Astrocytic HILPDA promotes lipid droplets generation to drive cognitive dysfunction in mice with sepsis-associated encephalopathy.

AIMS: Sepsis-associated encephalopathy (SAE) is manifested as a spectrum of disturbed cerebral function ranging from mild delirium to coma. However, the pathogenesis of SAE has not been clearly elucidated. Astrocytes play important roles in maintaining the function and metabolism of the brain. Most recently, it has been demonstrated that disorders of lipid metabolism, especially lipid droplets (LDs) dyshomeostasis, are involved in a variety of neurodegenerative diseases. The aim of this study was to investigate whether LDs are involved in the underlying mechanism of SAE. METHODS: The open field test, Y-maze test, and contextual fear conditioning test (CFCT) were used to test cognitive function in SAE mice. Lipidomics was utilized to investigate alterations in hippocampal lipid metabolism in SAE mice. Western blotting and immunofluorescence labeling were applied for the observation of related proteins. RESULTS: In the current study, we found that SAE mice showed severe cognitive dysfunction, including spatial working and contextual memory. Meanwhile, we demonstrated that lipid metabolism was widely dysregulated in the hippocampus by using lipidomic analysis. Furthermore, western blotting and immunofluorescence confirmed that LDs accumulation in hippocampal astrocytes was involved in the pathological process of cognitive dysfunction in SAE mice. We verified that LDs can be inhibited by specifically suppress hypoxia-inducible lipid droplet-associated protein (HILPDA) in astrocytes. Meanwhile, cognitive dysfunction in SAE was ameliorated by reducing A1 astrocyte activation and inhibiting presynaptic membrane transmitter release. CONCLUSION: The accumulation of astrocytic lipid droplets plays a crucial role in the pathological process of SAE. HILPDA is an attractive therapeutic target for lipid metabolism regulation and cognitive improvement in septic patients.

[Cannabinoid receptor 1 agonist arachidonyl-2'-chloroethylamide (ACEA) improves sepsis-associated encephalopathy by inhibiting inflammatory factors].

Objective To investigate the impact of the cannabinoid receptor agonist arachidonyl-2'-chloroethylamide (ACEA) on cognitive function in mice with sepsis-associated encephalopathy (SAE). Methods C57BL/6 mice were randomly divided into artificial cerebrospinal fluid (ACSF) and lipopolysaccharide (LPS) groups. The SAE model was established by intraventricular injection of LPS. The severity of sepsis in mice was assessed by sepsis severity score (MSS) and body mass changes. Behavioral paradigms were used to evaluate motor ability (open field test) and cognitive function (contextual fear conditioning test, Y-maze test). To evaluate the effects of ACEA intervention on SAE, mice were randomly assigned to ACSF group, ACEA intervention combined with ACSF group, LPS group, and ACEA intervention combined with LPS group. The dosage of ACEA intervention was 1.5 mg/kg. Real-time quantitative PCR was used to measure the mRNA expression levels of interleukin 1ß (IL-1ß), IL-6, and tumor necrosis factor α (TNF-α) in mouse hippocampal tissues. Western blot analysis was used to assess the protein levels of IL-6 and TNF-α in the hippocampus. Nissl staining was performed to examine neuronal damage in the CA1 region of the mouse hippocampus. Behavioral paradigms were again employed to evaluate motor ability and cognitive function. Results Three days after intraventricular LPS injection, mice exhibited significant cognitive dysfunction, confirming SAE modeling. Compared to the control group, the LPS group showed significant increases in mRNA of inflammatory factors such as IL-6, TNF-α, and IL-1ß, together with significant increases in IL-6 and TNF-α protein levels in the hippocampus, a decrease in Nissl bodies in the CA1 region, and significant cognitive dysfunction. Compared to the LPS group, the ACEA intervention group showed a significant decrease in the mRNA of IL-6, TNF-α, and IL-1ß, a significant reduction in IL-6 and TNF-α protein levels, an increase in Nissl bodies, and improved cognitive function. Conclusion ACEA improves cognitive function in SAE mice by inhibiting the expression levels of inflammatory factors IL-6 and TNF-α.

The cytosolic DNA-sensing cGAS-STING pathway in neurodegenerative diseases.

BACKGROUND: With the widespread prevalence of neurodegenerative diseases (NDs) and high rates of mortality and disability, it is imminent to find accurate targets for intervention. There is growing evidence that neuroimmunity is pivotal in the pathology of NDs and that interventions targeting neuroimmunity hold great promise. Exogenous or dislocated nucleic acids activate the cytosolic DNA sensor cyclic GMP-AMP synthase (cGAS), activating the stimulator of interferon genes (STING). The activated STING triggers innate immune responses and then the cGAS-STING signaling pathway links abnormal nucleic acid sensing to the immune response. Recently, numerous studies have shown that neuroinflammation regulated by cGAS-STING signaling plays an essential role in NDs. AIMS: In this review, we summarized the mechanism of cGAS-STING signaling in NDs and focused on inhibitors targeting cGAS-STING. CONCLUSION: The cGAS-STING signaling plays an important role in the pathogenesis of NDs. Inhibiting the cGAS-STING signaling may provide new measures in the treatment of NDs.

Estrogen receptor GPR30 in the anterior cingulate cortex mediates exacerbated neuropathic pain in ovariectomized mice.

Menopausal women experience neuropathic pain 63% more frequently than men do, which may attribute to the estrogen withdrawal. However, the underlying mechanisms remain unclear. Here, the role of estrogen receptors (ERs) in ovariectomized (OVX) female mice following chronic constriction injury (CCI) was investigated. With 17ß-estradiol (E2) supplemented, aggravated mechanical allodynia in OVX mice could be significantly alleviated, particularly after intra-anterior cingulate cortex (ACC) E2 delivery. Pharmacological interventions further demonstrated that the agonist of G-protein-coupled estrogen receptor 30 (GPR30), rather than ERα or ERß in the ACC, exhibited the similar analgesic effect as E2, whereas antagonist of GPR30 exacerbated allodynia. Furthermore, OVX surgery reduced GPR30 expression in the ACC, which could be restored with estrogen supplementation. Selective downregulation of GPR30 in the ACC of naïve female mice induces mechanical allodynia, whereas GPR30 overexpression in the ACC remarkedly alleviated OVX-exacerbated allodynia. Collectively, estrogen withdrawal could downregulate the ACC GPR30 expression, resulting in exacerbated neuropathic pain. Our findings highlight the importance of GPR30 in the ACC in aggravated neuropathic pain during menopause, and offer a potential therapeutic candidate for neuropathic pain management in menopausal women.

[Mechanism of myeloid differentiation factor 2 on mediating sepsis-associated encephalopathy].

OBJECTIVE: To investigate the role and underlying mechanism of human myeloid differentiation protein 2 (MD2) in the process of neuronal death induced by lipopolysaccharide (LPS) by establishing an in vitro model of sepsis-associated encephalopathy (SAE) by LPS. METHODS: Healthy C57BL/6J mice at 14-18 days of gestation were selected, and brain cortical tissue was taken from fetal mice. Neurons were stimulated with 0 (control), 1, 5 and 10 g/L of LPS for 24 hours. The release of lactate dehydrogenase (LDH) was detected and the death of neurons was observed. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of inflammatory factors interleukins (IL-6, IL-1ß), in order to determine the optimal dose of LPS for establishing an in vitro neuroinflammation model of SAE. The cells were divided into blank control group and LPS group. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling (TUNEL) was used to discover apoptosis. Western blotting was used to detect the expression of the relevant protein markers activated caspase-3, necroptosis-associated protein neuronal receptor-interacting serine/threonine-protein kinase 3 (RIPK3) and phosphorylated RIPK3 (p-RIPK3). Immunofluorescence chemical staining was used to detect the expressions of p-RIPK3 and microtubule-associated protein 2 (MAP2) to evaluate the type of cell death and the degree of neuronal death. Western blotting was used to detect MD2 expression. Immunofluorescence chemical staining was performed to observe the expression and distribution of p-RIPK3 and MD2 in neurons to assess whether MD2 was involved in the inflammatory response promoting neuronal death. In addition, the cells were divided into blank control group, LPS group, and MD2 interfering peptide group (LPS+TC group), and the levels of IL-6, IL-1ß and LDH were detected to evaluate whether interfering with MD2 can alleviate LPS induced neuroinflammation. RESULTS: 10 g/L LPS induced notable neuronal death, and the release of LDH in neurons stimulated with this concentration for 24 hours was significantly higher than that in the blank control group (relative release: 1.45±0.04 vs. 1.00±0.00, P < 0.01), indicating apoptosis and necroptosis occurred in neurons, and the levels of inflammatory factors IL-6 and IL-1ß were remarkable increased [IL-6 (relative level): 1.94±0.04 vs. 1.00±0.00, IL-1ß (relative level): 1.53±0.09 vs. 1.00±0.00, both P < 0.01]. Compared with the blank control group, the apoptosis of cells, cleaved-caspase-3 expression, the p-RIPK3/RIPK3 ratio, and p-RIPK3 expression around neurons in the LPS group were significantly increased [cleaved-caspase-3/GAPDH: 1.55±0.10 vs. 1.00±0.00, P < 0.01; p-RIPK3/RIPK3 ratio (relative value): 1.54±0.06 vs. 1.00±0.00, P < 0.05], which suggested that typical apoptosis and necroptosis apoptosis occurred in neurons in the septic environment. Furthermore, MD2 expression was significantly increased in the LPS group compared with the blank control group (MD2/GAPDH: 1.91±0.07 vs. 1.00±0.00, P < 0.01), and MD2 expression around neurons was increased, indicating that LPS-induced MD2 upregulation may play a key role in neuroinflammation and induction of neuronal death in sepsis. In addition, compared with the LPS group, the MD2-interfering peptide could reduce the expression levels of inflammatory factors IL-6 and IL-1ß [IL-6 (relative level): 1.16±0.08 vs. 1.94±0.04, IL-1ß (relative level): 1.15±0.05 vs. 1.75±0.09, both P < 0.01] and decrease LDH release (relative release: 1.09±0.01 vs. 1.44±0.04, P < 0.05). CONCLUSIONS: LPS induced neuronal inflammatory responses via MD2, which ultimately leads to apoptosis and necroptosis. Interfering with MD2 reduces inflammation and inhibits neuronal death.

Screening Serum Biomarkers for Rats Preconditioned with Hyperbaric Oxygen: Potential of Predicting Prognosis for Stroke.

BACKGROUND: Stroke is a major health concern and a leading cause of mortality and morbidity. We and other groups have documented that hyperbaric oxygen preconditioning could significantly alleviate neuronal damage in ischemia‒reperfusion models through various mechanisms. However, we found that some of the subjects did not benefit from preconditioning with hyperbaric oxygen. The preconditioning phenomenon is similar to vaccination, in which the endogenous survival system is activated to fight against further injuries. However, with vaccine inoculations, we could test for specific antibodies against the pathogens to determine if the vaccination was successful. Likewise, this experiment was carried out to explore a biomarker that can reveal the effectiveness of the preconditioning before neuronal injury occurs. METHODS: Middle cerebral artery occlusion (MCAO) was used to induce focal cerebral ischemia-reperfusion injury. 2D-DIGE-MALDI-TOF-MS/MS proteomic technique was employed to screen the differentially expressed proteins in the serum of rats among the control (Con) group (MCAO model without hyperbaric oxygen (HBO) preconditioning), hyperbaric oxygen protective (HBOP) group (in which the infarct volume decreased after HBO preconditioning vs. Con), and hyperbaric oxygen nonprotective (HBOU) group (in which the infarct volume remained the same or even larger after HBO preconditioning vs. Con). Candidate biomarkers were confirmed by western blot and enzyme linked immunosorbent assay (ELISA), and the relationship between the biomarkers and the prognosis of cerebral injury was further validated. RESULTS: Among the 15 differentially expressed protein spots detected in the HBOP group by Two-dimensional fluorescence difference gel electrophoresis (2D-DIGE), 3 spots corresponding to 3 different proteins (haptoglobin, serum albumin, and haemopexin) products were identified by MALDI-TOF-MS/MS. Serum albumin and haemopexin were upregulated, and haptoglobin was downregulated in the HBOP group (p < 0.05 vs. Con and HBOU groups). After the western blot study, only the changes in haemopexin were validated and exhibited similar changes in subjects from the HBOP group in accordance with MALDI-TOF-MS/MS proteomic analysis and enzyme linked immunosorbent assay (ELISA) analysis. The serum level of the hemopexin (HPX) at 2 h after HBO preconditioning was correlated with the infarct volume ratio after MCAO. CONCLUSIONS: Haemopexin may be developed as a predictive biomarker that indicated the effectiveness of a preconditioning strategy against cerebral ischaemic injury.

Autophagy Suppresses Ferroptosis by Degrading TFR1 to Alleviate Cognitive Dysfunction in Mice with SAE.

Sepsis-associated encephalopathy (SAE) is a serious complication of sepsis that is characterized by long-term cognitive impairment, which imposes a heavy burden on families and society. However, its pathological mechanism has not been elucidated. Ferroptosis is a novel form of programmed cell death that is involved in multiple neurodegenerative diseases. In the current study, we found that ferroptosis also participated in the pathological process of cognitive dysfunction in SAE, while Liproxstatin-1 (Lip-1) effectively inhibited ferroptosis and alleviated cognitive impairment. Additionally, since an increasing number of studies have suggested the crosstalk between autophagy and ferroptosis, we further proved the essential role of autophagy in this process and demonstrated the key molecular mechanism of the autophagy-ferroptosis interaction. Currently, we showed that autophagy in the hippocampus was downregulated within 3 days of lipopolysaccharide injection into the lateral ventricle. Moreover, enhancing autophagy ameliorated cognitive dysfunction. Importantly, we found that autophagy suppressed ferroptosis by downregulating transferrin receptor 1 (TFR1) in the hippocampus, thereby alleviating cognitive impairment in mice with SAE. In conclusion, our findings indicated that hippocampal neuronal ferroptosis is associated with cognitive impairment. In addition, enhancing autophagy can inhibit ferroptosis via degradation of TFR1 to ameliorate cognitive impairment in SAE, which shed new light on the prevention and therapy for SAE.

Chaperone-mediated autophagy (CMA) alleviates cognitive impairment by reducing neuronal death in sepsis-associated encephalopathy (SAE).

Sepsis-associated encephalopathy (SAE) is a common and severe complication of sepsis, which causes long-term neurological deficits, such as cognitive impairment. Despite extensive research, there is still lack of specific treatments for SAE. Chaperone-mediated autophagy (CMA), a selective type of autophagy, has been reported to be related to cognitive dysfunctions in many neurodegenerative diseases. The aim of this study was to investigate the alteration of CMA activity in the hippocampus of SAE mice and explore the neuroprotective effect of enhanced CMA. Cecal ligation and puncture (CLP) was conducted to induce SAE. In the contextual fear conditioning test, the ratio of freezing time of CLP mice significantly decreased compared with that of the mice in the Sham group, indicating cognitive impairment in SAE mice. The expression of lysosome-associated membrane protein type 2A (Lamp2a) and chaperone heat shock cognate 71 kDa protein (Hsc70), positive markers for CMA activity, decreased in hippocampal neurons of SAE mice. Although overexpression of Lamp2a in neurons via adeno-associated virus injection in the hippocampus had little effect on the mortality of septic mice, this intervention significantly alleviated the memory impairments in contextual fear conditioning test, Y-maze test and novel objective recognition test, and attenuated the neural death observed in SAE mice. We further demonstrated that the overexpression of Lamp2a in the hippocampus increased the expression of phosphorylated cyclic-AMP response element binding protein (p-CREB), brain-derived neurotrophic factor (BDNF) and B-cell lymphoma-2 (Bcl-2), and suppressed the expression of cleaved caspase-3. Taken together, our study results suggested that the upregulation of CMA activity ameliorated cognitive impairments and neuron loss in SAE mice partially through the p-CREB-BDNF/Bcl-2 signaling pathways, providing a potential therapeutic target for SAE.

The Role of Ferroptosis in Nervous System Disorders.

Ferroptosis is distinct from other apoptotic forms of programmed cell death and is characterized by the accumulation of iron and lipid peroxidation. Iron plays a crucial role in the oxidation of lipids via the Fenton reaction with oxygen. Hence, iron accumulation causes phospholipid peroxidation which induces ferroptosis. Moreover, detoxification by glutathione is disrupted during ferroptosis. A growing number of studies have implicated ferroptosis in nervous system disorders such as depression, neurodegenerative disease, stroke, traumatic brain injury, and sepsis-associated encephalopathy. This review summarizes the pathogenesis of ferroptosis and its relationship with various nervous system disorders.

Electroacupuncture pretreatment induces ischemic tolerance by neuronal TREM2-mediated enhancement of autophagic flux.

The mechanism of electroacupuncture (EA) pretreatment-induced neuroprotection remains unclear. In this study, we found that neuronal Triggering receptor expressed on myeloid cells 2 (TREM2) expression was increased and peaked at 48 h and 72 h after ischemia/reperfusion. After specific knockdown of TREM2 in excitatory neurons, neurological function was damaged, and the infarct volume was enlarged. Furthermore, the expression of LC3II/LC3I and Beclin1 was decreased, while the expression of p62 was increased. EA pretreatment enhanced TREM2, LC3II/LC3I and Beclin1 expression while reducing p62 in the ischemic penumbra area. The EA-induced neuroprotective effects and improvements in autophagic flux were abolished by specific knockdown of TREM2 in excitatory neurons. Taken together, our findings provide novel mechanistic insight into EA-induced ischemic tolerance and suggest a promising therapeutic strategy of targeting neuronal TREM2 to treat brain ischemia.

[Rapamycin alleviates cognitive impairment in mice with sepses-associated encephalopathy by promoting autophagy].

Objective To investigate the role of rapamycin in alleviating cognitive dysfunction by promoting autophagy in mice with sepsis-associated encephalopathy (SAE). Methods The model of SAE mice was established by caecal ligation and perforation (CLP). Murine sepsis score (MSS) was used to evaluate the severity of sepsis in SAE mice. And the cognitive function was tested by the contextual fear conditioning test. The expression levels of microtubule-associated protein 1 light chain 3 (LC3) and P62 in the hippocampus of the SAE mice were detected by Western blot analysis. Furthermore, the expression and distribution of LC3 in the hippocampal neurons were observed by immunofluorescence. Results The mortality of CLP-induced mice reached 41.7% with 14 days after the procedure, and significant cognitive dysfunction was detected in the surviving mice. Meanwhile, autophagy in the hippocampal tissue was impaired 14 days after CLP. The cognitive dysfunction of SAE mice was alleviated by promoting autophagy via rapamycin. Conclusion Rapamycin alleviated the cognitive dysfunction of SAE mice by promoting autophagy in the hippocampal neurons.

Combined-Acupoint Electroacupuncture Induces Better Analgesia via Activating the Endocannabinoid System in the Spinal Cord.

Electroacupuncture (EA) therapy has been widely reported to alleviate neuropathic pain with few side effects in both clinical practice and animal studies worldwide. However, little is known about the comparison of the therapeutic efficacy among the diverse EA schemes used for neuropathic pain. The present study is aimed at investigating the therapeutic efficacy discrepancy between the single and combined-acupoint EA and to reveal the difference of mechanisms behind them. Electroacupuncture was given at both Zusanli (ST36) and Huantiao (GB30) in the combined group or ST36 alone in the single group. Paw withdrawal mechanical threshold (PWMT) was measured to determine the pain level. Electrophysiology was performed to detect the effects of EA on synaptic transmission in the spinal dorsal horn of the vGlut2-tdTomato mice. Spinal contents of endogenous opioids, endocannabinoids, and their receptors were examined. Inhibitors of CBR (cannabinoid receptor) and opioid receptors were used to study the roles of opioid and endocannabinoid system (ECS) in EA analgesia. We found that combined-acupoint acupuncture provide stronger analgesia than the single group did, and the former inhibited the synaptic transmission at the spinal level to a greater extent than later. Besides, the high-intensity stimulation at ST36 or normal stimulation at two sham acupoints did not mimic the similar efficacy of analgesia in the combined group. Acupuncture stimulation in single and combined groups both activated the endogenous opioid system. The ECS was only activated in the combined group. Naloxone totally blocked the analgesic effect of single-acupoint EA; however, it did not attenuate that of combined-acupoint EA unless coadministered with CBR antagonists. Hence, in the CCI-induced neuropathic pain model, combined-acupoint EA at ST36 and GB30 is more effective in analgesia than the single-acupoint EA at ST36. EA stimulation at GB30 alone neither provided a superior analgesic effect to EA treatment at ST36 nor altered the content of AEA, 2-AG, CB1 receptor, or CB2 receptor compared with the CCI group. Activation of the ECS is the main contributor of the better analgesia by the combined acupoint stimulation than that induced by single acupoint stimulation.

Machine Learning Prediction Models for Postoperative Stroke in Elderly Patients: Analyses of the MIMIC Database.

Objective: With the aging of populations and the high prevalence of stroke, postoperative stroke has become a growing concern. This study aimed to establish a prediction model and assess the risk factors for stroke in elderly patients during the postoperative period. Methods: ML (Machine learning) prediction models were applied to elderly patients from the MIMIC (Medical Information Mart for Intensive Care)-III and MIMIC-VI databases. The SMOTENC (synthetic minority oversampling technique for nominal and continuous data) balancing technique and iterative SVD (Singular Value Decomposition) data imputation method were used to address the problem of category imbalance and missing values, respectively. We analyzed the possible predictive factors of stroke in elderly patients using seven modeling approaches to train the model. The diagnostic value of the model derived from machine learning was evaluated by the ROC curve (receiver operating characteristic curve). Results: We analyzed 7,128 and 661 patients from MIMIC-VI and MIMIC-III, respectively. The XGB (extreme gradient boosting) model got the highest AUC (area under the curve) of 0.78 (0.75-0.81), making it better than the other six models, Besides, we found that XGB model with databalancing was better than that without data balancing. Based on this prediction model, we found hypertension, cancer, congestive heart failure, chronic pulmonary disease and peripheral vascular disease were the top five predictors. Furthermore, we demonstrated that hypertension predicted postoperative stroke is much more valuable. Conclusion: Stroke in elderly patients during the postoperative period can be reliably predicted. We proved XGB model is a reliable predictive model, and the history of hypertension should be weighted more heavily than the results of laboratory tests to prevent postoperative stroke in elderly patients regardless of gender.

Neuronal MD2 induces long-term mental impairments in septic mice by facilitating necroptosis and apoptosis.

Sepsis-associated encephalopathy (SAE) is a complication of sepsis with high morbidity rates. Long-lasting mental health issues in patients with SAE result in a substantial decrease in quality of life. However, its underlying mechanism is unclear, and effective treatments are not available. In the current study, we explored the role of apoptosis and necroptosis related to mental dysfunction in sepsis. In a mouse model of sepsis constructed by cecal ligation and puncture (CLP), altered behavior was detected by the open field, elevated-plus maze and forced swimming tests on the fourteenth day. Moreover, apoptosis- and necroptosis-associated proteins and morphological changes were examined in the hippocampus of septic mice. Long-lasting depression-like behaviors were detected in the CLP mice, as well as significant increases in neuronal apoptosis and necroptosis. Importantly, we found that apoptosis and necroptosis were related according to Ramsay's rule in the brains of the septic mice. Inhibiting myeloid differentiation factor 2 (MD2), the crosstalk mediator of apoptosis and necroptosis, in neurons effectively reduced neuronal loss and alleviated depression-like behaviors in the septic mice. These results suggest that neuronal death in the hippocampus contributes to the mental impairments in SAE and that inhibiting neuronal MD2 is a new strategy for treating mental health issues in sepsis by inhibiting necroptosis and apoptosis.

Pioglitazone attenuates ischaemic stroke aggravation by blocking PPARγ reduction and inhibiting chronic inflammation in diabetic mice.

Diabetes can cause vascular remodelling and is associated with worse outcome after ischaemic stroke. Pioglitazone is a commonly used anti-diabetic agent. However, it is not known whether pioglitazone use before ischaemia could reduce brain ischaemic injury. Pioglitazone was administered to 5-week-old db+ or db/db mice. Cerebral vascular remodelling was examined at the age of 9 weeks. Expression of peroxisome proliferator-activated receptor-γ (PPARγ), p-PPARγ (S112 and S273), nucleotide-binding domain (NOD)-like receptor protein 3 (Nlrp3), interleukin-1ß (IL-1ß) and tumour necrosis factor-α (TNF-α) was evaluated in the somatosensory cortex of mice. Neurological outcome was evaluated 24 h after brain ischaemia. Results showed that early pioglitazone treatment provided a long-lasting effect of euglycaemia but enhanced hyperlipidaemia in the db/db mice. Diabetic mice exhibited increased vascular tortuosity, narrower middle cerebral artery (MCA) width and IgG leakage in the brain. These changes were blocked by early pioglitazone treatment. In diabetic animals, PPARγ expression was reduced, and p-PPARγ at S273 but not S112, Nlrp3, IL-1ß and TNF-α were increased in the somatosensory cortex. PPARγ decrease and Nlrp3 increase were mainly in the neurons of the diabetic brain, which was reversed by early pioglitazone treatment. Pioglitazone attenuated the aggravated neurological outcome after stroke in diabetic mice. But this protective effect was abolished through restoring cerebral inflammation by intracerebroventricular administration of IL-1ß and TNF-α in pioglitazone-treated diabetic mice before MCAO. In summary, early pioglitazone treatment attenuates cerebral vascular remodelling and ischaemic brain injury possibly via blocking chronic neuroinflammation in the db/db mice.

Roles and Molecular Mechanisms of Physical Exercise in Sepsis Treatment.

Physical exercise is a planned, purposeful action to keep a healthy lifestyle and improve physical fitness. Physical exercise has been widely used as a non-pharmacological approach to preventing and improving a wide range of diseases, including cardiovascular disease, cancer, metabolic disease, and neurodegenerative disease. However, the effects of physical exercise on sepsis have not been summarized until now. In this review, we discuss the effects of physical exercise on multiple organ functions and the short- and long-time outcomes of sepsis. Furthermore, the molecular mechanisms underlying the protective effects of physical exercise on sepsis are discussed. In conclusion, we consider that physical exercise may be a beneficial and non-pharmacological alternative for the treatment of sepsis.

Neuronal GPER Participates in Genistein-Mediated Neuroprotection in Ischemic Stroke by Inhibiting NLRP3 Inflammasome Activation in Ovariectomized Female Mice.

Estrogen replacement therapy (ERT) is potentially beneficial for the prevention and treatment of postmenopausal cerebral ischemia but inevitably increases the risk of cerebral hemorrhage and breast cancer when used for a long period of time. Genistein, a natural phytoestrogen, has been reported to contribute to the recovery of postmenopausal ischemic stroke with reduced risks. However, the underlying mechanism of genistein-mediated neuroprotection remains unclear. We reported that genistein exerted significant neuroprotective effects by enhancing the expression of neuronal G protein-coupled estrogen receptor (GPER) in the ischemic penumbra after cerebral reperfusion in ovariectomized (OVX) mice, and this effect was achieved through GPER-mediated inhibition of nod-like receptor protein 3 (NLRP3) inflammasome activation. In addition, we found that peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) was the pivotal molecule that participated in GPER-mediated inhibition of NLRP3 inflammasome activation in OVX mice after ischemia/reperfusion (I/R) injury. Our data suggest that the neuronal GPER/PGC-1α pathway plays an important role in genistein-mediated neuroprotection against I/R injury in OVX mice.

Sex-specific cannabinoid 1 receptors on GABAergic neurons in the ventrolateral periaqueductal gray mediate analgesia in mice.

Sex differences in analgesic effects have gradually attracted public attention in preclinical and clinical studies. Both human and animal females are more sensitive to cannabinoid antinociception than males. Expression of the cannabinoid 1 receptor (CB1 R) and the function of the endocannabinoid system have been explored in both male and female mice and CB1 Rs in the ventrolateral periaqueductal gray (vlPAG) participate in antinociception. However, whether there are cell-type- and sex-specific patterns of vlPAG CB1 R expression that affect analgesia is unknown. In the current study, we either activated or inhibited CB1 Rs in the vlPAG and found that female mice produced stronger analgesia or developed more robust mechanical allodynia than males did. Specific deletion of GABAergic CB1 Rs in the vlPAG promoted stronger mechanical allodynia in female mice than that in male mice. However, no sex differences in cannabinoid antinociception were found following chemogenetic inhibition of GABAergic neurons. Using fluorescence in situ hybridization, we found that the sex difference in cannabinoid antinociception was due to females having higher expression of GABAergic CB1 Rs in the vlPAG than males. Furthermore, activation of CB1 Rs in the vlPAG significantly reduced the frequency of GABA-mediated spontaneous inhibitory postsynaptic currents recorded in vGlut2-tdTomato positive neurons in both sexes. This effect was greater in females than males and this reduction was closely related to CB1 R expression difference between sexes. Our work indicates that vlPAG GABAergic CB1 Rs modulate cannabinoid-mediated analgesia in a sex-specific manner, which may provide a potential explanation of sex difference found in the analgesic effect of cannabinoids.

MD2 contributes to the pathogenesis of perioperative neurocognitive disorder via the regulation of α5GABAA receptors in aged mice.

BACKGROUND: Perioperative neurocognitive disorder (PND) is a long-term postoperative complication in elderly surgical patients. The underlying mechanism of PND is unclear, and no effective therapies are currently available. It is believed that neuroinflammation plays an important role in triggering PND. The secreted glycoprotein myeloid differentiation factor 2 (MD2) functions as an activator of the Toll-like receptor 4 (TLR4) inflammatory pathway, and α5GABAA receptors (α5GABAARs) are known to play a key role in regulating inflammation-induced cognitive deficits. Thus, in this study, we aimed to investigate the role of MD2 in PND and determine whether α5GABAARs are involved in the function of MD2. METHODS: Eighteen-month-old C57BL/6J mice were subjected to laparotomy under isoflurane anesthesia to induce PND. The Barnes maze was used to assess spatial reference learning and memory, and the expression of hippocampal MD2 was assayed by western blotting. MD2 expression was downregulated by bilateral injection of AAV-shMD2 into the hippocampus or tail vein injection of the synthetic MD2 degrading peptide Tat-CIRP-CMA (TCM) to evaluate the effect of MD2. Primary cultured neurons from brain tissue block containing cortices and hippocampus were treated with Tat-CIRP-CMA to investigate whether downregulating MD2 expression affected the expression of α5GABAARs. Electrophysiology was employed to measure tonic currents. For α5GABAARs intervention experiments, L-655,708 and L-838,417 were used to inhibit or activate α5GABAARs, respectively. RESULTS: Surgery under inhaled isoflurane anesthesia induced cognitive impairments and elevated the expression of MD2 in the hippocampus. Downregulation of MD2 expression by AAV-shMD2 or Tat-CIRP-CMA improved the spatial reference learning and memory in animals subjected to anesthesia and surgery. Furthermore, Tat-CIRP-CMA treatment decreased the expression of membrane α5GABAARs and tonic currents in CA1 pyramidal neurons in the hippocampus. Inhibition of α5GABAARs by L-655,708 alleviated cognitive impairments after anesthesia and surgery. More importantly, activation of α5GABAARs by L-838,417 abrogated the protective effects of Tat-CIRP-CMA against anesthesia and surgery-induced spatial reference learning and memory deficits. CONCLUSIONS: MD2 contributes to the occurrence of PND by regulating α5GABAARs in aged mice, and Tat-CIRP-CMA is a promising neuroprotectant against PND.

Adiponectin Treatment Attenuates Cerebral Ischemia-Reperfusion Injury through HIF-1α-Mediated Antioxidation in Mice.

Adiponectin (ADPN) plays an important role in cerebral ischemia-reperfusion injury. Although previous studies have confirmed that ADPN pretreatment has a protective effect on ischemic stroke, the therapeutic effect of ADPN on ischemic stroke and the underlying mechanism are still unclear. In order to clarify these questions, focal transient cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) in mice and ADPN was administered for three times at 6 h, 24 h, and 48 h after reperfusion. Meanwhile, a virus-delivered HIF-1α siRNA was used before ADPN administration. The infarct volume, neurological score, cellular apoptosis, and oxidative stress were assessed at 72 h after reperfusion. The long-term outcome of mice after stroke was recorded as well. The results indicated that ADPN treatment reduced the infarct volume (P = 0.032), neurological deficits (P = 0.047), cellular apoptosis (P = 0.041), and oxidative responses (P = 0.031) at 72 h after MCAO. Moreover, ADPN increased both the protein level and transcriptional activity of HIF-1α as evidenced by the transcription levels of VEGF (P = 0.046) and EPO (P = 0.043) at 72 h after MCAO. However, knockdown of HIF-1α partially reversed the antioxidant and treatment effect of ADPN after cerebral ischemia. In the observation of long-term outcome after ADPN treatment, it demonstrated that ADPN not only prevented the cerebral atrophy (P = 0.031) and the neurological function decline (P = 0.048), but also promoted angiogenesis (P = 0.028) after stroke. In conclusion, our findings suggest that ADPN is effective in treatment of ischemic stroke which could be attributed to the increased antioxidant capacity regulated by HIF-1α.