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Screen Genealogies: From Optical Device to Environmental Media
In response to Het Nieuwe Instituut's Screen Spaces project one might ask why today we would emphasize the screen in talking about video culture?
Phantasmagoria, Étienne-Gaspard Robertson.
The screen might seem self-evident, but it is not. Video practitioners and critics in the 1960s and 1970s stressed very different terms, for instance. Terms such as feedback, topology, narcissism, and broadcasting, to name only a few of the most common, were more important than screen. More than the screen, it was the ability to watch a moving image recorded, played back, manipulated, or delayed in 'real time' that were sources of interest.
The concern for the screen is not an essential or self-evident quality of video, it belongs to the cultural and political questions we bring to these technologies today. Whatever we understand the screen to be, it is only one element of video, film, gaming, computer monitors, or mobile phones. Each of those technologies may be perceived through a screen, but how that screen relates to the other technological, spatial, formal, and interactive elements, and how users relate to it, can differ quite significantly.
In this sense, one reason why the screen may rise up as a term for thinking video today is that it is a feature allowing us think very different kinds of media in relationship to one another. In that sense, screens are symptomatic of what media theorists have called convergence, the conversion of previously distinct media forms—print, video, film, computer graphics—into code that can be accessed through the same graphical user interface. It also speaks to the rapid proliferation and transformation of screens in our daily life world, from movie and television screens, to phones, tablets, monitors, GPS devices, billboards, facades, kiosks, and more. For that very reason, it seems important to find ways of thinking carefully about screens, in ways that do not simply mix them all up. One of the ways in which that might be done is to examine more carefully the screen as a spatial, architectural, and environmental technology, rather than a strictly optical one, which is often how we think of it. There is no screen without a space and an architecture, and conversely, an attention to the spatial and architectural relationships required by screens, or configured by screens, can be an important means of distinguishing different operations of screening, together with the deeper histories to which they belong.
The spatial and environmental emphasis has been central to a project that we have been working on at Yale University. Entitled "Screen Genealogies," it has been running, with support from the Andrew W. Mellon foundation, over the past two years. The collaborators, Craig Buckley, (History of Art), Rüdiger Campe (in German Languages and Literatures), and Francesco Casetti (Film and Media Studies Program) have hosted a series of the seminars bringing together scholars from different disciplines and institutions to explore spatial and environment approaches to the screen.
To emphasize the screen's spatial functions, also means thinking about this entity in very different historical terms. As we have sought to argue, the common notion of the screen as a surface that displays impermanent images and which disappears under these images—an understanding epitomized by movie and television screens—is a recent one. It is only in the early nineteenth century, in connection with new period spectacles like the phantasmagoria, that the word screen was firmly linked to optical experiences. Before the 19th century, screens meant things like "fire screens," which warded off the heat and sparks from fires. It meant a "partition of wood or stone, pierced by one or more doors, dividing a room or building in two parts, or "a wall thrown out in front of a building and masking the façade." It also meant devices useful for concealing objects or people from view. In military contexts, it described a group of soldiers detached from their units in order to cover the movement of an army. It also described apparatuses used in the sifting and filtering, whether of grain, coal, or air, etc. […] For most of its history, a screen was a filter, a divide, a shelter, or a form of camouflage. Only the advent of cinema, and later of television and video, made the screen acquire its most common identity as a surface supporting a changing representation.
Diorama, Louis-Jacques-Mandé Daguerre.
The current explosion and transformation of screens paradoxically brings about the reappearance of these older meanings. New media expand the screen's function beyond the optical. Surveillance cameras provide protection and defence, their very display functions defining differences between inside and outside, security and insecurity. In retrieving information, computers filter vast reservoirs of data by combining a user's query with a search engine's secretive algorithms. Hand-held devices enable users to create existential bubbles in which they can find intimacy and refuge in any range of situations. Global Positioning Systems parse territory and identify the right routes. Digital interfaces underline the separation between two worlds, and maintain control over the passages between them. Illuminated digital façades promise to make buildings more conspicuous and responsive, while also hiding and dissimulating their underlying structures from view. Screens have again become filters, shelters, divides, and means of camouflage. They remain surfaces that display images and data, yet their opticality has been deeply affected by their reference to, and connection with, and intervention into the various spaces they inhabit.
In the project and its forthcoming book, Screen Genealogies: From Optical Device to Environmental Media (Amsterdam University Press, 2019) a range of objects that we might not typically think of in terms of screens are theorized in these terms. These include such things as Casetti returning to Siegfried Kracauer's description of the film screen as the polished shield of Athena. To conceive of the screen as a shield is to understand it as something that reflects the world but which also protects the hero from being destroyed by the deadly gaze of medusa, opening up the screen's latent status as a site of spatial conflicts within film theory. It includes Yuriko Furuhata's analysis of smoke screens in terms of past and contemporary geo-engineering, in particular the link between Nakaya Fujiko's fog environments at the Pepsi pavilion at Osaka 70, and its relationship to efforts by her father to create artificial weather during wartime military research. It includes Buckley's effort to outline a genealogy of the media façade, one which does not begin with new lighting technologies and advertisement in the 1970s, but with a much older dialectical conflict over the façade's role as a facial media between inside and outside. It also includes recent work by Noam Elcott on the media archaeology of screen formats and by Nanna Verhoeff on the "spectatorial territories" created by recent interactive screen projects for public spaces.
To borrow a notion from the philosopher and historian of science Ian Hacking: screens today are not only devices of representation but also devices for intervening in the world. Such an approach aims to rediscover the history of screens in places where we don't expect to find them; it also seeks to comprehend the ways in which an optical understanding of the screen came to dominate other historical possibilities.
To approach screens genealogically means that something other than looking back to the find "origins," "precursors," or "anticipations," which has been the focus of much media archaeology to date. Rather it would insist that screens are never pre-existing objects nor are they the inventions of particular individuals or groups. What is essential is to highlight the processes and struggles where a set of technologies, spaces, spectators, and operations come together in order to become a screen. A surface becomes a screen through an arrangement of apparatuses and by virtue of the struggle between forces and practices. What is always at stake is an ensemble of elements—an assemblage—characterized by certain dispositions and sustaining certain types of operations. A genealogy of the screen would thus emphasize processes of transformation and emergence, rather than moments of invention or historical culmination.
Craig Buckley, Rüdiger Campe, and Francesco Casetti.
The book Screen Genealogies: From Optical Device to Environmental Media is published with Amsterdam University Press in 2019 and available online.
Screen and Screenscapes with Francesco Casetti
Professor Francesco Casetti reads the screen in its diverse instantiations. "Screens not only display images; they define the modes of access to these images. In this, they are inherently political."
Research and curation Marina Otero Verzier, Director of Research Katía Truijen, Senior Researcher
New York Vere van Gool, curator Emma Macdonald, curatorial assistant
Rotterdam Shay Kreuger, moderator Koos Breen, set design and graphic design Jeannette Slütter, set design Tanja Busking, film production
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A share is a type of ownership of a company. When you buy shares you become a part-owner of a company and share in the future profitability of that company.
You can buy shares directly via the stock market or indirectly, through a managed fund which invests in shares on your behalf.
Direct Shares can provide regular income through the payment of dividends and may also provide capital growth through increases in share prices.
Indirect ownership - where you buy units in a managed investment which invests in shares.


When it's time to chow down on pizza, this is the ideal emoticon for you to post on Facebook. This smiley has a personality all its own and your friends will share in its delight. We have a vast array of fun images like this one that you can share or use in a message.


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Khusro Mumtaz
Rangreza ** ½
Dir: Amir Mohiuddin
Starring: Bilal Ashraf, Urwa Hocane, Gohar Rasheed, Ghana Ali, Saleem Mairaj, Seemi Pasha, Imran Peerzada, Tanveer Jamal
A rock star, Ali (Bilal Ashraf) falls in love with the daughter, Reshmi (Urwa Hocane) of a qawwal (Tanveer Jamal) who is betrothed to her cousin, Waseem (Gohar Rasheed). The expected complications ensue until the movie reaches its expected denouement. The prosaicness of the plot - exacerbated by sloppy editing and scripting and uneven acting (ranging from the good to the bad to the ugly) – is a pity because writer Akhtar Qayyum and director Amir Mohiuddin had a potentially absorbing drama on their hands and that potential ends up being wasted. Achi Manzil, where the qawwal family reside, has a fascinating backdrop with the two sons of the dying patriarch (Akbar Subhani, whose emaciated appearance, contrasted cleverly with fading pictures from his television hero days, strikes an appropriately melancholic – and meta - chord) and their respective families. Qayyum and Mohiuddin could have used this milieu to explore not only familial grievances and ancestral secrets but also to comment on the state of qawwali and our musical (and otherwise) heritage in present day Pakistan. Instead, by concentrating on a truly unremarkable and unoriginal romance across class divides, the movie wastes a golden opportunity to give us something truly original and
A rock star, Ali (Bilal Ashraf) falls in love with the daughter, Reshmi (Urwa Hocane) of a qawwal (Tanveer Jamal) who is betrothed to her cousin, Waseem (Gohar Rasheed). The expected complications ensue until the movie reaches its expected denouement. The prosaicness of the plot - exacerbated by sloppy editing and scripting and uneven acting (ranging from the good to the bad to the ugly) – is a pity because writer Akhtar Qayyum and director Amir Mohiuddin had a potentially absorbing drama on their hands and that potential ends up being wasted. Achi Manzil, where the qawwal family reside, has a fascinating backdrop with the two sons of the dying patriarch (Akbar Subhani, whose emaciated appearance, contrasted cleverly with fading pictures from his television hero days, strikes an appropriately melancholic – and meta - chord) and their respective families. Qayyum and Mohiuddin could have used this milieu to explore not only familial grievances and ancestral secrets but also to comment on the state of qawwali and our musical (and otherwise) heritage in present day Pakistan. Instead, by concentrating on a truly unremarkable and unoriginal romance across class divides, the movie wastes a golden opportunity to give us something truly original and absorbing.
absorbing.
However, on the plus side, the music is very good though we really should have been given at least one good qawwali. Also, in the boorish, uneducated but talented tabla nawaz Waseem, Gohar Rasheed, manages to create a memorable, larger-than-life character without (just) tipping over into caricature. The movie (rightly or wrongly) belongs to him. Tanveer Jamal is also very good. Bilal Ashraf, on the other hand is truly weak and Imran Peerzada, playing his politician father, is possibly even worse.
Cut to chase: A potentially absorbing movie is buried beneath an underwhelming romantic plotline.
Chupan Chupai **
Dir: Mohsin Ali
Starring: Ahsan Khan, Neelam Muneer, Faizan Khawaja, Ali Rizvi, Adnan Jaffar, Rehan Sheikh, Talat Hussain, Vajdaan Shah
A quartet of inexperienced kidnappers led by Babu (Ahsan Khan, quite good) get in over their heads when they get embroiled in a scheme to abduct a politician's son. This comedy crime caper certainly has its moments which elicit a few chuckles, including a neat twist concerning Babu's girlfriend, Pari (Neelam Muneer). But, really, director/co-writer Mohsin Ali can't claim much credit because Chupan Chupai is a remake of Tamil hit Soodhu Kavvum and most of the movie's "novelty" comes to us thanks to the original. That said, while the movie's first half keeps chugging along well enough, Ali loses control in the second. The film turns repetitive – despite the addition of a new character, Khan/Doctor (Rehan Sheikh) a fixer with filmmaking aspirations – the editing becomes a bit messy and the plot unnecessarily complicated without ever becoming as clever as it thinks it is. The Khan/Doctor character adds nothing to the plot, for instance.
The mostly young cast do what they can with what they are given but I wish Mohsin Ali had tightened up the script, remained less faithful to the original and riffed mightily off the nifty idea of the truth behind Pari (he had a potential goldmine there). If Chupan Chupai had been a completely original film I might - repeat, might – have awarded it an extra half star but being a remake I really can't bring myself to be that generous.
Cut to chase: This remake of a Tamil hit flounders in its second half.
Kmumtaz1@hotmail.com; Twitter: @KhusroMumtaz

redpajama_set_name: RedPajamaCommonCrawl


Thanks to you and Uwe for the answers!
> Ivan can you describe more about your application?
> won't result in a very large speedup for your application.
>> No, we do not use StandardAnalyser or StandardTokenizer.
>> covering the phrase token.
>> that now is generated by JFlex instead of JavaCC.
>> THEY ARE ADDED 5 TIMES FASTER!!!
>> the reason the total process of indexing to be not so reasonably faster.
>>> This week I switched the lucene library version on one customer system.
>>> including optimisation. Great Job!
>>> Absender zuruckschicken. Bitte loschen Sie danach diese Email.
>>> return email and delete the document.
>>> This message was checked by NOD32 antivirus system.
>> Absender zurückschicken. Bitte löschen Sie danach diese Email.
>> return email and delete the document.


text:
Q: When asked to find all solutions to a differential equation, what's left implicit? Suppose the following problem were to appear in an introductory textbook on differential equations.

Find all solutions to $y'=y$.

What's left implicit in such a statement? My interpretation of the problem would be:
Find the set of all differentiable functions $y : X \rightarrow \mathbb{R}$ such that

*

*For all $x \in X$ it holds that $y'(x) = y(x)$

*$X$ is connected.

*No function with properties (1) and (2) is a proper extension of $y$.

Edit: An alternative opinion was advanced in the comments, that the problem is to find the set of all differentiable functions $y : \mathbb{R} \rightarrow \mathbb{R}$ such that for all $x \in \mathbb{R}$ it holds that $y'(x) = y(x).$

A: Often, finding the largest X is part of the problem. For example with linear DE where coefficients are functions with discontinuities.
Sometimes you have two sets of solutions on intervals $]-\infty, a[$ and $]a,+\infty[$, and you have to find solutions on $\mathbb{R}$ : there may be infinitely many, only some, or none, depending on cases.
You may also have to look for singular solutions.

redpajama_set_name: RedPajamaStackExchange


In the course of our lives, we have different kinds of experiences. Some give us happiness while some others leave us with bitter memories. All through such moments of our life, we convey our emotions and experiences through our faces and this takes a toll on our skin after a certain age. But with Juvéderm, we can fight back.
Juvéderm is an injectable gel made of hyaluronic acid, a substance found in human skin. Our skin is made of a collagen, elastic fibers and hyaluronic acid. When any of these components of the skin breaks down we get lines and wrinkles. If we can replace the hyaluronic acid then we can fill the lines and wrinkles in your skin.
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Bauma, the largest trade show in the world hides some MBI branded record. An unbelievable number of visitors at the MBI stand; people coming from all over the world to discover the latest news in the brilliant MBI product range.
The demolition multisystem MS140, the biggest demolition attachment displayed at bauma thanks to its 4.5 meters height, has been protagonist of a huge number of selfies and dominated the social profiles of our fans.
The new combi crusher line CC has been the most exciting news for demolition professionals, thrilled to see these great tool at work on their machines soon.
In addition, the new EAGLESHEARS brand caught the eyes of our visitors, ready to surf the new dedicated website; www.eagleshears.com to discover every detail of the tool together with its endless applications.


text:
Contributing to the COVID-19 Fight One Stitch at a Time, Esther Muna's Story
"I got involved with Amref Health Africa back in 2015 at the Dagoretti Centre through the Divas – a program for young mothers. Each of the Divas had a young child, and we had dropped out of school. We were taught business skills, and through a partnership with Kenya Kanga, we were equipped with tailoring skills.
When COVID-19 hit Kenya, we had no source of income. Things were not easy for us.
In May, Amref called on us, and we were taken through a week's training on what personal protective equipment (PPEs) are and the process of producing surgical face masks and gowns. We are excited because through our work to produce these PPEs, we are among those contributing to the fight against COVID-19.
I am also glad to have a sustained income to take care of my son, who is now in Grade 2.
Amref has continued to train us, and we have more skills in business, fashion and design. With these skills, we can even eventually set up our boutiques," concludes Esther.
'Divas' is a Young Mothers' group facilitated through a partnership between Amref Health Africa and Kenya Khanga, where a group of 12 young mothers between the ages of 17 and 32 years were equipped with tailoring skills. The young mothers participated in a two-day weekly tailoring workshop where they worked on selected items and were paid according to work done, earning Kshs 1,500-2,500 (US$15-25) per month, which many of them used to pay rent, buy food and some pay for their children's daycare. They gained life skills, and 50% of them adopted family planning to delay more pregnancies, they gained alternative skills in fashion, design and urban gardening.
Since the year 2000, Amref Health Africa has been running youth empowerment programmes in Dagoretti which have impacted over 100,000 lives through various interventions focusing on vulnerable children and young people. One such venture is the Dagoretti Fashion and Design Centre – a social impact-driven enterprise in fashion and design to produce high-quality market-driven products to socially and economically transform the lives of young women.
Due to the COVID-19 pandemic, there has been a great demand for Personal Protective Equipment (PPE) globally, and shortages are leaving health workers and the general public dangerously exposed to the disease. Like many other countries, the Kenyan government issued a directive for the public to wear face masks at all times when in public places to avert the spread and transmission of COVID-19. In light of this, there has been an exponential demand for PPE, and countries have had to innovate and activate industry capacity to support the local manufacturing amid the global and national lockdowns.
With support from The Coca-Cola Foundation, Amref Health Africa has commenced the process of training 50 young mothers and women from Dagoretti (who had previously received training in fashion and design) on mass production of PPE at the Dagoretti Fashion and Design Centre.
Amref currently has a production capacity of 20,000 masks and 100 surgical gowns per month, with the capacity to increase production based on demand. This presents an opportunity for young women to earn a living by using their skills to address the dire need for PPE. Amref will also continue to equip these young women with important information and knowledge on sexual reproductive health and rights.
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[Free eBooks] Freebies, Lunchtime, More Freebies, Let's Eat!
I am hungry for lunch and a few freebies. Let's EAT!!
A girl abandoned to the streets.A brother brutally murdered.Can she find the killer before he finds her?
YOUNG JAMES FROST just knows, deep in his bones, that he's a writer. He writes far into early mornings, after his wearying hours of scrubbing toilets and sweeping floors. He loves writing that much.
But it's not only the joy of words that keeps him grinding; it's his desire to retire the janitor's mop. He sees being published as the key to living an improved life. James has another deep-seated conviction: that he's not good enough. He secretly longs to be accepted. However, the conventional others in his life seem all too willing to remind him that he's wasting his time.
Then he meets and falls in love with Leigh, the one bright spot in his endless misery of self-doubt. A quiet but resolutely religious girl, she has to fight off disapproval of her own from overly critical parents, whose insults are countered by James's often-voiced admiration of her. Likewise, Leigh's faith in his talents begins to build his confidence, eventually allowing her to introduce him to a different way to help himself: relying on God.
Ultimately, James's newfound faith is sorely tested to the point of doubt when his dream to be published seems to melt into a mirage, smothered by countless rejection slips from agents and publishers. His faith is also battered by having to fight highly emotional battles and suffer fear and loss. Just when James appears hopelessly sapped by devastating events, one last door opens, and he's rocked by an epiphany.
Revenge. Romance. Murder. A gorgeous Blonde, a kidnapped Sister, a handsome Stranger, and a vengeful Killer. The desperate race is on to find the sister before both women wind up dead. A Romantic Suspense novel that will curl your toes.
He has the power to Walk between worlds. His captors think they can hold him. They're wrong.
What do you say when offered a lifelong position in the most powerful and corrupt secret organization in mankind's history? If you want your life to last longer than the job interview, you say yes. In exchange for wealth, power, and training for his newly discovered abilities, all Daniel has to do is help them reach a new, unsuspecting world. To make things worse, a group of desperate escapees pin their thin hopes on him, willing to risk everything on a miracle that Daniel has no idea how to provide.
He's a wealthy Italian celebrity used to dating beautiful women. She's a guesthouse cook without a penny to her name. When tragedy brings them together for a few weeks, an unlikely romance blossoms. But he has commitments that take him back to his jet-setting life, while her future lies in a remote English village. Is it possible for two people from such different walks of life to find a way to be together?
"Lost in Infinity" is a fictional psychological thriller wrapped in the factual memoir of a chronic insomniac suffering from apeirophobia (the fear of infinity). The novel unfolds the history of his life as he tries to unlock repressed memories through a near schizophrenic relationship with his own splintered subconscious.
The novel offers a no holds barred look into the private confessions of a man on the precipice of a mental breakdown. The recurring theme of déjà vu leads the reader through the work giving glimpses of a dark past while offering anecdotes that will relate to every reader. Mixing in humor and satire with the tragedy of a childhood spent under the microscope of therapists keeps the mood light while he digs deeper into his past looking for the root of his problems.
The author pulls back the curtain and reveals the dark inner workings of his mind as he fears a slow deliberate path toward schizophrenia. Suicide, insomnia, stress, parenthood, relationships, careers and redemption will keep you reading and waiting for the ultimate reveal.
You'll feel as if you've read this book before. Maybe you have? All is not as it seems. The feeling of déjà vu may be much more than a feeling. What if his fears are correct? What if the world is stuck in an infinite cycle set to skip and repeat for eternity? What if we are all simply lost in infinity?
What would you do if your son went missing? How far would you go to get him back?
Michael Barton comes home to find his infant son gone. There is no trace of the kidnappers, and the police have no leads. Michael is driven by a promise and he won't stop.
Detective Jason Strong led the task force to find Michael's son and has become his friend. But as Michael's darkness grows, Jason fears he may be the only one who can stop him.
It is a race against time, as the detective tries to stop the events that will change everyone, before it's too late.
Most of Zoë's friends are dead, but she doesn't mind because they died long before she met them. Then one Tuesday night an angel takes her salsa dancing and turns her world upside down.
Grim reality closes in when she discovers a body in her company's boiler room and Higher Angels accuse her best ghost friend of murder. Knowing she's the only one who can stand against them, Zoë resorts to lying, stealing and summoning, all while trying to determine if what she's feeling is real. In the end, getting blood on her hands forces Zoë to question herself, and her angel to question her.
Even with her mother's death days away, even as her marriage is falling apart, Euly Winger leaves home, returning to the place she grew up in order to uncover a dark secret about her parent's divorce.
On her search for answers, Euly meets up with an old family friends, some offer help, some disaster. Euly returns home just hours before her mother dies. As they sit alone together, Euly asks her mother for the truth.
Deep under Arctic waters lies an ice kingdom carved into a glacier. Those who dwell within it possess magnificent biological secrets. Due to the dangers of impending war, the Princess of Adlivun is forced to flee her undersea utopia and regroup with her sisters in Alaska.Captain Trevain Murphy is a successful king crab fisherman who has spent his life building his empire above the sea, and knows nothing of the empire beneath it. When he meets a mysterious dancer whose father has recently died, he extends kindness towards her, unaware of her unique genetics and royal lineage. Trevain's attraction to the enigmatic Aazuria Vellamo will involve him in dangerous designs that will forever change his life, and his perspective on himself and his world. He embarks on perilous journeys in which he will need to release all of his insecurities and inhibitions in order to survive.
This is a short story prequel to the Heven and Hell series. It is approx. twenty pages plus bonus material of the first chapter of Masquerade!
What if your life was charmed and everything in it was perfect…Before.
This is the story of my past. Of what things were like for me when everything was normal. Of what every teenagers life is like. Clothes. Parties. Boys and summer vacation. What's so wrong with that? I liked it. I was happy. At least I thought I was. I didn't know that lies and secrets were about to take over my existence. I didn't know that there was someone out there, someone meant for me. I didn't know that I was about to go on a journey, a journey that would lead me to the girl I am today. This is the story that leads up to one of the worst years of my life. Would I go back and change things? Erase everything that has happened to go back into these moments? Not a chance. This is a story of before.


About this journey The average travel time between Toulouse and Sete is 2h48 minutes. The quickest route is 1h48 minutes. The first train leaving Toulouse is at 05:50, the last at 19:49. There is an average of 40 trains a day between Toulouse and Sete, leaving approximately every 23 minutes.
"We enjoyed travelling on the train and would recommended it as a great way to travel on holiday, it was very relaxing, the trains were comfortable and always on time. "


Companies around the world implement ERP systems for a wide variety of reasons: some to eliminate manual efforts, some to run their businesses in an integrated environment, some for regulatory reasons, and on and on. I think that if you ask many people why they undertake an ERP implementation, you might get an answer like "So we can improve".
True as it is that an integrated ERP system can help a company gain efficiencies, it seems like that improvement sought is not always clearly understood or realized.
There is an old saying that goes something like this: "When you automate a bad process, you can now produce junk at the speed of light". I do not remember who originally said it and the first time I heard it, it was stated more colorfully. The point needs to be well taken though.
Does implementing an ERP system necessarily mean that the business is going to run better and more efficiently? Will it be more profitable? Will the customers be happier? Will cycle times be quicker? Will the company be more agile?
Maybe yes, maybe no. Without fundamental process improvement, no real change occurs.
Hand-in-hand with a new system implementation should be an aggressive effort to analyze and improve the business processes the new system is to support. Too many times, we see companies who deploy a new system with the expectation that it will simply be used to duplicate the current or legacy processes.
It is only through the relentless process of continually analyzing the current business processes, identifying the waste and inefficiency in those processes and then working diligently to re-structure them that real improvement occurs.
There is much to say about how process improvement can be achieved and sustained. Believe me – much more and you will see more on this topic in our blog. For now, though, keep in mind that the system by itself, does not necessarily make for improved practices.
I leave you with a quote by W. Edwards Deming who said, "Improvement is not mandatory but neither is survival".


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Research ArticleCardiology Free access | 10.1172/JCI33372
Combined deficiency of ABCA1 and ABCG1 promotes foam cell accumulation and accelerates atherosclerosis in mice
Laurent Yvan-Charvet, Mollie Ranalletta, Nan Wang, Seongah Han, Naoki Terasaka, Rong Li, Carrie Welch, and Alan R. Tall
Division of Molecular Medicine, Department of Medicine, Columbia University College of Physicians and Surgeons, New York, New York, USA.
Address correspondence to: Laurent Yvan-Charvet, Division of Molecular Medicine, Department of Medicine, Columbia University, 630 West 168th Street, New York, New York 10032, USA. Phone: (212) 305-5789; Fax: (212) 305-5052; E-mail: ly2159@columbia.edu.
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First published November 8, 2007 - More info
Published in Volume 117, Issue 12 on December 3, 2007
J Clin Invest. 2007;117(12):3900–3908. https://doi.org/10.1172/JCI33372.
© 2007 The American Society for Clinical Investigation
First published November 8, 2007 - Version history
Received: July 23, 2007; Accepted: September 19, 2007
HDLs protect against the development of atherosclerosis, but the underlying mechanisms are poorly understood. HDL and its apolipoproteins can promote cholesterol efflux from macrophage foam cells via the ATP-binding cassette transporters ABCA1 and ABCG1. Experiments addressing the individual roles of ABCA1 and ABCG1 in the development of atherosclerosis have produced mixed results, perhaps because of compensatory upregulation in the individual KO models. To clarify the role of transporter-mediated sterol efflux in this disease process, we transplanted BM from Abca1–/–Abcg1–/– mice into LDL receptor–deficient mice and administered a high-cholesterol diet. Compared with control and single-KO BM recipients, Abca1–/–Abcg1–/– BM recipients showed accelerated atherosclerosis and extensive infiltration of the myocardium and spleen with macrophage foam cells. In experiments with isolated macrophages, combined ABCA1 and ABCG1 deficiency resulted in impaired cholesterol efflux to HDL or apoA-1, profoundly decreased apoE secretion, and increased secretion of inflammatory cytokines and chemokines. In addition, these cells showed increased apoptosis when challenged with free cholesterol or oxidized LDL loading. These results suggest that the combined effects of ABCA1 and ABCG1 in mediating macrophage sterol efflux are central to the antiatherogenic properties of HDL.
Plasma HDL levels have a strong inverse relationship to the incidence of heart attack and stroke (1), but the underlying mechanisms are poorly understood. The ability of HDL to promote efflux of cholesterol from peripheral tissues and transport to the liver was proposed long ago as an antiatherogenic mechanism (2). However, HDL also has antiinflammatory, antioxidant, antithrombotic, and vasodilating properties that could all potentially be involved in its antiatherogenic effects (3, 4). In vivo proof of the relevance of these various potential antiatherogenic mechanisms is still lacking.
HDL and its major apolipoprotein, apoA-1, have the ability to promote the efflux of cholesterol from cholesterol-loaded macrophages and smooth muscle cells, the major cellular components of atherosclerotic plaques (5). Several different pathways of cellular cholesterol efflux have been described, such as aqueous diffusion (6), efflux of cholesterol associated with apoE secretion (7, 8), scavenger receptor–BI–mediated (SR-BI–mediated) cholesterol efflux (9), and active cholesterol efflux via the ATP-binding cassette transporters ABCA1 and ABCG1 (10, 11). ABCA1 promotes cholesterol and phospholipid efflux from cells to lipid-poor apoA-1 but not to mature HDL particles, while ABCG1 promotes cholesterol efflux to HDL and other lipoprotein particles but not to lipid-poor apoA-1 (12–14), suggesting that the 2 transporters could have complementary activities in vivo. ABCA1 is the mutant molecule in Tangier disease, characterized by low HDL levels, by macrophage foam cell accumulation in the tonsils and spleen, and probably by modestly increased atherosclerosis (15–18). However, the increased risk of atherosclerosis in patients with Tangier disease does not appear as dramatic as one would expect in individuals with an almost complete absence of HDL (18), and targeted inactivation of ABCA1 did not affect the development of atherosclerosis in apoE–/– and Ldlr–/– mice, 2 mouse models of atherosclerosis (19). Transplantation of BM from Abca1–/– mice into Ldlr–/– or apoE–/– recipients causes a moderate increase in atherosclerosis (19, 20), but there are conflicting reports on the effects of ABCA1 overexpression in transgenic mouse models of atherosclerosis (21, 22). Abcg1–/– mice accumulate macrophage foam cells in various tissues such as the lung (23), but the effects of ABCG1 deficiency on atherosclerosis are controversial and poorly understood. BM transplantation from Abcg1–/– mice into apoE–/– or Ldlr–/– recipients resulted in either a modest increase (24) or decrease in atherosclerosis (25, 26). The decreased atherosclerosis was attributed to increased macrophage apoptosis in response to oxidized LDL (26) or to compensatory upregulation of ABCA1 and increased apoE secretion in Abcg1–/–macrophages (25).
In an attempt to resolve these controversies (24–26) and to test the hypothesis that effects of single transporter deficiency are masked by compensatory upregulation of the other transporter, we carried out BM transplantation from mice with single or combined deficiencies of ABCA1 or ABCG1 into Ldlr–/– mice. Consistent with our hypothesis, the Abca1–/–Abcg1–/– BM recipients showed increased atherosclerosis and massive myocardial foam cell accumulation. Mechanistic studies showed a major defect in cholesterol efflux in Abca1–/–Abcg1–/– macrophages, decreased apoE secretion, increased apoptosis in response to loading with free cholesterol or oxidized LDL, and increased expression of inflammatory and chemokine genes. Thus, many of the proposed atheroprotective properties of HDL appear to be secondary to macrophage sterol efflux mediated by these 2 transporters.
Massive myocardial foam cell accumulation in Abca1–/–Abcg1–/– BM recipients. We bred mice with single or combined KO of Abca1 and Abcg1, transplanted BM from these mice into irradiated Ldlr+/– mice, and, after a recovery period, placed the recipients on a high-cholesterol diet for 12 weeks. Upon sacrificing the mice, we noticed a striking phenotype that was seen in all recipients of Abca1–/–Abcg1–/– BM (n = 9) but not in mice receiving WT, Abca1–/–, or Abcg1–/– BM. In the Abca1–/–Abcg1–/– BM recipients, the hearts were smaller and were pale compared with mice of the other genotypes (Figure 1A). Pallor was also observed in the small intestine, and the spleen was enlarged, in Abca1–/–Abcg1–/– recipients. Microscopic examination of the hearts of Abca1–/–Abcg1–/– mice revealed massive infiltration with cells containing foamy cytoplasm that stained positively with Mac-3, indicating that they were macrophage foam cells (Figure 1, B and C). In addition, these infiltrates contained empty spaces of similar appearance to the cholesterol clefts commonly seen in atherosclerotic lesions (Figure 1C). There were also a moderate number of neutrophils in the infiltrate as shown by MCA771G staining (Figure 1D). TUNEL staining revealed clusters of apoptotic cells in the midst of the inflammatory infiltrate and this was observed only in Abca1–/–Abcg1–/– BM recipients (Figure 1E). Previous studies have shown accumulation of foam cells and other inflammatory cells in the lungs of Abcg1–/– mice (23). We also found foci of foam cells and other inflammatory cells including neutrophils in the lungs of Abcg1–/– BM recipients; interestingly, this phenotype was not increased in Abca1–/–Abcg1–/– recipients beyond what was seen in Abcg1–/– recipients (Figure 1F). In Abca1–/–Abcg1–/– recipients, the spleen also showed an increase in foam cells compared with control and single-KO recipients (Figure 1G).
Foam cell infiltration of the myocardium in Abca1–/–Abcg1–/– BM recipients but not WT or single-KO recipients. (A) Representative hearts obtained from Ldlr+/– mice transplanted with BM derived from WT, Abcg1–/–, Abca1–/–, or Abca1–/–Abcg1–/– mice and fed a high-cholesterol diet for 12 weeks. Hearts from all Abca1–/–Abcg1–/– recipients were visibly smaller and pale compared with WT and single-KO controls. Scale bar: 1 cm. (B–G) Paraffin-embedded serial sections obtained from the myocardium surrounding the proximal aorta, lung, and spleen of mice transplanted with BM of all 4 genotypes. Original magnification, ×200. (B) H&E staining revealed an extensive cellular infiltrate in the myocardium of Abca1–/–Abcg1–/– recipients (9 of 9) but not controls (WT, 0 of 12; Abcg1–/–, 0 of 8; Abca1–/–, 0 of 16). The infiltrate resembled foam cell lesions of the vessel wall, including apparent cholesterol clefts (arrows). (C) Mac-3 immunostaining (dark brown) confirmed that the majority of cells were macrophages. (D) MCA771G immunostaining (dark brown) revealed a moderate number of neutrophils (arrows) as well. (E) TUNEL staining revealed pockets of apoptotic cells (red) in the infiltrated regions of myocardium from Abca1–/–Abcg1–/– recipients (7 of 9) only. (F and G) H&E staining revealed an extensive cellular infiltrate in the lungs and spleens of Abca1–/–Abcg1–/–recipients as well as in the lungs of Abcg1–/– recipients. Arrows indicate foam cells.
Accelerated atherosclerosis in Abca1–/–Abcg1–/– BM recipients. After 12 weeks on the high-cholesterol diet, control mice receiving WT BM had developed only sparse, foam cell–containing atherosclerotic lesions in their proximal aortas (Figure 2B). Atherosclerotic lesion development was similar to control mice in Abcg1–/– BM recipients and was moderately increased in recipients of Abca1–/– BM (Figure 2A). In contrast, atherosclerosis was more dramatically increased in mice receiving Abca1–/–Abcg1–/– BM (Figure 2). Further characterization of these atherosclerotic lesions revealed complex plaques containing macrophages, necrotic cores, fibrous caps (Figure 2), and small numbers of neutrophils and TUNEL-positive apoptotic cells (6–8 cells per high-power field; data not shown). TUNEL-positive cells were rarely observed in lesions from mice of the other genotypes.
Atherosclerotic lesion development in the proximal aorta. (A) Quantification of proximal aortic root lesion area by morphometric analysis of H&E-stained sections in Ldlr+/– mice transplanted with WT (n = 12), Abcg1–/– (n = 8), Abca1–/– (n = 16), or Abca1–/–Abcg1–/– (n = 9) BM after 12 weeks of high-cholesterol diet. Values for individual mice are shown as open circles, representing the average of 5 sections per mouse. Horizontal bars indicate the group medians: control, 3,994 μm2/section; Abcg1–/–, 3,724 μm2/section; Abca1–/–, 40,070 μm2/section; Abca1–/–Abcg1–/–, 186,182 μm2/section. Mean lesion areas (mean ± SEM) were as follows: control, 9,090 ± 3,872 μm2/section; Abcg1–/–, 15,412 ± 1,640 μm2/section; Abca1–/–, 72,675 ± 28,879 μm2/section; Abca1–/–Abcg1–/–, 174,958 ± 36,269 μm2/section. ANOVA was performed with square root–transformed data. (B) Representative H&E-stained proximal aortas from mice receiving BM of all the 4 genotypes after 12 weeks of high-cholesterol diet. Original magnification, ×100.
Plasma lipoprotein profiles. We next carried out studies to elucidate potential mechanisms responsible for increased macrophage foam cell accumulation in arteries, myocardium, and other tissues in mice receiving Abca1–/–Abcg1–/– BM. The atherogenic diet resulted in moderate hyperlipidemia in all groups of mice (Supplemental Table 1; supplemental material available online with this article; doi:10.1172/JCI33372DS1). Plasma lipoprotein analysis by fast protein liquid chromatography revealed slightly increased VLDL and LDL cholesterol levels (less than 30%) in mice receiving Abca1–/–Abcg1–/– BM (Supplemental Figure 1A) and no major difference in the lipoprotein profile of animals receiving Abca1–/– or Abcg1–/– compared with WT BM (data not shown). Decreased macrophage apoE secretion (see below) could explain the slightly higher VLDL and LDL levels (27, 28) and lower levels of apoE in these fractions in Abca1–/–Abcg1–/– BM recipients (Supplemental Figure 1B). There were no changes in HDL-cholesterol levels, most likely because after 12 weeks on the atherogenic diet, mice showed a redistribution of apoE among plasma lipoproteins and less than 10% associated with the HDL fraction (29). Although the increase in VLDL and LDL could have contributed to increased atherosclerosis, it is unlikely to have been the major causative factor.
Defective cholesterol efflux and foam cell formation in Abca1–/–Abcg1–/– macrophages. To test the hypothesis of a defect in cholesterol efflux as the underlying cause of increased atherosclerosis, we carried out cholesterol efflux studies and measured the cholesterol content of freshly isolated peritoneal macrophages. Abcg1–/– cells displayed a decrease in cholesterol efflux to serum, polyethyleneglycol-HDL (PEG-HDL), and human HDL isolated by ultracentrifugation (20%, 30%, and 30% decreases, respectively), while in Abca1–/–Abcg1–/– macrophages there was a more profound decrease (more than 2-fold) in the ability of serum and HDL to promote cholesterol efflux compared with single-KO and control macrophages (Figure 3). Cholesterol efflux to HDL was slightly increased in Abca1–/– cells (Figure 3C), likely reflecting a compensatory induction of ABCG1 (see below). Cholesterol efflux studies were also performed using media devoid of acceptors or containing lipid-poor apoA-1 or apoE (Figure 3D). This showed a profound defect in cholesterol efflux under all 3 conditions in Abca1–/–Abcg1–/– macrophages, even below the level observed for Abca1–/– cells. Defective cholesterol efflux led to a marked increase in cholesterol content primarily in the form of cholesteryl esters in peritoneal macrophages isolated from Abca1–/–Abcg1–/– mice fed standard chow (Figure 4A) or high-cholesterol diet (Figure 4B). In contrast, single-KO mice did not show increased cholesterol content when fed standard chow and only modestly increased cholesterol content compared with controls when fed high-cholesterol diet (Figure 4, A and B). These studies suggest synergistic roles of ABCA1 and ABCG1 in promoting macrophage cholesterol efflux, consistent with recent in vivo studies of macrophage reverse cholesterol transport showing additive effects of deficiencies of ABCG1 and ABCA1 in macrophages (30). Together these data indicate that a major defect in cholesterol efflux in Abca1–/–Abcg1–/– macrophages is the underlying mechanism responsible for increased atherosclerosis.
Cholesterol efflux from peritoneal macrophages isolated from WT, Abca1–/–, Abcg1–/–, and Abca1–/–Abcg1–/– mice. (A, B, and D) Thioglycollate-elicited macrophages were cultured for 24 h in DMEM plus 0.2% BSA containing 50 μg/ml acLDL, 3 μmol/l T0901317, and 2 μCi/ml [3H]-cholesterol. Then, a pool of human serum (2.5%; A), a pool of human serum devoid of apoB-containing particles (2.5%; B), and media devoid of acceptors or containing lipid-poor apoA-1 or apoE (D) were added as acceptor and incubated for 6 h before the media and cells were collected for analysis. (C) Cholesterol efflux was also confirmed by cholesterol mass analysis. Cells were cultured overnight with 50 μg/ml acLDL and 3 μmol/l T0901317 and then incubated for 6 hours with 50 μg/ml human HDL isolated by ultracentrifugation. The HDL-mediated net cholesterol efflux (ΔTC) was calculated as the difference of cholesterol mass of the medium with and without cells. Values are mean ± SEM. *P < 0.05 versus WT. ΧP < 0.05 versus single-KOs.
Cellular cholesterol mass in peritoneal macrophages from WT, Abcg1–/–, Abca1–/–, and Abca1–/–Abcg1–/– macrophages. Thioglycollate-elicited macrophages were harvested from WT, Abcg1–/–, Abca1–/–, and Abca1–/–Abcg1–/– mice fed standard chow (A) or high-cholesterol diet for 2 weeks (B). After a 1-hour incubation at 37°C, nonadherent cells were removed and adherent cells consisting of macrophages were directly used to estimate cellular cholesterol and cholesteryl ester mass content by gas-liquid chromatography. TC, total cholesterol; FC, free cholesterol; CE, cholesteryl esters. Values are mean ± SEM. *P < 0.05 versus WT. ΧP < 0.05 versus single-KOs.
Decreased apoE secretion in Abca1–/–Abcg1–/– macrophages. Next we assessed apoE secretion in transporter-deficient macrophages. Previous studies have suggested that ABCA1 deficiency impairs apoE secretion most likely secondary to the role of ABCA1 in adding phospholipids and cholesterol to apoE (25, 31, 32) and have shown increased apoE secretion in Abcg1–/– macrophages associated with upregulation of ABCA1 (25). Macrophage apoE accumulation in media was measured in freshly isolated peritoneal macrophages or following loading of macrophages with acLDL plus liver X receptor (LXR) activator (T0901317) to maximally upregulate apoE expression and secretion (33). ABCG1 protein was increased in Abca1–/– cells, while ABCA1 protein was increased in Abcg1–/– cells (Figure 5), reflecting compensatory induction of their cognate mRNAs (data not shown). There was a small decrease in apoE secretion from Abca1–/– cells under basal conditions and an increase in apoE secretion in Abcg1–/– cells (Figure 5), as expected (25, 32). ApoE secretion was profoundly decreased in Abca1–/–Abcg1–/– cells compared with control and Abcg1–/– cells and was even significantly lower than in Abca1–/– cells (Figure 5). These effects were posttranscriptional: apoE mRNA was not changed in Abcg1–/– cells and slightly increased in Abca1–/– and Abca1–/–Abcg1–/– macrophages compared with WT cells (data not shown). Cell lysates from Abca1–/– and Abca1–/–Abcg1–/– macrophages showed amounts of apoE protein similar to those of WT cells, consistent with a defect in secretion rather than intracellular apoE degradation as the underlying cause (Figure 5). These findings suggest that the increase in apoE secretion in Abcg1–/– cells is secondary to induction of ABCA1 and indicate a possible role of ABCG1 in supporting residual apoE secretion in Abca1–/– cells. The profound defect in apoE secretion in Abca1–/–Abcg1–/– macrophages likely contributes to the increased atherosclerosis in Abca1–/–Abcg1–/– BM recipients.
Western blots showing ApoE protein in media and cell lysates of peritoneal macrophages derived from WT, Abcg1–/–, Abca1–/–, and Abca1–/–Abcg1–/–mice. Thioglycollate-elicited peritoneal macrophages were loaded with 50 μg/ml acLDL and treated with LXR activator (3 μmol/l T0901317) for 24 h and then incubated for 16 h in DMEM plus 0.2% BSA. Equivalent volumes of media normalized for cellular protein levels and equal amounts of cellular proteins (20 μg) from each sample were used for western blot analysis. Numbers below blots indicate change in KO cells compared with loaded control cells from 3 independent experiments. *P < 0.05 versus WT. ΧP < 0.05 versus single KOs.
Increased secretion of inflammatory cytokines and chemokines by Abca1–/–Abcg1–/– macrophages. In order to understand the increased infiltration of inflammatory cells in the tissues of Abca1–/–Abcg1–/– mice, we measured the secretion of inflammatory cytokines and chemokines into media by freshly isolated peritoneal macrophages not treated with exogenous inflammatory stimuli during 6 h in cell culture. This showed increased secretion of a variety of inflammatory cytokines and chemokines in Abca1–/–Abcg1–/– macrophages, notably TNF-α, IL-6, IL-1β, and IL-12 as well as MIP-1α, MIP-2, growth-regulated oncogene α, and, to a lesser extent, MCP-1 (Figure 6). There was also significantly increased secretion of some inflammatory cytokines and chemokines in Abcg1–/– cells (i.e., TNF-α, IL-1β, IL-12, MIP-1α, MIP-2, and MCP-1), but, except for a slight increase in MIP-2 and IL-12, no major difference in Abca1–/– cells (Figure 6). The increased secretion of inflammatory cytokines and chemokines paralleled mRNA expression changes as determined by real-time PCR (data not shown) and likely explains the increased tissue inflammatory responses in Abca1–/–Abcg1–/– BM recipients.
Inflammatory and chemokine gene expression in WT, Abcg1–/–, Abca1–/–, and Abca1–/–Abcg1–/– macrophages. Peritoneal macrophages were harvested from WT, Abcg1–/–, Abca1–/–, and Abca1–/–Abcg1–/– mice fed high-cholesterol diet for 2 weeks. After a 1-h incubation at 37°C, nonadherent cells were removed and adherent cells consisting of macrophages were incubated in 0.2% BSA DMEM. After 6 h, media were used for secretion analysis. (A) Secretion of chemokines MIP-1α, MIP-2, and MCP-1. KC, growth-regulated oncogene α. (B) Secretion of inflammatory cytokines IL-6, TNF-α, and IL-1β. Secretion levels were normalized to cellular protein amount and expressed as a percentage of WT. Values are mean ± SEM. *P < 0.05 versus WT. ΧP < 0.05 versus single-KOs.
Increased apoptosis after loading with free cholesterol or oxidized LDL in Abca1–/–Abcg1–/– macrophages. The Abca1–/–Abcg1–/– BM recipients showed increased numbers of apoptotic macrophages in the heart and atherosclerotic lesions. Apoptosis of macrophages in atherosclerotic lesions involves different mechanisms and may be brought about by accumulation of oxysterols derived from oxidized LDL or by accumulation of free cholesterol related to uptake of modified LDL and a failure of ACAT-mediated cholesterol esterification (34, 35). In order to understand the accumulation of apoptotic cells in heart and atherosclerotic lesions, we compared the apoptotic responses of macrophages of the 4 different genotypes using oxidized LDL or a standardized free cholesterol loading procedure (acLDL plus ACAT inhibitor 58035) in the presence or absence of HDL (Figure 7). Free cholesterol–induced apoptosis was dramatically increased in Abca1–/–Abcg1–/– macrophages compared with cells of the other genotypes (Figure 7A). Apoptosis induced by oxidized LDL was also markedly increased in Abca1–/–Abcg1–/– and Abcg1–/– cells, but not in cells of the other genotypes (Figure 7B). Treatment with HDL ameliorated apoptotic responses induced by oxidized LDL in WT and Abca1–/– cells, but not in Abcg1–/– or Abca1–/–Abcg1–/– macrophages. The enhanced susceptibility of Abca1–/–Abcg1–/– macrophages to cholesterol- and oxysterol-induced apoptosis likely explains the in vivo findings of increased apoptotic cells in the inflammatory infiltrate in the heart and atherosclerotic lesions.
Apoptosis of peritoneal macrophages after loading with free cholesterol loading or oxidized LDL. Peritoneal macrophages were cultured in DMEM plus 0.2% BSA containing 100 μg/ml acLDL plus ACAT inhibitor (compound 58035; A) or 100 μg/ml Cu-oxidized LDL (oxLDL; B) for 24 h in the presence or absence of 100 μg/ml human HDL. Apoptosis of macrophages was determined by annexin V staining. Results are mean ± SEM. *P < 0.05, genotype effect. #P < 0.05, HDL effect.
Our studies suggest that the combined activities of ABCA1 and ABCG1 in macrophages are central to the antiatherogenic effects of HDL and also account, at least in part, for some of the associated mechanisms that have been considered as potential atheroprotective properties of HDL. Together these transporters have a synergistic role mediating macrophage cholesterol efflux and protecting against macrophage foam cell formation, inflammatory cell infiltration, apoptosis, and atherosclerosis. Because ABCA1 and ABCG1 are both targets of LXR transcription factors (36, 37), when one transporter is deficient, the other is upregulated as a result of sterol accumulation and LXR activation. The role of the individual transporters in these effects is masked by mutual compensation, likely explaining some of the mixed results of previous studies (19–26) and also why individual deficiencies of ABCA1 and ABCG1 produce relatively modest phenotypes.
The elucidation of ABCA1 as the defective molecule in Tangier disease (15–17), explaining the underlying defect in cholesterol and phospholipid efflux from cells to apoA-1 (12, 13), was a major advance supporting the idea that macrophage cholesterol efflux was central to the antiatherogenic effects of HDL (1, 38). The lack of a dramatic atherosclerosis phenotype in Tangier disease patients and in Abca1–/– mice has traditionally been explained by the low levels of VLDL- and LDL-cholesterol that accompany markedly reduced HDL levels. Our findings suggest that another factor is likely to be the compensatory upregulation of ABCG1 in Abca1–/– macrophages. Even though Abca1–/– mice have very low HDL levels, ABCG1 can support cholesterol efflux to other lipoprotein particles (14, 23), and our present studies revealed a further role of ABCG1 in supporting residual apoE secretion and cholesterol efflux to apoA-1 and apoE in Abca1–/– cells.
Tangier disease cells have relatively normal cholesterol efflux to HDL (39), and ABCA1 is almost inactive in mediating cholesterol efflux to plasma HDL particles (13, 14); thus activity of ABCA1 could not readily account for the inverse relationship between HDL levels and atherosclerosis observed in epidemiological studies. The idea that there might be an LXR-activated active transport pathway for cholesterol involving HDL particles (40) led to the discovery that ABCG1 could promote net efflux of cholesterol from macrophages to HDL particles (14, 23). Disappointingly, however, while Abcg1–/– mice accumulated foam cells in lungs and other tissues (23), 2 laboratories found that ABCG1 deficiency in BM-derived cells actually protected mice from atherosclerosis (25, 26) and another found that enhanced expression of ABCG1 in Ldlr–/– mice increased atherosclerosis (41). The results of the present study confirmed that atheroprotection in the former studies (25, 26) is related to compensatory upregulation of ABCA1 in Abcg1–/– BM recipients. Along with recent studies of in vivo macrophage reverse cholesterol transport (30), our findings indicate additive or synergistic effects of ABCA1 and ABCG1 in protecting against macrophage foam cell formation and atherosclerosis. The enhanced sterol accumulation in Abca1–/–Abcg1–/– macrophages led to markedly increased inflammatory and chemokine gene expression and secretion and profoundly increased apoptotic responses, explaining the dramatic in vivo phenotype.
We measured cholesterol efflux from macrophages to isolated HDL or to serum and documented an additive or synergistic role of ABCA1 and ABCG1 in mediating macrophage cholesterol efflux. In the presence of HDL there was still a considerable level of residual mass and isotopic efflux that was not explained by these 2 transporters (Figure 3). A number of other pathways of macrophage cholesterol efflux to HDL particles have been described. These include passive or diffusional cholesterol efflux and SR-BI–mediated cholesterol efflux. These 2 pathways only result in net cholesterol efflux when HDL composition is altered such that there is a relative depletion in unesterified cholesterol relative to phospholipids, for example as may occur with lecithin cholesterol acyltransferase–mediated (LCAT-mediated) cholesterol esterification in HDL. SR-BI is expressed at low levels in murine macrophages, and SR-BI–/– macrophages show no defect in net cholesterol efflux to HDL (42, 43) or in vivo reverse cholesterol transport (30). Moreover, in contrast to synergistic effects on cholesterol efflux resulting from coexpression of ABCA1 and ABCG1 (44, 45), when ABCA1 and SR-BI are coexpressed in the same cell there is inhibition of ABCA1-mediated cholesterol efflux to apoA-1, as a result of a futile cycle of cholesterol efflux to apoA-1, followed by reuptake into the cell via SR-BI (46). These studies speak against a significant role of SR-BI in macrophage cholesterol efflux. However, a number of studies have shown an increase in atherosclerosis in apoE–/– or Ldlr–/– mice transplanted with SR-BI–deficient BM, and a potential in vivo role of SR-BI in macrophage cholesterol efflux cannot be definitively excluded.
A role for macrophage ABCA1 in promoting macrophage apoE secretion has been documented in several studies (25, 31, 32), and in the present study we confirm that induction of ABCA1 in Abcg1–/– cells is responsible for their increased apoE secretion (25). Moreover, compared with Abca1–/– cells, Abca1–/–Abcg1–/– macrophages showed a further decrease in cholesterol efflux to lipid-poor apoA-1 or apoE, in spontaneous cholesterol efflux (Figure 3), and in apoE secretion (Figure 5). These observations provide a hint that ABCG1 has additional cellular activities beyond its ability to promote cholesterol efflux to lipoprotein particles, which might be involved in supporting apoE and cholesterol secretion. The profound decrease in apoE secretion in Abca1–/–Abcg1–/– macrophages is likely an important factor in accelerated atherosclerosis in Abca1–/–Abcg1–/– BM recipients. Macrophage apoE deficiency has marked proatherogenic consequences (29), which are thought to reflect antiinflammatory effects of macrophage apoE secretion as well as the role of macrophage apoE in cholesterol efflux (47).
There was a marked accumulation of apoptotic cells, most likely macrophages, in the heart and atherosclerotic lesions of Abca1–/–Abcg1–/– BM recipients. In an earlier study Abcg1–/– macrophages were shown to have increased susceptibility to apoptosis induced by oxidized LDL, and lesions contained increased numbers of TUNEL-positive cells (26). Recently, we have shown that this reflects a specific role of ABCG1 in mediating efflux of 7-ketocholesterol and related oxysterols from macrophages to HDL (48). The decrease in atherosclerosis was attributed to increased apoptosis of macrophages in early lesions (26), because some mutations that increase macrophage apoptosis result in smaller foam cell lesion formation (49). In contrast, in the present study an increase in apoptosis in Abca1–/–Abcg1–/– BM recipients was associated with increased lesion formation. This does not signify that conclusions from the earlier study (26) were necessarily incorrect, because multiple factors such as decreased cholesterol efflux, decreased apoE secretion, and increased inflammatory gene expression could have been causative for increased atherosclerosis in the Abca1–/–Abcg1–/– BM recipients. However, it is important to point out that macrophage apoptosis has been associated with the critical clinical events associated with atherosclerosis, i.e., plaque rupture and thrombus formation. Our findings suggest that ABCA1 and ABCG1 may have an important role in suppressing macrophage apoptosis in advanced atherosclerotic plaques and protecting against critical plaque complications.
A striking finding was the massive accumulation of foam cells and, to a lesser extent, neutrophils in the heart. HDL has been shown to have antiinflammatory properties, and these have been proposed to underlie antiatherogenic effects of HDL (4). We documented increased secretion of a variety of inflammatory cytokines and chemokines in freshly isolated peritoneal macrophages from Abca1–/–Abcg1–/– mice. MIP-1α and MIP-2 belong to the 2 best-understood classes of chemokines, namely the CXC and CC families, and are potent neutrophil and macrophage chemokines (50). Increased TNF-α, IL-6, and IL-1β secretion from Abca1–/–Abcg1–/– foam cells is likely to lead to induction of genes such as VCAM-1 and ICAM-1 in lesional endothelial cells, leading also to increased recruitment of monocytes and possibly neutrophils into lesions. Previous studies have documented increased expression of inflammatory and chemokine genes as a result of loading of unesterified cholesterol or oxysterols in monocytes and macrophages (35, 51). Together with the efflux and sterol accumulation data, our findings suggest that increased secretion of inflammatory cytokines and chemokines is secondary to sterol accumulation in Abca1–/–Abcg1–/– macrophages. Increased foam cell accumulation in the spleen was also found in Abca1–/–Abcg1–/– BM recipients. It is not clear why foam cells accumulate in certain tissues but not others. Interestingly, Abcg1–/– BM recipients showed foci of mixed inflammatory cells in the lung, as reported previously (26). However, this phenotype was not increased further in Abca1–/–Abcg1–/– BM recipients. This could be related to the specific role of ABCG1 in promoting efflux of oxysterols from cells (48) and our present finding that oxidized LDL-induced apoptosis occurred at fairly similar levels in Abcg1–/– and Abca1–/–Abcg1–/– macrophages. These findings suggest that ABCG1 has a special role in protecting lung cells from damage mediated by oxidative modification of sterols.
Recently, there has been intense interest in the development of novel therapies directed at HDL, such as CETP inhibitors, niacin with flushing inhibitors, infusion of apoA-1 or apoA-1–like peptides, and LXR activators (52, 53). Our studies suggest that the interaction of HDL and its apolipoproteins with ABCA1 and ABCG1 in macrophages is the key to their antiatherogenic properties, involving relatively small changes in sterol pools in specific inflammatory cells, and that such atheroprotective effects probably do not require an overall stimulation of reverse cholesterol transport to the liver. Treatment with niacin or CETP inhibitors leads to increased HDL, apoA-1, and cholesterol levels and is associated with prominent accumulation of large HDL-2 type particles. Importantly, large apoE- and LCAT-rich HDL particles accumulating in genetic CETP deficiency or in subjects treated with CETP inhibitors have increased ability to promote cholesterol efflux from macrophages via ABCG1 (54). While the CETP inhibitor torcetrapib failed in a recent clinical trial, this may have been related to underdosing and off-target toxicity (55). Our results confirmed the role of ABCG1 in protecting against atherosclerosis and were consistent with the idea that enhancing macrophage sterol efflux via ABCG1 could have antiatherogenic consequences. While our studies suggest that the beneficial effects of increasing sterol efflux via ABCG1 could be partly offset by decreased ABCA1 expression, it is interesting to note that ABCG1 appears to have a unique role in protecting against macrophage apoptosis induced by oxidized LDL (48) and in suppressing inflammatory and chemokine gene expression (Figure 6). Thus, increasing HDL by niacin or CETP inhibitors could have antiinflammatory and antiapoptotic effects in advanced atherosclerotic lesions, and perhaps in some inflammatory lung diseases, as a result of promotion of cholesterol and oxysterol efflux via ABCG1. In principle, strategies that increase macrophage sterol efflux by upregulating both ABCA1 and ABCG1, or by increasing overall levels of particles or peptides that increase efflux via both transporters, could ultimately prove to be the most effective therapies for increasing macrophage sterol efflux and decreasing atherosclerosis.
Mice. Abca1–/– mice in DBA background were obtained from The Jackson Laboratory, and Abcg1–/– backcrossed into the C57BL/6 background were originally purchased from DeltaGen. Because we were unable to breed mice with combined deficiency in the C57BL/6 background, we bred them in a mixed C57BL/6 × DBA background and transplanted BM from these animals into the genetically uniform F1 generation obtained by crossing C57BL/6 Ldlr–/–mice with DBA mice (The Jackson Laboratory). The malformation of the placenta in C57BL/6 Abca1–/– mice resulting in severe embryo growth retardation, fetal loss, and neonatal death was most likely related to the difficulty of breeding the Abca1–/– trait into the C57BL/6 background (56). Animal protocols were approved by the Institutional Animal Care and Use Committee of Columbia University. Animals had ad libitum access to both food and water.
BM transplantation. BM transplantation was performed as previously described (57). The atherosclerosis studies were conducted in moderately hyperlipidemic (Supplemental Table 1) F1 hybrid C57BL/6 × DBA Ldlr+/– mice fed high-fat/cholesterol/bile salt diet containing 1.25% cholesterol, 7.5% cocoa butter, and 0.5% cholic acid (TD 88051; Harlan Teklad). A similar approach has been used previously (58–60). Combined results of males and females were shown for each genotype: similar results were obtained with data divided by sex. Mice were allowed to recover for 5 weeks after irradiation and BM transplantation before diet studies were initiated.
Histological analysis of the myocardium and proximal aortas. The myocardium and proximal aortas were serially paraffin sectioned and stained with H&E as previously described (57). Aortic lesion size of each animal was calculated as the mean of lesion areas in 5 sections from the same mouse. Mac-3 and MCA771G antibodies (BD Biosciences — Pharmingen) were used to stain macrophages and neutrophils in paraffin sections, respectively. Apoptotic cells were detected using TUNEL assays (57).
Isolation of HDL from plasma. HDLs (density, 1.063–1.21 g/ml) were isolated by preparative ultracentrifugation from normolipidemic human plasma and stored in PBS containing 1 mM EDTA. ApoB-containing particles was precipitated from serum by adding 100 μl serum to 40 μl 20% polyethyleneglycol (PEG; P-2139 in 200 mM glycine, pH 10; Sigma-Aldrich) solution. This mixture was incubated at room temperature for 15 min. After this incubation, the solution was centrifuged at 1,900 g for 20 min. The supernatant, containing HDL fractions, was removed and used for experiments.
Isolation of mouse peritoneal macrophages. Fresh peritoneal macrophage cells were harvested from WT, Abcg1–/–, Abca1–/–, and Abca1–/–Abcg1–/– mice 3 days after receiving an i.p. injection of thioglycollate and plated in 10% FBS in DMEM as previously described (25). After a 1-h incubation at 37°C, nonadherent cells were removed and adherent cells consisting of macrophages were used for the experiment as described in the Figure 6 legend.
Isotopic cholesterol efflux assay. Thioglycollate-elicited macrophages were cultured for 24 h in DMEM plus 0.2% BSA containing 50 μg/ml acLDL, 3 μmol/l T0901317, and 2 μCi/ml of [3H]-cholesterol. Cholesterol efflux was performed for 6 h in 0.2% BSA DMEM containing different acceptors as described in the Figure 3 legend. The cholesterol efflux was expressed as the percentage of the radioactivity released from the cells in the medium relative to the total radioactivity in cells plus medium (14).
Cholesterol mass analysis. Thioglycollate-elicited macrophages were first incubated in 0.2% BSA DMEM with LXR agonist (T0901317, 3 μmol/l) and with 50 μg/ml acLDL for 24 h. Then, cholesterol efflux was performed for 6 h in DMEM containing 0.2% BSA in presence or absence of HDL (50 μg protein/ml). After incubation with HDL, the lipid fractions were extracted from the collected media with hexane or from cells using hexane/isopropanol (3:2, vol/vol) in presence of stigmasterol (5 μg per sample) added as the internal standard. Total cholesterol was determined after saponification by gas-liquid chromatography. The HDL-mediated net cholesterol efflux was calculated as the difference of cholesterol mass of the medium cultured with and without cells (14). The cholesterol mass in cells was determined from briefly cultured macrophages harvested from mice fed standard chow or high-cholesterol diet for 2 weeks.
Western blot analysis. The expression of ABCG1, ABCA1, and apoE were measured in peritoneal macrophage cells by Western blot analysis. Primary antibodies for ABCG1, ABCA1, and apoE were purchased from Santa Cruz Biotechnology Inc., Abcam, and Biodesign, respectively.
Secretion analysis. Mouse peritoneal macrophages were harvested and cultured for 6 h in DMEM containing 0.2% BSA. Cytokines and chemokines were measured by Luminex assay (Cytokine Core Laboratory).
Apoptosis analysis. Macrophages were incubated in 0.2% BSA supplemented with acLDL (100 μg/ml) plus ACAT inhibitor (compound 58035) or Cu-oxidized LDL (100 μg/ml) for 24 h with or without HDL (100 μg/ml). Apoptosis of macrophages was determined by annexin V staining as previously described (57). Free cholesterol–induced apoptosis was not prominent in WT macrophages because we used thioglycollate-elicited macrophages in these experiments (I. Tabas, personal communication).
Statistics. Data are shown as mean ± SEM. Comparison of mean values between groups was evaluated by 2-tailed Student's t test or ANOVA with square root–transformed data. A P value less than 0.05 was considered significant.
View Supplemental data
This work was supported by grants from the NIH (HL 54591) and the Philippe Foundation.
Nonstandard abbreviations used: LXR, liver X receptor; SR-BI, scavenger receptor–BI.
Conflict of interest: A.R. Tall has received consulting fees from Pfizer, Merck, AstraZeneca, and Roche; lecture fees from Merck; and grant support from Merck and Pfizer.
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A large part of the hold that organized baseball has on the American imagination derives from the concept of loyalty to the franchise rather than to individuals. In a world reeling from the rural to the urban, in a country where one does not live where he grew up, in the ever changing megalopolis, loyalty to a baseball franchise offers the same assurance and stability that 'home from the holidays' does.
I'm sure there are several ballparks around the MLB that look similar to this right now, especially in the Northeast. Fortunately, pre-season ball is played in much warmer climes than this. Three weeks to Spring Training!
I stumbled across this graph this morning depicting Ted Williams's batting average in various parts of the strike zone. I wish the numbers were bigger, but if you can make them out, it is pretty mind-blowing. No pitch was safe anywhere near the plate. This is how you hit .400 for a season.
On January 26, 1957, Joe Cairnes became president of the Braves, replacing Lou Perini. Perini would best be remembered as the man who moved the Braves from Boston to Milwaukee in 1953, citing financial losses and poor attendance.
This may be a reality sooner than we realize.
I've always approached spring training as I have something to prove.
I like the bald honesty of this piece. As much as many folks don't like to think about it, race and ethnicity have played a role in the history of baseball. Fans sometimes, albeit unfairly, have preconceived notions about what a baseball hero ought to look like. This poem is also beautifully written; full of imagery, evoking feelings of nostalgia.
As a comb of cells for the honey.
And blonds could be left-handed.
In Long Branch, New Jersey.
Of the sky. Tight red stitches.
At rest and of blank paper.
Than Whitey Ford: A Dodger.
People were amazed by him.
He refused to pitch on Yom Kippur.


text:
Popular Tesco store in Weymouth to be sold
Negotiations for sale of Tesco and Subway in Weymouth
By Ellie Maslin @dorsetechoellie Reporter
Tesco Express in Weymouth is up for sale along with a takeaway next door Picture: Finnbarr Webster
A BUSY convenience store in Weymouth is up for sale along with a takeaway and residential flats - with negotiations currently underway.
A prospective buyer has come forward after the Wyke Regis Tesco Express and adjoining Subway sandwich restaurant at 67 Portland Road went on the market for £2.5 million last year. The sale also includes four residential apartments above the Tesco building, which are let privately.
Estate agents Savills, marketing the property, says it is of 'far superior quality compared to local competition' due to its prominent location. The convenience store and takeaway unit is situated on the only route through to Portland - 'ideally positioned to service the surrounding population of Wyke Regis, as well as Portland and tourists to the area,' the estate agents note.
Tesco, Subway and four residential flats on Portland Road, Weymouth are up for sale
Yesterday a spokesman for Savills confirmed that negotiations for the sale are underway, "with an announcement expected by end of January."
The 0.4 acre site, including the flats, represents a total size of 7,520 square foot and includes the customer car park comprising 18 parking spaces, four of which are reserved for the residential flats.
The complex was purpose-built on land occupied by the former Texaco petrol station, which was demolished in 2018 to make way for the development.
The former Texaco petrol station on Portland Road was demolished to make way for the complex
The building is described as brick-built, rendered in part with Trespa 'slate' cladding, featuring Juliet balconies on the flats along the front and north west side of the property.
On the first floor are two two-bedroomed and two one-bedroomed flats, 'all fitted to a high specification' and with separate access to the shops.
READ MORE: Wyke Stores to close after 160 years over competition with nearby Tesco
The site is offered as a modern freehold investment opportunity, with the Tesco branch let on a 20 year lease until 2038, currently at a rate of £85,000 per year, while Subway is let on a 15 year lease until 2033, currently at £15,000 per year.
The one-bed flats fetch £600 pcm while a rent of £775 pcm is charged for each of the two-bedroomed properties. They are valued at around £160,000, £165,000, £190,000 and £200,000
According to the listing, a total commercial income of £100,000 per annum is generated from the site - 85 per cent from Tesco and 15 per cent coming from the Subway branch.


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