text:
Fraud, money laundering and embezzlement, often referred to as 'economic crimes', cost UK businesses an estimated £40bn a year, according to a new study endorsed by the Home Office. Research by BDO Stoy Hayward shows that the cost of fraud is certainly high and increasing: frauds worth more than £400m were registered in the first six months of this year, compared with just over £330m for the whole of 2003 (although the number of frauds remained constant).
Ministers are concerned, particularly because some of the most prominent fraud prosecutions have led to acquittals, and are planning to introduce new legislation (including, for the first time, a statutory definition of 'fraud') to make it easier to convict because of the impression that fraudsters are laughing all the way to their Swiss banks.
In many cases, criminal prosecutors use crimes that are easier to prove but which carry lower sentences (for example theft and deception). Further weapons in the prosecutors' armoury are expected to be included in the Queen's Speech next month, with the definition of 'fraudulent trading' being extended to individuals, and there will be a new offence of possessing equipment to commit frauds (such as possessing personal financial information belonging to others). Following pressure, the Government has also decided to retain the offence of 'conspiracy to fraud' – the second most frequent charge levied against fraudsters (after the offence of 'fraudulent evasion of VAT').
This impression is partly misleading, as the Serious Fraud Office (SFO) has a relatively good success rate for most frauds it prosecutes. Similarly, and without fanfare, the civil courts have created a hugely effective set of procedures that allow businesses in the UK to don the investigator's mantle and seek speedy recovery and justice against fraudsters.
Civil fraud prosecutions are a growth area and judges are surprisingly supportive of businesses that make use of this avenue to thwart fraudsters.
The courts recognise that going to the police with this sort of crime is not always effective because of the hurdles faced by businesses. These include:
- Often the police are not that interested in prosecuting corporate fraud. With limited resources, not surprisingly most police officers will concentrate on the crimes where the victims are human and the effects of the crime personally devastating. Well-resourced businesses come near the bottom of the force's priority list for police time. Only in the last couple of years has corporate fraud been recognised as one of the priorities for the SFO, and it will only become seriously involved in frauds of more than £5m.
- If the police do investigate, it will be highly disruptive, probably with many files and computers seized for forensic analysis. Also, the business will not see many of them back for months, whether the case requires them or not.
- It will take several years to recover the money (for instance, following the collapse of Versailles in 1999, victims have only this year been able to start proceedings to get a recovery from the directors).
By contrast, English civil law makes this country an excellent jurisdiction for bringing civil cases and also as the base for tracking and recovering assets internationally. The civil route allows victims (usually companies, but occasionally private individuals) to secure court orders that are almost draconian in their scope to seize assets and raid premises. Also, the English courts are prepared to open up suspects' bank accounts to help the victim find their stolen money and recover it.
The English court orders are treated with a high degree of credibility in most overseas jurisdictions (which is inevitable, with the more complex frauds invariably having an international element), and the UK benefits from reciprocal enforcement treaties which allow it to enforce its orders in many of the offshore jurisdictions where crooks stash their loot.
In addition to allowing a fraudster's assets to be frozen within days, often a civil judgment will be obtained within 12-15 months, with success more likely because of the lower burden of proof.
There will also be a punishment element, as the fraudster will have to pick up the costs if found guilty. Similarly, they will go to jail through contempt of court if they lie or destroy evidence, while the evidence can (with the judge's permission) subsequently be passed to the police for a follow-up criminal case. One client was able to get back more than £6m of defrauded money within six weeks through the civil courts. The subsequent criminal case landed the fraudster a sentence of over four years.
The corollary of this encouragement of civil cases is that English judges are very tolerant of clients instructing investigators to seek evidence for the prosecutions. This has seen a corresponding growth in tertiary industries, including accounting and computing forensics, surveillance, private investigators undertaking covert investigations, telephones being tapped (in an employment context) and computer traffic being monitored.
However, there has to be some care with how the material is obtained, both from the legal and reputational viewpoints. For instance, courts do not particularly like people going to the extent of trampling over someone's human rights and will be particularly upset if there is a failure to disclose to the judge any relevant investigative methods.
Whatever new legislation emerges from the Government to combat fraud, for most businesses the first avenue for the prosecution of fraud will continue to be the civil courts to, at the very least, maximise the recovery of assets and secure evidence. The criminal route should, for most cases, come second because of its inevitable slow pace.
The Government is concerned about the ability for the criminal authorities in the UK to secure convictions for fraud, and in its forthcoming Fraud Bill it is reported to be adding further weapons for prosecutors, with the definition of 'fraudulent trading' being widened as well as a new offence coming into being of possessing equipment to commit frauds (such as possessing personal financial information belonging to others).
This may help, as research by BDO Stoy Hayward shows that prosecutors currently frequently resort to charging fraudsters with easier to prove crimes carrying lower sentences (such as theft and deception).
BDO's research into all criminal fraud cases reported since January 2003 also raises the question of whether sentences are sufficient, given both the severity of the crime and in order to have a deterrent effect. Certainly, it is the norm for a convicted fraudster to receive a custodial sentence.
However, few receive the seven years handed out earlier this year to former Goldman Sachs PA Joyti De-Laurey for stealing £4.4m from her bosses' private accounts. As the attached chart shows, most fraudsters will escape such a sentence. Above average also are the sentences recently given to the main directors of Versailles, of six years each, for what judge Mr Justice Jackson described as "a massive fraud" and "a grand and evil scheme", which cost its associated traders £23m and others many millions too. It is hard to see a US judge finding that a six-year sentence fitted such a large fraud. For example, in March tax specialist James Olis, a mid-ranking executive at Dynegy (Enron's smaller rival), was jailed for 24 years over his part in a $300m (£162.4m) accounting fraud. (And in the US the sentence you get is typically the sentence you serve.)
Given that fraud, including software and branded goods counterfeiting, are areas increasingly being taken up by organised crime (partly to avoid the risks and sentences associated with the traditional areas of robbery and drugs), making it easier to prosecute fraudsters is unlikely to stem the rising problem of fraud unless the penalties do ensure that it is a crime that does not pay.
Certainly, the typical fraud at the moment continues to be centred around directors or senior employees stealing from their own businesses, with personal greed or domestic difficulties their driver.
However, sophisticated criminals realise that more money can be made by donning a suit and tie, rather than a balaclava. The signs of this trend towards a greater undertaking of economic crimes by organised criminals are already present. Given that a fraud can be at least as devastating to its victims as a robbery, sentencing policy needs to be seriously reviewed to ensure the penalties in this growth area are in line with other crimes of the same magnitude.


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X FACTOR judge Nicole Scherzinger shrugged off rumours she has split with Lewis Hamilton.
Me and my lovely man are lovely, that's what's going on. We're great
The ex-Pussycat Doll, 34, and Formula 1 ace's romance was said to be on the rocks after recent photos emerged of her in an embrace with singer Chris Brown.
But Nicole denied their relationship was over, saying: "Me and my lovely man are lovely, that's what's going on. We're great."
US star Nicole has just found a pad in London to live in while she works on The X Factor.
And Lewis, 27, her when he returns from Japan.
Nicole giggled: "I love cooking for my boyfriend. He likes my cooking. His favourite meal is so easy. It's just chicken fajitas."
But her move from the US is not long-term, as she said: "The weather would stop me moving here."
Nicole will be on TV tomorrow with her boys Rylan Clark, 23, Jahmene Douglas, 22, and James Arthur, 23, who will be singing for survival on the first live show where the theme is Heroes.
● SIMON Cowell claims his talent shows actually "benefit" from going up against strong rivals like The Voice and Strictly.
He said yesterday: "I see no end in sight for these shows."

subset: commoncrawl


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The big story of Week 5 was no doubt the backup or, in the case of Calgary's Bo Levi Mitchell, the backup to the backup quarterback.
Four passers who were expected to carry a clipboard all season started games over the last six days, and most of them enjoyed plenty of success.
In fact, the only backup who didn't win lost to another backup. That's how good of a week it was for the reserve pivots.
Now, of course, the natural reaction is to try and figure out which of these backup quarterbacks the Ottawa RedBlacks are going to covet four months from now. We need to remember, however, that it's only one game.
Bombers backup quarterback Ryan Dinwiddie threw for 450 yards in his first regular-season game — it came six months after his surprise start in the 2007 Grey Cup — and got the win. Interestingly, he never won another game.
So while it's too early to be making proclamations about which quarterback the RedBlacks should grab, the early projections look good. Here's a gander at the four backup performances from Week 5 and the odds they'll even be available for Ottawa:
1. Bo Levi Mitchell
The 23-year-old Stampeder dissected the Bombers' secondary, completing 29 of 33 attempts for 376 yards and three TDs, and was named the CFL's offensive player of the week on Wednesday. He had a fantastic safety valve in running back Jon Cornish, but Mitchell showed plenty of poise in the pocket.
Ottawa Odds? Not good, as Calgary is likely to protect him. Drew Tate can't stay healthy, while Kevin Glenn isn't someone to build a franchise around at this stage of his career.
2. Zach Collaros
The former Cincinnati Bearcat juked and jived and threw his way to a 38-12 drubbing of the B.C. Lions on Tuesday night. It was an impressive performance, for sure. In my eyes, though, he's the one who has to follow it up with another strong showing or two for me to be convinced. There were a couple balls that could have been picked, but he does have the athleticism.
Ottawa Odds? Probably not. He could be the guy who takes over for Ricky Ray at some point down the road, however.
3. Drew Willy
The 26-year-old, who bounced around the NFL for a few years, took over for Darian Durant and guided the Roughriders to their fifth straight victory. Willy basically managed the offence instead of being the star, but it was a polished and mature showing nonetheless. He got the ball to the right people.
Ottawa Odds? You never know. If he has a few more games like that, Marcel Desjardins might take a second look.
4 Justin Goltz
The fourth-year Bombers pivot finally made his first start against the Stampeders, and he was by far the second best backup on the field next to Mitchell. That being said, the man who looks to be in the process of becoming Winnipeg's No. 1 gun didn't have too terrible a night. He threw two touchdowns against only one pick, and he has the arm and the legs.
Ottawa Odds? There's a good chance the Bombers will protect him. They have no one else to protect.
If the Riders can win one more game, they will match their franchise record for best start at 6-0 … The CFL announced its Week 5 fines on Wednesday. Alouettes receiver Brandon London got fined for his comments about the officials after Montreal's loss to Calgary, while Saskatchewan special teamer Levi Steinhauer got dinged for drilling Ticats returner Lindsey Lamar with the crown of his helmet … In case you missed it, Calgary kicker Rene Paredes broke the CFL record for consecutive field goals made. He was 5-for-5 on Friday in Winnipeg and has now connected on 34 straight. B.C.'s Paul McCallum held the previous mark, which was 30 … The CFL players of the week are Calgary quarterback Bo Levi Mitchell (offence), Montreal linebacker Kyries Hebert (defence), Montreal kicker Sean Whyte (special teams) and Montreal linebacker Shea Emry (Canadian) … The Alouettes have lost defensive tackle Scooter Berry for the rest of the season after he tore two knee ligaments, including his ACL, in last Thursday's game against Edmonton … The Lions are facing more pressure on their offensive line after rookie guard Kirby Fabien probably tore a few ligaments in his knee on Wednesday night in Toronto … Brandon Isaac's debut with the Hamilton Tiger-Cats didn't last long, as he pulled up lame with a leg injury against the Roughriders and is expected to miss Friday's contest in Edmonton.
We've said it before, and we'll say it again: STOP HANDING THE BALL OFF WHEN ONLY ONE YARD IS NEEDED — ESPECIALLY OUT OF THE SHOTGUN!
Yes, we're talking about Edmonton head coach Kavis Reed and offensive co-ordinator Doug Sams, who somehow let Eskimos quarterback Mike Reilly take a snap out of the shotgun and hand it off to John White on the last play of last Thursday's game in Montreal.
Shockingly, White got gobbled up well short of the end zone and the Esks suffered a five-point loss. If only they had lined up under centre and had Reilly plunge …
Also, Reed said a few days after the game that there was confusion because they thought the CFL command centre was going to review the previous play, during which White almost made it into the end zone.
Sorry, but a team should always go on the notion that the play isn't going to be reviewed. Coaches always talk about taking care of the things they can control. Getting a play reviewed in the final three minutes is not in their control, so they should have been thinking more about the final play instead of whether they were going to get a booth review.
Had they done that, they probably would NOT HAVE HANDED THE BALL OFF WHEN ONLY ONE YARD WAS NEEDED!!!!!!
Good for Matt Nichols, the Eskimos quarterback who is spreading the good word of the CFL while rehabilitating his injured knee.
Here's what PGA Tour player Hunter Mahan put out on Twitter last Friday night during the Bombers-Stampeders game: "I'm watching Canadian football and a team scored a single point. Someone let me know how that's possible."
Mahan was in Toronto for the RBC Canadian Open, and he was actually leading on Friday night, but he flew home on Saturday to be with his wife for the birth of their first child.
Anyway, before Mahan left the country, Nichols dropped some CFL knowledge on the 30th ranked golfer in the world: "@HunterMahan Any punt or FG that doesn't get brought out of the endzone is a rouge (single) for kicking team. #CFL"
Responded Mahan: "Perfect explanation! Thank you."
1. (1) Saskatchewan
Doesn't matter who plays quarterback, the Green and White get it done.
2. (2) Calgary
Third-string pivot Bo Levi Mitchell sliced and diced the Blue and Gold.
3. (4) Toronto
CFL teams take note: Chris Jones' defence looks like a Chris Jones defence again.
Young receiving corps and a weakened offensive line couldn't keep up with Jones.
5. (5) Montreal
Lucky to eke out the win over the Esks, but there are signs of improvement.
6. (7) Edmonton
Very well could have escaped Molson Stadium with a victory, but they didn't.
7. (8) Hamilton
Got beat at home by a backup quarterback, and their defence is awful.
8. (6) Winnipeg
Got beat at home by a third-string quarterback, and their defence is awful.
Hamilton at Edmonton
The Eskimos showed a little something last week in Montreal, while the battered Tabbies barely have enough players to put a team on the field. Eskimos by 10.
Winnipeg at B.C.
The Leos will be baring their teeth after getting smoked by the Boatmen, while the Bombers are still trying to find their way. Lions by 15.
Calgary has got to beat the Riders next week. If they don't, Saskatchewan wins the head-to-head series in only seven weeks. If that happens, the Riders and Toronto might have the division championships wrapped up before October … Lyle Bauer wants back into the CFL and Winnipeg is his choice destination. The Stampeders let him go after being bought by the Flames, and he has deep roots in Manitoba … Other than the Argos, I don't think another East team will win more than six games. Maybe seven … As for Ottawa's quarterback situation, the Argos have two young kids (Collaros and Trevor Harris). (Thomas) DeMarco in B.C. could be a dark horse, and (Drew) Willy because he's done well … Ottawa will have Henry Burris and/or Kevin Glenn with a couple of other guys (Tate/Willy). They have to win games out of the chute — hence Henry or Kevin … Montreal barely beat the Eskimos. They were lucky to hold on at the end. Have you seen their schedule? The next six games, they play Toronto three times, Saskatchewan twice and B.C. one time. Montreal is 1-4 right now. It's going to get worse over there … Based on last year, B.C.'s defensive line is underachieving so far … Edmonton needs a big back to help that offensive line they've got. If Calvin McCarty gets hurt, they are screwed. (Hugh) Charles and (John) White are quick-footed little guys. Good with the football, but they can't block. The Sedin brothers are more physical … Ottawa is going to be good pretty fast. Not like last time. In that dispersal draft, Ottawa only got Dan Crowley, who was terrible. Tillman found a good one in Joseph, but then the Gliebermans bought the team and Paopao had no chance. Everyone is saying the new Ottawa owners are great. Don't know them, but hear they are real good. Ottawa will get a couple of real good quarterbacks this time. Tate or that Bo Levi Mitchell. Willy or Collaros. They might be better than Winnipeg and Hamilton their first year. Maybe Montreal, too … The Bombers have not reached out for trades at all. They're saying they plan to stick with their own and sign off the neg list after NFL cuts begin … Ottawa needs a quarterback with experience in the new offensive system. Think that will go into who they hire as head coach. It might help (Dave) Dickenson or (George) Cortez get that job if they are going to pick quarterbacks from Calgary and Saskatchewan … Tom Higgins used to be one of us. He's not a coach like us anymore. He protects them (the officials) and doesn't help us.


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Written on: 15/08/2011 by Woltato (1 review written)
How on earth did Pinnacle deem this piece of software ready for release. A lot of the key features don't work properly and it constantly crashes.
Stablilty is a major issue. Sometimes when it crashes you'll get the blue screen of death and have to completely restart your computer. Other times just the studio application crashes and this launches the pinnacle error reporting system which expects you to sit and wait for about 10 minutes while it's "gathering system information" then wants you to write an essay about what causes the crash. Doubt anyone could be bothered using this after the first couple of crashes. The absolute worst thing is that sometimes it will crash when you click on "Save project" At this point it completely wipes out the project and any previously saved work.
For some reason when Studio is open it takes over and hogs the computers audio driver. There is no sound in any other application while Pinnacle is running.
Creating a movie from a finished project doesn't work properly. After carefully editing frame by frame in edit mode I discover that the created movie doesn't match what I created in edit mode. there seems to be black frames inserted inbetween the clips that aren't there in edit mode. Also it often seem to cut a second or to off the beginning of the video.
These are the major issues but there are literally hundreds of little bugs and glitches too numerous to list them all.
This is an absolutely terrible piece of software. Wouldn't recommend it even if it were free.
" Excellent Transfer Service " Read More
" Best video converter I've used " Read More
" VSO? Or, Avangate? " Read More
" Best Youtube Video Downloader " Read More
Written by davidbarel
" Clever Piece of Kit " Read More
Written by dalbeattie


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tRUMPETER 1/200 BISMARK r.c fit
To moderators, if you feel this is in the wrong thread please feel free to move.
Project To design and construct a r/c system to fit in to the new 1/200 Trumpeter Bismark kit.
First buy the kit and look in the box, everthing that has been said about the kit is true, and more
we will update the thread as we progress,even when we get it wrong ?
Deans Marine
Include convenient turret animation, and you'll have a real winner!
1/200 bismark
Saturday evening, what a way to get away from work, build a model boat ?
up date, trial fit deck and mark in hull access areas,
First assembly and trials will be with twin screws, independant motors to test handling, will trial fit 3rd shaft later
Open out the proptube exits with a file
Trial fit the tubes in place
check equal angle on both tubes
Drill our the rudder tubes
Turn up 2mm bearings and push fit into drilled rudder tubes.
Assemble twin tiller arms and locking wires.
Trial fit in to rudder tubes.
Drill centre of rudders
Fit rudders on to shafts with epoxy
ensure all is central and square, wait for epoxy to dry.
Open bottle of red wine and go and relax
Deans Marine
the way to insert straight shafts to rudders and glue with epoxy is not going to last.. I'd say it will free turn in just anout one or two runs..
I you insist to use the original rudder then one way is to drill a 1mm hole on the rudder surface right into the shart and insert a 1mm steel/brass rod ... nevertheless, the original plastic rudder is not meant for R/C running for such a boat size...
Todays progress
Today project, how to spend a sunday
Drill out the A frames 2.75 mm
Fit a 2.1mm ID brass bearing in to the frames
Trial fit in to the hull with pattermakers wax
Fit cast brass 25mm 2mm 162 type warship props to the shaft with
lock nut and nylon washers
Fit 2mm shafts in to the hull with wax, and lose fit A frames,
in to hull to ensure they line up and run smooth
(will add oiling tubes later if run is a success)
Make a motor mount from hardwood and vacform in 1.5mm Hi Impact styrene
Sand to fit in to the hull bottom and line motor up with shafts
Trial fit motor in mount and temp hold in place with
D /S carpet tape for the test run. was going to use 108s to ease lining
up problems but have concerns will not have enough tourque for a heavy
boat with small props
Fit 2mm / 2.3 MM EXL couplings and lubricate with vaseline.
Repeat with opposite side.
Temp fit rudder servo in place with servo tape for the test run.
Make pattern for a battery boxe and vacform x 2 in Hi Impact plastic.
Wire up and fit 2 x 3800 ah Nimh packs
Test in pond, it takes 3.8Kgs lead ballast to bring to bottom of waterline,
this is some very heavy boat.
Fit 4 channel radio with 2 x 10amp ESC and rudder servo, bench test all.
Lightly lubricte the bearings with teflon so none gets on the boat,
Put batterys on charge and wait for the day to warm up and off to the
lake this afternoon.
Have coffee with shot of scotch ready for anti freeze purpose only.
I don't know if we get to help with the design or not (your build here is great marketing regardless) but here is some unsolicited thoughts and pictures.
Why not vacu-mold the motors, battery trays, and radio box as one piece instead of separate? (example below)
Detailed metal rudders and R/C components, maybe larger rudders if needed.
Simple turrent animation (as previously mentioned).
Coffee anti-freeze! Awesome.
Follow up from the test run
It was a lovely day but so cold, had to break the ice on the water first.
The long prop tubes need some support on the inboard end as they vibrated when under power, so a quick fix was made with some plastic strips.
It is now back to the work bench, The trials were not a great success, this is a very heavy ship and the props do not deliver the power to drive her at acceptable speed, looks good on the water, but she has no reserves of torque, so in any breeze she would struggle a bit.
Differential steering was slow, but again in any breeze she would be a hand full, of more concern to me was that she had no "brakes" at all stopping distance we 3-4 boat lengths and this would not be safe.
The 25mm 1292 type brass props have a pitch of 25.6mm, I feel that the cure will be increase the prop to 30mm, but the problem will be that a prop this size will have a M4 shaft size.
That will not be acceptable as the 4mm shaft dia will not fit in the A frames or the exits in the hull. I will try making up a stainless steel shaft on 2mm with an adaptor on the prop end to convert to M4.
I may also try to fit a prop tube with roller bearings at the inboard end as a experiment while I am waiting for the shafts to be produced, and play around with drive only on the centre engine so see how she handles.
Apart from the power problems she ran very well and looked most impressive on the water. With a third engine fitted this will give a bit more power and the twin rudders will be more effective as the will be in the wash of the centre engine, but this will make the design more complex and expensive with the separate drive to the centre engine and this is not the brief we have from the customer.
As an aside the "anti freeze" coffee was the best taste ever, it was VERY cold on Sunday, and when you drop the little allen key in the water, you know it will be hard to find when you roll up your sleeve have to start rooting around in the bottom of the pond.
More to follow when the new parts arrive, and I get some time in the workshop
I may make the mounts in one piece later, but we have tried out three different motors up to date, two couplings, and now shafts, seperate mounts are sensible at this early stage, also we may cast a scale rudder, or a oversize one, but then we may need to increase the size, this is why we have trials and test run first. The shafts will be more the man enough for this boat, we have runn 2mm shafts in far larger and heavier boats before, as long as they are well supported with good bearing.
Deans Marine
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"Jintsuriki" is the character theme for Bankotsubo throughout the series (except for the arcade version of Dead or Alive 2) and the final boss theme for the console versions of Dead or Alive 2. It is also the ending theme for Ayane in Dead or Alive 2.
- 神通力 (Jintsūriki) literally means either "supernatural [an occult] power", "divine power" or "magical power".
See alsoEdit


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Category Archives:New York
Update on New York's Driver License Suspension for the Failure to Pay Back Taxes
In August 2013, I wrote a blog regarding New York's new enforcement procedure to collect state back taxes by suspending drivers' licenses. The link to my blog post is: https://www.irsproblemsolve.com/2013/08/new-york-back-tax-penalty/. Below discusses a recently published case regarding the license suspension program. The case illustrates how simple it is for NY to suspend a driver's license for the failure to pay back taxes.
In order to suspend a New York drivers license for the failure to pay taxes, Tax Law §171-v [3], requires that the taxpayer owe $10,000 or more of NY back taxes. The statute further requires that NY send notification to the delinquent taxpayer of inclusion into the driver's license suspension program. Upon receipt of the notification, the delinquent taxpayer is provided with 60 days to respond. The taxpayer's response is limited to the following:
- Pay the back taxes or establish a payment plan
- Notify the Division of a tax exemption; or
- Protest the suspension by filing a request for a conciliation conference with the Bureau of Conciliation and Mediation Services (BCMS) or a petition with the Division of Tax Appeals.
Todd S. Unger, Esq. Discusses Offer In Compromise In New York Times
Contrary to popular belief, the IRS wants to work with taxpayers. Owing the IRS money can be intimidating, but you have options. One option when you owe back taxes, is to try to negotiate less than the total amount owed. This is called an offer in compromise, and can be an excellent way for you to catch up financially and move forward with your life.
In some cases, you won't be able to pay all the taxes you owe. If you can prove to the IRS that you do not have the ability to pay your owed taxes within 10 years, the offer in compromise could be worth trying. Tax Attorney, Todd Unger was recently quoted in a New York Times article that dealt with the IRS and its offer in compromise program. Unger stated that "when the IRS accepts an offer it can be the deal of a lifetime for the taxpayer needing assistance."Continue Reading...
Do You Have a NY Back Tax Problem? Resolve it or call a Cab
If you owe back taxes that exceed $10,000 in NY State, your driver's license may be suspended.
The New York State Department of Taxation and Finance is sending 16,000 suspension notices to delinquent taxpayers. Delinquent taxpayers will then have 60 days from the mailing date to work out a payment plan. The failure to act will result in the NY Department of Motor Vehicles sending a warning letter. If the taxpayer fails to work out a tax payment plan, within 15 days from the date of the second letter, their driver's license will be suspended. In order to reinstate their license, full payment or a plan to full pay is necessary.Continue Reading...
IRS Hardship Loans and Distribution Relief for Hurricane Sandy Victims
In order to the alleviate hardships of individuals affected by Hurricane Sandy, the IRS has announced that it will relax procedural and administrative rules that apply to retirement plan loans. The IRS is providing this relief in addition to the postponement of various tax filing and payment deadlines to FEMA-designated areas such as New Jersey, New York, and Connecticut. <!–more–>
Continue Reading...
To provide disaster relief, the IRS is minimizing the amount of red tape that eligible retirement plan participants would undergo in order to access their money. The standard six-month ban on 401(k) and 403(b) contributions that would normally affect employees who take hardship loans will no longer apply.
IRS Provides Tax Relief to Victims of Hurricane Sandy
The IRS announced tax relief to individuals and business affected by Hurricane Sandy.
The IRS tax relief postponed various tax filing and payment deadlines. Hurricane Sandy victims have until Feb. 1, 2013 to file all tax returns and pay any taxes due. Tax payments and tax returns, include but are not limited to, fourth quarter individual estimated tax payment, payroll and excise tax returns and accompanying payments for the third and fourth quarters. Additionally, tax-exempt organizations required to file Form 990 series returns has been postponed.Continue Reading...
IRS Gives Addition Time to Taxpayers and Preparers affected by Hurricane Sandy
On October 31, 2012, the Internal Revenue Service (IRS) announced that taxpayers and tax preparers have been given until November 7 to file returns and accompanying payments. The relief was issued specifically for those in the areas of the Mid-Atlantic and Northeastern United States that were affected by the storm or by Hurricane Sandy. The announced relief is automatic and does not require filing waivers or extensions.Continue Reading...
The IRS May Be a Red Sox Fan...
The U.S. Justice Department is suing the New York Yankees' managing general partner, Hal Steinbrenner, son of George Steinbrenner, over what it calls an "erroneous" tax refund.
According to Bloomberg News, citing the complaint, the refund originated from an audit involving the New York Yankees' parent company for the tax years 2001 and 2002. At the conclusion of the tax audit, the late owner, George Steinbrenner and the IRS settled. The settlement resulted in adjustments to the tax returns of the beneficiaries; Hal Steinbrenner had a 25% interest, of a family trust. Hal Steinbrenner paid taxes in 2008, and then filed an amended 2001 tax return in 2009 seeking a refund because of a $6.8 million net operating loss carried back from 2002.
The IRS is claiming the refund claim was filed late and has sued Steinbrenner to recover $670,493.78.
Refund Claims Must Be Filed Timely, Though Amount of Refund May Be Limited
Generally, speaking the timeliness of a refund claim depends on whether a tax return was filed. If a return was filed, the claim must be filed within the later of: (a) 3 years from the time the return was filed, or (b) 2 years from the time the tax was paid.
Even if a claim is timely filed, the amount of the credit of refund may be limited. In general, the amount cannot exceed the tax paid within the 3 year period immediately preceding the filing of the claim.
Suits By United States for Recovery of Erroneous Refunds Must be Timely.
The recovery of an erroneous refund by suit is allowed only if the suit has commenced within 2 years after the making of such refund unless it appears the refund was induced by fraud or misrepresentation. In that case, the suit may be brought at any time within 5 years from the making of the refund.
The case is U.S. v. Harold Z. Steinbrenner and Christina L. Steinbrenner, 11-02840, U.S. District Court for the Middle District of Florida (Tampa).Continue Reading...
New Jersey and New York Tax Relief for Victims of Hurricanes
Because of Hurricane Irene, President Obama declared the following New York and New Jersey counties a federal disaster area:
Atlantic, Bergen, Burlington, Camden, Cape May, Cumberland, Essex, Gloucester, Hudson, Hunterdon, Mercer, Middlesex, Monmouth, Morris, Ocean, Passaic, Salem, Somerset, Sussex, Union and Warren.
Albany, Clinton, Columbia, Delaware, Dutchess, Essex, Greene, Montgomery, Nassau, Orange, Putnam, Otsego, Rensselaer, Rockland, Saratoga, Schenectady, Schoharie, Suffolk, Sullivan, Ulster, Warren, Washington and Westchester.
Treasury Regulation §301.7508A-1 discusses the postponement of tax-related deadlines because of a federally declared disaster. Taxpayers affected by Hurricane Irene may be eligible for the postponement of time to file returns, pay taxes and other time-sensitive acts.
If you are a New York or New Jersey taxpayer affected by the Hurricane and receive an IRS penalty notice, contact the IRS to request that the IRS abate any interest and any late filing or late payment penalties. On the IRS's website it states that "Penalties or interest will be abated only for taxpayers if an original or extended filing, payment or deposit due date, including an extended filing or payment due date, that falls within the postponement period." The IRS also states that "it is waiving the failure-to-deposit penalties for employment (payroll) and tax deposits due on or after Aug. 27, and on or before Sept. 12, as long as the deposits are made by Sept. 12, 2011."Continue Reading...


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Slingerland BDP Super Gene Krupa Radio King Snare Drum
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From 1957 on Slingerland dropped the words "Radio King" and simply called these snare drums the Super Gene Krupa model. I replaced one missing tension rod washer and of course both heads. Other than that everything else is original. The top head is new coated ambassador and the bottom is a new ambassador snare head. The wrap and all chrome have justed been cleaned and look great. Even the tension rods were cleaned and lightly lubed. The shell is Slingerlands famous solid maple shell with maple reenforcing rings. The BDP wrap is in great shape and the seam is tight. There are NO extra holes. Please look at all pics and ask any questions you have prior to bidding. Shipping in lower 48 states will be a flat $20. International bidders welcome, please email for shipping quote. I will not fake value for customs docs. PayPal accepted.


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Just do a NAND backup(on a SD)and be careful when replacing or leave it to someone.
These may be stupid questions but I'd like to know. When I replace my Wiis disc drive (sadly it seems I need to do that) it won't affect my system memory with all my game saves and everything will it? I backed them all up to a DVD anyway but it'll be a hassle to copy them back (assuming I can or would have to). And secondly, is it possible to upgrade the 512mb to something bigger? Say, 2gb?
Just do a NAND backup(on a SD)and be careful when replacing or leave it to someone.
"The Wind... It is blowing...." -Ganondorf
The Mightest Hero is slain by a single arrow
<--Press Thanks If I've Helped!
I already go my NAND backup, first thing I did after hacking lol. But to clarify, would installing a new disc drive affect the flash memory?
I can't find any clear info on if the memory is located inside the disc drive or somewhere else in the system.
No - swapping dvd drive will not affect your NAND. It's mostly impossible to upgrade 512mb NAND to something bigger. Maybe if you will be ale to resolid memory chip with motherboard but i guess it's to hard to do without having yuour own motherboard factory
If you want to have big nand try to use virtual nand on sd or usb (sneek or uneek).
I'm not even bold enough to try upgrading the memory lol, I was just curios cause I've read the disc drive can be replaced with HDD (which is pretty cool if you don't intend on using your retail games at all anymore).


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I think (and I guess everyone in here agrees) Michael Bay is visually sensitive filmmaker.
I'm interested in cinema grammer, visual & aural image, something like that.
I re-watched The Rock a week ago. It's hardly necessary to say, but there are all around stylish moments. Bay makes even plain moments unforgettable.
Anyway, do you remember any stylish moments in bay's films?
And I wonder, what's memorable aural image in bay's films? I can't remember this.


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Saturday's letter was typical of the misinformation out there about efforts to remove Obamacare ("The real death panels," May 18). As a fellow senior, I can clearly see that our Medicare needs reworking. Just as Obamacare is clearly unsustainable without unreasonable taxation, Medicare needs adjustment to continue to help our nation's seniors. Fixing a problem with Medicare is much different than having our government handling our medical needs and deciding if a medical procedure is needed. My experience has been that Medicare allows the doctors to make the decisions.4 comments on this story
The Congress did not vote to "end Medicare," nor did they "vote for my death." That statement is untrue and causes fear. These are people with the good of the country at heart, having families themselves who will be or are on Medicare. They want a solution where Medicare will continue responsibly. In addition, it's becoming more obvious that Obamacare will cause a lot of problems, be extremely more expensive than ever dreamed (Medicare rates have already climbed) and will ruin some individuals' currently working insurance plans. I just wish enough of our senators had the foresight and strength to also vote to end this program before it starts causing more chaos.
Kathy Boyer


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Results 1 to 5 of 7
May 21st, 2013 12:22 PM #1
Thinking of the People of Oklahoma
Hi my fellow Berries, I've been watching the terrible devastation in the US state of Oklahoma with a heavy heart. I just wanted to post to say that the victims and their families are in my thoughts and prayers at this difficult time. I hope that any Nameberries who live in this area or who know people in Oklahoma are safe and sound.All the best,
May 21st, 2013 01:08 PM #3
Yes it is very sad and terrifying. My prayers go out to them.
May 21st, 2013 01:53 PM #5
I'm glad to hear from you, Rollo! I've been thinking of you and keeping you in my daily prayers! Keep strong!All the best,
May 21st, 2013 02:44 PM #7
Thank you Mischa!
I cannot believe the utter destruction or fully grasp the scope of the disaster that occurred.
What people need right now is money and blood. If you are able to give either please assist (the Red Cross has made it very easy to text a $10 donation). The Salvation Army has a text donation option too. Assistance and prayers are greatly appreciated.
Last edited by poppy528; December 31st, 2013 at 03:17 PM. Reason: partial post redaction#TWINMOM #BOYMOM
"And remember, my sentimental friend, that a heart is not judged by how much you love, but by how much you are loved by others." L. Frank Baum, The Wizard of Oz
May 21st, 2013 02:51 PM #9


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First look: iStora 4000 from Breece Hill
The iStora 4000 offers idiot-proof disk-to-disk-to-tape backup that smaller customers can afford.
This Article Covers
Backup tools
Integration is the key to D2D2T
The iStora 4000, a disk-to-disk-to-tape (D2D2T) appliance from Breece Hill, Louisville, Colo., automates backup to disk and then to tape, while the integrated data management software can replicate, migrate and purge data based on policies.
SearchStorage writers evaluate backup's role in ransomware detection
WannaCry, NotPetya, the list of dangerous ransomware viruses goes on and on, well beyond the most highly-publicized attacks. Use this guide to learn about the extent of the threat posed by ransomware, and bolster protection of your data, apps, and systems by using the backup strategies detailed within.
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The 4U appliance consists of a server with a 2.4 GHz Intel CPU with 512 MB memory (expandable to 4 GB) running Windows XP Pro or Windows Server 2003, Gigabit and 10/100 NICs, six 250GB Serial ATA (SATA) disks and the capacity for one to 10 tapes. The product is shipping with LTO-1 or LTO-2 support, but will soon support SDLT-600, Sony SAIT and VS160. For companies with compliance requirements, there's a write once, ready many (WORM) media option for unalterable tape and optical storage.
W. Curtis Preston, vice president of service development, GlassHouse Technologies, says he likes the self-contained design of this integrated D2D2T product, and adds that it "provides a great value for smaller customer and branch offices."
The appliance includes iStora software that adds policy-based replication, migration, purging and file management. With client agents, iStora can back up to disk within the iStora 4000, then migrate to tape.
For users with existing backup applications like those from Veritas and Legato, the iStora 4000 can integrate by first configuring part of its available disk (or other network storage) as a standard Windows share. The share is then identified as the backup destination for the backup-to-disk functionality in the existing backup application. The data backed up to this share can be further managed by the iStora software to migrate it to secondary disk or to tape or both, freeing up the resources on the share for future backups.
The value of the iStora 4000, for Scott Leif, president of Globalstor Data Corp., Chatsworth, CA, a storage reseller, is that "Breece Hill has removed the complexity of integrating disk, tape and backup software ... the product is virtually idiot-proof."
The iStora software allows organizations of varying sizes to groom their data retention to the appropriate balance of online, nearline and offline storage.
For example, a company can establish policies for automating client backup to the iStora 4000 disk. If the company requires 30 days of online backups, it can set a migration policy on the iStora to move data from disk to tape every 30 days before purging it from disk. It could also allow the data to remain on disk for a longer period of time before purging.
While the iStora 4000 is positioned for smaller shops, it may potentially be used in larger companies. But there are a few caveats: The system board and processor are single points of failure, and its lack of tape emulation could restrict its entry into the enterprise.
About the author: Lawrence Didsbury is the managing director of Channel Play Corporation, a marketing consultancy focused on storage and computing technologies.
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Message:So was the time he walked into the Blimpie sub shop (sauced up, of course) and walked out with the huge potted tree right in front of all the employees and customers. Tree lived in his apartment for the year and was returned with aplogizes after graduation. The Blimpie people were humored by his absurd activity.
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A FORMER general secretary of the Scottish Secondary Teachers' Association has died following a long illness.
David Eaglesham, 63, who held the post for 12 years from 1996 to 2008, was described as an"outstanding" general secretary who had made a "huge contribution" to the organisation.
Alan McKenzie, the SSTA's acting general secretary, said: "David brought to the post a sense of tirelessness and, sometimes in the face of much opposition, a determination to change the SSTA and increase its profile nationally.
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"Those of us who worked with him remember his huge contribution. We are all deeply saddened by his death following a long illness".
Tributes to Mr Eaglesham, a former principal teacher of modern studies at Cathkin High, Cambuslang, were led by Cabinet Secretary for Education Michael Russell.
He said: "I was saddened to learn of the passing of David Eaglesham, with whom I worked closely when I was in opposition, and whose advice I valued greatly, as did his union members.
"I would offer my condolences to his family and friends. He will be greatly missed. David was committed to serving his fellow teachers through his work with the SSTA and his interests more broadly in Scottish education."
And Larry Flanagan, general secretary of the Educational Institute of Scotland (EIS) teaching union, added: "Everyone associated with the EIS was saddened to hear of the death of former SSTA general secretary David Eaglesham and everyone at the EIS offers condolences to his family, friends and colleagues.
"David was committed to the work of his association, as well as to the cause of Scottish education more generally.
"The EIS has always enjoyed a positive working relationship, both with David personally as well as with the SSTA on issues of mutual interest, through his term as general secretary and beyond."


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Bio & Incident Details
Age: 32
Tour: 13 years
Badge # 137
Cause: Automobile crash
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Sergeant Bobby Crapse was killed when his patrol vehicle was struck head-on by a wrong way driver on I-95, near mile mark 97 in Chatham County, at approximately 2:15 am. The subject was driving without lights.
He had just completed a traffic control assignment and was driving back to the north part of Bryan County when the crash occurred. Sergeant Crapse's canine partner sustained very minor injuries in the crash.
Sergeant Crapse had served with the Bryan County Sheriff's Office for 13 years and was posthumously promoted to Sergeant. He is survived by his wife and three children.


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Picture: Renewable Energy Learning Centre our newest addition to the CSI
Lakeland College's Centre for Sustainable Innovation encompasses research in three of the College's four areas of strategic priority and academic excellence: Agriculture, Environmental Sciences, and Energy. These areas are supported by significant strengths in construction, manufacturing, and business and are approached with an interdisciplinary mindset. The College's fourth priority area is Fire and Emergency Medicine.
On May 31, 2012 the College celebrated the opening of the Centre for Sustainable Innovation and the key pieces of infrastructure that form the backbone of the applied research activities. The dedicated land base and facilities will be used for the College's Natural Sciences and Engineering Research Council of Canada (NSERC) College and Community Innovation project focused on integrating multiple renewable energy and building science technologies together to identify best practical applications of these technologies in Western Canada. This research on distributed energy generation (wind, solar, solar thermal, geothermal, cogeneration) and storage has significant potential for our rural region.
The Renewable Energy Centre and the Research Fabrication Shop are the first stops on what we hope will become a Centre showcasing a multitude of technologies. We are looking forward to growing our relationships relating to bioenergy and advancing Alberta's bioeconomy and we have already made great progress with our partners at Alberta Innovates Technology Futures, Alberta Innovates Bio Solutions, Western Economic Diversification and the Canada Foundation for Innovation.
We look forward to inviting you back to explore and learn with us as we grow the Centre.
Read the press release on the grand opening of the Centre for Sustainable Innovation.


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An analysis of satellite images released Tuesday shows North Korea has ramped up work at its nuclear test site over the last month.
The news comes a day after the top US envoy for North Korea, Glyn Davies, warned Pyongyang that an atomic test would result in "swift and sure" punishment, the Associated Press reported.
International concern has risen that North Korea may follow a failed April 13 long-range rocket test with its third nuclear test — a pattern that happened in 2006 and 2009 — according to the LA Times.
The Unha-3 rocket flew for less than two minutes before breaking up over the Yellow Sea west of South Korea.
The US believes the rocket was a test of long-range missile-firing capability.
Meanwhile, satellite images taken by DigitalGlobe and GeoEye over the past month show heightened activity at the Punggye-ri nuclear test site in North Korea's northeast, the AP reported in a separate article Tuesday.
South Korean officials said last month that satellite images showed North Korea was digging a new tunnel in what appeared to be preparation for another nuclear test at the site.
Existing tunnels probably caved in and became contaminated with radioactive material after previous tests, the intelligence officers reportedly said.
The US-Korea Institute at Johns Hopkins School of Advanced International Studies said the satellite images of North Korea's Tonghae Satellite Launching Ground "strongly suggest that this new pad is designed to launch rockets larger than the recently tested Unha, either more capable, liquid-fueled space launch vehicles or missiles with intercontinental ranges," according to Reuters.
According to Reuters, experts say the reclusive country already has enough fissile material for at least six nuclear bombs.
More from GlobalPost: North Korea resumes work on nuclear reactor
Davies, meantime, said Monday: "It is very important that North Korea not miscalculate again and engage in any future provocation."
Davies described it as "an uncertain period with North Korea" after the rocket launch and the leadership handover to Kim Jong Un, the young son of Kim Jong Il, who has vowed to place top priority on North Korea's military.
Davies' comments came after he met in Seoul with representatives from the other nations of the stalled six-party talks on North Korea's nuclear programs.
He plans to travel to China and Japan later this week.
More from GlobalPost: UN blacklists North Korea firms over rocket launch


text: Hamilton is a major centre located in the south-west of Victoria, south of Horsham and north of the coastal towns of Portland and Port Fairy. Hamilton, both the geographical and business hub of the region has been the centre of a wealthy pastoral industry for over 160 years. It has grown to become a thriving city, exhibiting style and quality, with excellent shopping, diverse architecture, an outstanding art gallery and stunning botanic gardens.


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The barrier function of the human mammary gland collapses if challenged with cationic drugs, causing their accumulation in milk. However, underlying molecular mechanisms are not well understood. To gain insight into the mechanism, we characterized transport of organic cations in the MCF12A human mammary gland epithelial cells, using carnitine and tetraethylammonium (TEA) as representative nutrient and xenobiotics probes, respectively. Our results show that the mammary gland cells express mRNA and proteins of human (h) novel organic cation transporters (OCTN) 1 and hOCTN2 (a Na+-dependent carnitine carrier with Na+-independent xenobiotics transport function), which belong to the solute carrier superfamily (SLC) of transporters. Other SLC OCTs such as hOCT1 and extraneuronal monoamine transporter (EMT)/hOCT3 are also expressed at mRNA levels, but hOCT2 was undetectable. We further showed mRNA expression of ATB0+ (an amino acid transporter with a Na+/Cl−-dependent carnitine transport activity), and Fly-like putative transporter 2/OCT6 (a splice variant of carnitine transporter 2: a testis-specific Na+-dependent carnitine transporter). TEA uptake was pH dependent. Carnitine uptake was dependent on Na+, and partly on Cl−, compatible with hOCTN2 and ATB0+ function. Modeling analyses predicted multiplicity of the uptake mechanisms with the high-affinity systems characterized by Km of 5.1 μM for carnitine and 1.6 mM for TEA, apparently similar to the reported hOCTN2 parameter for carnitine, and that of EMT/hOCT3 for TEA. Verapamil, cimetidine, carbamazepine, quinidine, and desipramine inhibited the carnitine uptake but required supratherapeutic concentrations, suggesting robustness of the carnitine uptake systems against xenobiotic challenge. Our findings suggest functional roles of a network of multiple SLC organic cation/nutrient transporters in human mammary gland drug transfer.
- organic cation transporter
- milk
- human
- novel organic cation transporter 2
- amino acid transporter B0+
many mothers choose breastfeeding, which is the best feeding method for neonates. However, the majority of these women receive medications in the postpartum period (22). As a result, newborns have increasing likelihood of being breastfed by mothers on medications.
The mammary gland is regarded not only as a milk-processing organ but also as a de facto protective barrier for the infant, enhancing accumulation of nutrients and endogenous bioactive factors in milk, while limiting transfer of nonnutritional, potentially toxic substances. Indeed, in contrast to excreta such as urine, xenobiotic/toxicant concentrations in milk are usually lower than those of maternal serum (16). However, this barrier function breaks down if challenged with certain compounds of cationic nature, which causes their accumulation in milk.
Cationic drugs constitute an important group of drugs that tend to be concentrated in milk in relation to maternal plasma levels, causing sometimes toxicity in breastfed infants (21 and 31; reviewed in Ref. 18). Accumulation of cationic drugs in milk was believed to be mainly the result of ion trapping as a result of relative acidity of milk compared with maternal plasma. A concept behind this is the long-held view that drugs are excreted in milk almost exclusively by passive diffusion. However, McNamara et al. (23, 24) showed that excretion of organic cations such as cimetidine and ranitidine (24) in milk of rats is more than expected from simple diffusion and ion trapping. An in vivo human study by the same group has also shown that accumulation of cimetidine in milk is significantly higher than expected from passive diffusion (29). However, the molecular mechanisms are not fully understood.
Recently, the breast cancer resistance protein BCRP (ABCG2), which belongs to the ATP-binding cassette transporter family, has been identified as a mammary drug/toxin transporter (19). Namely, the Abcg2−/− mice showed significantly reduced milk excretion of certain drugs and toxins such as nitrofurantoin, cimetidine, acyclovir, and topotecan (19, 25). However, roles of other drug-transporting proteins for a wide range of cationic compounds remain to be elucidated in the human mammary gland.
Organic cation transporters (OCTs) are essential in eliminating nonnutrient, often toxic, cationic compounds that are ingested as food or drug. These proteins are classified as the solute carrier superfamily (SLC). Grundemann et al. (12) cloned the first member of the OCT family from the rat kidney. In humans, two homologous transporters, human (h) OCT1 (SLC22A1) and OCT2 (SLC22A2), were the first OCTs cloned (11, 44). OCT3 (SLC22A3) was then identified from a rat placental cDNA library (20). Its human ortholog was cloned as an extraneuronal catecholamine transporter (13) and was shown to be identical to the extraneuronal monoamine transporter (uptake2) or EMT (41). Subsequently, the following two distinct OCTs were characterized and coined novel OCTs: human OCTN1 (SLC22A4; see Ref. 34) and OCTN2 (SLC22A5; see Refs. 35 and 40). OCTN1 functions as a pH-dependent proton/cation antiporter (43) and is strongly expressed in tissues such as kidney, trachea, and bone marrow.
OCTN2 is a unique transporter with a dual mode of transport (one as a Na+-independent OCT and the other as a Na+-dependent, high-affinity carnitine transporter). Human milk is rich in carnitine (30), which is essential in the β-oxidation of long-chain fatty acids. Neonates are developmentally immature for carnitine biosynthesis, depending on breast milk as an exogenous source of carnitine. Moreover, a congenital defect of hOCTN2 causes a lethal metabolic condition known as primary systemic carnitine deficiency (27). This implies an important role of carnitine transporters in the human mammary gland.
In accordance with the in vivo data suggesting carrier-mediated excretion of cationic drugs in milk, mammary gland mRNA expressions of the SLC OCTs have been reported recently in rats (9) and humans (1). However, the function of these gene products in the human mammary gland remains to be demonstrated. Moreover, it is intriguing that the mammary gland expresses xenobiotic transporters of detoxification function, including BCRP, because the notion apparently contradicts the nutritional role that the mammary gland plays for the offspring. It is possible that nutrient carriers such as OCTN2 are hijacked by potentially toxic cationic xenobiotics, causing collapse of the barrier function of the mammary gland epithelia.
To gain insight into an integrated mechanism of drug excretion in the human mammary gland, our objective in the present study was to demonstrate and characterize carrier-mediated transport of organic cations in the MCF12A human mammary gland epithelial cells, which are widely used as an in vitro system of noncancerous human mammary gland epithelia. In particular, we asked questions whether OCTN2 as a representative SLC transporter of dual roles [i.e., transport of a nutrient (l-carnitine) and cationic xenobiotics] is expressed and functioning in the human mammary gland and, if so, whether therapeutic compounds interfere with its function. Our data indicate mammary expression of multiple SLC OCTs, including OCTN2, and corresponding functional multiplicity for uptake of carnitine and tetraethylammonium (TEA). Overall, carnitine transport in the MCF12A cells is a robust process, requiring supratherapeutic concentrations of cationic drugs for its inhibition. In addition to the high-affinity carnitine uptake compatible with OCTN2 function, partial Cl− dependency of the uptake suggests a functional role of a Na+/Cl−-dependent low-affinity carnitine transporter, human amino acid transporter B0+ (ATB0+; see Refs. 26 and 33). In agreement with the notion, the MCF12A cells show specific uptake of glycine, an ATB0+ substrate, which is inhibited by carnitine and cationic drugs such as verapamil and desipramine.
Cell culture.
The MCF12A human mammary gland epithelial cell line was obtained from American Type Culture Collection (Rockville, MD) at a passage 53. They were cultured, as specified by ATCC, in a 1:1 mixture of Ham's F-12 medium and DMEM with 5% horse serum, supplemented with 2 mM glutamine, 0.1 μg/ml cholera enterotoxin, 10 μg/ml insulin, 0.5 μg/ml hydrocortisone, and 20 ng/ml epidermal growth factor. The cells were maintained under an atmosphere of 95% air-5% CO2 at 37°C and subcultured weekly using 0.02% EDTA and 0.05% trypsin. Media were changed every 2–3 days. For the uptake experiments, the MCF12A cells were used in the passages 56–60.
A human mammary myoepithelial cell line (HMEC) was obtained at the passage 7 from Clonetics (San Diego, CA). The cells were maintained in the Mammary Epithelial Cell Basal Medium (Clonetics) supplemented with 52 μg/ml bovine pituitary extract, 10 ng/ml human recombinant epidermal growth factor, 5 μg/ml insulin, 0.5 μg/ml hydrocortisone, 50 μg/ml gentamicin, and 50 ng/ml amphotercin B. The cell line was maintained under an atmosphere of 95% air-5% CO2 at 37°C and subcultured two times weekly using 0.02% EDTA and 0.05% trypsin. The HMEC cells were used at the passage 10.
Human tissue preparation.
Discarded mammary gland tissue was obtained from women of child-bearing age undergoing cosmetic breast reduction operation after the consent form was obtained. The study protocol was approved by the respective Institutional Research Boards. We processed the tissue as previously described (38). Briefly, the tissue was dissected, immediately after surgery; excess adipose tissue was removed, and the parenchyma containing the ductal-lobular-alveolar structures was isolated and processed for total RNA extraction. The remaining tissue was snap-frozen in liquid nitrogen and stored at −80°C until used for protein detection. Mammary tissue used for immunohistochemistry was fixed with 10% formalin in PBS immediately after surgery, subsequently embedded in paraffin, and sectioned to ∼5 μm thickness. Human kidney tissue samples were provided by the Cooperative Human Tissue Network, which is funded by the National Cancer Institute.
Total RNA was extracted from cultured cells (e.g., MCF12A cells and HMEC cells), dissected glandular tissue of the mammary gland, and other tissue samples, using the TRIzol reagent (GIBCO-BRL, Grand Island, NY), and cDNA was synthesized by reverse transcription using Moloney murine leukemia virus RT. For RNA extraction from the mammary tissue, homogenized tissue was centrifuged at 12,000 g for 10 min at 4°C. The supernatant was then processed for total RNA extraction as described above. Purity and integrity of the RNA samples were verified using an ultraviolet spectrophotometer and agarose gel visualization of ribosomal bands, respectively. The respective transcripts were then amplified using the primers. For OCT1 and OCT2, we used the primers reported by Gorboulev et al. (11). We designed specific primers for hOCTN1 (forward: 5′-CTG GAT GCT CCT AAT TTA CAT GG-3′; reverse: 5′-AGG AGA CTC TCT AGA AAT GGT TGG-3′), hOCTN2 (forward: 5′-GTG CTG TTG GGC TCC TTC ATT TA-3′; reverse: 5′-AGC TGC ATG AAG AGA AGG ACA CTG-3′), EMT/hOCT3 (forward: 5′-ACT CCA CCA TCG TCA GCG AG 3′; reverse; 5′-CTG GAT TCC TTG CCA GTT GG-3′), and ATB0+ (forward: 5′ -ACA CAT GGC CCA TAT ATC TGG-3′; reverse: 5′ -ATG GAA TTG CGC CAT AAT TAG G-3′). The expected amplification products are as follows: hOCTN1, 785 bp, corresponding to the nucleotide positions 1227–2011 on the hOCTN1 cDNA sequence; hOCTN2, 800 bp, covering the nucleotide positions between 580 and 1379; EMT/hOCT3, 404 bp, covering the nucleotide positions between 437 and 840; and ATB0+, 541 bp, corresponding to the positions 1197–1738. In addition, we designed a specific primer for Fly-like putative transporter (Flipt) 2/carnitine transporter (CT) 2 (forward: 5′-GGA GTC ACC CCT CAT CAT GT-3′; reverse: 5′-CAA AAG TCA CCG ATC CCA GT-3′). It can distinguish CT2 and its splice variant, Flipt2/OCT6, because the primer covers the Flipt2/OCT6-specific sequence, which is an in-frame retention of the third intron of CT2 gene (6, 7, 10). The expected amplification products for Flipt2/OCT6 and CT2 are 385 and 289 bp, respectively. Because the primer does not encompass an intronic sequence of Flipt2/OCT6, we ruled out genomic contamination by performing PCR without using reverse trascriptase.
The identity of each PCR product was verified by sequencing (Center for Applied Genomics, the Hospital for Sick Children, Ontario, Canada).
Immunoblot analyses for human OCTN1 and OCTN2.
Protein was extracted from the mammary tissue, the kidney, and the cell-membrane fraction of the MCF12A cells. Briefly, the mammary gland and the kidney tissues were homogenized in ice-cold RIPA buffer containing phenylmethylsulfonyl fluoride and proteinase inhibitors. After incubation for 1 h on ice, the lysate was centrifuged at 10,000 g for 10 min at 4°C, and the supernatant was collected. For MCF12A cells, cell membrane fractions were made as previously described (2), with some modifications. Briefly, MCF12A cells (50 × 106) were suspended in 10 mM Tris·HCl, pH 7.4, 10 mM KCl, 1.5 mM MgCl2, 200 μg/ml EDTA with protease inhibitors, 25 μg/ml RNase, and 50 μg/ml DNase I. After 10 min of incubation, cells were homogenized using a glass homogenizer with 100 strokes. The homogenate was centrifuged at 800 g for 15 min at 4°C. The supernatant was centrifuged at 100,000 g for 20 min at 4°C. The pellet containing the enriched membrane fraction was then resuspended in 10 mM Tris·HCl, pH 7.6, with 125 mM sucrose and protease inhibitors. Protein concentration was measured by the BCA Protein Assay kit (Pierce) using BSA as a standard.
We used specific, site-directed polyclonal antibodies raised against the COOH termini of human OCTN1 and OCTN2. The amino acid sequences of the antigenic peptides were WFRSGKKTRDSMETEENPK (corresponding to amino acid residues 527–545) for hOCTN1 and HRKTPSHTRMLKDGQERPT (corresponding to amino acid residues 532–550) for hOCTN2 (37).
Protein samples (50–100 μg) were loaded on a 7.5% SDS-polyacrylamide gel, subjected to electrophoresis, and transferred to the nitrocellulose membranes that were then blocked with 4% skim milk overnight at 4°C. The membranes were incubated with the primary antibodies (1:1,000 dilution) for 3 h at room temperature. After thorough rinsing, the membranes were incubated with horseradish peroxidase-conjugated goat anti-mouse antibody (0.4 mg/ml; Santa Cruz Biotechnology) for 1 h at room temperature. The proteins were detected using enhanced chemiluminescence (ECL detection system; Amersham). In addition, to confirm the specificity of the antibodies, corresponding antigenic peptides were incubated in a 5:1 ratio with the primary antibodies at 4°C overnight. The preadsorbed antibodies were then used with the protein-loaded membranes.
Immunohistochemical detection of human OCTN2.
Deparaffinized and rehydrated human breast tissue sections were treated to inactivate endogenous peroxidase and biotin. Slides were washed and incubated for 1 h at room temperature with the rabbit polyclonal antiserum (37) at 1:80 dilution in Tris-buffered saline containing 0.1% Tween 20 (TBST). After being washed in TBST, the slides were incubated with horseradish peroxidase-conjugated goat anti-rabbit IgG (CSA rabbit link; Dako) for 45 min, washed in TBST, and developed for 1–3 min in diaminobenzidine solution and hydrogen peroxide. After the color was developed, the slides were counterstained with Harris Hematoxylin stain. Preimmune rabbit serum was used as controls.
Uptake experiments.
MCF12A cells in the passages between 56 and 60 were grown on six-well tissue culture plates at an initial seeding density of 5 × 105 cells/well until 80–90% confluency in the same culture condition as described above. After the media was removed, the cells were immersed in uptake media. For uptake experiments, the media (pH 7.5) consisted of 25 mM Tris/HEPES, 140 mM NaCl, 5.4 mM KCl, 1.8 mM CaCl2, 0.8 mM MgSO4, and 5 mM glucose (8). After preincubation at 37°C in room air for 60 min, 65 μM [14C]TEA with 25 nM [3H]mannitol or 0.1 μM l-[3H]carnitine with 0.86 μM [14C]mannitol (to allow dual counting at the probe-mannitol disintegrations/min ratio of ∼10:1) were added to the uptake media in the presence (nonspecific uptake) and absence (total uptake) of an excess amount of the unlabeled substrates at various concentrations of inhibitors. One hundred-fold excess of unlabeled carnitine or TEA (up to 10 mM because of solubility) was used to determine the nonspecific uptake for the labeled substrates. The nonspecific uptake values were subtracted from the total uptake values to obtain the specific uptake. At a given incubation time, the incubation medium was removed, and the cells were rinsed two times in ice-cold PBS for 30 s each. The cell membranes were then solubilized using 0.1% Triton X-100 (BDH, Toronto, Ontario), and the radioactivity was counted as disintegrations/min. The cellular protein content was measured using the BCA protein assay kit (Pierce, Rockford, IL) and BSA as the standard. For Na+-free experiments, NaCl was substituted with equimolar LiCl, KCl, or choline chloride. Similarly, for Cl−-free experiments, Cl salts were replaced with equimolar sodium gluconate, sodium thiocyanate, or sodium phosphate. For pH-dependency experiments, the pH was adjusted appropriately using HCl or NaOH (36). [3H]glycine (1 μM) uptake was examined similarly in the presence and absence of various inhibitors using the uptake buffer reported previously (26). All experiments were done in triplicate.
Data analyses.
Data were compared using Student t-test or ANOVA (followed by Tukey's test), where appropriate. In the text, results were expressed as means ± SE of 3–9 independent measurements. In Figs. 1–7, error bars were specifically designated whether they represent SD or SE.
Estimation of kinetic parameters.
Kinetic parameters of carnitine and TEA uptake were determined by fitting the pooled data on the total and nonspecific initial uptake rates at various substrate concentrations to the Michaelis-Menten models. We tested three different models with one, two, or three Michaelis-Menten terms, all of which had a linear term to take into account a nonspecific diffusion/uptake component. The two-system model as a representative example is shown below where suffixes 1 and 2 denote system 1 and 2, respectively, [C] is the substrate concentration, and A represents the nonspecific uptake constant. The experimentally determined nonspecific uptake was also fit to the following equation simultaneously, so that the parameters satisfying both equations at the same time can be estimated
The nonlinear data fitting was performed using MLAB (Civilized Software, Bethesda, MD), as described elsewhere (17). Briefly, after confirming that there is no systematic deviation in data fitting, and that errors for individual parameter estimates are relatively small, the statistical criteria were used for the selection of a best-fit model. The residual sum-of-square error (RSS) was compared between models using the F-test. If there is a statistically significant difference (i.e., P < 0.05), the model with a greater sum of square error was rejected in favor of the other. A best-fit model requires the fewest parameters, while providing a reasonably good fitting that is statistically indistinguishable from that of more complex models. This aspect was assessed by two parsimony criteria; namely, the Akaike's information criterion (AIC) and the Schwartz criterion (SC). In the case where RSSs of two models are not statistically different, a model with lower AIC and SC was selected as a better model.
Insulin, epidermal growth factor, cholera toxin, and amino acids were purchased from Sigma-Aldrich (Oakville, Ontario, Canada). Quinidine was a gift from AstraZeneca. Cell culture-related materials were obtained from GIBCO-BRL. l-[N-methyl-3H]carnitine hydrochloride (80 Ci/mmol) was purchased from American Radiolabeled Chemicals (St. Louis, MO). d-[1-14C]mannitol (58.1 mCi/mmol), [1-14C]TEA bromide (2.4 mCi/mmol), and d-[1-3H(N)]mannitol (26.3 Ci/mmol) were purchased from New England Nuclear Life Science Products (Boston, MA). [2-3H]glycine (60 Ci/mmol) was purchased from Perkin-Elmer (Boston, MA).
mRNA expression of OCTs.
We examined the expression of OCTs in the three representative models of human mammary gland: the mammary gland epithelial cell line (MCF12A), the mammary cell line with myoepithelial cell characteristics (HMEC), and the human mammary gland tissues obtained from women undergoing breast reduction surgery. In all three, we detected PCR products for hOCT1, hOCTN2, hOCTN1, and hOCT3/EMT (Fig. 1A). ATB0+, a Na+/Cl−-dependent amino acid transporter, was also detectable in MCF12A, the breast tissue, and in kidney (Fig. 1B). Sequence analysis further confirmed the identity (data not shown). However, RT-PCR for hOCT2 produced no amplicon (Fig. 1A).
Another high-affinity carnitine transporter (CT2) has been recently reported as the testis-specific protein by Enomoto et al. (6). Interestingly, CT2 mRNA was undetectable in the MCF12A cells and the mammary gland tissues, but its splice variant showed ample expression (Fig. 1C), which was independently reported and designated as Flipt2 by Eraly and Nigam (7) because of its homology to Drosophila putative transporter (Fly-like putative transporter 2), and OCT6 by Gong et al. (10). PCR without RT was negative, indicating no genomic DNA contamination (data not shown).
The affinity-purified OCTN1 and -2 antibodies against the corresponding peptides stained bands with a relative molecular mass of ∼60 kDa (Fig. 2, A-C). The estimated sizes of OCTN1 and -2 are ∼63 kDa, but the detected bands were consistently smaller than 63 kDa, suggesting posttranscriptional modification and/or degradation. Noteworthy is that a recent study on hOCTN2 expression in Caco-2 intestinal cells reported similar results (5).
Immunohistochemistry with hOCTN2 antibody.
To determine mammary tissue localization of hOCTN2, normal human mammary tissues were stained with the specific hOCTN2 antibody (36, 37). In accordance with the data obtained by RT-PCR and Western blotting, staining was evident in the ductular-lobular-alveolar structures in the mammary gland parenchyma, especially in the alveolar glands (Fig. 2D). Specificity of immunostaining was established by observing a lack of staining with the preimmune rabbit serum in the same tissue. These results indicate that OCTN2 expression is mainly localized in the alveolar epithelia of the mammary gland. Immunohistochemistry using the hOCTN1 antibody was unsuccessful.
l-carnitine and TEA uptake by MCF12A.
The time course of l-carnitine uptake (Fig. 3A) indicated a linear phase for the initial 120 min. TEA uptake in MCF12A cells was time dependent, with the initial linear phase of ∼60 min (Fig. 3B). On the basis of these results, subsequent experiments for initial uptake rate of l-carnitine and TEA were conducted at 60 and 30 min, respectively.
l-Carnitine uptake in MCF12A cells was Na+ dependent. When Na+ in the medium was replaced with other monovalent cations, the uptake was inhibited significantly [ANOVA: F = 7.75; degrees of freedom (df) = 6, P < 0.001; Fig. 4A]. Choline exerted the highest inhibition (P < 0.001), followed by potassium (P = 0.006) and lithium (P = 0.018). When Cl− was replaced (Fig. 4A), ∼40% inhibition was observed. However, only sodium phosphate reached statistical significance (P = 0.035), which is likely to be because of lack of power of the sample size. Overall, the results suggest that the carnitine uptake function of the MCF12A cells is at least in part dependent on Na+ and Cl−. TEA uptake was pH dependent (Fig. 4B), increasing by ∼60% as media pH was increased from 6 to 8 (ANOVA: F = 9.96; df = 2, P = 0.012).
Kinetics of organic cation uptake in MCF12A.
To gain insight into the functional roles of these transporters, different Michaelis-Menten models were fit to the saturation data for carnitine and TEA. Because a model with more than three Michaelis-Menten systems did not converge to meaningful estimates, we examined the following two simpler models with a nonspecific diffusion term: a single-system model and a double-system model. None of the fitting results of the two models for either substrate was under active constraints at termination. Although the modeling analyses were successfully employed, it should be noted that experimental constraints did not allow us to increase the substrate concentrations to cover the Km values of the low-affinity systems.
For l-carnitine (Fig. 5A), the single-system model caused systematic deviation from the data points (the runs test; data not shown) and was rejected in favor of the double-system model (r2 = 0.999; RSS = 5.96). The double-system model predicted presence of a high- and a low-affinity system with Km of 5.1 ± 4.0 μM (Vmax = 832 ± 246 pmol·mg protein−1·h−1) and 15.9 ± 6.9 mM (Vmax = 170 ± 59 nmol·mg protein−1·h−1), respectively. Inhibition of 0.1 μM l-[3H]carnitine uptake in MCF12A cells was demonstrated using various cationic compounds (Fig. 5B). Verapamil and l-carnitine itself at a 20–50 μM range significantly inhibited l-carnitine uptake. Other therapeutic drugs such as cimetidine, carbamazepine, quinidine, and desipramine showed >50% inhibition at supratherapeutic concentrations. l-Carnitine transport by ATB0+, a Na+/Cl−-dependent transporter for neutral and cationic amino acids, is known to be inhibited by zwitterionic and cationic amino acids such as alanine, glycine, arginine, and glutamine (26, 33). As shown in Fig. 5C, alanine, glycine, arginine, and glutamine at 1 mM significantly inhibited carnitine uptake (ANOVA: F = 256.2; df = 4, P < 0.001). Coupled with partial Cl− dependency of carnitine uptake (Fig. 4A), this suggests contribution of hATB0+ to carnitine uptake in the MCF12A cells.
For TEA (Fig. 6A), the difference in goodness-of-fit of the two models did not reach statistical significance (F = 6.44, P = 0.056), although the double-system model resulted in substantially lower RSS (0.82 vs. 3.48). Given the statistically indistinguishable fitting performance, the parsimony criteria (AIC and SC) were used to select the best model. AIC and SC of the double-system model (AIC, 8.3; and SC, 9.2) were nearly one-half those of the single-system model (AIC, 17.2; and SC, 17.8); the AIC and SC differences between the two models were about eight, indicating the strong support for the former. Therefore, we considered the double-system model to also be the best model for TEA. This model predicted that the high-affinity system has Km of 1.63 ± 0.51 mM and Vmax of 8.9 ± 2.4 nmol·mg protein−1·0.5 h−1, and that a low-affinity system has Km of 73.6 ± 21.6 mM and Vmax of 139.5 ± 20.0 nmol·mg protein−1·0.5 h−1. The uptake of 65 μM [14C]TEA was inhibited by various drugs (Fig. 6B). Of the therapeutic drugs tested, desipramine and disopyramide were the most potent, inhibiting >50–80% of the uptake at concentrations of 10–100 μM, followed by cimetidine. In contrast to carnitine uptake inhibition, TEA uptake was not inhibited by carnitine at 1–5 mM.
Glycine uptake in MCF12A.
If both hOCTN2 and ATB0+ contribute to the mammary carnitine uptake as our results suggest, it is intriguing that drugs such as verapamil and desipramine exert nearly complete inhibition of carnitine uptake (Fig 5B). This implies that these drugs may interfere not only with hOCTN2 but also with ATB0+. Therefore, we have examined inhibitor specificity of ATB0+ in the MCF12A cells, using glycine as a probe substrate (15, 26). As shown in Fig. 7, 1 μM [3H]glycine uptake was significantly reduced by all inhibitors but aspartic and glutamic acid (ANOVA: df = 13, F = 47.2, P < 0.001; pairwise comparison with no inhibitor without 1 mM unlabeled glycine by Tukey's test, P < 0.001). Unlabeled glycine (1 mM) decreased the uptake of 1 μM [3H]glycine by ∼60%, indicating presence of the specific glycine uptake component in the MCF12A cells, supposedly through ATB0+. In the presence of 1 mM unlabeled glycine, however, carnitine and verapamil did not show significant additional inhibition on 1 μM [3H]glycine uptake, indicating that they inhibit the glycine-specific component (i.e., through ATB0+). Asparagine and glutamine also decreased the 1 μM [3H]glycine uptake (P < 0.001), but, in the presence of 1 mM unlabeled glycine, they further decreased the 1 μM [3H]glycine uptake significantly (P = 0.003 and 0.015, respectively), probably because of their relatively low affinity to human ATB0+ (33) and/or interaction with a non-ATB0+ low-affinity glycine transporter. Consistent with the data on mouse and human ATB0+ expression systems (26, 33), aspartate and glutamate showed virtually no inhibition. Interestingly, 1 mM desipramine showed substantial inhibition, even compared with 1 mM unlabeled glycine.
Our study showed expression of several SLC OCTs, and corresponding multiplicity of carnitine/TEA uptake kinetics in the MCF12A human mammary gland epithelial cells. Although vectoral transport of drugs across a polarized cell layer of the mammary gland epithelia remains to be examined, our data suggest that excretion of cationic drugs in human milk is most likely mediated by a network of multiple drug-transporting proteins.
It should be noted that our model-derived Km values are probably hybrid constants and that further experimental validation of the low-affinity systems was not possible. In addition, phosphorylation status of the transporters such as OCT1 influences substrate affinity (3), making Km comparison difficult between different experimental systems. Nevertheless, estimated kinetic parameters warrant some discussion. Shennan et al. (32) characterized carnitine transport in lactating rat mammary tissues, demonstrating a Na+-dependent component especially in the early lactation stage. Na+-dependent l-carnitine uptake through hOCTN2 is characterized by Km values of 3.5 to 4.8 μM in various expression systems (8, 28, 35, 39). Our estimate of the Km value (5.1 μM) of the high-affinity system for carnitine (Fig. 5) is consistent with these reported values. Together with hOCTN2 protein expression (Fig 2) and the inhibitor specificity (Fig. 5B), our data suggest that Na+-dependent uptake of carnitine in MCF12A cells is mediated at least in part by hOCTN2.
Sloan and Mager (33) discovered ATB0+, a neutral/cationic amino acid transporter, which is expressed in the lung, trachea, salivary gland, and mammary gland. Nakanishi et al. (26) recently reported the Na+/Cl−-dependent carnitine transport mediated by the mouse ATB0+. In accordance with the work by Sloan and Mager (33), we observed human ATB0+ mRNA expression in the mammary gland epithelia (Fig. 1B). Our findings further indicate a Cl−-dependent component and amino acid inhibition of the carnitine uptake, which are compatible with the putative function of human ATB0+.
If both hOCTN2 and ATB0+ play a role in the mammary carnitine transport, nearly complete inhibition of carnitine uptake by verapamil and desipramine (Fig. 5B) merits discussion. Using glycine as a probe substrate, we were able to show that uptake of glycine, an ATB0+ substrate, is significantly inhibited by these drugs (Fig. 7). At present, nitric oxide synthase inhibitors are known to interact with glycine via ATB0+, probably because most nitric oxide synthase inhibitors are structurally related to ATB0+ substrate amino acids (15). Although we do not know if the inhibition by verapamil and desipramine is competitive or not, our data added another dimension to drug-amino acid interaction.
The Km value (1.6 mM) of the high-affinity uptake system we predicted for TEA was substantially higher than those reported for hOCT1 [160–260 μM (4, 45, 46)], hOCTN1 [436 μM (34)] and hOCTN2 [304 μM (28)]. There are conflicting reports as to whether EMT/hOCT3 mediates TEA transport. Grundeman et al. (14) reported that EMT/hOCT3 does not transport TEA. On the other hand, EMT/hOCT3-mediated TEA uptake was reported by Wu et al. (42), as well as rat and mouse OCT3-mediated TEA transport (20, 42). Given the reciprocal inhibition of hOCTN2 by carnitine and TEA (28), the lack of inhibition by 5 mM carnitine on TEA uptake in the mammary gland (Fig. 6B) strongly suggests the presence of a TEA uptake system in the MCF12A cells other than OCT1, OCTN1, or OCTN2. Interestingly, the EMT/OCT3-mediated transport of TEA in those studies provides similar Km values to our estimate (mouse: Km, 1.9 mM; rat: Km, 2.5 mM; and human: Ki, 1.3 mM; see Refs. 20 and 42).
Alcorn et al. (1) reported that hOCTN2 mRNA expression in the epithelial cells dislodged from the mammary gland in milk is substantially lower than that seen in the mammary gland tissues at nonlactating stage. Presently, it is not known if its expression and function are decreased similarly in the plasma membrane of the intact mammary gland epithelia at the lactating stage. It is critical to examine protein expression of various transporters, including OCTN2 in the in vivo lactating human mammary gland epithelia.
The inhibitor specificity assay in this study showed that several therapeutic drugs with cationic nature interact with the carnitine/TEA transporters in the mammary gland. However, their inhibitory constants, especially for the carnitine transport, appear relatively high compared with their therapeutic plasma concentrations, suggesting that the likelihood of in vivo drug-carnitine interaction is remote. Overall, carnitine transport in the mammary gland seems to be a robust process, which is not readily affected by exogenous xenobiotics.
Interestingly, Flipt2/OCT6 (7, 10), a splice variant of CT2 (6), was detectable in the human mammary gland. CT2 is a high-affinity Na+-dependent carnitine transporter with an estimated uptake Km of ∼20 μM in a Xenopus oocyte expression system (6). A carnitine transport parameter has not been reported for Flipt2/OCT6. Further studies are needed to reveal its exact functional role.
In summary, the human mammary gland epithelial cells, MCF12A, express multiple functional SLC transporters, suggesting their contribution to drug excretion in human milk. The MCF12A cells have a potential to be an in vitro mammary model for organic cation transfer in human milk, which circumvents ethically challenging drug studies in breastfeeding mother-infant dyads. To this end, it would be important to determine if the MCF12A cells undergo lactogenesis and highly express other transporters such as BCRP, which is known to be significantly upregulated during lactation.
This work is supported by Canadian Institute of Health Research (CIHR) Grant MT13747. Dr. W. Gao was supported by a fellowship from the Canadian Society for Clinical Pharmacology, and Dr. S. Ito was a recipient of a Career award from the CIHR/Rx&D Health Research Foundation.
We thank Drs. Jacintha Shenton and Herman Yeger and the staff of the Breastfeeding program at the Hospital for Sick Children for advice and support.
Part of this work was presented at the American Society for Clinical Pharmacology and Therapeutics meeting in Orlando, FL, March 6–10, 2001.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
- Copyright © 2006 the American Physiological Society


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Eminent Domain Lawyers and Law Firms in Norwalk, CT
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Harris, Harris & Schmid
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Dear Editor:
I have prepared this letter in support of the reelection of Ken Fox to the LPEA Board of Directors. During his tenure, Mr. Fox has been an active participant in board activities and has always been ready to engage any of his neighbors and respond to questions about the utility. In an era when many electrical utilities are headlong adopting costly sources of alternative energy, Ken has always taken a balanced approach to exploiting proven sources of renewable energy while being ever mindful of maintaining economical rates for his neighbors. After almost seven years of economic hardship here in Archuleta County, the last thing we need is anyone who wants to crusade for green energy at the cost of our community's economic recovery.
Hence, I would like to urge my friends and neighbors to think about Ken's record and his vision for the future when you cast your LPEA ballot.
H. Pat Artis
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How to smooth out year-end tax planning
Though the year is almost over, there are still new ways to smooth out the 2016 tax planning process.
For starters, says Robert Fishbein, a vice president and corporate counsel in Prudential's Tax Department, taxpayers can plan for 2016 and 2017 together to reduce their overall tax liability by shifting income and deductions across the two years. One way is to pay January mortgage payments now to make the interest deductible this year. Another is to accelerate charitable deductions or medical expenses into 2016. Those over age 70½ can consider making charitable gifts using required minimum distribution funds, up to $100,000 a year.
"It's usually better than an outright gift to charity because the deduction would probably not offset the income from the required minimum distributions due to limitations on deductions," Fishbein says.
He also notes if taxpayers have unused capital losses from prior years they can sell investments at a gain to make those losses work. "You won't be taxed on those gains, up to the amount of the losses," he explains.
Another tax strategy wrinkle? Converting a traditional IRA to a Roth IRA. Although the conversion would trigger a gain, says Fishbein, there are numerous benefits from holding Roth IRA assets over the longer term—from hedging against increased tax rates to growing without forced distributions. And distributions, he says, "won't increase your taxable income for purposes of the 3.8 percent Medicare surcharge, calculating the taxable amount of your Social Security benefits or determining the Medicare Part B premium surcharge."
Interested in learning more about year-end tax planning strategies? Read Robert's opinion piece. Want to speak to Robert? Contact Lisa Bennett.
Neither Prudential Financial, its affiliates, nor its financial professionals, renders tax or legal advice. Please consult with an attorney, accountant, and/or tax advisor for advice concerning your particular circumstances.