Effect of glycerol, n, n-dimethylformamide and n-methyl-2-pyrrolidone on rabbit sperm stored at 4 °C and 16 °C

Artificial insemination with cooled semen is the most common practice in rabbit farms and any improvement on it helps to increase the efficiency and productivity of rabbit meat farms. Therefore, the aim of this study was to assess whether different cryoprotectant agents (CPA) as glycerol, N, N-Dimethylformamide (DMF) and N-Methyl--2-Pyrrolidone (NMP) can improve cooled rabbit sperm quality stored at 4C and 16C. Sperm samples were diluted with INRA 96® (Extender A), INRA 96® with 6% glycerol (Extender B) or 6% DMF (Extender C) or 6% NMP (Extender D) respectively and stored at 4C and 16C. Samples were then analysed at 4, 24, 48 and 72 hours after refrigeration by integrated sperm analysis system (ISAS), eosin-nigrosin stain (vitality), hypo-osmotic swelling test (HOS test) and acrosome integrity test. Extender C showed higher percentage of motility, vitality and HOS test than extender B and D (p<0.05). Whereas sperm quality decreased over time (p<0.05), data showed that the addition of DMF kept the motility and sperm plasma membrane integrity after 24 hours of storage better than other diluents. These results suggest that the addition of DMF to INRA 96® exerts a protective effect on the membrane of spermatozoa improving seminal quality

Long-term preservation of freeze-dried rabbit sperm by adding rosmarinic acid and different chelating agents

Freeze-drying (FD) technique has been applied as an alternative technology to preserve gene resources to allow simple sperm preservation and shipment at 4 degrees C. Nevertheless, DNA sperm might be damaged by mechanical or oxidative stress throughout FD procedure. Therefore, suitable protection to maintain DNA integrity is required. The aim of this study was to determine the effect of rosmarinic acid (RA) as an antioxidant and two chelating agents (EGTA and EDTA) on the DNA integrity of freeze-dried rabbit sperm after storage of the samples at 4 degrees C and room temperature for 8 months. Rabbit sperm were freeze-dried in basic medium (10 mM Tris-HCl buffer and 50 mM NaCl) supplemented with 50 mM EGTA (1), 50 mM EGTA plus 105 mu M RA (2), 50 mM EDTA (3) or 50 mM EDTA plus 105 mu M RA (4). Semen samples were kept at 4 degrees C and room temperature during 8 months. After rehydration, DNA integrity was evaluated with Sperm Chromatin Dispersion test observing that DNA fragmentation was higher when semen samples were freeze-dried with EGTA (10.9%) than with EDTA (4.1%) (p < 0.01). Furthermore, RA acted better under adverse conditions and no significant differences were found in temperature storage. Summarizing, FD is a method that can allow simple gene resources preservation among 4 degrees C to 25 degrees C during 8 months and transportation without the need for liquid nitrogen or dry ice. EDTA chelating agent is the most suitable media for freeze-dried rabbit sperm and the addition of RA protects the DNA against the oxidative stress caused during FD procedure

Repurposing Vandetanib plus Everolimus for the Treatment of ACVR1-Mutant Diffuse Intrinsic Pontine Glioma.

Somatic mutations in ACVR1 are found in a quarter of children with diffuse intrinsic pontine glioma (DIPG), but there are no ACVR1 inhibitors licensed for the disease. Using an artificial intelligence-based platform to search for approved compounds for ACVR1-mutant DIPG, the combination of vandetanib and everolimus was identified as a possible therapeutic approach. Vandetanib, an inhibitor of VEGFR/RET/EGFR, was found to target ACVR1 (K d = 150 nmol/L) and reduce DIPG cell viability in vitro but has limited ability to cross the blood-brain barrier. In addition to mTOR, everolimus inhibited ABCG2 (BCRP) and ABCB1 (P-gp) transporters and was synergistic in DIPG cells when combined with vandetanib in vitro. This combination was well tolerated in vivo and significantly extended survival and reduced tumor burden in an orthotopic ACVR1-mutant patient-derived DIPG xenograft model. Four patients with ACVR1-mutant DIPG were treated with vandetanib plus an mTOR inhibitor, informing the dosing and toxicity profile of this combination for future clinical studies. SIGNIFICANCE: Twenty-five percent of patients with the incurable brainstem tumor DIPG harbor somatic activating mutations in ACVR1, but there are no approved drugs targeting the receptor. Using artificial intelligence, we identify and validate, both experimentally and clinically, the novel combination of vandetanib and everolimus in these children based on both signaling and pharmacokinetic synergies.This article is highlighted in the In This Issue feature, p. 275

Effects of seminal plasma and different cryoprotectants on rabbit sperm preservation at 16°c

The purpose of this research was to assess whether the presence of seminal plasma (SP) can improve sperm quality of rabbit spermatozoa stored at 16°C for 72 h and moreover evaluate the cryoprotectant effects of glycerol, N-N-Dimethylformamide (DMF), and N-methyl-2-pyrrolidone (NMP). Semen samples were pooled and divided in eight fractions. Four of them were diluted with INRA (extender A), INRA with 6% glycerol (extender B), INRA with 6% DMF (extender C), or INRA with 6% NMP (extender D), respectively. The other four fractions were centrifuged, and the supernatant was removed in order to eliminate SP. Each sample was then resuspended with extender A, B, C, or D, respectively. All samples were stored at 16°C and analysed at 4, 24, 48, and 72 h by ISAS®, vitality test, HOS test, and acrosome integrity test. After analyse of the results, SP samples showed a significantly higher percentage (P=0.020) in the HOS test (71.9 ± 1.6%) than non-SP samples (66.5 ± 1.6%). Non-SP samples had better results for kinematic parameters. Extenders A and C showed great results for the percentage of motile spermatozoa (63.1 ± 4.3% and 63.4 ± 3.7%, respectively), vitality (88.9 ± 2.6% and 87.7 ± 2.7%, respectively), and HOS test (68.9 ± 1.4% and 75.2 ± 1.4%, respectively). Extenders B and D showed worse data for sperm quality. These results suggest that SP has a protective effect on rabbit sperm membranes and maintains better sperm motility. The addition of glycerol and NMP to INRA does not improve rabbit sperm quality; nevertheless, the DMF cryoprotectant exerts a protective effect on the membrane of spermatozoa, improving seminal quality during rabbit sperm preservation at 16°C

Clinical implementation and rapid commissioning of an EPID based in-vivo dosimetry system.

Using an Electronic Portal Imaging Device (EPID) to perform in-vivo dosimetry is one of the most effective and efficient methods of verifying the safe delivery of complex radiotherapy treatments. Previous work has detailed the development of an EPID based in-vivo dosimetry system that was subsequently used to replace pre-treatment dose verification of IMRT and VMAT plans. Here we show that this system can be readily implemented on a commercial megavoltage imaging platform without modification to EPID hardware and without impacting standard imaging procedures. The accuracy and practicality of the EPID in-vivo dosimetry system was confirmed through a comparison with traditional TLD in-vivo measurements performed on five prostate patients. The commissioning time required for the EPID in-vivo dosimetry system was initially prohibitive at approximately 10 h per linac. Here we present a method of calculating linac specific EPID dosimetry correction factors that allow a single energy specific commissioning model to be applied to EPID data from multiple linacs. Using this method reduced the required per linac commissioning time to approximately 30 min. The validity of this commissioning method has been tested by analysing in-vivo dosimetry results of 1220 patients acquired on seven linacs over a period of 5 years. The average deviation between EPID based isocentre dose and expected isocentre dose for these patients was (-0.7 +/- 3.2)%. EPID based in-vivo dosimetry is now the primary in-vivo dosimetry tool used at our centre and has replaced nearly all pre-treatment dose verification of IMRT treatment

In aqua vivo EPID dosimetry

Item does not contain fulltextPURPOSE: At the Netherlands Cancer Institute--Antoni van Leeuwenhoek Hospital in vivo dosimetry using an electronic portal imaging device (EPID) has been implemented for almost all high-energy photon treatments of cancer with curative intent. Lung cancer treatments were initially excluded, because the original back-projection dose-reconstruction algorithm uses water-based scatter-correction kernels and therefore does not account for tissue inhomogeneities accurately. The aim of this study was to test a new method, in aqua vivo EPID dosimetry, for fast dose verification of lung cancer irradiations during actual patient treatment. METHODS: The key feature of our method is the dose reconstruction in the patient from EPID images, obtained during the actual treatment, whereby the images have been converted to a situation as if the patient consisted entirely of water; hence, the method is termed in aqua vivo. This is done by multiplying the measured in vivo EPID image with the ratio of two digitally reconstructed transmission images for the unit-density and inhomogeneous tissue situation. For dose verification, a comparison is made with the calculated dose distribution with the inhomogeneity correction switched off. IMRT treatment verification is performed for each beam in 2D using a 2D gamma evaluation, while for the verification of volumetric-modulated arc therapy (VMAT) treatments in 3D a 3D gamma evaluation is applied using the same parameters (3%, 3 mm). The method was tested using two inhomogeneous phantoms simulating a tumor in lung and measuring its sensitivity for patient positioning errors. Subsequently five IMRT and five VMAT clinical lung cancer treatments were investigated, using both the conventional back-projection algorithm and the in aqua vivo method. The verification results of the in aqua vivo method were statistically analyzed for 751 lung cancer patients treated with IMRT and 50 lung cancer patients treated with VMAT. RESULTS: The improvements by applying the in aqua vivo approach are considerable. The percentage of gamma values </=1 increased on average from 66.2% to 93.1% and from 43.6% to 97.5% for the IMRT and VMAT cases, respectively. The corresponding mean gamma value decreased from 0.99 to 0.43 for the IMRT cases and from 1.71 to 0.40 for the VMAT cases, which is similar to the accepted clinical values for the verification of IMRT treatments of prostate, rectum, and head-and-neck cancers. The deviation between the reconstructed and planned dose at the isocenter diminished on average from 5.3% to 0.5% for the VMAT patients and was almost the same, within 1%, for the IMRT cases. The in aqua vivo verification results for IMRT and VMAT treatments of a large group of patients had a mean gamma of approximately 0.5, a percentage of gamma values </=1 larger than 89%, and a difference of the isocenter dose value less than 1%. CONCLUSIONS: With the in aqua vivo approach for the verification of lung cancer treatments (IMRT and VMAT), we can achieve results with the same accuracy as obtained during in vivo EPID dosimetry of sites without large inhomogeneities

Targeted drug distribution in tumor extracellular fluid of GD2-expressing neuroblastoma patient-derived xenografts using SN-38-loaded nanoparticles conjugated to the monoclonal antibody 3F8

Neuroblastoma is a pediatric solid tumor with high expression of the tumor associated antigen disialoganglioside GD2. Despite initial response to induction therapy, nearly 50% of high-risk neuroblastomas recur because of chemoresistance. Here we encapsulated the topoisomerase-I inhibitor SN-38 in polymeric nanoparticles (NPs) surface-decorated with the anti-GD2 mouse mAb 3F8 at a mean density of seven antibody molecules per NP. The accumulation of drug-loaded NPs targeted with 3F8 versus with control antibody was monitored by microdialysis in patient-derived GD2-expressing neuroblastoma xenografts. We showed that the extent of tumor penetration by SN-38 was significantly higher in mice receiving the targeted nano-drug delivery system when compared to non-targeted system or free drug. This selective penetration of the tumor extracellular fluid translated into a strong anti-tumor effect prolonging survival of mice bearing GD2-high neuroblastomas in vivo.Fil: Monterrubio, Carles. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Paco, Sonia. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Olaciregui, Nagore G.. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Pascual Pasto, Guillem. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Vila Ubach, Monica. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Cuadrado Vilanova, Maria. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Ferrandiz, M. Mar. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Castillo Ecija, Helena. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Glisoni, Romina Julieta. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Nanobiotecnología. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Nanobiotecnología; ArgentinaFil: Kuplennik, Nataliya. Technion-Israel Institute of Technology; IsraelFil: Jungbluth, Achim. Memorial Sloan-Kettering Cancer Center; Estados UnidosFil: de Torres, Carmen. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Lavarino, Cinzia. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Cheung, N. K. V.. Memorial Sloan-Kettering Cancer Center; Estados UnidosFil: Mora, Jaume. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; EspañaFil: Sosnik, Alejandro Dario. Technion-Israel Institute of Technology; IsraelFil: Montero Carcaboso, Angel. Institut de Recerca Sant Joan de Deu; España. Hospital Sant Joan de Deu Barcelona; Españ