Fingolimod improves diffuse brain injury by promoting AQP4 polarization and functional recovery of the glymphatic system.

BACKGROUND: Diffuse brain injury (DBI) models are characterized by intense global brain inflammation and edema, which characterize the most severe form of TBI. In a previous experiment, we found that fingolimod promoted recovery after controlled cortical impact injury (CCI) by modulating inflammation around brain lesions. However, it remains unclear whether fingolimod can also attenuate DBI because of its different injury mechanisms. Furthermore, whether fingolimod has additional underlying effects on repairing DBI is unknown. METHODS: The impact acceleration model of DBI was established in adult Sprague-Dawley rats. Fingolimod (0.5 mg/kg) was administered 0.5, 24, and 48 h after injury for 3 consecutive days. Immunohistochemistry, immunofluorescence analysis, cytokine array, and western blotting were used to evaluate inflammatory cells, inflammatory factors, AQP4 polarization, apoptosis in brain cells, and the accumulation of APP after DBI in rats. To evaluate the function of the glymphatic system (GS), a fluorescent tracer was injected into the cistern. The neural function of rats with DBI was evaluated using various tests, including the modified neurological severity score (mNSS), horizontal ladder-crossing test, beam walking test, and tape sensing and removal test. Brain water content was also measured. RESULTS: Fingolimod administration for 3 consecutive days could reduce the levels of inflammatory cytokines, neutrophil recruitment, microglia, and astrocyte activation in the brain following DBI. Moreover, fingolimod reduced apoptotic protein expression, brain cell apoptosis, brain edema, and APP accumulation. Additionally, fingolimod inhibited the loss of AQP4 polarization, improved lymphatic system function, and reduced damage to nervous system function. Notably, inhibiting the GS weakened the therapeutic effect of fingolimod on the neurological function of rats with DBI and increased the accumulation of APP in the brain. CONCLUSIONS: In brief, these findings suggest that fingolimod alleviates whole-brain inflammation and GS system damage after DBI and that inhibiting the GS could weaken the positive effect of fingolimod on nerve function in rats with DBI. Thus, inhibiting inflammation and regulating the GS may be critical for the therapeutic effect of fingolimod on DBI.

Inactivation of ERK1/2 signaling mediates dysfunction of basal meningeal lymphatic vessels in experimental subdural hematoma.

Rationale: Meningeal lymphatic vessels (MLVs) are essential for the clearance of subdural hematoma (SDH). However, SDH impairs their drainage function, and the pathogenesis remains unclear. Herein, we aimed to understand the pathological mechanisms of MLV dysfunction following SDH and to test whether atorvastatin, an effective drug for SDH clearance, improves meningeal lymphatic drainage (MLD). Methods: We induced SDH models in rats by injecting autologous blood into the subdural space and evaluated MLD using Gadopentetate D, Evans blue, and CFSE-labeled erythrocytes. Whole-mount immunofluorescence and transmission electron microscopy were utilized to detect the morphology of MLVs. Phosphoproteomics, western blot, flow cytometry, and in vitro experiments were performed to investigate the molecular mechanisms underlying dysfunctional MLVs. Results: The basal MLVs were detected to have abundant valves and play an important role in draining subdural substances. Following SDH, these basal MLVs exhibited disrupted endothelial junctions and dilated lumen, leading to impaired MLD. Subsequent proteomics analysis of the meninges detected numerous dephosphorylated proteins, primarily enriched in the adherens junction, including significant dephosphorylation of ERK1/2 within the meningeal lymphatic endothelial cells (LECs). Subdural injection of the ERK1/2 kinase inhibitor PD98059 resulted in dilated basal MLVs and impaired MLD, resembling the dysfunctional MLVs observed in SDH. Moreover, inhibiting ERK1/2 signaling severely disrupted intercellular junctions between cultured LECs. Finally, atorvastatin was revealed to protect the structure of basal MLVs and accelerate MLD following SDH. However, these beneficial effects of atorvastatin were abolished when combined with PD98059. Conclusion: Our findings demonstrate that SDH induces ERK1/2 dephosphorylation in meningeal LECs, leading to disrupted basal MLVs and impaired MLD. Additionally, we reveal a beneficial effect of atorvastatin in improving MLD.

The inflammatory cellular feature in the peripheral blood of chronic subdural hematoma patients.

BACKGROUND: Chronic subdural hematoma (CSDH) is a common neurosurgical disease with an increasing incidence. The absorption route of CSDH is not clear. Whether inflammatory factors enter the peripheral blood and cause systemic reactions is unknown. METHODS: We screened 105 CSDH patients and 105 control individuals. Their clinical characteristics and blood routine results were collected and compared. The blood routine changes of CSDH patients before and after treatment were compared. Age-stratified analysis was performed due to age may affect the inflammatory markers. RESULTS: The white blood cell count, absolute neutrophil count, neutrophil percentage, neutrophil-lymphocyte count ratio (NLR), and platelet to lymphocyte count ratio (PLR) of CSDH patients before treatment were within the normal range, while were significantly higher than the control individuals (p < 0.001). The absolute lymphocyte count and lymphocyte percentage of control individuals were higher than those of patients (p < 0.001). The inflammatory cells in patients of different age groups were similar. After the patient was cured, the white blood cell count, the absolute value and percentage of neutrophils decreased (p < 0.05), while the number of monocytes increased. CONCLUSIONS: CSDH caused slight systemic inflammatory responses in the peripheral blood, implying that there is a non-hematologic route for the absorption of hematoma.

Possible mechanism and Atorvastatin-based treatment in cupping therapy-related subdural hematoma: A case report and literature review.

Subdural hematoma (SDH) is one of the most lethal types of traumatic brain injury. SDH caused by Intracranial Pressure Reduction (ICPR) is rare, and the mechanism remains unclear. Here, we report three cases of SDH that occurred after substandard cupping therapy and are conjected to be associated with ICPR. All of them had undergone cupping treatments. On the last cupping procedure, they experienced a severe headache after the cup placed on the occipital-neck junction (ONJ) was suddenly removed and were diagnosed with SDH the next day. In standard cupping therapy, the cups are not usually placed on the ONJ. We speculate that removing these cups on the soft tissue over the cisterna magna repeatedly created localized negative pressure, caused temporary but repeated ICPR, and eventually led to SDH development. The Monro-Kellie Doctrine can explain the mechanism behind this - it states that the intracranial pressure is regulated by a fixed system, with any change in one component causing a compensatory change in the other. The repeated ICPR promoted brain displacement, tearing of the bridging veins, and development of SDH. The literature was reviewed to illustrate the common etiologies and therapies of secondary ICPR-associated SDH. Despite the popularity of cupping therapy, its side effects are rarely mentioned. This case is reported to remind professional technicians to fully assess a patient's condition before cupping therapy and ensure that the cups are not placed at the ONJ.

Effects of increased intracranial pressure on cerebrospinal fluid influx, cerebral vascular hemodynamic indexes, and cerebrospinal fluid lymphatic efflux.

The glymphatic-lymphatic fluid transport system (GLFTS) consists of glymphatic pathway and cerebrospinal fluid (CSF) lymphatic outflow routes, allowing biological liquids from the brain parenchyma to access the CSF along with perivascular space and to be cleaned out of the skull through lymphatic vessels. It is known that increased local pressure due to physical compression of tissue improves lymphatic transport in peripheral organs, but little is known about the exact relationship between increased intracranial pressure (IICP) and GLFTS. In this study, we verify our hypothesis that IICP significantly impacts GLFTS, and this effect depends on severity of the IICP. Using a previously developed inflating balloon model to induce IICP and inject fluorescent tracers into the cisterna magna, we found significant impairment of the glymphatic circulation after IICP. We further found that cerebrovascular occlusion occurred, and cerebrovascular pulsation decreased after IICP. IICP also interrupted the drainage of deep cervical lymph nodes and dorsal meningeal lymphatic function, enhancing spinal lymphatic outflow to the sacral lymph nodes. Notably, these effects were associated with the severity of IICP. Thus, our findings proved that the intensity of IICP significantly impacts GLFTS. This may have translational applications for preventing and treating related neurological disorders.

Dexamethasone enhances the efficacy of atorvastatin in inhibiting excessively inflammation-induced abnormal angiogenesis by regulating macrophages.

BACKGROUND: We have recently showed that atorvastatin (ATO) combined with low dose of dexamethasone (DEX) was more efficacious in treating patients with chronic subdural haematoma (CSDH) than ATO monotherapy. This study was designed to investigate the underlying mechanisms of the improved efficacy of this combined therapy. METHODS: Mass spectrometry was performed to quantitatively detect drugs in haematoma fluids and serum samples from CSDH patients and also in cultured macrophages after treatment with either ATO alone or in combination with DEX. The differentiation and apoptosis of macrophages were evaluated using flow cytometry. The expression of cytokines, chemokines and angiogenesis-related proteins was evaluated using proteome profile arrays, immunoblots and ELISA, respectively. RESULTS: ATO was detected in haematoma fluids and serum samples, whose levels were increased significantly in samples collected from patients treated with both ATO and DEX. ATO was also increased in cultured macrophages treated with ATO and DEX. The numbers of M1-polarized macrophages were higher than the M2 phenotype in the haematoma fluids of patients. Cultured macrophages treated with ATO and DEX had reduced numbers of M1-polarized macrophages, increased numbers of M2-polarized macrophages as compared to monotherapies, and decreased rate of apoptosis induced by high-dose DEX. DEX enhanced the anti-inflammatory and anti-angiogenic activity of ATO by suppressing VEGFA and other inflammatory angiogenic factors. Consistent with the finding, patients responded well to the drug treatments had lower serum levels of VEGFA. CONCLUSIONS: We have shown for the first time that ATO given orally was detected in CSDH haematoma fluids. DEX enhances the anti-inflammatory and anti-angiogenic effects of ATO, primarily by increasing the presence of ATO in haematoma and macrophages and by regulating the functions of macrophages.

Risk Factors for Atorvastatin as a Monotherapy for Chronic Subdural Hematoma: A Retrospective Multifactor Analysis.

Chronic subdural hematoma (CSDH) is a common form of intracranial hemorrhage in the aging population. We aimed to investigate the predictive factors for atorvastatin efficacy as a monotherapy for moderate CSDH. We retrospectively reviewed the medical records of patients who were diagnosed with moderate CSDH and received atorvastatin monotherapy between February 5, 2014, and November 7, 2015, in multiple neurosurgical departments. Univariate, multivariate and receiver operating characteristic curve analyses were performed to identify the potential significant factors indicative of the good therapeutic efficacy or poor therapeutic efficacy of atorvastatin for mild CSDH, such as age, sex, history of injury, Markwalder grading scale-Glasgow Coma Scale (MGS-GCS), Activities of Daily Life-the Barthel Index scale (ADL-BI), American Society of Anesthesiologists Physical Status classification system (ASA-PS), blood cell counts, serum levels and computed tomography findings. A total of 89 patients (75 men and 14 women) aged 24-88 years (mean age 61.95 ± 15.30 years) were followed-up for 24 weeks. Computed tomography findings at admission showed mixed-density hematoma in 22 patients, isodense hematoma in 13 patients, high-density hematoma in 26 patients, and low-density hematoma in 28 patients. In total, 3, 80, and 6 patients had MGS-GCS grades of 0, 1, and 2, respectively. The efficacy rate at 6 months was 87.6% (78/89). Eleven patients were switched to surgery due to a worsened neurological condition, of whom 8, 1, 1, and 1 had high-density, low-density, isodense and mixed-density hematomas, respectively. These patients were switched to surgery over a range of 2-27 days, with a median interval of 12 days after the medication treatment. Univariate and multivariate analyses, confirmed by ROC curves, revealed that high-density hematoma, basal cistern compression, and hematoma volume to be independent risk factors for the efficacy of atorvastatin monotherapy in patients with moderate CSDH. Atorvastatin is an effective monotherapy for the treatment of mild CSDH. High-density hematoma, basal cistern compression, and hematoma volume are independent predictors of the efficacy of atorvastatin as a non-surgical treatment. The results suggested that ADL-BI was more sensitive than the MGS-GCS and ASA-PS for determining patient outcomes in our moderate CSDH cohort.

Atorvastatin Plus Low-Dose Dexamethasone May Be Effective for Leukemia-Related Chronic Subdural Hematoma but Not for Leukemia Encephalopathy: A Report of Three Cases.

We are not aware of any reports regarding conservative treatment for leukemia-related chronic subdural hematoma (CSDH). We report our experience with 3 men who were admitted with subdural masses and abnormal leukocyte counts. In two patients, leukemia and CSDH were confirmed on the basis of medical records, mild head trauma, and neuroimaging features. Both patients experienced reduced CSDH and neurological symptoms after receiving atorvastatin (20 mg/day) plus low-dose dexamethasone. However, this combined conservative treatment was ineffective in the third patient, who was diagnosed as having leukemia and showed an increased hematoma volume after two weeks of therapy. Magnetic resonance imaging findings suggested dural metastasis, which prompted a switch from statin-based conservative treatment to chemotherapy. Complete remission of the leukemia and resolution of the subdural mass were observed after chemotherapy, which supported a diagnosis of leukemia encephalopathy. The 5-month follow-ups did not reveal CSDH relapse in all 3 cases. Thus, atorvastatin-based conservative treatment may be effective for leukemia-related CSDH but not for leukemia encephalopathy.

Subdural haematomas drain into the extracranial lymphatic system through the meningeal lymphatic vessels.

Subdural haematomas (SDHs) are characterized by rapidly or gradually accumulated haematomas between the arachnoid and dura mater. The mechanism of haematoma clearance has not been clearly elucidated until now. The meningeal lymphatic vessel (mLV) drainage pathway is a novel system that takes part in the clearance of waste products in the central nervous system (CNS). This study aimed to explore the roles of the mLV drainage pathway in SDH clearance and its impacting factors. We injected FITC-500D, A488-fibrinogen and autologous blood into the subdural space of mice/rats and found that these substances drained into deep cervical lymph nodes (dCLNs). FITC-500D was also observed in the lymphatic vessels (LYVE+) of the meninges and the dCLNs in mice. The SDH clearance rate in SDH rats that received deep cervical lymph vessel (dCLV) ligation surgery was significantly lower than that in the control group, as evaluated by haemoglobin quantification and MRI scanning. The drainage rate of mLVs was significantly slower after the SDH model was established, and the expression of lymphangiogenesis-related proteins, including LYVE1, FOXC2 and VEGF-C, in meninges was downregulated. In summary, our findings proved that SDH was absorbed through the mLV drainage pathway and that haematomas could inhibit the function of mLVs.

Hematoma-derived exosomes of chronic subdural hematoma promote abnormal angiogenesis and inhibit hematoma absorption through miR-144-5p.

Exosomes are small (30-150 nm diameter) lipid bilayer-enclosed vesicles found in all bodily fluids. We investigated whether exosomes play a role in chronic subdural hematoma (CSDH). Exosomes were identified and characterized using transmission electron microscopy and NanoSight particle tracking. The functions of hematoma-derived exosomes were evaluated in a rat model of acute subdural hematoma (SDH). The hematoma-derived exosomes inhibited hematoma absorption and exacerbated neurological deficits in SDH rats. We examined the effects of the exosomes on angiogenesis and cell permeability in human umbilical vein endothelial cells (HUVECs). Co-culture of exosomes with HUVECs revealed that the hematoma-derived exosomes were taken-in by the HUVECs, resulting in enhanced tube formation and vascular permeability. Additionally, there was a concomitant increase in ANG-2 expression and decrease in ANG-1 expression. Exosomes were enriched with microRNAs including miR-144-5p, which they could deliver to HUVECs to promote angiogenesis and increase membrane permeability. Overexpression of miR-144-5p in HUVECs and in SDH rats promoted abnormal angiogenesis and reduced hematoma absorption, which mimicked the effects of the hematoma-derived exosomes both in vitro and in vivo. Thus, hematoma-derived exosomes promote abnormal angiogenesis with high permeability and inhibit hematoma absorption through miR-144-5p in CSDH.