Ceftriaxone Relieves Trigeminal Neuropathic Pain Through Suppression of Spatiotemporal Synaptic Plasticity via Restoration of Glutamate Transporter 1 in the Medullary Dorsal Horn.

Using a rat model of trigeminal neuropathic pain (TNP) produced by chronic compression of the infraorbital nerve (CCI-ION), we investigated the analgesic effect and the underlying mechanisms of ceftriaxone (Cef), a ß-lactam antibiotic, that is thought to be a potent stimulator of glutamate transporter 1 (GLT-1). First, repeated intraperitoneal (i.p.) injections of Cef (200 mg/kg) for 5-days since Day 1 of CCI-ION could significantly relieve both mechanical and thermal pain hypersensitivity from day 10 after drug administration. Western blot and immunofluorescent results demonstrated that 5-days administration of Cef resulted in the restoration of GLT-1 expression to a level equivalent to the sham control which was dramatically lost under the TNP condition. Moreover, multi-electrode (8 × 8) array recordings of network field excitatory postsynaptic potentials (fEPSPs) were performed on the acutely dissociated medullary dorsal horn slice evoked by electrical stimulation of the trigeminal spinal tract. The results showed that the increased number of fEPSPs, induction rate, and maintenance of long-term potentiation caused by CCI-ION were significantly suppressed by 5-days administration of Cef. Taken together, the results indicate that Cef can relieve TNP through suppression of spatiotemporal synaptic plasticity via GLT-1 restoration in the medullary dorsal horn of the trigeminal nerve.

Complete structure and variation of the chloroplast genome of Agropyron cristatum (L.) Gaertn.

Agropyron cristatum (L.) Gaertner, a perennial grass in the tribe Triticeae (Poaceae), is a wild relative of cereal crops that is suitable for genetic improvement. In this study, we first sequenced the complete chloroplast (cp) genome of Ag. cristatum using Hiseq4000 PE150. The Ag. cristatum chloroplast genome is 135,554bp in length, has a typical quadripartite structure and contains 76 protein-coding genes, 29 tRNA genes and four rRNA genes. The cp genome of Ag. cristatum was used for comparison with other seven Triticeae species. One large variable region (800bp), which primarily contained the rpl23 (non-reciprocally translocated from IRs) and accD genes, was detected between rbcL gene and psaI gene within LSC region. The deletion of the accD and translocated rpl23 genes in Ag. cristatum indicated an independent gene-loss events or an additional divergence in Triticeae. Analyses of the dn/ds ratio and K2-P's genetic distance for 76 protein-coding genes showed that genes with evolutionary divergence might suffer from the effect of sequence regional constraints or gene functional constraints in Triticeae species. Our research will generally contribute to the knowledge of plastid genome evolution in Triticeae.

Correlations between plasma endothelin-1 levels and breakthrough pain in patients with cancer.

Endothelin-1 (ET-1) may be involved in driving pain in patients with advanced cancer. However, a few studies focus on the role of ET-1 in breakthrough pain (BP). The aim of this pivotal study was to explore the correlation between the plasma (ET-1) level and BP intensity. A total of 40 patients were enrolled in the study, and they were divided into two groups: BP group and non-BP group. Moreover, 20 healthy adults were used as the normal control group. Pain intensity was measured using visual analog scale (VAS) scores of 1-10. Plasma ET-1 levels were detected by an ET radioimmunoassay kit. Subsequently, the correlation of ET-1 level with the VAS score and cancer types was analyzed by Pearson's correlation coefficient. The plasma ET-1 level in the BP group (35.31±8.02 pg/mL) was higher than that in the non-BP group (29.51±6.78 pg/mL) and the normal control group (24.77±10.10 pg/mL, P<0.05). In addition, the VAS score in the BP group (7.45±0.82) was higher than that in the non-BP group (2.80±1.23, P<0.05). The plasma ET-1 level was positively correlated with the VAS score of the BP group (Pearson's r=0.42). There was no significant correlation between the plasma ET-1 level and VAS score of the non-BP group (Pearson's r=-0.22) or/and cancer types (P>0.05). The elevated plasma ET-1 levels were positively related to BP, and targeting ET-1 may provide a novel pain-reducing therapeutic treatment in BP.

Gene silencing of NR2B-containing NMDA receptor by intrathecal injection of short hairpin RNA reduces formalin-induced nociception in C57BL/6 mouse.

Spinal NR2B-containing N-methyl-D-aspartate receptors (NR2B) play a critical role in the formation of central sensitization and persistent pain. Previous studies show that gene silencing of the spinal NR2B subunit by small interfering RNA (siRNA) could alleviate nociception in animals. The siRNA is a 19- to 23-nt RNA duplex, which can be synthesized in vitro or derived from short hairpin RNAs (shRNAs). In the present study, we investigated whether intrathecal injection of shRNAs targeting NR2B (GRIN2B shRNA) could affect nociception on formalin-induced pain in mice. Our results showed that intrathecal injection of GRIN2B shRNA could decrease NR2B mRNA and protein expression levels and hence effectively relieve formalin-induced nociception in mice, suggesting that intrathecal delivery of GRIN2B shRNA can be an efficient way to silence the target gene and provide new insights into the treatment of chronic pain.

Surgery upregulates high mobility group box-1 and disrupts the blood-brain barrier causing cognitive dysfunction in aged rats.

AIM: Postoperative cognitive dysfunction (POCD) is a growing and largely underestimated problem without defined etiology. Herein, we sought to determine the relationship between cognitive decline, blood-brain barrier (BBB) permeability, and inflammation, namely high mobility group box-1 (HMGB1), after surgery in aged rats. METHODS: Aged rats were randomly assigned as surgery group (n = 45, splenectomy under general anesthesia), anesthesia (n = 45, 2% isoflurane for 2 h), and naïve control (n = 15). Markers of inflammation were measured in plasma and brain. Blood-brain barrier ultrastructure and permeability were measured by transmission electron microscope (TEM) and IgG immunohistochemistry. Cognitive function was assessed in a reversal learning version of the Morris water maze (MWM). RESULTS: Surgical trauma under general anesthesia caused distinct changes in systemic and central proinflammatory cytokines. Levels of HMGB1 and the receptor for advanced glycation end products (RAGE) were significantly upregulated in the hippocampus of operated animals. Immunohistochemistry and TEM showed BBB disruption induced by surgery and anesthesia. These molecular changes were associated with cognitive impairment in latency with the MWM up to postoperative day 3. CONCLUSIONS: HMGB1 and RAGE signaling appear pivotal mediators of surgery-induced cognitive decline and may contribute to the changes in BBB permeability after peripheral surgical trauma.

[Changes of prostaglandin D2,carboxypeptidase A3 and platelet activating factor in guinea pig in anaphylactic shock].

OBJECTIVE: To detect the changes of leukotriene E4(LTE4), prostaglandin D2(PGD2), carboxypeptidase A3(CPA3) and platelet activating factor (PAF) in guinea pigs died from anaphylactic shock. METHODS: Guinea pigs were used for establishing anaphylactic shock models. The levels of LTE4, PGD2 and CPA3, and PAF were detected in urine, plasma, and brain tissues with ELISA kit, respectively. The significant biomarkers were selected comparing with control group. The changes of PGD2, CPA3 and PAF in the guinea pigs at time zero, 12 and 24 hours after death were observed and compared respectively. The effect of platelet activating factor acetylhydrolase (PAF-AH) to PAF in guinea pig brain was examined and compared. RESULTS: There were no statistically differences of LTE4 levels in urine observed between experimental group and control group. The levels of CPA3, PGD2 and PAF in the experimental group were significantly higher than that in the control group at 0 h. The levels of PAF at 12 and 24 hours after anaphylactic shock were significantly higher than that in the control group. The levels of PAF decreased significantly after pretreatment with PAF-AH. CONCLUSION: LTE4 in urine cannot be selected as a biomarker to determine the anaphylactic shock. PGD2 and CPA3 in plasma, and PAF in brain tissue may be used as biomarkers to determine the anaphylactic shock. PAF-AH may be potentially useful for clinical treatment of anaphylactic shock.

Effects of linear-polarized near-infrared light irradiation on chronic pain.

In order to study the efficacy of linear-polarized near-infrared light irradiation (LPNIR) on relieving chronic pain in conjunction with nerve block (NB) or local block (LB), a 3-week prospective, randomized, double-blind, controlled study was conducted to evaluate the pre- and post-therapy pain intensity. Visual analogue scales (VASs) were measured in all patients before and 6 months after therapy visiting the pain clinic during the period of August 2007 to January 2008. A total of 52 patients with either shoulder periarthritis or myofascial pain syndrome or lateral epicondylitis were randomly assigned into two groups by drawing lots. Patients in Group I were treated with NB or LB plus LPNIR; Group II patients, for their part, were treated with the same procedures as in Group I, but not using LPNIR. In both groups, the pain intensity (VAS score) decreased significantly immediately after therapy as compared to therapy. There was a significant difference between the test and control groups immediately after therapy (P < 0.05), while no effect 6 months later. No side effects were observed. It is concluded that LPNIR is an effective and safe modality to treat various chronic pains, which has synergic effects with NB or LB.

Involvement of neuronal nitric oxide synthase in cognitive impairment in isoflurane-treated rats.

The underlying causes of post-operative cognitive dysfunction (POCD) in elderly patients remain to be elucidated. In order to explore possible contributory mechanisms, we tested the effects of isoflurane anesthesia on (i) expression of hippocampal neuronal nitric oxide synthase (nNOS) and (ii) the relationship of changes in nNOS expression to cognitive dysfunction in isoflurane-treated aged rats. Our results indicate that isoflurane treatment leads to significant changes in correct reactions (F=28.35, p<0.001), initiative avoidances (F=29.33, p<0.001), and total reaction time (TRT) (F=6.99, p<0.05) of treated rats in the Y-maze test. Isoflurane-treated rats had fewer correct reactions and initiative avoidances in the Y-maze test 24 and 48 h after 2 h of isoflurane anesthesia compared with control group rats (p<0.05). TRTs to complete 20 trials of the Y-maze test increased significantly 48 h after 2 h anesthesia. The number of nNOS-positive hippocampal neurons decreased 24 h after anesthesia, corresponding to an increased mean immunostaining grey-scale value. These data show that isoflurane causes a transient decrease in expression of hippocampal nNOS in aged rats during early post-anesthesia stages, and that the transient decrease of nNOS is closely correlated with cognitive impairment in isoflurane-treated aged rats.

[Establishment and evaluation of a bone cancer pain model].

OBJECTIVE: To explore the feasibility of a bone cancer pain model by injecting the Lewis lung carcinoma cells into the femur bone marrow cavity of C57BL/6 mice. METHODS: Sixty clear grade male C57BL/6 mice (body weight 18 approximately 20 g) were randomly divided into 4 groups(15 in each group). Cancer cell inoculated group: 2*10(6) Lewis lung carcinoma cells in 10 microL PBS were injected into the left femur bone marrow cavity, and the other 3 control groups were injected the heat inactivated Lewis cells, PBS, or a false operation respectively. Spontaneous lifting time and mechanical allodynia threshold of the mice hind paw were measured in the alternative days throughout the experiment. The structural damage of the femur was monitored by radiogram on the 7th,15th, and 23rd day respectively,and the pathohistological changes of the femur bones were observed by HE staining on the same days. RESULTS: Those mice that received intra-femur innoculation of Lewis lung carcinoma cells gradually developed the spontaneous pain, which was began on the 11th day after the innoculation, and followed by mechanical allodynia. The course of flinch lasted in the later experimental session. The 50% Von Frey threshold was significantly decreased on the 13th day after the innoculation, and the mechanical allodynia lasted the whole experimental period. On the 23rd day after the innoculation, X-ray film showed that the medullary cavity of ipsilateral distal femur was filled with tumor cells, and the cortical bone became thick; furthermore, the tumor cells invaded the peripheral muscles. CONCLUSION: Injecting the Lewis lung carcinoma cells into the femoral medullary cavity of C57BL/6 mice can successfully establish a murine bone cancer pain model, and the murine model shows much resemblance compared with the human bone cancer pain.

Down-regulation of beta1-adrenoceptors gene expression by short interfering RNA impairs the memory retrieval in the basolateral amygdala of rats.

The influence of basolateral amygdala (BLA) on memory is known to depend critically on adrenergic neurotransmission. However, the roles of noradrenergic receptors on memory retrieval have been elusive and controversial. Here, we investigated the effect of beta(1)-adrenoceptor (beta(1)-AR) on auditory fear memory in the rat BLA. We attenuated the expression of beta(1)-AR by RNA interference, a popular means to specific suppress gene expression. Bilaterally microinjection of beta(1)-AR short interfering RNA (siRNA) could reach a satisfying transfection in the BLA: beta(1)-AR protein expression was reduced transiently by siRNA in vivo at day 3. The behavioral tests indicated that memory retrieval was impaired as beta(1)-AR protein expression was prevented, and the memory was restored when the beta(1)-AR protein got back to normal level. The results suggested that beta(1)-AR might be critical for the retrieval of auditory fear memory.

[Correlation between dynamic contrast-enhanced MRI and expression of microvascular density in the glioma].

OBJECTIVE: To explore the correlation between microvascular density (MVD) and dynamic contrast-enhanced MRI in the glioma. METHODS: We examined 35 patients with histologically verified glioma. Gadolinium-enhanced dynamic TurboFLASH imaging was performed preoperatively in all patients followed by conventional MRI. The steepest slope (SSmax) of curve and corresponding Tm1 in "first-pass" phase were obtained by analyzing time-signal curve. All specimens were immunostained with anti-human Factor VIII relative antigen monoclonal antibody postoperatively by streptavidin-peroxidase method to obtain the MVD. The correlation between SSmax, Tm1, and MVD was analyzed. RESULTS: SSmax was positively correlated with MVD (r = 0.640, P < 0.01). Tml was negatively correlated with MVD (r = -0.671, P < 0.01). CONCLUSION: The MVD correlates obviously with SSmax and Tml in the glioma. Analyzing the time-signal curve of dynamic contrast-enhanced MRI is helpful to predict the angiogenesis in the glioma.

Syntheses of acetylated steroid glycosides and selective cleavage of O-acetyl groups in sugar moiety.

Acetylated 3beta-O-beta-glycosyl steroid derivatives were synthesized by the reaction of a new polyhydroxysteroid 3beta,5alpha,6beta-trihydroxypregn-16-en-20-one (2) with the peracetylated 1-bromo derivatives of d-glucose and d-galactose, respectively. Subsequent protection by excess acetic anhydride in pyridine selectively gave the 6beta-O-acetylated steroid glycosides. Deprotection of the acetylated steroid glycosides separately with moderate catalysis dibutyltin oxide in methanol selectively removed all acetyl groups of sugar moiety, whereas the acetyl group of the steroid part was retained. The structures of the steroid glycosides were confirmed by mass spectrometry, NMR and IR. The complete protocol was shown to be non-destructive at all stages to the sugar moieties and the steroid nucleus. These regioselective reactions open a route to the synthesis of a series of closely related isomers of 2 and other widespread polyhydroxysteroids and steroid glycosides in marine organisms and some terrestrial species.