1. The Absence of a Microbiota Enhances TSLP Expression in Mice with Defective Skin Barrier but Does Not Affect the Severity of their Allergic Inflammation 
Laura J. Yockey, Shadmehr Demehri, Mustafa Turkoz, Ahu Turkoz, Philip P. Ahern, Omar Jassim, Sindhu Manivasagam, John F. Kearney, Jeffrey I. Gordon, Raphael Kopan 
Journal of Investigative Dermatology 
Volume 133, Issue 12, Pages (December 2013)
DOI: /jid
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2. Figure 1
Epidermal and hair follicle structure are unaffected in germ-free (GF) mice compared with their conventionally raised (CONV-R) counterparts. Hematoxylin and eosin (H&E) staining at (a) P9 and (c) P30 shows skin morphology and inflammation levels in GF and CONV-R RBPjCKO and wild-type mice. Bar, 50μm. Immunocytochemical staining for keratin-14 (K14), involucrin (INV), and filaggrin (FLG) at (b) P9 and (d) P30 shows skin architecture in GF and CONV-R wild-type and RBPjCKO animals. Both GF and CONV-R RBPjCKO animals exhibit similar barrier defects. Bar, 10μm.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Skin inflammation and atopic dermatitis (AD)–like disease persist in germ-free (GF) RBPjCKO animals. (a) Conventionally raised (CONV-R) and GF P80 mice. Scale bar, 1cm. Hematoxylin and eosin (H&E), CD45 immunohistochemistry, and toluidine blue staining reveal skin inflammation and mast cell infiltration in both CONV-R and GF wild-type (Wt) and RBPjCKO (Mut) mice. Bar, 50μm. (b) Quantification of mast cells viewed in three randomly selected microscopic fields at × 200 (CONV-R Wt, n=4; CONV-R Mut, n=15; GF Wt, n=4; GF Mut, n=15). (c) Serum IgE levels in P80 Wt mice (CONV-R, n=10; GF, n=6) and RBPjCKO animals (CONV-R, n=23; GF, n=23). (d) Flow cytometry gated on CD4+ T cells in the spleen to detect effector (CD44hi CD62Llo) and naive (CD44lo CD62Lhi) cells in RBPjCKO (CONV-R, n=9; GF, n=9) and Wt mice (CONV-R, n=4; GF, n=4). A compilation of results from four independent experiments is shown. (e) Flow cytometry gated on CD4+ T cells detects regulatory T cells (Treg, Foxp3+) in RPBjCKO (GF and CONV-R, n=6) and Wt mice (CONV-R and GF, n=3). A compilation of three independent experiments is shown. Asterisks mark statistical significance; *P<0.05, **P<0.01, ***P<0.001 (unpaired Student’s t-test).
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Asthma in germ-free (GF) RBPjCKO mice is not more severe but thymic stromal lymphopoietin (TSLP) levels are higher than in their conventionally raised (CONV-R) counterparts. (a) Tslp mRNA levels in P9 CONV-R wild-type (Wt; n=6), CONV-R RBPjCKO (n=7), GF Wt (n=10), and GF RBPjCKO (n=10) mice. (b) Serum TSLP levels in P9 RBPjCKO (CONV-R, n=7; GF, n=10), P30 (CONV-R, n=7; GF, n=14), and P80 (CONV-R, n=19; GF, n=24). (c) Hematoxylin and eosin staining of the lungs after ovalbumin (OVA) challenge at 6–8 weeks shows increased inflammatory infiltrate (arrows) in the RBPjCKO mice. Bar, 50μm. (d) White blood cell count in bronchial alveolar lavage fluid (BAL) of Wt (CONV-R, n=6; GF, n=10) and RBPjCKO (CONV-R, n=8; GF, n=8) mice. (e) The percentage of eosinophils, macrophages, lymphocytes, or neutrophils in BAL from RBPjCKO (CONV-R, n=11; GF, n=9) or Wt (CONV-R, n=3; GF, n=4) mice. The differences between RBPjCKO and wild-type animals remain evident in a GF state. *P<0.05, **P<0.01, ***P<0.001 (unpaired Student’s t-test). PBS, phosphate-buffered saline.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
B-cell expansion persists in germ-free (GF) RBPjCKO animals. (a) White blood cell (WBC) counts in P30 RBPjCKO mice (conventionally raised (CONV-R), n=7; GF, n=14) and P80 (CONV-R, n=33; GF, n=43). (b–c) Spleen size mirrored WBC levels, reported as a percentage of body weight (wild-type (Wt), CONV-R n=13; GF, n=19; Mut CONV-R, n=34; GF, n=43). Bar, 1cm. (d) B-cell (B220+ CD45+), myeloid (Ly6G+), and T cell (CD3+) expansion in 10- to 12-week-old mice, quantified by flow cytometry (Wt CONV-R, n=5; GF, n=4; RBPjCKO CONV-R, n=13, GF, n=12). (e) The percentage of immature B220+ cells (IgM+ IgDlo) in GF RPBjCKO animals. Data compiled from five independent experiments (Wt CONV-R, n=5, GF, n=4; RBPjCKO CONV-R and GF, n=11). (f) Granulocyte colony-stimulating factor (G-CSF) levels in serum from GF and CONV-R mice (Wt CONV-R, n=5, GF, n=10; RBPjCKO CONV-R, n=11; GF n=14). (g) Anti-phosphorylcholine (anti-PC) IgM levels (Wt CONV-R, n=6; GF, n=10; RBPjCKO CONV-R, n=9; GF n=12). (h) RBPjCKO mice show lower hematocrit than Wt counterparts (Wt CONV-R, n=13; GF, n=17; RBPjCKO CONV-R, n=34; GF n=43). (i) Serum lactate dehyrogenase (LDH) levels (Wt CONV-R and GF, n=6; RBPjCKO CONV-R and GF, n=12). (j) Serum haptoglobin levels (Wt CONV-R and GF, n=6; RBPjCKO CONV-R and GF, n=12). *P<0.05, **P<0.01, ***P<0.001 (unpaired Student’s t-test).
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions