Intravoxel Incoherent Motion Metrics as Potential Biomarkers for Survival in Glioblastoma.

Intravoxel incoherent motion (IVIM) is an MRI technique with potential applications in measuring brain tumor perfusion, but its clinical impact remains to be determined. We assessed the usefulness of IVIM-metrics in predicting survival in newly diagnosed glioblastoma. Fifteen patients with glioblastoma underwent MRI including spin-echo echo-planar DWI using 13 b-values ranging from 0 to 1000 s/mm2. Parametric maps for diffusion coefficient (D), pseudodiffusion coefficient (D*), and perfusion fraction (f) were generated for contrast-enhancing regions (CER) and non-enhancing regions (NCER). Regions of interest were manually drawn in regions of maximum f and on the corresponding dynamic susceptibility contrast images. Prognostic factors were evaluated by Kaplan-Meier survival and Cox proportional hazards analyses. We found that fCER and D*CER correlated with rCBFCER. The best cutoffs for 6-month survival were fCER>9.86% and D*CER>21.712 x10-3mm2/s (100% sensitivity, 71.4% specificity, 100% and 80% positive predictive values, and 80% and 100% negative predictive values; AUC:0.893 and 0.857, respectively). Treatment yielded the highest hazard ratio (5.484; 95% CI: 1.162-25.88; AUC: 0.723; P = 0.031); fCER combined with treatment predicted survival with 100% accuracy. The IVIM-metrics fCER and D*CER are promising biomarkers of 6-month survival in newly diagnosed glioblastoma

Orthogonal Regulatory Circuits for Escherichia coli Based on the gamma-Butyrolactone System of Streptomyces coelicolor

Chemically inducible transcription factors are widely used to control gene expression of synthetic devices. The bacterial quorum sensing system is a popular tool to achieve such control. However, different quorum sensing systems have been found to cross-talk, both between themselves and with the hosts of these devices, and they are leaky by nature. Here we evaluate the potential use of the γ-butyolactone system from Streptomyces coelicolor A3(2) M145 as a complementary regulatory circuit. First, two additional genes responsible for the biosynthesis of γ-butyrolactones were identified in S. coelicolor M145 and then expressed in E. coli BL21 under various experimental conditions. Second, the γ-butyrolactone receptor ScbR was optimised for expression in E. coli BL21. Finally, signal and promoter crosstalk between the γ-butyrolactone system from S. coelicolor and quorum sensing systems from Vibrio fischeri and Pseudomonas aeruginosa was evaluated. The results show that the γ-butyrolactone system does not cros..

The Impact of Small Molecule Binding on the Energy Landscape of the Intrinsically Disordered Protein C-Myc

Intrinsically disordered proteins are attractive therapeutic targets owing to their prevalence in several diseases. Yet their lack of well-defined structure renders ligand discovery a challenging task. An intriguing example is provided by the oncoprotein c-Myc, a transcription factor that is over expressed in a broad range of cancers. Transcriptional activity of c-Myc is dependent on heterodimerization with partner protein Max. This protein-protein interaction is disrupted by the small molecule 10058-F4 (1), that binds to monomeric and disordered c-Myc. To rationalize the mechanism of inhibition, structural ensembles for the segment of the c-Myc domain that binds to 1 were computed in the absence and presence of the ligand using classical force fields and explicit solvent metadynamics molecular simulations. The accuracy of the computed structural ensembles was assessed by comparison of predicted and measured NMR chemical shifts. The small molecule 1 was found to perturb the composition of the apo equilibrium ensemble and to bind weakly to multiple distinct c-Myc conformations. Comparison of the apo and holo equilibrium ensembles reveals that the c-Myc conformations binding 1 are already partially formed in the apo ensemble, suggesting that 1 binds to c-Myc through an extended conformational selection mechanism. The present results have important implications for rational ligand design efforts targeting intrinsically disordered proteins

Correlation between Topographic Vessel Density and Retinal Thickness Changes in Patients with Diabetic Macular Edema Treated with Anti-VEGF Therapy: Is It a Suitable OCTA Biomarker?

The objective of this study was to determine the correlation between topographic vessel density (VD) and retinal thickness (RT) reductions induced by vascular endothelial growth factor inhibitors (anti-VEGF) in patients with diabetic macular edema (DME) using optical coherence tomography angiography (OCTA). This was a prospective, interventional case series. VD and RT measurements were separately taken in four parafoveal subfields at baseline and after six months of treatment. This correlation was statistically assessed using Spearman's rho correlation coefficient after adjustment for multiple comparisons. The study included a total of 48 eyes in the final analysis. Mean VD decreased from baseline to month 6 (from 45.2 (+/- 3.5) to 44.6% (+/- 3.2) in the superficial capillary plexus and from 50 (+/- 3.3) to 49% (+/- 3.9) in the deep capillary plexus). Statistically significant reductions in RT were observed in all ETDRS sectors (p < 0.0001). No significant association was found between RT and VD, even when analyzing responders and non-responders separately. After six months of anti-VEGF treatment, no significant correlation was observed between the topographic VD and RT values. These findings suggest that reductions in VD values may not solely result from a reduction in microaneurysms, also being affected by the repositioning of displaced vessels due to edema and a reduction in their caliber. Therefore, VD changes may not be a suitable indirect OCTA biomarker of microaneurysm turnover and treatment response

Naive T lymphocytes chemotax long distance to CCL21 but not to a source of bioactive S1P

Naive T lymphocytes traffic through the organism in search for antigen, alternating between blood and secondary lymphoid organs. Lymphocyte homing to lymph nodes relies on CCL21 chemokine sensing by CCR7 receptors, while exit into efferent lymphatics relies on sphingolipid S1P sensing by S1PR1 receptors. While both molecules are claimed chemotactic, a quantitative analysis of naive T lymphocyte migration along defined gradients is missing. Here, we used a reductionist approach to study the real-time single-cell response of naive T lymphocytes to CCL21 and serum rich in bioactive S1P. Using microfluidic and micropatterning ad hoc tools, we show that CCL21 triggers stable polarization and long-range chemotaxis of cells, whereas S1P-rich serum triggers a transient polarization only and no significant displacement, potentially representing a brief transmigration step through exit portals. Our in vitro data thus suggest that naive T lymphocyte chemotax long distances to CCL21 but not toward a source of bioactive S1P

Combined Small Interfering RNA Therapy and In Vivo Magnetic Resonance Imaging in Islet Transplantation

OBJECTIVE Recent advances in human islet transplantation are hampered by significant graft loss shortly after transplantation and inability to follow islet fate directly. Both issues were addressed by utilizing a dual-purpose therapy/imaging small interfering RNA (siRNA)-nanoparticle probe targeting apoptotic-related gene caspase-3. We expect that treatment with the probe would result in significantly better survival of transplanted islets, which could be monitored by in vivo magnetic resonance imaging (MRI). RESEARCH DESIGN AND METHODS We synthesized a probe consisting of therapeutic (siRNA to human caspase-3) and imaging (magnetic iron oxide nanoparticles, MN) moieties. In vitro testing of the probe included serum starvation of the islets followed by treatment with the probe. Caspase-3 gene silencing and protein expression were determined by RT-PCR and Western blot, respectively. In vivo studies included serial MRI of NOD-SCID mice transplanted with MN-small interfering (si)Caspase-3–labeled human islets under the left kidney capsule and MN-treated islets under the right kidney capsule. RESULTS Treatment with MN-siCaspase-3 probe resulted in decrease of mRNA and protein expression in serum-starved islets compared with controls. In vivo MRI showed that there were significant differences in the relative volume change between MN-siCaspase-3–treated grafts and MN-labeled grafts. Histology revealed decreased caspase-3 expression and cell apoptosis in MN-siCaspase-3–treated grafts compared with the control side. CONCLUSIONS Our data show the feasibility of combining siRNA therapy and in vivo monitoring of transplanted islets in mice. We observed a protective effect of MN-siCaspase-3 in treated islets both in vitro and in vivo. This study could potentially aid in increasing the success of clinical islet transplantation

Keratocytes migrate against flow with a roly-poly-like mechanism.

peer reviewedWhile cell migration can be directed by various mechanical cues such as force, deformation, stiffness, or flow, the associated mechanisms and functions may remain elusive. Single cell migration against flow, repeatedly reported with leukocytes, is arguably considered as active and mediated by integrin mechanotransduction, or passive and determined by a mechanical bias. Here, we reveal a phenotype of flow mechanotaxis with fish epithelial keratocytes that orient upstream or downstream at shear stresses around tens of dyn cm-2. We show that each cell has an intrinsic orientation that results from the mechanical interaction of flow with its morphology. The bulbous trailing edge of a keratocyte generates a hydrodynamical torque under flow that stabilizes an upstream orientation, just as the heavy lower edge of a roly-poly toy generates a gravitational torque that stabilizes an upright position. In turn, the wide and flat leading edge of keratocytes destabilizes upstream orientation, allowing the existence of two distinct phenotypes. To formalize these observations, we propose a simple mechanical model that considers keratocyte morphology as a hemisphere preceded by a wide thin sheet. Our findings show that this model can recapitulate the phase diagram of single cell orientation under flow without adjustable parameters. From a larger perspective, this passive mechanism of keratocytes flow mechanotaxis implies a potential absence of physiological function and evolution-driven process

Expression of the NH2-Terminal Fragment of RasGAP in Pancreatic β-Cells Increases Their Resistance to Stresses and Protects Mice From Diabetes

OBJECTIVE: Our laboratory has previously established in vitro that a caspase-generated RasGAP NH(2)-terminal moiety, called fragment N, potently protects cells, including insulinomas, from apoptotic stress. We aimed to determine whether fragment N can increase the resistance of pancreatic beta-cells in a physiological setting. RESEARCH DESIGN AND METHODS: A mouse line, called rat insulin promoter (RIP)-N, was generated that bears a transgene containing the rat insulin promoter followed by the cDNA-encoding fragment N. The histology, functionality, and resistance to stress of RIP-N islets were then assessed. RESULTS: Pancreatic beta-cells of RIP-N mice express fragment N, activate Akt, and block nuclear factor kappaB activity without affecting islet cell proliferation or the morphology and cellular composition of islets. Intraperitoneal glucose tolerance tests revealed that RIP-N mice control their glycemia similarly as wild-type mice throughout their lifespan. Moreover, islets isolated from RIP-N mice showed normal glucose-induced insulin secretory capacities. They, however, displayed increased resistance to apoptosis induced by a series of stresses including inflammatory cytokines, fatty acids, and hyperglycemia. RIP-N mice were also protected from multiple low-dose streptozotocin-induced diabetes, and this was associated with reduced in vivo beta-cell apoptosis. CONCLUSIONS: Fragment N efficiently increases the overall resistance of beta-cells to noxious stimuli without interfering with the physiological functions of the cells. Fragment N and the pathway it regulates represent, therefore, a potential target for the development of antidiabetes tools

Nutritional status and quality of life of patients with advanced gastroenteropancreatic neuroendocrine neoplasms in Spain: the NUTRIGETNE (GETNE-S2109) study

Patients with advanced gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs) have impaired nutritional and physical performance due to the cancer pathophysiology and its treatment. The NUTRIGETNE study sought to characterize the nutritional status of patients with advanced GEP-NENs in Spain. This is a cross-sectional study that included patients with advanced GEP-NENs receiving active oncological treatment. Patients had a complete physical examination, anthropometry, bioelectrical impedance, dynamometry, laboratory analysis, and a comprehensive nutritional risk assessment. Malnutrition was defined according to Global Leadership Initiative on Malnutrition (GLIM) criteria. The study included 399 patients out of the 400 planned (Pearson's chi 2; alpha 0.05). Median age was 62 years (22-83). Tumors most commonly originated in the small intestine (43.9%) and the pancreas (41.6%), 94.7% were metastatic, and 36.7%, 49.4%, and 12.5% were G1, G2, and G3, respectively. Malnutrition prevalence was 61.9% (25.8% moderate; 36.1% severe), mainly due to low muscle mass (50.9%), which was the most prevalent GLIM phenotypic criteria. Moreover, malnutrition showed a correlation with decreased hand grip strength (mean 23 vs 31.9 kg; P <.001) and phase angle (median 5o vs 5.6o; P <.001). The prevalence of sarcopenia was 15%. Malnutrition was more frequent in patients with diabetes (74.4% vs 56.7%; P <.001), NECs (82.1% vs 60.3%; P =.062), and in those treated with chemotherapy (71.2% vs 59.7%; P =.058), whereas it did not correlate with tumor origin (P =.507), histological grade (P =.781), or functionality (P =.465). Malnutrition was correlated to body mass index (BMI) (P =.015), although it was also diagnosed in a high proportion of patients with no weight loss (63%, 54.1%, and 65.1% of patients with normal BMI, overweight, and obesity, respectively). Cachexia was present in 109 (27.3%) patients. Malnutrition is very prevalent and commonly underdiagnosed in patients with GEP-NENs. It is associated with sarcopenia and a worse QoL, requiring a multifactorial nutritional assessment. Certain factors such as the presence of diabetes may require closer monitoring due to a higher risk of malnutrition