Single-cell transcriptomics analysis of cellular heterogeneity and immune mechanisms in neurodegenerative diseases.

Single-cell transcriptomics analysis is an advanced technology that can describe the intracellular transcriptome in complex tissues. It profiles and analyses datasets by single-cell RNA sequencing. Neurodegenerative diseases are identified by the abnormal apoptosis of neurons in the brain with few or no effective therapy strategies at present, which has been a growing healthcare concern and brought a great burden to society. The transcriptome of individual cells provides deep insights into previously unforeseen cellular heterogeneity and gene expression differences in neurodegenerative disorders. It detects multiple cell subsets and functional changes during pathological progression, which deepens the understanding of the molecular underpinnings and cellular basis of neurodegenerative diseases. Furthermore, the transcriptome analysis of immune cells shows the regulation of immune response. Different subtypes of immune cells and their interaction are found to contribute to disease progression. This finding enables the discovery of novel targets and biomarkers for early diagnosis. In this review, we emphasize the principles of the technology, and its recent progress in the study of cellular heterogeneity and immune mechanisms in neurodegenerative diseases. The application of single-cell transcriptomics analysis in neurodegenerative disorders would help explore the pathogenesis of these diseases and develop novel therapeutic methods.

Relationship between TyG index and the degree of coronary artery lesions in patients with H-type hypertension.

BACKGROUND: The TyG index, a prominent metric for assessing insulin resistance, has gained traction as a prognostic tool for cardiovascular disease. Nevertheless, the understanding of the prognostic significance of the extent of coronary artery stenosis in individuals afflicted with H-type hypertension remains limited. METHODS: A retrospective study was conducted at Wuhan Third Hospital, including a cohort of 320 inpatients who were diagnosed with hypertension in combination with coronary artery disease. The study period spanned from January 1, 2021, to February 1, 2023. The study cohort was stratified based on the severity of stenosis into three distinct groups: low stenosis, medium stenosis, and high stenosis, as determined by the Gensini score derived from coronary angiography findings. The present study aimed to investigate the association between the severity of coronary stenosis and the number of lesion branches, utilizing the TyG index as a testing indicator. The predictive ability of TyG for coronary lesion severity was assessed using logistic regression analysis. RESULTS: The results of our study indicate a positive correlation between elevated levels of TyG and an increased susceptibility to severe stenosis in individuals diagnosed with H-type hypertension. Upon careful consideration of potential confounding variables, it has been observed that the TyG index exhibits a robust association with the likelihood of severe stenosis in individuals with H-type hypertension (odds ratio [OR] = 4000, 95% confidence interval CI 2.411-6.635, p = 0.0001), as well as the prevalence of multivessel disease (OR = 1.862, 95% CI 1.036-3.348, p < 0.0001). The TyG index demonstrated superior predictive ability for severe coronary stenosis in patients with H-type hypertension compared to those without H-type hypertension (area under the curve [AUC] = 0.888, 95% confidence interval CI 0.838-0.939, p < 0.0001, versus AUC = 0.615, 95% CI 0.494-0.737, p < 0.05). CONCLUSION: The TyG index is an independent risk factor for the degree of coronary stenosis and a better predictor in patients with H-type hypertension combined with coronary artery disease.

Potentials of circSOBP in the diagnosis and prognosis of gastric cancer.

OBJECTIVES: Gastric cancer (GC) causes no symptoms at early stages. However, with the progression, GC causes symptoms mimicking normal gastrointestinal issues, such as irritable bowel syndrome (IBS), gastroesophageal reflux disease (GERD), gastritis, or peptic ulcers (PU). CircRNA circSOBP has been characterized as a critical regulator in prostate cancer. The present study aimed to study its involvement in GC. MATERIALS AND METHODS: Plasma samples were collected from GC patients (n = 64), IBS patients (n = 64), GERD patients (n = 64), gastritis patients (n = 64), PU patients (n = 64), and healthy controls (HCs, n = 64). Paired GC and non-tumor samples were from all GC patients (n = 64). Tissue and plasma samples were subjected to RT-qPCR to determine circSOBP expression. The role of circSOBP in distinguishing GC patients from other patients was analyzed by ROC curve. The 64 patients were followed up for 5 years to study the role of circSOBP in predicting the survival of GC patients. RESULTS: Decreased circSOBP RNA accumulation was observed in GC tissues compared to normal tissue samples. Decreased plasma circSOBP accumulation was only observed in GC patients, but not other patients, compared to HCs. With plasma circSOBP as a biomarker, GC patients were separated from other patients and HCs. Patients with high plasma or tissue levels of circSOBP showed better survival conditions. In addition, plasma and tissue circSOBP levels were only closely correlated with GC patients' tumor metastasis, but not other clinical factors. CONCLUSIONS: Decreased circSOBP accumulation may be applied in clinical practice to improve the diagnosis and prognosis of GC.

Multiple organ injury on admission predicts in-hospital mortality in patients with COVID-19.

Multiorgan injury has been implicated in patients with coronavirus disease 2019 (COVID-19). We aim to assess the impact of organ injury (OI) on prognosis according to the number of affected organs at admission. This is a retrospective cohort study of patients with confirmed COVID-19 in Wuhan Third Hospital & Tongren Hospital of Wuhan University from February 17 to March 22, 2020. We classified the patients according to the presence and number of damaged organs (heart, liver, and kidney). The percentage of patients with no, one, two, or three organs affected was 59.75%, 30.46%, 8.07%, and 1.72%, respectively. With the increasing number of OI, there is a tendency of gradual increase regarding the white blood cell counts, neutrophil counts, levels of C-reactive protein (CRP), lactate dehydrogenase, D-dimer, and fibrinogen as well as the incidence of most complications. In a Cox regression model, individuals with OI, old age, and an abnormal level of CRP were at a higher risk of death compared with those without. Patients with three organ injuries had the highest mortality rate (57.9%; hazard ratio [HR] with 95% confidence interval [CI] vs. patients without OI: 22.31 [10.42-47.77], those with two [23.6%; HR = 8.68, 95% CI = 4.58-16.48], one [8.6%; HR = 3.1, 95% CI = 1.7-5.7], or no OI [2.6%]; p < .001). The increasing number of OI was associated with a high risk of mortality in COVID-19 infection.

Neutrophil to lymphocyte ratio as prognostic and predictive factor in patients with coronavirus disease 2019: A retrospective cross-sectional study.

This retrospective study was designed to explore whether neutrophil to lymphocyte ratio (NLR) is a prognostic factor in patients with coronavirus disease 2019 (COVID-19). A cohort of patients with COVID-19 admitted to the Tongren Hospital of Wuhan University from 11 January 2020 to 3 March 2020 was retrospectively analyzed. Patients with hematologic malignancy were excluded. The NLR was calculated by dividing the neutrophil count by the lymphocyte count. NLR values were measured at the time of admission. The primary outcome was all-cause in-hospital mortality. A multivariate logistic analysis was performed. A total of 1004 patients with COVID-19 were included in this study. The mortality rate was 4.0% (40 cases). The median age of nonsurvivors (68 years) was significantly older than survivors (62 years). Male sex was more predominant in nonsurvival group (27; 67.5%) than in the survival group (466; 48.3%). NLR value of nonsurvival group (median: 49.06; interquartile range [IQR]: 25.71-69.70) was higher than that of survival group (median: 4.11; IQR: 2.44-8.12; P < .001). In multivariate logistic regression analysis, after adjusting for confounding factors, NLR more than 11.75 was significantly correlated with all-cause in-hospital mortality (odds ratio = 44.351; 95% confidence interval = 4.627-425.088). These results suggest that the NLR at hospital admission is associated with in-hospital mortality among patients with COVID-19. Therefore, the NLR appears to be a significant prognostic biomarker of outcomes in critically ill patients with COVID-19. However, further investigation is needed to validate this relationship with data collected prospectively.

Interleukin-1beta released by microglia initiates the enhanced glutamatergic activity in the spinal dorsal horn during paclitaxel-associated acute pain syndrome.

Patients receiving paclitaxel for cancer treatment often develop an acute pain syndrome (paclitaxel-associated acute pain syndrome, P-APS), which occurs immediately after paclitaxel treatment. Mechanisms underlying P-APS remain largely unknown. We recently reported that rodents receiving paclitaxel develop acute pain and activation of spinal microglial toll like receptor 4 (TLR4) by paclitaxel penetrating into the spinal cord is a critical event in the genesis of P-APS. Our current study dissected cellular and molecular mechanisms underlying the P-APS. We demonstrated that bath-perfusion of paclitaxel, at a concentration similar to that found in the cerebral spinal fluid in animals receiving i.v. paclitaxel (2 mg/kg), resulted in increased calcium activity in microglia instantly, and in astrocytes with 6 min delay. TLR4 activation in microglia by paclitaxel caused microglia to rapidly release interleukin-1ß (IL-1ß) but not tumor necrosis factor α, IL-6, or interferon-γ. IL-1ß release from microglia depended on capthepsin B. IL-1ß acted on astrocytes, leading to elevated calcium activity and suppressed glutamate uptake. IL-1ß also acted on neurons to increase presynaptic glutamate release and postsynaptic AMPA receptor activity in the spinal dorsal horn. Knockout of IL-1 receptors prevented the development of acute pain induced by paclitaxel in mice. Our study indicates that IL-1ß is a crucial molecule used by microglia to alter functions in astrocytes and neurons upon activation of TLR4 in the genesis of P-APS, and targeting the signaling pathways regulating the production and function of IL-1ß from microglia is a potential avenue for the development of analgesics for the treatment of P-APS.

Chronic pain and impaired glial glutamate transporter function in lupus-prone mice are ameliorated by blocking macrophage colony-stimulating factor-1 receptors.

Systemic lupus erythematosus (SLE) is a multi-organ disease of unknown etiology in which the normal immune responses are directed against the body's own healthy tissues. Patients with SLE often suffer from chronic pain. Currently, no animal studies have been reported about the mechanisms underlying pain in SLE. In this study, the development of chronic pain in MRL lupus-prone (MRL/lpr) mice, a well-established lupus mouse model, was characterized for the first time. We found that female MRL/lpr mice developed thermal hyperalgesia at the age of 13 weeks, and mechanical allodynia at the age of 16 weeks. MRL/lpr mice with chronic pain had activation of microglia and astrocytes, over-expression of macrophage colony-stimulating factor-1 (CSF-1) and interleukin-1 beta (IL-1ß), as well as suppression of glial glutamate transport function in the spinal cord. Intrathecal injection of either the CSF-1 blocker or IL-1 inhibitor attenuated thermal hyperalgesia in MRL/lpr mice. We provide evidence that the suppressed activity of glial glutamate transporters in the spinal dorsal horn in MRL/lpr mice is caused by activation of the CSF-1 and IL-1ß signaling pathways. Our findings suggest that targeting the CSF-1 and IL-1ß signaling pathways or the glial glutamate transporter in the spinal cord is an effective approach for the management of chronic pain caused by SLE.

Blocking the GABA transporter GAT-1 ameliorates spinal GABAergic disinhibition and neuropathic pain induced by paclitaxel.

Paclitaxel is a chemotherapeutic agent widely used for treating carcinomas. Patients receiving paclitaxel often develop neuropathic pain and have a reduced quality of life which hinders the use of this life-saving drug. In this study, we determined the role of GABA transporters in the genesis of paclitaxel-induced neuropathic pain using behavioral tests, electrophysiology, and biochemical techniques. We found that tonic GABA receptor activities in the spinal dorsal horn were reduced in rats with neuropathic pain induced by paclitaxel. In normal controls, tonic GABA receptor activities were mainly controlled by the GABA transporter GAT-1 but not GAT-3. In the spinal dorsal horn, GAT-1 was expressed at presynaptic terminals and astrocytes while GAT-3 was only expressed in astrocytes. In rats with paclitaxel-induced neuropathic pain, the protein expression of GAT-1 was increased while GAT-3 was decreased. This was concurrently associated with an increase in global GABA uptake. The paclitaxel-induced attenuation of GABAergic tonic inhibition was ameliorated by blocking GAT-1 but not GAT-3 transporters. Paclitaxel-induced neuropathic pain was significantly attenuated by the intrathecal injection of a GAT-1 inhibitor. These findings suggest that targeting GAT-1 transporters for reversing disinhibition in the spinal dorsal horn may be a useful approach for treating paclitaxel-induced neuropathic pain. Patients receiving paclitaxel for cancer therapy often develop neuropathic pain and have a reduced quality of life. In this study, we demonstrated that animals treated with paclitaxel develop neuropathic pain, have enhancements of GABA transporter-1 protein expression and global GABA uptake, as well as suppression of GABAergic tonic inhibition in the spinal dorsal horn. Pharmacological inhibition of GABA transporter-1 ameliorates the paclitaxel-induced suppression of GABAergic tonic inhibition and neuropathic pain. Thus, targeting GAT-1 transporters for reversing GABAergic disinhibition in the spinal dorsal horn could be a useful approach for treating paclitaxel-induced neuropathic pain.

Paclitaxel induces acute pain via directly activating toll like receptor 4.

Paclitaxel, a powerful anti-neoplastic drug, often causes pathological pain, which significantly reduces the quality of life in patients. Paclitaxel-induced pain includes pain that occurs immediately after paclitaxel treatment (paclitaxel-associated acute pain syndrome, P-APS) and pain that persists for weeks to years after cessation of paclitaxel treatment (paclitaxel induced chronic neuropathic pain). Mechanisms underlying P-APS remain unknown. In this study, we found that paclitaxel causes acute pain in rodents in a dose-dependent manner. The paclitaxel-induced acute pain occurs within 2 hrs after a single intravenous injection of paclitaxel. This is accompanied by low levels of paclitaxel penetrating into the cerebral spinal fluid and spinal dorsal horn. We demonstrated that an intrathecal injection of paclitaxel induces mechanical allodynia in a dose-dependent manner. Paclitaxel causes activation of toll like receptor 4 (TLR4) in the spinal dorsal horn and dorsal root ganglions. Through activating TLR4, paclitaxel increases glutamatergic synaptic activities and reduces glial glutamate transporter activities in the dorsal horn. Activations of TLR4 are necessary in the genesis of paclitaxel-induced acute pain. The cellular and molecular signaling pathways revealed in this study could provide rationales for the development of analgesics and management strategies for P-APS in patients.

Activation of toll like receptor 4 attenuates GABA synthesis and postsynaptic GABA receptor activities in the spinal dorsal horn via releasing interleukin-1 beta.

Toll like receptor 4 (TLR4) is an innate immune pattern recognition receptor, expressed predominantly on microglia in the CNS. Activation of spinal TLR4 plays a critical role in the genesis of pathological pain induced by nerve injury, bone cancer, and tissue inflammation. Currently, it remains unknown how synaptic activities in the spinal dorsal horn are regulated by TLR4 receptors. Through recording GABAergic currents in neurons and glial glutamate transporter currents in astrocytes in rodent spinal slices, we determined whether and how TLR4 modulates GABAergic synaptic activities in the superficial spinal dorsal horn. We found that activation of TLR4 by lipopolysaccharide (LPS) reduces GABAergic synaptic activities through both presynaptic and postsynaptic mechanisms. Specifically, LPS causes the release of IL-1ß from microglia. IL-1ß in turn suppresses GABA receptor activities at the postsynaptic site through activating protein kinase C (PKC) in neurons. GABA synthesis at the presynaptic site is reduced upon activation of TLR4. Glial glutamate transporter activities are suppressed by IL-1ß and PKC activation induced by LPS. The suppression of glial glutamate transporter activities leads to a deficiency of glutamine supply, which results in an attenuation of the glutamate-glutamine cycle-dependent GABA synthesis. These findings shed light on understanding synaptic plasticity induced by activation of TLR4 under neuroinflammation and identify GABA receptors, glial glutamate transporters, IL-1ß and PKC as therapeutic targets to abrogate abnormal neuronal activities following activation of TLR4 in pathological pain conditions.

Adenosine Monophosphate-activated Protein Kinase Regulates Interleukin-1ß Expression and Glial Glutamate Transporter Function in Rodents with Neuropathic Pain.

BACKGROUND: Neuroinflammation and dysfunctional glial glutamate transporters (GTs) in the spinal dorsal horn are implicated in the genesis of neuropathic pain. The authors determined whether adenosine monophosphate-activated protein kinase (AMPK) in the spinal dorsal horn regulates these processes in rodents with neuropathic pain. METHODS: Hind paw withdrawal responses to radiant heat and mechanical stimuli were used to assess nociceptive behaviors. Spinal markers related to neuroinflammation and glial GTs were determined by Western blotting. AMPK activities were manipulated pharmacologically and genetically. Regulation of glial GTs was determined by measuring protein expression and activities of glial GTs. RESULTS: AMPK activities were reduced in the spinal dorsal horn of rats (n = 5) with thermal hyperalgesia induced by nerve injury, which were accompanied with the activation of astrocytes, increased production of interleukin-1ß and activities of glycogen synthase kinase 3ß, and suppressed protein expression of glial glutamate transporter-1. Thermal hyperalgesia was reversed by spinal activation of AMPK in neuropathic rats (n = 10) and induced by inhibiting spinal AMPK in naive rats (n = 7 to 8). Spinal AMPKα knockdown (n = 6) and AMPKα1 conditional knockout (n = 6) induced thermal hyperalgesia and mechanical allodynia. These genetic alterations mimicked the changes of molecular markers induced by nerve injury. Pharmacological activation of AMPK enhanced glial GT activity in mice with neuropathic pain (n = 8) and attenuated glial glutamate transporter-1 internalization induced by interleukin-1ß (n = 4). CONCLUSIONS: These findings suggest that enhancing spinal AMPK activities could be an effective approach for the treatment of neuropathic pain.

Endogenous interleukin-1ß in neuropathic rats enhances glutamate release from the primary afferents in the spinal dorsal horn through coupling with presynaptic N-methyl-D-aspartic acid receptors.

Excessive activation of glutamate receptors and overproduction of proinflammatory cytokines, including interleukin-1ß (IL-1ß) in the spinal dorsal horn, are key mechanisms underlying the development and maintenance of neuropathic pain. In this study, we investigated the mechanisms by which endogenous IL-1ß alters glutamatergic synaptic transmission in the spinal dorsal horn in rats with neuropathic pain induced by ligation of the L5 spinal nerve. We demonstrated that endogenous IL-1ß in neuropathic rats enhances glutamate release from the primary afferent terminals and non-NMDA glutamate receptor activities in postsynaptic neurons in the spinal dorsal horn. Myeloid differentiation primary response protein 88 (MyD88) is a mediator used by IL-1ß to enhance non-NMDA glutamate receptor activities in postsynaptic neurons in the spinal dorsal horn. Presynaptic NMDA receptors are effector receptors used by the endogenous IL-1ß to enhance glutamate release from the primary afferents in neuropathic rats. This is further supported by the fact that NMDA currents recorded from small neurons in the dorsal root ganglion of normal rats are potentiated by exogenous IL-1ß. Furthermore, we provided evidence that functional coupling between IL-1ß receptors and presynaptic NMDA receptors at the primary afferent terminals is mediated by the neutral sphingomyelinase/ceramide signaling pathway. Hence, functional coupling between IL-1ß receptors and presynaptic NMDA receptors at the primary afferent terminals is a crucial mechanism leading to enhanced glutamate release and activation of non-NMDA receptors in the spinal dorsal horn neurons in neuropathic pain conditions. Interruption of such functional coupling could be an effective approach for the treatment of neuropathic pain.

Interleukin-1 beta enhances endocytosis of glial glutamate transporters in the spinal dorsal horn through activating protein kinase C.

Excessive activation of glutamate receptors in spinal dorsal horn neurons is a key mechanism leading to abnormal neuronal activation in pathological pain conditions. Previous studies have shown that activation of glutamate receptors in the spinal dorsal horn is enhanced by impaired glial glutamate transporter functions and proinflammatory cytokines including interleukin-1 beta (IL-1ß). In this study, we for the first time revealed that spinal glial glutamate transporter activities in the neuropathic animals are attenuated by endogenous IL-1ß. Specifically, we demonstrated that nerve injury results in an increased expression of IL-1ß and activation of PKC in the spinal dorsal horn as well as suppression of glial glutamate uptake activities. We provided evidence that the nerve-injury induced suppression of glial glutamate uptake is at least in part ascribed to endogenous IL-1ß and activation of PKC in the spinal dorsal horn. IL-1ß reduces glial glutamate transporter activities through enhancing the endocytosis of both GLT-1 and GLAST glial glutamate transporters. The IL-1ß induced trafficking of glial glutamate transporters is through the calcium/PKC signaling pathway, and the dynamin-dependent endocytosis, which is dependent on the integrity of actin filaments. The signaling pathway regulating glial glutamate transporters revealed in this study provides novel targets to attenuate aberrant activation of glutamate receptors in the spinal dorsal horn, which could ultimately help the development of analgesics.

Ranolazine attenuates hypoxia- and hydrogen peroxide-induced increases in sodium channel late openings in ventricular myocytes.

Ranolazine attenuates cardiac arrhythmic activity associated with hypoxia and hydrogen peroxide (H2O2) by inhibition of late sodium current (late INa). The mechanism of ranolazine's action on Na channels was investigated using whole-cell and single-channel recording from guinea pig isolated ventricular myocytes. Hypoxia increased whole-cell late INa from -0.48 ± 0.02 to -3.99 ± 0.07 pA/pF. Ranolazine at 3 and 9 µmol/L reduced the hypoxia-induced late INa by 16% ± 3% and 55% ± 3%, respectively. Hypoxia increased the mean open probability and open time of Na-channel late openings from 0.016 ± 0.001 to 0.064 ± 0.007 milliseconds and from 0.693 ± 0.043 to 1.081 ± 0.098 milliseconds, respectively. Ranolazine at 3 and 9 µmol/L attenuated the hypoxia-induced increase of open probability by 19% ± 7% and 61% ± 1%, and increase of open time by 26% ± 19% and 74 ± 21%, respectively. H2O2 increased the mean open probability and open time of Na-channel late openings from 0.013 ± 0.002 to 0.107 ± 0.015 milliseconds and from 0.689 ± 0.075 to 1.487 ± 0.072 milliseconds, respectively. Ranolazine at 3 and 6 µmol/L reduced the H2O2-induced increase of mean open probability by 60% ± 7% and 95% ± 2%, and the increase of mean open time by 31% ± 21% and 82% ± 8%. In conclusion, the inhibition by ranolazine of hypoxia- and H2O2-stimulated late INa is due to reduction of both the open probability and open time of Na-channel late openings.

Quercetin Promotes the Repair of Mitochondrial Function in H9c2 Cells Through the miR-92a-3p/Mfn1 Axis.

OBJECTIVE: Cardiocerebrovascular disease is a severe threat to human health. Quercetin has a wide range of pharmacological effects such as antitumor and antioxidant. In this study, we aimed to determine how quercetin regulates mitochondrial function in H9c2 cells. METHODS: An H9c2 cell oxygen glucose deprivation/reoxygenation (OGD/R) model was constructed. The expression of miR-92a-3p and mitofusin 1 (Mfn1) mRNA in the cells was detected using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Changes in the mitochondrial membrane potential of cells were examined by JC-1 staining. ATP production in the cells was detected using a biochemical assay. Mitochondrial morphological changes were observed using transmission electron microscopy. Detection of miR-92a-3p binding to Mfn1 was done using dual luciferase. Western blotting was used to detect the protein expression of Mfn1 in the cells. RESULTS: miR-92a-3p is essential in regulating cell viability, apoptosis, and tumor cell metastasis. OGD/R induced miR-92a-3p expression, decreased mitochondrial membrane potential and mitochondrial ATP production, and increased mitochondrial damage. Mitochondria are the most critical site for ATP production. Continued opening of the mitochondrial permeability transition pore results in an abnormal mitochondrial transmembrane potential. Both quercetin and inhibition of miR-29a-3p were able to downregulate miR-29a-3p levels, increase cell viability, mitochondrial membrane potential, and ATP levels, and improve mitochondrial damage morphology. Furthermore, we found that downregulation of miR-29a-3p upregulated the protein expression of Mfn1 in cells. Additionally, miR-92a-3p was found to bind to Mfn1 in a luciferase assay. miR- 29a-3p overexpression significantly inhibited the protein expression level of Mfn1. Quercetin treatment partially reversed the effects of miR-29a-3p overexpression in H9c2 cells. CONCLUSION: Quercetin promoted the recovery of mitochondrial damage in H9c2 cells through the miR-92a-3p/Mfn1 axis.

Endogenous activation of presynaptic NMDA receptors enhances glutamate release from the primary afferents in the spinal dorsal horn in a rat model of neuropathic pain.

Activation of N-methyl-D-aspartate (NMDA) receptors (NMDARs) is a crucial mechanism underlying the development and maintenance of pain. Traditionally, the role of NMDARs in the pathogenesis of pain is ascribed to their activation and signalling cascades in postsynaptic neurons. In this study, we determined if presynaptic NMDARs in the primary afferent central terminals play a role in synaptic plasticity of the spinal first sensory synapse in a rat model of neuropathic pain induced by spinal nerve ligation. Excitatory postsynaptic currents (EPSCs) were recorded from superficial dorsal horn neurons of spinal slices taken from young adult rats. We showed that increased glutamate release from the primary afferents contributed to the enhanced amplitudes of EPSCs evoked by input from the primary afferents in neuropathic rats. Endogenous activation of presynaptic NMDARs increased glutamate release from the primary afferents in neuropathic rats. Presynaptic NMDARs in neuropathic rats were mainly composed of NR2B receptors. The action of presynaptic NMDARs in neuropathic rats was enhanced by exogenous D-serine and/or NMDA and dependent on activation of protein kinase C. In contrast, glutamate release from the primary afferents in sham-operated rats was not regulated by presynaptic NMDARs. We demonstrated that the lack of NMDAR-mediated regulation of glutamate release in sham-operated rats was not attributable to low extracellular levels of the NMDAR agonist and/or coagonist (D-serine), but rather was due to the insufficient function and/or number of presynaptic NMDARs. This was supported by an increase of NR2B receptor protein expression in both the dorsal root ganglion and spinal dorsal horn ipsilateral to the injury site in neuropathic rats. Hence, suppression of the presynaptic NMDAR activity in the primary sensory afferents is an effective approach to attenuate the enhanced glutamatergic response in the spinal first sensory synapse induced by peripheral nerve injury, and presynaptic NMDARs might be a novel target for the development of analgesics.

A rapid analytical method for the quantification of paclitaxel in rat plasma and brain tissue by high-performance liquid chromatography and tandem mass spectrometry.

RATIONALE: Paclitaxel, an antitumor agent for the treatment of several types of cancers, has recently been reported to cause impaired cognitive function and neuropathic pain in humans. To assess the effects of paclitaxel on the central nervous system, a sensitive and accurate method is required to quantify paclitaxel concentrations in plasma and brain tissue obtained from rodents receiving paclitaxel. METHODS: The biological samples were prepared by liquid-liquid extraction and separated by a 3.5 min reversed-phase liquid chromatography (RPLC) method using a BDS Hypersil C8 column under isocratic conditions. Paclitaxel was quantified using multiple reaction monitoring (MRM) with a triple quadrupole tandem mass spectrometer working in the positive electrospray ionization (ESI+) mode. A stable isotope labeled analogue of paclitaxel was used as the internal standard (IS). RESULTS: The method was validated to be precise and accurate within the dynamic range of 0.5-100 ng/mL based on 100 µL plasma and 1.5-300 ng/g based on 33 mg of brain tissue in homogenate. This method was applied to samples from 2 mg/kg intravenously dosed rats. The plasma concentrations were observed to be 26.62 ± 8.93 ng/mL and brain concentrations 11.08 ± 4.18 ng/g when measured 4 h post-dose. CONCLUSIONS: This rapid LC/MS/MS method was validated to be sensitive, specific, precise and accurate for the quantification of paclitaxel in rat plasma and brain tissue homogenate. Application of the method to study samples provided sufficient proof of blood-brain barrier penetration of paclitaxel, allowing further investigation of its influence on the central nervous system.

Activation of Protease-Activated Receptor-1 Causes Chronic Pain in Lupus-Prone Mice Via Suppressing Spinal Glial Glutamate Transporter Function and Enhancing Glutamatergic Synaptic Activity.

Systemic lupus erythematosus (SLE) is an unpredictable autoimmune disease where the body's immune system mistakenly attacks healthy tissues in many parts of the body. Chronic pain is one of the most frequently reported symptoms among SLE patients. We previously reported that MRL lupus prone (MRL/lpr) mice develop hypersensitivity to mechanical and heat stimulation. In the present study, we found that the spinal protease-activated receptor-1(PAR1) plays an important role in the genesis of chronic pain in MRL/lpr mice. Female MRL/lpr mice with chronic pain had activation of astrocytes, over-expression of thrombin and PAR1, enhanced glutamatergic synaptic activity, as well as suppressed activity of adenosine monophosphate-activated protein kinase (AMPK) and glial glutamate transport function in the spinal cord. Intrathecal injection of either the PAR1 antagonist, or AMPK activator attenuated heat hyperalgesia and mechanical allodynia in MRL/lpr mice. Furthermore, we also identified that the enhanced glutamatergic synaptic activity and suppressed activity of glial glutamate transporters in the spinal dorsal horn of MRL/lpr mice are caused by activation of the PAR1 and suppression of AMPK signaling pathways. These findings suggest that targeting the PAR1 and AMPK signaling pathways in the spinal cord may be a useful approach for treating chronic pain caused by SLE. PERSPECTIVE: Our study provides evidence suggesting activation of PAR1 and suppression of AMPK in the spinal cord induces thermal hyperalgesia and mechanical allodynia in a lupus mouse model. Targeting signaling pathways regulating the PAR1 and AMPK could potentially provide a novel approach to the management of chronic pain caused by SLE.

A bibliometric analysis on traumatic brain injury in forensic medicine of a half-century (1972-2021).

Traumatic brain injury (TBI) is among the most common injuries in forensic medicine, the identification of which is of particular importance in forensic practice. To reveal the circumstances and trends of TBI in the forensic field, we used the Web of Science (WoS) database for comprehensive retrieval. We made a metrological analysis of 1,089 papers in the past 50 years (1972-2021). The United States and Germany have the most forensic research on TBI. Diffuse axonal injury (DAI) has been the focus of attention for many years, and much effort has been devoted to its diagnosis in forensic pathology. Infants and children are the subgroups of most concern, especially in infant and child abuse cases. Research on identifying shaken baby syndrome has received increasing attention in recent years. Overall, our study provides a comprehensive list and analysis of the articles regarding TBI in legal medicine, which may shed light on recognizing the trends and research hotspots in this field.

Histone modifications in cocaine, methamphetamine and opioids.

Cocaine, methamphetamine and opioids are leading causes of drug abuse-related deaths worldwide. In recent decades, several studies revealed the connection between and epigenetics. Neural cells acquire epigenetic alterations that drive the onset and progress of the SUD by modifying the histone residues in brain reward circuitry. Histone modifications, especially acetylation and methylation, participate in the regulation of gene expression. These alterations, as well as other host and microenvironment factors, are associated with a serious of negative neurocognitive disfunctions in various patient populations. In this review, we highlight the evidence that substantially increase the field's ability to understand the molecular actions underlying SUD and summarize the potential approaches for SUD pharmacotherapy.