. Higher expressions of serum IFN-γ, IL-17, IL-21, and IL-22 were observed in CHC patients than in HCs, but the differences were not significantly different in CHC patients and HCV-related MC patients. The frequency of Th1 cells was associated with activated memory B cells in HCV-related MC patients, and the frequency of Th1 cells and activated memory B cells was closely related to HCV RNA in HCV-related MC patients.
The increased frequencies of Th17 cells, Tfh cells, Th22 cells, Treg cells, cytokines IL-17, IL-21, IL-22, and tissue-like B cells, were related to HCV infection but not type III asymptomatic MC. Higher frequencies of Th1 cells and activated memory B cells were associated with type III asymptomatic MC in HCV infection.
Chronic hepatitis C virus (HCV) infection is not only responsible for hepatic diseases but is also associated with several extrahepatic manifestations. Mixed cryoglobulinemia (MC), an autoimmune disease characterized by the formation of cold-precipitable cryoglobulin complexes composed of immunoglobulins, is recognized as the most common extrahepatic disease induced by HCV infection [1, 2]. Based on the immunoglobulin composition, MC has two types of cryoglobulins: Type II cryoglobulins consisting of polyclonal IgG and monoclonal IgM with rheumatoid factor (RF) activity, and type III cryoglobulins characterized by polyclonal IgG with polyclonal IgM. In HCV-related MC, the majority of patients have asymptomatic MC, and only 10 –15 % of these patients will develop cryoglobulinemic symptoms, characterized by small vessel vasculitis, glomerulo-nephritis, and neuropathy due to immune complex deposition and activation of the complement cascade in small blood vessels [3, 4].
To date, the involvement of host immune responses mediated by T cells and B cells with MC in chronic hepatitis C (CHC) patients is poorly understood. Several cytokines and chemokines, including IL-6, TNF-α, IL-1β, IFN-γ, MIP-1α, CXCL9, CXCL10, and CXCL11 associated with T helper (Th) 1 immune responses, were found to be involved in HCV-related type II MC [5–13], especially in patients with autoimmune thyroiditis or active vasculitis. Theoretically, agents that selectively neutralize molecules such as CXCL10, might increase patient responsiveness to HCV therapy [14]. Recent evidence also showed a deficiency of CD4 + CD25+ regulatory T (Treg) cells in HCV-related MC patients with active vasculitis [15]. In addition, a highly restricted T cell receptor variable β gene repertoire in liver and peripheral blood was found in HCV-related MC patients, suggesting that the clonally expanded TCR Vβ repertoire was associated with HCV-related MC [16].
MC is considered to be a benign B-cell proliferative disorder. The expansion of IgM + CD27 + CD21-B cells, which express VH1-69 and VH4-34 genes, are present in type II MC patients with HCV infection [17, 18], and effective treatment for HCV-related MC decreased the number of VH1-69 + B cells [19]. In addition, the expansion of immature B cell subsets and activated memory B cells with distinct activated markers such as CD86 and CD71 were noted in HCV-related MC patients [20]. In contrast, naive B cells in MC patients with HCV infection are decreased because they have a higher sensitivity to apoptosis, and treatment of MC with rituximab restored normal B cell compartments [21]. This suggested that B cells play an important role in the pathogenesis of MC with HCV infection.
Th cell subsets such as Th2 cells (IL-4 + CD4 + T cells), Th17 cells (IL-17 + CD4 + T cells), Th22 cells (IL-22 + CD4+ T cells) and Treg cells are associated with HCV infection [22–25]. To our knowledge, no study has evaluated the immune phenotype of Th cell subsets and their relationship with B cells in CHC patients with type III asymptomatic MC. The identification of immunological markers associated with T and B cell immune responses in type III asymptomatic MC may help to improve clinical monitoring and disease management. To characterize the features and relationships of peripheral Th cell and B cell subsets in CHC patients with type III asymptomatic MC, 26 CHC patients with type III asymptomatic MC, 28 CHC patients without MC and 23 sex- and age-matched healthy controls (HCs) were enrolled in the study. We analyzed different immune cell subsets and Th cell cytokines to identify distinct phenotypes associated with type III asymptomatic MC in HCV infection.
Increased percentage of Th1 cells in type III asymptomatic MC with HCV infection
Among 54 CHC patients, 26 CHC patients were tested positive for type III MC, and none of these MC patients had cryoglobulinemic syndrome (Table 1). To assess the potential role of different subsets of T cells, especially Th cell subsets, in type III asymptomatic MC with HCV infection, total T cells, CD4+ T cells, and CD4+ T cell subsets, including Th1 cells (IFN-γ + CD4+ T cells), Th2 cells, Th17 cells, Th22 cells, and Treg cells (CD4 + CD25 + Foxp3+ cells) were analyzed in CHC patients, HCV-related MC patients, and HCs. We did not note any difference in the frequencies of CD3+ T cells and CD4+ T cells in the three groups (Fig. 1b-c). The frequency of Th1 cells in total CD4+ T cells was significantly higher in MC patients compared with HCs (Fig. 1d). Though a higher frequency of Th1 cells was found in CHC patients, the difference was not significant between CHC patients and HCs. The percentages of Th2 cells, Th17 cells, and Th22 cells in total CD4+ T cells were increased in both CHC patients and HCV-related MC patients compared with HCs (Fig. 1e-g). The frequencies of Th2 cells, Th17 cells, and Th22 cells were not significantly different between CHC patients and HCV-related MC patients, suggesting that higher frequencies of Th2 cells Th17 cells, and Th22 cells were associated with HCV infection but were not related to MC. CD4+ T cells are a main source of IL-21, a cytokine responsible for the activation and differentiation of T cells and B cells [26, 27]. We also measured the frequency of IL-21 + CD4+ T cells in three groups, and found their percentage was increased in CHC patients and HCV-related MC patients (Fig. 1h). However, there was no correlation between the frequency of IL21 + CD4+ T cells and MC. We next detected the frequency of Treg cells in three groups, and found their frequency was higher in CHC patients than in HCV-related MC patients and HCs. However, the frequency of Treg cells was not significantly different between HCV-related MC patients and HCs (Fig. 1i). The ratio of Th1 cells/Th2 cells was also examined in three groups, but there was no significant difference among these groups (Fig. 1j). Taken together, these results indicated that Th2 cells, Th17 cells, Th22 cells, IL21 + CD4+ T cells, and Treg cells were only associated with HCV infection but not MC, while Th1 cells were related to type III MC in HCV infection.
Table 1 Baseline characteristics of CHC patients, HCV-related MC patients, and HC
Frequencies of CD4 + T cell subsets in CHC patients and HCV-related asymptomatic MC patients. a The gating strategy for T cell subsets by flow cytometric analysis. b The frequencies of total T cells. c The frequency of CD4 + T cells in total T cells. d, e, f g, h, and i show the frequencies of IFN-γ + CD4+ T cells, IL-4 + CD4+ T cells, IL-17 + CD4+ T cells, IL-22 + CD4+ T cells, IL-21 + CD4+ T cells, and Treg cells (CD4 + CD25 + Foxp3+ cells) in total CD4+ T cells. j Shows the ratio of Th1 cells to Th2 cells. *p <0.05
Increased frequency of activated memory B cells in type III asymptomatic MC with HCV infection
Current studies have reported abnormal B cells in type II MC with HCV infection [17, 18, 20], while few studies have investigated the characteristics of B cells in HCV-related type III asymptomatic MC. As shown in Fig. 2b, the frequencies of total B cells and memory B cells (CD27+ B cells) were decreased in CHC patients. The frequencies of total B cells and memory B cells were comparable between CHC patients and HCV-related MC patients. Measurement of activated B cells, including tissue-like B cells (CD27-CD21- B cells) and activated memory B cells (CD27 + CD21- B cells) in three groups indicated the frequency of tissue-like B cells were increased in both CHC patients and HCV-related MC patients (Fig. 2c), but the difference of percentages of tissue like B cells in CHC patients and HCV-related MC patients was not significant. The percentage of activated memory B cells was only increased in HCV-related CHC patients compared with HCs. Because the increased expression of CD86 and CD95 is associated with B cell activation [28], we detected the expression of CD86 and CD95 in B cell subsets. We found that the increased frequencies of CD86 + CD27- B cells and CD95 + CD27- B cells were comparable in CHC patients and HCV-related MC patients, compared to HCs (Fig. 2d-e). Increased percentages of CD86 + CD27+ B cells and CD95 + CD27+ B cells were observed in HCV-related MC patients but not in CHC patients when compared with HCs. Moreover, in HCV-related MC patients, the frequency of activated memory B cells was positively correlated with CD95 + CD27+ B cells (Fig. 2f). These findings indicate that the increased frequencies of tissue-like B cells are associated with HCV infection, while activated memory B cells with an increase percentage of CD95 + CD27+ B cells is related to type III asymptomatic MC in HCV infection.
Frequencies of B cell subsets in CHC patients and HCV-related asymptomatic MC patients. a The gating strategy for B cell subsets by flow cytometric analysis. b The frequencies of total B cells and memory B cells. The frequencies of c CD21- B cells in CD27- B cells and CD27+ B cells; d CD86+ B cells in CD27- B cells and CD27+ B cells; and e CD95+ B cells in CD27- B cells and CD27+ B cells. f The relationships between the frequency of CD27 + CD21- B cells, CD86 + CD27 + IgG+ B cells and CD95 + CD27 + IgG+ B cells.*p <0.05
Increased frequency of circulating Tfh cells is associated with HCV infection but not type III asymptomatic MC
Tfh cells (CXCR5 + CD4+ T cells), a major subset of effector CD4 + T cells, are associated with the production of IL-21 and are responsible for the activation and differentiation of B cells [29]. In peripheral blood, circulating Tfh cells were characterized by CXCR5 with ICOS or PD-1, and could be divided into different subsets based on the expression of IFN-γ, IL-4 and IL-17 [30]. Previous studies showed that Tfh cells were associated with HCV infection and autoimmune diseases [28, 31, 32]. Whether Tfh cells are related to type III asymptomatic MC in HCV infection is well understood. We examined the frequencies of different Tfh cell subsets in the three patient groups. As shown in Fig. 3b-d, the frequencies of CXCR5 + CD4+ T cells, PD-1 + CXCR5 + CD4+ T cells and ICOS + CXCR5 + CD4+ T cells in CHC patients and HCV-related MC patients were significantly higher than in HCs, although the frequencies of these Tfh cell subsets were not statistically different in CHC patients and HCV-related MC patients. We further measured different Tfh subsets including IFN-γ + Tfh cells, IL-4+ Tfh cells, IL-17+ Tfh cells, and IL-21 + Tfh cells (Fig. 3e). The results showed that IFN-γ + Tfh cells, IL-4+ Tfh cells, and IL-21+ Tfh cells were associated with HCV infection but not MC. In addition, the frequencies of IL-17+ Tfh cells were comparable in the three groups. Taken together, these results suggest that Tfh cell subsets are associated with HCV but are not involved in type III asymptomatic MC with HCV infection.
Frequencies of circulating Tfh cell subsets in CHC patients and HCV-related asymptomatic MC patients. a The gating strategy for circulating Tfh cell subsets by flow cytometric analysis. b Frequency of circulating Tfh cells in CD4 + T cells. c Frequency of PD-1+ Tfh cells in CD4+ T cells. d Frequency of ICOS+ Tfh cells in total CD4+ T cells. e Frequencies of Tfh1 (IFN-γ + Tfh) cells, Tfh2 (IL-4+ Tfh) cells, Tfh 17 (IL-17+ Tfh) cells, and IL-21+ Tfh in total Tfh cells. *p <0.05
Increased expressions of IFN-γ, IL-17, IL-21, and IL-22 are associated with HCV infection but not type III asymptomatic MC
We next detected the cytokines IFN-γ, IL-4, IL-17, IL-21, IL-22, and TGF-β, which are associated with immune responses of Th1 cells, Th2 cells, Th17 cells, Tfh cells, Th22 cells, and Treg cells. As shown in Fig. 4a-d, the expressions of IFN-γ, IL-17, IL-21, and IL-22 were higher in CHC patients and HCV-related MC patients than in HCs, although this was not significant between CHC patients and HCV-related MC patients. Compared with HCs, the expression of TGF-β was lower in CHC patients and HCV-related MC patients, but was not significantly different between CHC patients and MC patients with HCV infection (Fig. 4e). Because the expressions of IL-4 in most subjects from the three groups were below the detection limit of the enzyme-linked immunosorbent assay (ELISA) kit (15.6 pg/mL), and we could not distinguish differences in IL-4 expression levels (data not shown). In summary, these results suggested that abnormalities of IFN-γ, IL-17, IL-21, IL-22, and TGF-β production were related to HCV infection but not associated with type III asymptomatic MC.
Expressions of serum cytokines in CHC patients and HCV-related asymptomatic MC patients. a, b, c, d and e show the expression of serum IFN-γ, IL-17, IL-21, IL-22, and TGF-β detected by ELISA. *p <0.05
Frequency of Th1 cells is positively correlated with the frequency of activated memory B cells in HCV-related asymptomatic MC
Next we analyzed the relationship of the frequencies of different subsets of CD4+ T cells with B cell subsets in HCV-related asymptomatic MC. The frequency of Th1 cells was positively related to the percentage of activated memory B cells in MC patients with HCV infection (Fig. 5a). However, there were no statistically significant associations of Th1 cells with CD86 + CD27+ B cells or CD95 + CD27+ B cells in HCV-related MC patients (Fig. 5b-c). Together, the results indicated that Th1 cells were associated with the activation of memory B cells, but were not related to the expression of activation markers CD86 and CD95 in asymptomatic MC with HCV infection.
Relationship between frequency of Th1 cells and activated memory B cells in HCV-related asymptomatic MC patients. The relationship of a IFN-γ + CD4+ T cells with CD27 + CD21- B cells; b IFN-γ + CD4+ T cells with CD86 + CD27+ B cells; and c IFN-γ + CD4+ T cells with CD95 + CD27+ B cells
Frequency of Th1 cells, activated memory B cells are associated with HCV RNA in type III MC
We further measured the relationship of Th1 cells and activated memory B cells with clinical parameters in HCV-related asymptomatic MC patients. We found the frequencies of Th1 cells and activated memory B cells were positively related to HCV RNA in MC patients with HCV infection (Fig. 6a-b). In HCV-related MC patients, the frequencies of CD86 + CD27+ B cells and CD95 + CD27+ B cells were not significantly associated with HCV RNA (Fig. 6c-d). In CHC patients without MC, we failed to detect significant associations of Th1 cells, activated memory B cells, CD86 + CD27+ B cells and CD95 + CD27+ B cells with HCV RNA. We also tested the relationship of alanine transaminase (ALT), a marker of inflammatory necrosis in the liver, with the frequencies of Th1 cells and activated memory B cells in CHC patients and HCV-related asymptomatic MC patients, but the results showed no significant association (Fig. 6e-h). In summary, these results suggested that HCV may have a significant influence on the presence of abnormal Th1 cells and activated memory B cells in type III asymptomatic MC.
Relationships between frequencies of Th1 cells and activated memory B cells with HCV RNA in CHC patients and HCV-related asymptomatic MC patients. a Relationship of Th1 cells with HCV RNA in CHC patients and MC patients with HCV infection. b Relationship of CD27 + CD21- B cells with HCV RNA in CHC patients and HCV-related MC patients. c Relationship of CD86 + CD27+ B cells with HCV RNA in CHC patients and MC patients with HCV infection. d Relationship of CD95 + CD27+ B cells with HCV RNA in CHC patients and HCV-related MC patients. e Relationship of Th1 cells with ALT in CHC patients and MC patients with HCV infection. f Relationship of CD27 + CD21- B cells with ALT in CHC patients and HCV-related MC patients. g Relationship of CD86 + CD27+ B cells with ALT in CHC patients and MC patients with HCV infection. h Relationship of CD95 + CD27+ B cells with ALT in CHC patients and HCV-related MC patients
Although MC in HCV infection is associated with abnormal immune responses mediated by T cells and B cells, the relationships of different subsets of CD4+ T cells and B cells with type III asymptomatic MC in HCV infection is not well explored. We found increased frequencies of Th1 cells and activated memory B cells in HCV-related type III asymptomatic MC. In addition, the frequency of Th1 cells was positively related to activated memory B cells in HCV-related MC. Moreover, the two target cell subsets were associated with HCV RNA in MC patients with HCV infection.
MC is considered an autoimmune disease, and immune disorders mediated by T cells have been investigated by different research groups. Although reports from Cacoub et al. and Sansonno et al. showed no significant difference in the frequency and activation of T cells in CHC patients and MC patients with HCV infection [33, 34], it is still unknown whether Th cell subsets are involved in the pathogenesis of HCV-related type III asymptomatic MC. In this study, we investigated the relationship of Th cell subsets with HCV-related type III MC without cryoglobulinemic symptoms, and found that Th2 cells, Th17 cells, Th22 cells and IL-21 + CD4+ T cells, were only associated with HCV infection but not MC. A higher frequency of Th1 cells was observed in HCV-related MC patients but not CHC patients in comparison to HCs, implying that immune responses mediated by Th1 cells might have an important role in the progression of HCV-related MC. Previous studies showed that Treg cells were increased in CHC patients [25], while Boyer et al. observed a deficiency of CD4 + CD25+ Treg cells in HCV-related MC patients with active vasculitis compared with CHC patients without MC and HCs [15]. In this study, we found a higher frequency of Treg cells in CHC patients, while a reduced the frequency of Treg cells was observed in MC. In addition, we did not observe a deficiency of Treg cells in HCV-related MC patients without cryoglobulinemic symptoms compared to HCs. The difference between the study by Boyer et al. and our results might be due to the presence or absence of cryoglobulinemic symptoms, which might enhance the inhibition of Treg cells in HCV-related MC patients.
Abnormalities of different subsets of B cells, including immature B cells, tissue-like B cells and memory B cells have been described in MC [17, 18, 20, 21], in studies largely focused on type II MC with cryoglobulinemic symptoms in HCV infection. We investigated different B cell subsets in HCV-related type III asymptomatic MC, and found that the frequency of memory B cells was reduced, while the percentage of tissue-like B cells was increased in HCV infection. These results are consistent with previous studies [35–37]. In the current study, only activated memory B cells with increased expression of activation markers CD86 and CD95 were associated with HCV-related type III asymptomatic MC. Although the features and functions of activated memory B cells, specifically IgM+ memory B cells, has been elucidated in type II MC with HCV infection [17, 18], their exact function in MC patients is still unclear. The composition of cryoglobulins in type III MC is polyclonal IgG and polyclonal IgM, and IgG from memory B cells with class switching [38]. Thus, we hypothesized that activated memory B cells might be a main source of polyclonal IgG in cryoglobulins in type III MC patients with HCV infection.
Considering Tfh cells are responsible for the activation and differentiation of B cells [29], and are involved in HCV infection and autoimmune diseases [28, 31, 32, 39], we speculated that abnormal memory B cell activation may be associated with Tfh cells in HCV-related MC. Unexpectedly, we found higher frequencies of Tfh cell subsets in CHC patients and HCV-related MC patients, and Tfh cells were not related to type III asymptomatic MC. In vivo activation of B cells is dependent on BCR stimulation, coupled with help from T cells via coupling of CD40L with CD40 on B cells and cytokine secretion by T cells, or relies on endogenous TLR ligand stimulation [40]. Studies by Charles et al. and Terrier et al. showed that activated memory B cells were anergic to BCR- and CD40-mediated stimulation but not TLR9 triggering in HCV-related MC patients [18, 41], implying endogenous TLR ligands but not Tfh cells were responsible for the aberrant activation of memory B cells in HCV-related MC.
Because the diverse functions of Th cells are mainly determined by the cytokines they produce, we investigated cytokines associated with Th1 cells, Th2 cells, Th17 cells, Tfh cells, Th22 cells, and Treg cells in the three patient groups. We found that the expression of IL-17, IL-21, and IL-22 was increased in CHC patients and HCV-related MC patients, consistent with the frequencies of Th17 cells, Tfh cells and Th22 cells in these patients. We also found the expression of IFN-γ was higher in both CHC patients and HCV-related MC patients, but it was not related to the frequency of Th1 cells in these patients compared to HCs. Furthermore, the expression of TGF-β was lower in CHC patients and HCV-related MC patients, which was also not related to the frequencies of Treg cells in these patients. However, IFN-γ and TGF-β are not only produced by T cells, but can also be secreted by many different types of cells such as professional antigen-presenting cells [42, 43]; thus, IFN-γ and TGF-β from other type cells might have a significant effect on the concentration of serum IFN-γ and TGF-β in CHC patients. Although the prevalence of IFN-γ and other cytokines associated with Th1 immune responses have been observed in CHC patients with MC, other studies mainly focused on MC with cryoglobulinemic symptoms. Our results suggested that type III asymptomatic MC is only associated with a higher frequency of Th1 cells but was not related to IFN-γ in HCV infection. To better understand the effect of cryoglobulinemic symptoms on the expression of Th1 cells, IFN-γ and cytokines associated with Th1 immune responses in HCV-related MC patients should be investigated further by enrolling CHC patients with symptomatic MC in future studies. Our results are consistent with Zhang et al., who found the expression of TGF-β was lower in CHC patients [44], although other studies detected no difference or a higher expression of serum TGF-β levels in CHC patients compared to HCs [45, 46]. This suggested that the divergent genetic background of the study population might have an effect on serum TGF-β levels in HCV infection. MC inhibits the expression of Treg cells, but the exact mechanism is still unknown. TGF-β has critical roles in the differentiation and function of Treg cells [47]; however, a relationship between TGF-β and Treg cells in HCV-related MC patients is not well understood. In the study, we did not find a similar change of TGF-β expression with frequency of Treg cells in HCV-related MC patients compared to CHC patients, implying that MC might induce the reduction and dysfunction of Treg cells in a non-TGF-β-dependent manner.
We next evaluated the relationships among Th1 cells, activated memory B cells, and clinical parameters in HCV-related asymptomatic MC. The frequency of Th1 cells was associated with activated memory B cells in MC patients with HCV infection, suggesting the immune response mediated by Th1 cells might contribute to memory B cell activation in HCV-related asymptomatic MC. In addition, the percentages of Th1 cells and activated memory B cells were related to HCV RNA but not ALT levels in HCV-related MC patients. In CHC patients without MC, the relationship of Th1 cells and activated memory B cells with HCV RNA load was not statistically significant. In chronic HCV infection, changes in Th1 cells and activated memory B cells may gradually increase to levels that contribute to the development of MC disorder and until the stage where type III MC develops. Therefore, the effect of virus infection on Th1 cells and activated memory B cells should be determined.
This study highlights that a higher frequency of Th1 cells is associated with memory B cell activation in HCV-related type III MC without cryoglobulinemic symptoms. HCV RNA load is closely associated with Th1 cells and activated memory B cells in asymptomatic MC patients with HCV infection. Although a small sample number of patients is tested, our observations indicate that alterations in the frequency of Th1 cells and activated memory B cells might play an important role in the development of type III asymptomatic MC with HCV infection. Th1 cells and activated memory B cells in HCV-infected type III asymptomatic MC patients might also be used as biomarkers for clinical monitoring and disease management. To understand the complex function of Th cells and B cells in MC with HCV infection, further investigations are warranted, by using a larger population and enrolling HCV-related MC patients with cryoglobulinemic syndrome in our future studies.
Patients and controls
Twenty-six CHC patients with type III MC and 28 CHC patients without MC, and 23 HCs were enrolled in the study (Table 1). All CHC patients were infected with genotype 1b HCV. Cryoglobulinemia was defined as the presence of circulating cryoglobulins [48], and the type of cryoglobulins was assessed by western blot assay. Clinical manifestations of cryoglobulinemic syndrome such as vessel vasculitis, glomerulo-nephritis, and neuropathy were not observed in these MC subjects. Liver fibrosis was evaluated by liver stiffness measurement (FibroScan®, Echosens, Paris, France) [49], and none had severe fibrosis or cirrhosis. All subjects were negative for hepatitis B virus or human immunodeficiency virus infection. This study was approved by the ethics committee of Peking University People's Hospital and written informed consent was obtained from all subjects.
Clinical tests and peripheral blood mononuclear cell (PBMC) preparation
HCV antibodies, HCV RNA, and HCV genotypes were tested as described previously [49, 50]. PBMCs were isolated by density gradient centrifugation with Ficoll-Paque Plus (GE Health Bio-science, AB, Sweden) as described by Guo et al. [51]. PBMCs were stored in liquid nitrogen until ready for analysis.
Detection and characterization of cryoglobulins
To detect cryoglobulins, blood was drawn into prewarmed vacutainer tubes and maintained at 37 °C until the serum was separated. Then serum was incubated at 4 °C for one week and then examined daily for the presence of cryoprecipitate. After the serum containing cryoprecipitate was spun in a centrifuge for 15 min at 4 °C, the supernatant was removed, and the pellet was vortexed in ice-cold PBS three times. The sample was then resuspended in 1 mL PBS and re-dissolved by incubation at 37 °C for 1 h. To determine the type of cryoglobulins, western blot was performed, following an established protocol, with antibodies for IgM, IgG, IgA, Lambda and Kappa chains (SouthernBiotech, Birmingham, USA) [52].
Flow cytometric analysis
Phenotypic analyses of T cells and B cells were performed with anti-human monoclonal antibodies (mAbs): anti-human CD3-PerCP, CD4-FITC, CD19-PerCP and CD21-APC were from BD Biosciences (San Jose, CA, USA). CXCR5-APC, ICOS-PE, PD-1-PE, IFN-γ-PE, IL-4-PE, IL-17-PE, IL-21-PE, IL-22-PE, CD27-FITC, CD86-PE, CD95-PE, CD25-APC, and Foxp3-PE antibodies were obtained from eBiosciences (San Diego, CA, USA). Cells were analyzed by flow cytometry (BD FACSCalibur, San Jose, CA) and data was analyzed by FlowJo software (Tree Star, San Carlos, CA).
Intracellular cytokine staining
PBMCs were stimulated with 50 ng/mL of phorbol-12-myristate-13-acetate (PMA) and 1 μg/mL of ionomycin (Sigma-Aldrich, St. Louis, USA), in the presence of 1 μl/ml GolgiStop™ (BD Biosciences) at room temperature for 4 h. After washing twice with PBS containing 0.5 % fetal bovine serum, the cells were stained with CD3-PercP, CD4-FITC with or without CXCR5-APC at room temperature in the dark for 20 min. Cells were fixed and permeabilized with Fixation/Permeabilization solution (BD Biosciences) at 4 °C for 20 min. After washing two times in 1× BD Perm/Wash™ buffer, the cells were incubated at room temperature for 20 min with IFN-γ-PE, IL-4-PE, IL-17A-PE, IL-21-PE, and IL-22-PE antibodies or isotype controls. For staining of Foxp3 in CD4 + CD25+ T cells sorted from PBMCs, Foxp3/Transcription Factor Staining Buffer Set (eBioscience) was used, and detection followed the manufacturer's instructions.
The serum concentration of cytokines IFN-γ, IL-4, IL-17, IL-22, and TGF-β were detected by human ELISA kit (UCallM Biotech Co., Ltd, Wuxi, China). The detection range of IFN-γ, IL-4, IL-17, and IL-22 was from 15.6 to 1000 pg/mL. The detection range of TGF-β was from 31.3 to 2000 pg/mL. Human IL-21 ELISA kit was from eBiosciences, and the detectable concentration was 78 to 5000 pg/mL. The detection of each cytokine was in accordance with the manufacturer's instructions.
Statistical analysis was by Student's t-test, ANOVA or Mann–Whitney U-test according to data distribution using version 16.0 SPSS software (SPSS Inc. Chicago, IL, USA). The data are presented as the mean ± standard deviation (SD). Analysis of correlation between variables was performed by linear regression analysis with Prism Version 5 software (GraphPad). A p value <0.05 was considered statistically significant.
PBMCs:
Peripheral blood mononuclear cells
CHC:
RF:
Rheumatoid factor
Th:
T helper
HCs:
Health controls
HCV:
Hepatitis C virus
Mixed cryoglobulinemia
Treg:
Immunoregulatory T
Tfh:
Follicular helper T cells
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Tangye SG, Ma CS, Brink R, Deenick EK. The good, the bad and the ugly - TFH cells in human health and disease. Nat Rev Immunol. 2013;13:412–26.
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The work was approved by the National Science and Technology Major Project for Infectious Diseases Control during the 12th Five-Year Plan (grant no. 2012ZX10002003). We thank all CHC patients who generously provided blood samples, and thank Hong Qin, Qian Jin, Ying Hu, Yanhui Chen, Kai Deng, Xingliang Zhao and Hechao Li for help in the experimental process.
Peking University People's Hospital, Peking University Hepatology Institute, Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, No.11 Xizhimen South Street, Beijing, 100044, China
Fanyun Kong
, Wei Zhang
, Bo Feng
, Henghui Zhang
, Huiying Rao
, Jianghua Wang
, Xu Cong
& Lai Wei
Department of Pathogenic biology and Laboratory of Infection and Immunology, Xuzhou Medical College, 84 West Huaihai Road, Xuzhou, 221002, Jiangsu Province, China
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Correspondence to Lai Wei.
FK contributed to the study design, experimental process, statistical analysis, and drafted the manuscript. WZ, BF, HZ and LW contributed to the study design and data analysis. HR and JW contributed to the study planning and data analysis, XC contributed to the experimental process. All authors read and approved the final manuscript.
Type III asymptomatic cryoglobulinemia
CD4+ T helper cells
Activated memory B cells
Hepatitis viruses


This summer marks a milestone anniversary for the 1940's WWII Era Ball. The annual sell-out destination event will celebrate its tenth year of transporting guests back to simpler times through music, costumes, a pin-up girl contest, food, drinks and dancing. Returning for this momentous occasion is the world's most popular and sought after big band, the Glenn Miller Orchestra. The event will be held on Saturday, June 16 from 6 p.m. to 1 a.m. at the Boulder Airport. General admission is $69.
The Colorado Tiny House Festival returns for its second year from June 22 to 24. Doubling in size, the festival will feature a uniquely diverse assortment of small living structures from tiny houses on wheels to container homes, a semi-trailer conversion, yurts and more. Hosted at the Adams County Fairgrounds, this festival will blend the spirit of the tiny living movement with the most up-to-date information on simplifying your life and living greener. Attendees will also enjoy expert speaker presentations and hands-on workshops.
For more information, visit ColoradoTInyHouseFestival.com.
Denver Zoo's after-hours adult lecture series is back! On July 12 from 6:30 to 9:30 p.m., the Denver Zoo is hosting an adult-only event you won't want to miss. You're in for a wild evening, with insider perspective on how Denver Zoo cares for its animals and works to conserve their cousins around the world. Each lecture in the series will have a fun and unique theme. Light appetizers will be served and a cash bar will be available. Tickets on sale to the public four weeks from the event.
The Chamber of Commerce of Highlands Ranch invites the business community to participate in its 17th annual golf tournament on Friday, July 27 starting at 7:30 a.m. at the Arrowhead Golf Club. Enjoy meeting other business professionals, owners and community members all while playing 18-holes and enjoying a breakfast and awards luncheon. Cost is Entry fee includes 18 holes, cart, breakfast and awards luncheon. Go to HighlandsRanchChamber.org to register.


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By Jodie Michalak
The best time to buy a car is a question that occupies many consumers' minds. Whether it's a small coupe or a luxury sedan, a new car is a big expense. LoveToKnow is here to help narrow down your options and save you money.
During the holiday season, most people are pre-occupied with purchasing Christmas gifts, traveling, and visiting relatives. Edmunds.com, SmartMoney.com, and other reputable sources all seem to agree that the best time to buy a car is in December, especially close to the end of the year. Discounts and incentives are widely available as car dealerships work to achieve an annual sales goal; December is their last chance to make their required numbers. However, sometimes you have to be a bit careful since car dealerships may decrease the number of vehicles they order in anticipation of a slow period.
2. When New Models are Released
Historically, the fall season is when new vehicles for the upcoming year become available for sale. Once a new line is released, there is a big push to sell all vehicles from the previous year. As a result, higher rebates, incentives, and dealer cash programs become available to the consumer. However, new models are not just released in the fall season but throughout the year. Stay on the lookout for advertising for the new "future year" vehicle releases. Visit car manufacturer websites, including Chevrolet, Toyota, Hyundai and Honda, for the latest rebate and discount information.
3. Month's End Sales Quota
Every month, the dealership sales department work toward a quota of vehicles they must sell in order to receive a bonus or to earn a higher dealership ranking. At the end of the month, salespeople who have not met their personal monthly quotas may feel pressured to work with sales managers to bend even farther on pricing. This is a good time to find a car deal. Remember, you have the option of walking away, and that's the last thing any sales person wants to happen. Keep is in mind when you begin to negotiate on a better price.
4. Middle of the Week
During the week, most people are working their day jobs and do not have time to car shop. You can use this to your advantage since many salespeople won't have as many distractions and are eager to assist. During the week, you might feel less pressured by time and will make better decisions. Take advantage of the salesperson's time and take a couple of test drives on different models.
5. Saturday Morning
If the dealership has experienced a slow week, then morale will be a bit low and the pressure to sell increases. Saturday often becomes one of the best times to buy a car. With the stress to sell, salespeople will have more flexibility on pricing. If the dealership has its own website, check for specially-priced vehicles such as weekend specials. If you see something you like, print it out and take it with you so the salesperson knows exactly what you're looking for in a vehicle.
6. End of a Vehicle's Design Cycle
Sometimes car manufacturer's decide to discontinue a particular model. It can be due to a variety of reasons, such as a lack of interest by the consumer, high manufacturring costs, design issues or vehicle defects. These vehicles are deeply discounted, so you can save a lot of money. Research carefully, since you want to make sure the reason the vehicle is no longer available is not due to defect. After researching, should you consider one of these vehicles, don't wait too long; demand for the vehicle sometimes increases due to scarcity, and dealerships often raise prices on their remaining stock.
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Why Rotary? Here's 20 reasons!
Rotary? Isn't it that club for old guys?
Why get involved with THE Ellicott City Rotary Club?
Rotarian or not do, together we can make Ellicott City and the world better places for all. Tell us what you do that builds goodwill, friendship, understanding, and peace?


Deliver at least one Texas Watershed Stewards training program, which is a broad-based training program for all watershed stakeholders that addresses both quantity and quality issues.
Deliver two additional educational programs targeting agricultural producers that focus on soil and water resource management and distribution of educational resources, including possible field demonstrations on water conservation practices.
Develop factsheets and release a Web-based curriculum, regarding water resource protection and management.


Those working within homeland security can find employment across both government (local, state and federal) and nongovernmental organizations. Individuals working within homeland security earn on average between 40,000 and 80,000 U.S Dollars a year. Income varies depending on educational qualifications, level of training and experience as well the type of organization where one finds employment.
An Associate of Arts in homeland security receives foundational knowledge relating to this field. In particular he or she learns about the internal and external threats that society faces. At the end of this two year program students often pursue on the job training. A Bachelor of homeland security is a four year program where students acquire extensive knowledge on the subject. A Bachelor of homeland security is better positioned to enter into careers within this field. A Master's in homeland security enables students to advance their careers further and to enter the public policy making arena.
In the course of a program in homeland security, students acquire greater knowledge and skills invaluable to advancing their careers in this field. Students are able to consolidate their knowledge of issues, concepts and theories relevant to law enforcement and homeland security. In addition, students are able to expand their practical skills and are trained in managing disasters/ emergencies, the art of negotiation and apprehension protocol. Areas of study include international and domestic terrorism, information security, cyber-security, terrorism and security, weapons of mass destruction, emergency management, criminal justice, public safety and security, intelligence and homeland security, homeland security and ethics, strategic planning and homeland security policy.


AutoClear I PCSS Stainless Steel Downspout Cleanouts Leaf and Debris Diverters in custom Silver Tone color.
These fit 4″ Round Downspouts of Any Material.
Powder Coated Stainless Steel to Last Forever.
While these are what we call "White Primer Coated" they should very closely match most "White" Downspouts by various Manufacturers.
The base color in this case is "White". It is Powder Coated to the Stainless Steel.
Mill Finish Stainless Steel is not easy to paint directly. It would require a Bonding Primer and then a paint application.


My heart is heavy this fall morning in Texas. My best friend in the Chemo room, went to be with the Lord this week. She is healed, happy and whole, but we all miss her smiling face.
When I walked into that room to begin my treatment in April, God knew I needed a friend. Someone special, that I would connect with immediately. That would be Vicki Davis.
I think I drove her crazy with my positive outlook on life. Nothing she could say would dampen my spirit. She tried to be grumpy with me because she had been down a very long road with this disease. I would smile at her, and just love on her until she smiled in return.
God has us scheduled for Chemo on the same day, Tuesdays, at about the same time. Somewhere around noon. When you are sitting in a Chemo chair, you can either laugh, cry, or sleep. We chose to laugh. Anyone who has heard my laugh knows, it can rock a room. Vicki had a laugh that matched. Get both of us laughing and it was quite disturbing.
One time, we sat in a semi private part of the room, away from everybody else. You could look across the room and see everyone lined up sleeping through their treatment. Vicki and I were trying to outdo one another with stories of stupid things we had done in our lives. We busted out laughing at the same time, and startled the sleeping souls lined up across the way.
Like a row of dominoes, they jumped, looked around, relaxed and then smiled. They were relieved it was just us having more fun than we should have been. That was life with Vicki.
My last day of Chemo, as I was getting ready, I felt God tug at my heart. It was time to let go of something very precious to me. I was given a bracelet last Christmas by another lady God placed in my life. Little did this lady know, I would be wearing it during Chemo treatments, a few months later. My Chemo was coming to an end, but I thought Vicki's wasn't quite yet.
It was engraved with one of my favorite verses, and encompassed my wrist as a reminder of what I was supposed to be doing. Psalm 46:10, "Be still and know that I am God." It was a challenge for Vicki and me to sit still in those chairs. It was long hours, but being with Vicki made the time fly by. On my last day, the nurses had saved a chair for me to sit beside her.
Vicki was growing weary in her journey, and she had every right to be. I gazed at her resting in her chair, and sat down in mine. There was silence between the two of us, but the love was always there. Her IV bag was almost empty, as the nurse had just hung mine. I got out of my chair, kneeled down beside her, slid the bracelet off my wrist, and slid it onto hers.
Her eyes opened and she smiled at me groggily. She touched the bracelet, and regretted not having her reading glasses, so she could see it more clearly. I told her what is was, and what it said, and that I wanted her to have it for the remainder of her journey. That was not even three weeks ago. It was her last Chemo that day too. Thank you God for bringing her home.


Richman says it's unusual for the "X Gon' Give It To Ya" rapper not to notify his probation officer of his travel plans. He believes the judge might put stricter conditions on X and require an ankle bracelet, but Richman is reportedly ready to push against it.
U.S District Attorney spokesman Nicholas Biase said X is to appear in court later this week.
X was arrested in July and reportedly faces 14 counts of evading over $1 million in federal taxes.
After walking out of a Manhattan courthouse on Monday (July 17), DMX (real name Earl Simmons) dropped a random freestyle about his trouble with the IRS when approached by reporters. During his verse, Simmons names his lawyer Murray Richman and seems confident he'll beat the 14 counts of tax evasion he's facing.
"We don't worry with Murray, he get you outta jail in a hurry/When the government trying to bury you, you good—early," he spits.
Although Richman reportedly declined to discuss the case after the hearing, he did say Simmons plans to hire a larger legal team, including tax accountants and lawyers who specialize in tax law.
Late last week, Simmons was arrested for allegedly cheating the IRS out of $1.7 million. He was freed after posting $500,000 bail. If convicted, he could end up in prison for around 40 years.
Watch his freestyle video above.
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Blade reboot
by Marvel is approaching
James Wan, one of the most conspicuous talents in Hollywood, has recently made a Blade reboot instead of Aquaman. Wan is a man behind such masterpieces as Saw, Insidious and The Conjuring franchises.
Over the last weeks, James Wan has been promoting his latest work Aquaman, which has been positively reviewed in advance, before the premiere. Hopefully for him, this project will let him get rid of the previously obtained negative attitude after the Justice League failed to impress the audience. In one of his recent interviews, he said that he was negotiating with Marvel about relaunching Blade. That is what Wan had to say about it.
"Years back, a couple of years back, I think it was right after Conjuring 1 that I went into a particular studio and told them 'Hey, I'm a big fan of Blade. Would you guys be interested in rebooting Blade?' So that was one. Obviously, nothing came out of that! But that was my first intro into looking at potential comic book properties."
James Wan is known to really be into the horror genre, and Blade fits this idea perfectly well. Just like the first X-Men, The Blade is a type of an old-school kickstarters for comic book era of movie adaptations. As we can see, it also holds huge potential for future.
Not only Wesley Snipes, the person who gave birth to Blade long ago, but also The President of Marvel Studios Kevin Feig, both have claimed that they would like to bring the vampire hunter back to the big screen, no matter how exactly.
This year, Snipes said that he had 2 different Blade projects on the way in cooperation with Marvel. Sadly, we have no clue about whether they are moving forward or not. Nonetheless, we definitely will see the character in future.
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