"There are just so many ladies," I said to myself whilst criss-crossing the expo at the ESPN Wide World of Sports Complex. Only being there one month earlier for the Dopey Challenge, the vendors' tables were now littered with tiaras, sparkly skirts and shoe jewelry as far as the eye could see.
I was completely out of my element.
How did I get here? It stems back to my wife and her best friend, Sally. On one of their many phone calls together, Sally mentioned how'd she'd love to run a half marathon. My wife, no stranger to half marathons or distances agreed to run one with her, and they both decided the 2016 Princes Half would be high noon. Not to skip out on a Disney race, I happily obliged.
While running every other runDisney event, the Princess had always been off my radar as I had heard horror stories of past events being more of a massive bachelorette party and less of a race. But, a race is a race and a trip to Disney is a trip to Disney - no matter how much pink I'd have to endure. Luckily, Sally's husband, Kirby, had made the trip, doubling the testosterone contingent.
This weekend was a quick turnaround event for me as we have just settled into our new city, Seattle and I didn't want to be away from work too long. By the way, Seattle to Orlando for a weekend trip should only be reserved for the most passionate sadist; it was brutal. Given the short time at WDW, we all decided to stay at the moderate resort, Port Orleans Riverside, a departure from our usual weeklong stays at Boardwalk and Wilderness Lodge.
We had requested adjoining rooms, but at check-in we were told we had reserved a Royal Suite. Now, a Royal Suite is a room that has been slathered with all kinds of Disney magic and is reserved for those true Disneyphiles who just need to have every inch of vacation cloaked in Disney - certainly not our thing. After explaining we didn't reserve a Royal room, we were told we were upgraded, something that happens fairly regularly with us, but we declined for a room in the same block as Sally and Kirby.
Port Orleans, in my opinion, is one of the better moderate resorts. It's tucked away from all the crazy and has a lot to offer in terms in eating and entertainment. The only downside is the transportation, as they make guests wait sometimes for 45 minutes for a bus, and then the bus will stop at every Riverside stop and its sister resort, French Quarter.
After hitting the expo, we all clowned around in the Magic Kingdom for awhile, catching one more scrumptious meal at the Skipper Canteen, which is becoming one of my favorite restaurants. We called it an early evening since the following day would be the 10k. My wife, Devin would be running the Glass Slipper Challenge, which included the 10k. Kirby was also running the 10k, so we hit the hay early in preparation for the following morning's race.
Acting my age is not one of my strong suits.
Saturday morning, we hit the race early to get Devin and Kirby ready for their 10k. Sally and I played the race supporter role, something new for me as I'm usually the one racing. I give you spectators a ton of credit, all that standing around is mind-numbing. Happily, we got to see the two of them start the race and again at the finish line. Both were in good spirits, Devin was now inching closer to finishing her 19.3 weekend journey.
The morning was especially sweet for Devin and I as we were able to catch some quality time with one of the greatest people on the planet, Jillian, who always makes my heart sing.
Later that day, we headed over to Disney Springs to catch up with some great people at the #WeRunSocial meetup. If you've read my past posts, you're probably familiar with #WRS. If not, we're just a bunch of runners who like to get together and hang out. That's it! It's always a positive vibe and it's a great opportunity to visit with friends and the folks who follow me on social media. As per usual, the whole gang was there: Gelcys, Dani, Christina, Sarah, among others. And, I finally had the pleasure of connecting with a longtime social media friend, Lauren, who is as baller in real life as she is on the socials. Great seeing you lovelies!
Because we do this every time (Dani & I).
Sunday morning brought my race day, the half marathon. Similar to WDW Marathon Weekend, the half would start outside of Epcot, head towards and through the Magic Kingdom and then back. Now, I'm not one to ever run in a costume. It's not that I don't like them, rather as a running purist, my thoughts on racing is just that, a race and not a parade. However, playing to the theme of the event, I dolled up in as much pink as I could stand: pink Pro Compression high calf socks, pink arm sleeves, a hot pink Dirtbag Runners trucker's cap and pink(ish) New Balance 1400's. This would have to do, ladies.
I felt like a bottle of Pepto bismol.
While my bib number was pretty low, Corral B was the earliest corral male runners could start - while male runners could participate, this was a female race, meaning no winners for the fellas. Looking around me, I deemed Corral B the "Sausage Corral" due to the sheer amount of dudes in the paddock.
Time to let the princesses out.
Shooting past the start line, I noticed the pace of this race was slower than Wine & Dine and January's half which pacing from the onset a challenge. I tried to run my normal race, but felt like I was going out too fast given the runners I was passing. Once on World Drive, it was easier to settle into a groove, but the damage was already done and I knew a PR would be out of the question. So, for the rest of the race I dug in, but held back and enjoyed the scenic pieces of the course and thought about the 19,000 princesses chasing me.
Also, for the first time ever in a race, I experienced what we runners call "digestive distress." From Mile 1 to about Mile 5, I experienced a hot, sharp stabbing in my intestines. During those miles all I wanted to do was finish as fast as possible. Luckily, the "running" gods and not the "runs" gods were on my side and the feeling subsided around Mile 6.
As I've stated numerous times before, running through the Magic Kingdom is such an awesome feeling, with the castle and all the spectators cheering us runners on. It's indescribable, really and one really has to experience it to understand it. Certainly, it's worth the long stretch of boring to get to it and after the mind-numbing stretch after it.
For all the wondrousness the Magic Kingdom provides, the final three miles of the race are ball busters. Starting at Mile 10, there is a mile-long curving, uphill monster that chews runners up. Miles 11 and 12 spit us out with some rather ugly hills - well, overpasses actually. Finally, 12.5 puts us back in Epcot and onto greatness. Like the races in January, a gospel choir greeted me just before the finish line, giving me a little more gas to motor through the finish in a lackluster 1:51:00.
I caught up with Kirby and then watched as the girls victoriously rolled in. Devin with her Glass Slipper 19.3 Challenge in the bag, Sally pocketing her first ever half marathon. That night we feasted like royalty at Boatwright's. The following day was reserved for celebrating the best way possible, drinking around the world at Epcot because, adulting.
Hearing horror stories prior to arriving at the weekend, my fears went unfounded. Overall, the experience was a lot of fun and something I'd do again. Truth be told, because I was in a front corral, my experience was probably pretty different than say, those in mid/late corrals. I had a ton of room, and when coming back on World Drive towards Epcot, the runners yet to hit the Magic Kingdom must have numbered in the thousands. However, I really enjoyed myself. Dev ad I had the opportunity to spend time with some of our best friends in a place we love to visit. That made me feel like a king for a weekend.
Until next time. Run fast. Run smart.


Can R510 cause interference with wireless subwoofer?
Wireless subwoofer dropping signal when R510 is nearby. Seems to be causing some interference, wondering if channelfly or background scanning would make a difference.
I think it's all related to which channel the AP is on, and what freq the wireless subw is using.


text:
The Gateway: Run the Press
Home Editor's Pick "Eminent Domain" hits close to home
"Eminent Domain" hits close to home
photo courtesy Omaha Community Playhouse
Cassie Wade
Family conflict, religion and small town Nebraska values dominant the stage at the world premiere of "Eminent Domain" performed at the Omaha Community Playhouse.
Written by Omaha native Laura Leininger-Campbell and set in Nance County, Nebraska, "Eminent Domain" tells the story of the MacLeod family and their battle to protect the family ranch from a Canadian oil pipeline.
"Eminent Domain" is performed by a nine-member cast and directed by Amy Lane. Its production is an official event of the Nebraska 150 Celebration, which is a yearlong celebration marking the 150 years of Nebraska's statehood.
The feeling and culture of Nebraska is well-embodied throughout the play. Everything from the set – an old farmhouse complete with a creaking porch – to the way the actors are dressed in jeans, flannel shirts and work boots is representative of the state.
References to the MacLeod's Scottish heritage highlights Nebraskans' tendency to hold on tightly to their ancestry. The sword holstered on the farmhouse's porch and actor's kilts are nice touches.
However, the play relies too heavily on Nebraska stereotypes at times. While Nebraska is a red state, having ranch owner Rob MacLeod, played by Bill Hutson, referring to Californians as communists is over the top.
Additionally, the characters' long-winded speeches about the importance of family may have been designed to reflect Nebraska's family-centric values but quickly becomes corny and unnecessary.
Issues relevant to Nebraskan families are tackled, though. For example, Adair MacLeod, played by Erika Hall Seiff, faces backlash for leaving the family farm behind to become a lawyer while her brother, Bart, played by Jeremey Estill, gives up his dreams to stay and help out.
The resulting conflict between the siblings and additional inter-family conflict makes the MacLeod family relatable and lovable. Be prepared to sit through several intense screaming matches during the show.
Audience members are warned the play contains adult language. Several "f-bombs" and many other curse words are frequently dropped during the show.
While the cursing adds to Rob MacLeod's character as a tough, no-nonsense old man, it prevents the show from being kid-friendly.
Besides verbal conflict between the family and the legal battle between the MacLeod's and the oil pipeline, the plot of "Eminent Domain" takes on other conflicts. This includes Theresa MacLeod's infidelity and the consequences that being unfaithful have on her marriage.
Rod Macleod's loss of religion resulting from the death of his wife is another conflict resolved over the course of the play.
All of the conflicts in "Eminent Domain" make the play about more than a Nebraska family fighting an oil pipeline. "Eminent Domain" is also a love story and a story of forgiveness.
The Omaha Community Playhouse's production of "Eminent Domain" is worth seeing, especially for Nebraska natives. The plot is action-packed, timely and relatable.
The show runs from Aug. 25 through Sept. 17. Tickets are available through the Omaha Community Playhouse Box Office, OmahaPlayhouse.com and TicketOmaha.com.
Tickets cost $36. Reduced pricing is available for large groups.
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Discussion in 'Support' started by Zeo, Dec 8, 2013.
Ok i am making this a separate thread because it will get all the info and accounts i have observed since suffering with this strange and in my case seemingly specialist-baffling-condition.
I started to play the game GTA5 and in doing so got a bluetooth in ear headset (just goes over one ear/in one ear) it was in my left ear to begin with. It was super, super quiet this thing even with the volume turned to max, however when it was on max you could hear the sort of 'white noise' hiss that some technology makes when on max volume, though this hiss was not loud in any way.
After playing for a few hours and removing the device the hiss sort of 'stuck' in my ear, but not bad. I repeated this over a week or so and the hiss would get slightly louder, but it was more what its like when you have been on the phone for a few hours to someone, not painful or even noticeable really in most cases.
I then switched the ear i used it on because i was concerned of damage. So i switched to my right ear but only for a few days and then stopped playing altogether. I also went on a training course with work which involved being on a moor which was seriously cold, my ears were painfully cold (as was my nose!) but nothing bad happened,or so it would seem.
I went to my GP who said it was a huge amount of wax, and not to worry at all. I was booked in for syrnging however not for almost another month! so i couldn't bare it after 10 days (in that time i used olive oil as a treatment) it was very bad in my left ear and less severe in my right so both needed doing.
Went to a specialist who used a suction, went very well and cleared 100% of wax out of my ears, told me my ears are healthy and free of wax. The sound went by 70-80%. Until i got home some 2hrs later, when it returned full blast.
However now my ears would get hot, especially the right one, really hot! So i then went to another doc whom said my left ear had a perforated ear drum that had healed however it was a very old injury! (i had previously had that ear syrnged some 3 years ago but with no problems) and my right ear had redness and swelling! In the day its fine, i can cope, in the evening the sound is unbearable. My ear gets hot and then the sound comes. What is also strange is the ringing has shifted from my left ear to my right, same intensity just a different ear, has been this way for 3 days straight now. My left ear feels free of sound, though when i use my headphones to go to sleep with rain sounds etc i can still hear it ringing, just less noticeable than the right one.
As a point of note, stress/anxiety does not seem to effect it, the sound will vary in intensity as does the heat in my right ear (no longer my left) and does not seem to be effected by either hot or cold temperatures.
My fiancé is certain that this is a result of an impacted wisdom tooth (i did have some pain in my mouth in october this year and swelling/sort of infection in my gum from a wisdom tooth coming through on my right hand side of my mouth) but i don't want to cling to hope that may not be there. I also find myself grinding my teeth in my sleep a lot. For this to be the case would be a coincidence that the ringing started when i used the headset then i suddenly get wax AND wisdom tooth pain? lots of coincidence!
Same. Just got it, bang, one day, no idea why. My affected ear is also constantly hot, no idea why. I also hear a crackling bubbling sound when I talk or swallow, no idea why. Been to a million doctors, no idea why. Great. Guess some people are just destined to get it. Which is not the best news you want to hear. Still, everyone seems to say that it gets better with time. I'm still not convinced as mine hasnt gotten any better in 4 months if anything, its worse. Try neuromonics. its expensive but has a good success rate. I dont care how much it is, i'd sell all my possessions and go into debt if i could find something that works!
Neenie, sorry but thats quite not the same and its not what i was asking for. I didn't suddenly get it BANG, like that, it happened gradually over a period of 5 days or so, mine switched ears too AND I had wax.
You're saying you had all the above too? if not then its not the same, I need to know of people that have this as a result of a similar set of circumstances as well a similar or recognised relation to some of the unusual symptoms from normally documented cases.
I get that your hot ear is similar however i feel that for me this is in relation to the redness/soreness from the suctioning, i added that info here because, for me, it is the indication of an 'attack' about to happen (sound gets noticeable/loud when it gets hot) and might explain how its 'switched' ears, perhaps the large volume of wax in the left caused it there and now its gone (seems like there is no screeching at all anymore there) the soreness in the right is causing it in the right? i don't know, but it would be good to find others here that might know.
I am sorry to hear of your tinnitus and i hope you can get relief for it, i know how bad it can truly be. But thank you for the reply.
I am trying to get an appointment with an ent for a hearing test. Harder than you think. I only had wax for 4-5 days max.
I have perfect hearing, seriously, even before the wax, perfect!
I'm curious to know more about your thoughts on this, cause I have also wondered to what extent my hearing tests are good proof of me NOT having T cause of hearing loss. I've had two independent hearing tests that covered the familiar 250-8000hz range with no sign of hearing loss. But why don't you think hearing tests are a good way of excluding or including hearing loss as a main cause of T? Because the loss could be hidden? The audiologist I went to in addition to the two different ENTs said tests over 8000hz is pointless cause we are not able to detect sound on those frequencies, the same person also told me that the test was conclusive, it did not show any sign of loss at all except from the normal loss range due to age. I guess they have an age template table and I was inside the normal range. I did a threshold test that clearly showed signs of severe H however, so I guess the audiologist became more occupied with that.
Almost needless to say none of the people in charge of the tests could even faintly suggest why I have mad loud T levels. When they don't see hearing loss they are totally clueless as usual. It's the only cause of T for most of them, although alot of people have T with no hearing loss they still stick to the same theories. It puzzles me.
another element i wish to add, again this is about UNUSUAL symptoms or less documented issues i want addressing and focus on in this thread.
My tinnitus in the morning is SUPER quiet and over a period of hours increases in volume to the very loudest it can get.
Nobody seems to be saying this happens to them.
This happens every morning, every day, no matter how I've slept or how much stress i have been under. By the evening my ear is hot and the tinnitus is very loud.
Another interesting point: tinnitus is usually balanced in both ears, but mine is not, mine is always in ONE ear at a time (or actually both but very reduced in the opposite ear) this is NOT normal nor is it documented very well here or ANYWHERE.
UPDATE: visited my dentist this morning and has performed an X-ray as well as listening to my symptoms. What showed up on the X-Ray is an Impacted wisdom tooth, he even thought that the saliva gland(s) may have been infected as well.
He believes it is this that is caused the tinnitus (sort of makes sense as a month or two ago i had really bad infections in my mouth, soreness and aching in the extreme which i was told at the time was my wisdom tooth coming through) I am being referred to a facial specialist in hospital and might result in surgery to remove the tooth and fix it. (if not it may sort itself out or i may need antibiotics to perhaps alleviate symptoms) Though still not sure why it started in my left ear, but maybe as a result of the wax in that ear? hmm.
I also asked if removal of the tooth can cause the tinnitus and was told that it cannot or is very very unlikely. It will however take 6 weeks for me to be seen at the earliest!
Can you keep us up to date after the tooth removal please Zeo? Thanks.
I will however i have a further development.
UPDATE 2: went to another doc today after the dentists that said he would be surprised if it turned out to be my tooth! He did however prescribe me some Amitriptyne Tablets which will help me sleep and MAY stop the noise or reduce it.
He also checked the ear that is making the loudest sounds, i.e. my right ear and said there was no swelling or redness now and when he opened my ear the sound got much louder! this puzzled him.
Its a total circus this, i have seen 6 doctors, 2 nurses and a dentist and ALL giving me different diagnosis!
How my symptoms are now: the tinnintus used be a 8-9 (volume wise) during the day, its now a rough 7 or so, when my ear is hot and maybe a 5 or so when its not hot. I wonder if its reducing somehow? the frequency seems to fluctuate a little as well, its not quite as constant a 'tone' anymore?
Just to clarify, theres almost no such thing as perfect hearing. Tests are not that rigid and we all have degrees of degraded hearing compared to when we were kids (if you didn't have a damage back then of course) I think we need better tests cause I'm a little skeptic based on experiences. My tests are all fine, meaning no severe hearing loss. I don't think the test detects slight losses or minor losses, so the audiologist only compares the result to a template table, your either inside or outside the normal levels as far as I know. If there's any audiologists here then please correct me.
In addition to this we have the almighty frequency debate of course. Is 250-8000hz a good enough range to exclude hearing loss etc. But that's a separate debate almost. Point is I'm not sure if the standard hearing tests represents a good enough tool to detect hearing loss.
Tell me about it. The more people you see the greater variety of fluffy explanations and theories you will get, except from this one thing: They don't know what caused it and they don't know how to get rid of it. For your sake I hope the latter isn't the case, but seeing more docs could give you more headaches. I hope it settles.
I tried a tablet just now and after a short while the heat has gone out of my ear and the sound is much quieter!
Just got it, bang, one day, no idea why.
Same here Neenie. There's alot of people sharing the same stories. From no T to mad T overnight. Makes you wonder how and why etc. Not knowing the cause is a second burden.
@Zeo Actually, I have similar tinnitus in my left ear. It hasn't swapped ears, but it is majority on left and a little on the right. The sound about doubles by 9 hours through my day. Check and see if jaw or eye movements change the frequency or loudness of your T. They can be subtle.
Lisel yours is rare then like mine, check your teeth! How long you had it?
@Zeo, only about a week. I've had ringing in my other ear much longer, not sure if it's related. But yes, I knowingly have TMJ issues and hopefully will see some improvement with T along with the TMJ.
Re: impacted wisdom teeth, I had all 4 impactions removed after the onset of my T. Was told by a lazy ENT that it might help the problem - did not make any change with T levels, but I was experiencing a LOT of pain from the wisdom teeth so it definitely helped tremendously in that respect.
Temporomandibular joint. Basically where you lower jaw connects to your skull. TMJ disorder usually stems from having an uneven bite. Grinding your teeth at night is actually a symptom of it. As are tinnitus, toothaches, and headaches as a few other major indicators.
Also, funny timing. I got into the car after a long long night in lab and barely being able to focus on my T. It shifted to my right ear. And as I type this, is shifting more back to the left. So huh! More similar than I thought, even.
Its cause for concern if it shifts ears arrantly, it should always be balanced.
I have always had issues with my teeth, i had too many teeth for my mouth and had to have lots removed, then i didn't brush them so i have lots of fillings, root canals and even a gold tooth. I ALWAYS grind my teeth at night, I'm talking 10 years+ i wake up with numb or sore gums/teeth. Im getting a mouth guard to sort that out.
I would be surprised that all this is not my wisdom tooth as I've had lots of infections/swelling and agony in my right side of my mouth recently and may even be infected as my dentist said.
Is there a cure for Temporomandibular joint issues?
A mouth guard is actually normally what they start you off with to treat TMJ issues. According to the orthodontist I am seeing, it is enough for most people. I'm not sure if the mouth guard you're getting for the teeth grinding is the same type.
I don't think there is a "cure" per say. Depends on the cause, I suppose. But wearing a mouth guard at night is no big deal, I can't complain. They can also do surgery and replace the joint, but the success rate on those is not so good.
Can i just add that my tinnitus sounds like something i couldn't quite figure out, but now i know!! remember those old CRT's Tv's? (old big, fat TV's) you recall when you would turn them off or sometimes when the tube was dying in the tv, the high pitched whine? its exactly that sound but right in my head/ear and super loud!
If the sound doesn't get me then the hotness of my ear does, which in turn reminds me of my sound, I'm still disgusted by the lack of knowledge, understanding and help for those with this condition and I'm not standing for it.
All those things are common with T. The hypersensitivity is called hyperacusis (H).
For a more detailed description of H see http://www.hyperacusis.net/ and read the different subjects.
Cool, is that a permanent thing? again because i won't be able to work at all anymore if it is! My only solution is to have earphones on all day/night every day/night with rain sounds on it full blast. I also experience it worse when i grind my teeth or my jaw hurts….. mainly when lying down.
EDIT: in the causes section of that site it does not mention tinnitus at all.
Do a search on H here on TT and you will find different posts on this subject. H is treatable to a certain level, at least its known to decrease by time, I hope thats true in my case as well but its a long stretch it seems. According to the research I have read about 40% of all people with T also have H, the severity of the condition varies greatly of course.
UPDATE 3: I've been in communication with the specialist ENT that did my suctioning and he is now passing my info on to very prominent tinnitus specialists which is awesome but i have also noted something interesting.
"I have noticed that i catch myself in moments of being anxious or tension (i.e. most of everyday life) clenching my teeth together or grinding, i do this subconsciously. When i wake, perhaps over the last 10 years or more i have numb teeth/gums/jaw and often headaches too (i thought the headaches may have been as result of my neck/nerves) looking back though they could be tension headaches.
The last few months have been quite anxious ones but was slowly calming down. Today i tried a little test out which was to see if it was a TMJ/TMD related thing and the tinnitus in now in my left ear with my jaw on that side aching a lot more than the right.
I have had a sore jaw all day today in fact and feel my jaw is somehow not right? When i grind now to test, the sound is somewhat louder too (on left side) i could be imagining all this however if it was dental related it would not surprise me, i have a long history of teeth related issues; having too many teeth in my mouth, root canal, gold tooth, infection/swelling recently from the wisdom tooth pushing the other teeth, neuralgia, lots of fillings and much more."
As i type this now i notice my jaw aching again just being where it is and in doing so the sound increases! when i grind on one particular side of my jaw, the sound increases in that ear, but gradually returns to a certain level.
The ENT is pretty sure (from his email) that its either my wisdom teeth and/or jaw related physical issues relating to TMJ/TMD.
Its always quiet in the morning then gets louder…. ENT recommends i use a hearing aid with a special frequency to stop the noise but will be very costly.


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Tag: kenya
African Broadband
[As ever, this piece is on the BBC News website, where they called it 'How Broadband is Changing Africa', though I think the real message is that a broadband Africa will change the Internet…]
Norman Borlaug, whose work in Mexico and India led to the 'green revolution' in agricultural production, died last week and was widely commemorated for his important work.
While the introduction of new crops and the use of irrigation, fertilisers and pesticides certainly enabled us to feed millions of people, the crops were delivered at a price, and we should not forget it and Borlaug's green revolution, like every revolution, had a negative as well as a positive side.
New farming practices create dependencies on machinery and chemicals, and the the patents that protect genetically modified crops limit the ability of farmers to do things like retain seed from one harvest to plant next year, forcing them instead to buy anew each year.
I was reminded of the debate over the green revolution this week as I stood outside my hotel in Nairobi, Kenya, and watched a work gang lay fibre optic cable in a trench they were laboriously digging with pickaxes on the other side of the busy road.
Continue reading "African Broadband"
Making Digital Planet in Nairobi
We've got another couple of days in Kenya, with some interviews lined up for future episodes of Digital Planet, but we finished off next week's Connected Africa show last night.
Rory Cellan Jones is in Mombasa, on the coast, preparing his own reports, but I've been in Nairobi.
It's been a remarkable few days, with some great people and even the odd giraffe
At the Giraffe Centre
A nice hotel
Nairobi Safari Club
And hard work for Gareth, writing the script
Gareth at work
In Kenya for Digital Planet
One of the best things about working on Digital Planet is that I get the opportunity to visit places that I wouldn't normally get to, and when I'm there I get to meet a lot of interesting people and talk about their work and their lives.
Yesterday I arrived in Nairobi, Kenya, to meet up with Gareth and Michelle, and after a trip to the BBC bureau we spent some time at KBC, the main Kenyan broadcaster.
But the afternoon was spent at Kibera, the slum in southern Nairobi. It's the largest slum in Africa, with a population of over one million, and being there was astonishing.
I don't have time now to write about it, or what I thought as we wandered through the space where a million people carve out a life. But I will.


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That's Amore Top Public Vote
Following on from the success of The Scottish Italian Awards, The Italian Awards team have now reached out to the English Italian community to compete in what will be the toughest contest England has seen in years.
The public were invited to vote for their favourite Italian venues/businesses across England and BedfordBID is very proud to announce that one of our very own restaurants, That's Amore, has made the top 10 in the Best Pizza category and Best family run business. Rosanna, owner, now faces her next challenge to be judged by an independent panel including the famous Aldo Zilli.
All winners will be revealed at the Gala Final on 18th September.
www.thats-amore.org.uk


Showing results by author "Graham Higgins"
Many people look at survival medicine as stocking up on some helpful over-the-counter medicines and coverings for wounds for their survival kit. However, survival medicine entails much more. To be adequately prepared, you must know what to do once your supplies run out. Survival medicine teaches you about common ailments, good over-the-counter medications to stock up on, and how to use your surroundings to make medicine in an emergency, by relying on herbs, berries, and other plants.
When the next natural disaster or the apocalypse comes around, those who are prepared are the ones who will survive. Look around your pantry right now- do you even have enough non-perishable foods stored to feed your family for a week? Unless you have been practicing other food preservation methods or have stocked up at the store, chances are you do not.


text:
var Class = require('../../class')

module.exports = Class.create({
  isAnimal: true
})

redpajama_set_name: RedPajamaGithub


text:
Desperate final moments to save Australian model Sinead McNamara on luxury superyacht
Sinead McNamara was working on billionaire Alberto Bailleres' $200 million superyacht. Photo / via Instagram
By: Andrew Koubaridis
The man who found popular Australian Instagram model Sinead McNamara on the luxury superyacht has revealed how he desperately tried to save her life.
The captain of a neighbouring boat made the horror discovery on the palatial Mayan Queen IV, where McNamara was working off the Greek island of Kefalonia.
According to a friend of the captain, the skipper was returning to his boat about 1.45am on Friday after a night in one of the many bars that surround Argostoli harbour when he saw the young woman.
Port authorities grounded the Mayan Queen IV and its crew to take witness statements and conduct forensic tests. Photo / Supplied
The friend said the captain shouted "What are you doing?" and when she didn't answer, he began frantically calling for help.
Port police officers and the Mayan Queen's own security were then alerted.
The captain has made a statement to the Argostoli Port Authority, which is investigating McNamara's sudden death.
There is no suggestion of foul play and while authorities have not confirmed her cause of death it is believed the young woman took her own life.
The captian said tragically a routine police patrol had just passed the superyacht and not seen the young woman in peril.
The captain described a frantic scramble by police and medics to save McNamara until, after about 30 minutes, they made the decision to fly her to a hospital in Athens.
"We were actually celebrating at first because we thought she had been saved. It was later we heard she had died," the friend told news.com.au.
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The 20-year-old from Port Macquarie in NSW died in hospital after hours in a coma.
The captain who witnessed the incident said he was deeply traumatised by the shock encounter.
The superyacht, owned by billionaire Mexican businessman Alberto Bailleres, has now left the Greek island. Mr Bailleres and his family had left two days before McNamara was found on the yacht, with only crew members on board during the incident.
Investigators have footage taken on the yacht's security cameras, suggesting they captured her final moments.
They also are understood to be examining her mobile phone for any messages she may have sent or received last Thursday night.
The captain of a neighbouring boat reportedly saw Sinead McNamara about 1.45am. Photo / via Instagram
McNamara's death came on what was said to be her final day as a crew member on the Mayan Queen. The model, who had more than 14,000 Instagram followers, had been working on the boat for four months and was soon to be reunited with her sister Lauren and mum Kylie, who were on their way to meet her in Greece.
She posted excitedly about the visit only a week ago. "Happy birthday to the most amazing big sister I could ever ask for, 5 days till I get to see you!! Excitement is an understatement," she wrote.
It followed a much darker message on her Instagram profile, in which she wrote: "My head is all over the shop today." The post was accompanied by emojis of a volcano, a tornado and a blood-soaked needle.
She then shared a picture of herself sitting carefree on a quad bike with a caption that said: "Take me back to this where my only worry was not cracking my skull open."
McNamara was active on social media and loved adventure — no more so than in the final weeks of her life.
Her last post detailed how she had been jet skiing in Komitata, Greece.
Greek authorities have said they are investigating the circumstances of McNamara's death and ordered an autopsy. A Port Authority official told news.com.au last night the autopsy results were not yet available.
The 20-year-old has more than 14,000 followers on Instagram. Photo / via Instagram
The Mayan Queen had initially been forbidden from leaving the port, however it left with the blessing of police on Sunday. All 23 crew members had been interviewed prior to the departure.
One local, who had seen the crew members, said they were quiet and visibly upset before they left. Another spoke of seeing several of them "in tears, they were all crying".
If you are worried about your or someone else's mental health, the best place to get help is your GP or local mental health provider. However, if you or someone else is in danger or endangering others, call police immediately on 111.
OR IF YOU NEED TO TALK TO SOMEONE ELSE:
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HONOLULU – In an effort to save lives, the Hawaii Department of Transportation (HDOT), four county police departments, and our private partners Toyota Hawaii and DTRIC Insurance are working together to educate Hawaii's roadway users about the dangers of distracted driving throughout National Distracted Driving Awareness Month in April. Free events will be held throughout April on Oahu, Kauai, and Hawaii Island. Event information is below.
Distracted driving is dangerous and claimed 3,166 lives nationwide in 2017 alone. In an effort to reduce this number, the National Highway Traffic Safety Administration (NHTSA) will be promoting their U Drive. U Text. U Pay. campaign from April 11, 2019 to April 15, 2019.
According to NHTSA, distracted driving involves anything that takes your attention away from the task of safe driving. This includes talking on your phone, eating and drinking, talking to people in your vehicle, fiddling with the stereo, entertainment or navigation system, and texting, the most alarming distraction of all.
In addition to conducting stepped-up high-visibility enforcement during April's National Distracted Driving Awareness Month, county police departments will continue its year-round enforcement of Hawaii's distracted driving law. Based on the 14,500 distracted driving citations police issued statewide last year, distracted driving continues to be a serious traffic safety priority.
Hawaii's law prohibits the use of mobile electronic devices while operating a motor vehicle, making it illegal for drivers to text or engage in other hand-held uses of mobile electronic devices such as cell phones, mp3 players, personal digital assistants, and navigation devices. The law also prohibits drivers from using a hand-held mobile electronic device when stopped at a red light or stop sign. Furthermore, no person under the age of 18 may use a hands-free mobile electronic device while operating a motor vehicle. The fine for violating this law starts at $297. Violations in school zones or construction areas are subject to a higher amount.
"We have learned that increased enforcement combined with public education have proven to be an effective method to reduce distracted driving and, more importantly, save lives," said Butay.
Locally, HDOT and our private partners Toyota Hawaii and DTRIC Insurance are coordinating efforts to educate Hawaii's roadway users about the dangers of distracted driving. HDOT's Distracted Driving Squad will conduct presentations, at high schools and shopping centers across the state, using a lightweight and portable virtual reality video system that can be attached to any motor vehicle. The presentations provide attendees the opportunity to experience the dangers and adverse effects of distracted driving in a simulate and safe environment. The distracted driving simulator system is the only one of its kind in Hawaii.


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Xenotransplantation, defined as the transfer of cells, tissues or organs between species, has been a subject of significant interest for decades as a response to the increasing demand for biological materials to treat patients. In this review, the history and recent progress in xenotransplantation research will be discussed, including the immunological challenges that need to be overcome and the molecular biological methods which are required to allow the complex genome engineering to meet the critical need for organs.
Over the course of the past 100 years, the rapid progress in drug development and surgical techniques has created a paradox for the area of transplantation medicine. Surgical protocols have become more successful and medicines have overcome many mechanisms of chronic rejection and allowed increased survival of transplant patients. However, the number of organs available for transplant has remained essentially constant. In addition, not all organs available through donation are viable for transplant. Organs such as lung, which are more prone to damage due to trauma, disease or deterioration, are available in drastically reduced numbers compared with heart or kidneys. Therefore, while there is an increasing number of patients who would survive and thrive long-term after organ transplantation, the limited number of organs available means a smaller percentage of eligible patients can actually undergo transplant surgery.
A hard truth about human organ donation is that even with exponential increases in donor numbers, it is unlikely that the organ shortage would be relieved. The diversity of the human species, paired with the efficiency of the immune system, significantly reduces the chances that a given organ will be compatible with the patients in greatest need. Although immunosuppressive drugs can enhance length of survival, chronic rejection remains a risk the greater the mismatch between organ and patient. Furthermore, the donor geographic proximity, organ size and timing of availability of a compatible organ with a matching patient may be limiting. Thus, even as human organ donation continues to be optimized, there remains an immense need for additional organs above and beyond the availability of human donors.
In order to address the above concerns and provide sufficient numbers of compatible organs, a number of approaches, both biological and mechanical, are being actively explored. Use of animal organs provides solutions to the challenges of availability and function. Multiple mammalian species possess organs which may substitute effectively for their human analogs and, in the case of agricultural species, can be rapidly bred in sufficient numbers to overcome organ shortages. Through use of controlled facilities, production of animals can be regulated and disease exposure eliminated. Additionally, careful breeding schedules can provide organs of appropriate size for any given patient on a predictable schedule for optimal timing of surgery. Finally, recent advancements in DNA sequencing and assembly and genome engineering technologies, paired with the advanced understanding of the cellular and molecular immunology responses in transplant rejection, allow the creation of animals which could provide an unlimited supply of rejection-free organs.
Examples of xenotransplantation can be found recorded as early as the seventeenth century, in which the transfusion of blood from animals into human patients was described . In the eighteenth century, more complex tissues such as skin were tested as grafts in human patients . In 1905, Princeteau transferred rabbit kidney sections into a child with immediate positive results, however, after 16 days the child died of pulmonary complications . Soon thereafter, two kidney xenotransplants were attempted, with one patient receiving an organ from goat, the other from pig. Unlike Princeteau's experiment, neither organs functioned and both apparently failed due to thrombosis . Similarly, an attempt by Unger in 1910 to transplant kidneys from a chimpanzee into humans led to failure due to thrombosis in about a day . In 1923, Neuhof transplanted a kidney from a lamb into a human patient, allowing the patient to survive 9 days .
In the early twentieth century, an odd offshoot of xenotransplantation was created due to interest in "rejuvenation" via transplant of animal testis in human males, as demonstrated by Voronoff in Russia  and Brinkley in the US  using chimpanzee or goat testis, respectively. So popular was the use of goat testis in the US, an entire radio empire was built around advertising the services, with many patients claiming enhanced fertility and sexual function .
The field of immunology developed in parallel with surgical approaches to xenotransplantation. As the mechanisms of immune rejection were better defined, the enormity of the challenges facing transplant of organs between members of the same species were recognized. During and after WWII, pharmaceutical companies created a series of increasingly effective immunosuppressive drugs which could inhibit some rejection responses, renewing interest in xenotransplantation.
During 1963–1964, Reemstma carried out a series of transplants into 13 human patients using chimpanzee kidneys, with one patient surviving 9 months after transplant surgery . The need for these experiments was driven in part by the desperate human organ shortage and lack of alternatives. Cadaveric organs often proved insufficient in quality, and volunteer human kidney donation, high risk at the time, was untenable for ethical and legal reasons. Although chronic dialysis had been demonstrated by the early 1960s, it was not widely available for patient treatment . Therefore, despite the risks, xenotransplantation was considered a potentially viable solution.
Reemstma was not alone in exploring xenotransplantation as a means to overcome critical organ shortages. Hume attempted transplanting a chimpanzee kidney into a human, but the organ failed to show renal function . Hardy and team focused on heart, observing survival for only 2 hours after transplanting a chimpanzee heart into a human patient . Starzl carried out a series of transplants in human patients with baboon kidney  and livers, with variable results . These seminal attempts at xenotransplantation showed that although surgical techniques and immunosuppressive drug treatments had greatly improved, they were insufficient to address the multitude of challenges in overcoming the xenorejection response. Indeed, it was nearly a generation later before Bailey used a baboon heart for transplantation into an infant, who survived several weeks after receiving the organ .
Although the close evolutionary relationship between non-human primates and humans would suggest an advantage in using chimpanzee or baboon organs for xenotransplantation, clinical, practical and ethical considerations prevent them from being a viable option. Non-human primate organs do indeed function almost identically to human organs, but are subject to a variety of diseases which are readily transmissible to humans . Given the relatively fragile state of patients receiving multiple immunosuppressive drugs, the risk of primate zoonoses is too great. In addition, chimpanzees, baboons and many other non-human primates are impractical for large scale breeding. The low numbers of progeny of non-human primates limits the production of large numbers of animals by natural breeding or in vitro fertilization compared with agricultural species. Finally, use of non-human primates as organ donors faces insurmountable ethical barriers.
A much more viable approach is the use of pig organs for xenotransplantation. Porcine organs are structurally and physiologically close to humans, and therefore can functionally substitute for analogous human organ functions. Unlike non-human primates, pigs are more evolutionarily distant from humans and thus have a greatly reduced risk of transmission of diseases to human patients, which can be essentially eliminated through genetic manipulation . Husbandry techniques for pigs are extremely well-understood, with large litter sizes and rapid cycle times, allowing production of populations that could overcome organ shortages in a much shorter timeframe than possible with non-human primates. Furthermore, a suite of genome engineering technologies is available for use in pigs to make critical changes to enhance survival and function of the pig organ, while avoiding the human immune rejection response. In fact, the complex engineering approaches now available may actually provide organs with advantages over even closely-matched human organs.
A variety of academic, clinical and industrial institutions have made substantial progress in recent years in the understanding of the molecular mechanisms of the xenorejection response and the genetic modification of pigs to overcome these mechanisms . Professional organizations are working with the FDA to develop guidelines for clinical use , with several groups indicating their intention to initiate clinical trials in the near term with porcine organs .
For xenotransplantation to become a viable routine human therapeutic option, there are a number of challenges that still need to be overcome. These challenges fall into two broad categories; the biology of the xenorejection responses and the engineering technologies needed to restructure the porcine genome to overcome these responses.
The immune system is an evolutionarily ancient collection of structures, mechanisms and cells that detect and eliminate harmful organisms from the host. In older texts, the immune system is often described as distinguishing "self" from "non-self," but more recent research demonstrates that there are a variety of roles for the microbiome ("non-self" micro-organisms resident within, on or around the host) in maintaining the health of the host organism. Thus, the host immune system must be able to not only identify and eliminate harmful pathogens, it must also tolerate the presence of a variety of beneficial bacteria, fungi and yeast . Because transplantation of cells, tissues and organs is an unnatural situation created through deliberate medical intervention, the human immune response uses incredible precision to identify even closely related human cells as "non-self" and efficiently removing them, a process referred to as "immune rejection." In general, the strength of the response is proportional to the degree of difference between the host and donor materials, therefore, when exposed to materials from an animal, the rejection response is much faster and stronger, increasing the challenge in controlling the immune response.
Xenorejection is a much more exaggerated and rapid form of the allorejection response. Four overlapping and inter-related reactions occur temporally; hyperacute rejection (HAR), acute vascular rejection/acute humoral xenograft rejection (AVR/AHXR), acute cellular rejection (ACR) and chronic rejection (CR) (Figure 1) . Although these processes can be characterized as distinct stages based on histological and clinical data, each is due to highly interconnected pathways and mechanisms that are challenging to separate. In fact, these responses were defined as pathologic observations prior to development of more detailed analyses of the molecular immunology mechanisms.
Overview of the xeno-rejection response. The human immune response to xeno-organs initiates within minutes to hours with hyperacute rejection (HAR, upper right), in which pre-existing antibodies (Ab) in human serum bind to xenoantigens (xeno Ag) on the surface of the pig cells. This results in cell destruction and presentation of porcine antigens to human helper (TH) and cytolytic (TC) T cells, as well as groups of pro-inflammatory and pro-immunity cytokines and other soluble mediators. Activation of human helper T cells (TH) stimulates human B cells (B) over the course of days to weeks, resulting in production of induced antibodies (induced Ab) as part of the acute vascular rejection/acute humoral xenograft rejection (AVR/AHXR, lower right) response. These secondary antibodies are often more specific and higher affinity than the pre-existing human serum antibodies, and also cause cellular destruction of the xenograft. in parallel with AVR/AHXR, the acute cellular rejection response (ACR, lower left) is carried out by human NK cells (NK), macrophages (M) and the xenograft-specific cytolytic T cells (TC) recruited to the xeno-organ within days to weeks. The activated cells express a variety of molecules to attack the porcine cells, as well as secrete additional cytokines to recruit more human immune cells. After weeks to months, the human immune response may be again induced to react to the xeno-organ during chronic rejection (CR, upper left), leading to specific antibody (Ab) responses from B cells (B) or cytolytic T cell (TC) destruction. A large collection of cytokines, chemokines, complement and coagulation factors (center) play a key role in regulating the complex set of reactions occurring in every aspect of rejection.
HAR is primarily due to immediate binding of pre-existing host natural antibodies specific for xenoantigens expressed by the donor tissues. Antibody binding can activate the endothelial cells, causing the release of immune activators, as well as inducing the complement-mediated destruction of the endothelial layer, reducing the barrier function and allowing host cells to infiltrate the organ. Cell debris released by the damage to the endothelium and products of the complement cascade also stimulate coagulation and the innate inflammatory response. These pathways synergize during rejection to create stronger responses that are more pathogenic and can be less amenable to control.
AVR/AHXR, like HAR, is also mediated by host antibodies. However, instead of pre-existing natural antibodies, AVR/AHXR is often the result of humoral responses which lead to production of antigen-specific antibodies. The AVR/AHXR is delayed due to the time it takes to induce an adaptive immune response via germinal center reactions, typically days to weeks. Much like natural antibodies, the induced antibodies recognize components of the xeno-organ and, similar to HAR cause activation of the endothelial cells and their destruction via the complement system. The specific antibody binding also attracts multiple elements of the cellular immune system, such as NK cells and phagocytes, creating further damage of the target tissues and secreting soluble factors, such as cytokines and chemokines, which further enhance immune responses.
ACR includes predominantly cellular responses to the graft, such as T cell activation, which occur within days to weeks of organ transplant. Although ACR is well-established in allotransplant, the importance of ACR in the xenorejection response is not entirely clear. This may be due to either the more rapid activation of hematopoietic populations in HAR and AVR/AHXR compared with allotransplantation. However, some groups have proposed that reduction of the HAR and AVR/AHXR in earlier stages of xenorejection would unmask ACR which would otherwise be unnoticed amidst the earlier more pathogenic responses. In either case, ACR is expected to be substantially similar between allo- and xenotransplantation and thus more readily controlled by immunosuppressive drugs already in use for allotransplant.
CR is longer term, occurring within months or even years after transplantation. CR can be due to complications due to other immune activity, such as infection, or escape of humoral or cellular responses from immunosuppressive drug control. CR is well-understood in allotransplant and effective treatments are available for control and reversal of CR.
HAR and AVR/AHXR are the most unique and most critical to address in xenotransplantation. These earlier reactions can greatly enhance later reactions, with some of the mechanistic elements of the xenorejection response initiated even before the transplant surgery itself occurs. Therefore, it is essential to control the initiating events as early as possible in order to reduce the course of later responses. Much like the layers of an onion, removing one layer reveals the next, but as each layer is removed the overall size may be diminished.
The latter two responses, ACR and CR, are mechanistically similar between xeno- and allorejection responses . Use of currently-available immunosuppressive drugs are believed to be able to control both responses as evidenced by the extensive data from allotransplants in humans. However, the speed and violence of the HAR and AVR reactions against xeno-organs can greatly accelerate and strengthen ACR and CR. Thus, even well-established treatments for allorejection may need to be reviewed as xenotransplantation proceeds toward clinical trials.
The immune system has two inter-related arms; the innate and the adaptive immune systems, both of which contribute to the rejection of xenotransplanted cells, tissues and organs. Although often described as separate, the systems have a large network of connections which are interdependent, and thus are not completely distinct. Both systems utilize multiple mechanisms to protect the host, creating a series of defense layers of increasing specificity. When functioning properly, a given layer may not be 100% efficient, but in aggregate will capture the overwhelming majority of pathogens. In addition, the ability to detect subtle differences between highly related cells has the potential benefit of identifying and eliminating cells with oncogenic mutations, preventing tumors before they have a chance to establish themselves .
The innate immune system is evolutionarily ancient, with related mechanisms found in both plants and animals. The innate immune system consists of relatively invariant mechanisms for the identification of pathogens, and, although less specific, is extremely rapid and strong in response. The rapidity of the innate immune system provides an immediate barrier to pathogen infiltration and infection of the host, limiting the pathogen burden and giving the adaptive immune system time to develop more specific responses .
The use of physical barriers is one of the most critical elements of innate immunity. Although organ transplant bypasses the skin as a protective layer, the endothelium of the blood vessels, which connect the organ to the host circulatory system, remains as the main interface between the human hematopoietic system and xeno-organ tissues. As such, many of the immediate mechanisms of the innate response are greatly influenced by the interactions between the human immune cells and the porcine endothelial cells. Once the human innate immune system attacks the porcine endothelium, the barrier function is quickly lost, followed by rapid influx of human immune cells, pro-inflammatory infiltrates and edema, and then necrosis and destruction of the xeno-organ. It is important to note that the endothelium is an extremely active part of the immune response, which responds to soluble factor and cellular interactions to induce a variety of immune and inflammatory responses. Therefore, any efforts to improve the engraftment of xeno-organs must take into account the functional role of the endothelium in regulating the rejection response .
Inflammation is one of the earliest innate responses, driven by pattern recognition receptors found on human immune cells which recognize damage-associated molecular patterns (DAMPs). The binding and signaling of DAMPs causes the immediate secretion of proinflammatory mediators, such as cytokines and chemokines, which attract additional innate immune cells and induce a variety of local responses which would be highly beneficial during an infection but destructive to xeno-organs. For example, vasodilation and increased vascular permeability, which would normally allow host immune cells greater access to tissue to rapidly eliminate pathogens, instead causes the xeno-tissue to be more quickly infiltrated by human innate immune cells, which in turn leads to more inflammation and destruction. Similarly, there are blood-borne proteinaceous biochemical cascades activated by inflammation, such as the coagulation and the complement systems, which further degrade xeno-organ function and survival .
The genes encoded by the porcine genome can encode proteins that are substantially different from their human counterparts or may carry post-translational modifications which are not present in humans. Interestingly, some of these molecules, referred to as "xenoantigens", are recognized by pre-existing natural antibodies found in human serum. One subset of these antigens is the swine leukocyte antigens (SLA), which are the physical and functional equivalent of the human leukocyte antigens (HLA). Much like the case for human allotransplant, the SLA genes are highly diverse and individual patients will have a variable level of cross-reactive antibodies in their serum for a given set of SLA genes . A separate group of xenoantigens are glycan molecules, such as Gal alpha (1,3) Gal and Neu5Gc, which are expressed in porcine, but not human, cells .
Although specific induced antibodies are produced by B cells as part of the adaptive response, the presence of pre-existing antibodies in human serum contributes to the innate response. The specific reasons for the existence of these human natural antibodies are not entirely clear. In the case of glycan structures, one hypothesis is that the molecules are related to those found in pathogens, and that the natural antibodies are cross-reactive to each. Alternately, consumption of porcine materials in the human diet may induce antibody formation. Regardless of the specific source in human serum, xenotransplantion of porcine cells and tissues in humans leads to binding of these pre-existing natural antibodies, activation of complement and eventual destruction of target cells carrying the xenoantigens.
Several approaches have been taken to address xenoantigens, including cross-matching donors and recipients for reduced immunoreactivity, removal or modification of the xenoantigen from the donor pig, or the reduction of the ability of the antibodies to induce the complement cascade. In the first case, typing of patients and porcine donors to find the best matches would be very similar to the current system used for determining allotranplant cross-reactivity . Use of gene targeting or editing technologies can eliminate the genes encoding SLA or the enzymes required for expression of the relevant glycan. This has been proven to be highly effective for ablating the genes GGTA1 (the gene encoding alpha 1,3-galactosyltransferase essential for Gal alpha (1,3) Gal), CMAH (cytidine monophosphate-N-acetylneuraminic acid hydroxylase critical for Neu5Gc biosynthesis) and B4GALNT2 (beta 1,4 N-acetylgalactosaminyltransferase). In each case, the elimination of the glycan leads to greatly reduced recognition of porcine cells by natural antibodies in human serum, and reduction in complement-mediated destruction . Unfortunately, as the number of antibody targets increases there is a risk that one or more of the xenoantigens alone or in combination may have essential functions which cannot be eliminated without damaging the development or function of the pig. Therefore, efforts to introduce more subtle mutation in SLA which remove immunogenic epitopes while leaving critical antigen-presentation functions intact, or even replacement of SLA with HLA, may be more effective.
The second approach, which is often used in combination with the first, is to reset the threshold at which the complement cascade is activated, making it more difficult for the binding of human natural antibodies to targets on porcine cells to induce the complement cascade. There are a series of "complement regulatory proteins" (CRPs), such as CD46, CD55 and CD59, expressed on the cell surface which prevent complement activation by the inadvertent non-specific binding of human antibody to human cells . By overexpressing one or more of the CRP molecules on the porcine endothelium, the amount of antibody binding required for complement activation is increased, which reduces the amount of antibody-mediated cell destruction due to human natural antibodies .
Inflammation and vascular leakage, due to loss of endothelial barrier function, both induce coagulation, which normally is required to repair localized endothelial damage. In the case of xenotransplantion, the attack of the endothelium is rapidly occurring at multiple sites, therefore, coagulation spreads throughout the blood vessels in the xeno-organ and can overcome the normal control mechanism. The thrombosis produced by the procoagulant environment leads to occlusion of the vessels within the graft, known as thrombotic microangiopathy (TM). The lack of blood flow results in hypoxia and tissue damage and necrosis, further complicating transplant function. The relatively greater amount of endothelial injury and coagulation in xenotransplant therefore creates more frequent and extensive TM and contributes to the more rapid destruction of the graft .
In addition to physiological pathways induced by human innate immune responses, there are non-physiological activities caused by mismatches between porcine and human constituents of the coagulation cascade . For example, porcine von Willebrand factor (vWF) has been shown to bind more avidly to the human GP1b receptor and activate human platelets, leading to coagulation and rapid loss of platelets from the circulation . Ongoing efforts seek to engineer porcine vWF to eliminate the inappropriate interactions with GP1b, while maintaining normal coagulative phenotypes. In addition, porcine proteins which provide positive and negative feedback to control the coagulation cascade do not function as efficiently upon the human coagulation targets, leading to dysregulation of the cascade. The targeting the porcine genome to express human regulatory proteins in porcine cells has been shown to help control human coagulation in response to exposure to the modified porcine materials .
Macrophages and neutrophils are two of the earliest host cell types to infiltrate xeno-organs. Both cell types are instrumental in the phagocytosis and destruction of pathogens during infection. During a xenorejection response, the damaged porcine cells release a variety of DAMPs which are recognized by the human innate cells, inducing phagocytic functions which further damage the xeno-organ and increasing production of additional proinflammatory and other immune mediators which attract more innate immune cells [36, 37].
Similar to the molecular mismatch described above for vWF and coagulation, macrophages express the SIRPA receptor, which must interact with the surface receptor CD47 to prevent the target cell destruction by the macrophage. Thus, the CD47 receptor expressed on the cell surface binds to SIRPA to instruct the macrophage not to consume the target cell. In the case of porcine CD47, the interaction with human SIRPA appears to be unproductive and cannot inhibit the macrophage activity. Expression of the human form of CD47 in porcine cells has been shown to greatly reduce human macrophage activity directed against the porcine cells .
NK cells are functionally analogous to cytolytic T cells, and even share some mechanistic pathways for targeted cell destruction. NK cells express a collection of stimulatory and inhibitory receptors on the cell surface, which engage conserved targets on the surface of target cells. The balance of activation and inhibition via combinatorial signaling determines whether the NK cells are stimulated to kill or ignore the target cell. The target cell receptors, such as HLA-E, may be perturbed by pathogens or tumorigenesis, which is detected by the NK cells and the target cells eliminated .
In the case of xenotransplantation, the porcine cell receptors, although expressed normally, are not sufficiently well-conserved with their human counterparts and thus cannot inhibit NK cell attack. By expressing on porcine cells the human versions of receptors which stimulate the inhibitory receptors on NK cells, the damage may be averted. With careful genetic modification, the normal mechanisms for detection of infection or other dysfunction may be maintained, allowing normal NK functions while eliminating the xeno-specific destruction [40, 41].
There are a variety of mechanisms used to resolve innate immune reactions. Many of the soluble mediators of innate immunity have extremely short half-lives which allows them to dissipate quickly. In addition, immune receptors become increasingly desensitized to further stimulation during the course of the innate response, reducing reactions. A variety of negative regulators are also produced to further inhibit the innate effectors. All of these mechanisms are in place to prevent over-reaction of the immune system and the destruction that it can cause once the pathogenic threat has been eliminated . In the case of a xenorejection response, however, the "threat" that is recognized comes from every porcine cell and thus the innate response is never fully resolved without intervention. It may be possible to take advantage of these resolution mechanisms to create porcine cells with an enhanced ability to curtail or end a human inflammatory response through careful genetic modification.
The adaptive immune system is comprised of cellular and antibody components which recognize pathogens and develop highly specific responses, which can increase in specificity and effectiveness over time and exposure. The adaptive immune response also creates immunological "memory" to allow more rapid reactions should similar pathogens be encountered in the future. Because of the time required to develop specific responses, the adaptive immune system generally becomes more critical after the initial innate immune response .
Antigen presentation is a crucial mechanistic part of the adaptive immune response and plays a major role in the decision between immunity and tolerance for a given target. There are two main routes for antigen presentation to the immune system, reflective of the different classes of pathogen antigens, intracellular or extracellular.
Intracellular antigens, either natural cellular proteins or those derived from viral or bacterial infection of cells, are enzymatically cleaved into peptides which bind to the ubiquitously expressed class I human lymphocyte antigens (HLA). The peptide-HLA class I complex is displayed on the cell surface where it can be surveyed by the binding of cytolytic T cells expressing T cell receptors (TCR) and CD8 co-receptors on the T cell surface. Similar to antibodies, TCRs are assembled combinatorially, creating a diversity of specificities for HLA-peptide complexes, with only a small subset of TCRs binding to a given complex. Should a given CD8 T cell be activated by the HLA-peptide complex, it will express a series of cytolytic molecules which kill the target cell. This system works due to the efficient T cell selection mechanisms applied during T cell development. After initial production of a rearranged TCR, the nascent T cell is tested in the thymus for inappropriate reactivity against cellular antigens. If the T cell survives the selection process, it exits to the body and theoretically will only be activated when it encounters and antigen that does not naturally exist in body, such as a peptide from a pathogenic organism, or a mutant peptide from an oncogenic cell .
Extracellular antigens can be any molecule taken up by a cell from its environment and degraded in lysosomes intracellularly. The resulting peptides are then loaded onto HLA class II molecules which, unlike HLA class I, are expressed on only a subset of immune-related cells. The class II HLA-peptide complex is recognized by a different T cell subset expressing TCRs with the co-receptor CD4. The CD4 T cell subset also undergoes thymic selection as observed with CD8 T cells, to eliminate recognition of self-antigens . However, CD4 T cells can be induced to create different phenotypes once they specifically recognize class II HLA-peptide complexes. A large variety of T cell subsets have been described, including production of helper T cells, which participate in the activation of B cells for the production of antigen-specific antibodies, or regulatory T cells, which act to inhibit the immune response . The choice of outcomes is driven by the soluble mediators, such as cytokines, found in the local environment, and the collection of co-receptors expressed on the antigen presenting cells.
HLA itself is a significant direct contributor to rejection responses outside of its role in antigen presentation. As described above, T cells are selected for lack of recognition of self-antigens. This not only includes the recognition of self-peptides bound to HLA molecules, but of the HLA molecules themselves. Normally, T cells bearing TCRs with inappropriately high affinity for binding HLA molecules, even in the absence of peptide, are eliminated early in T cell development. Because the human T cells have not been exposed to, or selected by, the class I or II swine lymphocyte antigens (SLA), a subset of human TCRs will bind to SLA and induce strong T cell activation, regardless of the peptide presented in the SLA . As porcine cells are attacked by the human immune system, donor peptides are efficiently presented by human cells via HLA to human T cells as part of the normal human adaptive response. Conversely, depending upon the organ transplanted, there can also be donor T cells and antigen presenting cells transferred which result in donor immune responses against the host tissues, referred to as graft versus host disease (GvHD) . In all cases, the immune cells are responding normally, but in the setting of xenotransplantation can be extremely pathogenic due to the artificially high concentration of immunogenic targets present.
Because HLA matching is part of organ selection in allotransplantation, a frequent question is whether introduction of human HLA in place of porcine SLA would help overcome rejection. Although SLA ablation may be helpful in averting antibody-dependent damage, this approach does not resolve some of the challenges related to antigen presentation in xenorejection responses . It is true that the human T cell binding directly to pig SLA could be eliminated by substitution of SLA with HLA, however, the HLA genes are highly polymorphic, hence the need to HLA match human patients. This means that for a given patient, a donor pig would need to be engineered to specifically express the HLA homologous to that patient, which would be limiting given the timelines necessary for production and validation of genetically-modified pigs. In addition, in the normal situation human cells display human peptides in the HLA, the overwhelming majority of which will be conserved between a human donor and recipient, and thus much less likely to induce a response. If pigs were engineered to express human HLA which is perfectly matched to the patient, the donor porcine cells could now be significantly more efficient at displaying porcine peptides to the human immune system and more rapidly induce T cell activation. Therefore, introduction of human HLA in place of porcine SLA may not provide a benefit without additional engineering.
Humans possess a number of pre-existing antibodies specific for porcine antigens which can contribute to the xeno-organ damage during HAR. As the donor tissue is damaged, the antigens are released and presented to T cells as described above, causing the activation of helper T cells. These T cells interact with B cells in lymphoid organs, inducing the activation of any B cells which express antibodies specific for the xeno-antigens. This initiates the germinal center reaction, in which antigen-specific B cells rapidly proliferate and mutate their antibody sequences and are then progressively selected for improved antibody function. The resulting B cells expressing the affinity-matured antibodies exit the germinal center and can differentiate further to plasma cells, which act as factories that can produce extraordinarily high levels of serum antibody . These induced antibodies, like natural antibodies, further amplify AVR/AHXR and contribute to the destruction of the xeno-organ.
The de novo production of antibodies can be quite rapid and are a risk for the lifetime of the transplant whether for allo- or xenotransplantation. There are a number of drugs available for the control of B cell reactions. One of the most effective approaches is the depletion of B cells using antibody therapeutics such as Rituxan, specific for the CD20 surface molecule . However, the constitutive ablation of host B cells will create long term immunosuppression and could be prohibitively expensive. Although highly related to CR in allotransplant, the B cell responses in xenotransplant are stronger and more challenging and likely to require more stringent therapeutic control.
Advancements in understanding of immune mechanisms in immune rejection have elucidated a number of targets and pathways for intervention, and discovered a variety of small molecule and protein therapeutics for the suppression and manipulation of the immune system. However, the restraint of the immune system required to prevent xeno-organ rejection places the patient at significant risk of infections, tumors and other diseases which are preventable by an intact immune system. Therefore, there is a growing interest in the application of immune tolerance mechanisms in the transplant setting.
Immune tolerance is the natural unresponsiveness of the immune system to targets which may otherwise create an immune response. As mentioned previously, there are many mechanisms used by the immune system to identify non-self-antigens to prevent autoimmune diseases. As the body of literature regarding the molecular basis of immune tolerance has grown, interest in testing tolerance mechanisms in xenotransplantation has also increased .
Mixed chimerism is one route that has shown significant promise in both allo- and xenotransplant settings. This approach combines the transfer to the recipient of both the organ and hematopoietic cells from the donor. Typically, the patient is pre-treated with radiation or drugs to allow hematopoietic cell engraftment prior to the organ transplant. The combination of hematopoietic cells from host and donor allows cross-tolerance of host immune cells to donor tissue as well as donor immune cells to host tissue. Therefore, the resulting immune system is a combination of the donor and host, or a "mixed chimera," which recognizes the donor organ and host tissue as "self" despite the differences in genetic origin .
A further refinement of mixed chimerism includes transplant of donor thymus into the recipient, allowing selection of host T cells via donor antigen presentation , suggesting that tolerance is T cell dependent. A large body of evidence points to the role of regulatory T cells (Treg) as a driver of immune tolerance. Treg cells are antigen-specific but upon binding of the specific HLA-peptide complex on antigen-presenting cells will produce a variety of immune inhibiting and tolerogenic factors. The Treg cells may be derived from either thymus selection (central tolerance) or selection in tissues (peripheral tolerance), with central tolerance believed to be more durable, and the conceptual basis for donor thymus transplantation in mixed chimerism .
A critical factor in the maintenance of tolerance is the balance between Treg and effector T cells over time. Any imbalance that increases the number of effector T cells can rapidly lead to immune rejection. If indeed the Treg population is the main active component of immune tolerance, then it may be desirable to specifically bolster the numbers of Treg cells transferred to the recipient to more greatly ensure that the balance is biased firmly toward tolerance. A number of groups have established protocols for the generation of Treg cells that are specific for xeno-organs and tissues through in vitro selections and expansions . While this has been shown to have positive effects in allograft tolerance, the durability is variable and, worse, some studies have described conversion of Treg to effector T cells which then contribute to rejection . Despite these concerns, mixed chimerism, with or without Treg supplementation, remains a potentially valuable approach to immune tolerance.
The progress of xenotransplantation research in recent times has closely paralleled the advancement in genome engineering technologies. As the complexity of the engineering toolsets has increased, so too has the complexity of porcine genomic manipulations increased to address the immunological challenges described in previous sections.
Complex mammalian genome engineering has advanced much more rapidly in mice than in virtually any other species, including pigs. The reason for the rapid progress in mice is the availability of embryonic stem (ES) cells which can be maintained in culture for extended periods time and undergo extensive transfection/transduction protocols and drug selection without losing the ability to produce large numbers of fertile progeny via blastocyst injection . Although several labs have made strides in this area, similarly manipulable and viable ES cells are not currently available for routine use in the generation of cloned pigs .
The most common approach for production of genetically-modified pigs is very similar to the protocol described in the creation of "Dolly the sheep." Briefly, the nucleus from a pig cell carrying the desired genome changes is extracted and introduced into a pig oocyte, which has previously had its own nucleus removed, and then induced to initiate embryogenesis using electrical and chemical induction, a process referred to as somatic cell nuclear transfer (SCNT). The newly-created cells are implanted into surrogate female pigs and allowed to develop to birth. Compared to genetically-modified mouse production, this process is significantly less efficient and more costly, limiting the number of facilities capable of effectively carrying out this complex process .
A key factor in the success of SCNT is the source of the donor nucleus. These cells are typically primary cells derived from fetal sources. Extended culture, transfection, or drug selection of these donor cells can all cause a significant loss of viability for subsequent productive SCNT. Therefore, the approaches commonly used for mouse ES cell manipulation such as multigenic targeting and selections with various drugs over long periods in culture would not allow for production of modified pigs using SCNT. Similarly, any genome manipulations of pig cells must also maintain the viability of the cells for SCNT, which alters the approaches available compared with mice.
One of the earliest genome engineering approaches applied to pigs was introduction of gene knockouts (KO). For any given gene, mutations which remove or disrupt the coding sequence can eliminate the expression of the gene and, provided that the KO is not lethal, create an organism which is entirely missing the gene product. The introduction of gene KO technology has been a key factor in the rapid advancement of the field of xenotransplantation .
As discussed above, there are several glycan molecules present in pigs which are absent in humans. These glycans are recognized by antibodies present in human serum which leads to rapid and extensive antibody-mediated damage to the porcine cells. Therefore, the elimination of the specific carbohydrate structures should help prevent human antibody recognition of the pig tissues. Unlike protein antigens which are directly coded by the DNA, glycosylation is due to the action of enzymes which create post-translational modifications of a variety of proteins produced by the cell. Therefore, glycosylation pathways must be examined to identify the key enzyme that creates the immunogenic glycan while otherwise leaving cellular metabolism intact.
The GGTA1 gene is responsible for creating the Gal alpha (1,3) Gal epitope in pigs. Although the specific reasons for this are unclear, human patients can express high levels of antibody specific for the Gal alpha (1,3) Gal epitope, presenting a major challenge to xenotransplantation . The KO of the GGTA1 gene is one of the earliest genetic modifications of pigs for application in xenotransplantation, and results in greatly reduced human antibody recognition of porcine cells . However, elimination of the GGTA1 gene alone has been shown to be insufficient due to a variety of other xenoantigens present in pig cells which are recognized by antibodies present in human serum. Generation of KO of CMAH , B4GALNT2  and other xenoantigen genes have further decreased the reactivity of porcine cells to human serum. However, it is important to keep in mind that the greater the number of gene KO, especially when made in combination, may lead to detrimental effects on pig health.
The use of gene KO approaches is highly useful for eliminating xenoantigens but does not address the need for expression of human or synthetic versions of genes necessary for control or proper function of dysregulated pathways. This requires the ability to permanently introduce heterologous DNA into the genome in a manner which maintains gene function.
The initial approach to gene insertions was simply random integration of DNA into the target genome. These genes are introduced from elsewhere and thus termed "transgenes" (TG). Once the ability to introduce DNA into mammalian cells was established using a variety of technological approaches, it became clear that over long-term culture a subset of cells could be isolated which have permanently incorporated the heterologous DNA. Because many transgenes do not provide a straightforward means to identify cells which have incorporated foreign DNA from the population that have not, TGs often include genes encoding drug resistance markers. In order for cells to survive drug treatment they must incorporate the resistance gene, greatly reducing the population to be screened, and increasing the chance of identifying cells which incorporate the TG of interest along with the drug resistance gene .
The integration of transgenes is rapid but relatively uncontrolled. Although there may be some preferences for integration site based upon chromatin accessibility, these are hard to predict and may be related to DNA breakage sites at which repair mechanisms fortuitously insert the transgene DNA . The random nature of the insertions can create risks. For example, the same transgene inserted at different sites can yield highly variable results in expression. Furthermore, some insertions may be deleterious to cell function, causing them to grow more slowly or die off, or, if these cells are used for generation of animals in vivo, there is a possibility of insertions creating mutations, instability or even lethality.
Due to the risks of random integration, significant effort has focused on protocols to create targeted integration, or gene knock-in (KI), of heterologous DNA into the genome. This is accomplished in mice by taking advantage of ES cells which undergo homologous recombination. In this approach, the transgene of interest is flanked by DNA sequences that are identical to regions of the genome to be targeted. After introduction of the heterologous DNA, the regions of DNA sequence identity are aligned with the target sequence and the homologous recombination machinery creates crossover events to switch the endogenous sequence with the heterologous sequence. This approach is much less efficient than random integration of TG, therefore drug selection schemes often need to be employed to identify the relatively rare targeting events .
Homologous recombination is well-established for targeting in mice but requires ES cells which express the enzymes necessary for the targeting event. Unfortunately, porcine ES cells are not available that both possess homologous recombination function and can reliably generate cloned animals. For reasons that are not entirely clear, generation of ES cells competent for homologous recombination and cloning seems to be challenging for most species other than mice . Therefore, alternate approaches are required for targeted integration in the pig genome .
A number of novel enzymatic molecules have been created which help resolve the dilemma of targeted integration in porcine cells. Zinc Finger Nucleases (ZFN), Transcription Activator-Like Effector Nuclease (TALEN) and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) are all synthetic molecules based on genuine proteins which allow the precise targeting of genomic DNA based upon sequence [66, 67, 68]. In each case there are two functional components, a targeting module which recognizes a specific genomic sequence, and an enzymatic module which introduces a double-stranded DNA break at the target site. In the case of ZFN and TALEN, the targeting module is a complex array of protein sequences which have previously been shown to recognize specific DNA sequences and can be mixed in a modular way to bind to any desired sequence. Although both technologies have shown great success, the effort and cost required to identify a single functional molecule can be significant. In contrast, the relatively more recently recognized CRISPR, and related prokaryotic systems, is much more easily applied in mammalian cells. The DNA binding module in this case is RNA base-pairing to provide sequence specificity. The enzymatic module cleaves DNA, creating a double strand break similar to ZFN and TALEN. When heterologous DNA is present, the cellular repair machinery may use the synthetic DNA to repair the break, inserting the TG at the desired genomic site. It is important to note that all of these systems, ZFN, TALEN or CRISPR, are essentially the same in that they introduce double strand DNA breaks at a selected site in the genome and do not directly affect the rate of DNA insertion. Therefore, it is often necessary to include selection schemes for identification of the modified cells. The greater efficiency and ease of use of these systems, CRISPR in particular, has allowed targeted insertion of DNA into genomes that were not previously able to be modified .
Due to the challenges of creating genomic modifications in porcine primary cells while maintaining their viability for SCNT, more efficient engineering methods are desirable. One approach to enhance efficiency is to target a specific region of DNA, called a landing pad, with multiple genes at once. By inserting a DNA vector bearing multiple therapeutic genes at once, a large amount of breeding and testing can be circumvented using a single event. This approach has the added advantage of avoiding inefficient crossbreeding necessary to bring loci from distinct chromosomes together in one lineage. When combined with the use of tools such as ZFN, TALEN and CRISPR more rapid progress in the genetic modification of animals has been greatly facilitated .
The increasing sophistication and accessibility of genome engineering toolsets and deeper understanding of immunological rejection mechanisms has allowed greater advancement in xenotransplantation than ever before. A key question is just how many genetic changes are required in order to make a pig organ suitable for transplantation? While the critical experimental data needed for such an assessment is still accumulating, it is clear that the number of alterations required for one organ may be different from another. For example, xeno-hearts with relatively minimal genetic modifications have demonstrated months to years survival in transplantation studies with non-human primates, whereas xeno-lungs with more extensive modifications have yet to survive more than a few weeks. This is due to the relative differences in structure and function of organs, the resilience to trauma, and susceptibility to rejection responses. Furthermore, tolerance mechanisms may be able to supplant the need for some genetic modifications, and thus the specific protocols and treatments will govern the ultimate complement of alterations.
The immediate need in xenotransplantation is to define the specific genetics required for xeno-organ survival, however, it is possible to project further enhancements such that porcine organs may be superior to human organs for human transplant. Synthetic biology approaches have created novel genetic circuits which can react in real time to human immune responses, inducing counter-reactions in the porcine cells to circumvent and tolerize the xeno-organ against human rejection. Furthermore, xeno-organs may be engineered to express protein therapeutics to further control human immunity while saving hundreds of thousands of dollars in expensive biotherapeutic treatments. Thus, the first version of pigs appropriate for xenotransplantation are likely to be further refined and improved to create increasingly useful rejection-free organs.


text:
Aymard ist der Familienname folgender Personen:

 Antoine Aymard (1773–1861), französischer General
 Édouard Aymard (1820–1878), französischer General

Siehe auch:

 Aimard
 Aymardus

redpajama_set_name: RedPajamaWikipedia


text:
Michael Steinlechner (* 24. April 1987 in Schwaz) ist ein österreichischer Fußballspieler.

Karriere
Steinlechner begann seine Karriere in seiner Heimatstadt beim SC Schwaz. 2006 wechselte er zum Regionalligisten WSG Wattens. In seiner Debütsaison wurde Wattens Dritter in der Regionalliga West. In den folgenden Jahren spielte man immer um den Aufstieg mit, scheiterte jedoch in den Saisonen 2010/11 und 2011/12 in der Relegation. Im Sommer 2013 wurde er an den Bundesligisten FC Wacker Innsbruck verliehen. Für Wacker gab er im August 2013 sein Debüt in der Bundesliga, als er am 7. Spieltag der Saison 2013/14 im Spiel gegen den FK Austria Wien in der Schlussphase eingewechselt wurde. Nachdem er mit Wacker in die Zweitklassigkeit abgestiegen war, kehrte er zu Wattens zurück. Mit Wattens konnte er 2015/16 schließlich Meister in der Regionalliga West werden und somit in den Profifußball aufsteigen.

Zur Saison 2018/19 wechselte er zum fünftklassigen SV Fügen.

Persönliches
Sein Vater Hermann (* 1957) war ebenfalls Fußballspieler.

Weblinks

 Michael Steinlechner in der Datenbank von fussballoesterreich.at

Einzelnachweise

Fußballspieler (SC Schwaz)
Fußballspieler (WSG Tirol)
Fußballspieler (FC Wacker Innsbruck)
Fußballspieler (SV Fügen)
Österreicher
Geboren 1987
Mann