17-Allylamino-Demethoxygeldanamycin Ameliorate Microthrombosis Via HSP90/RIP3/NLRP3 Pathway After Subarachnoid Hemorrhage in Rats.

BACKGROUND: Subarachnoid hemorrhage (SAH) is a severe and emergent cerebrovascular disease, the prognosis of which usually very poor. Microthrombi formation highlighted with inflammation occurs early after SAH. As the main cause of DCI, microthrombosis associated with the prognosis of SAH. The aim of this study was to show HSP90 inhibitor 17-AAG effect on microthrombosis after SAH in rats. METHODS: Ninety-five SD rats were used for the experiment. For time course study, the rats were randomly divided into five groups: sham group and SAH group with different time point (1d, 2d, 3d, 5d). Endovascular perforation method was conducted for SAH model. Neurological score, SAH grade, and mortality were measured after SAH. The samples of the left hemisphere brain were collected. The expression of HSP90 was detected by Western blot. The microthrombosis after SAH in rats' brain was detected by immunohistochemistry. For mechanism study, rats were randomly divided into three groups: sham, SAH + vehicle, and SAH +17-AAG (n = 6/group). 17-AAG was given by intraperitoneal injection (80 mg/kg) 1 h after SAH. Neurological function were measured at 24 h after SAH. The expression of RIP3, NLRP3, ASC, and IL-1ß was measured by Western blot. Microthrombosis was detected by immunohistochemistry. RESULTS: Our results showed that the HSP90 protein level increased and peaked at 2 days after SAH. Microthrombosis caused by SAH was increased in 1 day and peaked at 2 days after SAH. Administration HSP90 specific inhibitor 17-AAG reduced expression of RIP3, NLRP3, ASC, and IL-1ß, reduced microthrombosis after SAH, and improved neurobehavior when compared to vehicle group. CONCLUSIONS: 17-AAG can ameliorate microthrombosis via HSP90/RIP3/NLRP3 pathway and improve neurobehavior after SAH.

Subarachnoid hemorrhage enhances the expression of TDP-43 in the brain of experimental rats and human subjects.

The transactive response DNA-binding protein of 43 (TDP-43) may be involved in neurodegenerative disease and in the response to brain injury; however, alterations in the expression of TDP-43 following subarachnoid hemorrhage (SAH) require further investigation. The present study reported a notable elevation in the expression of TDP-43 within the cerebrospinal fluid (CSF) of patients with aneurysmal SAH and increased brain expression of TDP-43 in a rat model of SAH. The TDP-43 protein and a derivative migrated at 43 and 24 kDa, respectively, as observed via the immunoblotting of concentrated CSF samples obtained from patients with SAH; no signal was detected in the CSF from healthy controls. SAH in rats was induced by intravascular suture puncture. The expression levels of TDP-43 in rat cortical lysates following SAH were increased at 0.5 h, peaked at 48 h and remained significantly elevated at 72 h post-injury, compared with sham controls. TDP-43 immunolabeling indication localization within neurons, astrocytes and microglia in the experimental rats. Collectively, the findings of the present study indicated the early involvement of TDP-43 in the brain in response to SAH, and that expression levels of TDP-43 in the CSF may serve as a prognostic biomarker among patients with this condition.

Milk Fat Globule-Epidermal Growth Factor-8 Pretreatment Attenuates Apoptosis and Inflammation via the Integrin-ß3 Pathway after Surgical Brain Injury in Rats.

Iatrogenic brain injury inevitably occurs in neurosurgical operations, leading to brain edema, ischemia, intracranial hematoma, and other postoperative complications, eventually worsening neurological outcomes of patients. If apoptotic cells are not rapidly eliminated by phagocytic engulfment, they may communicate with surrounding cells to undergo secondary necrosis and releasing toxic signals. Recent studies have shown that milk fat globule-epidermal growth factor-8 (MFGE8), which promotes phagocytosis and inhibits inflammation, is an endogenous protective factor in response to brain infarction, Alzheimer's disease, subarachnoid hemorrhage, and prion disease. In the present study, we sought to investigate the different effects of both pretreated and posttreated recombinant milk fat globule-epidermal growth factor-8 (rhMFGE8) for the surgical brain injury (SBI) rat model and potential involvement of its receptor integrin ß3 for apoptosis and neuroinflammation after SBI. One hundred and sixty-seven male rats were employed in the preset study. Experiment 1 was performed to evaluate neurological scores and MFGE8, cleaved caspase-3 (CC3), and interleukine-1 beta (IL-1ß) levels at 3, 24, and 120 h after SBI. Experiment 2 was performed to evaluate the effects of rhMFGE8 pretreatment (10 min before SBI) and rhMFGE8 posttreatment (6 h after SBI) on brain edema at 24 and 72 h after SBI. Experiment 3 was performed to evaluate the potential anti-apoptotic and anti-inflammatory effects of rhMFGE8 pretreatment and posttreatment. Experiment 4 sought to investigate the involvement of the integrin-ß3 signal in the effects of MFGE8 pretreatment. Our data showed rhMFGE8 pretreatment alleviated neurological deficits and decreased brain water content and apoptotic cells in the SBI model, which exhibited neurological dysfunction, apoptosis, and inflammation. Meanwhile, MFGE8 siRNA, which inhibited endogenous MFGE8 expression, significantly increased IL-1ß, TUNEL positive cells, and CC3. Furthermore, knockdown of its receptor integrin ß3 by siRNA abolished the effects of rhMFGE8 in the SBI model. In conclusion, we found that rhMFGE8 pretreatment effectively alleviated neurological deficits and decreased brain water content and apoptotic cells in the SBI model through the MFGE8/integrin-ß3 pathway, and treatment time was an important factor in achieving curative effects. Therefore, MFGE8 pretreatment may serve as a promising therapeutic strategy for SBI patients.