Sirolimus in combination with low-dose extended-release tacrolimus in kidney transplant recipients.

Introduction: Many challenges remain for long-term survival of renal allografts. Once-daily sirolimus (SRL) combined with low-dose extended-release tacrolimus (LER-TAC) may improve medication adherence and reduce the potential nephrotoxicity of calcineurin inhibitors (CNI) compared with standard immunosuppression regimens, thus potentially improving long-term graft survival. Methods: This retrospective, observational, single-center, propensity score matching (PSM) study compared conversion to SRL combined with low-dose ER-TAC and mycophenolic acid (MPA) combined with standard-dose TAC in kidney transplant recipients. After PSM, there were 56 patients in each group. Efficacy, safety, and medication adherence were evaluated over 12 months. Results: There was no significant difference between the two groups in terms of graft and recipient survival and incidence of biopsy-proven acute rejection (p = 1.000), and none of the recipients developed dnDSA after conversion. The mean eGFR improved in SRL + LER-TAC group after conversion compared to before conversion (51.12 ± 20.1 ml/min/1.73 m2 vs. 56.97 ± 19.23 ml/min/1.73 m2, p < 0.05). The medication adherence at 12 months after conversion was superior to before conversion (p = 0.002). Discussion: Our findings suggest that an immunosuppressive regimen of SRL combined with low-dose ER-TAC is no less effective and safe than standard immunosuppressive regimens for renal transplant recipients and may improve graft renal function and medication adherence.

Methyl eugenol protects the kidney from oxidative damage in mice by blocking the Nrf2 nuclear export signal through activation of the AMPK/GSK3ß axis.

Disrupted redox homeostasis contributes to renal ischemia-reperfusion (IR) injury. Abundant natural products can activate nuclear factor erythroid-2-related factor 2 (Nrf2), thereby providing therapeutic benefits. Methyl eugenol (ME), an analog of the phenolic compound eugenol, has the ability to induce Nrf2 activity. In this study, we investigated the protective effects of ME against renal oxidative damage in vivo and in vitro. An IR-induced acute kidney injury (AKI) model was established in mice. ME (20 mg·kg-1·d-1, i.p.) was administered to mice on 5 consecutive days before IR surgery. We showed that ME administration significantly attenuated renal destruction, improved the survival rate, reduced excessive oxidative stress and inhibited mitochondrial lesions in AKI mice. We further demonstrated that ME administration significantly enhanced Nrf2 activity and increased the expression of downstream antioxidative molecules. Similar results were observed in vitro in hypoxia/reoxygenation (HR)-exposed proximal tubule epithelial cells following pretreatment with ME (40 µmol·L-1). In both renal oxidative damage models, ME induced Nrf2 nuclear retention in tubular cells. Using specific inhibitors (CC and DIF-3) and molecular docking, we demonstrated that ME bound to the binding pocket of AMPK with high affinity and activated the AMPK/GSK3ß axis, which in turn blocked the Nrf2 nuclear export signal. In addition, ME alleviated the development of renal fibrosis induced by nonfatal IR, which is frequently encountered in the clinic. In conclusion, we demonstrate that ME modulates the AMPK/GSK3ß axis to regulate the cytoplasmic-nuclear translocation of Nrf2, resulting in Nrf2 nuclear retention and thereby enhancing antioxidant target gene transcription that protects the kidney from oxidative damage.

Phase Ib study of avadomide (CC-122) in combination with rituximab in patients with relapsed/refractory diffuse large B-cell lymphoma and follicular lymphoma.

The multicenter, phase Ib CC-122-DLBCL-001 dose-expansion study (NCT02031419) explored the cereblon E3 ligase modulator (CELMoD) agent avadomide (CC-122) plus rituximab in patients with relapsed/refractory (R/R) diffuse large B-cell lymphoma (DLBCL) or follicular lymphoma (FL). Patients received avadomide 3 mg/day 5 days on/2 days off plus rituximab 375 mg/m2 on day 8 of cycle 1, day 1 of cycles 2 through 6, and day 1 of every third subsequent cycle for 2 years. Primary endpoints were safety and tolerability; preliminary efficacy was a secondary endpoint. A total of 68 patients were enrolled (DLBCL [n = 27], FL [n = 41; 31 lenalidomide-naïve, 10 lenalidomide-treated]). Median age was 62 years (range, 33-84 years), and patients had received a median of 3 (range, 1-8) prior regimens. Among patients with DLBCL, 66.7% had primary refractory disease (partial response or less to initial therapy). Among patients with FL, 65.9% were rituximab-refractory at study entry and 10.0% were lenalidomide-refractory. The most common any-grade avadomide-related adverse events (AEs) were neutropenia (63.2%), infections/infestations (23.5%), fatigue (22.1%), and diarrhea (19.1%). The most common grade 3/4 avadomide-related AEs were neutropenia (55.9%) infections/infestations (8.8%), and febrile neutropenia (7.4%). In patients with DLBCL, overall response rate (ORR) was 40.7% and median duration of response (mDOR) was 8.0 months. In patients with FL, ORR was 80.5% and mDOR was 27.6 months; response rates were similar in lenalidomide-naïve and -treated patients. Avadomide plus rituximab was well tolerated, and preliminary antitumor activity was observed in patients with R/R DLBCL and FL, including subgroups with typically poor outcomes. These results support further investigation of novel CELMoD agents in combination with rituximab in R/R DLBCL and FL.

Mouse strain specificity of DAAO inhibitors-mediated antinociception.

D-Amino acid oxidase (DAAO) specifically catalyzes the oxidative deamination of neutral and polar D-amino acids and finally yields byproducts of hydrogen peroxide. Our previous work demonstrated that the spinal astroglial DAAO/hydrogen peroxide (H2 O2 ) pathway was involved in the process of pain and morphine antinociceptive tolerance. This study aimed to report mouse strain specificity of DAAO inhibitors on antinociception and explore its possible mechanism. DAAO inhibitors benzoic acid, CBIO, and SUN significantly inhibited formalin-induced tonic pain in Balb/c and Swiss mice, but had no antinociceptive effect in C57 mice. In contrast, morphine and gabapentin inhibited formalin-induced tonic pain by the same degrees among Swiss, Balb/c and C57 mice. Therefore, mouse strain difference in antinociceptive effects was DAAO inhibitors specific. In addition, intrathecal injection of D-serine greatly increased spinal H2 O2 levels by 80.0% and 56.9% in Swiss and Balb/c mice respectively, but reduced spinal H2 O2 levels by 29.0% in C57 mice. However, there was no remarkable difference in spinal DAAO activities among Swiss, Balb/c and C57 mice. The spinal expression of glutathione (GSH) and glutathione peroxidase (GPx) activity in C57 mice were significantly higher than Swiss and Balb/c mice. Furthermore, the specific GPx inhibitor D-penicillamine distinctly restored SUN antinociception in C57 mice. Our results reported that DAAO inhibitors produced antinociception in a strain-dependent manner in mice and the strain specificity might be associated with the difference in spinal GSH and GPx activity.

[Effect of methyl eugenol on hypoxia/reoxygenation injury of human renal tubular epithelial cells and its mechanism].

This study aimed to investigate the effect of methyl eugenol(ME) on hypoxia/reoxygenation(H/R)-induced injury of human renal tubular epithelial HK-2 cells and its mechanism. The viability of HK-2 cells cultured with different concentrations of ME and exposed to H/R was detected by cell counting kit-8(CCK-8) assay. The effect of ME on the morphology of HK-2 cells was observed under an inverted microscope. The content of intracellular reactive oxygen species in different groups was detected after 2',7'-dichlorodihydrofluorescein diacetate(DCFH-DA) fluorescence staining. Cell apoptosis was determined by flow cytometry. Changes in mitochondrial membrane potential were monitored by JC-1 dye. The concentrations of nuclear factor erythroid 2 related factor 2(Nrf2), heme oxygenase-1(HO-1), and nicotinamide adenine dinucleotide phosphatase oxidase 4(Nox4) were measured by Western blot, followed by the assay of Nrf2 concentration changes in cytoplasm and nucleus by confocal fluorescence staining. The results showed that when the concentration of ME was 0-40 µmol·L~(-1), the activity of HK-2 cells was not affected. Compared with the model group, ME enhanced the activity of HK-2 cells and the cell morphology was normal. As revealed by further experiments, ME inhibited the release of reactive oxygen species and the decline in mitochondrial membrane potential of HK-2 cells after H/R injury, promoted Nrf2/HO-1 expression and Nrf2 translocation to the nucleus, and down-regulated the expression of Nox4, thereby significantly reducing apoptosis. This protective effect of ME could be reversed by the specific Nrf2 inhibitor ML385. These findings have preliminarily proved that ME effectively protected HK-2 cells against H/R injury, which might be related to its promotion of Nrf2/HO-1 signaling pathway and inhibition of Nox4. Such exploration on the possible mechanism of ME in the treatment of renal ischemia-reperfusion injury(IRI) and protection of organ function from the perspective of antioxidant stress has provided reference for related research on the treatment of acute kidney injury with traditional Chinese medicine.

IOUS and CE-IOUS during hepatic resection for patients with hepatocellular carcinoma in liver cirrhosis1.

PURPOSE: To retrospectively evaluate the role of intraoperative ultrasonography (IOUS) and contrast-enhanced IOUS (CE-IOUS) for the patients with hepatocellular carcinoma (HCC) undergoing hepatic resection (HR). METHODS: Twenty-one consecutive patients who had undergone HR for HCC were included in this study. The patients were subject to preoperative imaging modalities including preoperative ultrasonography (Pre-US) and preoperative contrast-enhanced ultrasonography (Pre-CEUS). All the patients then underwent intraoperative ultrasonography (IOUS) and contrast-enhanced intraoperative ultrasonography (CE-IOUS) during surgery. The visualization of primary HCC and additional lesions of all patients were analyzed. RESULTS: Twenty-one HCCs were detected during Pre-US and the remaining six lesions (28.6%) were detected during IOUS and CE-IOUS. Thus the treatment plan was changed in 28.6% of patients. Twenty-one HCCs (diameter, 0.6-3.0 cm; mean±SD, 1.98±0.85 cm) were measured on Pre-US and remeasured on IOUS (diameter, 0.9-3.3 cm; mean±SD, 2.19±0.84 cm) (p < 0.001). The 6 additional lesions consisted of three moderately differentiated HCCs, one cholangiocarcinoma (ICC), and two high-grade dysplastic nodules (DNs). The mean maximal diameter of the 6 additional lesions was 0.83 cm (range: 0.6-1.1 cm). The malignancy associated features such as capsule interruption, echo heterogeneity, hypo-echoic rim, and a nodule in nodule pattern were more often depicted on IOUS than on Pre-US (all p < 0.01). On CEUS, 19 (90.5%) of 21 HCCs were hyper-enhanced in the arterial phase and washed out from the portal phase to the late phase; the remaining two (9.5%) were hypoenhanced. On CE-IOUS, tumor vasculatures were classified as four patterns: 11 (52.4%) exhibited netlike pattern, 7 (33.3%) annular pattern, 2 (9.5%) mixed pattern, and 1 (4.8%) radial pattern. 3 mHCCs and 2 DNs of six additional nodules had similar greyscale imagining features on IOUS, but they showed different enhancement patterns on CE-IOUS. The ICC showed slightly heterogeneous enhancement during the arterial phase and hypo-enhancement during the portal phase. CONCLUSIONS: IOUS detects more lesions and the treatment plan is changed in 28.6% of patients. HCCs were larger on IOUS than on Pre-US. The typical imaging features of HCCs were better depicted on IOUS in comparison with Pre-US. CE-IOUS can catch the details of microcirculation perfusion of HCCs more sensitively than CEUS. Both IOUS and CE-IOUS were able to provide more decision information during surgery.

Dynamic changes of phenotypes and secretory functions during the differentiation of pre-DCs to mature DCs.

The dynamic expression of cytokines and phenotypes during the differentiation process of dendritic cell precursors (pre-DCs) to mature dendritic cells (DCs) was investigated, and the effects of inflammatory stimulation with lipopolysaccharide (LPS) on DCs differentiation were understood. The differentiation of bone marrow cells isolated from Balb/c mice was induced to DCs in an 8-day cell culture system with RPMI-1640 complete culture medium containing 10% FBS, 20 ng/mL recombinant mouse granulocyte-macrophage colony-stimulating factor (rmGM-CSF) and 10 ng/mL recombinant mouse interleukin-4 (rmIL-4). On the day 3, 6 and 7 after culture, DCs were divided into non-LPS group and LPS group, given 500 ng/mL LPS for 24 h stimulation and no stimulation respectively. The expression levels of CD11c+, MHC-II+, CD40+, CD80+ and CD86+ were detected by flow cytometry, and those of IL-2, IL-4, IL-10, IL-12 p70 and IFN-γ in the supernatant by ELISA. On the day 3 and 6 after culture, the expression of IL-2, IL-4, IL-10 and IFN-γ in DCs showed no significant differences between non-LPS group and LPS group, whereas the differences were significant at day 7. The expression levels of cytokines (for IL-2, IL-4, IL-10, IFN-γ and IL-12 p70: 152.86±6.91, 778.33±8.35, 44.55±2.54, 58.26±1.09 and 2423.00±57.21 pg/mL respectively) in LPS group were higher than those in non-LPS group, especially IL-12 p70 increased obviously at day 7. It was concluded that during the differentiation process of pre-DCs to mature DCs, LPS stimulates DCs producing large amounts of IL-12 p70 and Th1-type cytokines as compared with Th2-type cytokines, and day 7 may be a key time-point for DCs polarization.

Dezocine exhibits antihypersensitivity activities in neuropathy through spinal µ-opioid receptor activation and norepinephrine reuptake inhibition.

Dezocine is the number one opioid painkiller prescribed and sold in China, occupying 44% of the nation's opioid analgesics market today and far ahead of the gold-standard morphine. We discovered the mechanisms underlying dezocine antihypersensitivity activity and assessed their implications to antihypersensitivity tolerance. Dezocine, given subcutaneously in spinal nerve-ligated neuropathic rats, time- and dose-dependently produced mechanical antiallodynia and thermal antihyperalgesia, significantly increased ipsilateral spinal norepinephrine and serotonin levels, and induced less antiallodynic tolerance than morphine. Its mechanical antiallodynia was partially (40% or 60%) and completely (100%) attenuated by spinal µ-opioid receptor (MOR) antagonism or norepinephrine depletion/α2-adrenoceptor antagonism and combined antagonism of MORs and α2-adenoceptors, respectively. In contrast, antagonism of spinal κ-opioid receptors (KORs) and δ-opioid receptors (DORs) or depletion of spinal serotonin did not significantly alter dezocine antiallodynia. In addition, dezocine-delayed antiallodynic tolerance was accelerated by spinal norepinephrine depletion/α2-adenoceptor antagonism. Thus dezocine produces antihypersensitivity activity through spinal MOR activation and norepinephrine reuptake inhibition (NRI), but apparently not through spinal KOR and DOR activation, serotonin reuptake inhibition or other mechanisms. Our findings reclassify dezocine as the first analgesic of the recently proposed MOR-NRI, and reveal its potential as an alternative to as well as concurrent use with morphine in treating pain.

Neuregulin-1/ErbB4 Signaling Regulates Visual Cortical Plasticity.

Experience alters cortical networks through neural plasticity mechanisms. During a developmental critical period, the most dramatic consequence of occluding vision through one eye (monocular deprivation) is a rapid loss of excitatory synaptic inputs to parvalbumin-expressing (PV) inhibitory neurons in visual cortex. Subsequent cortical disinhibition by reduced PV cell activity allows for excitatory ocular dominance plasticity. However, the molecular mechanisms underlying critical period synaptic plasticity are unclear. Here we show that brief monocular deprivation during the critical period downregulates neuregulin-1(NRG1)/ErbB4 signaling in PV neurons, causing retraction of excitatory inputs to PV neurons. Exogenous NRG1 rapidly restores excitatory inputs onto deprived PV cells through downstream PKC-dependent activation and AMPA receptor exocytosis, thus enhancing PV neuronal inhibition to excitatory neurons. NRG1 treatment prevents the loss of deprived eye visual cortical responsiveness in vivo. Our findings reveal molecular, cellular, and circuit mechanisms of NRG1/ErbB4 in regulating the initiation of critical period visual cortical plasticity.

Fluorescence Resonance Energy Transfer-based Biosensor Composed of Nitrogen-doped Carbon Dots and Gold Nanoparticles for the Highly Sensitive Detection of Organophosphorus Pesticides.

The present article reports a novel biosensor for organophosphorus pesticides based on fluorescence resonance energy transfer (FRET) between nitrogen-doped carbon dots (NC-dots) and gold nanoparticles (AuNPs). The effective NC-dots/AuNPs assembly through the Au-N interaction results in good fluorescence quenching. Active acetylcholinesterase (AChE) catalyzes the hydrolysis of acetylthiocholine into -SH containing thiocholine to replace the NC-dots and trigger the aggregation of AuNPs. In the presence of paraoxon, the activity of AChE is inhibited, and thus preventing the generation of thiocholine, causing fewer NC-dots to be replaced. As a consequence, the fluorescence intensity gradually decreases with increasing amount of paraoxon. This biosensor does not require any complex synthesis or modification, and the results show a wide detection range of from 10(-4) to 10(-9) g/L with a detection limit of 1.0 × 10(-9) g/L (3.6 × 10(-12) mol/L). Two linear response regions have been reported with a turning point at about 10(-6) g/L and three different factors that would influence the response behavior. These phenomena discussed in detail so as to explain the special response mechanism.

Peptidic exenatide and herbal catalpol mediate neuroprotection via the hippocampal GLP-1 receptor/ß-endorphin pathway.

Both peptidic agonist exenatide and herbal agonist catalpol of the glucagon-like peptide-1 receptor (GLP-1R) are neuroprotective. We have previously shown that activation of spinal GLP-1Rs expresses ß-endorphin in microglia to produce antinociception. The aim of this study was to explore whether exenatide and catalpol exert neuroprotection via activation of the hippocampal GLP-1R/ß-endorphin pathway. The rat middle cerebral artery occlusion model was employed, and the GLP-1R immunofluorescence staining and ß-endorphin measurement were assayed in the hippocampus and primary cultures of microglia, neurons and astrocytes. The immunoreactivity of GLP-1Rs on microglia in the hippocampus was upregulated after ischemia reperfusion. Intracerebroventricular (i.c.v.) injection of exenatide and catalpol produced neuroprotection in the rat transient ischemia/reperfusion model, reflected by a marked reduction in brain infarction size and a mild recovery in neurobehavioral deficits. In addition, i.c.v. injection of exenatide and catalpol significantly stimulated ß-endorphin expression in the hippocampus and cultured primary microglia (but not primary neurons or astrocytes). Furthermore, exenatide and catalpol neuroprotection was completely blocked by i.c.v. injection of the GLP-1R orthosteric antagonist exendin (9-39), specific ß-endorphin antiserum, and selective opioid receptor antagonist naloxone. Our results indicate, for the first time, that the neuroprotective effects of catalpol and exenatide are GLP-1R-specific, and that these effects are mediated by ß-endorphin expression probably in hippocampal microglia. We postulate that in contrast to the peripheral tissue, where the activation of GLP-1Rs in pancreas islet ß-cells causes secretion of insulin to perform glucoregulation, it leads to ß-endorphin expression in microglial cells to produce neuroprotection and analgesia in the central nervous system.

Contributions of spinal D-amino acid oxidase to chronic morphine-induced hyperalgesia.

Spinal D-amino acid oxidase (DAAO) is an FAD-dependent peroxisomal flavoenzyme which mediates the conversion of neutral and polar D-amino acids (including D-serine) to the corresponding α-keto acids, and simultaneously produces hydrogen peroxide and ammonia. This study has aimed to explore the potential contributions of spinal DAAO and its mediated hydrogen peroxide/D-serine metabolism to the development of morphine-induced hyperalgesia. Bi-daily subcutaneous injections of morphine to mice over 7 days induced thermal hyperalgesia as measured by both the hot-plate and tail-immersion tests, and spinal astroglial activation with increased spinal gene expression of DAAO, glial fibrillary acidic protein (GFAP) and pro-inflammatory cytokines (interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α)). Subcutaneous injections of the potent DAAO inhibitor CBIO (5-chloro-benzo[D]isoxazol-3-ol) prevented and reversed the chronic morphine-induced hyperalgesia. CBIO also inhibited both astrocyte activation and the expression of pro-inflammatory cytokines. Intrathecal injection of the hydrogen peroxide scavenger PBN (phenyl-N-tert-butylnitrone) and of catalase completely reversed established morphine hyperalgesia, whereas subcutaneous injections of exogenous D-serine failed to alter chronic morphine-induced hyperalgesia. These results provided evidence that spinal DAAO and its subsequent production of hydrogen peroxide rather than the D-serine metabolism contributed to the development of morphine-induced hyperalgesia.

Comparison of the effectiveness and safety of ultrasound- and CT-guided percutaneous radiofrequency ablation of non-operation hepatocellular carcinoma.

To retrospectively compare the effectiveness and safety of ultrasound (US)- and computer tomography (CT)-guided percutaneous radiofrequency ablation (PRFA) in treating patients with non-operation hepatocellular carcinoma (HCC). Forty patients with non-operation HCC who were treated with US-guided PRFA (20 patients with 24 HCC lesions) or CT-guided PRFA (20 patients with 27 HCC lesions) were enrolled in this study. Follow-up was performed with US and CT/MRI. Complete ablation rate, local recurrence rate, and overall survival rate were used to evaluate the efficacy of the two therapeutic choices. The PRFA-related complications including hilar bile duct injury, sepsis, liver failure, renal dysfunction, peritoneal hemorrhage, and skin burn were assessed. The operation time of CT-guided group was significantly longer than that of the US-guided group (P < 0.05). The single ablation times for tumors with similar size showed no significant difference between the two groups (P > 0.05). The differences in complete ablation rate (79.2 vs. 88.9 %, P > 0.05) and local recurrence rate (16.7 vs. 14.8 %, P > 0.05) between US- and CT-guided groups were not statistically significant. In the US-guided group, the 1-, 2-, and 3-year overall survival rates were 85, 74, and 68 %, respectively, while they were 84, 72, and 58 % in the CT-guided group. The differences were not statistically significant (P > 0.05). No severe complications were found in the two groups. Both US- and CT-guided PRFA are safe and effective therapies for patients with HCC when surgical options are precluded.

The non-peptide GLP-1 receptor agonist WB4-24 blocks inflammatory nociception by stimulating ß-endorphin release from spinal microglia.

BACKGROUND AND PURPOSE: Two peptide agonists of the glucagon-like peptide-1 (GLP-1) receptor, exenatide and GLP-1 itself, exert anti-hypersensitive effects in neuropathic, cancer and diabetic pain. In this study, we have assessed the anti-allodynic and anti-hyperalgesic effects of the non-peptide agonist WB4-24 in inflammatory nociception and the possible involvement of microglial ß-endorphin and pro-inflammatory cytokines. EXPERIMENTAL APPROACH: We used rat models of inflammatory nociception induced by formalin, carrageenan or complete Freund's adjuvant (CFA), to test mechanical allodynia and thermal hyperalgesia. Expression of ß-endorphin and pro-inflammatory cytokines was measured using real-time quantitative PCR and fluorescent immunoassays. KEY RESULTS: WB4-24 displaced the specific binding of exendin (9-39) in microglia. Single intrathecal injection of WB4-24 (0.3, 1, 3, 10, 30 and 100 µg) exerted dose-dependent, specific, anti-hypersensitive effects in acute and chronic inflammatory nociception induced by formalin, carrageenan and CFA, with a maximal inhibition of 60-80%. Spinal WB4-24 was not effective in altering nociceptive pain. Subcutaneous injection of WB4-24 was also antinociceptive in CFA-treated rats. WB4-24 evoked ß-endorphin release but did not inhibit expression of pro-inflammatory cytokines in either the spinal cord of CFA-treated rats or cultured microglia stimulated by LPS. WB4-24 anti-allodynia was prevented by a microglial inhibitor, ß-endorphin antiserum and a µ-opioid receptor antagonist. CONCLUSIONS AND IMPLICATIONS: Our results suggest that WB4-24 inhibits inflammatory nociception by releasing analgesic ß-endorphin rather than inhibiting the expression of proalgesic pro-inflammatory cytokines in spinal microglia, and that the spinal GLP-1 receptor is a potential target molecule for the treatment of pain hypersensitivity including inflammatory nociception.

Lamiophlomis rotata, an orally available Tibetan herbal painkiller, specifically reduces pain hypersensitivity states through the activation of spinal glucagon-like peptide-1 receptors.

BACKGROUND: Lamiophlomis rotata is an orally available Tibetan herb prescribed for the management of pain, with shanzhiside methylester (SM) and 8-O-acetyl-SM as quality control ingredients. This study aimed to evaluate the antinociceptive properties of L. rotata, determine whether SM and 8-O-acetyl-SM are principle effective ingredients, and explore whether L. rotata produces antinociception through activation of spinal glucagon-like peptide-1 receptors (GLP-1Rs). METHODS: Formalin test, neuropathic pain, and bone cancer pain models were used, and the animal sample size was 5 to 6 in each group. Hydrogen peroxide-induced oxidative damage was also assayed. RESULTS: The L. rotata aqueous extract blocked formalin-induced tonic hyperalgesia and peripheral nerve injury- and bone cancer-induced mechanical allodynia by 50 to 80%, with half-effective doses of 130 to 250 mg/kg, close to the human dosage. The herb was not effective in alleviating acute nociceptive pain. A 7-day gavage with L. rotata aqueous extract did not lead to antiallodynic tolerance. Total iridoid glycosides, rather than total flavonoids, were identified by the activity-tracking method as effective ingredients for antihyperalgesia, whereas both SM and 8-O-acetyl-SM were principal components. Further demonstrations using the GLP-1R antagonist and gene silencer against GLP-1R at both the spinal and the cellular levels indicated that L. rotata inhibited pain hyperactivity by activation of spinal GLP-1Rs, and SM and 8-O-acetyl-SM appeared to be orthosteric, reversible, and fully intrinsic agonists of both rat and human GLP-1Rs. CONCLUSIONS: Results support the notion that the activation of spinal GLP-1Rs leads to specific antinociception in pain hypersensitivity and further suggest that GLP-1R is a human-validated target molecule for the treatment of chronic pain.

Activation of spinal glucagon-like peptide-1 receptors specifically suppresses pain hypersensitivity.

This study aims to identify the inhibitory role of the spinal glucagon like peptide-1 receptor (GLP-1R) signaling in pain hypersensitivity and its mechanism of action in rats and mice. First, GLP-1Rs were identified to be specifically expressed on microglial cells in the spinal dorsal horn, and profoundly upregulated after peripheral nerve injury. In addition, intrathecal GLP-1R agonists GLP-1(7-36) and exenatide potently alleviated formalin-, peripheral nerve injury-, bone cancer-, and diabetes-induced hypersensitivity states by 60-90%, without affecting acute nociceptive responses. The antihypersensitive effects of exenatide and GLP-1 were completely prevented by GLP-1R antagonism and GLP-1R gene knockdown. Furthermore, exenatide evoked ß-endorphin release from both the spinal cord and cultured microglia. Exenatide antiallodynia was completely prevented by the microglial inhibitor minocycline, ß-endorphin antiserum, and opioid receptor antagonist naloxone. Our results illustrate a novel spinal dorsal horn microglial GLP-1R/ß-endorphin inhibitory pathway in a variety of pain hypersensitivity states.

Geniposide and its iridoid analogs exhibit antinociception by acting at the spinal GLP-1 receptors.

We recently discovered that the activation of the spinal glucagon-like peptide-1 receptors (GLP-1Rs) by the peptidic agonist exenatide produced antinociception in chronic pain. We suggested that the spinal GLP-1Rs are a potential target molecule for the management of chronic pain. This study evaluated the antinociceptive activities of geniposide, a presumed small molecule GLP-1R agonist. Geniposide produced concentration-dependent, complete protection against hydrogen peroxide-induced oxidative damage in PC12 and HEK293 cells expressing rat and human GLP-1Rs, but not in HEK293T cells that do not express GLP-1Rs. The orthosteric GLP-1R antagonist exendin(9-39) right-shifted the concentration-response curve of geniposide without changing the maximal protection, with identical pA2 values in both cell lines. Subcutaneous and oral geniposide dose-dependently blocked the formalin-induced tonic response but not the acute flinching response. Subcutaneous and oral geniposide had maximum inhibition of 72% and 68%, and ED50s of 13.1 and 52.7 mg/kg, respectively. Seven days of multidaily subcutaneous geniposide and exenatide injections did not induce antinociceptive tolerance. Intrathecal geniposide induced dose-dependent antinociception, which was completely prevented by spinal exendin(9-39), siRNA/GLP-1R and cyclic AMP/PKA pathway inhibitors. The geniposide iridoid analogs geniposidic acid, genipin methyl ether, 1,10-anhydrogenipin, loganin and catalpol effectively inhibited hydrogen peroxide-induced oxidative damage and formalin pain in an exendin(9-39)-reversible manner. Our results suggest that geniposide and its iridoid analogs produce antinociception during persistent pain by activating the spinal GLP-1Rs and that the iridoids represented by geniposide are orthosteric agonists of GLP-1Rs that function similarly in humans and rats and presumably act at the same binding site as exendin(9-39).

Gelsemine, a principal alkaloid from Gelsemium sempervirens Ait., exhibits potent and specific antinociception in chronic pain by acting at spinal α3 glycine receptors.

The present study examined the antinociceptive effects of gelsemine, the principal alkaloid in Gelsemium sempervirens Ait. A single intrathecal injection of gelsemine produced potent and specific antinociception in formalin-induced tonic pain, bone cancer-induced mechanical allodynia, and spinal nerve ligation-induced painful neuropathy. The antinociception was dose-dependent, with maximal inhibition of 50% to 60% and ED50 values of 0.5 to 0.6 µg. Multiple daily intrathecal injections of gelsemine for 7 days induced no tolerance to antinociception in the rat model of bone cancer pain. Spinal gelsemine was not effective in altering contralateral paw withdrawal thresholds, and had only a slight inhibitory effect on formalin-induced acute nociception. The specific antinociception of gelsemine in chronic pain was blocked dose-dependently by the glycine receptor (GlyR) antagonist strychnine with an apparent ID50 value of 3.8 µg. Gelsemine concentration-dependently displaced H(3)-strychnine binding to the membrane fraction of rat spinal cord homogenates, with a 100% displacement and a Ki of 21.9µM. Gene ablation of the GlyR α3 subunit (α3 GlyR) but not α1 GlyR, by a 7-day intrathecal injection of small interfering RNA (siRNA) targeting α3 GlyR or α1 GlyR, nearly completely prevented gelsemine-induced antinociception in neuropathic pain. Our results demonstrate that gelsemine produces potent and specific antinociception in chronic pain states without induction of apparent tolerance. The results also suggest that gelsemine produces antinociception by activation of spinal α3 glycine receptors, and support the notion that spinal α3 glycine receptors are a potential therapeutic target molecule for the management of chronic pain.

Epstein-Barr virus negative primary hepatic leiomyoma: case report and literature review.

Primary hepatic leiomyoma is a neoplasm of mesenchymal origin and occurs only rarely. Secondary to benign smooth muscle proliferation, it is usually found in adult women and is associated with Epstein-Barr virus (EBV) infection. Here, we report the 29(th) case of primary hepatic leiomyoma with its unique features related to diagnosis, treatment and developmental biology. A 48-year-old man, with an immunocompromised status, complained of pain in the upper quadrant of the abdomen. Serological analysis indicated no presence of hepatitis virus, no human immunodeficiency virus, and no EBV infection. The levels of α-fetoprotein and carcinoembryonic antigen were normal. A mass was detected in segment III of the hepatic lobe by ultrasonography and an abdominal computed tomography scan. Endoscopy had negative findings. Exploratory laparotomy found no existing extrahepatic tumor and left lateral lobectomy was performed. Pathological examination showed the mass to be a typical leiomyoma. The cells were positive for α-smooth muscle actin and desmin, and negative for the makers of gastrointestinal stromal tumor (GIST), including CD117, CD34 and DOG1 (discovered on GIST1). In situ hybridization revealed negative status for EBV-encoded small RNA. After left lateral lobectomy, the patient was not given chemotherapy or radiotherapy. During a 2-year follow-up, no sign of local recurrence or distant metastasis was observed. In conclusion, we report a rare case of primary hepatic leiomyoma in a male patient without EBV infection. Hepatic resection was curative. This case presents data to expand our knowledge concerning the complex and heterogeneous nature of primary liver leiomyoma, indicating that EBV infection is important but neither necessary nor sufficient for the development of primary liver leiomyoma.