Department of Pathology and Laboratory Medicine

IQGAP1-Dependent Signaling Pathway Regulates Endothelial Cell Proliferation and Angiogenesis

2012-01-12T07:01:42Z

IQGAP1-Dependent Signaling Pathway Regulates Endothelial Cell Proliferation and Angiogenesis Meyer, Rosana D.; Sacks, David B.; Rahimi, Nader BACKGROUND. Vascular endothelial growth factor receptor-2 (VEGFR-2) signaling is an obligate requirement for normal development and pathological angiogenesis such as cancer and age-related macular degeneration. Although autophosphorylation of tyrosine 1173 (Y1173) of VEGFR-2 is considered a focal point for its angiogenic signal relay, however, the mechanism of phosphorylation of Y1173, signaling proteins that are recruited to this residue and their role in angiogenesis is not fully understood. METHODOLOGY/PRINCIPAL FINDINGS. In this study we demonstrate that c-Src kinase directly through its Src homology 2 (SH2) domain and indirectly via c-Cbl binds to phospho-Y1057 of VEGFR-2. Activation of c-Src kinase by a positive feedback mechanism phosphorylates VEGFR-2 at multi-docking site, Y1173. c-Src also catalyzes tyrosine phosphorylation of IQGAP1 and acts as an adaptor to bridge IQGAP1 to VEGFR-2. In turn, IQGAP1 activates b-Raf and mediates proliferation of endothelial cells. Silencing expression of IQGAP1 and b-Raf revealed that their activity is essential for VEGF to stimulate angiogenesis in an in vivo angiogenesis model of chicken chorioallantoic membrane (CAM). CONCLUSIONS/SIGNIFICANCE. Angiogenesis contributes to the pathology of numerous human diseases ranging from cancer to age-related macular degeneration. Determining molecular mechanism of tyrosine phosphorylation of VEGFR-2 and identification of molecules that are relaying its angiogenic signaling may identify novel targets for therapeutic intervention against angiogenesis-associated diseases. Our study shows that recruitment and activation of c-Src by VEGFR-2 plays a pivotal role in relaying angiogenic signaling of VEGFR-2; it phosphorylates VEGFR-2 at Y1173, facilitates association and activation of IQGAP1 and other signaling proteins to VEGFR-2. IQGAP1-dependent signaling, in part, is critically required for endothelial cell proliferation, a key step in angiogenesis. Thus, Y1057 of VEGFR-2 serves to regulate VEGFR-2 function in a combinatorial manner by supporting both diversity of recruitment of angiogenic signaling proteins to VEGFR-2, and its ability to promote angiogenesis.

Oral Tolerance Inhibits Pulmonary Eosinophilia in a Cockroach Allergen Induced Model of Asthma: A Randomized Laboratory Study

2012-01-12T07:01:42Z

Oral Tolerance Inhibits Pulmonary Eosinophilia in a Cockroach Allergen Induced Model of Asthma: A Randomized Laboratory Study Vaickus, Louis J; Bouchard, Jacqueline; Kim, Jiyoun; Natarajan, Sudha; Remick, Daniel G BACKGROUND. Antigen desensitization through oral tolerance is becoming an increasingly attractive treatment option for allergic diseases. However, the mechanism(s) by which tolerization is achieved remain poorly defined. In this study we endeavored to induce oral tolerance to cockroach allergen (CRA: a complex mixture of insect components) in order to ameliorate asthma-like, allergic pulmonary inflammation. METHODS. We compared the pulmonary inflammation of mice which had received four CRA feedings prior to intratracheal allergen sensitization and challenge to mice fed PBS on the same time course. Respiratory parameters were assessed by whole body unrestrained plethysmography and mechanical ventilation with forced oscillation. Bronchoalveolar lavage fluid (BAL) and lung homogenate (LH) were assessed for cytokines and chemokines by ELISA. BAL inflammatory cells were also collected and examined by light microscopy. RESULTS. CRA feeding prior to allergen sensitization and challenge led to a significant improvement in respiratory health. Airways hyperreactivity measured indirectly via enhanced pause (Penh) was meaningfully reduced in the CRA-fed mice compared to the PBS fed mice (2.3 ± 0.4 vs 3.9 ± 0.6; p = 0.03). Directly measured airways resistance confirmed this trend when comparing the CRA-fed to the PBS-fed animals (2.97 ± 0.98 vs 4.95 ± 1.41). This effect was not due to reduced traditional inflammatory cell chemotactic factors, Th2 or other cytokines and chemokines. The mechanism of improved respiratory health in the tolerized mice was due to significantly reduced eosinophil numbers in the bronchoalveolar lavage fluid (43300 ± 11445 vs 158786 ± 38908; p = 0.007) and eosinophil specific peroxidase activity in the lung homogenate (0.59 ± 0.13 vs 1.19 ± 0.19; p = 0.017). The decreased eosinophilia was likely the result of increased IL-10 in the lung homogenate of the tolerized mice (6320 ± 354 ng/mL vs 5190 ± 404 ng/mL, p = 0.02). CONCLUSION. Our results show that oral tolerization to CRA can improve the respiratory health of experimental mice in a CRA-induced model of asthma-like pulmonary inflammation by reducing pulmonary eosinophilia.

Allergens Induce Enhanced Bronchoconstriction and Leukotriene Production in C5 Deficient Mice

2012-01-12T07:01:43Z

Allergens Induce Enhanced Bronchoconstriction and Leukotriene Production in C5 Deficient Mice McKinley, Laura; Kim, Jiyoun; Bolgos, Gerald L; Siddiqui, Javed; Remick, Daniel G BACKGROUND. Previous genetic analysis has shown that a deletion in the complement component 5 gene-coding region renders mice more susceptible to allergen-induced airway hyperresponsiveness (AHR) due to reduced IL-12 production. We investigated the role of complement in a murine model of asthma-like pulmonary inflammation. METHODS. In order to evaluate the role of complement B10 mice either sufficient or deficient in C5 were studied. Both groups of mice immunized and challenged with a house dust extract (HDE) containing high levels of cockroach allergens. Airways hyper-reactivity was determined with whole-body plesthysmography. Bronchoalveolar lavage (BAL) was performed to determine pulmonary cellular recruitment and measure inflammatory mediators. Lung homogenates were assayed for mediators and plasma levels of IgE determined. Pulmonary histology was also evaluated. RESULTS. C5-deficient mice showed enhanced AHR to methylcholine challenge, 474% and 91% increase above baseline Penh in C5-deficient and C5-sufficient mice respectively, p < 0.001. IL-12 levels in the lung homogenate (LH) were only slightly reduced and BAL IL-12 was comparable in C5-sufficient and C5-deficient mice. However, C5-deficient mice had significantly higher cysteinyl-leukotriene levels in the BAL fluid, 1913 +/- 246 pg/ml in C5d and 756 +/- 232 pg/ml in C5-sufficient, p = 0.003. CONCLUSION. These data demonstrate that C5-deficient mice show enhanced AHR due to increased production of cysteinyl-leukotrienes.

Weight, Blood Pressure, and Dietary Benefits After 12 Months of a Web-based Nutrition Education Program (DASH for Health): Longitudinal Observational Study

2012-01-12T07:01:42Z

Weight, Blood Pressure, and Dietary Benefits After 12 Months of a Web-based Nutrition Education Program (DASH for Health): Longitudinal Observational Study Moore, Thomas J; Alsabeeh, Nour; Apovian, Caroline M; Murphy, Megan C; Coffman, Gerald A; Cullum-Dugan, Diana; Jenkins, Mark; Cabral, Howard BACKGROUND The dietary habits of Americans are creating serious health concerns, including obesity, hypertension, diabetes, cardiovascular disease, and even some types of cancer. While considerable attention has been focused on calorie reduction and weight loss, approaches are needed that will not only help the population reduce calorie intake but also consume the type of healthy, well-balanced diet that would prevent this array of medical complications. OBJECTIVE To design an Internet-based nutrition education program and to explore its effect on weight, blood pressure, and eating habits after 12 months of participation. METHODS. We designed the DASH for Health program to provide weekly articles about healthy nutrition via the Internet. Dietary advice was based on the DASH diet (Dietary Approaches to Stop Hypertension). The program was offered as a free benefit to the employees of EMC Corporation, and 2834 employees and spouses enrolled. Enrollees voluntarily entered information about themselves on the website (food intake), and we used these self-entered data to determine if the program had any effect. Analyses were based upon the change in weight, blood pressure, and food intake between the baseline period (before the DASH program began) and the 12th month. To be included in an outcome, a subject had to have provided both a baseline and 12th-month entry. RESULTS After 12 months, 735 of 2834 original enrollees (26%) were still actively using the program. For subjects who were overweight/obese (body mass index >25; n = 151), weight change at 12 months was -4.2 lbs (95% CI: -2.2, -6.2; P< .001). For subjects with hypertension or prehypertension at baseline (n = 62), systolic blood pressure fell 6.8 mmHg at 12 months (CI: -2.6, -11.0; P<.001; n = 62). Diastolic pressure fell 2.1 mmHg (P = .16). Based upon self-entered food surveys, enrollees (n = 181) at 12 months were eating significantly more fruits, more vegetables, and fewer grain products. They also reduced consumption of carbonated beverages. Enrollees who had visited the website more often tended to have greater blood pressure and weight loss effect, suggesting that use of the DASH for Health program was at least partially responsible for the benefits we observed. CONCLUSIONS We have found that continued use of a nutrition education program delivered totally via the Internet, with no person-to-person contact with health professionals, is associated with significant weight loss, blood pressure lowering, and dietary improvements after 12 months. Effective programs like DASH for Health, delivered via the Internet, can provide benefit to large numbers of subjects at low cost and may help address the nutritional public health crisis.