Stress Affects Mast Cell Proteases in Murine Skin in a Model of Atopic Dermatitis-like Allergic Inflammation.

Stress exposure worsens allergic inflammatory diseases substantially. Mast cells (MCs) play a key role in peripheral immune responses to neuroendocrine stress mediators such as nerve growth factor (NGF) and substance P (SP). Mast cell proteases (MCPs) and cholinergic factors (Chrna7, SLURP1) were recently described to modulate MC stress response. We studied MCPs and Chrna7/SLURP1 and their interplay in a mouse model for noise induced stress (NiS) and atopic dermatitis-like allergic inflammation (AlD) and in cultured MC lacking Chrna7. We found that the cholinergic stress axis interacts with neuroendocrine stress mediators and stress-mediator cleaving enzymes in AlD. SP-cleaving mMCP4+ MC were upregulated in AlD and further upregulated by stress in NiS+AlD. Anti-NGF neutralizing antibody treatment blocked the stress-induced upregulation in vivo, and mMCP4+ MCs correlated with measures of AlD disease activity. Finally, high mMCP4 production in response to SP depended on Chrna7/SLURP1 in cultured MCs. In conclusion, mMCP4 and its upstream regulation by Chrna7/SLURP1 are interesting novel targets for the treatment of allergic inflammation and its aggravation by stress.

[Effects of catgut embedding and PGLA embedding at "Zusanli" (ST 36) on skin mast cells, substance P and histamine in healthy rats].

OBJECTIVE: To observe the effects of catgut embedding and polyglycolic acid/poly-lactic acid (PGLA) embedding at "Zusanli" (ST 36) on the activation of local skin mast cells (MC), and expression of substance P (SP) and histamine (HA), and to explore the mechanism of the temporal stimulation effect of acupoint catgut embedding and provide a foundation for further research on the initiation mechanism of acupoint catgut embedding. METHODS: One hundred and sixty male SPF-grade SD rats were randomly divided into a blank group (10 rats), a sham-embedding group (50 rats), a catgut group (50 rats), and a PGLA group (50 rats). Each intervention group was further randomly divided into five subgroups according to the time points after intervention: 8 hours, 3 days, 7 days, 14 days, and 21 days, with 10 rats in each subgroup. One-time sham-embedding, catgut embedding and PGLA embedding was given at left "Zusanli" (ST 36) in each intervention group, respectively. The skin and subcutaneous connective tissue of the left "Zusanli" (ST 36) were collected at the corresponding time points after intervention, except for the blank group (only one day before intervention). Toluidine blue staining was used to detect MC count and degranulation, and immunohistochemical staining was used to detect the expression of SP and HA positive cells. RESULTS: There was no significant difference in MC count between the subgroups of each intervention group and the blank group (P>0.05). There was no significant difference in MC count between the subgroups of the catgut group and the PGLA group (P>0.05). The MC count in the 8-hour subgroup of PGLA group was higher than that in the 8-hour subgroup of catgut group (P<0.05), while the MC count in the 21-day subgroup of PGLA group was lower than that in the 21-day subgroup of catgut group (P<0.05). Compared with the blank group, the degranulation rates of MC were increased in the 8-hour and 3-day subgroups of sham-embedding group, 8-hour, 3-day, and 7-day subgroups of catgut group, and 8-hour, 3-day, 7-day, and 14-day subgroups of PGLA group (P<0.01, P<0.05, P<0.001). There was no significant difference in the degranulation rate of MC between the subgroups of the catgut group and the PGLA group (P>0.05), and no significant difference in the degranulation rate of MC between the two embedding groups at the same time point (P>0.05). Compared with the blank group, the expression of SP positive cells was increased in the 8-hour subgroup of sham-embedding group, 8-hour, 3-day, 7-day, and 14-day subgroups of catgut group, and 3-day, 7-day, and 14-day subgroups of PGLA group (P<0.001, P<0.05). The expression of SP positive cells in the 7-day subgroup of catgut group was higher than that in the 8-hour subgroup of catgut group (P<0.05), while the expression of SP positive cells in the 14-day subgroup of catgut group was lower than that in the 7-day subgroup of catgut group (P<0.001). The expression of SP positive cells in the 7-day subgroup of PGLA group was higher than that in the 3-day subgroup of PGLA group (P<0.05), while the expression of SP positive cells in the 14-day subgroup of PGLA group was lower than that in the 7-day subgroup of PGLA group (P<0.01). There was no significant difference in the expression of SP positive cells between the subgroups of the two embedding groups at the same time point (P>0.05). Compared with the blank group, the expression of HA positive cells was increased in the 8-hour, 3-day subgroups of sham-embedding group, 8-hour, 3-day, 7-day, and 14-day subgroups of catgut group, and 8-hour, 3-day, 7-day, 14-day, and 21-day subgroups of PGLA group (P<0.001, P<0.01, P<0.05). The expression of HA positive cells in the 14-day subgroup of catgut group was lower than that in the 7-day subgroup of catgut group (P<0.05), while the expression of HA positive cells in the 3-day subgroup of PGLA group was higher than that in the 8-hour subgroup of PGLA group (P<0.05), and the expression of HA positive cells in the 14-day subgroup of PGLA group was lower than that in the 7-day subgroup of PGLA group (P<0.05). The expression of HA positive cells in the 3-day subgroup of PGLA group was higher than that in the 3-day subgroup of catgut group (P<0.05). CONCLUSION: Catgut and PGLA embedding at "Zusanli" (ST 36) in healthy rats could induce changes in local skin MC, SP, and HA, which may be one of the mechanisms of the temporal stimulation effect after acupoint embedding. There are certain differences between different suture materials. A moderate inflammatory response in the acupoint area, mediated by MC and involving SP and HA, may be one of the initiating factors for the effect of acupoint catgut embedding.

Involvement of Substance P (SP) and Its Related NK1 Receptor in Primary Sjögren's Syndrome (pSS) Pathogenesis.

Primary Sjögren's Syndrome (pSS) is a systemic autoimmune disease that primarily attacks the lacrimal and salivary glands, resulting in impaired secretory function characterized by xerostomia and xerophthalmia. Patients with pSS have been shown to have impaired salivary gland innervation and altered circulating levels of neuropeptides thought to be a cause of decreased salivation, including substance P (SP). Using Western blot analysis and immunofluorescence studies, we examined the expression levels of SP and its preferred G protein-coupled TK Receptor 1 (NK1R) and apoptosis markers in biopsies of the minor salivary gland (MSG) from pSS patients compared with patients with idiopathic sicca syndrome. We confirmed a quantitative decrease in the amount of SP in the MSG of pSS patients and demonstrated a significant increase in NK1R levels compared with sicca subjects, indicating the involvement of SP fibers and NK1R in the impaired salivary secretion observed in pSS patients. Moreover, the increase in apoptosis (PARP-1 cleavage) in pSS patients was shown to be related to JNK phosphorylation. Since there is no satisfactory therapy for the treatment of secretory hypofunction in pSS patients, the SP pathway may be a new potential diagnostic tool or therapeutic target.

Gut microbiota modulates visceral sensitivity through calcitonin gene-related peptide (CGRP) production.

Abdominal pain is common in patients with gastrointestinal disorders, but its pathophysiology is unclear, in part due to poor understanding of basic mechanisms underlying visceral sensitivity. Accumulating evidence suggests that gut microbiota is an important determinant of visceral sensitivity. Clinical and basic research studies also show that sex plays a role in pain perception, although the precise pathways are not elucidated. We investigated pain responses in germ-free and conventionally raised mice of both sexes, and assessed visceral sensitivity to colorectal distension, neuronal excitability of dorsal root ganglia (DRG) neurons and the production of substance P and calcitonin gene-related peptide (CGRP) in response to capsaicin or a mixture of G-protein coupled receptor (GPCR) agonists. Germ-free mice displayed greater in vivo responses to colonic distention than conventional mice, with no differences between males and females. Pretreatment with intracolonic capsaicin or GPCR agonists increased responses in conventional, but not in germ-free mice. In DRG neurons, gut microbiota and sex had no effect on neuronal activation by capsaicin or GPCR agonists. While stimulated production of substance P by DRG neurons was similar in germ-free and conventional mice, with no additional effect of sex, the CGRP production was higher in germ-free mice, mainly in females. Absence of gut microbiota increases visceral sensitivity to colorectal distention in both male and female mice. This is, at least in part, due to increased production of CGRP by DRG neurons, which is mainly evident in female mice. However, central mechanisms are also likely involved in this process.

[Effects of moxibustion on serum levels of ß-EP, SP and expression of IL-1ß and COX-2 protein in brainstem in rats with migraine].

OBJECTIVE: To observe the effects of moxibustion at "Baihui" (GV 20) and "Dazhui" (GV 14) at different time points on the serum level of ß-endorphin (ß-EP), substance P (SP) and expression of interleukin-1ß (IL-1ß) and cyclooxygenase-2 (COX-2) protein in brainstem in rats with migraine, and to explore the effect and mechanism of moxibustion in preventing and treating migraine. METHODS: Forty male SD rats were randomly divided into a blank group, a model group, a prevention+treatment (PT) group and a treatment group, 10 rats in each group. Except the blank group, the rats in the remaining groups were injected with nitroglycerin subcutaneously to prepare migraine model. The rats in the PT group were treated with moxibustion 7 days before modeling (once a day) and 30 min after modeling, while the rats in the treatment group were treated with moxibustion 30 min after modeling. The "Baihui" (GV 20) and "Dazhui" (GV 14) were taken for 30 minutes each time. The behavioral scores in each group were observed before and after modeling. After intervention, ELISA method was used to detect the serum level of ß-EP and SP; the immunohistochemistry method was used to detect the number of positive cells of IL-1ß in brainstem; the Western blot method was used to detect the expression of COX-2 protein in brainstem. RESULTS: Compared with the blank group, the behavioral scores in the model group were increased 0-30 min, 60-90 min and 90-120 min after modeling (P<0.01); compared with the model group, in the treatment group and the PT group, the behavioral scores were decreased 60-90 min and 90-120 min after modeling (P<0.01). Compared with the blank group, in the model group, the serum level of ß-EP was decreased (P<0.01), while the serum level of SP, the number of positive cells of IL-1ß in brainstem and the expression of COX-2 protein were increased (P<0.01). Compared with the model group, in the PT group and and the treatment group, the serum level of ß-EP was increased (P<0.01), while the serum level of SP, the number of positive cells of IL-1ß and the expression of COX-2 protein in brainstem were decreased (P<0.01, P<0.05). Compared with the treatment group, in the PT group, the serum level of ß-EP was increased and COX-2 protein expression was decreased (P<0.05). CONCLUSION: Moxibustion could effectively relieve migraine. The mechanism may be related to reduce the serum level of SP, IL-1ß and COX-2 protein expression in brainstem, and increase the serum level of ß-EP, and the optimal effect is observed in the PT group.

Effects of catgut embedding and PGLA embedding at "Zusanli" (ST 36) on skin mast cells, substance P and histamine in healthy rats / 中国针灸

OBJECTIVE@#To observe the effects of catgut embedding and polyglycolic acid/poly-lactic acid (PGLA) embedding at "Zusanli" (ST 36) on the activation of local skin mast cells (MC), and expression of substance P (SP) and histamine (HA), and to explore the mechanism of the temporal stimulation effect of acupoint catgut embedding and provide a foundation for further research on the initiation mechanism of acupoint catgut embedding.@*METHODS@#One hundred and sixty male SPF-grade SD rats were randomly divided into a blank group (10 rats), a sham-embedding group (50 rats), a catgut group (50 rats), and a PGLA group (50 rats). Each intervention group was further randomly divided into five subgroups according to the time points after intervention: 8 hours, 3 days, 7 days, 14 days, and 21 days, with 10 rats in each subgroup. One-time sham-embedding, catgut embedding and PGLA embedding was given at left "Zusanli" (ST 36) in each intervention group, respectively. The skin and subcutaneous connective tissue of the left "Zusanli" (ST 36) were collected at the corresponding time points after intervention, except for the blank group (only one day before intervention). Toluidine blue staining was used to detect MC count and degranulation, and immunohistochemical staining was used to detect the expression of SP and HA positive cells.@*RESULTS@#There was no significant difference in MC count between the subgroups of each intervention group and the blank group (P>0.05). There was no significant difference in MC count between the subgroups of the catgut group and the PGLA group (P>0.05). The MC count in the 8-hour subgroup of PGLA group was higher than that in the 8-hour subgroup of catgut group (P<0.05), while the MC count in the 21-day subgroup of PGLA group was lower than that in the 21-day subgroup of catgut group (P<0.05). Compared with the blank group, the degranulation rates of MC were increased in the 8-hour and 3-day subgroups of sham-embedding group, 8-hour, 3-day, and 7-day subgroups of catgut group, and 8-hour, 3-day, 7-day, and 14-day subgroups of PGLA group (P<0.01, P<0.05, P<0.001). There was no significant difference in the degranulation rate of MC between the subgroups of the catgut group and the PGLA group (P>0.05), and no significant difference in the degranulation rate of MC between the two embedding groups at the same time point (P>0.05). Compared with the blank group, the expression of SP positive cells was increased in the 8-hour subgroup of sham-embedding group, 8-hour, 3-day, 7-day, and 14-day subgroups of catgut group, and 3-day, 7-day, and 14-day subgroups of PGLA group (P<0.001, P<0.05). The expression of SP positive cells in the 7-day subgroup of catgut group was higher than that in the 8-hour subgroup of catgut group (P<0.05), while the expression of SP positive cells in the 14-day subgroup of catgut group was lower than that in the 7-day subgroup of catgut group (P<0.001). The expression of SP positive cells in the 7-day subgroup of PGLA group was higher than that in the 3-day subgroup of PGLA group (P<0.05), while the expression of SP positive cells in the 14-day subgroup of PGLA group was lower than that in the 7-day subgroup of PGLA group (P<0.01). There was no significant difference in the expression of SP positive cells between the subgroups of the two embedding groups at the same time point (P>0.05). Compared with the blank group, the expression of HA positive cells was increased in the 8-hour, 3-day subgroups of sham-embedding group, 8-hour, 3-day, 7-day, and 14-day subgroups of catgut group, and 8-hour, 3-day, 7-day, 14-day, and 21-day subgroups of PGLA group (P<0.001, P<0.01, P<0.05). The expression of HA positive cells in the 14-day subgroup of catgut group was lower than that in the 7-day subgroup of catgut group (P<0.05), while the expression of HA positive cells in the 3-day subgroup of PGLA group was higher than that in the 8-hour subgroup of PGLA group (P<0.05), and the expression of HA positive cells in the 14-day subgroup of PGLA group was lower than that in the 7-day subgroup of PGLA group (P<0.05). The expression of HA positive cells in the 3-day subgroup of PGLA group was higher than that in the 3-day subgroup of catgut group (P<0.05).@*CONCLUSION@#Catgut and PGLA embedding at "Zusanli" (ST 36) in healthy rats could induce changes in local skin MC, SP, and HA, which may be one of the mechanisms of the temporal stimulation effect after acupoint embedding. There are certain differences between different suture materials. A moderate inflammatory response in the acupoint area, mediated by MC and involving SP and HA, may be one of the initiating factors for the effect of acupoint catgut embedding.

Effect of Magnolia officinalis Cortex Before and After Sweating on Distribution of Intestinal Flora and Levels of 5-HT and SP in IBS-C Rats / 中国实验方剂学杂志

ObjectiveTo investigate the mechanism of Magnolia officinalis cortex for constipation-type irritable bowel syndrome（IBS-C） rats before and after sweating. MethodIBS-C rat model was established by gavage of ice water， and rats were randomly divided into the blank group， model group， mosapride group（1 mg·kg-1）， M. officinalis cortex group（10 g·kg-1） and sweated M. officinalis cortex group（10 g·kg-1）. The changes of body weight， fecal number and fecal water content of rats were observed， 16S rRNA sequencing was used to detect the structural changes of fecal intestinal flora in rats， the levels of 5-hydroxytryptamine（5-HT） and substance P（SP） in colonic tissues of rats were determined by enzyme-linked immunosorbent assay（ELISA）. ResultCompared with the model group， the fecal water content and fecal number of mosapride group， M. officinalis cortex group and sweated M. officinalis cortex group were significantly increased（P<0.05）. At the phylum level， the top four species of flora abundance were Firmicutes， Bacteroidetes， Spirochaetes and Proteobacteria. Compared with the blank group， the proportion of Firmicutes in the model group was significantly reduced（P<0.05）， while the proportion of Spirochaetes was significantly increased（P<0.05）. Compared with the model group， the proportion of Firmicutes and Spirochaetes in M. officinalis cortex group and sweated M. officinalis cortex group tended to be similar to that in the blank group， and the proportion of Spirochaetes in sweated M. officinalis cortex group was lower than that of M. officinalis cortex group. At the family level， the top four species of flora abundance were Lactobacillaceae， S24_7， Ruminococcaceae， Bacteroidaceae， compared with the blank group， the proportion of Lactobacillaceae in the model group decreased significantly（P<0.05）， and its proportion in the M. officinalis cortex group and sweated M. officinalis cortex group increased significantly after administration（P<0.05）， and the flora structure of the two groups tended to be similar to that of the blank group. At the genus level， the top four species of flora abundance were Lactobacillus， Unspecified_S24_7， Bacteroides and Treponema. Compared with the blank group， the proportion of Lactobacillus in the model group decreased significantly（P<0.05）， while the proportion of Treponema increased significantly（P<0.05）. Compared with the model group， ratio of bacterial structure of Lactobacillus and Treponema in the M. officinalis cortex group and sweated M. officinalis cortex group tended to be similar to those in the blank group， indicating that M. officinalis cortex could restore the intestinal microbial structure of IBS-C rats before and after sweating. Compared with the model group， the 5-HT content in mosapride group was significantly reduced（P<0.05）， the contents of 5-HT and SP in the M. officinalis cortex group and sweated M. officinalis cortex group were significantly increased（P<0.01）， and the sweated M. officinalis cortex group was higher than the M. officinalis cortex group. ConclusionM. officinalis cortex can play a therapeutic role on IBS-C rats by regulating 5-HT pathway and intestinal flora structure before and after sweating.

Effects of moxibustion on serum levels of β-EP, SP and expression of IL-1β and COX-2 protein in brainstem in rats with migraine / 中国针灸

OBJECTIVE@#To observe the effects of moxibustion at "Baihui" (GV 20) and "Dazhui" (GV 14) at different time points on the serum level of β-endorphin (β-EP), substance P (SP) and expression of interleukin-1β (IL-1β) and cyclooxygenase-2 (COX-2) protein in brainstem in rats with migraine, and to explore the effect and mechanism of moxibustion in preventing and treating migraine.@*METHODS@#Forty male SD rats were randomly divided into a blank group, a model group, a prevention+treatment (PT) group and a treatment group, 10 rats in each group. Except the blank group, the rats in the remaining groups were injected with nitroglycerin subcutaneously to prepare migraine model. The rats in the PT group were treated with moxibustion 7 days before modeling (once a day) and 30 min after modeling, while the rats in the treatment group were treated with moxibustion 30 min after modeling. The "Baihui" (GV 20) and "Dazhui" (GV 14) were taken for 30 minutes each time. The behavioral scores in each group were observed before and after modeling. After intervention, ELISA method was used to detect the serum level of β-EP and SP; the immunohistochemistry method was used to detect the number of positive cells of IL-1β in brainstem; the Western blot method was used to detect the expression of COX-2 protein in brainstem.@*RESULTS@#Compared with the blank group, the behavioral scores in the model group were increased 0-30 min, 60-90 min and 90-120 min after modeling (P<0.01); compared with the model group, in the treatment group and the PT group, the behavioral scores were decreased 60-90 min and 90-120 min after modeling (P<0.01). Compared with the blank group, in the model group, the serum level of β-EP was decreased (P<0.01), while the serum level of SP, the number of positive cells of IL-1β in brainstem and the expression of COX-2 protein were increased (P<0.01). Compared with the model group, in the PT group and and the treatment group, the serum level of β-EP was increased (P<0.01), while the serum level of SP, the number of positive cells of IL-1β and the expression of COX-2 protein in brainstem were decreased (P<0.01, P<0.05). Compared with the treatment group, in the PT group, the serum level of β-EP was increased and COX-2 protein expression was decreased (P<0.05).@*CONCLUSION@#Moxibustion could effectively relieve migraine. The mechanism may be related to reduce the serum level of SP, IL-1β and COX-2 protein expression in brainstem, and increase the serum level of β-EP, and the optimal effect is observed in the PT group.

The amino-terminal heptapeptide of the algesic substance P provides analgesic effect in relieving chronic neuropathic pain.

Of painful conditions, somatic pain of acute nociceptive origin can be effectively managed clinically, while neuropathic pain of chronic neuropathy origin is difficult to control. For molecules involved in pain sensation, substance P (SP) is algesic, exacerbating painful sensation, while its amino-terminal fragment, heptapeptide SP(1-7), confers biological activities different from its full-length parent neuropeptide precursor. We previously demonstrated SP(1-7) interaction with pain processing to alleviate chronic pain. Here we evaluated SP(1-7) and its C-terminal amidated analogue SP(1-7)amide, together with SP and opioid agonist DAMGO. We tested mouse behaviors of both acute somatic pain in tail-flick latency assay, and neuropathic pain in sciatic nerve injury model of chronic constriction injury (CCI). DAMGO produced dose-dependent analgesia for somatic pain as expected, so did both SP(1-7) and its analogue SP(1-7)amide, while SP yielded the opposite effect of algesia, in a phenomenon we termed 'contrintus', meaning 'opposite from within' to denote that two peptides of the same origin (SP and its metabolic fragment SP(1-7)) produced opposite effects. In CCI model, DAMGO showed a general reduction in allodynia sensitivity for both nerve-injured and normal paws, without selective effect for neuropathic pain, consistent with clinical observation that opioids are less effective for chronic neuropathic pain. On the other hand, both SP(1-7) and SP(1-7)amide displayed dose-dependent anti-allodynia effect that is selective for neuropathic pain. These findings suggest that SP(1-7) and its analogue may be useful for developing pharmaceuticals to treat neuropathic pain.

The effects of two different intensities of aerobic training protocols on pain and serum neuro-biomarkers in women migraineurs: a randomized controlled trail.

OBJECTIVES: We have a weak understanding of how aerobic training may influence migraine, and the optimal parameters for exercise regimens as migraine therapy are not clear. The objectives of this study were to assess, first, effects of two different intensities of aerobic exercise on migraine headache indices; second, serum neuro-biomarker in women migraineurs. METHODS: A total of 45 non-athlete female migraine patients were selected by a neurologist and randomly divided into three groups: control (CON), moderate-intensity aerobic training (MOD T), and high-intensity aerobic training (HIGH T). Before and after the training protocol, body composition factors, migraine pain indices, VO2max, and serum Adenylate-Cyclase Activating Polypeptide (PACAP) and Substance P (SP) were measured. Exercise training protocol includes two different intensities of aerobic exercise: Moderate (13-15 Borg Scale, 60-80% HRmax) and High (15-17 Borg Scale, 65-95% HRmax). RESULTS: Moderate-intensity aerobic training (MOD T) reduced headache intensity, frequency, and duration in women with migraine (p < 0.001, for all). Also, high-intensity aerobic training (HIGH T) reduced headache intensity, frequency, and duration (p < 0.001, for all). However, for headache intensity and duration, MOD T was effective rather than HIGH T (p < 0.001; p ≤ 0.05, respectively). In addition, neither MOD T nor HIGH T could not alter PACAP and SP contents (p = 0.712; p = 0.249, respectively). CONCLUSIONS: Our results demonstrated that either MOD T or HIGH T could modify migraine pain indices but neither MOD T nor HIGH T could not alter the PACAP and SP contents in women with migraine.

Inhibitors of neuropeptide peptidases engaged in pain and drug dependence.

Owing to a broad spectrum of functions performed by neuropeptides, this class of signaling molecules attracts an increasing interest. One of the key steps in the regulation of biological activity of neuropeptides is proteolytic conversion or degradation by proteinases that change or terminate biological activity of native peptides. These enzymes, in turn, are regulated by inhibitors, which play integral role in controlling many metabolic pathways. Thus, the search for selective inhibitors and detailed knowledge on the mechanisms of binding of these substances to enzymes, could be of importance for designing new pharmacological approaches. The aim of this review is to summarize the current knowledge on the inhibitors of enzymes that convert selected groups of neuropeptides, such as dynorphins, enkephalins, substance P and NPFF fragments. The importance of these substances in pathophysiological processes involved in pain and drug addiction, have been discussed. This article is part of the special issue on Neuropeptides.

Targeting Neuropathic Pain: Pathobiology, Current Treatment and Peptidomimetics as a New Therapeutic Opportunity.

There is a huge need for pharmaceutical agents for the treatment of chronic Neuropathic Pain (NP), a complex condition where patients can suffer from either hyperalgesia or allodynia originating from central or peripheral nerve injuries. To date, the therapeutic guidelines include the use of tricyclic antidepressants, serotonin-noradrenaline reuptake inhibitors and anticonvulsants, beside the use of natural compounds and non-pharmacological options. Unfortunately, these drugs suffer from limited efficacy and serious dose-dependent adverse effects. In the last decades, the heptapeptide SP1-7, the major bioactive metabolite produced by Substance P (SP) cleavage, has been extensively investigated as a potential target for the development of novel peptidomimetic molecules to treat NP. Although the physiological effects of this SP fragment have been studied in detail, the mechanism behind its action is not fully clarified and the target for SP1-7 has not been identified yet. Nevertheless, specific binding sites for the heptapeptide have been found in brain and spinal cord of both mouse and rats. Several Structure-Affinity Relationship (SAR) studies on SP1-7 and some of its synthetic analogues have been carried out aiming to developing more metabolically stable and effective small molecule SP1-7-related amides that could be used as research tools for a better understanding of the SP1-7 system and, in a longer perspective, as potential therapeutic agents for future treatment of NP.

Intraarticular injection of processed lipoaspirate cells has anti-inflammatory and analgesic effects but does not improve degenerative changes in murine monoiodoacetate-induced osteoarthritis.

BACKGROUND: Previous basic research and clinical studies examined the effects of mesenchymal stem cells (MSCs) on regeneration and maintenance of articular cartilage. However, our pilot study suggested that MSCs are more effective at suppressing inflammation and pain rather than promoting cartilage regeneration in osteoarthritis. Adipose tissue is considered a useful source of MSCs; it can be harvested easily in larger quantities compared with the bone marrow. The present study was designed to evaluate the anti-inflammatory, analgesic, and regenerative effects of intra-articularly injected processed lipoaspirate (PLA) cells (containing adipose-derived MSCs) on degenerative cartilage in a rat osteoarthritis model. METHODS: PLA cells were isolated from subcutaneous adipose tissue of 12-week-old female Sprague-Dawley rats. Osteoarthritis was induced by injection of monoiodoacetate (MIA). Each rat received 1 × 106 MSCs into the joint at day 7 (early injection group) and day 14 (late injection group) post-MIA injection. At 7, 14, 21 days after MIA administration, pain was assessed by immunostaining and western blotting of dorsal root ganglion (DRG). Cartilage quality was assessed macroscopically and by safranin-O and H&E staining, and joint inflammation was assessed by western blotting of the synovium. RESULTS: The early injection group showed less cartilage degradation, whereas the late injection group showed cartilage damage similar to untreated OA group. The relative expression level of CGRP protein in DRG neurons was significantly lower in the two treatment groups, compared with the untreated group. CONCLUSIONS: Intra-articular injection of PLA cells prevented degenerative changes in the early injection group, but had little effect in promoting cartilage repair in the late injection group. Interestingly, intra-articular injection of PLA cells resulted in suppression of inflammation and pain in both OA groups. Further studies are needed to determine the long-term effects of intra-articular injection of PLA cells in osteoarthritis.

Subclinical lipopolysaccharide from Salmonella Enteritidis induces neuropeptide dysregulation in the spinal cord and the dorsal root ganglia.

BACKGROUND: Despite increasing evidence that lipopolysaccharide (LPS) affects the biological active substances of dorsal root ganglia (DRG) we have limited knowledge of the influence of a single low dose of LPS, which does not result in any clinical symptoms of disease (subclinical LPS) on neuropeptides connected with the sensory pathway. Accordingly, in this work, we investigated the influence of subclinical LPS from Salmonella Enteritidis on selected neuropeptides: substance P (SP), galanin (GAL), neuropeptide Y (NPY), vasoactive intestinal peptide (VIP) and somatostatin (SOM) in the cervical, thoracic, lumbar and sacral regions of the DRG and spinal cord. METHODS: This study was performed on immature female pigs of the Pietrain × Duroc breed. Seven days after the intravenous injection of saline solution for control animals (n = 5) and 5 µg/kg b.w. LPS from S. Enteritidis for the experimental group (n = 5), the DRG and the spinal cord were collected to extract the neuropeptides using solid-phase extraction technology. RESULTS: Our results demonstrated that subclinical LPS in DRG was able to change the levels of all studied neuropeptides except SOM, whereas in the spinal cord it down-regulated all studied neuropeptides in the sacral spinal cord, maintaining the concentration of all studied neuropeptides in other regions similar to that observed in the control animals. The significant differences in the intensity and character of observed changes between particular regions of the DRG suggest that the exact functions of the studied neuropeptides and mechanisms of responses to subclinical LPS action depend on specific characteristics and functions of each examination region of DRG. CONCLUSIONS: The mechanisms of observed changes are not fully understood and require further study of the molecular interactions between subclinical LPS from S. Enteritidis and neuronal and non-neuronal cells of DRG and spinal cord. The peripheral and central pain pathways must be analysed with the aspect of unknown long-term consequences of the influence of subclinical LPS from S. Enteritidis on neuropeptides in the spinal cord and the dorsal root ganglia.

NF-κB-Associated Pain-Related Neuropeptide Expression in Patients with Degenerative Disc Disease.

The role of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) has been highlighted in mechanisms underlying inflammatory and neuropathic pain processes. The present study was designed to investigate whether NF-κB signaling is associated with pain-related neuropeptide expression in patients with chronic back pain related to degenerative disc disease (DDD). Intervertebral disc (IVD) tissues were collected from forty DDD patients undergoing disc replacement or fusion surgery, and from eighteen postmortem (PM) control subjects. RELA, NFKB1, CGRP, TAC1, TRPV1, and MMP-3 gene expression were analyzed by RT-qPCR, while NF-κB subunit RelA and NF-κB1⁻DNA binding in nuclear extracts and calcitonin gene related peptide (CGRP), substance P (SP), and transient receptor potential, subfamily V, member 1 (TRPV1) protein levels in cytosolic extracts of tissues were assessed by enzyme-linked immunosorbent assay (ELISA). An upregulated NF-κB1⁻DNA binding, and higher CGRP and TRPV1 protein levels were observed in DDD patients compared to PM controls. In DDD patients, NF-κB1⁻DNA binding was positively correlated with nuclear RelA levels. Moreover, NF-κB1⁻DNA binding was positively associated with TRPV1 and MMP-3 gene and SP and TRPV1 protein expression in DDD patients. Our results indicate that the expression of SP and TRPV1 in IVD tissues was associated with NF-κB activation. Moreover, NF-κB may be involved in the generation or maintenance of peripheral pain mechanisms by the regulation of pain-related neuropeptide expression in DDD patients.

Neuropathic Pain and Itch Mechanisms Underlying Allergic Conjunctivitis.

OBJECTIVE: Among the constellation of symptoms that characterizes allergic conjunctivitis, many (eg, burning and stinging) can be attributed to chronic neuropathic pain. Cumulative data support that these hallmark symptoms might be linked to the effects of allergen-induced neuromodulation. This review investigates the key characteristics of neuropathic itch and pain in allergic conjunctivitis and their underlying pathogenic mechanisms. METHODS: A literature review was conducted using a PubMed search focusing on allergic conjunctivitis, neurogenic inflammation, neuropathic itch, and neuropathic pain. Articles were reviewed, and those discussing clinical course, pathophysiology, and neuronal regulation of chronic neuropathic symptoms as related to allergic disease were summarized. RESULTS: Recent evidence suggests that some symptoms of allergic conjunctivitis may be better represented as a chronic neuropathic disorder. We found that neurogenic mechanisms may have a significant role in chronic ocular surface inflammation from allergic inflammation. Manifestations may be associated with repeated ocular sensory nerve injury leading to an acute-to-chronic transition, which is in turn associated with neuropathologic changes (peripheral and central sensitization), neuronal dysfunction, and spontaneous ocular pain. CONCLUSION: Current goals in the management of allergic conjunctivitis aim to minimize the inflammatory cascade associated with the allergic response in the initial stages of the pathogenic mechanism. Based on the mechanistic data reviewed herein, the recognition that neuronal inflammation explains many of the symptoms in allergic conjunctivitis opens new frontiers for drug discovery.

Human acute myeloid leukemia cells express Neurokinin-1 receptor, which is involved in the antileukemic effect of Neurokinin-1 receptor antagonists.

The substance P/neurokinin-1 receptor system has been implicated in tumor cell proliferation. Neurokinin-1 receptor has been identified in different solid tumors but not frequently in hematopoietic malignant cells. We investigated the presence of the Neurokinin-1 receptor in acute myeloid leukemia cell lines (KG-1 and HL-60), demonstrating that acute myeloid leukemia cell lines overexpress the truncated Neurokinin-1 receptor isoform compared with lymphocytes from healthy donors. Using the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) method, we demonstrated that substance P induced cell proliferation in both acute myeloid leukemia cell lines. We also observed that four different Neurokinin-1 receptor antagonists (L-733,060, L-732,138, CP 96-345 and aprepitant) elicited inhibition of acute myeloid leukemia cell growth lines in a concentration-dependent manner, while growth inhibition was only marginal in lymphocytes; the specific antitumor action of Neurokinin-1 receptor antagonists occurs via the Neurokinin-1 receptor, and leukemia cell death is due to apoptosis. Finally, administration of high doses of daily intraperitoneal fosaprepitant to NOD scid gamma mice previously xenografted with the HL60 cell line increased the median survival from 4 days (control group) to 7 days (treated group) (p = 0.059). Taken together, these findings suggest that Neurokinin-1 receptor antagonists suppress leukemic cell growth and may be considered to be potential antitumor drugs for the treatment of human acute myeloid leukemia.

Activation of orexin-1 receptors in the ventrolateral periaqueductal grey matter (vlPAG) modulates pulpal nociception and the induction of substance P in vlPAG and trigeminal nucleus caudalis.

AIM: To characterize the role of orexin-1 receptors (OX1Rs) in ventrolateral periaqueductal grey matter (vlPAG) on modulation of capsaicin-induced pulpal nociception in rats. METHODOLOGY: Sixty-six adult male Wistar rats (2 months old) weighing between 230 and 260 g were used. The animals were cannulated for microinjection of drugs into the vlPAG matter. Pulpalgia was induced by intradental application of capsaicin solution (100 µg) into the incisor teeth of the rats. Ten min prior to capsaicin application, orexin-A (50, 100 and 150 pmol L-1 per rat) was administered. Orexin-A (150 pmol L-1 ) was also co-administrated with SB-334867 (40 nmol L-1 per rat), an OX1Rs antagonist; or bicuculline (1 µg per rat), a GABAA receptors antagonist. Moreover, treatment effects on the release of pro-nociceptive modulator substance P (SP) in vlPAG and trigeminal nucleus caudalis (Vc) of rats were explored using an immunofluorescence technique. One-way analysis of variance was used for the statistical analysis. RESULTS: Orexin-A dose-dependently decreased capsaicin-induced nociceptive behaviour. However, SB-334867 (40 nmol L-1 per rat) pretreatment (P < 0.05), but not bicuculline (1 µg per rat), attenuated the analgesic effect of orexin-A (150 pmol L-1 ). The level of SP was significantly increased in Vc and decreased in vlPAG of capsaicin-treated rats (P < 0.05). Capsaicin-induced changes in SP levels, however, were prohibited by orexin-A treatment (150 pmol L-1 ) (P < 0.05). CONCLUSIONS: Orexin-A administration into the vlPAG was associated with an inhibitory effect on capsaicin-induced pulpal nociception and bidirectional effects on the induction of SP in vlPAG and Vc of rats. Central activation of OX1Rs is a potential therapeutic tool for pulpalgia.

[Effect of transcutaneous electrical acupoint stimulation on postoperative analgesia of ureteroscopic holmium laser lithotripsy].

OBJECTIVE: To explore the effect of transcutaneous electrical acupoint stimulation (TEAS) on postoperative analgesia of ureteroscopic holmium laser lithotripsy. METHODS: One hundred and twenty adult patients, American Association of Anesthesiologists (ASA) Class Ⅰ or Ⅱ, scheduled to ureteroscopic holmium laser lithotripsy, were randomly assigned into an observation group and a control group, 60 cases in each one. The patients in the observation group were treated with TEAS for postoperative analgesia. TEAS was implemented at bilateral Shenshu (BL 23) and Yinlingquan (SP 9) at the time of back ward and postoperative 4 h, 8 h, 12 h. TEAS at 7:00, 11:00 and 15:00 at the above acupoints were used on the second and third days; while placebo (twice a day, 100 mg a time) was used. Tramadol hydrochloride tablets for postoperative analgesia were applied in the contnol group, twice a day, 100 mg a time, and electrode sheets without stimulation were put on Shenshu (BL 23) and Yinlingquan (SP 9). When analgesia was insufficient with the score of visual analogue scale (VAS)≥3, the patients were treated with tramadol tablets for remedy analgesia. The VAS score, the concentrations of serotonin (5-HT) and substance P (SP) in 3 mL venous blood at the time of back ward (T0), postoperative 4 h (T1), 12 h (T2), 24 h (T3), and 48 h (T4) were detected respectively. The total amount of medication for remedy analgesia and the incidence of adverse reactions, such as nausea and vomiting within postoperative 48 h were compared between the two groups. RESULTS: The VAS scores at T1 through T4 were lower than those at T0 in the two groups (all P<0.05). Compared with the control group, the VAS scores at T1 through T3 in the observation group were lower (all P<0.05). The total dose of remedy analgesic medicine within 48 h after operation in the observation group was lower than that in the control group (P<0.05). Compared with the control group, the concentrations of 5-HT at T1, T2, T4 and SP at T1 through T4 were lower (all P<0.05). The numbers of constipation, nausea and vomiting in the observation group were less than those in the control group (both P<0.05). CONCLUSION: TEAS can relieve the pain and reduce the total amount of analgesic medicine, the levels of substance causing pain and the incidence of adverse reactions after ureteroscopic holmium laser lithotripsy.

[Time-effect relationship and molecular mechanism of electroacupuncture for repair of gastric mucosal lesion].

OBJECTIVE: To dynamically observe the effects of electroacupuncture (EA) on repair of gastric mucosal lesion in rats with gastric ulcer, and to explore the time-effect relationship and molecular mechanism of EA for gastric ulcer. METHODS: A total of 72 SD rats were randomly assigned to a normal group, a model group, a acupoint group and a sham acupoint group, and each group were further divided into a 1-day subgroup, a 4-day subgroup and a 7-day subgroup, 6 rats in each subgroup. The rat model of gastric ulcer was established by using intragastric administration of ethyl alcohol. The rats in the acupoint group were treated with EA at"Zusanli"(ST 36) and"Liangmen"(ST 21); the rats in the sham acupoint group were treated with EA at points 5 mm next to"Zusanli"(ST 36) and"Liangmen"(ST 21); the EA was given 30 min per treatment, once a day. The rats in the normal group and model group were treated with immobilization for 30 min per day, and no EA was given. PR-PCR method was applied to test the expression of proliferating cell nuclear antigen (PCNA) and substance P (SP); Western blot method was applied to test the expression of neurotensin (NT). RESULTS: After 1-day treatment, the ulcer index in the model group was higher than that in the normal group (P<0.01), and the expression of PCNA, SP and NT was decreased (P<0.01, P<0.05); compared with the model group and sham acupoint group, the ulcer index was decreased in the acupoint group (both P<0.05), and the expression of PCNA and SP was up-regulated (all P<0.05) while that of NT was up-regulated (both P<0.01). After 4-day treatment, the ulcer index in the model group was reduced but still higher than that in the normal group (P<0.05), and the expression of PCNA, SP and NT was up-regulated and higher than that in the normal group (all P<0.01); the ulcer index in the acupoint group was similar to that in the normal group (P>0.05), and the expression of PCNA and SP was lower than that in the model group (P<0.01, P<0.05), and the expression of NT was not significantly different from that in the model group (P>0.05). After 7-day treatment, the differences of indexes above were not significant among the four groups (all P>0.05). CONCLUSION: EA at acupoints of stomach meridian has two-way regulation on PCNA and SP and improve the expression of NT in different pathological state of gastric ulcer, which could further improve the repair of gastric ulcer.