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Worthy is the Lamb - Hillsong United (Karaoke-Track) MP3
Christians Karaoke.
worthy is the Lamb - Karaoke MP3
Hillsong download instrumental http://www.aimini.net/view/?fid=ni8qSQN3piEYluT0umLN.
Worthy Is The Lamb Hillsong Karaoke MP3
I have cropped this video ti make the words appear bigger than the original posted on youtube.
Hillsong - Worthy Is The Lamb Karaoke Instrumental MP3
Download MP3: http://go.earlpeter.com/download/youtube/audio/WorthyIsTheLamb For pitch adjustments, send a personal email concerning your request to ...
Worthy is the Lamb - Minus One Instrumental MP3
AGNUS DEI - Lamb of God - Instrumental MP3
By Michael W. Smith. Alleluia! Alleluia! For the Lord God Almighty reigns. Alleluia! Alleluia! For the Lord God Almighty reigns. Alleluia! Holy, Holy are You, Lord ...
Agnus Dei (Worthy Is The Lamb) Instrumental MP3
I do not own this music nor is any copyright infringement intended you can download the music here ...
Worthy Is The Lamb, Instrumental, (Hillsong) MP3
I do not own this song and no copy right infringement is intended. You can purchase this music from worship-downloads.com.
Worthy is the lamb ♪ Jotta A- a real Talent ♫ MP3
Worthy is the lamb - Agnus Dei by Jotta A Kids talent show.
Worthy is the Lamb - Hillsong (Instrumental Cover) MP3
One take Worthy Is The Lamb originally by Hillsong Covered instrumentally. Hope you enjoy it :) Keyboard by Amos Ghui Mixing, recording by Amos Ghui ...
Handel - Messiah - Worthy is the Lamb + Amen (score) MP3
Darlene Zschech - Worthy is the Lamb (with Lyrics) MP3
Enjoy! Please Comment.
Worthy the Lamb [Live] MP3
Music video by Bill & Gloria Gaither performing Worthy the Lamb (feat. Gaither Vocal Band) [Live]. (P) (C) 2012 Spring House Music Group. All rights reserved.
Worthy Is The Lamb - Low key performance track w/ background vocals MP3
Provided to YouTube by Warner Music Group Worthy Is The Lamb - Low key performance track w/ background vocals · Worship Tracks Worship Tracks - Worthy ...
Worthy is the Lamb - Hillsong (karaoke vietnamese) MP3
Revelation Song - Instrumental with lyrics MP3
Maranatha Top 25 Praise songs instrumental (2012 edition) Itunes link: http://itunes.apple.com/us/album/top-25-praise-songs-instrumental/id507733950 Worthy ...
Digno es el Senor (Worthy is the Lamb Spanish Translation) - pista instrumental piano, karaoke MP3
Pueden descargar la pista instrumental del piano aqui: https://dl.dropboxusercontent.com/u/3033081/pistas/digno%20es%20el%20senor%20pista%20fx.mp3.
[Karaoke] Worthy is the Lamb - Tôn Vinh Chiên Con - Hillsong Lời Việt: Lê Đình Tuấn MP3
Ton Vinh Chien Con - Hillsong Worthy is the Lamb Hillsong Trinh bay: Phuong Trinh Loi Viet: Le Dinh Tuan Hòa âm & Phối khí: Quốc Thi Video Karaoke by: ...
Worthy The Lamb - Gaither Vocal Band MP3
Hear the cries of the shackled from the onset of time, For the chains of defeat theres no key. See the tears of the broken, the cries of the slaves: Is there no one ...
Worthy is the Lamb sung by me Karaoke MP3
Me singing with soundtrack. I love this song.


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Each month, Rated People connects over 50,000 homeowners with quality tradesmen - simply enter your job details in the form above to get started.
We'll match your job to the relevant tradesmen in your area - those interested will pay to see your contact details and call you to quote.
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After the work is complete, return to the site and rate your tradesman based on their quality of work, reliability and value for money.
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The Bescor X-672 is a 6 VDC battery specifically designed to work with the LED-60 light. It offers more run time than the X645 and comes with the BC-665R charger.


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The God Question: Deistic Relevancy and the Size of ChristSubmitted by KnockEmOuttt on November 24, 2012 - 11:31pm.
The God question is one we constantly find ourselves both asked and asking. Does God exist? It's quite a tricky one, especially since there are so many possible answers to such a simply complex question. However, this question can be made much easier once one realizes that there is not one, but two questions here; the big question and the small question. The big question is whether or not there is any kind of being, sentient or not, which can be considered solely responsible for the creation of the universe and everything in it. The small question is the more specific form, the one which must be asked on a religion by religion basis. Once we determine the question being asked, the answer becomes quite clear. The answer to the small question is no. Gods like Zeus, Thor, Yahweh, Amun-ra et al. almost certainly do not exist. Their nature and deeds are mythologies invented by man. As for the big question, the answer is largely irrelevant.
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I'm signing the petitions for the red states to secede from USASubmitted by Sapient on November 14, 2012 - 10:22am.
There are some people from the red states that are currently having a public temper tantrum to secede from America. Why not let them? The red states are a drain. They have higher rates of homicide, infant mortality, suicide, and unemployment. They tend to take higher amounts of federal funds, they are less educated, less diverse, less tolerant, have a lower IQ, and are more religious.
I'm all for embracing America, I believe we should try to work together rather than divide. I would never propose a secession for my state when I am frustrated. When George Bush won as a result of fraud and the Supreme Court selected him in another election you didn't see me proposing we secede. I knew the damage he would cause, I was disgusted, I lived every day of those 8 years upset at who was in the White House. I lived every day of those 8 years fearful of what he would do next and sad about the extreme damage he was causing to this country. I did not propose a secession. Instead I fought to inform the public. And in doing so I'd like to believe that I played a very small role in ensuring that we have a better leader now. Now it's their turn.
Open letter to Christians against gay marriageSubmitted by Sapient on November 13, 2012 - 12:35am.
I recently had an exchange with a Christian friend who was "disgusted" at the states that approved gay marriage. He believes the people who voted for it are "disgraceful." Christian disapproval of gay marriage was the impetus behind this open letter that will hopefully open the eyes of a few of my Christian readers.
Marriage was initially created by men many years ago to control women, at a time much more archaic than the time we live in now. It was created at a time that we would stone someone for being gay as well as for cursing at their parents. Many things have changed in society since marriage was first created. We now are able to traverse the planet to find a mate and no longer need to claim one within walking distance as our property. Today marriage is viewed much differently… so is stoning. In fact we are so far away from the original reasons behind the creation of marriage that nowadays it can feel like marriage is a means for women to control men.
Indoctrinated home schooled girl refuses life in the kitchen and seeks educationSubmitted by Sapient on November 12, 2012 - 1:13pm.
The atheist community is firmly united in assisting a young girl who was brought up in a religious home that seeks to keep women uneducated and in the kitchen. Despite her parents refusing to help in any way she has dedicated herself to getting an education and saving her siblings from the same fate. Please "vote" for her essay about this to help her get a scholarship. As of my writing of this blog post she has twice as many votes as the second place entry, let's keep it up!
In her essay she states:
Then, I would love to create a safe-house where children of religious homeschooling families can come to improve their lives and future and have hope.
From struggling with poverty, to being unable to get a job without a resume or employment history, and often a substandard education; the children of the Quiverfull movement have extreme challenges to overcome. A home like this would be a safe platform for them to transition into the world without being abandoned to struggle. Many of their parents wish this difficulty for their children, saying it will cause them to return and be content with the party line. I want to create a buffer for the pain parents cause when they shun and disinherit their children for religious reasons.
Because there are more of us every day, and so much pain and heartache.
Girl mystified at why she got fired?Submitted by digitalbeachbum on November 12, 2012 - 6:15am.
Stupid as a stupid does..
I'm all for people having their opinions but saying what this immature idiot said is one of those things you keep to yourself. You don't go on Facebook and blurt it out, "I hope is gets assassinated". Comments like these are immature and racist; she deserves to be investigated and she deserves to lose her job.
The hate coming forth from this country is ridiculous even if it is a minority voice. White people need to get a grip and realize that they will soon be a minority. They will soon be on the short end of the stick. If they are not careful they will soon be living a life like in the movie "White Man's Burden".
Time to take your heads out of your asses and realize that their is no place in this world for hate; hate stalls progress and is a regressive emotion.
Sen. Mitch McConnell is a...Submitted by digitalbeachbum on November 7, 2012 - 4:02pm.
I was searching Google for information on Sen. Mitch McConnell and I found the following... the first one is actually (douchebag)..
Why Romney LostSubmitted by digitalbeachbum on November 7, 2012 - 3:31pm.
I found this article http://t.news.msn.com/politics/why-mitt-romney-lost and thought it was worth the read.
The GOP has completely lost it and while this hijacking has been slowly taking place over the last twenty years, it has reach a point of either turning back toward the middle and dumping radicals/tea party idiots.
There is no place that the GOP can survive unless they return to the middle. No matter how much money the Koch brothers invest, radicalism eventually leads to the destruction of the country and the people. If that is what they want, then they should be considered enemies of the state.
Greetings From the Negative Zone! (Election Day Special)Submitted by Sage_Override on November 6, 2012 - 8:01pm.
Well, it's certainly been very interesting lately in this place. The people have pretty much accepted me without going through a thorough background check or asking me a lot of questions. They just seem to amalgamate anything and everything that seems non-threatening. Granted, I was KICKED through the temporal nexus so to speak that brought me here, but we're just splitting hairs at this point.
So, yes, I hope everything has been good for the rest of you. To those mocking my experience in this place as a bad acid trip, you can go fuck yourselves. Thank you. Now then, I know I haven't been updating at all or lurking around here, but I've been busy with some dignitaries here and they recently invited me to one of their "Civil Commons" gatherings which is what I think we should call ours if such a thing existed in politics today. No, what WE have is arrogance, greed, lobbyists, Wall Street banksters and liars. Ain't America grand?
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Go vote for Obama!Submitted by Sapient on November 5, 2012 - 9:18pm.
This is an appeal to those not planning on voting. Please vote! When polled those who claim they are not voting, if they had to vote would vote Obama 3:2. You (the non-voter) are the biggest voting block. Go to the polls non-voters!
"I can't promise you that your life will get better, easier under Barack Obama. I do think he cares and I know for sure that if the other guy is sitting in the Oval Office, I can guarantee you that not only will your life not get better, it will get much, much worse. Don't take my word for it. Just ask your parents what life was like before a 30-year pillage by the Republicans of the middle class. Your parents bought a house and eventually owned it outright. They weren't in debt. College was free. They bought a new car every 3 or 4 years. They took vacations and were home for dinner by 5 or 6 PM. They had a savings account in the bank. They didn't live in fear of not knowing if they'd even have a job next year.
That's all gone. I don't know if we can get it back, but I do know that Mr. Romney would love the chance to complete the final elimination of the middle class and the American Dream. " - Michael Moore
Read the entire appeal from Michael Moore to urge you to vote Obama.
Free Thinking, Free LearningSubmitted by digitalbeachbum on November 4, 2012 - 8:30am.
I've been a member of Khan Academy for about five months now. I occasionally log on and watch a few videos to see if I can further my awareness on subjects which I have never been exposed to; most of which seem to be things about finance.
If you have not yet been to Khan Academy then I suggest you visit it. It's free and you get to keep track of what you learn and test yourself with quizzes.
What is amazing about this project is how it got started. You can read a bit here.
But even more interesting is how much money we (as Americans) spend on education but are one of the worst countries in the world for learning.


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Institutional Diversity, Agency and Governance for Sustainable Value
Extant views of (shareholder) value, (corporate) governance, and competitiveness have a narrow view of 'agency', a poorly developed theory of value and pay little attention to sustainability. In this paper we develop a perspective on the determinants of value-wealth creation at the firm, meso-, and national levels, explore the limitations of extant theory of the firm, concerning governance and value in its context, and discuss some prerequisites of sustainability. We conclude that the pursuit of value is not antithetical to, but it derives from, the notion of sustainability, that sustainability requires both internal and external controls and that institutional diversity can help effect mutual 'stewardship' and monitoring. Moreover, for sustainable value creation, corporate governance needs to be aligned to national and global governance.
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Please report citation or reference errors to , or , if you are the registered author of the cited work, log in to your RePEc Author Service profile, click on "citations" and make appropriate adjustments.:
- Alchian, Armen A & Demsetz, Harold, 1972.
"Production , Information Costs, and Economic Organization,"
American Economic Review,
American Economic Association, vol. 62(5), pages 777-795, December.
- Armen A. Alchian & Harold Demsetz, 1971. "Production, Information Costs and Economic Organizations," UCLA Economics Working Papers 10A, UCLA Department of Economics.
- Young, Allyn A., 1928. "Increasing Returns and Economic Progress," History of Economic Thought Articles, McMaster University Archive for the History of Economic Thought, vol. 38, pages 527-542.
- Richardson, G B, 1972. "The Organisation of Industry," Economic Journal, Royal Economic Society, vol. 82(327), pages 883-896, September.
- Fama, Eugene F, 1980. "Agency Problems and the Theory of the Firm," Journal of Political Economy, University of Chicago Press, vol. 88(2), pages 288-307, April. Full references (including those not matched with items on IDEAS)
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Mal Olson writes adrenaline-kicked romantic suspense. When her consuming passion for writing allows time, she enjoys reading, flower gardening, jamming with friends on the mountain dulcimer, and hiking in a nearby state forest (or in the mountains somewhere). She has three grown children and one granddaughter and resides with her own special hero in southeast Wisconsin where she juggles writing time with her freelance landscape design business.
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on Oct. 20, 2012 :
(review of free book)
Review by:
Ellen Mansoor Collier
on Aug. 18, 2012 :
A great read that highlights the trials and troubles of military dogs in an entertaining, often humorous way.
Maybe more broken dogs will get adopted with this romantic, kind-hearted tale of lost and found.
(review of free book)


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Related terms for "confined space pump ":
confined space companies
We are one of the UK's leading manufacturers and installers of sewage pumps and packaged pumping stations. With over a decade of industry leading experience we are well known as `The Pump People`.Established over a decade ago our attention to detail, high quality products and professional approach to customer requirements has taken us to a nationwide service who provide a ...
based in Kent, UNITED KINGDOM.
24 hour spill response - sewer cleaning, storm drain cleaning and more. United Pumping is a family owned Minority Business (MBE) specializing in servicing hazardous and non-hazardous enviromental waste needs of private and public sector clients for over 37 years. Perform emergency spill response, vacuum trucks, vactor guzzlers, vactor jetters, portable jetters, ...
based in City of Industry, CALIFORNIA (USA).
Fast Flow Pump is American owned and operated. All hydraulic submersible pumps and hydraulic power units are assembled in the USA and are made from high quality components. Fast Flow provides superior pumping systems that outperform the competition. Our pumps provide maximum performance in a compact footprint. Fast Flow Pumps are lean, full performance pumps, built on ...
based in Moss Point, MISSISSIPPI (USA).
Pumps & Dewatering Equipment, Traffic Control, Pipeline Testing Equipment, Confined Space Entry, Shoring Equipment, Construction Lasers, Contractors Hardware, Training Services.
based in Ingleburn, AUSTRALIA.
Cat Pumps has proven to offer the most dependable high-pressure piston and plunger pumps available in the marketplace. Designed and built to the highest quality standards, Cat Pumps provide industry-leading life. With a performance range of 1 to 240 gpm and 100 to 10,000 psi, customers choose Cat Pumps due to the low life cycle cost, providing exceptional value over the ...
based in Minneapolis, MINNESOTA (USA).
STAS Safety Services have a reputation for providing top quality safety training and products to all industry sectors. Management and Supervision of confined spaces: Confined space standby rescue, confined space labour, compiling on-site risk assessments, compiling safe systems of work, tunnel work, assisted decommissioning of plant, culvert work, pumping out services, ...
based in Gloucester, UNITED KINGDOM.
HCP Pumps America is one of the most successful Submersible pump manufacturers in the world. Since the inception of our company, our staff is always enthusiastic in developing and implementing better hydrodynamic techniques with lower consumption, and offering the best customer service in an effort to gain the highest satisfaction for all of our customers.
based in Foley, ALABAMA (USA).
The Pump Technology Ltd, National Pumping Partner network has been providing customers with prompt and economical on-site support since 1994. For- Site Survey Installation Commissioning Routine Servicing / Maintenance Emergency Repair. 24/7. Refurbishment of Existing Equipment Identification & Replacement of Existing Pumps, Like for Like, or Equivalent.
based in Aldermaston, Berkshire, UNITED KINGDOM.
Afton specializes in making what other pump manufacturers consider a specialty item. With more than 50 years experience making quality pumps, Afton can take your specific application requirements, engineer and manufacture a pump suited for your industry. Most models are available with API-610 Hydraulic Institute conformance. Tell us how you want your pump built and we ...
based in Houston, TEXAS (USA).
Welcome to PD Pumps your first call for Submersible Pumps and Dry Installed Pumps At PD Pumps we specialise in the supply of Submersible Pumps and Dry Installed Pumps. Our operations are extremely flexible, ensuring that the whole team - sales, design, test, service and repair - is available at all times to provide a completely satisfactory result for our client.
based in Stockport, Cheshire, UNITED KINGDOM.
Pump Systems Ltd is one of Australia and New Zealand`s leading suppliers of chopper pumps and solids handling pumps for the combined municipal and industrial wastewater sectors. As the Authorised Representative for Vaughan Chopper Pumps in Australia and New Zealand, we provide specialist expertise for the most challenging solids pumping and tank mixing applications.
based in Christchurch, NEW ZEALAND.
SPP Pumps requires the highest standards of manufacturing excellence from its facilities around the world. This is crucial to the on-going growth and development of the company. At the main manufacturing facility located in the UK, SPP Pumps set the highest standards attainable in the industry for quality and reliability.SPP Pumps distinguishes its product split between ...
based in Theale Reading, UNITED KINGDOM.
Located in the north west of England, Centrifugal Pump Services offers a professional and considerate approach to their customers whether it be a multi-national company, local industry or the general public. Centrifugal Pump Services can offer a vast knowledge due to the many years of experience gained by its management and staff, whether it is sales, service or ...
based in Stockport, UNITED KINGDOM.
Pumping Services, Inc. is a stocking distributor and service facility for the world's leading manufacturers of equipment used in the treatment and transportation of water, wastewater and other fluids. We serve the municipal, industrial, construction, commercial and utility markets in New Jersey, New York and surrounding areas. We compete by providing excellent service, ...
based in Middlesex, NEW JERSEY (USA).
Detroit Pump & Mfg. Co. is a leading distributor of pumps and fluid handling products representing many of the leading pump manufacturers. The company provides services including local engineering support, inside and outside sales, pumps and parts inventory, fabrication, repair and warranty to our valued customers. The company pursues opportunities in the State of ...
based in Grand Rapids, MICHIGAN (USA).
Distrimex Pumps & Service Ltd is a manaufurer representative in the field of pumps and active throughout the Netherlands. We supply a wide range of pumps and brands, including Calpeda, Wangen, Pumpex, Ritz and Tsurumi.
based in Doetinchem, NETHERLANDS.
Established in 1992, Pump Pro`s has been continually expanding to encompass any fluid handling need that customers may have. In 2003 Pump Pro`s purchased Wagner-Smith Pumps and Systems (est. 1917) then in 2008 purchased Jenkins Equipment (est. 1952). More recently, Pump Pro`s has grown to include two new divisions, Municipal and Cryogenics.
based in Mason, OHIO (USA).
We specialise in working with companies that are involved in confined space and hazardous environments. If you need support with confined space training, rescue teams for dangerous situations or the purchase and servicing of your safety equipment then call us today for an appointment.. By 2011 due to the success of the confined space training sector the company moved ...
based in Huntingdon, UNITED KINGDOM.
Bale Group Ltd is a one-stop hazardous and non-hazardous liquid waste management solution, specialising in specific fields. These include, Bulk liquid Haulage, Deep suction and jetting, confined space & industrial Cleaning, Pollution control, drummed/packaged hazardous chemical disposal, oil & water recycling, organic waste recycling, tanker hire and road tanker ...
based in Honiton, UNITED KINGDOM.
We work safe. We work hard. We work smart. We stay current with training and technology. And we exercise good old-fashioned common sense in getting environmental maintenance and cleanup done right—safely, on time and on budget. Deep within confined industrial spaces and in the great outdoors, on highways and waterways, in business districts and along shorelines, on land ...
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Hi kids and welcome to Monday!
The Heat won, Magistrate Judge Vitunac retires in style, and it rained all night.
How can you stop the rain from falling down?
RIP Robin Gibb.
He will be missed.ReplyDelete
Three singers this month died from cancer: Adam Yauch, Donna Summer & Robin Gibb. Sometimes I wish cancer would get cancer and once and for all just die.ReplyDelete
Q: How can you mend a broken heart?
A: Time heals the wounds that no one can see.
I will definitely miss Robin Gibb. I am a fan of the Bee Gees, I grew up with their music.ReplyDelete
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Report summaryThis analysis details some of PERTAMINA EP's assets in West Papua, both own-operated and operated by other companies through Technical Assistance Contract (TAC) and Operation Cooperation (KSO) contracts. Although several gas discoveries exist in the area, the volumes discovered are not thought to be commercial given their size and the lack of available markets. The majority of the oil fields are now very mature, and are producing high levels of water cut. Most of the commercial activity ...
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We were still talking to each other normally when he suddenly stopped replying. I've no idea what led him to react this way.
How do I get him to talk to me again? Did I do something wrong? Could it be my personality? Or could he have lost interest in me suddenly?
by the way he's a guy friend whom I have a crush on. Not my boyfriend.
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just dont send him messages for a couple of days and see if he initiate anything. If after a week or two he doesn't message you he's probably not intressted.10|10|0
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(Me and Dually at Young Riders in 2007)
That, of course, is ridiculous. I certainly did not stop improving after my last Young Riders came and went without me; in fact, I like to think I am a considerably better all around rider and horseman than I was when I was trying to qualify. I certainly didn't lose any of my love for horses, and indeed being able to ride without the constant pressure of making Young Riders was refreshing and wonderful, and made me appreciate my equine partners even more. And I certainly became no less of a person for failing to qualify (though admittedly I was probably pretty hard to be around for a while there right when I realized it wasn't going to happen... sorry about that, World).
(Yeah... pretty sure I can ride a little better than I did then)
(Big D... the love of my life)
Maybe someday.


text: Carlyle established its South America Buyouts team in 2008. In Brazil, Carlyle has also invested in CVC, a tourism operator; Qualicorp, a health plan broker and administrator; Scalina, a lingerie manufacturer and retailer; Ri-Happy, a toy retailer; and Grupo Orguel, an equipment rental company.About The Carlyle Group The Carlyle Group (NASDAQ: CG) is a global alternative asset manager with $156 billion of assets under management across 99 funds and 63 fund of fund vehicles as of June 30, 2012. Carlyle's purpose is to invest wisely and create value. Carlyle invests across four segments – Corporate Private Equity, Real Assets, Global Market Strategies and Fund of Funds Solutions – in Africa, Asia, Australia, Europe, the Middle East, North America and South America. Carlyle has expertise in various industries, including: aerospace, defense & government services, consumer & retail, energy, financial services, healthcare, industrial, technology & business services, telecommunications & media and transportation. The Carlyle Group employs 1,300 people in 32 offices across six continents. The Carlyle Group – Website The Carlyle Group – YouTube Channel


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Richie Hayward
August 13, 2010 by staff
Richie Hayward, Little Feat drummer and founding member Richie Hayward died Thursday of complications surrounding liver cancer and pneumonia, which had been fighting since 2009. He was 64.
Playing drums since childhood, Hayward was best known for his 40 years of playing rock band based in Southern California. After auditioning for Frank Zappa's Mothers of Invention Factory then join the short term, Hayward formed Little Feat keyboardist Bill Payne with the original bass singer Bill Payne and Lowell George in 1969.
After releasing their debut album in 1971, the band made six albums, but split shortly after George's death in 1979. It was renovated in 1987 and Craig Fuller recruited as the lead singer. Hayward also recorded and performed with numerous musicians such as Eric Clapton, Bob Dylan, Carly Simon, Tom Waits, John Cale, Peter Frampton and Robert Plant.
Hayward remained as a member of Little Feat until 2009, when he was diagnosed with liver cancer. "] [Richie is hanging in there," said guitarist Paul Barrere Spinner in October. "He's trying to be as positive as possible. It's sad that he can not be on the road. One of us is called at least once a day just to maintain their morale."
Before Heyward died, his bandmates, and amateur musicians organized to raise funds to offset many medical expenses, she had no health insurance. He gave his last concert in July, when he joined Little Feat in the Vancouver Island Music Festival.
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Inicio > Search > YOYOGI UEHARA - Hogar para la comunidad musulmana
- Casa compartida
- Dormitorio
Opinión de residentes
Indonesia By Muhammad, Setyawan and Timbul [16/09/2013]
We would like to come back again and present this Angklung, an Indonesian musical instrument, to the house!
Francia By Anfal Seddik [12/03/2013]
I find myself so lucky to have found SAKURA HOUSE Yoyogi Uehara Muslim House, I think no other accommodation could have suited me more. Since it is made for Muslims, I really feel that my way of living, which is bound by my beliefs, is respected. At the same time, I have the opportunity to meet other Muslim people who come from other countries, which is just awesome! Last but not least, the share house is just 5 minutes walk from Yoyogi Uehara station, and SAKURA HOUSE has always been very kind and helpful. This has made my stay in Tokyo even more enjoyable!
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ITABASHI Muslim-Friendly Apartments
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Estación de Tabata.
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text:
I have a Canon Lide 80 scanner. According to Canon it is "he world's most advanced film-capable flatbed scanner". Yet not so advanced that when I bought my new 64 bit laptop last year they had drivers available. One of the reasons I shifted to Linux was the hope that there might be some way to get this working again outside the Windows environment. I have done a bit of searching and found a thread in German which leads to a page with a lot of downloads I don't understand:
http://www.linux-community.de/Community ... on-Lide-80
Has anyone here solved this problem?
Canon scanner drivers
Questions about hardware,drivers and peripherals
Forum rules
Before you post please read how to get help
Before you post please read how to get help
4 posts • Page 1 of 1
- Level 1
- Posts: 10
- Joined: Wed Sep 11, 2013 9:53 am
- Level 1
- Posts: 26
- Joined: Thu Sep 19, 2013 1:51 pm
- Location: Southwest Montana
Re: Canon scanner drivers
Shifting to Linux in hope that some piece of hardware will work that windows won't handle is a setup for disappointment. The quick Google search I did seems to suggest that it only works with Windows and OS X. My suggestion is to dual boot with XP or 7 and to use your scanner there. As for not being able to read the German site, just copy and past sections of text to Google's translate.
Linux Mint 15 Cinnamon on an Asus P5G41T-M LX Plus, Intel E6800 3.33Ghz Dual Core, 4gb DDR3 Memory, 500GB Hard drive.
- Level 1
- Posts: 31
- Joined: Fri Jul 19, 2013 10:41 am
Re: Canon scanner drivers
Unfortunately Windows is sometimes necessary and this is probably one of those cases. I have a CanoScan scanner that works in a virtualbox running windows 7. Doesn't work in linux however. By running in VB I can use the scanned image in linux. I have several programs that I need that I run under Windows in VB, wish I could completely do away with Windows but in today's world that's almost impossible. I'll give up Windows before I give up my Mint LMDE!
- Level 1
- Posts: 10
- Joined: Wed Sep 11, 2013 9:53 am
Re: Canon scanner drivers
That's annoying. Is Canon the only ones who can make these drivers?
I am wondering whether they are trying to control this or if people just haven't bothered generally?
I am wondering whether they are trying to control this or if people just haven't bothered generally?
4 posts • Page 1 of 1
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Hemoglobin (Hb) potently inactivates the nitric oxide (NO) radical via a dioxygenation reaction forming nitrate (NO3−). This inactivation produces endothelial dysfunction during hemolytic conditions and may contribute to the vascular complications of Hb-based blood substitutes. Hb also functions as a nitrite (NO2−) reductase, converting nitrite into NO as it deoxygenates. We hypothesized that during intravascular hemolysis, nitrite infusions would limit the vasoconstrictive properties of plasma Hb. In a canine model of low- and high-intensity hypotonic intravascular hemolysis, we characterized hemodynamic responses to nitrite infusions. Hemolysis increased systemic and pulmonary arterial pressures and systemic vascular resistance. Hemolysis also inhibited NO-dependent pulmonary and systemic vasodilation by the NO donor sodium nitroprusside. Compared with nitroprusside, nitrite demonstrated unique effects by not only inhibiting hemolysis-associated vasoconstriction but also by potentiating vasodilation at plasma Hb concentrations of <25 μM. We also observed an interaction between plasma Hb levels and nitrite to augment nitroprusside-induced vasodilation of the pulmonary and systemic circulation. This nitrite reductase activity of Hb in vivo was recapitulated in vitro using a mitochondrial NO sensor system. Nitrite infusions may promote NO generation from Hb while maintaining oxygen delivery; this effect could be harnessed to treat hemolytic conditions and to detoxify Hb-based blood substitutes.
- blood flow
- vascular endothelial function
- blood substitute
- vascular biology
endothelium-derived nitric oxide (NO) plays a central role in vascular physiology as an autocrine and paracrine vasodilator molecule that maintains basal vasomotor tone, mediates flow-mediated vasodilation, and inhibits platelet aggregation and endothelium adhesion molecule expression. NO's ability to diffuse as an endocrine molecule in blood is limited by an extremely rapid dioxygenation reaction with oxy-hemoglobin (oxy-Hb) to form the inert nitrate (NO3−) (20, 21, 35, 36, 42, 61, 62), as follows: (1) Despite the near diffusion limited nature of this reaction in vitro, the rate of the NO dioxygenation reaction in vivo is limited by the compartmentalization of Hb within the erythrocyte (2, 6–8, 30, 42, 46–48, 72–74). This compartmentalization of Hb in the red blood cell reduces the rate of the inactivation reaction of NO by Hb by ∼1,000-fold, allowing NO to regulate vasodilation despite the presence of 10 mM intravascular red cell Hb.
During intravascular hemolysis, normal NO physiology is disrupted by the release of cell-free plasma Hb, leading to rapid scavenging of NO, vasoconstriction, and organ dysfunction (52, 79). Repeated episodes of intravascular hemolysis with NO scavenging may play a role in the long-term complications of sickle cell disease and other intravascular hemolytic disorders (1, 17, 19, 41, 59, 65). Similarly, the administration of cell-free Hb-based blood substitutes leads to excessive NO scavenging by Hb and may have contributed to the increased rates of pulmonary and systemic hypertension, decreased organ perfusion, gastrointestinal spasms, and increased mortality in clinical trials of cell-free Hb-based blood substitutes (14, 15, 28, 29, 77). In addition to NO scavenging, another potential mechanism that may contribute to the vasoconstrictive effects of cell-free Hb includes increased oxygen delivery to resistance vessels by cell-free Hb (9, 44).
In addition to endothelium-derived NO, accumulating evidence suggests that an intravascular NO storage molecule exists that can produce NO in hypoxic tissues independent of NO synthase pathways (10, 21, 25, 26, 31, 33, 39, 42, 56, 57, 71). Debate remains about the exact systemic storage form(s) of NO that can account for hypoxic vasodilation, but one potential candidate is the nitrite anion (NO2−) (10, 18, 21, 23, 27, 31, 33, 50, 60, 76). Multiple groups have recently reported vasodilatory effects of nitrite at both physiological and pharmacological doses (4, 10, 13, 34, 37, 43, 45, 49, 51, 53, 56). The proposed mechanisms of nitrite-induced vasodilation involve the reduction of nitrite to NO by acidic disproportionation, xanthine oxidoreductase activity, or the nitrite reductase activity of Hb (10, 11, 24, 31, 33, 49, 51, 54, 80). The first two mechanisms are likely to have limited physiological roles because they require a low pH and near anoxia (21). However, recent evidence suggests that the nitrite reductase activity of Hb likely plays an important role in hypoxic vasodilation and may have significant therapeutic implications for various human disease processes (10, 11, 31, 33).
The reduction of nitrite by Hb is enhanced during physiological hypoxia and as proton concentration increases (5, 16). (2) (3) Analogous to bacterial nitrite reductases (concerted electron and proton transfer to nitrite), the reduction of nitrite to NO requires a one-electron reduction of nitrous acid (33). Within the Hb tetramer, the proton dependence of nitrite reduction decreases from 1.0 to 0.88 secondary to a small stabilizing effect of the proton on the T state (tense; deoxy) of Hb (redox Bohr effect) (16). Hb functions as an allosteric nitrite reductase with a maximal NO generation rate around the intrinsic Hb P50 (50% Hb saturation with oxygen) due to the conformational change of Hb from the R (relaxed; oxy) state to the T state during deoxygenation from artery to vein (11, 31, 33). This dependence of the nitrite reductase activity of Hb on physiological hypoxia and acidosis allows for a metabolic "sensing" mechanism that leads to maximal NO generation in ischemic and hypoxic tissues (11, 21, 31, 33, 42). Supporting this theory, a number of laboratories have confirmed that deoxygenating Hb and myoglobin in the presence of nitrite generates NO in vivo and generates cGMP, causes vasodilation, and inhibits cytochrome c oxidase of the mitochondrial electron transport chain (an NO effect) in vitro (10, 11, 38, 56, 64, 66, 70). In a recent study, Rassaf and colleagues (64) infused nitrite in vivo in mice and measured the change in energy for ATP hydrolysis using MRI spectroscopy. They showed that nitrite generates NO in the heart and that this NO inhibits cellular respiration. Interestingly, this effect was only observed in myoglobin wild-type mice and not evident in myoglobin knockout mice, supporting a mechanism for nitrite reduction to NO that requires myoglobin (64, 70). Thus, Hb and myoglobin can be viewed as oxygen-regulated NO-processing molecules that act as NO scavengers under normoxia via the dioxygenase reaction (Eq. 1) and as NO generators during hypoxia via the nitrite reductase reaction (Eq. 3). We have recently found that this nitrite reductase reaction is coupled to a more complicated nitrite anydrase reaction that generates freely diffusible N2O3, which provides a mechanism for NO escape from the erythrocyte (3).
The allosteric nitrite reductase activity of deoxy-Hb may allow nitrite to have a specific therapeutic role during intravascular hemolysis and during the administration of cell-free Hb-based blood substitutes. According to this hypothesis, as Hb deoxygenates in the microcirculation, it may begin to generate NO as it releases oxygen. Such an effect could theoretically counter NO-scavenging effects of Hb and thus limit vasoconstriction and endothelial dysfunction during hemolysis or infusion of stroma-free Hb-based blood substitutes. To test these hypotheses, we used a previously developed canine model of acute intravascular hemolysis that is characterized by systemic NO scavenging, endothelial dysfunction, and vasculopathy (52). The sequella of intravascular hemolysis in this model manifests as systemic and pulmonary vasoconstriction, a process that reproduces the toxicities evident during the infusion of cell-free Hb-based blood substitutes and during episodes of acute intravascular hemolysis in patients with hemolytic disorders. We performed experiments to characterize the physiological effects of intravenous sodium nitrite and to determine if these effects of nitrite are influenced by the nitrite reductase activity of cell-free plasma Hb during intravascular hemolysis. Furthermore, we directly compared the effects of nitrite in this setting with a conventional NO donor medication, sodium nitroprusside.
Experimental Design
All experiments were approved by the Animal Care and Use Committee of the Clinical Center of the National Institutes of Health. Thirty-two purpose-bred beagles (12–28 mo, 9–12 kg) were studied.
All procedures were performed after the induction of anesthesia with halothane (1–4%) and the initiation of mechanical ventilation. Upon completion, the halogenated gas was terminated, and 100% oxygen was administered until the dog emerged from anesthesia and was extubated. Subsequently, the animal breathed room air spontaneously and was sedated throughout the duration of the experiments. Continuous infusions of medetomidine (sedation; 2–5 mcg·kg−1·h−1) and fentanyl (analgesia; 2.5–20 mcg·kg−1·h−1) were initiated postextubation and maintained for the study duration. Animals were monitored continuously, and signs of pain and distress were evaluated immediately and the infusions adjusted appropriately.
Nitrite infusion.
Using pilot experiments to characterize the beagle-specific pharmacokinetics of sodium nitrite, an infusion of 165 mg sodium nitrite over 6 h (27.5 mg/h) was chosen to be administered during the study to reach a targeted plasma nitrite concentration between 15 and 20 μM. Animals randomized to groups not receiving sodium nitrite received an equivalent rate and total volume infusion of 0.9% NaCl (normal saline) to serve as a placebo control.
Water-infusion intravascular hemolysis model.
A previously developed and validated canine model of water infusion-induced intravascular hemolysis was used in this study (52). Water-induced hemolysis produces direct intravascular hemolysis, thereby maintaining the same intravascular concentration of total Hb during hemolysis while altering the distribution of Hb between the red cell and plasma compartment. In this model, a 6-h infusion of water (rate: 16 ml·kg−1·h−1) produces clinically relevant levels of cell-free plasma Hb (20–300 μM heme), simulating an acute hemolytic episode. The extent of hemolysis increases over time, allowing for a graded physiological assessment of vasomotor dysfunction as plasma Hb levels rise. The final levels of plasma Hb would be analogous to those achieved following coronary bypass surgery or a hemolytic crisis induced by paroxysmal nocturnal hemoglobinuria or acute immune-mediated hemolysis. Control animals receive an equivalent rate and total volume infusion of 5% dextrose (D5W) to account for any potential hypotonic and volume effects of the water infusion on hemodynamics. We used a full-factorial study design with four groups of animals receiving either (1) D5W, (2) D5W plus intravenous sodium nitrite, (3) water, or (4) water plus intravenous sodium nitrite. This design allowed for the determination of the physiological effects of a sodium nitrite infusion, the physiological effects of intravascular hemolysis (water), and the assessment of an interaction between nitrite and hemolysis. Specifically, the interaction statistic tested if the effects of nitrite and Hb were influenced by the nitrite reductase activity of cell-free plasma Hb.
Paired experiments were performed in 20 animals (5 animals/group). In the first week, all animals underwent a baseline study and received a D5W infusion (16 ml·kg−1·h−1) through a central venous catheter to determine the physiological effects of the volume load in each animal. The D5W infusion did not cause hemolysis; it allowed each animal to serve as its own control for the effects of a hypotonic volume load in the model. One week later, animals underwent an intervention study and were randomized to receive a 6-h infusion through a central venous catheter of either (1) D5W (16 ml·kg−1·h−1), (2) D5W (16 ml·kg−1·h−1) plus nitrite (27.5 mg/h), (3) water (16 ml·kg−1·h−1), or (4) water (16 ml·kg−1·h−1) plus nitrite (27.5 mg/h). This paired experimental design allowed for the minimization of animal-to-animal variability by calculating the change for each measurement performed in each animal during the baseline and intervention experiments. Subsequent analyses calculate the differences across treatment groups by subtracting the previously calculated differences within animals (from baseline to intervention experiments) in one treatment group from another treatment group (i.e., comparison of the differences of the differences). This design allowed for analysis of the effects of hemolysis, the effects of sodium nitrite, and detection of any interaction between the two.
Preliminary analysis demonstrated a wider range of hemolysis than previously described secondary to the addition of salt-based therapies (sodium nitrite or sodium chloride) that affected the rate of hypotonic erythrocyte lysis. In these experiments, the 6-h water infusions produced low-rate hemolysis in 50% of the animals and rapid rate hemolysis in 50% of the animals. This created two equal-sized groups of animals with either low or high levels of cell-free plasma Hb, respectively. Both groups had peak cell-free plasma Hb levels that continued to be within a clinically relevant range (20–200 μM heme). Preliminary data analyses also suggested a possible interaction between sodium nitrite and hemolysis that was dependent on the amount of hemolysis (heme concentration <25 vs. >25 μM). Subsequently, we accounted for the variation in hemolytic rate and this potential interaction by calculating the number of animals needed to determine if there was an interaction between nitrite and hemolysis level (assuming a 1:1 ratio of low- to high-rate hemolysis in animals receiving a water infusion) and included the level of hemolysis in our final data analysis. The necessary additional paired experiments were then performed using the same treatment regimens with a weighted randomization scheme to the following groups: (1) D5W plus nitrite (n = 2), (2) water (n = 5), or (3) water plus nitrite (n = 5). Overall, these experiments used 32 animals.
Sodium nitroprusside challenge.
To determine the vascular responsiveness to exogenous NO in the presence and absence of hemolysis and sodium nitrite, all animals received a 20-min infusion of escalating doses of sodium nitroprusside, a direct NO donor (1, 3, 9, and 27 mcg·kg−1·min−1), at 5-min intervals before the conclusion of the study. These experiments allowed for comparison of a "traditional" NO donor with nitrite to determine if the observed nitrite effects (i.e., Hb-based nitrite reduction) were distinct from a pure NO vasodilatory effect.
Data Collection
Femoral arterial (20-gauge) and external jugular venous (8-French) catheters (Maxxim Medical, Athens, TX) were placed percutaneously under anesthesia using sterile techniques. Mean arterial pressure (MAP) and heart rate (HR) were obtained from the femoral artery catheter tracing. Additionally, a pulmonary artery thermodilution catheter (7-French, Abbott Critical Care, Chicago, IL) was introduced through the external jugular vein catheter to measure cardiac output (CO), mean pulmonary artery pressure (PAM), pulmonary artery occlusion pressure (PAOP), and central venous pressure (CVP). At the end of the first week's fluid control experiments, all catheters were removed and the animals recovered. At the end of the second week's intervention experiments, all animals were euthanized.
Hemodynamic measurements (MAP, CVP, PAM, CO, and PAOP) and laboratory measurements (hematocrit, Hb, serum chemistries, arterial blood gas analysis, spectrophotometric-based quantification of cell-free Hb concentration, and chemiluminescence-based assays of NO consumption and nitrite levels) were obtained at 0-, 1.5-, 3.0-, 4.5-, and 6.0-h time points. Hemodynamic measurements were also obtained at the end of each dose of sodium nitroprusside.
Plasma Nitrite and Hb Assays
Plasma nitrite levels were measured by an I-based chemiluminescent assay as previously described using the NO analyzer (model 280i NO analyzer, Seivers, Boulder, CO) (78). The total plasma Hb concentration (expressed in terms of heme groups, division by four gives the Hb concentration) was measured by visible absorbance spectrophotometry (HP8453 UV-Vis Diode Array Spectrophotometer, Hewlett Packard). The concentration of oxy-Hb and met-Hb were analyzed by deconvoluting the spectrum into components from basis spectra of canine Hb in PBS buffer using a least-square method as previously described, with subtraction of background plasma scattering (32).
In Vitro Mitochondrial Respiration Experiments
Male Sprague-Dawley rats (175–250 g) were used in accordance with the Animal Care and Use Committee of the National Heart Lung Blood Institute. Liver mitochondria were isolated by differential centrifugation in buffer consisting of sucrose (250 mM), Tris (10 mM), and EGTA (1 mM) as previously described (70). Mitochondrial respiration was measured by suspending isolated mitochondria (2 mg/ml) in respiration buffer (120 mM KCl, 25 mM sucrose, 10 mM HEPES, 1 mM EGTA, 1 mM KH2PO4, and 5 mM MgCl2) in a stirred sealed chamber fit with a Clark-type oxygen electrode (Instech) connected to a data recording device (DATAQ Systems). Mitochondria were supplemented with succinate (15 mM) and ADP (1 mM) to stimulate respiration. In experiments testing the effects of nitrite and Hb, sodium nitrite and human purified oxy-Hb (33) were incubated with mitochondria at the beginning of the experiment (70). In this experimental system, the rate of oxygen generation from the added Hb is less than the rate of oxygen consumption by the mitochondria so that the oxygen increase in the system after Hb addition (20 μM) is not detected by the oxygen electrode and not observed in the raw trace unless high concentrations of Hb are added, in which case a transient increase in the oxygen level may be detected. Note that in this system the chamber is opened to air and oxygen is diffusing into the system as well, but the rate of oxygen diffusion into the system is less than the rate of oxygen consumption by the mitochondria. Only after mitochondrial inhibition do the oxygen levels rise to detection by the electrode.
Statistical Analysis
Data were analyzed using ANOVA, with main effects for study (baseline and intervention), hemolysis (0 μM D5W, <25 μM heme, and >25 μM heme), nitrite, time, and animal (52). Two- and three-way interactions were included in the model. Analysis of responses to sodium nitroprusside were performed using ANOVA on percent changes in hemodynamic variables with increasing doses in the intervention experiments with main effects for hemolysis, nitrite, nitroprusside dose, and animal. Two-way interactions were included in the model. All values are depicted in the figures as means ± SE, and all Hb concentrations are expressed in terms of heme groups.
Physiological Effects of Intravenous Sodium Nitrite
While many groups have now confirmed that sodium nitrite is a potent vasodilator in vivo, to our knowledge no group has characterized more specifically its activity in vivo as a relative arterial versus venous vasodilator or its effects on inotropy and chronotropy. An intravenous infusion of sodium nitrite (27.5 mg/h) rapidly increased plasma nitrite levels to a steady-state concentration (range: 15–21 μM) that was maintained throughout the duration of the 6-h infusion (Fig. 1A). In animals receiving a D5W infusion, sodium nitrite increased the cardiac index (CI; P = 0.001) and decreased the systemic vascular resistance index (SVRI; P = 0.04), pulmonary vascular resistance index (PVRI; P = 0.001), MAP (P = 0.08), PAM (P = 0.09), CVP (P = 0.01), and PAOP (P = 0.65) compared with placebo (normal saline) (Fig. 1, B–H). These physiological effects suggest that low-dose sodium nitrite is a more potent arterial vasodilator than a venodilator and that nitrite increases cardiac performance by direct afterload reduction. Supporting evidence for the vasodilatory effects of nitrite can be derived by examining the components of CI in the log scale (Fig. 2) (68). In this format, the individual contributions of each component of CI are additive (normal scale: CI = SVI × HR, log scale: log CI = log SVI + log HR, where SVI is the stroke volume index). This transformation demonstrates that the nitrite-induced increase in CI is mediated predominantly through a sustained increase in the SVI and to a lesser extent by a chronotropic effect. This transformation also accounts for the rise in PAOP during the last 3 h of the experiment (Fig. 1H). The decrease in HR over time (an effect of anesthesia/analgesia seen in all groups in this model) (Fig. 2B) increases diastolic filling time in the ventricles, leading to higher end-diastolic volumes and pressures that translate into increases in PAOP and further increases in SVI. These data imply that nitrite enhances cardiac performance by afterload reduction through an arterial vasodilatory mechanism. These data also indicate that isolated measures of MAP in animal studies may fail to sensitively assess the magnitude of nitrite-dependent vasodilation because of the rise in the CI.
Physiological Effects of Sodium Nitrite During Intravascular Hemolysis
In previous intravascular hemolysis experiments (52) and in the current experiments, cell-free plasma Hb increased systemic arterial pressure, PAM, SVRI, PVRI, and PAOP (Table 1; P = 0.04, 0.14, 0.06, 0.42, and 0.21, respectively, for the interaction of hemolysis levels and the mean change in each physiological variable during baseline and intervention experiments). If nitrite functioned purely as an NO donor medication, one would expect the vasoconstrictive effects of intravascular hemolysis to attenuate the vasodilatory effects of nitrite because any NO generated from nitrite would be readily scavenged by cell-free plasma Hb. However, in these experiments, the physiological effects of nitrite were not simply inhibited by increasing levels of hemolysis. In fact, the effect of nitrite was dependent on the level of intravascular hemolysis in an unusual way (Fig. 3). We detected a consistent U-shaped relationship between the physiological effects of nitrite and the levels of cell-free plasma Hb suggesting an interaction between the effects of nitrite and the amount of intravascular hemolysis (Fig. 3; P = 0.01 for a differing effect of nitrite at low-level hemolysis compared with 0 and high-level hemolysis across the 7 physiological variables combined). At low levels of hemolysis (Hb < 25 μM), the vasodilatory effects of nitrite were apparently potentiated, whereas with higher levels of hemolysis (Hb > 25 μM), the expected inhibition of the vasodilatory effects of nitrite were observed. These results suggest that there are two reactions that regulate the availability of NO at the smooth muscle: the reaction of Hb with NO and an opposing reaction of nitrite with deoxy-Hb that generates NO. Our results demonstrate that at low levels of Hb, the physiologic effects of the latter reaction are detected; however, with increasing Hb concentrations, the former reaction dominates. These effects are examined more closely and compared with the NO donor sodium nitroprusside in additional experiments described below.
Nitrite Levels and Hb Species Formed During Intravascular Hemolysis
In animals receiving nitrite, plasma nitrite levels were similar and were maintained within a range of 16–20 μM throughout the 6-h experiment (Figs. 1A and 4). Intravascular hemolysis occurred at varying rates (Fig. 4). Animals receiving D5W and nitrite represent the zero-hemolysis control group with all measured cell-free plasma Hb levels < 5 μM (data not shown). In animals receiving water and nitrite infusions with low levels of hemolysis (Hb < 25 μM), the average peak cell-free plasma Hb level was 20 μM. In animals receiving water and nitrite infusions with high levels of hemolysis, the average peak cell-free plasma Hb level was 142 μM. In animals receiving D5W and nitrite (zero hemolysis), 81% of the measured cell-free plasma Hb was oxy-Hb (Fig. 4, values depicted as a dark gray reference line in B and E), consistent with observations in normal volunteers and sickle cell patients showing that plasma Hb is maintained largely in the reduced or ferrous-oxygen bound state (HbFe2+-O2). In hemolyzing animals, oxy-Hb accounted for 71% and 69% of the measured cell-free plasma Hb in animals with low and high levels of hemolysis, respectively (Fig. 4).
During nitrite infusions at low levels of hemolysis, nitrite reacted with Hb to form ∼30% met-Hb (Fig. 4B, values for the D5W + nitrite zero-hemolysis control are depicted as a light gray reference line in B and E). This reaction likely reflects two reactions of nitrite: the reaction of nitrite with oxy-Hb to form met-Hb and nitrate (NO2− + HbO2 → met-Hb + NO3−) and the reaction of nitrite with deoxy-Hb to form met-Hb and NO (as described in the Introduction, Eqs. 2 and 3). The former reaction decreases NO scavenging, and the latter reaction contributes to NO generation. Note that, paradoxically, there is sufficient oxy-Hb at the end of 6 h to almost completely scavenge and inhibit any NO that might form, yet the nitrite remains vasoactive and potentiated by low levels of Hb (Fig. 3). As shown in Fig. 4, B and E, with increasing Hb concentrations, the rate of met-Hb formation increases from 0 to 3 h (P = 0.0001), producing higher levels of met-Hb from 3 to 6 h (P = 0.0001) in animals with higher levels of hemolysis compared with animals with lower levels of hemolysis. This is because the overall reactions of nitrite and Hb are second order during their lag phases, meaning that as the Hb concentration increases, the rate of the reactions increases. Again, we see that the amount of oxy-Hb at the end of the reaction is sufficient to almost completely scavenge any NO that might be formed if nitrite acted as a pure NO donor.
Effects of Nitrite and Level of Hemolysis on Hemodynamic Responses to Sodium Nitroprusside, an Infused NO Donor
Sodium nitroprusside was administered to all animals to determine the physiological effects of a direct NO-donating agent in the setting of hemolysis with and without sodium nitrite. The physiological effects of sodium nitroprusside were dependent on the level (or dose) of hemolysis and the presence of nitrite. As expected, in animals that did not receive nitrite, sodium nitroprusside-induced increases in CI and decreases in SVRI and PVRI were progressively inhibited by increasing levels of hemolysis, suggesting progressive consumption of the donated NO by increasing levels of cell-free plasma Hb during intravascular hemolysis (Fig. 5). In contrast, the effects of sodium nitroprusside at the three levels of hemolysis were different in animals receiving nitrite compared with animals not receiving nitrite (Fig. 5). Compared with nonhemolyzing animals not receiving nitrite (zero hemolysis, no nitrite), nonhemolyzing animals receiving nitrite (zero hemolysis, nitrite) demonstrated blunted effects of sodium nitroprusside on CI, SVRI, and PVRI, suggesting a decreased vasodilator effect of donated NO in the presence of nitrite without hemolysis. If the effect of nitrite on the response to sodium nitroprusside during hemolysis was additive (i.e., the same effect at all levels of hemolysis), then the demonstrated relationship should be a similar linear relationship to the one demonstrated in animals not receiving nitrite, but starting at a smaller magnitude percent change due to the decreased vasodilator effect of donated NO from nitroprusside in the presence of nitrite (comparing zero hemolysis, no nitrite with zero hemolysis + nitrite). However, in animals receiving nitrite, the effects of sodium nitroprusside on CI, SVRI, and PVRI were accentuated with low levels of hemolysis (Hb < 25 μM, nitrite) and then attenuated with high levels of hemolysis (Hb > 25 μM, nitrite) compared with nonhemolyzing animals (zero hemolysis, nitrite) (Fig. 5; P = 0.09, 0.05, and 0.009 for the interaction, demonstrating a different relationship between level of hemolysis and nitrite on the effect of sodium nitroprusside for CI, SVRI, and PVRI respectively). Animals with low-level hemolysis demonstrated a similar or greater percent change on the physiological variables than zero hemolysis (instead of the expected smaller effect in an additive model) and animals with higher-level hemolysis demonstrated blunted physiological responses. This interaction is consistent with the U-shaped physiological effects of nitrite demonstrated during the 6-h hemolysis experiment; compared with the effect in animals with zero hemolysis, the physiological effect of nitrite was accentuated with low-level hemolysis and then attenuated at higher levels of hemolysis. This interaction may be explained by the additional nitrite reduction reaction with Hb contributing to vasodilation. Nitrite reacts with oxy- and deoxy-Hb to form met-Hb and met-Hb + NO, respectively (10). These nitrite reactions may lead to enhanced vasodilation by sodium nitroprusside in the setting of low levels of hemolysis by (1) minimizing the amount of oxy-Hb available in the plasma to consume the donated NO from sodium nitroprusside and (2) directly causing vasodilation secondary to the NO generated by the reaction of nitrite with deoxy-Hb. At higher levels of intravascular hemolysis, the nitrite reduction reaction with Hb may be overwhelmed by the large amounts of cell-free plasma Hb that consume any NO formed from the reaction. Consequently, the donated NO from sodium nitroprusside and the generated NO from the reaction of nitrite with deoxy-Hb are consumed by the excess oxy-Hb in the plasma.
Confirmatory In Vitro Mitochondrial Respiration Experiments
In vitro mitochondrial respiration experiments were performed with nitrite and cell-free Hb levels similar to those obtained in vivo to confirm that the effects of nitrite on vasoactivity during hemolysis are dependent on the reaction of nitrite and deoxy-Hb to generate NO. In these experiments, mitochondria serve as NO sensors because NO avidly binds to cytochrome c oxidase to inhibit respiration. In this experimental system, mitochondria suspended in a closed chamber respire until the chamber becomes anoxic (oxygen trace reads zero). Removal of the chamber lid allows oxygen diffusion into the chamber; however, the trace remains at zero due to rapid oxygen consumption by the respiring mitochondria. The oxygen trace deviates from zero only once the mitochondria stop respiring due to the exhaustion of substrate or inhibition (Fig. 6A). Time to inhibition (oxygen reaccumulation) should be dependent on the rate of NO production from reactions of nitrite with deoxy-Hb and the rate of NO consumption by excess oxy-Hb. With the addition of nitrite (18 μM) and low levels of Hb (10–20 μM), mitochondrial respiration was inhibited compared with mitochondria with nitrite or Hb alone. The shortest time to inhibition was observed with nitrite and 20 μM Hb, above which increasing concentrations of Hb resulted in longer times to inhibition (Fig. 6B). These mitochondrial inhibition experiments demonstrated a U-shaped relationship between nitrite and Hb levels consistent with the results of our in vivo experiments. Our animal experiments suggest an interaction between the effects of nitrite and the level of hemolysis such that low levels of hemolysis accentuate the vasodilatory effects of nitrite. These mitochondrial experiments demonstrate that NO generation and accumulation from nitrite reduction by Hb is maximal at low levels of hemolysis and decreases with higher levels of hemolysis. These results suggest that the in vivo accentuated vasodilatory effects of nitrite during low levels of hemolysis may be mediated by the generation of NO from the reduction of nitrite by Hb.
Under physiological conditions, our experiments demonstrate that low-dose sodium nitrite is a potent arterial vasodilator that increases cardiac performance by direct afterload reduction with mild chronotropic effects. During hemolysis, we detected a consistent U-shaped relationship between the effects of nitrite and cell-free plasma Hb levels across three experimental settings, suggesting an interaction between nitrite and the level of intravascular Hb.
Nitrite reacts with oxy-Hb and deoxy-Hb to form met-Hb and met-Hb + NO, respectively (5, 11, 16, 31, 33). During low-level hemolysis, these reactions will minimize the amount of oxy-Hb available in the plasma that can consume NO (via the dioxygenation reaction) and generate NO by the reaction of nitrite with deoxy-Hb. The net result is accentuated vasodilation compared with no hemolysis. At higher levels of intravascular hemolysis, the large amounts of cell-free plasma Hb overwhelms the nitrite reductase reaction of Hb and consumes both the NO formed by nitrite reduction with Hb and endothelium-derived NO. The net result is vasoconstriction compared with low-level hemolysis and no hemolysis. During a sodium nitroprusside infusion with low-level hemolysis and nitrite, the nitrite-oxy-Hb reaction minimizes the oxy-Hb concentration and allows the donated NO from nitroprusside to cause vasodilation. This vasodilation is further accentuated by the production of additional NO from reactions of nitrite with deoxy-Hb. At higher levels of hemolysis, the vasodilatory effects of sodium nitroprusside are attenuated by the high levels of oxy-Hb, which consume both the NO donated from nitroprusside and the NO generated from the reaction of nitrite with deoxy-Hb. During the mitochondrial experiments, maximal NO production and accumulation occurred with nitrite and low levels of Hb because the excess heme groups at higher levels of Hb consumed the NO generated by the nitrite-deoxy-Hb reaction, leading to decreased NO accumulation.
Traditional NO donors, such as sodium nitroprusside, produce dose-dependent vasodilation that is inhibited by cell-free plasma Hb (52). In contrast, nitrite led to accentuated vasodilation during low-level hemolysis despite the presence of oxy-Hb levels sufficient to scavenge any NO that might be formed if nitrite acted as pure NO donor. The vasodilatory effect of nitrite clearly differs from traditional NO donors in the presence of Hb and can in part be explained by the nitrite reductase activity of Hb (11, 31, 33). Generation of NO from nitrite and Hb requires both hypoxia and an acidic environment, which are present in hypoxic tissues. This allows for maximal NO generation by the deoxyheme-nitrite allosteric reaction as Hb deoxygenates within the circulation.
This study provides in vivo evidence supporting the theory that Hb possesses a functional nitrite reductase activity. The notable interaction between nitrite and Hb in this study was markedly different from the behavior of a traditional NO donor, sodium nitroprusside. While the latter was inhibited in a dose-dependent manner, the former was potentiated by Hb at concentrations that produced NO in our in vitro mitochondrial NO sensor experiments. In another study with the Kim-Shapiro laboratory (3), we examined how NO could be generated from the nitrite reductase activity of Hb and then be able to escape heme autocapture (via Eq. 1 or an analogously fast reaction with deoxy-Hb). We found that nitrite can also bind to met-Hb to form a nitrite-met-Hb intermediate that possesses nitrogen-dioxide-ferrous Hb character. The NO that forms from nitrite reduction can react rapidly in a radical-radical reaction with nitrogen dioxide to form N2O3 (3). N2O3 is an uncharged, highly lipophilic, and diffusible molecule that is more stable than authentic NO. N2O3 can nitrosate thiols to form vasodilatory S-nitrosothiols, can homolyze back into NO, or can regenerate nitrite. The apparent inhibition of the nitrite effect at higher Hb concentrations in these experiments indicates that at least part of the mechanism must involve the regeneration of NO, which can be in part scavenged by excess Hb.
The levels of cell-free plasma Hb in the low hemolysis group of this study are consistent with the levels observed during sickle cell vasoocclusive crisis (40, 58, 65) and during other clinically relevant human hemolytic conditions such as cardiopulmonary bypass, malarial infection, hemolytic-uremic syndrome/thrombotic thrombocytopenic purpura, paroxysmal nocturnal hemoglobinuria, alloimmune hemolytic anemia, and rhabdomyolysis (myoglobin) (12, 40, 55, 58, 63, 65, 69). All of these conditions have now been associated with progressive vasculopathy and pulmonary hypertension and are associated with systemic NO scavenging by plasma Hb (22, 52, 67). Furthermore, hemolysis is associated with platelet activation and inhibition of NO-cGMP signaling in platelets (75). These represent processes in which the allosteric nitrite reductase activity of Hb may make nitrite an ideal therapeutic agent to attenuate the effects of accelerated NO scavenging by cell-free Hb released during intravascular hemolysis (1, 17, 19, 41, 52, 59, 65). In these clinical scenarios, the administration of low-dose nitrite will have minimal physiological effects in normal tissues. However, in tissues that have become hypoxic secondary to vasoconstriction from accelerated NO scavenging by cell-free Hb, low-dose nitrite may cause vasodilation by (1) reacting with oxy-Hb to form met-Hb, thereby preventing NO scavenging; and (2) reacting with deoxy-Hb to generate NO and met-Hb. The net effect would be hypoxic vasodilation in local tissues that have become ischemic from the vasoconstrictive effects of accumulating cell-free plasma Hb from ongoing low-level intravascular hemolysis.
In addition to low-level intravascular hemolysis, nitrite may also have a therapeutic role in minimizing the vascular toxicities of more severe episodes of intravascular hemolysis (cell-free plasma Hb levels > 50 μM) and the administration of several types of cell-free Hb-based blood substitutes (cell-free plasma Hb levels > 600 μM) (14, 15, 28, 29, 77). In these clinical scenarios, the ability of nitrite to attenuate the physiological effects of cell-free plasma Hb will be overwhelmed by the accelerated NO consumption caused by the large amounts of plasma Hb. However, in these scenarios, the affected tissues and organs will subsequently develop areas of hypoxia and acidosis. Within these areas, there will be accelerated reduction of nitrite by deoxygenated cell-free plasma Hb leading to local NO generation and vasodilation. Therapeutic strategies to deliver Hb-based blood substitutes will either require increasing the molar ratio of nitrite to Hb or will require modulating the reaction kinetics by increasing the concentration of nitrite bound to met-Hb (to facilitate the formation of N2O3) (3), decreasing the Hb oxygen affinity (so that there is more deoxyheme to reduce nitrite), or decreasing the redox potential of the heme-based blood substitute (to increase the reactivity with nitrite). Therefore, despite not reversing global physiological changes during high-level canine hemolysis, nitrite may be able to limit organ damage and dysfunction during severe hemolytic episodes and during the administration of Hb-based blood substitutes. In these situations, the optimal dose of nitrite with Hb for resuscitation that maximizes NO generation and minimizes Hb oxidation (which would limit oxygen transport of Hb) has yet to be determined.
Nitrite is a unique vasodilator. Under normal physiological conditions, low-dose nitrite is potent arterial vasodilator that increases cardiac performance by direct afterload reduction. During hemolysis, the allosteric nitrite reductase activity of Hb produces an accentuated vasodilatory response to nitrite during low levels of hemolysis that is overwhelmed during high levels of hemolysis. These unique properties of nitrite and Hb may allow nitrite to be used as therapy during episodes of acute intravascular hemolysis or potentially to detoxify cell-free Hb-based blood substitutes.
This work was funded by the Intramural Research Division of the National Heart, Lung, and Blood Institute (NHLBI) and the Critical Care Medicine Department, Clinical Center, of the National Institutes of Health (NIH) and by extramural NHLBI Grant K22-HL-089041-01. Further support was provided from the Office of Rare Diseases and the Clinical Center of the NIH via a bench to bedside grant.
M. T. Gladwin is named as a coinventor on an NIH government provisional patent application for the use of nitrite salts in the treatment of cardiovascular diseases.
↵* P. C. Minneci and K. J. Deans contributed equally to this work. S. B. Solomon and M. T. Gladwin also contributed equally to this work.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
- Copyright © 2008 by the American Physiological Society


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James Gunn Wants You to Know He's Worked Real Real Hard on 'Guardians Vol. 2'
Dangerous business, making trailers these days. When a beloved franchise unveils the first look at its latest installment, they can be sure that the devoted fanbase will go through each and every frame with a fine-toothed comb, on the hunt for any context-free images that may hint at a new character, location, or plot development. In the worst cases, a trailer becomes a maddening game of keep away between the editor cutting it and the fans itching to strip-mine it for spoilers. James Gunn, as he's readied the first trailer for his upcoming Guardians of the Galaxy sequel, has had to learn this the hard way.


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Nordic skier injured, taken away by stretcher
KRASNAYA POLYANA, Russia
A scary moment for a Japanese Nordic skier at the Olympic Winter Games in Sochi.
Taihei Kato of Japan broke his left arm Tuesday when he fell hard while landing during the ski jumping component of the Nordic combined large hill event.
Kato had to be taken off the mountain by stretcher.
In steady rain, Kato landed awkwardly, lost his left ski and then fell hard on to his left elbow. He was conscious and appeared to be in pain as he was helped away by medical officials.
Last week, Russian ski jumper Mikhail Maksimochkin fractured two ribs and spent two days in the hospital after a fall on the large hill.


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