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DRAFT GDC Bylaw Changes
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US Congressional Record Daily Digest 17 October 2005
Industrial Worker - April 2009
Industrial Worker - Issue #1775, June 2015
NLCCC Community Kick Off Final
ASG Senate Resolution No. 1 The ASG Senate Standing Rules Act of 2010-2011
Rotation Note for Consultation
BOCW Act.pptx
Redistricting Janice Dysinger Testimony
Makabayan Bloc's petition on TRAIN
Act 556, Lembaga Akreditasi Negara Act 1996
Week of May 23
Constitutional Law 1 Reviewer Finals Back Up File
Eric Steltzer Slides 6.7.11
Muncipal Council Matters Under Jurisdiction
Letter to the Honorable Dave Camp Regarding CBO's Report Titled Options for Taxing U.S. Multinational Corporations
The 1987 Constitution
Spe-Anuc Chapter Constitution.
Healthcare Industry Money in Politics
Fellow Workers here is a DRAFT of the changes to the GDC Bylaws the Steering Committee aims to present to Referendum
m for the membership to vote on. PLEASE look it over carefully, the changes are fairly extensive and aimed at making it easier to run the GDC as we comtinue to grow. Any area with a strikethrough means we propose to delete it. Any area that is bold and underlinded means we propose to add it. YOUR FEEDBACK IS ESSENTIAL. We need to hear ALL of your recommendations before this goes to Referendum so we can ensure it reflects what the membership feels is best for the GDC. If you stay silent, or simply say NO without offering a reason why, then you're not really participating! Get involved in YOUR General Defense Committee, fellow workers! From today (September 12) there is just over a month until the deadline for Referendum. That should be plenty of time for all of us to have some input on these Bylaws and make them the best the IWW has ever seen. Amongst the highlights of the proposal: Create new funds under the GDC to aid our non-imprisoned membership (ULP Fund, Good and Welfare Fund) as well as a Prisoner Sponsorship fund to allow incarcerated comrades a way to join the organization. Changes GDC International Groups to GDC Locals; we're not sure why a separate name was ever chosen for international comrades. More clearly defines the relationship of the Steering Committee to the CST and vice-versa. Enables Locals to set their own dues rates. Establishes guidelines for Referendums that are currently missing from the GDC Bylaws. Currently, they are NO guidelines, which makes Referendum sort of the CSTs call. Establishes that GDC Locals are always subordinate to the IWW, and must have 5 IWW members in good standing for a charter. This ensures no one can take the GDC as a whole or a GDC Local. More clearly spells out the roles of CST, Steering Committee, and Local officers. Currently, the Bylaws are vague in spots on what exactly the rights, responsibilities, and privileges of those offices are. In Solidarity, Chuck Bailey and Eric Zenke Sept. 12, 2011
NEW LANGUAGE OLD LANGUAGE Bylaws of the General Defense Committee of the Industrial Workers of the World Established: October 5th, 1917 As amended through 2001 As amended for 2012 All labor involved in the production of this pamphlet document was DONATED. Table of Contents TO BE ADJUSTED WHEN FINAL VERSION IS LAID OUT FOR PRINTING Article I 2 Section 1 Name . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Section 2 Structure . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Section 3 Locals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Section 4 Internationals . . . . . . . . . . . . . . . . . . . . . . . . . 3 Section 5 At Large/Regional Delegates . . . . . . . . . . . . . . . . . 4 Article II 4 Section 1 Membership . . . . . . . . . . . . . . . . . . . . . . . . . . 4 Article III 5 Section 1 Goals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 Article IV 5 Section 1 General Officers . . . . . . . . . . . . . . . . . . . . . . . . 5 Section 2 Duties of the Central Secretary-Treasurer . . . . . . . . . . 5 Section 3 Election of the Central Secretary-Treasurer . . . . . . . . . 6 Section 4 Duties of the Steering Committee . . . . . . . . . . . . . . 7 Section 5 Election of the Steering Committee . . . . . . . . . . . . . 7 Section 6 Charges Against Officers . . . . . . . . . . . . . . . . . . . 8 Article V 8
Section 1 Funds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 Section 2 Dues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9 Article VI 10 Section 1 Clearing House . . . . . . . . . . . . . . . . . . . . . . . . . 10 Article VII 10 Section 1 Materials . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 Section 2 Referendums . . . . . . . . . . . . . . . . . . . . . . . . . . 11 Appendix I: Charges Against General Officers 12 Appendix II: Offenses 13
Preamble The aims of this organization shall be to provide defense and relief to members of the working class who are being persecuted for their activity in the class struggle. Coordinating our efforts will enable the General Defense Committee to oppose the vicious onslaught of the master class with a powerful defense, especially when labor is engaged in industrial strife. It has been found in past labor struggles that the lack of adequate defense has resulted in many of the most active fighters for labor being arrested and imprisoned for long periods of time or being legally murdered by the employing class. It shall be the purpose of the General Defense Committee to aid those who find themselves persecuted by the powers that be in their struggle for justice and freedom. It shall also be the purpose of the General Defense Committee to publish and distribute literature exposing the false and reactionary propaganda of the master class to victims of incarceration, their friends, family and allies. By organizing the many and centralizing their efforts in each instance, labor will be given the protection of the entire working class, thereby making an injury to one the concern of all.
Article I Section 1 Name This organization shall be known as the General Defense Committee of the Industrial Workers of the World, and shall be stated so on membership cards, literature, and letterheads. Section 2 Structure and Publications A The General Defense Committee shall be composed of GDC Central, which includes the CST and the Steering Committee, chartered GDC Locals initiated by Industrial Unions and/or General Membership Branches, and chartered GDC Regional Groups Locals initiated by ROCs and individuals in areas outside North America of the United States. B GDC locals may be known as the GDC of the IWW, or of a particular local number, ROC, IUB, geographic location, or GMB where applicable. C The official publication of this organization shall be called the Defiant Spirit, and distributed electronically by default. The CST is in charge of producing the Defiant Spirit, or appointing a Defiant Spirit editor to produce the publication under the direction of the Steering Committee and CST. The Steering Committee and CST are to establish a schedule for the production and distribution of the Defiant Spirit. Submission deadlines for each Defiant Spirit shall be announced 30 days prior to close. D The official means of communication between Defiant Spirit issues shall be the GDC Email List at gdc-list@lists.iww.org. All GDC members so indicating on their membership form shall be subscribed to this list. Members can also communicate with GDC Central via mail at the address printed at the end of this document. E Paper subscriptions to the Defiant Spirit are available for a subscription cost decided each year by the Steering Committee. This subscription cost will cover the costs of printing and mailing. Paper subscriptions shall be the default for any and all incarcerated GDC and IWW members at no cost. These subscriptions will be maintained and processed by GDC Central. F Upon request, incarcerated members may communicate by mail to the GDC email list, or be published in the Defiant Spirit by proxy of the Steering Committee or CST. G The Constitution and Bylaws of the Industrial Workers of the World hereby
supercede any conflicting language within the GDC Bylaws as the GDC is bound by the IWW Constitution. H Financial reports will be made to the GOB of the IWW by the CST every fiscal quarter. Section 3 Locals A A GDC local must consist of at least five IWW members in good standing, and membership shall be open to non-members of the IWW who subscribe to the general principles and aims of the IWW and GDC. B Charters for GDC locals shall be issued and approved by the Steering Committee or GEB. A local charter may be placed in suspension by a two-thirds vote of the body which has jurisidiction over said Local IU, ROC, or GMB with jurisdiction, pending a and any GDC charter may be revoked immediately by a majority vote decision by of the Steering Committee or GEB to revoke a charter. The GDC local may appeal the decision of the Steering Committee GEB regarding charters to the membership GEB of the IWW. Revocations remain in effect during the appeals process. C Quorum to meet for a for an official GDC local meeting shall be five GDC members in good standing, and at least one present member must be an IWW member in good standing. D Each chartered local shall elect a Secretary-Treasurer who shall be the is the responsible custodian of all branch records, funds and supplies, oversees coordinating efforts with other locals, and reporting reports all such business and activities to the local IU or GMB. and It is also the duty of the Local Secretary Treasurer and all Delegates of the GDC to continue making monthly quarterly reports to the CST of the GDC, even if the report is to inform your local or the CST that no dues were taken. The local Secretary-Treasurer will be is responsible for maintaining all minutes of meetings and his/her their own monthly financial report to his/her their branch and shall endeavor to keep all members in good standing and aware of all referenda. E The locals elected Secretary-Treasurer shall be a is the delegate for the local, and as such is resposible responsible for all initiation fees, dues and assessments taken in by the local, and seeing that all funds are properly maintained and properly recorded, and that the per capita set aside for the Central Defense Fund is properly deposited with the CST. The local may elect, and the CST may appoint, other delegates as necessary. . F All local officers elected shall be members in good standing of the GDC.
G Each local may modify their local dues structure while keeping the split with GDC Central 50% of dues collected at the rate established by GDC Central, and produce local assessments stamps. No GDC Local may modify their dues structure to be less than $5 per quarter dues unless it is for the membership of the incarcerated. A copy of any stamp or structure changes issued by a local must be sent either by mail or email to GDC Central. H No chartered local shall enact any legislation or motion conflicting with the constitution of the IWW. Refer to Article 1 Section 2G. I All Locals and Delegates will remit copies of financial records to any member of GDC Central, any member of the GEB or the GST immediately upon request. The requesting body will reimburse copy costs for locals and delegates in hardship, provided receipts are included with IWW Reimbursement Forms. Reimbursement Forms can be found at www.iww.org or be mailed by request of GDC Central. J All Delegates are responsible for maintaining his/her own monthly financial report to their branch or the CST and shall endeavor to keep all members within their jurisidiction in good standing and aware of all referenda. K Uncleared Delegates and dechartered locals will remit all GDC supplies and funds to GDC Central upon a passing motion of the Steering Committee or the GEB. Section 4 Internationals Locals Outside of the United States A A GDC International Group will be the equivalent of a regional local outside North America for the GDC GDC Local. Each GDC International Local must be sponsored by an ROC, consist of at least five members of the IWW GDC in Good Standing, and be open to non members of the IWW who subscribe to the general principles and aims of the IWW and GDC. B Charters of GDC International Groups Locals shall be must be issued and approved by the GEB or Steering Committee or GEB. Charters for GDC locals with that International Groups Region shall be issued and approved by the local ROC. C Initiation fees, dues, and assessments may be set by a GDC International Group in that regions currency, and taking into consideration that regions prevailing economic situation. All funds raised by the International Group will be maintained by that group for local defense work with the exception of an initial charter fee of $25 and a yearly sustainer fee of $10 $50 (US) to be deposited to the General Fund no later than February 1st of
each year.These fees may be negotiated or waived by the CST in cases where they represent an economic hardship to the International Group. D Each International Group shall elect a Secretary-Treasurer who shall be responsible for keeping records, coordinating efforts with other locals, and reporting activities to the local IU or GMB and making quarterly reports to the CST of the GDC. F International Groups will be responsible for their own Bylaws, membership, officers, and means of internal communication. They may modify their local dues structure, and produce local assessments stamps. A copy of any stamp, structural change, or bylaws issued by an International Group must be sent to GDC Central via mail or email. G No International Group shall enact any legislation or motion conflicting with the constitution of the IWW. H International Groups will be chartered the same as any Local and will be given a GDC Local number and official Charter issued by GDC Central. Section 5 At Large/Regional Delegates A In areas where there is no branch or local affiliate or the GDC, the The Central Secretary-Treasurer may issue credentials authorizing certain members to act as regional or at large delegates with the authority to initiate new at-large members of the GDC, and/or collect dues and assessments for the GDC International (also known as GDC Central). B Regional delegates must be members in good standing of the GDC. C Regional delegates of the GDC IWW may also serve as delegates for the IWW GDC upon requesting Delegate Credentials from GDC Central. , but no local branch or body of the IWW may have more than one regional or at-large delegate. D Each regional delegate will be responsible resposible to the GDC for all funds and supplies, including initiation fees, dues and assessments and will pass on such information along with all funds received to GDC Central Administration in a bi-monthly quarterly report. Article II Section 1 Membership A Any member of the IWW in good standing may be a member of the GDC.
B Non-members of the IWW, and those ineligible for membership in the IWW may still join the GDC, as long as they subscribe to the general principles and aims of the IWW and GDC. C All members shall agree to abide by the regulations of the GDC, and the Constitution and regulations of the IWW; to study the principles contain therein; and make themselves acquainted with its purpose. D No expelled member of the IWW shall be eligible for membership in the GDC. Article III Section 1 Goals A The GDC of the IWW wants most to provide support to any member of the working class who finds themselves in legal trouble due to their involvement in the class war. B Any member of the IWW or GDC in good standing may petition the GDC for direct aid in their legal defense on charges resulting from protests, picketing, or other acts related to job actions or activist causes. This aid may include bail, bond, or legal aid, and may take the form of a grant or a no interest loan. Members may also rest assured assurend of solidarity from the organization, including motions of support, letter writing, and public education as to the case (or cases) in question. C Non-members of the IWW and GDC who find themselves embattled with the law may also expect solidarity from the organization, including motions of support, letter writing, and public education as to the case (or cases) in question. D GDC Locals and International Groups will always try to engage in strike support and other activities consistent with the aims and principles of the IWW. E The GDC is also available to aid our organizers facing ULPs and other work related legal issues. Article IV Section 1 General Officers A The officers of the GDC shall consist of the Central Secretary-Treasurer (CST), the Chair of the General Executive Board, and General Defense Steering Committee. Section 2 Duties of the Central Secretary-Treasurer
A The CST of the GDC shall be to must take charge of all books, papers and effects of the GDC, shall will conduct the correspondence pertaining to his/her office, shall be is the custodian of the seal of the organization and shall will act as a clearing house for stamps, cards and other educational supplies. B The CST shall be is responsible for the financial finical accounting of all funds of the GDC, controlling the Central Defense Fund of the GDC, and all other expenses and special accounts. The CST will be is responsible for making a quarterly financial finical report available to the General Executive Board, the general membership of the IWW, and the membership of the GDC by submission to the IWW GOB. C The CST shall publish a quarterly circular for all North American members and International Groups of the GDC to be issued in Febuary, May, August, and November. D The CST will make himself themself available each year to the IWW and the GDC during the General Delegate Convention Assembly of the Union IWW, reporting on the state of the GDC and on its various projects. Any business, or discussion of the processes, procedures, or methods of the GDC may be handled by the union Convention Delegates at this time. E C The term of office shall be for 1 year and shall commence on January 1. D The CST of the GDC will have a voice and vote on all Steering Committee motions and will maintain all records of motions failed and passed. F The CST will remit any and all financial and communication records upon request by any member of the GDC Steering Committee, or any member of the GEB or the GST of the IWW. Section 3 Election of the Central Secretary-Treasurer A Nominees for the office if of the CST shall each be a member of the IWW, in good standing for two years, and a member of the GDC for at least 6 months one year prior to nomination. B In the event that no nominee for CST meets these requirements, then they shall be reduced to 1 years membership in the IWW, and 6 months membership in the GDC. C Nominations for the office of CST will be made at the General Assembly IWW Delegate Convention of the IWW, beginning in the year 2000. or through the Nominations may also be made via mail or e-mail, with nominations closed by the close final
adjournment of the General Convention GA. The election shall be by general referendum of the membership of the IWW. D Any person elected to the CSTs office must remain in good standing with both the IWW and the GDC while holding office. E No candidate for the CST will be permitted to run or hold other office within the General Administration at the same time, unless no other qualified candidates are available. F No candidate is eligible for more than three terms in succession. Members who have served three or more consecutive terms as CST shall only have their names placed on the ballot, in the event that two qualified nominees cannot otherwise be secured. G The General Executive Board, of the IWW shall have the power to appoint the Central Secretary-Treasurer of the General Defense Committee whenever the post is left vacant. G If the post of Central Secretary-Treasurer becomes vacant, the GDC Steering Committee shall have the power to appoint a short-term Central SecretaryTreasurer of the General Defense Committee by unanimous vote. This appointment shall become effective immediately but shall be reviewed by the GEB. The GEB shall have final authority over who the new long-term CST shall be. Section 4 Duties of the Steering Committee A The General Defense Steering Committee will be made up of three elected representatives of the GDC three members elected by GDC referendum and the CST. and The Chair of the General Executive Board will act as a deciding vote when a tiebreaker is neccessary. B The Steering Committee shall have general supervision of over all affairs of the GDC, and shall watch vigilantly over the interests throughout its jurisdiction. C The Steering Committee shall consider petitions of those requesting aid from the GDC, and have the full power to provide whatever help is available up to and including 50% 30%, of non-earmarked funds, of from the Central Defense Fund of the Committee GDC. Funds greater than 30% shall be approved by GEB vote. D The Steering Committee will have full editorial oversight over all GDC publications and guide their policy. E Elected Representatives to the Steering Committee shall each serve a term of office of 1 year, which shall commence on January 1.
F Each elected Representative to the Steering Committee is eligible for a maximum of five terms in succession. Members who have served five or more consecutive terms shall only have their names appear on the ballot, in the event that three other qualified nominees cannot otherwise be secured. G In the absence of elected members of the Steering Committee, the remaining participants, the CST, the GST of the IWW, and the Chair of the General Executive Board, may make any and all decisions on the Committees behalf. Section 5 Election of the Steering Committee A Nominees for the Steering Committee shall each be a member of the IWW for 1 year 2 years, and a member of the GDC for 6 months 1 year prior to nomination. B Members of the GDC who are not eligible for membership in the IWW may be nominated to serve on the Steering Committee, but must have maintained membership in good standing for at least 2 years prior to nomination. C In the event that no nominee meets these requirements, then they shall be reduced to 6 months membership in the IWW and the GDC at the time of nomination. D Nominations for the Steering Committee must be made by mail to the Central Secretary-Treasurer no later than 10 days before the IWW Delegate Convention General Assembly, The Central Secretary Treasurer and Steering Committee will contact nominees regarding their acceptance before reviewing nominees eligibility. A list of eligible nominees will be released with all proposed referendum resolutions via direct mailing to each GDC member in good standing. The election shall be by special referendum of the membership of the GDC only, mailed out with the November Quarterly. of the GDC. E Any person elected to the Steering Committee must remain in good standing with both the IWW and the GDC while holding office. F No elected member of the Steering Committee will be permitted to run for or hold other office within the General Administration as the same time. G In the event of a vacancy on the Steering Committee, with all duly elected members or alternates already serving or being unavailable, the Central Secretary-Treasurer and the GEB Chairperson shall appoint a new committee member until another shall be elected by referendum.
Section 6 Charges Against Officers A Charges against any Officers of the GDC should be filed in writing with the GEB or the General Assembly, following the protocols laid out for charges in the Constitution of the IWW. (Refer to Appendix I and II) Article V Section 1 Funds A Two Five funds will be maintained by GDC Central. The General Fund will be maintained as the operating fund for the GDC, for the purpose of expenses, printing, and other needs supplies that may arise related to the operation of GDC Central. The Central Defense Fund of the GDC shall be retained in a separate account accounted for separately and distributed for the purpose of legal defense work. The Prisoner Sponsorship fund will be maintained and dispensed by the CST and used for sponsoring incarcerated individuals seeking membership in the GDC and/or IWW, pending a majority vote of the Steering Committee. Prisoner Sponsorship funds will be accumulated through donations and assessments. The GDC Unfair Labor Practice fund will be maintained by the CST and distributed by petition from the IWW Organizing Department Board, released upon a majority vote of the Steering Committee. Funds will be accumulated through donations and assessments. The ULP fund is to be used toward legal costs affiliated with IWW organizers dealing with Unfair Labor Practices. Individuals must be members of the GDC to receive this funding. The Good and Welfare Fund (GaWF) will be maintained by the CST and distributed by majority vote of the Steering Committee. The GaWF is accumulated through donations and assessments. The Good and Welfare Fund exists to help organizers who find themselves facing hardship such as eviction, not being able to afford food and other basic neccessities, due to being fired for participating in union activity. Only 10% of the available funds in the GaWF may be dispersed to any one individual within a 12 month period. Individuals must be members of the GDC to receive funding. B Any member of the IWW or GDC in good standing may petition the GDC Steering
Committee for direct aid in their legal defense from the Central Defense Fund. This aid may take the form of bail, bond or legal aid, and make take the form of a grant or a no interest loan. This member should make an honest good faith effort to repay this no interest grant or loan to the Defense Fund within 24 months. C Such funds may normally only be drawn out of the Central Defense Fund at the direction of by the Steering Committee, after a proper petition, motion and a majority vote in favor. D In the interest of fulfilling one of the GDCs goals of strengthening Solidarity with all prisoners of the class war, the CST is permitted to issue checks for Gifts of Solidarity to other Defense efforts that are brought to the attention of the GDC. Such gifts may not exceed $25 each per recipient per year, and the total of all gifts in a calendar year may not exceed $200 $500, or %4 of the Defense Fund, whichever is greater. Such gifts may only be issued to Defense Funds and not or to incarcerated individuals who are members of the General Defense Committee or IWW. These gifts can also be given to prisoners as commissary gifts E In the event of an emergency, the CST may seek the approval of the Chair of the GEB to access up to 50% 75% of funds not yet earmarked the fund in the Central Defense Fund for legal aid, or other needs arising from state oppression. F The membership of the GDC may requisition up to 50% of the funds from the Central Defense Fund for any given initiative by means of a referendum of the entire GDC membership. Said referendum may be called by a signed petition of 15 members of the GDC, in good standing for at least one month prior to the month the petition is initiated, or by a chartered local making official petition by majority vote and balloting to be conducted by the office of the CST. G GDC Locals and International Groups may raise funds through assessments and other fundraising techniques to aid in the defense of the class war prisoners outside the IWW and GDC. These funds will be a separate trust maintained by the chartered local or international group. Delegates at large are required to submit all dues and assesments with their quarterly delegate report. H Donations sent to the GDC on behalf of the class war prisoners outside the IWW and GDC, will always be so earmarked and always be passed on in a timely fashion to a Defense Account for aforementioned prisoner. Donations that remain unclaimed after 6 months of attempting to establish contact with the prisoners Representative(s) will be allocated to the Defense Fund by the CST or to the Prisoners Commissary fund if possible.
I The General Fund is not to exceed $10,000. Funds exceeding $10,000 in the General Fund shall be transferred to the Defense Fund. J There shall be no limit to funds held in the GDC Defense Fund. K Funds over $10,000 in the GDC ULP fund will be transfered to the IWW ODB. L Funds over Section 2 Budget and Fiscal Year A At the start of each IWW fiscal year the Steering Committee and GEB Chair will discuss, create, and finalize a budget for the GDC. B The GDC Central budget must be completed and approved via GDC Steering Committee no later than 15 days after the start of the IWW Fiscal Year. C The GDCs fiscal reporting year shall coincide with that of the IWW. Currently each fiscal year starts July 1st and ends June 31st. Section 2 3 Dues A Initiation fees for GDC membership shall be are $5.00, all of which goes in the GDC General Fund, and dues shall be a sliding scale starting at no less than $5.00 per quarter. It is recommended that one quarters GDC dues be the same as one month of the members IWW dues. It is the policy of the GDC to put no financial barrier to prevent any worker from joining. Accordingly, initiations shall not exceed $5.00 nor quarterly dues exceed $27.00. B Each local shall retain 75% 50% of dues collected, and pass the remaining 25% to the Central Defense Fund and 25% to the General Fund. C At-large members, who are not part of a branch local shall pay all their dues directly to the CST, or an at-large or regional delegate. Of these dues, 75% 50% shall be deposited in the Central Defense Fund; the remaining 25% 50% may will be deposited in the General Fund. D Dues and other funds, such as assessments, raised by International Groups will be maintained by
that group for local defense work. The exception will be an initial charter fee of $25(US) and a yearly sustainer fee of $10$50 (US) to be deposited to the General Fund no later than February 1st. These fees may be negotiated or waived by the CST in cases where they represent an economic hardship to the International Group. Article VI Section 1 Clearing House A The CST and Steering Committee will maintain contact with each local of the GDC, and the CST will function as a Clearing House for all supplies and materials necessary to conduct the business of the GDC. B The Secretary-Treasurer of each local or group GDC Local or International Local will maintain contact with the Clearing House through the CST, and shall record all fees, dues, and assessments collected on forms by their locals delegate(s) provided by the Clearing House and return this along with all fees and dues money set aside for the General and Central Defense funds to the CST monthly. Quarterly. Quarterly delegate, branch, and body reports are due by the 1st of March, June, September, and December. C All regional delegates, delegates at large and or others bearing such credentials shall record all fees, dues, and assessments collected on forms provided by the Clearing House and return this along with all fees and dues money received to the CST bimonthly.shall record all fees, dues, and assessments collected on forms provided by the Clearing House, and must return this along with all fees and dues money allocated for GDC funds to the CST. Section 2 Clearing House and Reporting Deadlines A The due dates for all reporting to GDC Central each quarter are hereby the 1st of January, April, July, and October of each year. This shall remain effective for all delegates, Locals, Secretary-Treasurers, International Group locals and delegatesat-large. Article VII Section 1 Materials A There shall be a standard label for the GDC. It shall be of a crimson color and always the same in design. The use of the label shall only be vested in our organization. The Steering Committee and CST shall take any necessary steps to protect the label of the GDC.
B All branches of the GDC should procure and use such supplies as dues books, dues stamps, official buttons, labels and badges from GDC Central, whenever possible. All such supplies are to be of a uniform design. International Groups may print their own materials, dues books, dues stamps, official buttons, labels and badges, in their own language, if they desire. The general design and format of these materials should remain generally consistent with those produced by the Central Administration, and copies of these materials must should be forwarded on to the CST. C Assessments may be produced by the local and regional groups distributing them in various denominations. Likewise, assessments may be produced by GDC Central of various denominations, and types for various fundraising purposes. A copy, digital or otherwise, of any stamp issued by an International Group must be sent to GDC Central. Section 2 Referendums A A general referendum on any organization question, including constitutional amendments, may be initiated by the General Steering Committee of the GDC, by a petition of 20 GDC members in good standing, or by the GEB or the General Delgate Convention Assembly of the IWW. B All properly submitted referendum items shall be included on a ballot to be issued not more than 60 days after the end of the IWW Delegate Convention. Notification of all referendum items, including the full text of all questions to be voted upon and the names of all nominees for the Steering Committee who are eligible and have not declined, will be provided to all members via mailing not less than 30 days before ballots are mailed. Ballots shall be sent to all members in good standing and shall be open for voting for 31 days. Ballots shall be sent by mail to all members. The deadline for receipt of ballots at GDC Central shall be printed on the ballot. Votes shall be counted and the results released no later 32 days after ballots were mailed to the membership. Referendums shall take place annually, the ballot to me mailed with the November circular. All members in North America will receive their ballots in this manner, and will be responsible for returning their ballots to the CST by mail before the deadline set. International Groups will handle their own balloting as best suits their situation; reporting their vote totals to the CST by mail before the deadline set. C Bylaws, as well as future amendments and revisions of the Bylaws, must be approved by the GDC membership through referendum. D All changes ratified by a general referendum ballot shall take effect January 1, unless otherwise noted in the measure.
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text:
RESEARCH DESIGN AND METHODS
Immunology and Transplantation| February 01 2003
T-Cell Compartments of Prediabetic NOD Mice
Stuart P. Berzins;
Stuart P. Berzins
From the Section on Immunology and Immunogenetics, Joslin Diabetes Center, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts
Emily S. Venanzi;
Emily S. Venanzi
Christophe Benoist;
Christophe Benoist
Diane Mathis
https://doi.org/10.2337/diabetes.52.2.327
Stuart P. Berzins, Emily S. Venanzi, Christophe Benoist, Diane Mathis; T-Cell Compartments of Prediabetic NOD Mice . Diabetes 1 February 2003; 52 (2): 327–334. https://doi.org/10.2337/diabetes.52.2.327
Given the importance of the NOD mouse as a model of type 1 diabetes, there is a surprising lack of published information on the overall composition of the thymic and peripheral T-cell compartments. In this study, we revisited some earlier reports of T-cell abnormalities in this strain and examined a number of additional parameters to provide a global view of T-cells in prediabetic NOD mice. In some cases, we concur with past conclusions, but in other important areas, we find that NOD mice closely resemble nonautoimmune strains. Specifically, and contrary to published reports, the thymocyte subset distribution, the rate and composition of thymic export, and the composition of the peripheral T-cell pool, including the proportion of CD25+CD4+ T-cells, are essentially normal in prediabetic NOD mice. These factors are therefore unlikely to be involved in the loss of tolerance that leads to autoimmunity within this strain.
NOD mice develop an autoimmune form of type 1 diabetes that is similar to the human disease (1). The model provides an opportunity to study diabetes in a genetically and environmentally controlled setting, yet surprisingly few studies offer a global view of lymphocytes. Of particular note, the characteristics of the NOD T-cell compartment remain poorly defined, despite the involvement of T-cells in type 1 diabetes being universally accepted. Among the articles that do exist are reports of problems within the thymus and peripheral T-cell compartments that may predispose, or otherwise directly contribute, to the autoimmune phenotype of the NOD strain (1–5).
In the context of autoimmune diabetes, abnormalities within the NOD thymus are especially noteworthy because of the strong association between disorganization of the thymus structure and defective central tolerance (6,7). Abnormalities ascribed to the NOD thymus include unusually large perivascular spaces (5), a poorly defined demarcation between the cortex and medulla (8), increased numbers of mature T- and B-cells (9), a relative deficiency of NKT cells (10), and defects in negative selection and T-cell export to the periphery (4,11,12). A variety of peripheral defects have also been reported, some of which could stem from thymic abnormalities. For instance, one often-cited feature of NOD mice is an unusually high proportion (rather than number) of T-cells in peripheral lymphoid organs, possibly resulting from excessive thymic export (1,3,4,13,14). Other studies cite deficiencies within the peripheral immunoregulatory T-cell compartments of NOD mice more consistent with low rates of thymic export (5) or independent of thymic dysfunction (15,16). Unfortunately, causative links between the thymus and peripheral T-cell abnormalities in NOD mice remain speculative because thymus emigration has not been directly tested.
Abnormal thymocyte differentiation, thymic export, or T-cell pool composition could predicate autoimmune disease and are therefore deserving of careful analysis in NOD mice (6,17). Unfortunately, some studies cited in support of peculiarities appear rather unconvincing in hindsight. Sometimes, this is because of the unsophisticated analytical tools available at the time or because only a partial comparison, or comparison with a single purportedly normal strain, was reported. In other cases, mice were examined at an age likely to coincide with systemic autoimmunity, making it difficult to exclude the possibility that abnormalities were secondary to disease. The result is a vague characterization of T-cell compartments in the dominant animal model of type 1 diabetes and uncertainty about how much it really does differ from the norm. It is therefore imperative to reexamine the reported abnormalities and, more generally, to characterize the T-cell repertoire of NOD mice in the context of today's knowledge and tools.
The value of our study rests on three critical features. First, NOD mice were examined at 6 weeks of age, when the T-cell compartments were fully established, but before diabetes onset or even substantial insulitis. Second, comparisons were made with age- and sex-matched mice of usually three other strains, enabling us to distinguish between common strain-to-strain variability and a real difference between NOD and nonautoimmune animals. Third, we exploited modern technology, including multiparameter flow cytometry and direct tracking of peripheral T-cells. The result is a global view of the NOD T-cell compartment that should frame future discussions of the anti-β-cell T-cell reactivity characteristic of this strain.
Female NOD, BALB/c, C57BL/6, and CBA mice obtained from The Jackson Laboratories were maintained in clean conditions in the Joslin animal facility. NOD mice congenic (>10 backcrosses) for major histocompatibility complex (MHC) class II deficiency (I-Aβ0/0) were obtained from our colony.
Flow cytometry.
Lymphocyte suspensions were prepared and stained according to standard protocols using Pharmingen or Caltag antibodies. Cells were analyzed using Coulter instrumentation, and data were analyzed using EXPO or WinMDI software.
Bone marrow chimeras.
Chimeras were prepared as described previously (18). Briefly, lethally irradiated mice reconstituted with 1.5 × 107 bone marrow (BM) cells derived from the femurs of donor mice were analyzed by fluorescence-activated cell sorting (FACS) 8 weeks later.
Analysis of thymic export.
Details of this technique are described elsewhere (19). Briefly, thymic lobes were injected with ∼10 μl of 350 μg/ml fluorescein isothiocyanate (FITC), randomly labeling 30–60% of thymocytes. Mice were killed 24 h later, and lymphoid organs were analyzed by FACS. Emigrants were identified as live-gated FITC+ cells expressing CD4 or CD8 (omitting autofluoresing cells and doublets) and quantified by considering the size of the peripheral pool to be equal to the number of spleen cells plus twice the number of cells pooled from mesenteric, inguinal, brachial, and axillary lymph nodes (20). Cell counts were performed in duplicate using a hemocytometer and trypan blue.
Homeostatic proliferation.
Cell suspensions from pooled lymph nodes of B6 and NOD mice were resuspended to 1 × 107 cells/ml in 1 μmol/l carboxyfluoroscein diacetate succinimidyl ester (CFSE) in Dulbecco's modified Eagle's medium 10, incubated for 10 min at 37°, and washed. The 5 × 106 cells were intravenously injected into age-matched syngeneic recipients, sublethally irradiated 2 days prior. Recipients were analyzed by FACS on days 2, 5, 8, and 13.
Statistical significance.
For normally distributed data, Student's unpaired t tests were used to compare two groups; ANOVA tests were used for three or more groups. Mann-Whitney tests were used when normal distribution could not be demonstrated.
T-cell differentiation.
Thymic abnormalities reported to affect NOD mice include structural irregularities (9), accumulations and deficiencies of particular thymocyte subsets (10), and export dysregulation (4), yet the size and distribution of NOD thymocyte subsets is generally observed to be normal (21). In light of this apparent inconsistency and the recent suggestion of defective negative selection (12), we compared the size and relative proportions of different thymocyte subsets in prediabetic NOD mice versus control (nonautoimmune) strains.
The distribution and size of the CD4−CD8−, CD4+CD8+, CD4+CD8−, and CD4−CD8+ subpopulations (collectively spanning all stages of thymic T-cell differentiation) were similar for all strains (Fig. 1A and B). Within these compartments, the expression of other maturity markers (such as αβ-T-cell receptor, heat stable antigen, or peanut agglutinin and CD25, CD44, and CD69 (Fig. 1B; data not shown) on NOD thymocytes (and other strains) was within the boundaries of strain-to-strain variation and was therefore consistent with normal T-cell differentiation. NOD mice were unusual only in their thymic B-cell compartment (which is two to five times larger than that in other strains [Fig. 1B]) and αβ-TCR+CD4−CD8− subset, which contains regulatory NKT cells. This latter deficiency has been reported previously (10,21) and will be discussed further.
The curiously high level of thymic B-cells was consistent with previous reports of mature T- and B-cells accumulating in the perivascular regions of the thymus from 12 weeks of age (5,22). We found no evidence of additional mature T-cells, but our finding of a similar trend in 6-week-old mice demonstrates that this trait develops before disease onset. Small numbers of B-cells are found normally in the thymus (23), but accumulation is associated with thymic abnormalities in other autoimmune models (24), making the higher incidence in NOD mice worthy of further investigation. B-cells play an important role in the development of diabetes in NOD mice (25–27), but in the thymus, they represent an extremely minor population of cells (<1%), and their impact on T-cell development is generally regarded as insignificant.
Dynamics of T-cell differentiation.
Before export, mature thymocytes undergo a highly regulated cascade of differentiation and proliferative events. The kinetics of development are as predictable as the phenotypic changes that take place, so abnormal kinetics can indicate aberrant maturation. The unusual kinetics of T-cell differentiation in diabetes-prone BB rats, for example, coincide with thymic defects linked to spontaneous T-cell autoreactivity in that strain (28).
In vivo pulse-labeling with bromodeoxyuridine (BrdU) was used to examine the dynamics of T-cell differentiation in NOD mice. BrdU is incorporated by dividing cells and retained until passed to daughter cells, thereby allowing the progression and kinetics of differentiation to be tracked. The initial incorporation of BrdU by thymocytes was similar for NOD and control strains for the proportion and phenotype of cells stained, resulting in comparable profiles of BrdU staining 24 h after BrdU administration. Interestingly, a significantly lower proportion of BrdU+ cells was observed among NOD thymocytes at day 3 (Fig. 2) (and to a lesser extent, day 2 [not shown]). The phenomenon was most dramatic among CD4+CD8+ cells where the proportion of BrdU+ cells in NOD mice was two to three times lower than that for B6 mice. This could have been caused by the loss of BrdU+ cells or may reflect the dilution of BrdU to background levels due to increased turnover. The former is more likely because levels of incorporation at 24 h were similar in all compartments. However, this is such an early defect, it seems difficult to attribute great significance to it in the context of tolerance induction.
Thymocyte deletion mediated by BM-derived antigen-presenting cells.
Thymic tolerance is mediated by the deletion of thymocytes with high affinity for self-peptides. Defects in the stringency of negative selection can lead to the release of autoreactive T-cells and an increased risk of autoimmune disease. The presence of overtly autoreactive T-cells in NOD mice has led to suggestions of defective negative selection. Supporting evidence has been largely circumstantial (8,29), but Kishimoto and Sprent (12) recently reported impaired negative selection among semi-mature NOD thymocytes. Stimuli of signaling pathways associated with negative selection induced significantly less apoptosis among NOD thymocytes compared with other strains. No direct link to peripheral autoimmunity was demonstrated, and nonspecific stimulation was used in preference to physiological signals such as MHC-peptide complexes, but the data were consistent with a central tolerance defect.
We assessed the global extent of negative selection in NOD mice using a classic strategy developed by van Meerwijk et al. (18). Since BM-derived antigen-presenting cells are important mediators of negative selection (30), one can grossly measure the extent of negative selection by constructing BM chimeras that express MHC molecules on radio-resistant stromal cells but not on radio-sensitive BM-derived cells. For example, it is well established that B6 mice expressing MHC class II molecules on stromal, but not hemopoietic, cells have ∼50% more CD4-single-positive thymocytes than control animals, indicating that about one-third of positively selected class II-restricted cells are negatively selected by BM-derived cells in the thymus (18).
When lethally irradiated NOD recipients were reconstituted with BM from MHC class II-deficient NOD donors, the proportion of CD4+CD8− cells in the thymus was augmented by ∼50%, consistent with the level of deletion in the nonautoimmune B6 strain (Fig. 3). This does not exclude the possibility of more subtle defects, but at a gross level, the extent of deletion mediated by MHC class II-positive cells in the NOD thymus appears similar to that of B6 mice and does not support the concept of a general failure of negative selection. This conclusion is supported by the consistently similar distribution of CD4+CD8+, CD4+CD8−, and CD4−CD8+ thymocytes in NOD mice and other strains (Fig. 1) and the similar rate of conversion from CD4+CD8+ to CD4+CD8− and CD4−CD8+ thymocytes revealed using BrdU pulse-labeling (data not shown). A significant failure of negative selection in the NOD thymus would presumably permit higher-than-normal proportions of single-positive NOD thymocytes to survive, and the absence of this phenomenon argues for comparable levels of negative selection.
T-cell export from the thymus
Abnormal thymic emigration is causally linked to a variety of autoimmune diseases (31,32), and reports alluding to defective export in NOD mice make it a logical target for investigation (4,5,11). Although a confirmed defect in the emigration of T-cells from the NOD thymus would provide important clues about the initiation of diabetes, to date, assessments of NOD thymic export have been based on indirect histological studies. The abnormal congregation of mature lymphocytes in the perivascular spaces of the NOD thymus led to suggestions that emigration of mature thymocytes, or certain regulatory subsets, was retarded (5). Reduced export can induce peripheral lymphopenia and homeostatic expansion of peripheral T-cells, which can perturb normal subset distribution and is associated with the development of autoimmunity in clinical and experimental settings (32–35). Other groups proposed a contradictory defect in which thymocyte export might occur at higher-than-normal levels, thereby contributing to increased peripheral T-cell levels (4). A third possibility is that immature thymocytes were erroneously released from the NOD thymus, exposing the host to cells that would normally be negatively selected (36). Unfortunately, the confusion surrounding the possible dysregulation of the thymic export in NOD mice has remained unresolved. Before this study, export from the NOD thymus had not been directly tested and there was a complete lack of data relating to the maturity of recent thymic emigrants (RTEs).
FITC was intrathymically injected to randomly label thymocytes, thereby allowing identification of emigrant cells in the periphery (19). The number and proportion of FITC+ T-cells within the periphery were determined according to methods established in our laboratory (20) and other laboratories (36), and markers of maturity (e.g., HSA) were used to assess whether immature thymocytes were inappropriately released.
For 6-week-old mice, there was no difference in the rate of T-cell export from the NOD and B6 thymus (Fig. 4A and B). Previous reports, including our own, report the rate of thymic export for B6 and other nonautoimmune mice to be 1–2% of total thymocytes per day (19,20). Our results for 6-week-old B6 and NOD mice fall within this range (Fig. 4), as do preliminary results from 15-week-old mice (data not shown). Importantly, emigrants from the NOD thymus were almost exclusively CD4+CD8− or CD4−CD8+ and expressed low levels of HSA (data not shown), indicating that thymocytes were not prematurely released.
To ensure that the initial seeding of the T-cell pool was normal, we assessed thymus export in 3-day-old mice. NOD and B6 neonates had similar T-cell subset distributions in the thymus and spleen, and the phenotype of thymic emigrants resembled that of adult mice, with NOD RTEs almost entirely CD4+CD8− or CD4−CD8+ and HSAlo, consistent with the exclusive release of mature T-cells (data not shown). It is noteworthy, however, that the distribution of CD4+ and CD8+ T-cells within NOD RTEs was different from B6 mice (Fig. 4C). The CD4/CD8 ratio among RTEs, and in the periphery, is normally remarkably stable in mice and humans, reflecting the tight regulation of thymocyte differentiation and genetic controls on the relative sizes of the subsets (20,37,38). Typically, the CD4/CD8 ratio of RTE is slightly higher than that of mature thymocytes and peripheral T-cells, perhaps indicative of the independent regulation of the export process (20,39) and the expansion phase undertaken by many RTEs immediately before export (40). Although the number of RTEs released from the NOD thymus remained both normal and stable, the CD4/CD8 ratio was variable and typically contained higher proportions of CD4+ cells, suggesting that this aspect of thymic export was not under the tight control seen with other strains.
Peripheral T-cell compartment
The CD4/CD8 ratio among RTEs might be expected to reasonably determine the ratio within the periphery. However, NOD mice displayed similar CD4+ and CD8+ T-cell proportions to other strains (Fig. 5). This result indicates that a greater correction of the RTE CD4/CD8 ratio (through preferential division of CD8+ cells or loss of CD4+ cells) must occur for NOD mice to produce the homeostatically controlled peripheral CD4/CD8 ratio of ∼2. However, high CD4/CD8 ratios among NOD RTEs may offer an explanation for previous reports of a steadily increasing peripheral CD4/CD8 ratio during the onset of insulitis in NOD mice (41). Coupled with the variability within the export ratio of different NOD mice, further analysis is warranted to explicitly exclude the possibility that thymic export irregularities contribute to the onset of autoimmunity.
In addition to the normal distribution of CD4+ and CD8+ cells within the periphery of NOD mice, the relative proportions of T- and B-cells were also similar to other strains. This finding conflicts with previous reports that T-cell "hyper-accumulation" or "T-accumulation" characterizes the NOD strain, reflecting persistent T-cell hyperplasia. Abnormal proportions or numbers of T-cells in the peripheral pool could indicate dysregulation within the repertoire, and previous reports have speculated on whether T-lymphoaccumulation was caused by problems with apoptosis, proliferation, or thymic export (1,4,14). Each might conceivably predispose to autoimmunity or at the very least indicate problems associated with tolerance breakdown. Our findings, however, argue that no such problems exist in the periphery of NOD mice before disease onset. The number and relative proportion of CD4+ and CD8+ T- and B-cells in the NOD spleen were consistently similar to other strains. Higher lymphocyte numbers were observed in the NOD lymph nodes (Fig. 5A), but this is not indicative of T-cell hyperplasia as previously described because the increase applied equally to the T- and B-cell compartments. The reason for the increase in lymph node cell numbers was not immediately evident, but given that it was not observed in the spleen and was relatively minor, it seems likely to reflect simple strain-to-strain variation.
We also investigated the capacity for homeostatic expansion of T-cells in NOD mice. When T-cells are transferred into a lymphopenic recipient, the availability of space provokes the T-cells to proliferate (42). It is a recurring theme that T-cell homeostasis may be disrupted in NOD mice, perhaps leading to too many or too few T-cells within the overall pool or within particular T-cell subsets. To test homeostatic proliferation, we transferred CFSE-labeled lymphocytes from NOD or control mice into irradiated age-matched syngeneic recipients. Recipients were killed on day 2, 5, 8, or 13, with the extent of cell division revealed by CFSE dilution. On day 2, transferred cells had not proliferated significantly in either strain, but on day 5 and 8, extensive, but similar, levels of proliferation had occurred (Fig. 6). After 13 days, most cells had divided to the extent that peaks of CFSE staining were no longer apparent. Although the possibility of changes arising after day 13 cannot be excluded, we found no significant difference in the extent or rate of CD4+ or CD8+ T-cell proliferation between NOD and B6 mice up to this time point. This indicates that in addition to the largely normal number and distribution of T-cells within the peripheral pool, the homeostatic response to lymphopenia was also normal.
Regulatory T-cells
There are many reports of defects affecting the regulatory T-cell compartment of NOD mice. Some describe functional deficiencies, whereas others report abnormally low numbers of particular regulatory subsets (43). Most prominent are reported deficiencies of NKT cells (well represented within the αβ-TCR+CD4−CD8− compartment) (15) and CD4+CD25+ T-cells (16). Both subsets are thought to dampen autoreactivity. Reports of deficiencies within this compartment are especially interesting because they offer a credible, yet simple, explanation of how autoimmunity might develop in NOD mice and suggest strategies for resolving the dysfunction.
αβ-TCR+CD4−CD8− T-cells.
One well-documented defect in the NOD mouse is a relative paucity of NKT cells (10). The deficiency was first identified as a significantly reduced proportion of αβ-TCR+ cells within the CD4−CD8− compartment, with subsequent studies confirming that a large proportion of αβ-TCR+CD4−CD8− T-cells are NKT cells (15). The importance of NKT cells to the diabetes process remains controversial, particularly because their numbers appear to normalize with age, but the deficiency has been implicated as a contributing factor in NOD autoreactivity (44–46). Our analysis of NOD mice confirms the widely reported comparative deficiency of αβ-TCR+CD4−CD8− T-cells in the spleen, lymph nodes, and, most prominently, the thymus (Fig. 7). The deficiency was least obvious in the periphery and was not statistically significant when compared with the CBA spleen or BALB/c lymph node. Although beyond the scope of this study, analysis of subsets within the broader NKT cell population in different tissues is likely to be informative.
CD25+CD4+ T-cells.
The CD25+CD4+ T-cell compartment appears to contain cells with an important role in maintaining self-tolerance in the periphery (47). Significantly, some recent reports describe NOD mice as deficient for CD25+CD4+ T-cells (16,48,49). Given the thymic origin of these cells, a peripheral deficit could potentially arise from maturational defects in the thymus, during export to the peripheral pool, or in the periphery itself.
In the NOD thymus, ∼5% of CD4+CD8− T-cells expressed CD25, a fraction falling within the same range as nonautoimmune strains (Fig. 8A). The proportion fell to ∼3% among RTEs (at 24 and 72 h after export), but this also occurred in the control B6 strain, indicating no defect in the production or export of these cells in NOD mice (Fig. 8A).
The spleen, lymph nodes, and pancreatic lymph node were examined for evidence of a systemic or localized peripheral deficiency. Surprisingly, the proportion of CD25+CD4+ T-cells in NOD mice was no different from that of control strains at any site (Fig. 8B). Consistent with reported levels in nonautoimmune strains (50,51), ∼7% of CD4+ T-cells in the lymph nodes and spleen of CBA, BALB/c, and B6 mice expressed CD25. We observed the same incidence for NOD mice. Subtle differences sometimes occurred, but were rarely significant, and often took place between control strains.
Lastly, no differences were observed in the thymus, RTE (24 h after export), or spleen of 3-day-old NOD mice, arguing against any transient deficiency during the establishment of the NOD pool (data not shown). Although functional differences within the compartment cannot be ruled out, quantitative deficiencies in the CD25+CD4+ compartment do not appear to explain the loss of tolerance in the NOD mouse.
It is difficult to be certain why our results differ from others who report a CD4+CD25+ T-cell deficiency in NOD mice. At present, it is not possible to isolate the regulatory component of the CD4+CD25+ T-cell compartment, so activated peripheral cells provide a feasible source of variability. However, our stringent comparison of age-matched female mice found no indication of a CD4+CD25+ T-cell deficiency in the thymus or thymic emigrants of NOD mice, nor was an indication found in the spleen, lymph nodes, or pancreatic lymph nodes. It should be noted that an analysis of the NOD CD4+CD25+ compartment was not the primary focus of the earlier studies, and this aspect of the data was typically presented as not shown, or in a limited comparison with one control strain.
DX5+ and CD45RBlo cells.
In addition to CD4+CD25+ and NKT cells, other subsets of the T-cell repertoire proposed to have regulatory properties are DX5+ and CD45RBlo T-cells (45,52). We again detected no obvious deficiencies in these compartments of the NOD T-cell pool compared with control mice (data not shown).
In conclusion, our analysis of the NOD T-cell compartment examined subpopulations and locations where abnormalities could potentially predispose to autoimmunity. With some exceptions, the size and composition of thymic, RTE, and peripheral T-cell compartments in NOD mice were similar to other strains. This finding indicates that no broad quantitative defect precedes the onset of autoimmunity. At the level of regulatory T-cells, we confirmed previous reports of a deficiency within the NKT cell compartment of NOD mice, but found no evidence to support recent suggestions that a deficiency in the CD25+CD4+ compartment may also contribute to autoimmunity.
The use of prediabetic mice in our study was an important consideration because age (17) and autoimmunity (39) can induce significant changes within the T-cell repertoire that could collectively mask characteristics predisposing to autoimmunity. This issue of timing may be one reason why our global analysis found no evidence of abnormalities. In addition, in some instances, we were able to apply techniques unavailable to the original investigators and this is likely to be another reason why our findings disagree with some conclusions drawn from earlier work. In summary, NOD mice are clearly distinct from most other strains in their susceptibility to autoimmune attack, but the loss of tolerance does not appear to originate in general deficiencies within the T-cell compartment.
T-cell differentiation. Thymocytes from NOD mice were compared with three non-autommune-prone strains on the basis of subset distribution and cell number. The distribution of subsets defined by the differential expression of CD4 and CD8 was normal for all mice (A). With the exception of B-cell levels in the NOD thymus, other markers of maturity (e.g., αβ-TCR and HSA) also revealed no differences between strains (B). A minimum of five age-matched female mice were assessed for each strain.
Kinetics of T-cell differentiation. After two injections of 1 mg/ml BrdU (4 h apart), the thymi of mice were harvested and the incorporation of BrdU was determined by FACS. Levels were comparable for NOD and B6 mice at days 1, 5, and 7, but at day 3 (and day 2 not shown), two- to threefold fewer NOD thymocytes were positive for BrdU. The pooled results from one of three separate experiments are shown.
MHC class II-mediated negative selection. NOD mice were lethally irradiated and then reconstituted with BM from congenic donors that were either MHC class II positive (i.e., wild-type [WT]) or deficient (MHC IIo). Six weeks after reconstitution, the thymii were assessed for the presence of CD4+CD8− T-cells, with the difference between the two groups of a reflection of deletion mediated by MHC class II-positive BM-derived antigen presenting cells.
T-cell export from the thymus. T-cell export from the thymus was directly assayed by injecting FITC (0.3 mg/ml) into the thymic lobes. Mice were killed 24 h later, and emigrant cells were identified by virtue of fluorescence (A). B6 and NOD mice exported similar levels of T-cells (B), but the variability of the CD4/CD8 ratio among emigrants at both 24 and 72 h after export from the thymus was far greater in the NOD group (C). The CD4/CD8 ratio was similar for B6 ([cjs2108]) and NOD mice (□) among medullary thymocytes (Thy) and the overall peripheral pool (total). Pooled results from one representative experiment (of four) involving four to six mice per group are shown. d, day; LN, lymph node.
Peripheral lymphocyte pool. Peripheral T- and B-cell numbers and proportions were assessed for NOD mice and compared with three other strains. The spleen of all strains contained similar numbers and proportions of T- and B-cells. The lymph nodes of NOD mice were typically 20% larger than those of other strains, although this reflected increased numbers of both T- and B-cells, because these made up similar proportions of the pool in all strains. A minimum of five age-matched female mice were assessed per group. *A characteristic significantly different from the other three strains (P < 0.001).
Homeostatic proliferation. After the adoptive transfer of syngeneic CFSE-labeled lymphocytes to sublethally irradiated NOD and B6 recipient mice, mice were harvested at different time points to assess the extent of homeostatic proliferation. The similar levels of division occurring within the periphery of NOD and B6 mice are shown. The proliferation index represents the proportion of proliferating cells divided by the proportion of quiescent cells.
Assessment of the αβ-TCR+CD4−CD8− population and the prevalence of αβ-TCR+CD4−CD8− cells. Levels of αβ-TCR+CD4−CD8− T-cells in NOD mice were measured and compared with three nonautoimmune strains. The proportions were lowest in the NOD strain, particularly in the thymus. NOD levels were similar to that of CBA mice in the spleen and to BALB/c in the lymph nodes. DNs, double negatives.
Proportion of CD25+CD4+ T-cells. A: Assessment of the thymic CD25+CD4+ T-cell compartment. Levels of CD25+CD4+ T-cells in the thymus of NOD mice were measured and compared with three nonautoimmune strains. No deficiencies were noted in any lymphoid compartment. Levels for the RTE population were also determined to be similar for NOD and the control B6 strain. Each symbol represents one mouse. B: Assessment of the peripheral CD25+CD4+ T-cell compartment. Levels of CD25+CD4+ T-cells in the spleen and lymph nodes of NOD mice were measured and compared with three other strains. The pancreatic lymph node (PLN) was assessed separately. No deficiencies were noted. Each symbol represents one mouse.
This work was funded by grants from the Juvenile Diabetes Research Foundation (JDRF) to the JDRF Center for Islet Transplantation at Harvard (project numbers 36 and 40) and from the National Institutes of Health (1R01DK59658-01). S.P.B. is a Long-Term Fellow of the Human Frontiers Science Program and is a former Fellow of the Juvenile Diabetes Foundation, and E.S.V. is a National Science Foundation Graduate Research Fellow.
We thank Ella Hyatt for animal husbandry and Grigoriy Losyev for flow cytometry assistance.
Atkinson MA, Leiter EH: The NOD mouse model of type 1 diabetes: as good as it gets?
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Semin Immunol
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J Immunol
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Lab Invest
Gombert JM, Herbelin A, Tancrede-Bohin E, Dy M, Carnaud C, Bach JF: Early quantitative and functional deficiency of NK1+-like thymocytes in the NOD mouse.
Eur J Immunol
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Kishimoto H, Sprent J: A defect in central tolerance in NOD mice.
Nat Immunol
Serreze DV, Leiter EH: Defective activation of T suppressor cell function in nonobese diabetic mice: potential relation to cytokine deficiencies.
Prochazka M, Gaskins HR, Shultz LD, Leiter EH: The nonobese diabetic scid mouse: model for spontaneous thymomagenesis associated with immunodeficiency.
Hammond KJ, Pellicci DG, Poulton LD, Naidenko OV, Scalzo AA, Baxter AG, Godfrey DI: CD1d-restricted NKT cells: an interstrain comparison.
Salomon B, Lenschow DJ, Rhee L, Ashourian N, Singh B, Sharpe A, Bluestone JA: B7/CD28 costimulation is essential for the homeostasis of the CD4+CD25+ immunoregulatory T cells that control autoimmune diabetes.
Aspinall R, Andrew D: Immunosenescence: potential causes and strategies for reversal.
Biochem Soc Trans
van Meerwijk JP, Marguerat S, Lees RK, Germain RN, Fowlkes BJ, MacDonald HR: Quantitative impact of thymic clonal deletion on the T cell repertoire.
Scollay RG, Butcher EC, Weissman IL: Thymus cell migration: quantitative aspects of cellular traffic from the thymus to the periphery in mice.
Berzins SP, Boyd RL, Miller JF: The role of the thymus and recent thymic migrants in the maintenance of the adult peripheral lymphocyte pool.
Godfrey DI, Kinder SJ, Silvera P, Baxter AG: Flow cytometric study of T cell development in NOD mice reveals a deficiency in alphabetaTCR+CDR-CD8-thymocytes.
J Autoimmun
O'Reilly LA, Healey D, Simpson E, Chandler P, Lund T, Ritter MA, Cooke A: Studies on the thymus of non-obese diabetic (NOD) mice: effect of transgene expression.
Nango K, Inaba M, Inaba K, Adachi Y, Than S, Ishida T, Kumamoto T, Uyama M, Ikehara S: Ontogeny of thymic B cells in normal mice.
Cell Immunol
Adachi Y, Inaba M, Inaba K, Than S, Kobayashi Y, Ikehara S: Analyses of thymic abnormalities in autoimmune-prone (NZW x BXSB) F1 mice.
Serreze DV, Chapman HD, Varnum DS, Hanson MS, Reifsnyder PC, Richard SD, Fleming SA, Leiter EH, Shultz LD: B lymphocytes are essential for the initiation of T cell-mediated autoimmune diabetes: analysis of a new "speed congenic" stock of NOD.Ig mu null mice.
Greeley SA, Katsumata M, Yu L, Eisenbarth GS, Moore DJ, Goodarzi H, Barker CF, Naji A, Noorchashm H: Elimination of maternally transmitted autoantibodies prevents diabetes in nonobese diabetic mice.
Noorchashm H, Greeley SA, Naji A: B-cell deficiency and type 1 diabetes.
Whalen BJ, Weiser P, Marounek J, Rossini AA, Mordes JP, Greiner DL: Recapitulation of normal and abnormal BioBreeding rat T cell development in adult thymus organ culture.
Ridgway WM, Fathman CG: MHC structure and autoimmune T cell repertoire development.
Curr Opin Immunol
Stockinger B: T lymphocyte tolerance: from thymic deletion to peripheral control mechanisms.
Adv Immunol
Zadeh HH, Greiner DL, Wu DY, Tausche F, Goldschneider I: Abnormalities in the export and fate of recent thymic emigrants in diabetes-prone BB/W rats.
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Immunol Rev
Yagi H, Matsumoto M, Nakamura M, Makino S, Suzuki R, Harada M, Itoh T: Defect of thymocyte emigration in a T cell deficiency strain (CTS) of the mouse.
Gleeson PA, Toh BH, van Driel IR: Organ-specific autoimmunity induced by lymphopenia.
Linton P, Thoman ML: T cell senescence.
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–D261,
Gabor MJ, Godfrey DI, Scollay R: Recent thymic emigrants are distinct from most medullary thymocytes.
Rocha B, Dautigny N, Pereira P: Peripheral T lymphocytes: expansion potential and homeostatic regulation of pool sizes and CD4/CD8 ratios in vivo.
Amadori A, Zamarchi R, De Silvestro G, Forza G, Cavatton G, Danieli GA, Clementi M, Chieco-Bianchi L: Genetic control of the CD4/CD8 T-cell ratio in humans.
Gabor MJ, Scollay R, Godfrey CI: Thymic T cell export is not influenced by the peripheral T cell pool.
Penit C, Vasseur F: Expansion of mature thymocyte subsets before emigration to the periphery.
Zhang ZL, Constantinou D, Mandel TE, Georgiou HM: Lymphocyte subsets in thymus and peripheral lymphoid tissues of aging and diabetic NOD mice.
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(Published erratum appears in Immunity 8:531, 1998)
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Shevach EM: Regulatory T cells in autoimmmunity*.
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Green EA, Choi Y, Flavell RA: Pancreatic lymph node-derived CD4(+)CD25(+) Treg cells: highly potent regulators of diabetes that require TRANCE-RANK signals.
Itoh M, Takahashi T, Sakaguchi N, Kuniyasu Y, Shimizu J, Otsuka F, Sakaguchi S: Thymus and autoimmunity: production of CD25+CD4+ naturally anergic and suppressive T cells as a key function of the thymus in maintaining immunologic self-tolerance.
Maloy KJ, Powrie F: Regulatory T cells in the control of immune pathology.
Gonzalez A, Andre-Schmutz I, Carnaud C, Mathis D, Benoist C: Damage control, rather than unresponsiveness, effected by protective DX5+ T cells in autoimmune diabetes.
Address correspondence and reprint requests to D. Mathis, Section on Immunology and Immunogenetics, Joslin Diabetes Center, One Joslin Pl., Boston, MA 02215. E-mail: cbdm@joslin.harvard.edu.
Received for publication 9 July 2002 and accepted in revised form 5 November 2002.
BM, bone marrow; BrdU, bromodeoxyuridine; CFSE, carboxyfluoroscein diacetate succinimidyl ester; FACS, fluorescence-activated cell sorting; FITC, fluorescein isothiocyanate; MHC, major histocompatibility complex; RTE, recent thymic emigrant.

redpajama_set_name: RedPajamaCommonCrawl


These mushroom-themed slippers are so hip (or hippie, I should say!), and would be perfect for that nature-loving family. They are made from 100% cotton fabric, are lined with fleece*, and come with soles of your choice (i.e. either non-slip or soft faux-suede). They will keep baby's feet warm and cozy, and the elastic around the ankle will ensure that they aren't kicked off!


Welcome to the home of Jackie Kirby Art…….
Thank you for taking time to view my Gallery and for your interest in me and my art.
All artwork available to purchase is currently listed in the Art For Sale pages by subject matter.
Prints and merchandise of my work can also be purchased. If there is a particular image that you are interested in obtaining as a print, but it is not currently listed, please contact me via the contact form detailing the picture you are interested in.
If you are interested in commissioning me to complete a unique bespoke piece of artwork for your home, please in the first instance contact me via the contact form with the details of your enquiry.
The Gallery pages list all artwork past and present by the different media used.
For those of you who are interested in painting and drawing, please make sure to make use of the free video tutorials on this website. I am currently developing this to include more tutorials, tips and techniques especially for beginners.
If you have any questions regarding me or my art, please do not hesitate to get in touch. To receive updates on new artwork as it is released, please follow my blog and sign up with your email details at the bottom of any page.


I was reading . Okuyordum.
I was not reading. Okumuyordum.
He was not reading. O okumuyordu.
They were not reading. Onlar okumuyorlardı.
Was he reading? O Okuyor muydu?
Were you reading? Sen okuyor muydun?
Were they reading?, etc. Onlar okuyor muydular?
I was reading a book while you were sleeping. Sen uyu(yor)ken ben kitap okuyordum.
I was watching TV at ten and twelve o'clock yesterday evening. Dün akşam saat on ile on iki arasında TV seyrediyordum.
It was getting colder and colder. Hava gittikçe soğuyordu.
I was playing I was not playing Was I playing?
You were playing You were not playing Were you playing?
He was playing He wasn't playing Was he playing?
We were playing We weren't playing Were we playing?
They were playing They weren't playing Were they playing?
Past Continuous Tense geçmişi anlatan bir zaman olduğu için Simple Past Tense ( –di li geçmiş zaman ) ile birlikte sıklıkla kullanılan '' yesterday, ago, last'' gibi zaman ifadeleri bu zamanda da kullanılabilir.
Alice hurt herself while she ……………………………………………. (skate).
I met my neighbor while I ………………………………………. (walk) home from work.
Sally saw a friend while she …………………………….. (ride) her bicycle along Park St.
Peter fell asleep while he …………………………………… (study).
Bob stepped on Jane's feet while they ………………………………….. (dance) together.
I cut myself while I ……………………………………. (shave).
and Mrs. Brown burned themselves while they …………………………… (bake) cookies.
Tommy had a nightmare while he …………………………….. (sleep) at a friend's house.
He ………………………………….. (talk) with Mary, when Mrs. Smith came in.
They ………………………………… (study) two hours last night.
Jane …………………………………. (sleep) when the telephone rang.
As I …………………………………. (walk) to the lab, I met my friend.
We ………………………………….. (watch) TV last night.
The customer ………………………… (pay) his cheque when he dropped his credit card.
The barber …………………………………. (cut) my hair yesterday.
She ………………………………. (dance) when she hurt her ankle.
It …………………………………. (rain) hard when I got up.
It …………………………………. (rain) hard last night.


text:
Wild by Law: The Rise of Environmentalism and the Creation of the Wilderness Act è un documentario del 1991 diretto da Diane Garey e Lawrence R. Hott candidato al premio Oscar al miglior documentario.

Collegamenti esterni

Film documentari canadesi
Film documentari statunitensi

redpajama_set_name: RedPajamaWikipedia


An ever in demand collection of flannel skirts includes the attractive Staunch Black Pencil Skirt which comes in the most attractive textured base. The skirt with its all coordinated texture has an attractive black hue which stands forth as a truly top designer variant in its trues form. The super fit provided by the excellently stitched skirt is simply unparralled in or by any other variant.


← The Millennium Falcon Returns!
Looks like Mortal Kombat is back! Interesting trailer, although I'm not sure how much you can tell about the game based on this clip. If the game is anything like the clip shows, it could be pretty amazing. Decide for yourself and view it below.
This entry was posted in Trailers, Video Games. Bookmark the permalink.


text:
Here is 25 Kickass Random Facts List #171.
1-5 Kickass Random Facts
1. Alan Tudyk, Adam Baldwin, and Nathan Fillion all voiced marines in Halo 3. Their characters were given personalities resembling the characters they played in Firefly. – Source
2. The Speaker of the House during Watergate scandal could have stalled the VP confirmation and impeached Nixon, which would have made himself POTUS, but he concluded that, as a Democrat, he had no right to a Presidency that the American people had entrusted by election to a Republican. – Source
3. The shortest film nominated for an Oscar was under 2 minutes and a stop animation about making guacamole named "Fresh Guacamole by PES." – Source
4. Hazel Ying Lee, a Chinese-American WWII pilot, had two forced landings. One landing took place in a Kansas wheat field. A farmer with a pitchfork in hand chased her around the plane while shouting to his neighbors that the Japanese had invaded Kansas. – Source
5. Every night the Disney World maintenance staff hoses down the park with over 120 degree hot water so that all the gum will melt off of the mid-ways and cue-lines. – Source
6-10 Kickass Random Facts
6. The verb "to snipe" originated in the 1770s among soldiers in British India where a hunter skilled enough to kill the elusive snipe bird was dubbed a "sniper." – Source
7. In 1912, criminologist Arthur MacDonald suggested making it a misdemeanor to name assassins publicly, stating that "This would lessen the hope for glory, renown or notoriety, which is a great incentive to such crimes." – Source
8. There are still four teams in the NFL that, in 50 years, have never once been to a Super Bowl. They are the Cleveland Browns, Detroit Lions, Houston Texans, and Jacksonville Jaguars. – Source
9. Google avoided $2 billion in taxes in 2012, by transferring royalty payments from its Irish and Dutch subsidiaries to a Bermuda unit, which was simply headquartered in a local law firm. – Source
10. Jesus' foreskin is called the "Holy Prepuce" and several European churches have claimed to be in possession of it and that it has miraculous powers. – Source
11-15 Kickass Random Facts
11. In response to the rioting and looting that followed the San Francisco 1906 earthquake, Mayor Eugene Schmitz authorized federal troops and local officers to "kill any and all persons found engaging in looting or in the commission of any other crime." – Source
12. Joseph Smith, founder of Mormonism, was killed by an angry mob while he was incarcerated. Smith had tried to suppress a local that had exposed Smith's practice of polygamy (he had more than 25 wives), implying that he used his religion as a pretext to seduce and marry unassuming women, including the wives of some of his closest associates. – Source
13. Under the clergy letter project, over 13,000 Christian clergies have signed an open letter affirming that evolution and god can coexist. – Source
14. Mr. Burns' telephone greeting in The Simpsons, "Ah-Hoy!" was actually how Alexander Graham Bell thought people should answer the telephone when he invented it. – Source
15. Dry red kidney beans can be toxic if they are cooked in a slow cooker, which doesn't get hot enough to get rid of phytohaemagglutnin(Kidney Bean Lectin), causing severe symptoms. – Source
16. While not strictly legal, starting in the 1690s, British husbands used to sell their wives to end bad marriages. It was usually an auction announced by newspaper ad, to which the wife was led by a rope around her neck. Often the buyer was pre-arranged and the sale was a form of symbolic separation. – Source
17. Nairobi, Kenya has a thriving country music scene because Kenyans "can identify with the stories in the songs." Country Music CDs make up to 15% of all music sold there. – Source
18. New York Giants linebacker Lawrence Taylor would routinely send prostitutes to his opponents' hotel rooms the night before a game in an attempt to tire them out. – Source
19. Alexander Turner, father of American poet Daisy Turner, was a slave who escaped from his Virginia plantation during the Civil War, joined the Union Army, and guided his regiment back to the plantation where he killed his former overseer. – Source
20. Refried beans are only fried once. – Source
21. San Marino is the oldest surviving Republic, gaining independence from the Roman Empire in 301 A.D. – Source
22. In 1973, a Libyan passenger jet having navigational problems wandered into Israeli airspace. When two Israeli F-4s intercepted the jet and instructed the crew to land, they refused because of bad Libyan-Israeli relations at the time. The jet was then shot down, killing 108 people. – Source
23. The township of Thierville, France suffered no losses in country's last five wars: WW1, the Franco-Prussian War, WW2, the First Indochina War, and the Algerian War. All the soldiers who took part in these wars returned home. – Source
24. A recent study found that 43% of married people don't know how much money their spouse makes, and 10% of those who guessed wrong were off by more than $25,000. – Source
25. In 1989 a Los Angeles woman received a phone call from her brother-in-law saying that her ex-husband was coming over to kill her. She dialed 911 but was told nothing could be done unless he showed up at the house. Within 15 minutes he arrived, killing her and 4 others – Source
Tags Random
Torbernite [Educational Album]
15 Interesting Historical Photos – Part 2
25 Kickass Random Advice List #1
25 Interesting Facts About World War 2 – Part 4
23 Facts About Napoleonic Wars
39 Facts About Gulf War
Valannin says:
#8: Of course the Jacksonville Jaguars haven't been to a Superbowl in the last 50 years. They've only existed for 20 years. And the the Houston Texans have only been around since 2002. Technically, the Ravens, who were for all intents and purposes, WERE the Cleveland Browns, appeared in, and won, the Superbowl, twice.


Merry Christmas! Today, we're talking The Santa Clause! We discuss our first impression of the film then our second impression after seeing it again this week. In our second act (15:22), we discuss our favorite movies from 2016.


These are mildly spicy pumpkin muffins that are great for breakfast or a snack.
In a large bowl, mix together flour, brown sugar, baking powder, baking soda, salt, cinnamon, ginger, cloves nutmeg, and allspice.
Make a well in the center of the dry ingredients; add pumpkin mixture and stir just to combine. stir in nuts and raisins.
Grease 12 muffin cups or use paper liners. Fill the cup 3/4 -7/8 full (Use a #12 scoop which is 1/3 cup). Theses muffins will be nicely rounded when baked.
Bake 20 - -25 minutes or until a tooth pick inserted in the center comes out clean.
Remove from the oven and cool 5 minutes before removing from the cups. Cool completely and store in an airtight container at room temperature.
Note: If you double the recipe use the whole 15oz. can of pumpkin, it's a bit more but will work just fine.


Some of you may have noticed I've been busy doing everything else but blogging.
What I do for living (and past-time) is build/maintain/hack systems. Some of them running web applications such as this cloud Azure VM Nginx/PHP/PostgreSQL -thingie.
Sometimes just starting a tedious task of fixing/updating/repairing a server instead of playing The Division or Far Cry doesn't do the trick for me. I choose to play for entertainment and not update the server.
This blog has been running on 5.6 for years and to get that updated, I had to re-asses configurations to get 7.2 running. Not an easy task, but had to be done to go forward.
Most of you have never realized, I wouldn't touch Wordpress with a 9 foot pole. This blog runs on Serendipity (aka s9y).
Previous version of S9y 2.0 wouldn't run correctly with PHP 7.2, so I had to go for this upgrade too.
This blog gets reasonable Page Rank from Google, but it really suffers from not being very good when displayed on a mobile device.
See #1 above. This is what I do for living. It should be not a complicated task to strap Bootstrap boot on top of the theme.
What hasn't changed: I'm still in Microsoft Azure, running CentOS 7.
The mobile version of Brown Paper is still under work in my lab VM, but it will be out soon. I promise!


Onzie's Gypsy Pant is made from free flow jersey fabric, super light and feels great with roll down waistband. Baggy leg pant moves freely with your movement.
K-DEER's Yoga Tank Top with a built in soft low-impact shelf bra. Soft seams at the neck, arms and hem provide one less distraction in your active life.