- Email: [email protected]
Differential recruitment of CD49d+ Neutrophils by Toll-like Receptor Agonists
AB56 Abstracts
SATURDAY
212
Enhancer of Zeste Homolog 2 induces Pulmonary Artery Smooth Muscle Cell Proliferation and Migration S. A. Aljubran, R. Cox, P. Tamarapu Parthasarathy, G. Kr, S. M. Mohapatra, R. Lockey, N. Kolliputi; University of South Florida, Tampa, FL. INTRODUCTION: Pulmonary Arterial Hypertension (PAH) is a progressively devastating disease characterized by excessive proliferation of the Pulmonary Arterial Smooth Muscle Cells (PASMC’s). Studies suggest that PAH and cancers share an apoptosis-resistant state featuring excessive cell proliferation. The proliferation of cancer cells is mediated by increased expression of Enhancer of Zeste Homolog 2 (EZH2), a mammalian histone methyltransferase that contributes to the epigenetic silencing of target gene. In this study, it hypothesized that EZH2 could play a role in the proliferation of PASMC’s. METHODS: The expression patterns of EZH2 were investigated in normal and hypertensive mouse PASMC’s. The effects of EZH2 overexpression on the proliferation of human PASMC’s were tested. PASMC’s were transfected with EZH2 or GFP using Lonza 4D nucleofector system. The proliferation and cell cycle analysis were performed using flow cytometry; the state of apoptosis of the PASMC’s was determined using annexin V staining, and cell migration tested by wound healing assay. RESULTS: EZH2 protein expression levels were correlated with an increase in right ventricular systolic pressure and Right Ventricular Hypertrophy (RVH). The overexpression of EZH2 in PASMC’s enhances proliferation, migration, and decreases the rate of apoptosis when compared to GFP-transfected cells. In the G2/M phase of the EZH2 transfected cells, a 3.5 fold increase in proliferation and a 1.5 fold increase in the percentage of cells were observed, while there was a significant decrease in rate of apoptosis of the PASMC’s. CONCLUSION: EZH2 could play a role in development of PAH and serve as a potential target for new therapies for PAH.
213
Differential recruitment of CD49d+ Neutrophils by Toll-like Receptor Agonists D. S. Cheung, E. J. Buell, D. A. Hunter, S. J. Zemple, M. H. Grayson; Medical College of Wisconsin, Milwaukee, WI. RATIONALE: Polymorphonuclear neutrophils (PMN) are important innate immune cells. Using the Sendai virus (SeV) model, we have shown that CD49d+ PMNs are critical for the development of post-viral atopic disease. SeV infection, but not LPS (TLR4 agonist) administration was shown to recruit these CD49d+ PMNs to the airways. We undertook the current study to determine if any other toll-like receptors (TLR) were capable of recruiting CD49d+ PMN to the lung. METHODS: C57BL/6 mice were inoculated intranasally with PBS, SeV, or various TLR agonists. One day later, bone marrow, peripheral blood, lung tissue, and bronchoalveolar lavage (BAL) fluid were isolated and examined for CD49d expressing PMN by flow cytometry. RESULTS: Compared to control mice, SeV and a TLR7 agonist reduced the frequency of CD49d+ PMN in the peripheral blood (reduced by 4.661.7%, p50.08; and 6.960.8%, p<0.01, compared to PBS; n53), while TLR3 and 9 agonists increased the percentage of these cells (increased by 18.265.6%, p<0.05; and 28.661.0%, p<0.0001; n53). In the lungs, a TLR9 agonist increased the percentage of CD49d+ PMNs (48.862.1% TLR9 treated versus 23.764.8% for control, p<0.05, n53). As we previously showed, TLR4 stimulation failed to increase CD49d+ PMN, while markedly increasing the CD49d- fraction. SeV was the only agonist to increase the percentage of BAL PMN expressing CD49d (71.861.6% SeV versus 4.164.1% PBS, p<0.001, n53). CONCLUSIONS: Virally associated TLRs appear to increase CD49d+ PMN mobilization from the bone marrow and/or recruitment into the lungs. However, entry into the alveolar space depends upon currently undefined signals associated with viral infection.
J ALLERGY CLIN IMMUNOL FEBRUARY 2012
214
Participant Survey Results From the Starting Hizentra Administration With Resources and Education (SHARE) Program E. Murphy1, P. Riley1, A. Zampelli1, C. Duff2; 1CSL Behring, LLC, King of Prussia, PA, 2University of South Florida, Tampa, FL. RATIONALE: Collaboration between patients and nurse educators is key to successful self-administration with subcutaneous immunoglobulin (SCIG). The Starting Hizentra Administration with Resources and Education (SHARE) program is designed to optimize nurses’ education and training techniques. Nurses were surveyed following participation in the SHARE program to evaluate perceptions about Hizentra and SCIG selfadministration. METHODS: Nurses from diverse care settings attended SHARE programs across the United States. The survey topics included: experience treating or administering SCIG to primary immunodeficiency disease (PIDD) patients; most frequent questions about Hizentra (infusion regimen, physical/chemical properties, safety, efficacy, general SCIG challenges); benefits of SCIG attributes (IgG steady-state levels, patient independence/convenience, improved systemic tolerability over intravenous immunoglobulin); and future educational/outreach interests. RESULTS: Nurses represented practices with SCIG and PIDD experience _ 5 patients using SCIG. The ranging from none to extensive; 66% had < most frequent questions on Hizentra were SCIG-specific challenges (30% of 223 interpretable responses), infusion regimen (21%) and safety (18%); 23% of nurses had no questions. 51% of respondents (155 interpretable responses) considered all SCIG attributes beneficial; 31% found the combination of IgG steady-state levels and patient independence/convenience to be most beneficial. Nurses were most interested in future educational programs on diseases outside of PIDD (25% of 106 interpretable responses), nursing outreach/educational topics (23%), and more information or updates on Hizentra (19%). CONCLUSIONS: Although the benefits of SCIG are generally appreciated, 30% of nurses desire additional information regarding SCIG selfadministration. Nurses are interested in additional educational/outreach, particularly on non-PIDDs and Hizentra.
215
Factors Associated with Daily Stress and Asthma Stress in Caregivers of Children with Asthma C. L. Vibbert1, A. Butz2, M. Donithan2, M. E. Bollinger1; 1University of Maryland, Baltimore, MD, 2Johns Hopkins University, Baltimore, MD. RATIONALE: To determine if caregiver report of daily or asthma daily stress scores are associated with frequency of symptom days in children with persistent asthma. METHODS: As part of an inner-city asthma feedback study, caregivers of the 300 children enrolled were surveyed to include demographic, environmental tobacco exposure, daily and asthma stress levels (0-10 scale) and child asthma characteristics. Level of stress was primary outcome. Chi-square test was used for comparison of categorical variables and ANOVA for continuous variables by stress scores. RESULTS: Children were primarily male (60%), African American (96%), young (mean 5.7 years) and Medicaid insured (92%). Caregivers _ high-school educated (70%) and low-inwere biological mother (92%), > come <$20,000 (72%). Mean baseline caregiver stress was daily stress 5.66 (SD2.5) and daily asthma stress 4.29 (SD2.8). At 2 months, daily stress was significantly associated with having a sibling, visitor or relative as smoker in household (p50.05). A trend was noted for mean daily stress and increased symptom days/14 days scores: 0-2 days: 5.34; 3-5 days: 5.90 and 6+ days: 6.10 (p50.07). At 2 months, mean daily asthma stress was associated with younger child age (3-5 years: 4.58 versus 6-10 years: 3.92; p50.04) and increased symptom days: 0-2 days, 3.75; 3-5 days, 4.74; and 6+ days, 4.87; p50.004. CONCLUSIONS: Having a young child with asthma was associated with higher daily asthma stress scores suggesting newly diagnosed or newly managed children with asthma may be more stressful for caregivers. Asthma management plans need to address underlying caregiver stress to be effective.
SATURDAY
212
Enhancer of Zeste Homolog 2 induces Pulmonary Artery Smooth Muscle Cell Proliferation and Migration S. A. Aljubran, R. Cox, P. Tamarapu Parthasarathy, G. Kr, S. M. Mohapatra, R. Lockey, N. Kolliputi; University of South Florida, Tampa, FL. INTRODUCTION: Pulmonary Arterial Hypertension (PAH) is a progressively devastating disease characterized by excessive proliferation of the Pulmonary Arterial Smooth Muscle Cells (PASMC’s). Studies suggest that PAH and cancers share an apoptosis-resistant state featuring excessive cell proliferation. The proliferation of cancer cells is mediated by increased expression of Enhancer of Zeste Homolog 2 (EZH2), a mammalian histone methyltransferase that contributes to the epigenetic silencing of target gene. In this study, it hypothesized that EZH2 could play a role in the proliferation of PASMC’s. METHODS: The expression patterns of EZH2 were investigated in normal and hypertensive mouse PASMC’s. The effects of EZH2 overexpression on the proliferation of human PASMC’s were tested. PASMC’s were transfected with EZH2 or GFP using Lonza 4D nucleofector system. The proliferation and cell cycle analysis were performed using flow cytometry; the state of apoptosis of the PASMC’s was determined using annexin V staining, and cell migration tested by wound healing assay. RESULTS: EZH2 protein expression levels were correlated with an increase in right ventricular systolic pressure and Right Ventricular Hypertrophy (RVH). The overexpression of EZH2 in PASMC’s enhances proliferation, migration, and decreases the rate of apoptosis when compared to GFP-transfected cells. In the G2/M phase of the EZH2 transfected cells, a 3.5 fold increase in proliferation and a 1.5 fold increase in the percentage of cells were observed, while there was a significant decrease in rate of apoptosis of the PASMC’s. CONCLUSION: EZH2 could play a role in development of PAH and serve as a potential target for new therapies for PAH.
213
Differential recruitment of CD49d+ Neutrophils by Toll-like Receptor Agonists D. S. Cheung, E. J. Buell, D. A. Hunter, S. J. Zemple, M. H. Grayson; Medical College of Wisconsin, Milwaukee, WI. RATIONALE: Polymorphonuclear neutrophils (PMN) are important innate immune cells. Using the Sendai virus (SeV) model, we have shown that CD49d+ PMNs are critical for the development of post-viral atopic disease. SeV infection, but not LPS (TLR4 agonist) administration was shown to recruit these CD49d+ PMNs to the airways. We undertook the current study to determine if any other toll-like receptors (TLR) were capable of recruiting CD49d+ PMN to the lung. METHODS: C57BL/6 mice were inoculated intranasally with PBS, SeV, or various TLR agonists. One day later, bone marrow, peripheral blood, lung tissue, and bronchoalveolar lavage (BAL) fluid were isolated and examined for CD49d expressing PMN by flow cytometry. RESULTS: Compared to control mice, SeV and a TLR7 agonist reduced the frequency of CD49d+ PMN in the peripheral blood (reduced by 4.661.7%, p50.08; and 6.960.8%, p<0.01, compared to PBS; n53), while TLR3 and 9 agonists increased the percentage of these cells (increased by 18.265.6%, p<0.05; and 28.661.0%, p<0.0001; n53). In the lungs, a TLR9 agonist increased the percentage of CD49d+ PMNs (48.862.1% TLR9 treated versus 23.764.8% for control, p<0.05, n53). As we previously showed, TLR4 stimulation failed to increase CD49d+ PMN, while markedly increasing the CD49d- fraction. SeV was the only agonist to increase the percentage of BAL PMN expressing CD49d (71.861.6% SeV versus 4.164.1% PBS, p<0.001, n53). CONCLUSIONS: Virally associated TLRs appear to increase CD49d+ PMN mobilization from the bone marrow and/or recruitment into the lungs. However, entry into the alveolar space depends upon currently undefined signals associated with viral infection.
J ALLERGY CLIN IMMUNOL FEBRUARY 2012
214
Participant Survey Results From the Starting Hizentra Administration With Resources and Education (SHARE) Program E. Murphy1, P. Riley1, A. Zampelli1, C. Duff2; 1CSL Behring, LLC, King of Prussia, PA, 2University of South Florida, Tampa, FL. RATIONALE: Collaboration between patients and nurse educators is key to successful self-administration with subcutaneous immunoglobulin (SCIG). The Starting Hizentra Administration with Resources and Education (SHARE) program is designed to optimize nurses’ education and training techniques. Nurses were surveyed following participation in the SHARE program to evaluate perceptions about Hizentra and SCIG selfadministration. METHODS: Nurses from diverse care settings attended SHARE programs across the United States. The survey topics included: experience treating or administering SCIG to primary immunodeficiency disease (PIDD) patients; most frequent questions about Hizentra (infusion regimen, physical/chemical properties, safety, efficacy, general SCIG challenges); benefits of SCIG attributes (IgG steady-state levels, patient independence/convenience, improved systemic tolerability over intravenous immunoglobulin); and future educational/outreach interests. RESULTS: Nurses represented practices with SCIG and PIDD experience _ 5 patients using SCIG. The ranging from none to extensive; 66% had < most frequent questions on Hizentra were SCIG-specific challenges (30% of 223 interpretable responses), infusion regimen (21%) and safety (18%); 23% of nurses had no questions. 51% of respondents (155 interpretable responses) considered all SCIG attributes beneficial; 31% found the combination of IgG steady-state levels and patient independence/convenience to be most beneficial. Nurses were most interested in future educational programs on diseases outside of PIDD (25% of 106 interpretable responses), nursing outreach/educational topics (23%), and more information or updates on Hizentra (19%). CONCLUSIONS: Although the benefits of SCIG are generally appreciated, 30% of nurses desire additional information regarding SCIG selfadministration. Nurses are interested in additional educational/outreach, particularly on non-PIDDs and Hizentra.
215
Factors Associated with Daily Stress and Asthma Stress in Caregivers of Children with Asthma C. L. Vibbert1, A. Butz2, M. Donithan2, M. E. Bollinger1; 1University of Maryland, Baltimore, MD, 2Johns Hopkins University, Baltimore, MD. RATIONALE: To determine if caregiver report of daily or asthma daily stress scores are associated with frequency of symptom days in children with persistent asthma. METHODS: As part of an inner-city asthma feedback study, caregivers of the 300 children enrolled were surveyed to include demographic, environmental tobacco exposure, daily and asthma stress levels (0-10 scale) and child asthma characteristics. Level of stress was primary outcome. Chi-square test was used for comparison of categorical variables and ANOVA for continuous variables by stress scores. RESULTS: Children were primarily male (60%), African American (96%), young (mean 5.7 years) and Medicaid insured (92%). Caregivers _ high-school educated (70%) and low-inwere biological mother (92%), > come <$20,000 (72%). Mean baseline caregiver stress was daily stress 5.66 (SD2.5) and daily asthma stress 4.29 (SD2.8). At 2 months, daily stress was significantly associated with having a sibling, visitor or relative as smoker in household (p50.05). A trend was noted for mean daily stress and increased symptom days/14 days scores: 0-2 days: 5.34; 3-5 days: 5.90 and 6+ days: 6.10 (p50.07). At 2 months, mean daily asthma stress was associated with younger child age (3-5 years: 4.58 versus 6-10 years: 3.92; p50.04) and increased symptom days: 0-2 days, 3.75; 3-5 days, 4.74; and 6+ days, 4.87; p50.004. CONCLUSIONS: Having a young child with asthma was associated with higher daily asthma stress scores suggesting newly diagnosed or newly managed children with asthma may be more stressful for caregivers. Asthma management plans need to address underlying caregiver stress to be effective.