Journal: PLoS ONE

Article Title: Autotaxin and LPA Receptors Represent Potential Molecular Targets for the Radiosensitization of Murine Glioma through Effects on Tumor Vasculature

doi: 10.1371/journal.pone.0022182

Figure Lengend Snippet: (A) HUVEC or (B) bEnd.3 cells were treated with vehicle control (▪) or 5 µM BrP-LPA (▴) for 45 min prior to irradiation. (C) HUVEC were treated with 0.1% fatty acid free BSA (•), 10 µM LPA (○) or 10 µM LPA plus 5 µM BrP-LPA (▪) in serum free medium for 45 min prior to irradiation. The cells were then irradiated with 0, 2, 4 and 6 Gy and plated for clonogenic survival assay. After 2–3 wks, cells were stained with 1% methylene blue and colonies consisting of >50 cells were counted by microscopy. Surviving colonies were normalized for plating efficiency. Shown are average survival fractions and SEM from three experiments; * p<0.05. (D) Equal numbers of HUVEC were plated in 96 well plates and treated with carrier control 3% fatty acid free BSA, 10 µM LPA or 10 µM LPA with 5 µM BrP-LPA in serum free medium for 45 min prior to irradiation. After 96 h, the cell viability was determined using a colorimetric cell proliferation assay (Promega). Shown is the average absorbance at 490 nm with SEM from three experiments; * p<0.05.

Article Snippet: To inhibit ATX and LPA receptors, we used α-bromomethylene phosphonate LPA (BrP-LPA) which was purchased from Echelon (Salt Lake City, UT).

Techniques: Irradiation, Clonogenic Cell Survival Assay, Staining, Microscopy, Proliferation Assay

Journal: PLoS ONE

Article Title: Autotaxin and LPA Receptors Represent Potential Molecular Targets for the Radiosensitization of Murine Glioma through Effects on Tumor Vasculature

doi: 10.1371/journal.pone.0022182

Figure Lengend Snippet: (A, B) HUVEC or (C) bEnd.3 cells were plated on matrigel-coated 96 well plates and treated with vehicle control or 5 µM BrP-LPA for 45 min prior to irradiation with 3 Gy. (A) Shown are the representative photomicrographs of tubule formation taken 16 h after irradiation. (B) Tubule formation was quantified as number of tubules per high power field (HPF). Shown are bar graphs of mean number of tubules per HPF relative to control and SEM from three experiments; * p<0.05.

Article Snippet: To inhibit ATX and LPA receptors, we used α-bromomethylene phosphonate LPA (BrP-LPA) which was purchased from Echelon (Salt Lake City, UT).

Techniques: Irradiation

Journal: PLoS ONE

Article Title: Autotaxin and LPA Receptors Represent Potential Molecular Targets for the Radiosensitization of Murine Glioma through Effects on Tumor Vasculature

doi: 10.1371/journal.pone.0022182

Figure Lengend Snippet: (A) bEnd.3 or (B) HUVEC were plated on 60 mm plates and allowed to grow to 70% confluency. The semi-confluent cell layer was scraped using a sterile pipette tip to create a scratch devoid of cells. The remaining cells were treated with vehicle control or 5 µM BrP-LPA for 45 min prior to irradiation with 3 Gy. Migration was observed at 36 h. Cells were fixed with ethanol and stained with 1% methylene blue. Migrated cells were counted and normalized to surrounding cell density per HPF. Shown are representative photomicrographs and bar graphs representing the mean percentages of migrating cells relative to corresponding controls with SEM from three experiments, * p<0.05.

Article Snippet: To inhibit ATX and LPA receptors, we used α-bromomethylene phosphonate LPA (BrP-LPA) which was purchased from Echelon (Salt Lake City, UT).

Techniques: Transferring, Irradiation, Migration, Staining

Journal: PLoS ONE

Article Title: Autotaxin and LPA Receptors Represent Potential Molecular Targets for the Radiosensitization of Murine Glioma through Effects on Tumor Vasculature

doi: 10.1371/journal.pone.0022182

Figure Lengend Snippet: (A, B) Mouse glioma GL261 cells were plated on 60 mm plates and allowed to grow to 70% confluency. Plates were scraped with a pipette tip to create a scratch devoid of cells and treated with vehicle control or 5 µM BrP-LPA for 45 min before irradiation with 3 Gy. After 24 h, cells were fixed and stained with methylene blue. Migrated cells were counted and normalized to surrounding cell density per HPF. Shown are representative photomicrographs (A) and a bar graph (B) representing the mean percentages of migrating cells relative to corresponding controls with SEM from three experiments; * p<0.05. (C) For clonogenic survival assay, GL261 cells were plated and allowed to attach. After 6 h, cells were treated vehicle control or with 5 µM BrP-LPA for 45 min and irradiated with 0, 2, 4, and 6 Gy. After 10 days, surviving colonies (>50 cells) were counted and normalized for plating efficiency. Shown is the clonogenic survival curve and mean surviving fractions and SEM from three experiments; * p<0.05.

Article Snippet: To inhibit ATX and LPA receptors, we used α-bromomethylene phosphonate LPA (BrP-LPA) which was purchased from Echelon (Salt Lake City, UT).

Techniques: Transferring, Irradiation, Staining, Clonogenic Cell Survival Assay

Journal: PLoS ONE

Article Title: Autotaxin and LPA Receptors Represent Potential Molecular Targets for the Radiosensitization of Murine Glioma through Effects on Tumor Vasculature

doi: 10.1371/journal.pone.0022182

Figure Lengend Snippet: bEnd.3 and GL261 cells were grown in co-culture for 24 h. Cells were treated with vehicle or 5 µM BrP-LPA control for 45 min before treatment with 3 Gy. Cells were lysed at 5 min after irradiation. Shown are immunoblot analyses using specific antibodies to phospho-Akt Thr308/Ser473 , total Akt, and actin. Numbers represent the ratios of phospho-Akt to total Akt protein normalized to actin (fold change relative to control).

Article Snippet: To inhibit ATX and LPA receptors, we used α-bromomethylene phosphonate LPA (BrP-LPA) which was purchased from Echelon (Salt Lake City, UT).

Techniques: Co-Culture Assay, Irradiation, Western Blot

Journal: PLoS ONE

Article Title: Autotaxin and LPA Receptors Represent Potential Molecular Targets for the Radiosensitization of Murine Glioma through Effects on Tumor Vasculature

doi: 10.1371/journal.pone.0022182

Figure Lengend Snippet: GL261 cells were injected into the hind limbs of nude mice. Tumors were irradiated with 3 Gy for 5 consecutive days for a total of 15 Gy. Mice were treated with 3 mg/kg BrP-LPA or vehicle control for 45 min prior to irradiation on days 1, 3, and 5. (A) Shown are mean tumor volumes with SEM from each treatment group of 5 mice. (B) Tumor growth delay was calculated as the number of days for tumors to reach a 6-fold volume increase compared to control. Shown is a bar graph representing the mean tumor growth delay with SEM from each treatment group of 5 mice; * p<0.05.

Article Snippet: To inhibit ATX and LPA receptors, we used α-bromomethylene phosphonate LPA (BrP-LPA) which was purchased from Echelon (Salt Lake City, UT).

Techniques: Injection, Irradiation

Journal: Purinergic Signalling

Article Title: Lysophosphatidic acid generation by pulmonary NKT cell ENPP-2/autotaxin exacerbates hyperoxic lung injury

doi: 10.1007/s11302-015-9463-6

Figure Lengend Snippet: Hyperoxia increases lysophosphatidic acid (LPA) levels in pulmonary NKT cells in vitro. BrP-LPA diminishes level of LPA. Pulmonary iNKT cells were cultured and exposed to 95 % O2/5 % CO2 for 72 h with and without addition of the pan-LPA antagonist BrP-LPA. LPA levels in hyperoxia-exposed cells were significantly lower when cells were incubated with the pan LPA antagonist BrP-LPA during hyperoxia (p < 0.05). r.u. relative units (relative to the background-reaction in the absence of weight)

Article Snippet: Bromomethylene phosphonate LPA (BrP-LPA) was purchased from Echelon®.

Techniques: In Vitro, Cell Culture, Incubation

Journal: Purinergic Signalling

Article Title: Lysophosphatidic acid generation by pulmonary NKT cell ENPP-2/autotaxin exacerbates hyperoxic lung injury

doi: 10.1007/s11302-015-9463-6

Figure Lengend Snippet: Kaplan–Meier survival curves for (n > 25) and plus BrP-LPA animals after 72 h in 100 % oxygen demonstrate a clear survival benefit of those animals that received a BrP-LPA injection prior to oxygen exposure

Article Snippet: Bromomethylene phosphonate LPA (BrP-LPA) was purchased from Echelon®.

Techniques: Injection

Journal: Purinergic Signalling

Article Title: Lysophosphatidic acid generation by pulmonary NKT cell ENPP-2/autotaxin exacerbates hyperoxic lung injury

doi: 10.1007/s11302-015-9463-6

Figure Lengend Snippet: Histological assessment of hyperoxic lung injury. a H&E staining of control lung tissue and b plus BrP-LPA injection prior to hyperoxia. Animals were exposed to 100 % oxygen for 72 h. The hyperoxia-exposed lungs show severe injury with pulmonary edema and hemorrhage, whereas the BrP-LPA injected animals express milder injury

Article Snippet: Bromomethylene phosphonate LPA (BrP-LPA) was purchased from Echelon®.

Techniques: Staining, Injection

Journal: Purinergic Signalling

Article Title: Lysophosphatidic acid generation by pulmonary NKT cell ENPP-2/autotaxin exacerbates hyperoxic lung injury

doi: 10.1007/s11302-015-9463-6

Figure Lengend Snippet: Flow cytometry analysis of pulmonary NKT cell and PMN populations after hyperoxia. a Lungs extracted from oxygen-exposed mice with and without BrP-LPA treatment were analyzed. Baseline iNKT cell populations in the lung did not differ between BrP-LPA treated and untreated wild type mice under normoxic conditions. After 72 h of 100 % exposure, animals show significant increases of iNKT cells compared to their baseline. After BrP-LPA treatment, animals exhibit only a minimal increase of pulmonary iNKT cells in response to hyperoxia. b Populations of GR-1+/F4/80- PMN-cells increase massively after oxygen exposure in lungs. When animals were injected with BrP-LPA prior to oxygen exposure PMN-cells only increased marginally

Article Snippet: Bromomethylene phosphonate LPA (BrP-LPA) was purchased from Echelon®.

Techniques: Flow Cytometry, Injection