Genetically Encoded Biosensors Unveil Neuronal Injury Dynamics via Multichromatic ATP and Calcium Imaging.

When a cell sustains damage, it liberates cytosolic ATP, which can serve as an injury signal, affecting neighboring cells. This study presents a methodological approach that employs in vitro axotomy and in vivo laser ablation to simulate cellular injury. Specially tailored biosensors are employed to monitor ATP dynamics and calcium transients in injured cells and their surroundings. To simultaneously visualize extracellular and cytosolic ATP, we developed bicistronic constructs featuring GRABATP1.0 and MaLionR biosensors alongside the calcium sensor RCaMP, enabling multiparametric imaging. In addition to transducing primary neuron cultures, we developed another method where we cocultured dorsal root ganglion neurons together with specialized "sniffer" cell lines expressing the bicistronic biosensors. Exploiting these approaches, we successfully demonstrated the release of ATP from the injured neurons and its extracellular diffusion in response to cellular injury in vitro and in vivo. Axotomy triggered intracellular calcium mobilization not only in the injured neuron but also in the intact neighboring cells, providing new insights into ATP's role as an injury signal. The tools developed in this study have demonstrated remarkable efficiency in unraveling the intricacies of ATP-mediated injury signaling.

Development of a Chemogenetic Approach to Manipulate Intracellular pH.

Chemogenetic Operation of iNTRacellular prOton Levels (pH-Control) is a novel substrate-based enzymatic method that enables precise spatiotemporal control of ultralocal acidification in cultured cell lines and primary neurons. The genetically encoded biosensor SypHer3s showed that pH-Control effectively acidifies cytosolic, mitochondrial, and nuclear pH exclusively in the presence of ß-chloro-d-alanine in living cells in a concentration-dependent manner. The pH-Control approach is promising for investigating the ultralocal pH imbalance associated with many diseases.

Results of a three-year follow-up after endovascular therapy with multilayer flow modulator in complex thoracoabdominal aortic aneurysm: A case report.

OBJECTIVES: Studies on the short-, medium and long-term effects of flow guiding stents are still limited. In this case report, we present three-year follow-up of the multilayer flow modulator stent in a 55-year-old patient with Crawford Type 2 thoracoabdominal aortic aneurysm. METHODS: A 55-year-old male patient with Crawford Type 2 thoracoabdominal aortic aneurysm had applied to our medical center. The aneurysm involved coeliac truncus and superior mesenteric artery and extended to the renal artery ostia. Multilayer flow modulator stent was successfully placed, and follow-up CT (Computed tomography) angiographic examination images recorded intermittently (36 months). RESULTS: After three years, it was observed that the left renal artery was thrombosed and the left kidney went to atrophy. Other major vascular branches were observed to be open. During this time, the aneurysm was completely closed with thrombus, but the diameter of the aneurysm continued to increase. CONCLUSIONS: Multilayer flow modulator stents are safe in complex aortic aneurysms. The device increases the thrombus load in the aortic aneurysm and maintains the flow of the main vascular branches. But re-interventions, dilatation of the aneurysm sac and visceral branch obstructions are still challenging for multilayer flow modulator stents.

Effects of different biomaterials on augmented bone volume resorptions.

OBJECTIVES: The aim of this study was to assess the effects of platelet-rich fibrin (PRF), deproteinized bovine bone graft (DBBG), and biphasic calcium phosphate (BCP) biomaterials on total volume resorption levels and bone augmentation. MATERIALS AND METHODS: Nine New Zealand rabbits were divided into three groups (PRF, DBBG, and BCP). Two titanium barriers were fixed bilaterally to each rabbit's calvarium; a total of 18 titanium barriers were used. All barriers were removed on the 90th day. Computed tomography (CT) images of the animals were taken on the 90th, 120th, 150th, and 180th days. RESULTS: When the resorption level of the total volume in all groups on the 120th, 150th, and 180th days was compared with that of the 90th day, no significant differences among all groups were found in all intervals. According to the total volume on the 90th and 180th days, statistically significant differences between groups DBBG and BCP were not found; however, statistically significant differences were found between group PRF and the others groups (P < 0.001). On the other hand, there were statistically significant differences in regenerated bone area between group PRF and BCP. CONCLUSION: According to the results of this study, grafts such as DBBG or BCP must be used to augment bone volume sufficiently in guided bone regeneration.

The effect of local simvastatin application on critical size defects in the diabetic rats.

OBJECTIVES: To evaluate bone-healing effects of local simvastatin application to critical size defects (CSDs) in the experimental diabetes mellitus (DM) rat model. MATERIALS AND METHODS: A total of 35 male Sprague-Dawley rats with an average weight of 350 g and aged 3 months were used in this study. The rats were divided into five groups of seven animals each: passive control (group A), active control (group B), 0.5 mg simvastatin (group C), 1.0 mg simvastatin (group D), and 1.5 mg simvastatin (group E). Streptozotocin was used to induce Type 1 diabetes in all rats. Eight mm CSDs were created under anesthesia in each rat calvarium. CSDs were left empty in group A. Defects in group B were grafted alone with a gelatin sponge mixed with normal saline. Defects in the experimental groups (groups A, B, and C) were grafted with gelatin sponge mixed saline solutions contain 0.5, 1.0, 1.5 mg simvastatin. Rats were sacrificed after 1 month, and the defects were prepared for radiologic and histomorphometric assessment of regenerated bone. RESULTS: None of the specimens exhibited complete closure of new bone across the 8-mm defect. A correlation between computed tomography and histomorphometric analysis was not determined. Both amount of volume and area of regenerated bone were found higher in the experimental groups than in the control groups. However, these values were not found statistically significant degree (P < 0.05) for each groups. The density of regenerated bone in the region of interest was higher in the control groups in contrast to in the experimental groups. However, statistical significance was just found between groups C and A and between groups C and B (P < 0.05). CONCLUSION: The local simvastatin application enhanced healing of the bone defects in the diabetic rat model CSDs.