Association between low incidence of TP53 mutations and reduced early relapse rates in Uygur DLBCL.

Diffuse large B-cell lymphoma (DLBCL) demonstrates significant heterogeneity, investigations into the distinctions in clinical and molecular characteristics between Chinese Uygur and Han DLBCL patients remain unexplored. We retrospectively reviewed 279 DLBCL patients (105 Uygur and 174 Han patients), of which 155 patients underwent genetic profiling by NGS. Compared with Han patient, Uygur patients have better clinical prognostic indicators, including a higher proportion of patients with 0-1 extranodal involvement and I/II Ann Arbor staging. Consistently, Uygur patients were significantly associated with lower risk of relapse (P = 0.06), with a one-year relapse rate of 5% vs 17% and two-year relapse rate of 19% vs 36% compared to Han patients. At the molecular level, TP53 (21.3%) was among the top frequently altered gene in the cohort. Notably, the Uygur patients exhibited a significantly lower frequency of TP53 alterations and higher frequency of ASXL3 alterations. Logistic regression analysis showed that the lowered frequency of TP53 and enrichment of ASXL3 in the Uygur patients were independent of other factors. However, only patients with TP53 mutations had higher relapse rate than those with wild type TP53 (one-year, 20% vs 10%; two-year, 51% vs 21%). Our findings highlight the notable contribution of a low TP53 mutation frequency in Uygur patients as a pivotal factor associated with the favorable prognosis of this population.

Whole-brain mapping of monosynaptic afferent inputs to the CRH neurons in the medial prefrontal cortex of mice.

Corticotropin-releasing hormone (CRH) neurons are densely distributed in the medial prefrontal cortex (mPFC), which plays a crucial role in integrating and processing emotional and cognitive inputs from other brain regions. Therefore, it is important to know the neural afferent patterns of mPFCCRH neurons, which are still unclear. Here, we utilized a rabies virus-based monosynaptic retrograde tracing system to map the presynaptic afferents of the mPFCCRH neurons throughout the entire brain. The results show that the mPFCCRH neurons receive inputs from three main groups of brain regions: (1) the cortex, primarily the orbital cortex, somatomotor areas, and anterior cingulate cortex; (2) the thalamus, primarily the anteromedial nucleus, mediodorsal thalamic nucleus, and central medial thalamic nucleus; and (3) other brain regions, primarily the basolateral amygdala, hippocampus, and dorsal raphe nucleus. Taken together, our results are valuable for further investigations into the roles of the mPFCCRH neurons in normal and neurological disease states. These investigations can shed light on various aspects such as cognitive processing, emotional modulation, motivation, sociability, and pain.

Hippocampus Insulin Receptors Regulate Episodic and Spatial Memory Through Excitatory/Inhibitory Balance.

It is well known that the hippocampus is a vital brain region playing a key role in both episodic and spatial memory. Insulin receptors (InsRs) are densely distributed in the hippocampus and are important for its function. However, the effects of InsRs on the function of the specific hippocampal cell types remain elusive. In this study, hippocampal InsRs knockout mice had impaired episodic and spatial memory. GABAergic neurons and glutamatergic neurons in the hippocampus are involved in the balance between excitatory and inhibitory (E/I) states and participate in the processes of episodic and spatial memory. InsRs are located mainly at excitatory neurons in the hippocampus, whereas 8.5% of InsRs are glutamic acid decarboxylase 2 (GAD2)::Ai9-positive (GABAergic) neurons. Next, we constructed a transgenic mouse system in which InsR expression was deleted from GABAergic (glutamate decarboxylase 2::InsRfl/fl, GAD2Cre::InsRfl/fl) or glutamatergic neurons (vesicular glutamate transporter 2::InsRfl/fl,Vglut2Cre::InsRfl/fl). Our results showed that in comparison to the InsRfl/fl mice, both episodic and spatial memory were lower in GAD2Cre::InsRfl/fl and Vglut2Cre::InsRfl/fl. In addition, both GAD2Cre::InsRfl/fl and Vglut2Cre::InsRfl/fl were associated with more anxiety and lower glucose tolerance. These findings reveal that hippocampal InsRs might be crucial for episodic and spatial memory through E/I balance hippocampal regulation.

Identification of FAT4 as a positive prognostic biomarker in DLBCL by comprehensive genomic analysis.

The molecular landscapes of diffuse large B-cell lymphoma (DLBCL) remained to be comprehensively investigated with an urgent need to identify novel prognostic biomarkers guiding prognostic stratification and disease monitoring. Baseline tumor samples of 148 DLBCL patients were analyzed using targeted next-generation sequencing (NGS) for mutational profiling, whose clinical reports were retrospectively reviewed. In this cohort, the subgroup of old DLBCL patients (age at diagnosis > 60, N = 80) exhibited significantly higher Eastern Cooperative Oncology Group scores and International Prognostic Index than their young counterparts (age at diagnosis ≤ 60, N = 68). As revealed by the NGS results, PIM1 (43.9%), KMT2D (31.8%), MYD88 (29.7%), and CD79B (27.0%) were identified as the most frequently mutated genes. Aberrations of genes of the immune escape pathway were significantly enriched in the young subgroup, while the altered epigenetic regulators were more abundant in the old patients. FAT4 mutation was identified as a positive prognostic biomarker, associated with longer progression-free survival and overall survival in the entire cohort and the old subgroup, using the Cox regression analyses. However, the prognostic function of FAT4 was not reproduced in the young subgroup. We comprehensively analyzed the pathological and molecular characteristics of old and young DLBCL patients and demonstrated the prognostic value of FAT4 mutation, which requires further validation with sizable cohorts in future research.

Projections from the Rostral Zona Incerta to the Thalamic Paraventricular Nucleus Mediate Nociceptive Neurotransmission in Mice.

Zona incerta (ZI) is an integrative subthalamic region in nociceptive neurotransmission. Previous studies demonstrated that the rostral ZI (ZIR) is an important gamma-aminobutyric acid-ergic (GABAergic) source to the thalamic paraventricular nucleus (PVT), but whether the ZIR-PVT pathway participates in nociceptive modulation is still unclear. Therefore, our investigation utilized anatomical tracing, fiber photometry, chemogenetic, optogenetic and local pharmacological approaches to investigate the roles of the ZIRGABA+-PVT pathway in nociceptive neurotransmission in mice. We found that projections from the GABAergic neurons in ZIR to PVT were involved in nociceptive neurotransmission. Furthermore, chemogenetic and optogenetic activation of the ZIRGABA+-PVT pathway alleviates pain, whereas inhibiting the activities of the ZIRGABA+-PVT circuit induces mechanical hypersensitivity and partial heat hyperalgesia. Importantly, in vivo pharmacology combined with optogenetics revealed that the GABA-A receptor (GABAAR) is crucial for GABAergic inhibition from ZIR to PVT. Our data suggest that the ZIRGABA+-PVT pathway acts through GABAAR-expressing glutamatergic neurons in PVT mediates nociceptive neurotransmission.

Morphological investigations of endomorphin-2 and spinoparabrachial projection neurons in the spinal dorsal horn of the rat.

It has been proved that endomorphin-2 (EM2) produced obvious analgesic effects in the spinal dorsal horn (SDH), which existed in our human bodies with remarkable affinity and selectivity for the µ-opioid receptor (MOR). Our previous study has demonstrated that EM2 made synapses with the spinoparabrachial projection neurons (PNs) in the SDH and inhibited their activities by reducing presynaptic glutamate release. However, the morphological features of EM2 and the spinoparabrachial PNs in the SDH have not been completely investigated. Here, we examined the morphological features of EM2 and the spinoparabrachial PNs by using triple fluorescence and electron microscopic immunohistochemistry. EM2-immunoreactive (-ir) afferents directly contacted with the spinoparabrachial PNs in lamina I of the SDH. Immunoelectron microscopy (IEM) were used to confirm that these contacts were synaptic connections. It was also observed that EM2-ir axon terminals contacting with spinoparabrachial PNs in lamina I contained MOR, substance P (SP) and vesicular glutamate transporter 2 (VGLUT2). In lamina II, MOR-ir neurons were observed to receive direct contacts from EM2-ir varicosities. The synaptic connections among EM2, MOR, SP, VGLUT2, and the spinoparabrachial PNs were also confirmed by IEM. In sum, our results supply morphological evidences for the analgesic effects of EM2 on the spinoparabrachial PNs in the SDH.

Glial cells and neurologic autoimmune disorders.

Neurologic autoimmune disorders affect people's physical and mental health seriously. Glial cells, as an important part of the nervous system, play a vital role in the occurrence of neurologic autoimmune disorders. Glial cells can be hyperactivated in the presence of autoantibodies or pathological changes, to influence neurologic autoimmune disorders. This review is mainly focused on the roles of glial cells in neurologic autoimmune disorders and the influence of autoantibodies produced by autoimmune disorders on glial cells. We speculate that the possibility of glial cells might be a novel way for the investigation and therapy of neurologic autoimmune disorders.

CARD11 is a novel target of miR-181b that is upregulated in chronic lymphocytic leukemia.

Aim: To investigate the targets of miR-181b in patients with chronic lymphocytic leukemia (CLL). Materials & methods: The bioinformatic softwares were used to indicate the key target genes associated with miR-181b, and the results were verified in CLL patient samples and 293T cells. Results:CARD11 is a potential target gene of miR-181b, an inverse relationship was revealed between the expression of CARD11 and miR-181b in 104 CLL patients, and it was confirmed in vitro with luciferase assays and western blotting. Kaplan-Meier analysis showed that CLL patients with high CARD11 expression demonstrated poor survival. Conclusion:CARD11 is a novel target of miR-181b that is upregulated, which could be a poor prognostic indicator for CLL patients.

Analysis of 4 imaging features in patients with COVID-19.

BACKGROUND: The aim of this was to analyze 4 chest CT imaging features of patients with coronavirus disease 2019 (COVID-19) in Shenzhen, China so as to improve the diagnosis of COVID-19. METHODS: Chest CT of 34 patients with COVID-19 confirmed by the nucleic acid test (NAT) were retrospectively analyzed. Analyses were performed to investigate the pathological basis of four imaging features("feather sign","dandelion sign","pomegranate sign", and "rime sign") and to summarize the follow-up results. RESULTS: There were 22 patients (65.2%) with typical "feather sign"and 18 (52.9%) with "dandelion sign", while few patients had "pomegranate sign" and "rime sign". The "feather sign" and "dandelion sign" were composed of stripe or round ground-glass opacity (GGO), thickened blood vessels, and small-thickened interlobular septa. The "pomegranate sign" was characterized as follows: the increased range of GGO, the significant thickening of the interlobular septum, complicated with a small amount of punctate alveolar hemorrhage. The "rime sign" was characterized by numerous alveolar edemas. Microscopically, the wall thickening, small vascular proliferation, luminal stenosis, and occlusion, accompanied by interstitial infiltration of inflammatory cells, as well as numerous pulmonary interstitial fibrosis and partial hyaline degeneration were observed. Repeated chest CT revealed the mediastinal lymphadenectasis in one patient. Re-examination of the NAT showed another positive anal swab in two patients. CONCLUSION: "Feather sign" and "dandelion sign" were typical chest CT features in patients withCOVID-19; "pomegranate sign" was an atypical feature, and "rime sign" was a severe feature. In clinical work, accurate identification of various chest CT signs can help to improve the diagnostic accuracy of COVID-19 and reduce the misdiagnosis or missed diagnosis rate.

[Expression Level and Target Gene Prediction of miR-181b in Patients with Chronic Lymphocytic Leukemia].

OBJECTIVE: To investigate the expression level of miR-181b in CD19+ B lymphocytes of patients with chronic lymphocytic leukemia (CLL), to analyze the relationship between its expression and the prognosis of CLL patients, and to predict the potential target gene of miR-181b in CLL by using bioinformatics. METHODS: Eight-four patients with CLL treated in People's Hospital of Xinjiang Uygur Autonomous Region from June 2013 to June 2018 were selected. and 20 healthy people were selected as control group. RNA was extracted from CD19+B lymphocytes of peripheral blood by magnetic bead sorting, the expression level of miR-181b was detected, and it's expression differences in different IPI groups were analyzed. The correlation between the expression level of miR-181b and PFS of CLL patients also was analyzed. miR-181b target genes were predicted by online database and literatures, and gene annotation analysis and relevant signal pathway analysis were performed for candidate target genes. RESULTS: The expression level of miR-181b in CLL patients was significantly lower than that in control group (P＜0.01); The expression level of miR-181b in the low-risk group was higher than that in high-risk group and extremely high-risk group (P＜0.05), but there was no statistical difference between low-risk group and medium-risk group (P=1.00). The expression level of miR-181b in medium-risk group was higher than that in high-risk group and extremely high-risk group (P＜0.05), but there was no difference between high-risk group and extremely high-risk group (P=1.00). ROC curve results showed that the area under the curve (AUC) was 0.792 (P＜0.01).When the expression level of miR-181b was at the threshold value of 0.279, it showed a better sensitivity (62.9%) and specificity (91.8%). Survival analysis results suggested that compared with the high expression group, the miR-181b low expression group had poor PFS (log rank: P=0.047). Prediction of miR-181b by using the starBase, targetscan and picTar database and its combination with literature reports indicated that CARD11, ZFP36L1, RUNX1, NR4A3, ATP1B1, PUM1 and PLAG1 related with blood diseases, and up-regulated CARD11 and ZFP36L1 participated in lymphoid tumor formation by promoting cell proliferation and inhibiting cell aging. CONCLUSION: The expression level of miR-181b in CLL group are significantly lower than that in the controls group, and the low expression of miR-181b relates with poor prognosis of CLL patients. Through bioinformatics prediction and combined with literature reports, it is speculated that CARD11 and ZFP36L1 as target genes of miR-181b may be participated in the occurrence and development of CLL. Further experiments are needed to verify this result.

Enkephalinergic Circuit Involved in Nociceptive Modulation in the Spinal Dorsal Horn.

Enkephalin (ENK) has been implicated in pain modulation within the spinal dorsal horn (SDH). Revealing the mechanisms underlying ENK analgesia entails the anatomical and functional knowledge of spinal ENK-ergic circuits. Herein, we combined morphological and electrophysiological studies to unravel local ENK-ergic circuitry within the SDH. First, the distribution pattern of spinal ENK-ergic neurons was observed in adult preproenkephalin (PPE)-GFP knock-in mice. Next, the retrograde tracer tetramethylrhodamine (TMR) or horseradish peroxidase (HRP) was injected into the parabrachial nucleus (PBN) in PPE-GFP mice. Immunofluorescent staining showed I-isolectin B4 (IB4) labeled non-peptidergic afferents were in close apposition to TMR-labeled PBN-projecting neurons within lamina I as well as PPE-immunoreactivity (-ir) neurons within lamina II. Some TMR-labeled neurons were simultaneously in close association with both IB4 and PPE-ir terminals. Synaptic connections of these components were further confirmed by electron microscopy. Finally, TMR was injected into the PBN in adult C57BL/6 mice. Whole-cell patch recordings showed that δ-opioid receptor (DOR) agonist, [D-Pen2,5]-enkephalin (DPDPE, 1 µM), significantly reduced the frequency of miniature excitatory postsynaptic current (mEPSC) and decreased the activity of TMR-labeled neurons. In conclusion, spinal ENKergic neurons receive direct excitatory inputs from primary afferents, which might be directly recruited to release ENK under the condition of noxious stimuli; ENK could inhibit the glutamatergic transmission towards projecting neurons via presynaptic and postsynaptic DORs. These morphological and functional evidence may explain the mechanisms underlying the analgesic effects exerted by ENK within the SDH.

miR-34a and miR-29b as indicators for prognosis of treatment-free survival of chronic lymphocytic leukemia patients in Chinese Uygur and Han populations.

The abnormal expression of miRNAs may play critical roles in the occurrence, development and prognosis of chronic lymphocytic leukemia (CLL), with potential ethnic differences being involved. p53 and immunoglobulin heavy chain variable region gene (IGVH) mutations were monitored and miRNA profile screening of CD19 + cells from Uygur CLL patients was performed, analyzed by miRNA arrays and verified using real-time PCR. There were 68 differentially expressed miRNAs in CD19 + B lymphocytes obtained from 6 Uygur CLL patients, of which miR-1295, miR-29b, miR-34a, miR-21 and miR-29c were the 5 most upregulated, and miR-181a, miR-126, miR-181b, miR-125a-5p and miR199b the 5 most downregulated miRNAs. miR-15a/miR-16-1 which might be important drivers of the disease, were not eliminated by profile screening. From the 68 differentially expressed miRNAs, 5 previously-reported CLL-related miRNAs were selected for further confirmation analyses, from which expression levels of miR-29b, miR-34a and miR-155 were found to be increased while miR-181a and miR-181b decreased. However, there were no differences in the expression levels of miR-15a/miR-16-1 between CLL patients and healthy donors, but the expression levels of miR-15a/miR-16-1 in CLL patients with a 13q deletion was depressed. In addition, there was no difference in the expression level of the above 7 miRNAs between 44 Han and 40 Uygur CLL patients. The expression levels of miR-29b, miR-181a and miR-181b correlated with IGVH mutations, while the expression levels of miR-34a, miR-29b and miR-181b correlated with a p53 abnormality in 84 Uygur and Han CLL patients. Taking p53 abnormality as the cut-off value criteria, low expression levels of miR-34a (cut-off value 4.65, P = 0.02) and miR-29b (cut-off value 4.71, P = 0.009) hinted at a poor treatment-free survival (TFS) prognosis for all CLL patients. Thus miR-34a and miR-29b may represent useful indicators for the prognosis of both Uygur and Han CLL patients.

Endomorphin-2 Inhibits the Activity of the Spinoparabrachial Projection Neuron through Presynaptic Mechanisms in the Spinal Dorsal Horn in Rats.

BACKGROUND/AIMS: Spinal dorsal horn (SDH) is one of the most important regions for analgesia produced by endomorphin-2 (EM2), which has a higher affinity and specificity for the µ-opioid receptor (MOR) than morphine. Many studies have focused on substantia gelatinosa (SG, lamina II) neurons to elucidate the cellular basis for its antinociceptive effects. However, the complicated types and local circuits of interneurons in the SG make it difficult to understand the real effects of EM2. Therefore, in the present study, we examined the effects of EM2 on projection neurons (PNs) in lamina I. METHODS: Tracing, immunofluoresence, and immunoelectron methods were used to examine the morphological connections between EM2-immunoreactive (-ir) terminals and PNs. By using in vitro whole cell patch clamp recording technique, we investigated the functional effects of EM2 on PNs. RESULTS: EM2-ir afferent terminals directly contacted PNs projecting to the parabrachial nucleus in lamina I. Their synaptic connections were further confirmed by immunoelectron microscopy, most of which were asymmetric synapses. It was found that EM2 had a strong inhibitory effect on the frequency, but not amplitude, of the spontaneous excitatory postsynaptic current (sEPSC) of the spinoparabrachial PNs in lamina I, which could be reversed by MOR antagonist CTOP. However, their spontaneous inhibitory postsynaptic current (sIPSC) and intrinsic properties were not changed after EM2 application. CONCLUSION: Applying EM2 to the SDH could produce analgesia through inhibiting the activities of the spinoparabrachial PNs in lamina I by reducing presynaptic neurotransmitters release from the primary afferent terminals.

Endomorphins: Promising Endogenous Opioid Peptides for the Development of Novel Analgesics.

Endomorphin-1 (EM1) and endomorphin-2 (EM2) are two endogenous ligands that belong to the opioid peptide family and have the highest affinity and selectivity for the µ-opioid receptor (MOR). The neuroanatomical distribution, ultrastructural features and neural circuitry of EM-containing neuronal structures have been morphologically demonstrated. In addition, the modulation effects of the EMs in different areas reflect their potential endogenous roles in many major physiological processes, including their remarkable roles in the transmission and modulation of noxious information. The distinguished antinociceptive property of the EMs in acute and chronic pain, including neuropathic pain, cancer pain and inflammatory pain, has been revealed and investigated for therapeutic purposes. However, EMs exert adverse effects in the gastrointestinal, urinary, cardiovascular, and respiratory systems, which impede the development of EMs as new analgesics. Numerous studies have synthesized and investigated EM analogues and demonstrated that these EM derivatives had improved pharmacological properties, supporting their therapeutic perspectives. In the present review, the results of previous studies, particularly morphological and pharmacological studies, were summarized. Finally, EM modifications and their potential clinical implications were described. Applying this knowledge about EMs may provide information for further investigations in clinical application.

Decreased Endomorphin-2 and µ-Opioid Receptor in the Spinal Cord Are Associated with Painful Diabetic Neuropathy.

Painful diabetic neuropathy (PDN) is one of the most common complications in the early stage of diabetes mellitus (DM). Endomorphin-2 (EM2) selectively activates the µ-opioid receptor (MOR) and subsequently induces antinociceptive effects in the spinal dorsal horn. However, the effects of EM2-MOR in PDN have not yet been clarified in the spinal dorsal horn. Therefore, we aimed to explore the role of EM2-MOR in the pathogenesis of PDN. The main findings were the following: (1) streptozotocin (STZ)-induced diabetic rats exhibited hyperglycemia, body weight loss and mechanical allodynia; (2) in the spinal dorsal horn, the expression levels of EM2 and MOR decreased in diabetic rats; (3) EM2 protein concentrations decreased in the brain, lumbar spinal cord and cerebrospinal fluid (CSF) in diabetic rats but were unchanged in the plasma; (4) the frequency but not the amplitude of spontaneous excitatory postsynaptic currents (sEPSCs) was significantly higher in diabetic rats than in control rats; and (5) intrathecal injection of EM2 for 14 days in the early stage of PDN partially alleviated mechanical allodynia and reduced MOR expression in diabetic rats. Our results demonstrate that the EM2-MOR signal may be involved in the early stage of PDN.

Endomorphin-2 Inhibition of Substance P Signaling within Lamina I of the Spinal Cord Is Impaired in Diabetic Neuropathic Pain Rats.

Opiate analgesia in the spinal cord is impaired in diabetic neuropathic pain (DNP), but until now the reason is unknown. We hypothesized that it resulted from a decreased inhibition of substance P (SP) signaling within the dorsal horn of the spinal cord. To investigate this possibility, we evaluated the effects of endomorphin-2 (EM2), an endogenous ligand of the µ-opioid receptor (MOR), on SP release within lamina I of the spinal dorsal horn (SDH) in rats with DNP. We established the DNP rat model and compared the analgesic efficacy of EM2 between inflammation pain and DNP rat models. Behavioral results suggested that the analgesic efficacy of EM2 was compromised in the condition of painful diabetic neuropathy. Then, we measured presynaptic SP release induced by different stimulating modalities via neurokinin-1 receptor (NK1R) internalization. Although there was no significant change in basal and evoked SP release between control and DNP rats, EM2 failed to inhibit SP release by noxious mechanical and thermal stimuli in DNP but not in control and inflammation pain model. We also observed that EM2 decreased the number of FOS-positive neurons within lamina I of the SDH but did not change the amount of FOS/NK1R double-labeled neurons. Finally, we identified a remarkable decrease in MORs within the primary afferent fibers and dorsal root ganglion (DRG) neurons by Western blot (WB) and immunohistochemistry (IHC). Taken together, these data suggest that reduced presynaptic MOR expression might account for the loss of the inhibitory effect of EM2 on SP signaling, which might be one of the neurobiological foundations for decreased opioid efficacy in the treatment of DNP.

Neurochemical features of endomorphin-2-containing neurons in the submucosal plexus of the rat colon.

AIM: To investigate the distribution and neurochemical phenotype of endomorphin-2 (EM-2)-containing neurons in the submucosal plexus of the rat colon. METHODS: The mid-colons between the right and left flexures were removed from rats, and transferred into Kreb's solution. For whole-mount preparations, the mucosal, outer longitudinal muscle and inner circular muscle layers of the tissues were separated from the submucosal layer attached to the submucosal plexus. The whole-mount preparations from each rat mid-colon were mounted onto seven gelatin-coated glass slides, and processed for immunofluorescence histochemical double-staining of EM-2 with calcitonin gene-related peptide (CGRP), choline acetyltransferase (ChAT), nitric oxide synthetase (NOS), neuron-specific enolase (NSE), substance P (SP) and vasoactive intestinal peptide (VIP). After staining, all the fluorescence-labeled sections were observed with a confocal laser scanning microscope. To estimate the extent of the co-localization of EM-2 with CGRP, ChAT, NOS, NSE, SP and VIP, ganglia, which have a clear boundary and neuronal cell outline, were randomly selected from each specimen for this analysis. RESULTS: In the submucosal plexus of the mid-colon, many EM-2-immunoreactive (IR) and NSE-IR neuronal cell bodies were found in the submucosal plexus of the rat mid-colon. Approximately 6 ± 4.2 EM-2-IR neurons aggregated within each ganglion and a few EM-2-IR neurons were also found outside the ganglia. The EM-2-IR neurons were also immunopositive for ChAT, SP, VIP or NOS. EM-2-IR nerve fibers coursed near ChAT-IR neurons, and some of these fibers were even distributed around ChAT-IR neuronal cell bodies. Some EM-2-IR neuronal cell bodies were surrounded by SP-IR nerve fibers, but many long processes connecting adjacent ganglia were negative for EM-2 immunostaining. Long VIP-IR processes with many branches coursed through the ganglia and surrounded the EM-2-IR neurons. The percentages of the EM-2-IR neurons that were also positive for ChAT, SP, VIP or NOS were approximately 91% ± 2.6%, 36% ± 2.4%, 44% ± 2.5% and 44% ± 4.7%, respectively, but EM-2 did not co-localize with CGRP. CONCLUSION: EM-2-IR neurons are present in the submucosal plexus of the rat colon and express distinct neurochemical markers.

[Discrimination of Descurainiae Semen and Pantagirus Semen by HPLC fingerprints].

OBJECTIVE: To discriminate Descurainiae Semen and Pantagirus Semen. METHOD: A high-performance liquid chromatographic method was developed to establish the fingerprint of Descurainiae Semen, and hierarchical cluster analysis (HCA) and partial least squares discriminant analysis (PLS-DA) were applied to study HPLC fingerprinting and chemical recognition. RESULT: There exists large difference of chromatographic peaks and its relative peak area of HPLC fingerprints between Descurainiae Semen and Pantagirus Semen, and after conducting statistical analysis, the result demonstrated that all samples were classified into three categories: Descurainiae Semen, Pantagirus Semen and their mixtures. CONCLUSION: The developed HPLC fingerprint combined with chemometrics can be utilized to discriminate between Descurainiae Semen and Pantagirus Semen, which was quick, simple, accurate and reliable an can provide the basis for the characterization and quality assess of Descurainiae Semen and Pantagirus Semen.

Puerarin alleviates noise-induced hearing loss via affecting PKCγ and GABAB receptor expression.

Noise-induced hearing loss (NIHL) often results from prolonged exposure to high levels of noise. Our previous study revealed that during the development of NIHL, the expression of protein kinase C γ subunit (PKCγ) and GABAB receptor (GABABR) was changed within the cochlear nuclear complex (CNC), suggesting that these molecules might be the potential targets for the treatment of NIHL. As an extending study, here we focused on puerarin, a major isoflavonoid extracted from Pueraria lobota, which has been used in the treatment of cardiovascular and cerebrovascular diseases, and investigated whether it could protect against NIHL by acting on PKCγ and GABABR. Transgenic GAD67-GFP knock-in mice were subjected to the NIHL model and their auditory functions were evaluated by the auditory brainstem response thresholds and distortion product oto-acoustic emission signals. Our results showed that 200mg/kg puerarin treatment ameliorated the thresholds of auditory brainstem response of NIHL mice significantly. Triple immunofluorescence staining and electron microscopy results revealed that GFP-positive neurons in the superficial layers of CNC expressed both PKCγ and GABABR1, and GAD67-positive terminals contacted PKCγ- or GABABR1-positive neurons. Immunoblotting and RT-PCR results showed that NIHL increased the expression of PKCγ but decreased that of GABABR1 and GABABR2 at both protein and mRNA levels in the CNC. Puerarin significantly attenuated the increased expression of PKCγ but elevated the reduced expression of GABABR1 and GABABR2 after noise exposure. Thus, we provided the first evidence that puerarin ameliorated the auditory functions of NIHL mice, and this effect may be due to its ability to regulate the expression of PKCγ and GABABR.

Neural tissue engineering scaffold with sustained RAPA release relieves neuropathic pain in rats.

AIMS: To investigate the effect of locally slow-released rapamycin (RAPA) from bionic peripheral nerve stent to reduce the incidence of neuropathic pain or mitigate the degree of pain after nerve injury. MAIN METHODS: We constructed a neural tissue engineering scaffold with sustained release of RAPA to repair 20mm defects in rat sciatic nerves. Four presurgical and postsurgical time windows were selected to monitor the changes in the expression of pain-related dorsal root ganglion (DRG) voltage-gated sodium channels 1.3 (Nav1.3), 1.7 (Nav1.7), and 1.8 (Nav1.8) through immunohistochemistry (IHC) and Western Blot, along with the observation of postsurgical pathological pain in rats by pain-related behavior approaches. KEY FINDINGS: Relatively small upregulation of DRG sodium channels was observed in the experimental group (RAPA+poly(lactic-co-glycolic acid) (PLGA)+stent) after surgery, along with low degrees of neuropathic pain and anxiety, which were similar to those in the Autologous nerve graft group. SIGNIFICANCE: Autoimmune inflammatory response plays a leading role in the occurrence of post-traumatic neuropathic pain, and that RAPA significantly inhibits the abnormal upregulation of sodium channels to reduce pain by alleviating inflammatory response.