Higher CSF Levels of NLRP3 Inflammasome Is Associated With Poor Prognosis of Anti-N-methyl-D-Aspartate Receptor Encephalitis.

Anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis is accepted as an autoimmune disorder of the central nervous system (CNS). NLR family pyrin domain containing 3 (NLRP3) inflammasome, a potent innate inflammatory mediator, can activate IL-1ß and induce the migration of T helper cell into CNS. However, the possible role of NLRP3 inflammasome in the pathogenesis of anti-NMDAR encephalitis remains unclear. This study aims to determine the levels of NLRP3 and related cytokines (IL-1ß, IL-6, and IL-17) in the cerebrospinal fluid (CSF) of anti-NMDAR encephalitis patients and to assess whether NLRP3 influences the clinical outcomes of this disease. Twenty-five patients with anti-NMDAR encephalitis, 12 viral meningoencephalitis patients and 26 controls with non-inflammatory neurological diseases were recruited. CSF NLRP3 inflammasome, IL-1ß, IL-6, and IL-17 were measured by enzyme-linked immunosorbent assay. Thirteen out of 25 patients were re-examed for the concentrations of NLRP3 and cytokines 6 months later. Our results showed significant increases of CSF NLRP3 inflammasome, IL-1ß, IL-6, and IL-17 in anti-NMDAR encephalitis patients. There were positive correlations between CSF NLRP3 inflammasome and cytokines in anti-NMDAR encephalitis patients. There was also a positive correlation between maximum modified Rankin Scale (mRS) scores and CSF NLRP3 inflammasome in anti-NMDAR encephalitis patients. During follow-up, the decrease of mRS was positively correlated with the decrease of CSF NLRP3 inflammasomes. These results suggested that the level of CSF NLRP3 inflammasome could represent the severity of anti-NMDAR encephalitis and the reduction of CSF NLRP3 inflammasome could act as an indicator for the prognosis of this disease.

Inflammasome proteins as biomarkers of traumatic brain injury.

BACKGROUND: The inflammasome plays an important role in the inflammatory innate immune response after central nervous system (CNS) injury. Inhibition of the inflammasome after traumatic brain injury (TBI) results in improved outcomes by lowering the levels of caspase-1 and interleukin (IL)-1b. We have previously shown that inflammasome proteins are elevated in the cerebrospinal fluid (CSF) of patients with TBI and that higher levels of these proteins were consistent with poorer outcomes after TBI when compared to patients that presented these inflammasome proteins at lower levels. METHODS AND FINDINGS: Here we extend our work by analyzing serum from 21 TBI patients and CSF from 18 TBI patients compared to 120 serum samples and 30 CSF samples from no-TBI donor controls for the expression of caspase-1, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), interleukin(IL)-1b and IL-18. Analysis was carried out using the Ella Simple Plex system (Protein Simple) to determine the sensitivity and specificity of inflammasome proteins as biomarkers of TBI. Receiver operator characteristic (ROC) curves, confidence intervals and likelihood ratios for each biomarker was determined. ROC curves, confidence intervals, sensitivity and specificity for each biomarker examined revealed that caspase-1 (0.93 area under the curve (AUC)) and ASC (0.90 AUC) in serum and ASC (1.0 AUC) and IL-18 (0.84 AUC) in CSF are promising biomarkers of TBI pathology. Importantly, higher protein levels (above 547.6 pg/ml) of ASC (0.91 AUC) were consistent with poorer outcomes after TBI as determined by the Glasgow Outcome Scale-Extended (GOSE). CONCLUSION: These findings indicate that inflammasome proteins are excellent diagnostic and predictive biomarkers of TBI.

Inflammasome Proteins in Cerebrospinal Fluid of Patients with Subarachnoid Hemorrhage are Biomarkers of Early Brain Injury and Functional Outcome.

BACKGROUND: Aneurysmal subarachnoid hemorrhage (SAH) remains a devastating disease with significant morbidity and mortality. To date, clinical variables are still used to evaluate injury severity and prognosis. This study was designed to investigate the role of cerebrospinal fluid (CSF) inflammasome proteins as laboratory-based biomarkers of brain injury severity and outcome in SAH patients. METHODS: Ten control patients and 24 SAH patients were prospectively enrolled in this study. CSF samples were collected within 72 hours after SAH. Levels of inflammasome proteins in the CSF, including NLRP1, ASC, and caspase-1 were analyzed. RESULTS: Immunoblot analysis exhibited that levels of NLRP1, ASC and caspase-1 were elevated in the CSF of SAH patients (P < 0.0001, P = 0.0178, and P < 0.0001, respectively). In the group of SAH patients, these inflammasome proteins were significantly higher in the patients with cerebral edema on computed tomographic scans (P = 0.0002, P = 0.0005, and P = 0.0004, respectively), and in the patients with acute hydrocephalus (P = 0.0009, P = 0.00017, and P = 0.0016, respectively). Higher levels of inflammasome proteins were associated with severe SAH and poor outcome 3 months after the SAH. High level of NLRP1 was the independent risk factor of poor outcome after SAH (P = 0.049, odds ratio = 1.730). CONCLUSIONS: Inflammasome proteins are potential biomarkers to assess early brain injury and to predict functional outcome after SAH.

Exosome-mediated inflammasome signaling after central nervous system injury.

Neuroinflammation is a response against harmful effects of diverse stimuli and participates in the pathogenesis of brain and spinal cord injury (SCI). The innate immune response plays a role in neuroinflammation following CNS injury via activation of multiprotein complexes termed inflammasomes that regulate the activation of caspase 1 and the processing of the pro-inflammatory cytokines IL-1ß and IL-18. We report here that the expression of components of the nucleotide-binding and oligomerization domain (NOD)-like receptor protein-1 (NLRP-1) inflammasome, apoptosis speck-like protein containing a caspase recruitment domain (ASC), and caspase 1 are significantly elevated in spinal cord motor neurons and cortical neurons after CNS trauma. Moreover, NLRP1 inflammasome proteins are present in exosomes derived from CSF of SCI and traumatic brain-injured patients following trauma. To investigate whether exosomes could be used to therapeutically block inflammasome activation in the CNS, exosomes were isolated from embryonic cortical neuronal cultures and loaded with short-interfering RNA (siRNA) against ASC and administered to spinal cord-injured animals. Neuronal-derived exosomes crossed the injured blood-spinal cord barrier, and delivered their cargo in vivo, resulting in knockdown of ASC protein levels by approximately 76% when compared to SCI rats treated with scrambled siRNA. Surprisingly, siRNA silencing of ASC also led to a significant decrease in caspase 1 activation and processing of IL-1ß after SCI. These findings indicate that exosome-mediated siRNA delivery may be a strong candidate to block inflammasome activation following CNS injury. We propose the following signaling cascade for inflammasome activation in peripheral tissues after CNS injury: CNS trauma induces inflammasome activation in the nervous system and secretion of exosomes containing inflammasome protein cargo into cerebral spinal fluid. The inflammasome containing exosomes then fuse with target cells to activate the innate immune response in peripheral tissues. We suggest that these findings may be used to develop new therapeutics to treat the devastating inflammation and cell destruction evoked by CNS injuries. IL-1ß and IL-18 = pro-inflammatory cytokines.

Inflammasome proteins in cerebrospinal fluid of brain-injured patients as biomarkers of functional outcome: clinical article.

OBJECT: Traumatic brain injury (TBI), the third most common CNS pathology, plagues 5.3 million Americans with permanent TBI-related disabilities. To evaluate injury severity and prognosis, physicians rely on clinical variables. Here, the authors seek objective, biochemical markers reflecting molecular injury mechanisms specific to the CNS as more accurate measurements of injury severity and outcome. One such secondary injury mechanism, the innate immune response, is regulated by the inflammasome, a molecular platform that activates caspase-1 and interleukin-1ß. METHODS: The authors investigated whether inflammasome components were present in the CSF of 23 patients with TBI and whether levels of inflammasome components correlate with outcome. The authors performed an immunoblot analysis of CSF samples from patients who suffered TBI and nontrauma controls and assessed the outcomes 5 months postinjury by using the Glasgow Outcome Scale. Data were analyzed using Mann-Whitney U-tests and linear regression analysis. RESULTS: Patients with severe or moderate cranial trauma exhibited significantly higher CSF levels of the inflammasome proteins ASC, caspase-1, and NALP-1 than nontrauma controls (p < 0.0001, p = 0.0029, and p = 0.0202, respectively). Expression of each protein correlated significantly with the Glasgow Outcome Scale score at 5 months postinjury (p < 0.05). ASC, caspase-1, and NALP-1 were significantly higher in the CSF of patients with unfavorable outcomes, including death and severe disability (p < 0.0001). CONCLUSIONS: NALP-1 inflammasome proteins are potential biomarkers to assess TBI severity, outcome, and the secondary injury mechanisms impeding recovery, serving as adjuncts to clinical predictors.