Construction of a disulfidptosis-related glycolysis gene risk model to predict the prognosis and immune infiltration analysis of gastric adenocarcinoma.

BACKGROUND: The pattern of cell death known as disulfidptosis was recently discovered. Disulfidptosis, which may affect the growth of tumor cells, represents a potential new approach to treating tumors. Glycolysis affects tumor proliferation, invasion, chemotherapy resistance, the tumor microenvironment (TME), and immune evasion. However, the efficacy and therapeutic significance of disulfidptosis-related glycolysis genes (DRGGs) in stomach adenocarcinoma (STAD) remain uncertain. METHODS: STAD clinical data and RNA sequencing data were downloaded from the TCGA database. DRGGs were screened using Cox regression and Lasso regression analysis to construct a prognostic risk model. The accuracy of the model was verified using survival studies, receiver operating characteristic (ROC) curves, column plots, and calibration curves. Additionally, our study investigated the relationships between the risk scores and immune cell infiltration, tumor mutational burden (TMB), and anticancer drug sensitivity. RESULTS: We have successfully developed a prognosis risk model with 4 DRGGs (NT5E, ALG1, ANKZF1, and VCAN). The model showed excellent performance in predicting the overall survival of STAD patients. The DRGGs prognostic model significantly correlated with the TME, immune infiltrating cells, and treatment sensitivity. CONCLUSIONS: The risk model developed in this work has significant clinical value in predicting the impact of immunotherapy in STAD patients and assisting in the choice of chemotherapeutic medicines. It can correctly estimate the prognosis of STAD patients.

Nutritional Status and Incidence of Radiation-Induced Oral Mucositis in Nasopharyngeal Carcinoma Patients Treated with Chemoradiotherapy.

Malnutrition is prevalent among patients with nasopharyngeal carcinoma undergoing radiotherapy. This study examined the nutritional status and incidence of radiation-induced oral mucositis (RIOM) in patients with nasopharyngeal carcinoma. A retrospective analysis was conducted to compare the incidence of RIOM, Nutritional Risk Screening (NRS) 2002 score, weight, body mass index (BMI), and hemoglobin levels in 338 patients treated with induction chemotherapy (IC) plus concurrent chemoradiotherapy (CCRT) or treated with CCRT alone. The IC + CCRT group exhibited an increase in weight and BMI but a decrease in hemoglobin levels after IC compared with baseline (p < 0.001). Both groups showed differences in weight at Week 0 and BMI at Weeks 0-2 of radiotherapy (p < 0.05). The IC + CCRT group experienced an increase in NRS 2002 scores from Week 2 to Week 6 (p < 0.05). The hemoglobin levels of the IC + CCRT group were consistently lower throughout radiotherapy (p < 0.001). However, no significant difference was observed in the incidence of RIOM between the two groups (p = 0.246). Patients treated with IC + CCRT exhibited a higher nutritional risk during radiotherapy. Although the incidence of Grade III RIOM was high, no significant difference was found between the groups.

Anatomic prognostic factors and their potential roles in refining M1 classification for de novo metastatic nasopharyngeal carcinoma.

BACKGROUND AND PURPOSE: To identify anatomic prognostic factors and their potential roles in refining M1 classification for de novo metastatic nasopharyngeal carcinoma (M1-NPC). MATERIALS AND METHODS: All M1-NPC treated with chemotherapy and/or radiotherapy between 2010 and 2019 from two centers (training and validation cohort) were included. The prognostic value of metastatic disease extent and involved organs for overall survival (OS) were assessed by several multivariable analyses (MVA) models. A new M1 classification was proposed and validated in a separate cohort who received immuno-chemotherapy. RESULTS: A total of 197 M1-NPC in the training and 307 in the validation cohorts were included for M1 subdivision study with median follow-up of 46 and 57 months. MVA model with "≤2 organs/≤5 lesions" as the definition of oligometastasis had the highest C-index (0.623) versus others (0.606-0.621). Patients with oligometastasis had better OS versus polymetastasis (hazard ratio [HR] 0.47/0.63) while liver metastases carried worse OS (HR 1.57/1.45) in MVA in the training/validation cohorts, respectively. We proposed to divide M1-NPC into M1a (oligometastasis without liver metastases) and M1b (liver metastases or polymetastasis) with 3-year OS of 66.5%/31.7% and 64.9%/35.0% in the training/validation cohorts, respectively. M1a subset had a better median progress-free survival (not reach vs. 17 months, p < 0.001) in the immuno-chemotherapy cohort (n = 163). CONCLUSION: Oligometastasis (≤2 organs/≤5 lesions) and liver metastasis are prognostic for M1-NPC. Subdivision of M1-NPC into M1a (oligometastasis without liver metastasis) and M1b (liver metastasis or polymetastasis) depicts the prognosis well in M1-NPC patients who received immuno-chemotherapy.

Impaired geriatric 8 score is associated with worse survival after radical intensity modulated radiation therapy in elderly patients with nasopharyngeal carcinoma.

BACKGROUND: Geriatric 8 score (G8) was an independent prognostic factor for survival and toxicities in various malignancies, but it has never been tested in nasopharyngeal carcinoma (NPC). OBJECTIVES: To evaluate the value of G8 in predicting survival in elderly patients with NPC. MATERIAL AND METHODS: Patients with NPC aged ≥70 who received intensity-modulated radiation therapy were recruited into this study. The overall survival (OS), progression-free survival (PFS), locoregional recurrence rate (LRR), and distant metastasis rate (DMR) between the patients with G8 > 14 and G8 ≤ 14 were calculated using the Kaplan-Meier method and compared with the Log-rank test. Cox proportional hazards model was applied to perform univariate and multivariate analysis. RESULTS: G8 ≤ 14 had significantly reduced OS (p = .001) and PFS (p = .032) than those with G8 > 14 by log-rank test. G8 score remained an independent prognosticator for OS (HR = 0.490, 95% CI = 0.267-0.900, p = .021) and was a borderline significance towards PFS (HR = 0.639, 95% CI = 0.386-1.058, p = .082) in multivariate analysis. Grade 3-4 acute toxicities were significantly more common in patients with G8 ≤ 14 than in those with G8 > 14. CONCLUSIONS AND SIGNIFICANCE: G8 is useful in predicting the OS in elderly patients with NPC. Further prospective study stratified by G8 is needed to explore the value of CT in elderly patients with NPC.

Serum and urine metabolomics analyses reveal metabolic pathways and biomarkers in relation to nasopharyngeal carcinoma.

RATIONALE: Nasopharyngeal carcinoma (NPC) is a malignant tumor that is endemic in Southeast Asia, North Africa, and southern China. There is an urgent need for effective early diagnosis and treatment of this disease since NPC is currently often detected at advanced stages. METHODS: To reveal the underlying metabolic mechanisms and discover potential diagnostic biomarkers of NPC, we employed ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) and UHPLC-Q-Exactive Orbitrap MS, respectively, to analyze 54 serum samples and 54 urine samples from 27 patients with NPC and 27 healthy control individuals. RESULTS: A total of 1230 metabolites were determined in serum samples, and 181 of the 1230 metabolites were significantly changed in NPC patients. The 181 metabolites were enriched in 16 pathways, including biosynthesis of unsaturated fatty acids, cholesterol metabolism, and ferroptosis. A total of 2509 metabolites were detected in the urine samples. Among them, 179 metabolites were significantly altered in NPC patients, and these metabolites were enriched in eight pathways, including the tricarboxylic acid (TCA) cycle and caffeine metabolism. Seven metabolites, including creatinine and paraxanthine, were found to be significantly changed in both NPC serum and urine samples. Based on them, further biomarker analysis revealed that the panel of three serum metabolites, octanoylcarnitine, creatinine, and decanoyl-l-carnitine, displayed a perfect diagnostic performance (area under the curve [AUC] = 0.973) to distinguish NPC patients from controls, while the other three-metabolite biomarker panel, consisting of stachydrine, decanoyl-l-carnitine, and paraxanthine, had an AUC = 0.809 to distinguish NPC and control in urine samples. CONCLUSION: This work highlights the key metabolites and metabolic pathways disturbed in NPC and presents potential biomarkers for effective diagnosis of this disease.

Rac1 Signaling in Amygdala Astrocytes Regulates Fear Memory Acquisition and Retrieval.

The importance of astrocytes in behavior control is increasingly appreciated, but little is known about the effects of their dynamic activity in regulating learning and memory. In the present study, we constructed AAVs of photoactivatable and photoinactivatable Ras-related C3 botulinum toxin substrate 1 (Rac1) under the mGFAP promoter, which enabled the manipulation of Rac1 activity in astrocytes by optical stimulation in free-moving mice. We found that both up-regulation and down-regulation of astrocytic Rac1 activity in the basolateral amygdala (BLA) attenuated memory acquisition in a fear conditioning mouse model. Meanwhile, neuronal activation in the BLA induced by memory acquisition was inhibited under both the up- and down-regulation of astrocytic Rac1 activity during training. In terms of the impact on fear memory retrieval, we found both up- and down-regulation of BLA astrocytic Rac1 activity impaired memory retrieval of fear conditioning and memory retrieval-induced neuronal activation. Notably, the effect of astrocytic Rac1 on memory retrieval was reversible. Our results demonstrate that the normal activity of astrocytic Rac1 is necessary for the activation of neurons and memory formation. Both activation and inactivation of astrocytic Rac1 activity in the BLA reduced the excitability of neurons, and thereby impaired fear memory acquisition and retrieval.

COVID-19 outbreak prevention by early containment in Shantou, China.

INTRODUCTION: To report about the successful outbreak containment of COVID-19 in Shantou, one of the prefectural cities of Guangdong province in the mainland China. METHODOLOGY: All patients confirmed as having COVID-19 between 23 January and 25 March 2020 by RT-PCR assay in the clinical lab of Shantou CDC were included and divided into three groups based on the source of identification: hospital diagnosis, contact tracing, and community screening. Collected data was analyzed and compared among these three groups. RESULTS: A total of 25 COVID-19 cases were identified in Shantou. The first case was identified on 14 January 2020 at one of two COVID-19 dedicated hospitals in Shantou. The majority of the cases were either imported from Wuhan or linked to Wuhan/Hubei. The median lag time for diagnosis (i.e., the time between symptom onset and case confirmation) was 2 days (IQR, 2.0-4.0) for all cases, 9 days (IQR, 7.0-10.0) for the cases diagnosed in hospitals, 2 days (IQR, 1.5-2.0) for the cases in contact tracing, and 4 days (IQR, 2.5-4.5) for cases in community screening, with a significantly longer diagnosis lag time in hospitals (p = 0.003). Multivariate linear regression models showed larger family size and severe cases as the significant predictor for increasing number of close contacts. CONCLUSIONS: The current pandemic appears to exist for an uncertain period. The early containment measures applied in Shantou, a city with insufficient healthcare resources for COVID-19, seems to be appropriate for cities or areas with similar profiles.

LRCH1 deficiency enhances LAT signalosome formation and CD8+ T cell responses against tumors and pathogens.

CD8+ T cells play pivotal roles in eradicating pathogens and tumor cells. T cell receptor (TCR) signaling is vital for the optimal activation of CD8+ T cells. Upon TCR engagement, the transmembrane adapter protein LAT (linker for activation of T cells) recruits other key signaling molecules and forms the "LAT signalosome" for downstream signal transduction. However, little is known about which functional partners could restrain the formation of the LAT signalosome and inhibit CD8+ cytotoxic T lymphocyte (CTL)-mediated cytotoxicity. Here we have demonstrated that LRCH1 (leucine-rich repeats and calponin homology domain containing 1) directly binds LAT, reduces LAT phosphorylation and interaction with GRB2, and also promotes the endocytosis of LAT. Lrch1-/- mice display better protection against influenza virus and Listeria infection, with enhanced CD8+ T cell proliferation and cytotoxicity. Adoptive transfer of Lrch1-/- CD8+ CTLs leads to increased B16-MO5 tumor clearance in vivo. Furthermore, knockout of LRCH1 in human chimeric antigen receptor (CAR) T cells that recognize the liver tumor-associated antigen glypican-3 could improve CAR T cell migration and proliferation in vitro. These findings suggest LRCH1 as a potential translational target to improve T cell immunotherapy against infection and tumors.

Inflammatory Renin-Angiotensin System Disruption Attenuates Sensory Hyperinnervation and Mechanical Hypersensitivity in a Rat Model of Provoked Vestibulodynia.

Vestibulodynia is characterized by perivaginal mechanical hypersensitivity, hyperinnervation, and abundant inflammatory cells expressing renin-angiotensin system proteins. We developed a tractable rat model of vestibulodynia to further assess the contributions of the renin-angiotensin system. Complete Freund's adjuvant injected into the posterior vestibule induced marked vestibular hypersensitivity throughout a 7-day test period. Numbers of axons immunoreactive for PGP9.5, calcitonin gene-related peptide, and GFRα2 were increased. Numbers of macrophages and T cells were also increased whereas B cells were not. Renin-angiotensin-associated proteins were abundant, with T cells as well as macrophages contributing to increased renin and angiotensinogen. Media conditioned with inflamed vestibular tissue promoted neurite sprouting by rat dorsal root ganglion neurons in vitro, and this was blocked by the angiotensin II receptor type 2 receptor antagonist PD123319 or by an angiotensin II function blocking antibody. Sensory axon sprouting induced by inflamed tissue was dependent on activity of angiotensin-converting enzyme or chymase, but not cathepsin G. Thus, vestibular Complete Freund's adjuvant injection substantially recapitulates changes seen in patients with provoked vestibulodynia, and shows that manipulation of the local inflammatory renin-angiotensin system may be a useful therapeutic strategy. PERSPECTIVE: This study provides evidence that inflammation of the rat vestibule induces a phenotype recapitulating behavioral and cytological features of human vestibulodynia. The model confirms a crucial role of the local inflammatory renin-angiotensin system in hypersensitivity and hyperinnervation. Targeting this system holds promise for developing new nonopioid analgesic treatment strategies.

Fear Conditioning Downregulates Rac1 Activity in the Basolateral Amygdala Astrocytes to Facilitate the Formation of Fear Memory.

Astrocytes are well known to scale synaptic structural and functional plasticity, while the role in learning and memory, such as conditioned fear memory, is poorly elucidated. Here, using pharmacological approach, we find that fluorocitrate (FC) significantly inhibits the acquisition of fear memory, suggesting that astrocyte activity is required for fear memory formation. We further demonstrate that fear conditioning downregulates astrocytic Rac1 activity in basolateral amygdala (BLA) in mice and promotes astrocyte structural plasticity. Ablation of astrocytic Rac1 in BLA promotes fear memory acquisition, while overexpression or constitutive activation of astrocytic Rac1 attenuates fear memory acquisition. Furthermore, temporal activation of Rac1 by photoactivatable Rac1 (Rac1-PA) induces structural alterations in astrocytes and in vivo activation of Rac1 in BLA astrocytes during fear conditioning attenuates the formation of fear memory. Taken together, our study demonstrates that fear conditioning-induced suppression of BLA astrocytic Rac1 activity, associated with astrocyte structural plasticity, is required for the formation of conditioned fear memory.

Motor, Somatosensory, Viscerosensory and Metabolic Impairments in a Heterozygous Female Rat Model of Rett Syndrome.

Rett Syndrome (RTT), an autism-related disorder caused by mutation of the X-linked Methyl CpG-binding Protein 2 (MECP2) gene, is characterized by severe cognitive and intellectual deficits. While cognitive deficits are well-documented in humans and rodent models, impairments of sensory, motor and metabolic functions also occur but remain poorly understood. To better understand non-cognitive deficits in RTT, we studied female rats heterozygous for Mecp2 mutation (Mecp2-/x); unlike commonly used male Mecp2-/y rodent models, this more closely approximates human RTT where males rarely survive. Mecp2-/x rats showed rapid, progressive decline of motor coordination through six months of age as assessed by rotarod performance, accompanied by deficits in gait and posture. Mecp2-/x rats were hyper-responsive to noxious pressure and cold, but showed visceral hyposensitivity when tested by colorectal distension. Mecp2-/x rats ate less, drank more, and had more body fat resulting in increased weight gain. Our findings reveal an array of progressive non-cognitive deficits in this rat model that are likely to contribute to the compromised quality of life that characterizes RTT.

[Optogenetic activation of dorsal hippocampal astrocytic Rac1 blocks the learning of associative memory].

Rac1 belongs to the family of Rho GTPases, and plays important roles in the brain function. It affects the cell migration and axon guidance via regulating the cytoskeleton and cellular morphology. However, the effect of its dynamic activation in regulating physiological function remains unclear. Recently, a photoactivatable analogue of Rac1 (PA-Rac1) has been developed, allowing the activation of Rac1 by the specific wavelength of light in living cells. Thus, we constructed recombinant adeno-associated virus (AAV) of PA-Rac1 and its light-insensitive mutant PA-Rac1-C450A under the control of the mouse glial fibrillary acidic protein (mGFAP) promoter to manipulate Rac1 activity in astrocytes by optical stimulation. Primary culture of hippocampal astrocytes was infected with the recombinant AAV-PA-Rac1 or AAV-PA-Rac1-C450A. Real-time fluorescence imaging showed that the cell membrane of the astrocyte expressing PA-Rac1 protruded near the light spot, while the astrocyte expressing PA-Rac1-C450A did not. We injected AAV-PA-Rac1 and AAV-PA-Rac1-C450A into dorsal hippocampus to investigate the role of the activation of Rac1 in regulating the associative learning. With optical stimulation, the PA-Rac1 group, rather than the PA-Rac1-C450A group, showed slower learning curve during the fear conditioning compared with the control group, indicating that activating astrocytic Rac1 blocks the formation of contextual memory. Our data suggest that the activation of Rac1 in dorsal hippocampal astrocyte plays an important role in the associative learning.

A Local Inflammatory Renin-Angiotensin System Drives Sensory Axon Sprouting in Provoked Vestibulodynia.

Vestibulodynia is a form of provoked vulvodynia characterized by profound tenderness, hyperinnervation, and frequently inflammation within well-defined areas of the human vestibule. Previous experiments in animal models show that inflammatory hypersensitivity and hyperinnervation occur in concert with establishment of a local renin-angiotensin system (RAS). Moreover, mechanical hypersensitivity and sensory axon sprouting are prevented by blocking effects of angiotensin II on angiotensin II receptor type 2 (AT2) receptors. This case-control study assessed whether a RAS contributes to hyperinnervation observed in human vestibulodynia. Vestibular biopsies from asymptomatic controls or patients' nontender areas showed moderate innervation and small numbers of inflammatory cells. In women with vestibulodynia, tender areas contained increased numbers of mechanoreceptive nociceptor axons, T-cells, macrophages, and B-cells, whereas mast cells were unchanged. RAS proteins were increased because of greater numbers of T cells and B cells expressing angiotensinogen, and increased renin-expressing T cells and macrophages. Chymase, which converts angiotensin I to angiotensin II, was present in constant numbers of mast cells. To determine if tender vestibular tissue generates angiotensin II that promotes axon sprouting, we conditioned culture medium with vestibular tissue. Rat sensory neurons cultured in control-conditioned medium showed normal axon outgrowth, whereas those in tender tissue-conditioned medium showed enhanced sprouting that was prevented by adding an AT2 antagonist or angiotensin II neutralizing antibody. Hypersensitivity in provoked vestibulodynia is therefore characterized by abnormal mechanonociceptor axon proliferation, which is attributable to inflammatory cell-derived angiotensin II (or a closely related peptide) acting on neuronal AT2 receptors. Accordingly, reducing inflammation or blocking AT2 represent rational strategies to mitigate this common pain syndrome. PERSPECTIVE: This study provides evidence that local inflammation leads to angiotensin II formation, which acts on the AT2 to induce nociceptor axon sprouting in vulvodynia. Preventing inflammation and blocking AT2 therefore present potential pharmacological strategies for reducing vestibular pain.

Astroglial ß-Arrestin1-mediated Nuclear Signaling Regulates the Expansion of Neural Precursor Cells in Adult Hippocampus.

Adult hippocampal neurogenesis is crucial for preserving normal brain function, but how it is regulated by niche cells is uncertain. Here we show that ß-arrestin 1 (ß-arr1) in dentate gyrus (DG) regulates neural precursor proliferation. ß-arr1 knockout (KO) mice show reduced neural precursor proliferation in subgranular zone (SGZ) which could be rescued by selective viral expression of ß-arr1 but not its nuclear-function-deficient mutants under control of hGFAP promotor in DG. Compared with wild type astrocytes, ß-arr1 KO astrocytes nurture less neurospheres, and this may be attributed to changed activity of soluble, heat-sensitive excretive factors, such as BMP2. RNA-sequencing reveals that ß-arr1 KO DG astrocytes exhibit an aberrant gene expression profile of niche factors, including elevated transcription of Bmp2. Taken together, our data suggest that ß-arr1 mediated nuclear signaling regulates the production of excretive factors derived from niche astrocytes and expansion of neural precursors in DG, thus maintaining homeostasis of adult hippocampal neurogenesis.

Persistent genital hyperinnervation following progesterone administration to adolescent female rats.

Provoked vestibulodynia, a female pelvic pain syndrome affecting substantial numbers of women, is characterized by genital hypersensitivity and sensory hyperinnervation. Previous studies have shown that the risk of developing provoked vestibulodynia is markedly elevated following adolescent use of oral contraceptives with high progesterone content. We hypothesized that progesterone, a steroid hormone with known neurotropic properties, may alter genital innervation through direct or indirect actions. Female Sprague Dawley rats received progesterone (20 mg/kg subcutaneously) from Days 20-27; tissue was removed for analysis in some rats on Day 28, while others were ovariectomized on Day 43 and infused for 7 days with vehicle or 17beta estradiol. Progesterone resulted in overall increases in vaginal innervation at both Day 28 and 50 due to proliferation of peptidergic sensory and sympathetic (but not parasympathetic) axons. Estradiol reduced innervation in progesterone-treated and untreated groups. To assess the mechanisms of sensory hyperinnervation, we cultured dissociated dorsal root ganglion neurons and found that progesterone increases neurite outgrowth by small unmyelinated (but not myelinated) sensory neurons, it was receptor mediated, and it was nonadditive with NGF. Pretreatment of ganglion with progesterone also increased neurite outgrowth in response to vaginal target explants. However, pretreatment of vaginal target with progesterone did not improve outgrowth. We conclude that adolescent progesterone exposure may contribute to provoked vestibulodynia by eliciting persistent genital hyperinnervation via a direct effect on unmyelinated sensory nociceptor neurons and that estradiol, a well-documented therapeutic, may alleviate symptoms in part by reducing progesterone-induced sensory hyperinnervation.

Trophic factor and hormonal regulation of neurite outgrowth in sensory neuron-like 50B11 cells.

Sensory axon integrity and regenerative capacity are important considerations in understanding neuropathological conditions characterized by hyper- or insensitivity. However, our knowledge of mechanisms regulating axon outgrowth are limited by an absence of suitable high-throughput assay systems. The 50B11 cell line generated from rat embryonic dorsal root ganglion neurons offers a promising model for screening assays. Prior characterization shows that these cells express cytoskeletal proteins and genes encoding ion channels and neurotrophin receptors in common with sensory nociceptor neurons. In the present study we further characterized 50B11 cells in regard to their phenotypes and responsiveness to neurotrophic and hormonal factors. 50B11 cells express neuronal cytoplasmic proteins including beta-3 tubulin, peripherin (a marker of unmyelinated neurons), and the pan-neuronal ubiquitin hydrolase, PGP9.5. Only PGP9.5 immunoreactivity was uniformly distributed throughout soma and axons, and therefore presents the best means for visualizing the entire axon arbor. All cells co-express both NGF and GDNF receptors and addition of ligands increased neurite length. 50B11 cells also showed immunoreactivity for the estrogen receptor-α and the angiotensin receptor type II, and both 17-ß estradiol and angiotensin II increased outgrowth by differentiated cells. 50B11 cells therefore show features reported previously for primary unmyelinated nociceptor neurons, including responsiveness to classical neurotrophins and hormonal modulators. Coupled with their ease of culture and predictable differentiation, 50B11 cells represent a promising cell line on which to base assays that more clearly reveal mechanisms regulating axon outgrowth and integrity.

Angiotensin II receptor type 2 activation is required for cutaneous sensory hyperinnervation and hypersensitivity in a rat hind paw model of inflammatory pain.

UNLABELLED: Many pain syndromes are associated with abnormal proliferation of peripheral sensory fibers. We showed previously that angiotensin II, acting through its type 2 receptor (AT2), stimulates axon outgrowth by cultured dorsal root ganglion neurons. In this study, we assessed whether AT2 mediates nociceptor hyperinnervation in the rodent hind paw model of inflammatory pain. Plantar injection of complete Freund's adjuvant (CFA), but not saline, produced marked thermal and mechanical hypersensitivity through 7 days. This was accompanied by proliferation of dermal and epidermal PGP9.5-immunoreactive (ir) and calcitonin gene-related peptide-immunoreactive (CGRP-ir) axons, and dermal axons immunoreactive for GFRα2 but not tyrosine hydroxylase or neurofilament H. Continuous infusion of the AT2 antagonist PD123319 beginning with CFA injection completely prevented hyperinnervation as well as hypersensitivity over a 7-day period. A single PD123319 injection 7 days after CFA also reversed thermal hypersensitivity and partially reversed mechanical hypersensitivity 3 hours later, without affecting cutaneous innervation. Angiotensin II-synthesizing proteins renin and angiotensinogen were largely absent after saline but abundant in T cells and macrophages in CFA-injected paws with or without PD123319. Thus, emigrant cells at the site of inflammation apparently establish a renin-angiotensin system, and AT2 activation elicits nociceptor sprouting and heightened thermal and mechanical sensitivity. PERSPECTIVE: Short-term AT2 activation is a potent contributor to thermal hypersensitivity, whereas long-term effects (such as hyperinnervation) also contribute to mechanical hypersensitivity. Pharmacologic blockade of AT2 signaling represents a potential therapeutic strategy aimed at biologic mechanisms underlying chronic inflammatory pain.

Systemic and topical hormone therapies reduce vaginal innervation density in postmenopausal women.

OBJECTIVE: Menopause is often accompanied by vaginal discomfort including burning, itching, dryness, and spontaneous or provoked pain. Although the direct effects of estrogen withdrawal on vaginal cells are implicated, surgical menopause in rodents causes autonomic and sensory nerves to proliferate, suggesting that indirect effects mediated by changes in vaginal innervation may contribute. We assessed whether postmenopausal women display hormone-dependent changes in vaginal innervation. METHODS: Vaginal biopsies from 20 postmenopausal women undergoing surgery for stress urinary incontinence and pelvic organ prolapse were fixed and immunostained for the pan-neuronal marker protein gene product 9.5, sympathetic marker tyrosine hydroxylase, parasympathetic marker vasoactive intestinal polypeptide, and sensory nociceptor marker calcitonin gene-related peptide. Innervation density was measured as an apparent percentage of the section area occupied by immunofluorescent axons. Specimens were grouped according to whether participants received systemic hormone therapy (HT), topical (vaginal) HT, or no HT. RESULTS: Women not receiving HT showed relatively high levels of total innervation, with most axons expressing tyrosine hydroxylase or vasoactive intestinal polypeptide immunoreactivity. In women receiving systemic HT, overall innervation was reduced, as were presumptive parasympathetic, sympathetic, and sensory axon populations. Topical HT elicited more dramatic reductions in innervation than in systemic HT. CONCLUSIONS: Hormone therapy reduces autonomic and sensory vaginal innervation density, which may, in part, contribute to relief from vaginal discomfort. Moreover, topical therapy is more effective than systemic therapy, which may help explain the greater improvement reported with topical compared with systemic HT.

Adaptive plasticity of vaginal innervation in term pregnant rats.

Changes in reproductive status place varied functional demands on the vagina. These include receptivity to male intromission and sperm transport in estrus, barrier functions during early pregnancy, and providing a conduit for fetal passage at parturition. Peripheral innervation regulates vaginal function, which in turn may be influenced by circulating reproductive hormones. We assessed vaginal innervation in diestrus and estrus (before and after the estrous cycle surge in estrogen), and in the early (low estrogen) and late (high estrogen) stages in pregnancy. In vaginal sections from cycling rats, axons immunoreactive for the pan-neuronal marker protein gene product 9.5 (PGP 9.5) showed a small reduction at estrus relative to diestrus, but this difference did not persist after correcting for changes in target size. No changes were detected in axons immunoreactive for tyrosine hydroxylase (sympathetic), vesicular acetylcholine transporter (parasympathetic), or calcitonin gene-related peptide and transient receptor potential vanilloid type 1 (TRPV-1; sensory nociceptors). In rats at 10 days of pregnancy, innervation was similar to that observed in cycling rats. However, at 21 days of pregnancy, axons immunoreactive for PGP 9.5 and each of the subpopulation-selective markers were significantly reduced both when expressed as percentage of sectional area or after correcting for changes in target size. Because peripheral nerves regulate vaginal smooth muscle tone, blood flow, and pain sensitivity, reductions in innervation may represent important adaptive mechanisms facilitating parturition.

Regulation of cardiac sympathetic afferent reflex by GABA(A) and GABA(B) receptors in paraventricular nucleus in rats.

The aim of the present study was to determine the role of GABA(A) and GABA(B) receptors in paraventricular nucleus (PVN) in regulating cardiac sympathetic afferent reflex (CSAR). Under urethane (800 mg/kg) and alpha-chloralose (40 mg/kg) anesthesia, renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP) and heart rate (HR) were recorded in sinoaortic-denervated and cervical-vagotomized rats. CSAR was evaluated based in the response of RSNA to epicardial application of capsaicin (0.3 nmol) or bradykinin (1 nmol). Bilateral PVN microinjection of the GABA(A) receptor agonist isoguvacine (10 nmol) attenuated CSAR, while the GABA(B) receptor agonist baclofen (1 nmol) abolished CSAR. Both isoguvacine and baclofen greatly decreased baseline RSNA and MAP. The GABA(A) receptor antagonist gabazine (0.1 nmol) had no significant effect on CSAR, but the GABA(B) receptor antagonist CGP-35348 (10 nmol) enhanced CSAR. Gabazine caused greater increases in baseline RSNA and MAP than CGP-35348. Vigabatrin (10 nmol), a selective GABA-transaminase inhibitor which increases endogenous GABA level, abolished CSAR, and decreased baseline RSNA, MAP and HR. The effects of vigabatrin were antagonized by combined gabazine (0.1 nmol) and CGP-35348 (10 nmol). The results indicate that activation of either GABA(A) or GABA(B) receptors in the PVN inhibits CSAR, while blockage of GABA(B) receptors in the PVN enhances CSAR. Endogenous GABA in the PVN could have an important role in regulating CSAR.