Nerve Growth Factor mRNA Levels Research Articles

Inhibitory Effect of Evodiamine on Psoriasis Lesions and Itching in Mice.

This study seeks to investigate the effect of evodiamine on psoriasis and psoriatic pruritus. Imiquimod-induced psoriasiform dermatitis in mice was used as a model, and evodiamine was topically applied for seven days. The mice were observed daily for skin damage on the back, clinical score and their scratching behavior was recorded. Blood samples were collected on the final day of the experiment, and the serum levels of pruritus-associated inflammatory cytokines tumor necrosis factor (TNF) -α, interleukin (IL) -23, and IL-17A were measured using enzyme-linked immunosorbent assay. Histopathological changes were observed in Hematoxylin and Eosin-stained skin specimens. The expression levels of transient receptor potential vanilloid (TRPV) 1, TRPV3, TRPV4, and the pruritus-related mediators Substance P (SP), nerve growth factor (NGF), and calcitonin gene-related peptide (CGRP) in the skin lesions were analyzed using Western blot and qRT-PCR. The effect of evodiamine on the exploratory behavior, motor, and coordination abilities of mice was assessed using open field, suspension, and Rota-Rod experiments. Molecular docking was utilized to verify the binding of evodiamine to the residues of TRPV1, TRPV3, and TRPV4. Evodiamine reduced pruritus and inhibited inflammation by decreasing the levels of inflammatory mediators TNF-α, IL-23, and IL-17A in the serum of Imiquimod-induced mice and attenuated the mRNA and protein expression levels of SP, NGF, CGRP, TRPV1, TRPV3, and TRPV4 in the skin. Evodiamine is an effective treatment for psoriasis and pruritus, due to its ability to inhibit immune inflammation and pruritic mediators.

Effects of Chlorinated Water on Neurite Length of Cultured Dorsal Root Ganglion Neurons and Semaphorin 3A Content of Cultured Epidermal Keratinocytes

The tap water that we normally use contains certain concentrations of free residual chlorine to kill microorganisms and viruses and make it safe for use. Water containing free residual chlorine not only dries out our hair and skin but can also cause irritation and itching in some people—especially those with sensitive skin or reduced skin barrier function. We investigated the effects of free residual chlorine on cultured dorsal root ganglion neurons and cultured epidermal keratinocytes. First, we measured neurite length in cultured rat dorsal root ganglion neurons. Next, to evaluate the effects of chlorine on semaphorin 3A (Sema3A) and nerve growth factor (NGF) levels in cultured human epidermal keratinocytes, we used an enzyme-linked immunosorbent assay to measure NGF in the supernatant and polymerase chain reaction and Western blot to determine Sema3A and NGF levels. Chlorine elongated the neurite length and increased the number of projections in cultured rat dorsal root ganglion neurons. Although there were no changes in NGF mRNA or protein levels in the supernatant of cultured human epidermal keratinocytes in the presence of chlorine, Sema3A mRNA and protein levels decreased, and the ratio of Sema3A to NGF was also reduced.

Changes in nerve growth factor in vastus lateralis muscle after the first versus second bout of one-leg eccentric cycling.

Delayed onset muscle soreness (DOMS) develops after performing unaccustomed eccentric exercises. Animal studies have shown that DOMS is mechanical hyperalgesia through nociceptor sensitization induced by nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF) upregulated by cyclooxygenase-2 (COX-2). However, no previous study has investigated these in relation to DOMS in humans. This study compared the first and second bouts of one-leg eccentric cycling (ECC) for changes in NGF, GDNF, and COX-2 mRNA in the vastus lateralis (VL). Seven healthy adults (18-40 years) performed two bouts of ECC (10 sets of 50 contractions) with 80% maximal voluntary concentric peak torque separated by 2 weeks (ECC1, ECC2). Muscle soreness that was assessed by a visual analog scale and maximal voluntary isometric contraction (MVC) torque of the knee extensors were measured before, immediately after (MVC only), 24 and 48 h post-exercise. Muscle biopsy was taken from the VL before the first bout from nonexercised leg (control) and 24 h after each bout from the exercised leg, and analyzed for NGF, GDNF, and COX-2 mRNA. Peak DOMS was more than two times greater and MVC torque at 48 h post-exercise was approximately 20% smaller after ECC1 than ECC2 (p < 0.05), suggesting the repeated bout effect. NGF mRNA level was higher (p < 0.05) post-ECC1 (0.79 ± 0.68 arbitrary unit) than control (0.06 ± 0.07) and post-ECC2 (0.08 ± 0.10). GDNF and COX-2 mRNA did not show significant differences between control, post-ECC1, and post-ECC2. These results suggest that an increase in NGF is associated with the development of DOMS in humans.

Alleviating skin barrier disruption, skin inflammation and pruritus: a moisturizing spray containing β-glucan and panthenol

Abstract Objective: Chronic pruritus is induced by a variety of causes and is an extremely common clinical problem. Previous studies have shown that many inflammatory skin diseases are associated with dry skin-induced pruritus. However, the relationship between skin inflammation, skin barrier function, and pruritus remains unclarified. The present study aimed to explore this relationship using an acetone-ether-water (AEW) mouse model, and to investigate the effects of the combined application of β-glucan and panthenol in a moisturizing spray in this mouse model. Methods: A dry skin-induced chronic pruritus mouse model was established by repeated AEW treatment of the skin. The pruritic behavior, skin barrier function, and expression of molecules related to pruritus and inflammation in topical lesions were measured. The effects of a spray containing β-glucan and panthenol were observed. Results: AEW treatment triggered spontaneous scratching with significantly increased numbers of scratching bouts, and disrupted the skin barrier with a significant increase in transepidermal water loss (TEWL) values and a significant decrease in filaggrin gene (FLG) expression (P &lt; 0.01). AEW treatment also significantly increased the expression of NOD-like receptor family pyrin domain containing 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD (ASC), caspase-1, interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α), as well as the pruritis-associated molecules nerve growth factor (NGF), cathepsin S (CTSS), and thymic stromal lymphopoietin (TSLP) in the skin (P &lt; 0.01). Application of the spray containing β-glucan and panthenol significantly alleviated these responses, resulting in a reduced number of scratching bouts in 1 hour, reduced TEWL values, increased FLG mRNA expression, and reduced mRNA levels of NLRP3, ASC, caspase-1, IL-1β, TNF-α, CTSS, TSLP, and NGF in skin lesions (P &lt; 0.05). Conclusion: The present results suggest a relationship between damaged skin barrier function, pruritus, and inflammation. Adding β-glucan and panthenol to moisturizing skincare products may alleviate pruritus induced by dry skin, improve the damaged skin barrier function, and reduce the inflammatory state in local skin areas.

Pyrroloquinoline Quinone Regulates Enteric Neurochemical Plasticity of Weaned Rats Challenged With Lipopolysaccharide.

The enteric nervous system (ENS) is important for the intestinal barrier to defend and regulate inflammation in the intestine. The aim of this study was to investigate the effect of pyrroloquinoline quinone (PQQ) on regulating neuropeptide secretion by ENS neurons of rats challenged with lipopolysaccharide (LPS) to create enteritis. Thirty Sprague Dawley rats were divided into five groups, namely, basal (CTRL), basal plus LPS challenge (LPS), basal with 2.5 mg/kg b.w./day of PQQ plus challenge with LPS (PQQ 2.5), basal with 5.0 mg/kg b.w./day PQQ plus challenge with LPS (PQQ 5), and basal with 10.0 mg/kg b.w./day PQQ plus challenge with LPS (PQQ 10). After treatment with basal diet or PQQ for 14 days, rats were challenged with LPS except for the CTRL group. Rats were euthanized 6 h after the LPS challenge. Rats showed an increased average daily gain in PQQ treatment groups (P < 0.05). Compared with the LPS group, PQQ 5 and PQQ 10 rats showed increased villus height and villus height/crypt depth of jejunum (P < 0.05). In PQQ treatment groups, concentrations of IL-1β and TNF-α in serum and intestine of rats were decreased, and IL-10 concentration was increased in serum compared with the LPS group (P < 0.05). Compared with the LPS group, the concentration of neuropeptide Y (NPY), nerve growth factor (NGF), vasoactive intestinal peptide (VIP), substance P (SP), calcitonin gene-related peptide (CGRP), and brain-derived neurotropic factor (BDNF) in serum were decreased in PQQ treatment groups (P < 0.05). Compared with the LPS group, ileal mRNA levels of BDNF, NPY, and NGF were decreased in PQQ treatment groups (P < 0.05). Jejunal concentrations of SP, CGRP, VIP, BDNF, NPY, and NGF were decreased in PQQ treatment groups compared with the LPS group (P < 0.05). Compared with the LPS group, phosphor-protein kinase B (p-Akt)/Akt levels in jejunum and colon were decreased in PQQ treatment groups (P < 0.05). In conclusion, daily treatment with PQQ improved daily gain, jejunal morphology, immune responses. PQQ-regulated enteric neurochemical plasticity of ENS via the Akt signaling pathway of weaned rats suffering from enteritis.

Association of BDNF / TrkB and NGF / TrkA Levels in Postmortem Brain with Major Depression and Suicide.

Suicide Attempts are the main complications of Major Depressive Episodes and are difficult to predict. There is still a lack of knowledge about its neurochemical aspects. There is increasing evidence that Brain-derived neurotrophic factor (BDNF) and Nerve growth factor (NGF) play a role in the pathophysiology and treatment of depression by binding and activating cognate receptors Tyrosine Kinase B (TrkB) and Tyrosie Kinase A (TrkA), respectively. This study was conducted to examine whether BDNF and / or TrkB as well as NGF and / or TrkA expression profiles were changed in the hippocampus of postmortem brain of individuals with depression who committed suicide. This study was conducted with the brain tissue of 61 victims who died as a result of suicide due to depression and 25 people who died due to traffic accidents. The psychiatric history of the cases was determined by the psychological autopsy method. Samples were taken from the hippocampus region of the brain at the forensic medicine institution. After storage under appropriate conditions, protein and mRNA levels of BDNF, TrkB, NGF and TrkA were determined in the genetics laboratory. Average age of the suicide group was 30 and the average age of the control group was 24.5. The suicide group consisted of 70.5% male and 29.5% female cases. There was no significant difference between the groups in terms of age (p=0.062) and gender (p=0.718). BDNF, NGF, TrkA and TrkB values were found to be lower in the suicide group compared to the control group and there was a significant difference between the groups (p≤0.001; p=0.001; p=0.001; p=0.011). Given the importance of BDNF and NGF and their cognate receptors in mediating physiological functions, including cell survival and synaptic plasticity, our findings regarding decreased expression of BDNF, TrkB, NGF and TrkA in both protein and mRNA levels of postmortem brains of suicide victims suggests that it may play an important role in the pathophysiological aspects of its behavior. Further studies in this context may be useful both in understanding the molecular basis of suicide and in designing therapeutic models targeting these molecular pathways.

Effects of different implant surface properties on the biological behavior of Schwann cells.

This study investigated the effects of different implant surface properties on the biological behavior of Schwann cells. Schwann cells (SCs) were cultured on three types of implant surfaces including smooth polished (SMO), sand-blasted, large grit, acid-etched (SLA), and chemically-modified SLA (modSLA). At different time points, the morphology and adhesion of SCs on the implant surfaces were observed by scanning electron microscope. Cell proliferation activity was detected by MTT method. The expression levels of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) were detected by enzyme-linked immunosorbent assay. Changes in the mRNA levels of NGF and BDNF were detected by real-time fluorescent quantitative polymerase chain reaction (PCR). SCs adhered, stretched, and proliferated well on the three types of implant surfaces. On the 3rd, 5th, and 7th days, the OD values of the SMO group were higher than those of the SLA group and the modSLA group, and the difference was statistically significant (P<0.05). On the 3rd day, the expression and mRNA levels of NGF and BDNF in the SLA group and the modSLA group were higher than those in the SMO group (P<0.05); in particular, the levels in the modSLA group were higher than those in the SLA group (P<0.05). Different implant surface properties have different effects on the biological behavior of SCs. Proliferation of SCs is significantly promoted by smooth surface, while secretion and gene expression of neurotrophic factors are significantly promoted by modSLA surface at early stage.

Increased nerve growth factor expression in the synovial tissues of patients with rotator cuff tears.

BackgroundRotator cuff tears (RCTs) are often associated with severe shoulder pain. Non-steroidal anti-inflammatory drugs, not recommended for long-term use, do not effectively manage RCT-induced pain, resulting in reduced quality of life. To improve management, a better understanding of the fundamental properties of RCT pain is needed. Here, we aimed to compare the expression levels of nerve growth factor (NGF) and cyclooxygenase-2 (COX-2) mRNA in the synovial tissues of patients with RCT-induced pain and patients with non-painful recurrent shoulder dislocation (RSD).MethodsThe study included 32 patients with RCT who underwent arthroscopic rotator cuff repair and 28 patients with non-painful RSD who underwent arthroscopic Bankart repair. Synovial tissue samples were harvested from subacromial bursa and rotator interval of RCT patients and from the rotator interval of RSD patients. Samples were analyzed quantitatively expression levels for NGF and COX2 mRNA and NGF protein.ResultsNGF mRNA and protein levels were significantly higher in the rotator interval of RCT patients than in the rotator interval of RSD patients (p = 0.0017, p = 0.012, respectively), while COX2 mRNA levels did not differ significantly between the two patient groups. In RCT patients, COX2 mRNA was more highly expressed in the rotator interval than in the subacromial bursa (p = 0.038), whereas the mRNA and protein levels of NGF did not differ between the two tissues. The expression of NGF mRNA in the synovium of the rotator interval was significantly correlated with the numeric rating scale of pain (ρ = 0.38, p = 0.004).ConclusionNGF mRNA and protein levels were elevated in patients with painful RCT compared with those in patients with non-painful RSD, whereas COX-2 levels were comparable in the two patient groups. These findings provide insights into novel potential strategies for clinical management of RCT.

Mycobacterium vaccae nebulization protects Balb/c mice against bronchial asthma through neural mechanisms

Objectives Bronchial asthma can be effectively controlled but not be cured, its etiology and pathogenesis are still unclear, and there are no effective preventive measures. The key characteristic of asthma is chronic airway inflammation, and recent research has found that airway neurogenic inflammation plays an important role in asthma. We previously found that Mycobacterium vaccae nebulization protects against asthma. Therefore, this objective of this study is to explore the effect of M. vaccae nebulization on asthmatic neural mechanisms. Methods A total 18 of female Balb/c mice were randomized into normal, asthma control, and M. vaccae nebulization (Neb.group) groups, and mice in the Neb.group were nebulized with M. vaccae one month before the asthmatic model was established. Then, 1 month later, the mice were sensitized and challenged with ovalbumin. Twenty-four hours after the last challenge, mouse airway responsiveness; pulmonary brain-derived neurotropic factor (BDNF), neurofilament-medium length (NF-M, using NF09 antibody), and acetylcholine expression; and nerve growth factor (NGF) mRNA level were determined. Results We found that the BDNF, NF09, acetylcholine expression, and NGF mRNA level were decreased in the Neb.group compared with levels in the asthma control group. Conclusion M. vaccae nebulization may protected in Balb/c mice against bronchial asthma through neural mechanisms.

Role of interleukin-1β in nerve growth factor expression, neurogenesis and deep dyspareunia in endometriosis.

Does interleukin-1β (IL-1β) play a role in promoting nerve growth factor expression, neurogenesis and deep dyspareunia in endometriosis? IL-1β directly stimulates nerve growth factor (NGF) expression in endometriosis and is associated with local neurogenesis around endometriosis and more severe deep dyspareunia. Local nerve density around endometriosis (using the pan-neuronal marker PGP9.5) is associated with deep dyspareunia in endometriosis, mediated in part by NGF expression. This in vitro study included endometriotic tissue samples from 45 patients. This study was conducted in a university hospital affiliated research institute and included 45 women with surgically excised deep uterosacral/rectovaginal endometriosis (DIE, n = 12), ovarian endometriomas (OMA, n = 14) or superficial peritoneal uterosacral/cul-de-sac endometriosis (SUP, n = 19). Immunolocalisation of IL-1β, IL-1 receptor type 1 (IL-1R1), NGF and PGP9.5 in endometriotic tissues was examined by immunohistochemistry (IHC), and the intensity of IHC staining in the endometriotic epithelium and stroma was semi-quantitatively evaluated using the Histoscore method (H-score). For each case, deep dyspareunia was pre-operatively rated by the patient on an 11-point numeric rating scale (0-10). In addition, primary endometriosis stromal cells were isolated and cultured from surgically excised endometriosis. These cells were treated with IL-1β alone or in combination of Anakinra (an inhibitor of IL-1R1), small inference RNA (siRNA) against IL-1R1, siRNA against c-FOS or NGF neutralising antibody. The mRNA and protein levels of target genes (NGF and c-FOS) were assessed by reverse-transcription qPCR and western blot/ELISA, respectively. Furthermore, immunofluorescent microscopy was used to examine the neurite growth of rat pheochromocytoma PC-12 cells, as an in vitro model of neurogenesis. For IHC, IL-1β expression in the endometriosis epithelium was significantly associated with more severe deep dyspareunia (r = 0.37, P = 0.02), higher nerve fibre bundle density around endometriosis (r = 0.42, P = 0.01) and greater NGF expression by the endometriosis epithelium (r = 0.42, P = 0.01) and stroma (r = 0.45, P = 0.01). In primary endometriosis stromal cells, treatment with exogenous IL-1β significantly increased the mRNA and protein levels of NGF and c-FOS. Pre-treatment with Anakinra, siRNA against IL-1R1, or siRNA against c-FOS, each attenuated IL-1 β-induced increases of NGF expression. In addition, supernatants from IL-1β treated endometriosis stromal cells significantly stimulated PC-12 neurite growth compared to controls, and these effects could be attenuated by pre-treatment with NGF neutralising antibody or Anakinra. N/A. We did not have data from cultures of endometriosis glandular epithelium, due to the known difficulties with primary cultures of this cell type. Our study revealed a mechanism for deep dyspareunia in endometriosis, whereby IL-1β stimulates NGF expression, promoting local neurogenesis around endometriosis, which in turn leads to tender pelvic anatomic sites and thus deep-hitting dyspareunia. There may also be potential for drug targeting of IL-1β and/or NGF in the management of endometriosis-associated pain. This study was funded by grants from the Canadian Institutes of Health Research (MOP-142273 and PJT-156084). P.Y. is also supported by a Health Professional Investigator Award from the Michael Smith Foundation for Health Research. MB has financial affiliations with Abbvie and Allergan. Otherwise, there are no conflicts of interest to declare.

Expression of Neurotrophic Factors, Tight Junction Proteins, and Cytokines According to the Irritable Bowel Syndrome Subtype and Sex.

Background/AimsEmerging evidence shows that the mechanism of irritable bowel syndrome (IBS) is associated with neurotrophic factors and tight junction proteins (TJPs). It is known that there are sex differences in the pathophysiology of IBS. The aim of the present study is to determine expression levels of neurotrophic factors, TJPs, and cytokines according to IBS subtype and sex.MethodsFrom 59 IBS (33 IBS-constipation, 21 IBS-diarrhea, and 5 IBS-mixed) and 36 control patients, colonic mucosa mRNA expression levels of transient receptor potential vanilloid-1 (TRPV1), nerve growth factor (NGF), glial cell-derived neurotrophic factor (GDNF), and various TJPs were assessed by real-time polymerase chain reaction. Western blot was performed to determine levels of zonular occludens-1 (ZO-1). Serum levels of cytokines were measured by enzyme-linked immunosorbent assay.ResultsTRPV1, GDNF, and NGF mRNA levels were significantly increased in those with IBS-constipation compared to those in controls (all P < 0.05). However, they showed no significant difference between those with IBS-diarrhea and controls. Expression level of TRPV1 correlated with that of GDNF (r = 0.741, P < 0.001) and NGF (r = 0.935, P < 0.001). ZO-1 RNA expression levels were lower (P = 0.021) in female IBS-diarrhea than those in controls, although they showed no significant differences between male IBS-diarrhea and controls. Serum IL-1β levels in female IBS were significantly higher than those of male IBS, especially in IBS-constipation (P < 0.001).ConclusionOur results suggest that neurotrophic factors and IL-1β are closely related to IBS-constipation and that decrease of ZO-1 is an important factor in female with IBS-diarrhea.

Hypomethylation of nerve growth factor (NGF) promotes binding of C/EBP\u03b1 and contributes to inflammatory hyperalgesia in rats

BackgroundChronic pain usually accompanied by tissue damage and inflammation. However, the pathogenesis of chronic pain remains unclear.MethodsWe investigated the role of nerve growth factor (NGF) in chronic inflammatory pain induced by complete Freund’s adjuvant (CFA), explored the methylation status of CpG islands in the promoter region of the NGF gene, and clarified the function and mechanism of C/EBPα-NGF signaling pathway from epigenetic perspective in the chronic inflammatory pain model.ResultsCFA induced significant hyperalgesia and continuous upregulation of NGF mRNA and protein levels in the L4–6 dorsal root ganglions (DRGs) in rats. Hypomethylation of CpG islands occurred in the NGF gene promoter region after CFA treatment. At the same time, the miR-29b expression level was significantly increased, while the DNA methyltransferase 3b (DNMT3b) level reduced significantly. Moreover, CFA treatment promoted binding of C/EBPα to the NGF gene promoter region and C/EBPα siRNA treatment obviously decreased expression of NGF levels and also alleviate inflammatory hyperalgesia significantly in rats.ConclusionCollectively, the results indicated that CFA leads to the upregulation of miR-29b level, which represses the expression of DNMT3b, enhances the demethylation of the NGF gene promoter region, and promotes the binding of C/EBPα with the NGF gene promoter, thus results in the upregulation of NGF gene expression and maintenance of chronic inflammatory pain.

Increased expression of neurogenic factors in uterine fibroids.

Are selective markers for the neuronal differentiation such as microtubule-associated protein 2 (MAP-2) and synaptophysin (SYP) as well as the nerve growth factor (NGF) expressed by fibroids, myometrium and eutopic endometrium? Neuronal markers NGF, MAP-2 and SYP are highly expressed in fibroids compared with matched myometrium, and this neurogenic pathway is upregulated by tumor necrosis factor (TNF) alpha in cultured smooth muscle cells (SMCs). Uterine fibroids or leiomyomas are the most common benign tumors, accounting for approximately one-third of hysterectomies. The present trend is to improve the medical treatment avoiding surgery, also for fertility sparing; hence, the pathogenic mechanisms are investigated, aiming to develop new therapeutic strategy. This laboratory-based case-control study is focused on fibroids and myometrial specimens obtained between 2015 and 2017 from 15 women of reproductive age at the proliferative phase of the menstrual cycle. Leiomyomas, matched myometrium and endometrium from each woman were analyzed. Control endometrium was obtained from women undergoing surgery for ovarian cyst (n = 15). qRT-PCR, western blotting and immunostaining were applied to evaluate the expression of neurogenic markers; the effects of TNF on NGF, MAP-2 and SYP expression in cultured SMCs from leiomyomas and matched myometrium were analyzed. qRT-PCR analyses using tissues from clinical patients showed that the levels of NGF, MAP-2 and SYP mRNA were significantly higher in uterine leiomyomas compared with their matched myometrium (P < 0.05), whereas only NGF was significantly increased in eutopic endometrium compared with healthy endometrium. In primary SMCs, isolated from fibroids or from the adjacent myometrium, NGF, MAP-2 and SYP mRNA expression were significantly increased by TNF treatment (P < 0.05). Finally, human endometrial stromal cells prepared from the endometrium of patients affected by uterine fibroids display higher TNF expression (P < 0.001). qRT-PCR analysis and immunofluorescence validation are robust methods demonstrating a clear upregulation of neurogenic factors in leiomyomas, even though additional studies are needed to establish a correlation between increased neuronal gene expression and degree of pain, as well as the involvement of inflammation mediators in the development of the neurogenic unhinge. Therefore, more in vivo studies are needed to confirm the results achieved from primary cultured SMCs. The increased expression of neurogenic factors in uterine fibroids and endometrium may contribute to explain the painful stimuli. Accordingly, these neurogenic pathways may represent potential therapeutic avenues to treat the fibroid-related disorders. This study was supported by research grants from the University of Siena. The authors declare no conflict of interest. N/A.

Roman chamomile inhalation combined with clomipramine treatment improves treatment-resistant depression-like behavior in mice

It is well known that chamomile is one of the oldest known medicinal herbs and has been used to treat various disorders, but it is mainly German chamomile. The effects of Roman chamomile on depression still unclear. In this study, we used chronically stressed mice to investigate whether inhalation of Roman chamomile essential oil affects depression-like behavior. We previously reported that restraint and water immersion stress produce depression-like behavior and a blunted response to the tricyclic antidepressant clomipramine. Each mouse was exposed to restraint and water immersion stress for 15 days, and resistance to the effect of clomipramine was induced in a behavioral despair paradigm. In the present study, we found that cotreatment with clomipramine and inhalation of Roman chamomile attenuated depression-like behavior in a forced swim test. Next, we examined the hippocampal mRNA levels of two cytokines, tumor necrosis factor (TNF) alpha and interleukin-6 (IL-6); a neurotrophic factor, brain derived-neurotrophic factor (BDNF); and nerve growth factor (NGF). TNF alpha, IL-6 and BDNF mRNA levels did not change in the hippocampus of stressed mice. However, the NGF mRNA level was significantly decreased, and this decrease was not attenuated by treatment with clomipramine or inhalation of Roman chamomile alone. We also examined whether Roman chamomile combined with clomipramine treatment affects hippocampal neurogenesis and serum corticosterone levels. Stressed mice had fewer doublecortin (DCX)-positive cells in the subgranular zone of the dentate gyrus, but this was significantly attenuated by Roman chamomile and clomipramine treatment. In addition, the serum corticosterone level was also significantly decreased by treatment with Roman chamomile and clomipramine. These results suggest that Roman chamomile inhalation may enhance the antidepressant effect of clomipramine by increasing hippocampal neurogenesis and modulating corticosterone levels in patients with treatment-resistant depression.

Formation of gap junctions between adipose stem cells-derived Schwann cells in a rat model of dyskinesia induced by brain injury

To investigate the formation of gap junctions between Schwann cells derived from differentiated adipose stem cells implanted in a rat model of dyskinesia induced by brain injury and its positive effect in promoting functional recovery of the rats. In a rat model of hemiplegia induced by motor cortex injury, adipose stem cells or Schwann cells differentiated from adipose stem cells, either with or without RNAi-mediated silencing of Cx43, were transplanted orthotopically in the lesion. The recovery of the motor function of the rats was observed and scored after the transplantation. Rat brain tissues were sampled to detect the expressions of nerve growth factor (NGF) using Western blotting and RT-PCR. All the 3 cell transplantation therapies obviously improved the motor function scores of the rats as compared with the control rats. The expression of NGF in the brain tissue was significantly lower in the control group than in the cell transplantation groups. NGF expression in the brain tissues of rats receiving transplantation of Schwann cells with Cx43 gene silencing was lower than that in rats receiving Schwann cells without Cx43 silencing, and was similar with that in rats transplanted with adipose stem cells. The results of RT-PCR showed that NGF mRNA level in the control group was significantly lower than that in the other 3 groups. NGF mRNA expression was the highest in Schwann cell group without Cx43 silencing, followed by adipose stem cell group, and then by Schwann cell group with Cx43 silencing. In the rat model of dyskinesia induced by brain injury, transplantations of adipose stem cells and adipose stem cells-derived Schwann cells both promote the functional recovery of brain damage, in which gap junction protein Cx43 plays an important role to promote functional gap junction formation possibly by enhancing NGF expression.

High hydrostatic pressure induces apoptosis of retinal ganglion cells via regulation of the NGF signalling pathway.

High pressure is the most important factor inducing retinal ganglion cell (RGC) apoptosis. However, the underlying mechanisms remain obscure. The present study investigated the effects of different levels of hydrostatic pressure (HP) on RGCs and the potential mechanisms involved. Primary cultured rat RGCs were exposed to five levels of HP (0, 20, 40, 60 and 80 mmHg) for 24 h. Morphological changes in RGCs were observed. The viability and apoptosis rate of RGCs were detected using a Cell Counting Kit-8 assay and Annexin V-fluorescein isothiocyanate/propidium iodide flow cytometry, respectively. Western blotting, reverse transcription-quantitative polymerase chain reaction and immunofluorescence were used to detect the expression and mRNA levels of nerve growth factor (NGF), protein kinase B (AKT), apoptosis signal-regulating kinase 1 (ASK1), forkhead box O1 (FoxO1) and cAMP response element binding protein (CREB). In the 0- and 20-mmHg groups, there were no apoptotic morphological changes. In the 40 mmHg group, parts of the cell were shrunken or disrupted. In the 60 mmHg group, neurite extension was weakened and parts of the cells were disintegrating or dying. In the 80 mmHg group, the internal structures of the cells were not visible at all. The apoptosis rates of RGCs were significantly higher and the viability rates significantly lower under 40, 60 and 80 mmHg compared with under 0 or 20 mmHg (all P<0.01). The expression and mRNA levels of NGF, AKT and CREB decreased in a dose-dependent manner in the 40-, 60- and 80-mmHg groups (all P<0.05), but those of ASK1 and FoxO1 increased in a dose-dependent manner (all P<0.05). Interestingly, the alterations to the expression and mRNA levels of CREB were significantly larger compared with the changes in ASK1 or FoxO1 in the 40-, 60- and 80-mmHg groups (all P<0.01). The results of the present study demonstrate that elevated HP of 40, 60 or 80 mmHg reduces viability and induces apoptosis in RGCs, which may occur through effects on the NGF/ASK1/FoxO1 and NGF/AKT/CREB pathways, of which the latter is more strongly affected.

Effects of n-hexane on learning and memory and the expressions of nerve growth factor mRNA and nerve growth factor receptor mRNA of brain tissue in mice exposed to N-hexane

Objective: To Effects of n-hexane on learning and memory and the expressions of nerve growth factor (NGF) mRNA and nerve growth factor receptor (NGFR) mRNA of brain tissue in mice exposed to N-hexane. Methods: 40 Kunming mice were randomly divided into low-dose group, meddle-dose group, high-dose dose group and control group, with 10 mice in each group. All the groups were orally exposed to n-hexane in different doses: low-dose group with 43.5 mg/kg, middle-dose group with 86.5 mg/kg and high-dose group with 173.0 mg/kg, 1 time per day for 20 d. After the poisoning, the Y-arm test and the expressions of NGF mRNA and NGFR mRNA and the concentrations of NGF and NGFR in the brain tissues of each group were measured. Results: In the first Y-arm test, there existed a significant difference in correct reaction rate generally in all groups (P<0.05), and correct reaction rate in the middle-dose group and the high-dose group were low significantly compared with that in the control group(P<0.05). In the second Y-arm test, there existed a significant differences in total electric shock time and correct reaction rate generally in all groups (P<0.01), and the total electric shock time prolonged significantly and the correct response rate decreased significantly in 3 dose groups compared with those of the control group(P<0.05). The expression levels of NGF mRNA in brain tissues of low, meddle and high dose-groups were 0.81±0.66, 0.67±0.37 and 0.69±0.26, and the expression levels of NGFR mRNA were 1.22±0.42, 1.98±0.84 and 2.01±2.01, respectively. Compared with the control group, the expressions of NGF mRNA in the 3 dose groups decreased significantly (P<0.05), and the expression of NGFR mRNA in middle-and high-dose groups increased significantly (P<0.05). The concentrations of NGF in brain tissues of low,meddle and high dose-groups were 39.97±7.24 ng/L, 39.26±7.88 ng/L,31.70±8.21 ng/L,and the concentrations of NGFR were 17.37±6.82 ng/L,21.37±7.16 ng/L, 22.46±7.70 ng/L, respectively. Compared with the control group, the concentrations of NGF in high-dose groups decreased significantly(P<0.05), and the concentrations of NGFR in middle-and high-dose groups increased significantly (P<0.05). Conclusion: N-hexane exposure can result in decrease of learning and memory in mice, which may be related to abnormal expression of NGF mRNA and NGFR mRNA in brain tissue.

IL-1β mediates NGF and COX-2 expression through transforming growth factor-activating kinase 1 in subacromial bursa cells derived from rotator cuff tear patients

BackgroundIncreased interleukin (IL)-1β expression in the subacromial bursa (SAB) is associated with severe pain in rotator cuff tears (RCTs). Additionally, transforming growth factor (TGF)-β-activated kinase 1 (TAK1) is essential for cytokine-mediated cascades. TAK1 also regulates the expression of pain-associated molecules such as cycloxygenase-2 (COX-2) and nerve growth factor (NGF) in synovial fibroblasts; however, this regulation in the SAB is not fully understood. MethodsSAB samples were harvested from 18 subjects with RCTs. The expression and localization of NGF and COX-2 was determined using polymerase chain reaction (PCR) analysis and immunohistochemistry. Regulation of COX-2 and NGF by IL-1β in subacromial bursa cells (SABCs) was investigated by culturing and stimulating SABCs with vehicle control (culture medium), 50 ng/ml recombinant human IL-1β (rhIL1-β), 50 ng/ml rhIL-1β and 10 μM celecoxib (COX-2 inhibitor), or 10 μM prostaglandin E2 (PGE2) for 24 h. The effects of TAK1 inhibition on rhIL-1β stimulation were determined by culturing and treating SABCs with control, 50 ng/ml rhIL-1β, or 50 ng/ml rhIL-1β and 10 μM (5Z)-7-oxozeaenol (TAK1 inhibitor) for 24 h. NGF and COX-2 mRNA expression was monitored using quantitative PCR. ResultsCOX-2 and NGF mRNA expression was observed in all SAB specimens. Immunohistochemical analysis showed that COX-2-positive cells were in the lining and sublining layers. NGF-positive cells were observed in the sublining layer. rhIL-1β treatment significantly increased NGF and COX-2 mRNA levels compared with control cells. The COX-2 inhibitor did not suppress rhIL-1β-induced NGF expression, and PGE2 stimulation did not alter NGF mRNA expression. In contrast, the TAK1 inhibitor significantly reduced rhIL-1β-stimulated COX-2 and NGF mRNA expression. ConclusionIL-1β regulates the expression of NGF and COX-2, pain-related molecules in the SAB, through TAK1. Therefore, TAK1 may be one potential therapeutic target for reducing pain in patients with RCTs.

Mechanism of interaction between ocular and nasal neurogenic inflammation in allergic rhinoconjunctivitis.

The mechanisms of naso-ocular interaction in allergic rhinoconjunctivitis are not well understood. Neurogenic inflammation affects both eyes and nose via the same neurogenic factors. The purpose of this study was to investigate the effects of neurogenic inflammation on conjunctival inflammation following nasal allergen provocation. Sensitized rats were exposed to ovalbumin (OVA) via the nose. Parts of the nasal mucosa and conjunctivae were sliced and used for hematoxylin-eosin staining, immunohistochemical analysis, western blotting, and real-time polymerase chain reaction. The slides were observed under a light microscope, and the acquired images were analyzed. The levels of substance P (SP), vasoactive intestinal peptide (VIP), and nerve growth factor (NGF) were detected. The levels of SP, VIP, and NGF were increased in both nasal mucosa and conjunctivae 1h and 24h after OVA administration (p < 0.05). Higher levels of SP, VIP, and NGF expression were observed in the nasal mucosa and conjunctivae 24h after OVA administration (p < 0.05). Following damage of the nasal sensory nerves by capsaicin, the protein and mRNA levels of SP, VIP, and NGF were reduced. In conclusion, the increased levels of VIP, SP, and NGF might be responsible for the ocular reaction following nasal challenge with allergen in rats.

Abstract 424: Knockdown of Nerve Growth Factor in Spinal Cord Reduces Myocardial Reperfusion Injury by Suppressing Ischemic Nociceptive Signaling Transmission

Objective: To test the hypothesis that nerve growth factor (NGF) sensitized spinal cardiac afferent activity contributes to myocardial ischemia-reperfusion injury (IRI). Methods and Results: The expression levels of NGF were increased in both dorsal root ganglion (DRG) and spinal cord after myocardial IRI in rats. Whole-cell patch clamp in cultured DRG neurons revealed that NGF aggravated capsaicin-induced current which was eliminated by the transient receptor potential vanilloid 1 (TRPV1) antagonist. Intrathecal injection of lentivirus-mediated NGF shRNA into thoracic T1-T8 segments successfully suppressed NGF mRNA and protein levels. Knockdown of NGF in spinal cord significantly limited IRI-induced infarct size, lowered down the concentration of cardiac troponin I (cTnI), and improved arrhythmia score. NGF gene suppression markedly reduced IRI-caused elevation of NGF and its receptor tropomyosin receptor kinase A (TrkA), TRPV1 and phosphorylated TRPV1. The activation of protein kinase B (Akt) and extracellular signal-regulated kinase (ERK) was also inhibited by NGF shRNA. Upon immunofluorescence staining, substance P (SP) and calcitonin gene-related peptide (CGRP) were found strongly stained in DRG and spinal dorsal horn following myocardial IRI, while these immunoreactive staining were repressed due to NGF gene knockdown. Conclusions: NGF aggravated spinal cardiac afferent nerves activity by sensitizing TRPV1-mediated nociceptive signal transmission. Inhibition of spinal NGF expression effectively protected the heart from myocardial IRI, which may imply a new potential therapeutic approach in ischemic heart diseases.