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McNally, DO, FACP, FACG University Colorado School of Medicine Center for Human Simulation Aurora, Colorado 80045[Pronio A, Montesani C, Butteroni C, Vecchione S, Mumolo G, Vestri A, Vitolo D, Boirivant M. \[% %    Probiotic Administration in Patients with Ileal Pouch-Anal Anastomosis for Ulcerative Colitis Is Associated with Expansion of Mucosal Regulatory Cells. Inflamm Bowel Dis. 2008;14:1-7PH ! Sh. Introduction8Pouchitis is the #1 long term complication after Ileal Pouch Anal Anastomosis (IPAA) for ulcerative colitis (20% at 1 yr & 50% at 10 yr).1 Pouchitis is believed to be a result of an unstable microflora: ! lactobacilli & bifidobacteria ! anaerobes/aerobes ratio ! Clostridia perfringens Antibiotics (Ciprofloxicin, Metronidazole, Rifaximin) are an effective acute therapy for pouchitis, but relapse is high.1,2ZSZ}Z @ 3y 2 s (   Sm/ IntroductionThe probiotic, VSL#3, has been shown to regulate mucosal response, by reductions in mucosal levels of interleukin (IL-8), and cytokines IL-1 & INF-. 3,4,5 4 Mechanisms of Inflammation in IBD & Pouchitis: Regulatory T cells (Tregs = CD4+CD25+ T lymphocytes) are important in down regulation of inflammation, but are relatively ! in the inflammatory states of IBD and pouchitis -- inverse relationship. Lamina Propria Mononuclear Lymphocytes (LPML) & Peripheral Blood Mononuclear Cells (PBMC) directly correlate with inflammation. IL-1 cytokine directly correlates with inflammation. Foxp3 is a specific functional marker of Treg (! in inflammation).\PP "*2"" NwZDAimg To determine if the probiotic, VSL#3, would suppress clinical pouchitis (pouchitis disease activity index) among patients with IPAA for Ulcerative colitis when compared to untreated controls. Compare the effect of the probiotic, VSL#3, on mucosal markers of inflammation among patients with IPAA for Ulcerative Colitis when compared to untreated controls. TfPPP"e:   Study Design:Pts with IPAA for UC from 1984 to 2005 None had active pouchitis at enrollment Open-label randomization to VSL#3 or No Rx Study Medication VSL#3 (450 billion cfu/pkg) 2 packet/d Cohort VSL#3 n=16 Control n=12 P1 Evaluations: T 3, 6,12 mo PDAI (>7 = pouchitis) Endoscopy Biopsy of Lamina Propria: mononuclear cells (LPMC s), IL-1, CD25high T cell, LAP(+)T cells, & Foxp3 mRNA Blood: peripheral mononuclear cells (PBMC)f P#W"*&",XR/,G  Lactobacillus (20185A), Bifidobacteria (24704B), and Streptoccus (26702B) Images copyright Dennis Kunkel Microscopy, Inc. Reproduced with permission. P> Mi-EClinical Inflammatory Marker Pouchitis Disease Activity Index (PDAI) "F() n0Biological Inflammatory Markers (Immune staining for PBMC and LPMC Conjugated mouse antihuman CD3 Conjugated mouse antihuman CD4 Conjugated mouse antihuman CD25 Goat anti-LAP (specific CD3/CD4 populations) 6""o1Biological Inflammatory Markers (:FOX3p m-RNA and IL-1 Expression of each gene transcript in biopsies was analyzed by real time reverse-transcription PCR using primer sequences and antisense ,/Baseline Demographic & Clinical Characteristics00(/Baseline Demographic & Clinical Characteristics00( PResults: significant ! PDAI in VSL#3 pts,)%&%v4RResults: Significant ! IL-1 in VSL#3 pts$*"p2^Results: Significant ! in CD25high in VSL#3 ptsB0 "*"/ Percentage of CD25high T lymphocytes in the CD3/ CD4 gated cell population of lamina propria mononuclear cells in patients with IPAA for ulcerative colitis randomized to (A) VSL#3 (*P 0.04 versus baseline and at 3 months; **P 0.012 versus baseline and at 6 months) or (B) no treatment (control). B0 )HZ {@ ConclusionsVSL#3 450 billion CFU bid decreased PDAI in patients with IPaA for UC p < 0.005 VSL#3 decreased peripheral and mucosal immune response seen in pts wit IPAA for UC. :h}CReviewer CommentsPronio, et al, have demonstrated several immunologic mechanisms by which VSL#3 suppresses pouchitis. ! Regulatory T cells (Tregs = CD4+CD25+ T lymphocytes) ! Foxp3 a specific and functional marker of Treg. ! Concentrations of Lamina Propria Mononuclear Lymphocytes (LPML) & Peripheral Blood Mononuclear Cells (PBMC) ! IL-1 an inflammatory cytokine that directly correlates with inflammation.0eZ'Ze'"^uITFEReviewer CommentsYPronio, et al, have demonstrated VSL#3 is more effective than no treatment (control) in suppressing pouchitis as measured by PDAI score. The findings of Pronio, et al, emphasize the need to support further research of the immunologic mechanisms responsible for probiotics to influence gut microbiotica, immunity and gastrointestinal disorders.ZZZ6 +/jrs 9& 0` ̙33` ` ff3333f` 333MMM` f` f` 3>?" dU,x? 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Oa( 2_Vienna980 zr@ (    00 P    P*    0("     R*  d  c $ ?    0%  0  RClick to edit Master text styles Second level Third level Fourth level Fifth level!     S  6* _P   P*    6. _   R*  H  0޽h ? 3380___PPT10.Npfu  x(  x x 0Ȣ P    X*  x 0Ħ     Z*  x 6  _P   X*  x 6 _   Z* H x 0޽h ? 3380___PPT10.p0 <(  ~  s *4n  ~  s *4@p   H  0޽h ? 33___PPT10i.8dW+D=' = @B +}0  $(  x  c $Xg4`  x  c $0h40P     0|i P NDepartments of Surgery, Internal Medicine & Experimental Medicine, Rome, Italy"O|( _OH  0޽h ? Oa(___PPT10i.e+D=' = @B + 0 (  x  c $w4      0xp`PD<4___PPT9 ZShen B. Managing Pouchitis. AM J Gastroenterol. 2007;102:S60-64. Kiehne K, et al. Defensin expression in chronic pouchitis in pts with UC or FAP. World J Surg. 2006;21:1065-1062.&m _F H  Y8X1  Z(xaxao ?P  1Pronio A, et al . 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Inflamm Bowel Dis. 2008:14:1-7.2Z2$ ]  Npxaxa1?P gStudy Design: VSL#3 450 billion CFU/dose Lactobacilus (4 sp), Bifidobacteium (3 sp), Streptoccus (1 sp)hh>)   R   C *APicture1 R   C *APicture2 R   C *APicture3 H  0޽h ? 33___PPT10i.0(\+D=' = @B +8 0  886<7(  r  S Ir4P  r h3   # #">2  p#    `Proaoa 1?"6@ NNN?Nt MScore  @`(    `[roaoa 1?"6@`NNN?Nw t R   @`0    `]roaoa 1?"6@ NNN?Nw  ZClinical Parameter  @`    `llroaoa 1?"6@ NNN?Nt I0  @`#   `froaoa 1?"6@`NNN?Nw t Musual  @`$   `xwroaoa 1?"6@ NNN?Nw  NStool#  @`   ` roaoa 1?"6@ NNN?Nt I1  @`%   `Proaoa 1?"6@ NNN?Nw t Opresent  @`(   ``roaoa 1?"6@ NNN?Nw  R   @`   `̜roaoa 1?"6@ NNN?Nt  I0  @`$   `@roaoa 1?"6@ NNN?Nw t Nabsent  @`+   `roaoa 1?"6@ NNN?N w  U Fever (>100F)  @`   `roaoa 1?"6@ NNN?Nt   I2  @`#   `roaoa 1?"6@ NNN?Nw t  Musual  @`(   `roaoa 1?"6@ NNN?N w  R   @`   `roaoa 1?"6@ NNN?Nt   I1  @`(   `Lroaoa 1?"6@ NNN?Nw  t  R occasional   @`(   `(roaoa 1?"6@ NNN?N w  R   @`   `roaoa 1?"6@ NNN?NtX   I0  @`"   `roaoa 1?"6@ NNN?Nw X t  Lnone  @`2   `uoaoa 1?"6@ NNN?NX w   \Fecal urgency/Cramps  @`   `Luoaoa 1?"6@ NNN?Ntd X  I1  @`#    `Tuoaoa 1?"6@`NNN?Nw d tX  MDaily  @`( !  `d#uoaoa 1?"6@ NNN?Nd w X  R   @` "  `%uoaoa 1?"6@ NNN?Ntnd  I0  @`" #  `<5uoaoa 1?"6@`NNN?Nw ntd  LNone  @`- $  `|>uoaoa 1?"6@ NNN?Nnw d  WRectal Bleeding  @` %  `Guoaoa 1?"6@ NNN?Ntyn I2  @`: &  `Quoaoa 1?"6@`NNN?Nw ytn d>3/d.  @`( '  `t[uoaoa 1?"6@ NNN?Nyw n R   @` (  `@duoaoa 1?"6@ NNN?Nty I1  @`# )  `Hmuoaoa 1?"6@`NNN?Nw ty M1-2/d  @`( *  `Xvuoaoa 1?"6@ NNN?Nw y R   @`B + Zoaoao ?"0@NNN?NB , Toaoa1 ?"0@NNN?NyyB - Toaoa1 ?"0@NNN?NnnB . Toaoa1 ?"0@NNN?Nd d B / Toaoa1 ?"0@NNN?NX X B 0 Toaoa1 ?"0@NNN?N  B 1 Toaoa1 ?"0@NNN?N  B 2 Toaoa1 ?"0@NNN?N  B 3 Toaoa1 ?"0@NNN?NB 4 Zoaoao ?"0@NNN?NB 5 Zoaoao ?"0@NNN?NB 6 Toaoa1 ?"0@NNN?Nw w B 7 Toaoa1 ?"0@NNN?NttB 8 Zoaoao ?"0@NNN?NB 9 Toaoa1 ?"0@NNN?N`B : 01 ?> ; T |uxaxao?`` 4Sandborn WJ, et al: Pouchitis after ileal pouch-anal anastomosis: PDAI. Mayo Clin Proc. 1994;69:409-415. Pouchitis is defined as a total score of e"7.H  J1 < Z\uxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$ H  0޽h ? Oa(___PPT10i.2+D=' = @B + 0 0o(  x  c $8Ru4@  u ~  s *\u4 u 1  Z0 uxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$ H  0޽h ? Oa(___PPT10i.2+D=' = @B + 0 @o(  x  c $`u4p@  u ~  s *8u4` u 1  Z8uxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$ H  0޽h ? Oa(___PPT10i.2+D=' = @B + 0 !6@((  @~ @ s *u4P`    u 1 d 5@ #"*_NONPT u @ <u?1d P 1.6+/- 1.6   @` @ <\u?j1d O 1.5+/-0.9   @` @ <u?1jd Q PDAI at T0   @` @ <u? 1 G5  @` @ <u?j 1 G5  @` @ <  RB   s *D>p p LB   c $D  RB   s *D>`  ` RB  s *D>P PRB  s *DjJP0 0 ` LB  c $D    0wP 5VSL#3 2  0"w  7control 2  0L%w0@ 6P=0.03 2RB  s *DjJp `` H  0޽h ? Oa(___PPT10i.Э,+D=' = @B +  0 7/(  ~  s */w4 P  w   0 C xoxaxaA! ?14? 0   w~  s *pw4  w 1  Z6wxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$    0<9w P  Z*P 0.046 2  0w  \ **P 0.0126 2H  0޽h ? Oa(___PPT10i.E-+D=' = @B + 0   `X5 (  X  X0 S ~oxaxaA# ?14?@   w X s *ddw4P   w x8Results: Foxp3 ! VSL#3 pts$ " X s *hw4p   w BChange in Foxp3 mRNA isolated from mucosal biopsies of consecutive observations in patients with IPAA for ulcerative colitis randomized to VSL#3 or no treatment (control) quantified by real time-PCR (*P 0.008 versus control).D Z 1 X Zzwxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$ R X s * p P X  X 0 @` LB  X c $D RB X s *D> @` LB X c $D p0 LB X c $D ` RB X s *D> p RB X s *D>` p ` RB X s *D>@` X 0wpW 5VSL#3 2 X 00wp W 7control 2 X 0@wppW 6P=0.08 2LB X@ c $D` H X 0޽h ? Oa(___PPT10i.j-+D=' = @B + 0 P h](  hr h S w4P  w r h S w4 w 1 h Zwxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$ H h 0޽h ? Oa(___PPT10i.,+D=' = @B + 0  t](  tr t S tw4P  w r t S Lw4 w 1 t Ztwxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$ H t 0޽h ? Oa(___PPT10i.,h+D=' = @B + 0  i(  x  c $@w4P  w x  c $w40 w 1  Zwxaxao ?P  1Pronio A, et al . Inflamm Bowel Dis. 2008:14:1-7.2Z2$ H  0޽h ? Oa(___PPT10i.,h+D=' = @B +0 <(  <^ < S      < c $ 0   CD25high T lymphocytes in the CD3+/CD4+ gated pool LAP = latency-associated peptide positive T lymphocytes in C3+/C4+ gated cell populationH < 0޽h ? 3380___PPT10.MS1 0 ((  ^  S      c $; 0    H  0޽h ? 3380___PPT10. .0 /(  ^  S      c $@ 0   %immunosuppressionH  0޽h ? 3380___PPT10. 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McNally, DO, FACP, FACG.-@Arial-. @2 G&University Colorado School of Medicine.-@Arial-. 02 Q"Center for Human Simulation .-@Arial-. (2 Z+Aurora, Colorado 80045.-՜.+,0    On-screen Show 6 Arial Wingdings 2_Vienna98Microsoft Graph ChartLiterature Review\Pronio A, Montesani C, Butteroni C, Vecchione S, Mumolo G, Vestri A, Vitolo D, Boirivant M. Introduction IntroductionAimStudy Design:Slide 7FClinical Inflammatory Marker Pouchitis Disease Activity Index (PDAI) Biological Inflammatory Markers Biological Inflammatory Markers0Baseline Demographic & Clinical Characteristics0Baseline Demographic & Clinical Characteristics+Results: significant ↓ PDAI in VSL#3 pts-Results: Significant ↓ IL-1β in VSL#3 pts2Results: Significant ↑ in CD25high in VSL#3 ptsResults: Foxp3 ↑ VSL#3 pts ConclusionsReviewer CommentsReviewer Comments  Fonts UsedDesign TemplateEmbedded OLE Servers Slide Titles%_ Susan McNevinSusan McNevin  !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~Root EntrydO)PicturesTCurrent UserSummaryInformation('PowerPoint Document(ADocumentSummaryInformation8