Dynamic changes in hippocampal diffusion and kurtosis metrics following experimental mTBI correlate with glial reactivity

Diffusion magnetic resonance imaging biomarkers can provide quantifiable information of the brain tissue after a mild traumatic brain injury (mTBI). However, the commonly applied diffusion tensor imaging (DTI) model is not very specific to changes in the underlying cellular structures. To overcome these limitations, other diffusion models have recently emerged to provide a more complete view on the damage profile following TBI. In this study, we investigated longitudinal changes in advanced diffusion metrics following experimental mTBI, utilising three different diffusion models in a rat model of mTBI, including DTI, diffusion kurtosis imaging and a white matter model. Moreover, we investigated the association between the diffusion metrics with histological markers, including glial fibrillary acidic protein (GFAP), neurofilaments and synaptophysin in order to investigate specificity. Our results revealed significant decreases in mean diffusivity in the hippocampus and radial diffusivity and radial extra axonal diffusivity (RadEAD) in the cingulum one week post injury. Furthermore, correlation analysis showed that increased values of fractional anisotropy one day post injury in the hippocampus was highly correlated with GFAP reactivity three months post injury. Additionally, we observed a positive correlation between GFAP on one hand and the kurtosis parameters in the hippocampus on the other hand three months post injury. This result indicated that prolonged glial activation three months post injury is related to higher kurtosis values at later time points. In conclusion, our findings point out to the possible role of kurtosis metrics as well as metrics from the white matter model as prognostic biomarker to monitor prolonged glial reactivity and inflammatory responses after a mTBI not only at early timepoints but also several months after injury. Keywords: Mild traumatic brain injury, Diffusion magnetic resonance imaging, DTI, DKI, White matter mode

Adjuvant therapeutic potential of tonabersat in the standard treatment of glioblastoma : a preclinical F98 glioblastoma rat model study

Purpose Even with an optimal treatment protocol, the median survival of glioblastoma (GB) patients is only 12-15 months. Hence, there is need for novel effective therapies that improve survival outcomes. Recent evidence suggests an important role for connexin (Cx) proteins (especially Cx43) in the microenvironment of malignant glioma. Cx43-mediated gap junctional communication has been observed between tumor cells, between astrocytes and between tumor cells and astrocytes. Therefore, gap junction directed therapy using a pharmacological suppressor or modulator, such as tonabersat, could be a promising target in the treatment of GB. In this preclinical study, we evaluated the possible therapeutic potential of tonabersat in the F98 model. Procedures Female Fischer rats were inoculated with +/- 25.000 F98 tumor cells in the right frontal lobe. Eight days post-inoculation contrast-enhanced T1-weighted (CE-T1w) magnetic resonance (MR) images were acquired to confirm tumor growth in the brain. After tumor confirmation, rats were randomized into a Control Group, a Connexin Modulation Group (CM), a Standard Medical Treatment Group (ST), and a Standard Medical Treatment with adjuvant Connexin Modulation Group (STCM). To evaluate therapy response, T2-weighted (T2w) and CE-T1w sequences were acquired at several time points. Tumor volume analysis was performed on CE-T1w images and statistical analysis was performed using a linear mixed model. Results Significant differences in estimated geometric mean tumor volumes were found between the ST Group and the Control Group and also between the STCM Group and the Control Group. In addition, significant differences in estimated geometric mean tumor volumes between the ST Group and the STCM Group were demonstrated. No significant differences in estimated geometric mean tumor volumes were found between the Control Group and the CM Group. Conclusion Our results demonstrate a therapeutic potential of tonabersat for the treatment of GB when used in combination with radiotherapy and temozolomide chemotherapy

The H2S-Releasing Naproxen Derivative ATB-346 and the Slow-Release H2S Donor GYY4137 Reduce Intestinal Inflammation and Restore Transit in Postoperative Ileus

Objective: Intestinal inflammation triggers postoperative ileus (POI), commonly seen after abdominal surgery and characterized by impaired gastrointestinal transit; when prolonged, this leads to increased morbidity. Hydrogen sulfide (H2S) is recognized as an important mediator of many (patho)physiological processes, including inflammation, and is now investigated for anti-inflammatory application. Therefore, the aim of this study was to investigate the effect of the H2S-releasing naproxen derivative ATB-346, developed to reduce gastrointestinal injury by naproxen, and the slow-release H2S donor GYY4137 on intestinal inflammation and delayed gastrointestinal transit in murine POI.Methods: C57Bl6J mice were fasted for 6 h, anesthetized and after laparotomy, POI was induced by compressing the small intestine with two cotton applicators for 5 min (intestinal manipulation; IM). GYY4137 (50 mg/kg, intraperitoneally), ATB-346 (16 mg/kg, intragastrically) or naproxen (10 mg/kg, intragastrically) were administered 1 h before IM. At 24 h postoperatively, gastrointestinal transit was assessed via fluorescent imaging, and mucosa-free muscularis segments were prepared for later analysis. Inflammatory parameters and activity of inducible nitric oxide synthase (iNOS) and cyclo-oxygenase (COX)-2 were measured. Histological examination of whole tissue sections was done on hematoxylin-eosin stained slides.Results: Pre-treatment with GYY4137 (geometric center; GC: 7.6 ± 0.5) and ATB-346 (GC: 8.4 ± 0.3) prevented the delayed transit induced by IM (GC: 3.6 ± 0.5 vs. 9.0 ± 0.4 in non-operated controls) while naproxen only partially did (GC: 5.9 ± 0.5; n = 8 for all groups). GYY4137 and ATB-346 significantly reduced the IM-induced increase in muscular myeloperoxidase (MPO) activity and protein levels of interleukin (IL)-6, IL-1β and monocyte chemotactic protein 1; the reduction by naproxen was less pronounced and only reached significance for MPO activity and IL-6 levels. All treatments significantly reduced the increase in COX-2 activity caused by IM, whereas only GYY4137 significantly reduced the increase in iNOS activity. Naproxen treatment caused significant histological damage of intestinal villi.Conclusion: The study shows that naproxen partially prevents POI, probably through its inhibitory effect on COX-2 activity. Both ATB-346 and GYY4137 were more effective, the result with GYY4137 showing that H2S per se can prevent POI

Intra-individual dynamic comparison of 18F-PSMA-11 and 68Ga-PSMA-11 in LNCaP xenograft bearing mice

Recently, a 18F-labeled derivative of the widely used 68Ga-PSMA-11 was developed for PET imaging of prostate cancer. Although 18F-PSMA-11 has already been evaluated in a Phase I and Phase II clinical trial, preclinical evaluation of this radiotracer is important for further understanding its dynamic behavior. Saturation binding experiments were conducted by incubation of LNCaP cells with 18F-PSMA-11 or 68Ga-PSMA-11 for 1 h, followed by determination of the specific and aspecific binding. Mice bearing LNCaP or PC-3 xenografts each received ± 3.7 MBq 18F-PSMA-11 and 68Ga-PSMA-11 followed by dynamic acquisition of 2.5 h as well as ± 15 MBq 18F-FDG followed by static acquisition at 1 h post injection (p.i.). Uptake was evaluated by comparison of uptake parameters (SUVmean, SUVmax, TBRmean and TBRmax). Mice underwent ex vivo biodistribution where 18F-PSMA-11 activity was measures in excretory organs (kidneys, bladder and liver) as well as bone fragments (femur, humerus, sternum and skull) to evaluate bone uptake. The dissociation constant (Kd) of 18F-PSMA-11 and 68Ga-PSMA-11 was 2.95 ± 0.87 nM and 0.49 ± 0.20 nM, respectively. Uptake parameters were significantly higher in LNCaP compared to PC-3 xenografts for both 18F-PSMA-11 and 68Ga-PSMA-11, while no difference was found for 18F-FDG uptake (except for SUVmax). Tumor uptake of 18F-PSMA-11 showed a similar trend over time as 68Ga-PSMA-11, although all uptake parameter curves of the latter were considerably lower. When comparing early (60 min p.i.) to delayed (150 min p.i.) imaging for both radiotracers individually, TBRmean and TBRmax were significantly higher at the later timepoint, as well as the SUVmax of 68Ga-PSMA-11. The highest %ID/g was determined in the kidneys (94.0 ± 13.6%ID/g 1 h p.i.) and the bladder (6.48 ± 2.18%ID/g 1 h p.i.). No significant increase in bone uptake was seen between 1 and 2 h p.i. Both radiotracers showed high affinity for the PSMA receptor. Over time, all uptake parameters were higher for 18F-PSMA-11 compared to 68Ga-PSMA-11. Delayed imaging with the latter may improve tumor visualization, while no additional benefits could be found for late 18F-PSMA-11 imaging. Ex vivo biodistribution demonstrated fast renal clearance of 18F-PSMA-11 as well as no significant increase in bone uptake

Hemin reduces postoperative ileus in a heme oxygenase 1‐dependent manner while dimethyl fumarate does without heme oxygenase 1‐induction

Background Postoperative ileus (POI), the impairment of gastrointestinal motility after abdominal surgery, is mainly due to intestinal muscular inflammation. Carbon monoxide (CO)-releasing compounds were shown to exert an anti-inflammatory effect in murine POI partially through induction of heme oxygenase-1 (HO-1). The influence of hemin and dimethyl fumarate (DMF), currently used for multiple sclerosis (MS), was therefore tested in murine POI. Methods C57BL/6J mice were anesthetized and after laparotomy, POI was induced via intestinal manipulation (IM). Animals were treated with either 30 mg kg(-1) hemin intraperitoneally (ip), 30 mg kg(-1) DMF ip, or 100 mg kg(-1) intragastrically (ig) 24 hours before IM. Intestinal transit was assessed 24 hours postoperatively and mucosa-free muscularis or whole segments of the small intestine were stored for later analysis. Intestinal HO-1 protein expression was studied at 6, 12, and 24 hours after administration of hemin or DMF in non-manipulated mice. Key results Pretreatment with hemin and DMF, both ig and ip, prevented the delayed transit seen after IM. Concomitantly, both hemin and DMF significantly reduced the increased interleukin-6 levels and the elevated leukocyte infiltration in the muscularis. Hemin but not DMF caused a significant increase in intestinal HO-1 protein expression and co-administration of the HO-1 inhibitor chromium mesoporphyrin abolished the protective effects of hemin on POI; DMF reduced the IM-induced activation of NF-kappa B and ERK 1/2. Conclusions and Inferences Both hemin and DMF improve the delayed transit and inflammation seen in murine POI, but only hemin does so in a HO-1-dependent manner

Accurate Quantification of Homologous Recombination in Zebrafish: brca2 deficiency as a paradigm

Abstract Homologous Recombination (HR) repair is essential for repairing DNA double strand breaks (DSB) in dividing cells and preventing tumorigenesis. BRCA2 plays an important role in HR by recruiting the DNA recombinase RAD51 to the DSB. Despite being a popular model organism in genetic and cancer research, knowledge on the conservation of the HR pathway and function of zebrafish Brca2 is limited. To evaluate this, we developed a Rad51 foci assay in zebrafish embryos. We identified the zebrafish embryonic intestinal tissue as an ideal target for Rad51 immunostaining. After inducing DSB through irradiation, Rad51 foci were present in irradiated embryos but not in unirradiated controls. We present a method for accurate quantification of HR. Both morpholino-induced knockdown and knockout of Brca2 lead to almost complete absence of Rad51 foci in irradiated embryos. These findings indicate conserved function of Brca2 in zebrafish. Interestingly, a statistically significant decrease in Rad51 foci was observed in Brca2 heterozygous carriers compared to wild types, indicative of haploinsufficiency, a hypothesised cause of some tumours in patients with a germline BRCA2 mutation. In conclusion, we demonstrated the suitability of zebrafish as an excellent in vivo model system for studying the HR pathway and its functionality