Journal: Journal of the American Heart Association

Article Title: Elastin‐Specific Autoimmunity in Smokers With Thoracic Aortic Aneurysm and Dissection is Independent of Chronic Obstructive Pulmonary Disease

doi: 10.1161/jaha.118.011671

Figure Lengend Snippet: Figure 1. EFs-specific CD4+ T cell response in TAA/TAD. A, Representative intracytoplasmic cytokine of CD4+ T cells (59105) isolated from peripheral blood of healthy controls (HC) or thoracic aortic aneurysm and dissection (TAA/TAD) patients were co-cultured with irradiated CD1a+/CD14+ APCs (59104) with EFs (3, 10, or 30 lg/mL) or no EFs. After 3 days, interferon-c and IL-17A producing CD4+ T cells were detected using flow cytometry. Relative abundance and mean fluorescent intensity (MFI) shown in the right lower quadrants. Data are representative of 4 to 5 subjects in each group. B, Representative histogram of EFs- specific CD4+ T cell proliferation. CFSE-labeled CD4+ T cells (59105) were cultured with APCs (59104) in presence of EFs (30 lg/mL) for 3 days. 2% phytohemagglutinin was used as positive control. Data are representative of 4 to 5 subjects in each group. C, Supernatants from the same co-culture conditions described in (A); concentration of interferon-c, IL-1b, IL-10, IL-17, and IL-13 were measured and plotted as fold change over nil stimulation. Each dot represents a data point from an individual subject. *P<0.05, and ***P<0.001 as determined by the Mann–Whitney test; HC: n=37; TAA/TAD n=49. APCs indicates antigen presenting cells; CFSE, carboxyfluorescein succinimidyl ester; EFs, elastin fragments; HC, healthy control; IFN-c, interferon-c; IL, interleukin; MFI, mean fluorescent intensity; PHA, phytohemagglutinin; TAA/TAD, thoracic aortic aneurysm and dissection.

Article Snippet: Approximately 10 to 309106 peripheral blood mononuclear cells were isolated from whole blood and were labeled with MicroBeads conjugated to anti-CD4 antibody (Miltenyi Biotec) followed by AutoMACS (Miltenyi Biotec) positive selection.

Techniques: Isolation, Dissection, Cell Culture, Irradiation, Cytometry, Labeling, Positive Control, Co-Culture Assay, Concentration Assay, MANN-WHITNEY, Control

Journal: Journal of the American Heart Association

Article Title: Elastin‐Specific Autoimmunity in Smokers With Thoracic Aortic Aneurysm and Dissection is Independent of Chronic Obstructive Pulmonary Disease

doi: 10.1161/jaha.118.011671

Figure Lengend Snippet: Figure 2. EFs-specific CD4+ T cell expression of interferon-c and IL-1b are independent of airway obstruction in TAA/TAD. Expression of interferon-c (A) and IL-1b (B) in response against EFs was assessed in healthy smoker and non-smoker HC (n=37), and smoker and non-smoker TAA/TAD patients (n=49) or TAA/ TAD without chronic obstructive airway disease (COPD; n=37). Each dot represents a data point from an individual subject. *P<0.05, **P<0.01, as determined by the Kruskal–Wallis test with Dunn post-test for multiple comparisons. COPD indicates chronic obstructive airway disease; EFs, elastin fragments; HC, healthy control; IFN-c, interferon-c; IL, interleukin; NS, not significant; TAA/TAD, thoracic aortic aneurysm and dissection.

Article Snippet: Approximately 10 to 309106 peripheral blood mononuclear cells were isolated from whole blood and were labeled with MicroBeads conjugated to anti-CD4 antibody (Miltenyi Biotec) followed by AutoMACS (Miltenyi Biotec) positive selection.

Techniques: Expressing, Control, Dissection

Journal: Journal of the American Heart Association

Article Title: Elastin‐Specific Autoimmunity in Smokers With Thoracic Aortic Aneurysm and Dissection is Independent of Chronic Obstructive Pulmonary Disease

doi: 10.1161/jaha.118.011671

Figure Lengend Snippet: Figure 3. EFs-specific CD4+ T cell expression of interferon-c, but not IL-1b response is dependent on history of smoking in TAA/TAD. Expression of interferon-c (A) and IL-1b (B) in response against EFs was assessed in HC non-smoker (n=18), smoker (former n=13, current n=6), TAA/TAD non-smoker (n=15), TAA/TAD smokers (former n=20, current n=14). Each dot represents a data point from an individual subject. For 2 groups comparison, Mann–Whitney test was used. For multiple comparisons in >3 groups, the Kruskal–Wallis test with Dunn’s post-test for multiple comparisons was used. *P<0.05, **P<0.01. EFs indicates elastin fragments; HC, healthy control; IFN-c, interferon-c; IL, interleukin; NS, not significant; TAA/ TAD, thoracic aortic aneurysm and dissection.

Article Snippet: Approximately 10 to 309106 peripheral blood mononuclear cells were isolated from whole blood and were labeled with MicroBeads conjugated to anti-CD4 antibody (Miltenyi Biotec) followed by AutoMACS (Miltenyi Biotec) positive selection.

Techniques: Expressing, Comparison, MANN-WHITNEY, Control, Dissection

Journal: Blood

Article Title: Induction of acute GVHD by sex-mismatched H-Y antigens in the absence of functional radiosensitive host hematopoietic-derived antigen-presenting cells

doi: 10.1182/blood-2011-10-384057

Figure Lengend Snippet: Single H-Y Ag-specific T cells induce CD8+ and CD4+ T cell–mediated GVHD. (A) Survival in CD8+-mediated GVHD after infusion of 1 × 106 CD8+ T cells from MataHari donors. Data are from 1 of 4 similar experiments. (B-E) GVHD analysis on day 5. Data are from 1 of 3 similar experiments. (B) Histopathologic analysis of liver (i-ii) and GI tract (iii-iv) by H&E stain (left) and scores (right). (C) Serum levels of TNF-α, IFN-γ, and IL-17A (ND indicates not detected). (D) Allo-H-Y Ag-specific donor CD45.2+CD8+ T-cell expansion in spleen. (E) Donor CD45.2+CD8+IFN-γ+ in spleen with representative histogram of IFN-γ expression gated by CD45.2+CD8+ T cells (left) and the absolute number of CD45.2+CD8+IFN-γ+ T cells in spleen (right). (F-G) Survival CD4+ GVHD. Data shown are from 1 of 3 similar experiments.

Article Snippet: 10 Briefly, splenic T cells were enriched by autoMACS with anti-CD4, -CD8 microbeads (Miltenyi Biotec).

Techniques: Staining, Expressing

Journal: Blood

Article Title: Induction of acute GVHD by sex-mismatched H-Y antigens in the absence of functional radiosensitive host hematopoietic-derived antigen-presenting cells

doi: 10.1182/blood-2011-10-384057

Figure Lengend Snippet: Presentation by donor and host nonhematopoietic APCs. (A) Experimental schema: B6♂ animals underwent thymectomy and were used for generation of (class II−/−♀→B6♂) chimeras. Three months later, these chimeras were irradiated and transplanted with either B6 or Marilyn T cells and monitored for (B) survival. Data are from 2 combined experiments. (C) Donor APCs alone stimulate GVH responses. Experimental schema: TCD BM from either WT or class II−/− B6♀ were transplanted along with WT or Marilyn T cells into lethally irradiated BALB/c males. (D) Total donor H2Kd+CD4+ T-cell expansion, H2kd+CD4+IFN-γ+ T-cell number, and serum IFN-γ concentration from day 7. Data are from 1 of 2 similar experiments. (E-I) Induction of GVHD in the absence of donor and radiosensitive host hematopoietic-derived APCs. (E) Experimental schema: B6Ly5.2 female and male animals were lethally irradiated, treated with anti–natural killer (NK) Ab and transplanted with either WT B6 or β2m−/− BM. Three months later, these chimeras were transplanted with TCD BM from either WT or β2m−/− B6♀ animals along with MataHari T cells. The animals were monitored for (F) survival and (G) clinical score. Data from 1 of 3 similar experiments are shown. (H) Donor CD45.2+CD8+ T cells expansion in spleen and (I) histopathology scores of liver and GI tract. Data from 1 of 2 similar experiments are shown.

Article Snippet: 10 Briefly, splenic T cells were enriched by autoMACS with anti-CD4, -CD8 microbeads (Miltenyi Biotec).

Techniques: Irradiation, Concentration Assay, Derivative Assay, Histopathology

Journal: Blood

Article Title: Induction of acute GVHD by sex-mismatched H-Y antigens in the absence of functional radiosensitive host hematopoietic-derived antigen-presenting cells

doi: 10.1182/blood-2011-10-384057

Figure Lengend Snippet: Alloantigen presentation and GVHD in the absence of radiosensitive host hematopoietic-derived APCs. (A) Experimental schema: β2m−/−♂→B6♂ chimeras were generated as described in “Methods.” They were then irradiated with 9 Gy and transplanted with TCD BM from β2m−/− animals along with T cells from B6♂ or BALB/c♀ donors and analyzed for donor T-cell expansion. (B) Total donor CD45.1+CD8+ or H2kd+CD8+ T cells expansion. (C) Donor CD8+IFN-γ+ T cell number and (D) serum levels of IFN-γ. Data from 1 of 2 similar experiments are shown. (E) GVH responses induced by host nonhematopoietic APCs depend on T-cell precursor frequency. Survival and GVHD score. Data from 1 of 2 similar experiments are shown. (F-H) Male nonhematopoietic cells can stimulate both H-Y Ag-specific T cells and allogenic T cells. (F-G) Marilyn CD4 T cells against male liver endothelial cells (H) and epithelial cells of small intestine (G). (H) B6 male–derived liver endothelial cells can stimulate allogeneic BALB/c T cells. (I-J) Class II expression on liver endothelial cells gated by CD146+ cells, (right) naive and (left) IFN-γ stimulation for 48 hours (I). (J) Bar graph of class II expression on liver endothelial cells. Data are from 1 of 3 similar experiments.

Article Snippet: 10 Briefly, splenic T cells were enriched by autoMACS with anti-CD4, -CD8 microbeads (Miltenyi Biotec).

Techniques: Derivative Assay, Generated, Irradiation, Expressing