Substance P Research Articles

The beneficial effect of Sanhuang ointment and its active constituents on experimental hemorrhoids in rats

Ethnopharmacological relevanceSanhuang ointment (SHO) has been widely used in the traditional Chinese medical system for 1500 years and has efficacy in clearing away heat and dampness, reducing swelling, and alleviating pain. Hemorrhoids will damage the normal physiological function of the body, resulting in obstructed defecation, accompanied by massive hemorrhage and necrosis of tissues and cells, which is easy to breed bacteria and cause infection. SHO can promote lesion healing in hemorrhoid rats, but the pharmacological mechanism underlying this effect remains unknown. AimTo evaluate the effect of SHO on experimental hemorrhoids in rats induced by croton oil and glacial acetic acid. Materials and methodsIn this research, the effective components of SHO were analyzed in detail by High performance liquid chromatography (HPLC) and Liquid chromatography/mass spectrometry (LC/MS). Hemorrhoids were induced by 6% balsam and glacial acetic acid respectively in the anorectal region of rats. SHO was administered externally to the anorectal region of rats at doses of 185 mg/g (crude drug/ointment), 370 mg/g (crude drug/ointment) and 740 mg/g (crude drug/ointment) for 11 days. Mayinglong musk hemorrhoids ointment (1 g/kg) and Taining cream (1 g/kg) were used as reference anti hemorrhoids drugs. On the 11th day, hemorrhoids were evaluated by measuring the biochemical parameters of hemorrhoids in rats and the histology of anorectal tissues. ResultsUsing high performance liquid chromatography liquid chromatography mass spectrometry, 41 compounds, including phenylpropionic acids and alkaloids, were identified. the fingerprints of 18 common peaks were identified. In Hemorrhoids like rats, acetic acid induced inflammation was inhibited in a dose-dependent manner during SHO treatment. In addition, the detailed experimental results show that SHO can effectively improve hemorrhoids by inhibiting the production of inflammatory cytokines in serum, reversing the down-regulation of vanillin subtype 1 (TRPV1), calcitonin gene related peptide (CGRP) and substance P (SP) levels of pain related genes in anal tissues, and the up regulation of Vascular endothelial growth factor (VEGF) levels of vascular growth related genes. ConclusionThe results showed that SHO could alleviate the edema caused by the exudation of anorectal tissue fluid in rats by anti-inflammatory effect and reducing the Vascular permeability of rats. The study validates the traditional use of SHO in the treatment of hemorrhoids and demonstrates its anti-hemorrhoidal potential.

Abstract PO-088: TRPV1+ sensory neurons provide a tumor supportive environment through recruitment of MDSCs

Abstract Cancer neuroscience has become an increasing area of focus in cancer research as our understanding of the presence and function of intra-tumoral neurons continues to emerge, with studies indicating that tumor infiltrating neurons impact components of the tumor microenvironment promoting cancer initiation and progression. We have previously published that head and neck squamous cell carcinomas are innervated. We now focus on defining the contributions of intra-tumoral neurons to disease progression in HNSCC. Tumor implantation in a double transgenic mouse (TRPV1cre::DTAfl/wt) lacking the cancer-recruited TRPV1-expressing sensory neurons allows us to study the consequences of this neuronal loss on the local immune response. Importantly, comparison between wild type and TRPV1cre::DTAfl/wt mice demonstrated no significant immune population differences at baseline. Thus, wildtype and TRPV1cre::DTAfl/wt mice were orthotopically implanted with tumor and tumoral immune cell populations identified by flow cytometry. Cytokine and peptide concentrations were measured by ELISA. In vitro assays were preformed to molecularly define the neuronal signals and immune responses. Here, co-culture experiments of immortalized head and neck cancer cell lines and dorsal root ganglia (DRG) from wildtype mice were used to model tumor innervation in vitro. The media generated was applied to bone marrow derived (BMD) immune cells and the impact on immune cell phenotypes assayed by flow cytometry. Our orthotopic model demonstrates that the presence of HNSCC promotes a nerve-injury phenotype in ipsilateral trigeminal ganglion, demonstrated by increased expression of activating transcription factor 3 (ATF3). Utilizing a 12-color flow cytometry panel we show that tumors from TRPV1cre::DTAfl/wt mice harbor a significantly smaller population of myeloid derived suppressor cells (MDSCs) compared to those from wildtype animals. Further analysis demonstrates this population shift occurs primarily due to a decrease in the CD11b+ Ly6G+/Ly6Clo population. In vitro studies using BMD immune cells show that tumor cell-DRG co-culture conditioned media induces an expansion of the MDSC population compared to condition media from DRG or tumor cells alone. Additionally, transwell migration assays show that co-culture condition media also increases cellular migration compared to condition media from DRG or tumor cells alone. Moreover, ELISA analysis indicates that co-culture of tumor cells and DRG increases neuronal release of the neuropeptide, Substance P (SP), potentiating IL-6 release from tumor cells. In summary, we identified several mechanisms by which TRPV1-expressing intra-tumoral neurons promote MDSC infiltration at the tumor bed and promote disease progression. These mechanisms define targets for therapeutic intervention including SP and IL-6 signaling. Future studies will test the clinical utility of these targets. Together, our data suggests that intra-tumoral neurons contribute to cancer progression through generation of a tumor supportive microenvironment. Citation Format: Anthony C. Restaino, Maryam Ahmadi, Sebastien Talbot, Paola D. Vermeer. TRPV1+ sensory neurons provide a tumor supportive environment through recruitment of MDSCs [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Innovating through Basic, Clinical, and Translational Research; 2023 Jul 7-8; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2023;29(18_Suppl):Abstract nr PO-088.

Activation of vascular endothelial cells by synovial fibrosis promotes Netrin-1-induced sensory nerve sprouting and exacerbates pain sensitivity.

Synovial fibrosis is one of the most dominant histopathological changes in osteoarthritis of the knee (KOA), and activation of vascular endothelial cells in synovial fibrosis is both an important factor in mediating pain in KOA and a major contributor to the generation of pain signals. At the same time, angiogenesis and nerve fibres are more likely to underlie the pathology of pain induced by synovial fibrosis. In the present study, we established a co-culture model of human umbilical vein endothelial cells (HUVECs) with dorsal root ganglion (DRG) and detected tissue and cellular Netrin-1, vascular cell adhesion molecule-1 (VCAM-1), intercellular cell adhesion molecule-1 (ICAM-1), growth-associated protein-43 (GAP43), colorectal cancer deleted (DCC), uncoordinated 5 (UNC5), and the related expression of calcitonin gene-related peptide (CGRP), substance P (SP) and nerve growth factor (NGF) in supernatant by ELISA to investigate the intervention of vascular endothelial cell activation on sensory nerve sprouting exacerbating peripheral pain sensitivity and to investigate the effect of Netrin-1 from the perspective of Netrin-1 secretion to illustrate its effector mechanism.

C. difficile intoxicates neurons and pericytes to drive neurogenic inflammation.

Clostridioides difficile infection (CDI) is a major cause of healthcare-associated gastrointestinal infections1,2. The exaggerated colonic inflammation caused by C. difficile toxins such as toxin B (TcdB) damages tissues and promotes C. difficile colonization3-6, but how TcdB causes inflammation is unclear. Here we report that TcdB induces neurogenic inflammation by targeting gut-innervating afferent neurons and pericytes through receptors, including the Frizzled receptors (FZD1, FZD2 and FZD7) in neurons and chondroitin sulfate proteoglycan 4 (CSPG4)in pericytes. TcdB stimulates the secretion of the neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP) from neurons and pro-inflammatory cytokines from pericytes. Targeted delivery of the TcdB enzymatic domain, through fusion with a detoxified diphtheria toxin, into peptidergic sensory neurons that express exogeneous diphtheria toxin receptor (an approach we term toxogenetics) is sufficient to induce neurogenic inflammation and recapitulates major colonic histopathology associated with CDI. Conversely, mice lacking SP, CGRP or the SP receptor (neurokinin 1 receptor) show reduced pathology in both models of caecal TcdB injection and CDI. Blocking SP or CGRP signalling reduces tissue damage and C. difficile burden in mice infected with a standard C. difficile strain or with hypervirulent strains expressing the TcdB2 variant. Thus, targeting neurogenic inflammation provides a host-oriented therapeutic approach for treating CDI.

The Effect of Group Training or Spinal Orthosis on Quality of Life and Potential Plasma Markers of Pain in Older Women With Osteoporosis. A Randomized Controlled Trial

ObjectivePrimary purpose was to examine the effects of exercise and use of a spinal orthosis on quality of life (QoL). Secondary, to explore the effects of above-mentioned interventions on plasma levels of potential markers of pain: substance P (SP), calcitonin gene-related peptide (CGRP), and interleukin-6 (IL-6). DesignRandomized controlled trial. SettingCommunity-dwelling women in Stockholm. ParticipantsA total of 113 women aged 60-93 years suffering from back pain and self-reported osteoporosis (n=113). InterventionsThe randomized controlled trial was 3-armed: participation in an equipment exercise group, treatment with an activating spinal orthosis or controls. The intervention time was 6 months. Main Outcome Measure(s)QoL (QUALEFFO-41 and SF-36), plasma levels of SP, CGRP, and IL-6 measured at baseline and after 6 months in all 3 arms. ResultsNo improvement of QoL was found. Comparing change in mobility (QUALEFFO-41), the effect in least squares means was lower in the spinal orthosis group compared with controls. In the exercise group, the role emotional score (SF-36) deteriorated during the intervention. Effect size varied between 0.02 and 0.6. There was no change in the levels of CGRP or SP, while IL-6 levels were lower at 6 months in the spinal orthosis group compared with the other groups. At least 1 previous vertebral fracture was verified by X-ray in 46 women. ConclusionThe interventions showed none or negative effect on QoL, which was unexpected. The modest effect size may prompt a cautious interpretation. We found a lowering of IL-6 levels in the spinal orthosis group, but more studies are needed.

Methylcobalamin in Combination with Early Intervention of Low-Intensity Pulsed Ultrasound Potentiates Nerve Regeneration and Functional Recovery in a Rat Brachial Plexus Injury Model.

This study evaluated and compared the functional recovery and histopathological outcomes of treatment involving low-intensity pulsed ultrasound (LIPUS) and methylcobalamin (B12) on brachial plexus injury (BPI) in an experimental rat model. Three days after BPI, the rats were assigned to receive either LIPUS or methylcobalamin alone or in combination consecutively for 12 days. Serial changes in sensory and motor behavioral responses, as well as morphological and immunohistochemical changes for substance P (SP), ionized calcium-binding adapter molecule 1 (iba1), brain-derived neurotrophic factor (BDNF), and S100 were examined 28 days after BPI as the outcome measurements. Early intervention of LIPUS and methylcobalamin, whether alone or in combination, augmented the sensory and motor behavioral recovery as well as modulated SP and iba1 expression in spinal dorsal horns, BDNF, and S100 in the injured nerve. Moreover, the combined therapy with its synergistic effect gave the most beneficial effect in accelerating functional recovery. In view of the effective initiation of early recovery of sensory and motor functions, treatment with LIPUS and methylcobalamin in combination has a potential role in the clinical management of early-phase BPI.

A Biomimetic C-Terminal Extension Strategy for Photocaging Amidated Neuropeptides.

Photoactivatable neuropeptides offer a robust stimulus-response relationship that can drive mechanistic studies into the physiological mechanisms of neuropeptidergic transmission. The majority of neuropeptides contain a C-terminal amide, which offers a potentially general site for installation of a C-terminal caging group. Here, we report a biomimetic caging strategy in which the neuropeptide C-terminus is extended via a photocleavable amino acid to mimic the proneuropeptides found in large dense-core vesicles. We explored this approach with four prominent neuropeptides: gastrin-releasing peptide (GRP), oxytocin (OT), substance P (SP), and cholecystokinin (CCK). C-terminus extension greatly reduced the activity of all four peptides at heterologously expressed receptors. In cell type-specific electrophysiological recordings from acute brain slices, subsecond flashes of ultraviolet light produced rapidly activating membrane currents via activation of endogenous G protein-coupled receptors. Subsequent mechanistic studies with caged CCK revealed a role for extracellular proteases in shaping the temporal dynamics of CCK signaling, and a striking switch-like, cell-autonomous anti-opioid effect of transient CCK signaling in hippocampal parvalbumin interneurons. These results suggest that C-terminus extension with a photocleavable linker may be a general strategy for photocaging amidated neuropeptides and demonstrate how photocaged neuropeptides can provide mechanistic insights into neuropeptide signaling that are inaccessible using conventional approaches.

12. NEUROKININ A AND SUBSTANCE P SERUM CONCENTRATIONS CORRELATE WITH MOTILITY METRICS PREDISPOSING TO REFLUX, BUT NOT WITH ESOPHAGITIS PRESENCE IN GERD

Abstract Background Neurokinin A (NKA) and substance P (SP) may be responsible for airway hyper-reactivity and hypersensitization. Animal studies showed an increase of their blood concentration in response to acid installation into the esophagus, suggesting the presence of neurogenic inflammation in pathogenesis of asthma and chronic cough. Association between serum concentration of these cytokines and esophageal function testing parameters in patients with gastroesophageal reflux disease (GERD) is poorly studied. Aim of the study was to assess serum concentration of neurokinin A and substance P in patients with erosive esophagitis and non-erosive form of GERD and perform correlation analysis between concentrations of these substances and data of multichannel esophageal pH-impedance (MII-pH) and high-resolution esophageal manometry (HREM). Methods The data of examination of 88 subjects (31 with erosive esophagitis (EE); 57 with non-erosive form of GERD (NERD)) served as a source for the study. Diagnosis of GERD was established per Lyon criteria. Subjects with autoimmune disorders, receiving some medications able to affect esophageal motility were not eligible. Patients underwent HREM with the use of 36-channel solid-state catheters and MII-pH with 2-pH, 6-impedandce sensors. Serum concentrations of NKA and SP were measured with ELISA kits and biochemistry analyzer. Results Acid exposure time (AET), and number of high gastroesophageal refluxes were higher in EE compared to NERD, while number of refluxes and concentrations of NKA and SP were similar. Mean, minimal and residual lower esophageal sphincter resting pressures were lower in EE compared to NERD. There was direct correlation between serum concentration of SP and number of gastroesophageal refluxes (Spearman rank R = 0.26), AET (R = 0.25), and number of refluxes lasting >5 min (R = 0.35). NKA concentration correlated directly with number of refluxes >5 min long (R = 0.28), and inversely with levels of integrated relaxation pressure (R = –0.32), and contractile front velocity (R = –0.38). Conclusions In patients with GERD, serum concentrations of Neurokinin A and Substance P do not depend on the presence of esophagitis and probably are not caused by the influence of reflux content to sensory neurons of esophageal mucosa. Established correlations may support the involvement of Neurokinin A and Substance P in pathogenesis of gastroesophageal reflux disease.

[Efficacy and safety of high-voltage pulsed radiofrequency combined with pregabalin on severe thoracic postherpetic neuralgia].

Objective: To investigate the efficacy and safety of high-voltage pulse radiofrequency combined with pregabalin on severe thoracic postherpetic neuralgia (PHN). Methods: A total of 103 patients with PHN who were admitted to the Department of Pain Medicine of Henan Provincial People's Hospital from May 2020 to May 2022 were retrospectively selected, including 50 males and 53 females, and aged 40 to 79 (65.4±9.2) years. The patients were divided into two groups according to the treatment methods they received: the control group (n=51) and the study group (n=52). The patients in the control group were treated with oral pregabalin, and the patients in the study group received pregabalin plus high-voltage pulse radiofrequency therapy. The pain intensity and efficacy of the two groups were evaluated before treatment and 4 weeks after treatment. The pain intensity, the sleep quality and the efficacy of treatment was evaluated by visual analogue scale (VAS) score, Pittsburgh Sleep Quality Index (PSQI) score and nimodipine method, respectively. The levels of pain mediators including serum neuropeptide Y (NPY), prostaglandin E2 (PGE2), substance P (SP) and β-endorphin were measured. The differences of the above indicators and the incidence of adverse reactions were compared between the two groups. Results: The VAS scores of the study group and the control group before treatment were 7.94±0.76 and 8.20±0.81, and PSQI scores were 16.84±3.90 and 16.29±3.84, respectively, with no statistically significant differences (both P>0.05). After 4 weeks of treatment, the VAS scores of the two groups were 2.84±0.80 and 3.35±0.87, and PSQI scores were 6.78±1.90 and 7.98±2.40, respectively, and the VAS score and PSQI score in the study group were lower than those in the control group (both P<0.05). There were no significant differences of the serum levels of NPY, PGE2, SP and β-endorphin before treatment in the study group and control group (all P>0.05). After 4 weeks of treatment, the levels of NPY, PGE2, SP and β-Endorphin in the study group were (240.7±26.8) ng/L, (74.4±8.6) μg/L, (108.9±15.7) ng/L and (4.4±0.9) ng/L, which were lower than those in the control group [(268.1±29.4) ng/L, (79.7±8.3) μg/L, (115.2±16.2) ng/L, (5.2±1.3) ng/L, respectively], with statistically significant differences (all P<0.05). After treatment, 29 cases were cured, 16 cases were markedly effective and 6 cases were effective in the study group, while 16 cases, 24 cases and 8 cases were cured, markedly effective and effective in the control group, respectively. The overall efficacy of patients in the study group was better than that in the control group (Z=-2.32, P=0.018). The incidence of adverse reactions in the study group and control group was 11.5% (6/52) and 7.8% (4/51), respectively, with no statistically significant difference (χ2=0.40, P=0.527). Conclusion: High-voltage pulse radiofrequency combined with pregabalin can significantly improve the pain intensity and sleep quality of patients with severe thoracic PHN and reduce the levels of pain mediators, with a high safety profile.

Rehmannia alcohol extract inhibits neuropeptide secretion and alleviates osteoarthritis pain through cartilage protection

Osteoarthritis (OA) is a common joint disease characterized by chronic pain, and the perception of pain is closely associated with brain function and neuropeptide regulation. Rehmannia is common plant herb with anti-inflammatory and analgesic properties that is used to treat OA. However, it is unclear whether Rehmannia alleviates OA-related pain via regulation of neuropeptides and brain function. We examined the pain relief regulatory pathway in OA after treatment with Rehmannia by verifying the therapeutic effect of Rehmannia alcohol extract in vivo and vitro and exploring of the potential mechanism underlying the analgesic effect of Rahmanian using functional magnetic resonance imaging and measuring neuropeptide secretion. Our results showed that Rehmannia alcohol extract and the related active ingredient, Rehmannioside D, can delay cartilage degradation and alleviate inflammation in OA rats. The Rehmannia alcohol extract can also relieve OA pain, reduce the secretion of calcitonin gene-related peptide (CGRP) and substance P (SP), and reverse the pathological changes in the cerebral cortex and hippocampus. Our research results demonstrate that Rehmannia alleviates OA pain by protecting cartilage, preventing the stimulation of inflammatory factors on neuropeptide secretion, and influencing the relevant functional areas of the brain.

Effects of spray cryotherapy on cough receptors and airway microenvironment in a canine model of chronic bronchitis

The aim of this study was to explore the effects of spray cryotherapy (SCT) on cough receptors and airway microenvironment in a canine model of chronic bronchitis. We examined the expression of transient receptor potential vanilloid 1/4 (TRPV1/4) and the neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP) at the gene and protein levels before and after SCT. In addition, we explored whether TRPV1/4 could regulate inflammatory factors via mediator adenosine triphosphate (ATP). The levels of ATP and cytokines in alveolar lavage fluid and cell supernatant were measured using ELISA. SCT effectively downregulated the expression of TRPV1/4 and SP/CGRP in canine airway tissues with chronic bronchitis and reduced the levels of inflammatory mediators and cytokines that affect cough receptor sensitivity, achieving cough relief. TRPV1/4 - ATP - inflammatory cytokines axis has been demonstrated at the cellular level, which in turn modulate the milieu of the airways and promote the formation of a cough feedback loop. Our study has fully revealed the specific mechanism of SCT in treating cough in a canine model of chronic bronchitis, providing a solid theoretical basis for future clinical treatment.

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

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. 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. 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). 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.

The vasoactive effects of bradykinin, vasoactive intestinal peptide, calcitonin gene-related peptide and neuropeptide Y depend on the perivascular tissue in porcine retinal arterioles in vitro.

The retina contains a number of vasoactive neuropeptides and corresponding receptors, but the role of these neuropeptides for tone regulation of retinal arterioles has not been studied in detail. Porcine arterioles with preserved perivascular retinal tissue were mounted in a wire myograph, and the tone was measured after the addition of increasing concentrations of bradykinin, vasoactive intestinal peptide (VIP), neuropeptide Y (NPY), substance P (SP), calcitonin gene-related peptide (CGRP) and brain natriuretic peptide (BNP). The experiments were performed during inhibition of the synthesis of nitric oxide (NO), prostaglandins and dopamine and were repeated after removal of the perivascular retinal tissue. Bradykinin, VIP and CGRP induced significant concentration-dependent dilatation and NPY significant concentration-dependent contraction of the arterioles in the presence of perivascular retinal tissue (p < 0.03 for all comparisons) but not on isolated arterioles. BNP and SP had no effect on vascular tone. The NOS inhibitor L-NAME reduced bradykinin- and VIP-induced relaxation (p < 0.001 for both comparisons), whereas none of the other inhibitors influenced the vasoactive effects of the studied neuropeptides. The effects of neuropeptides on the tone of retinal arterioles depend on the perivascular retinal tissue and may involve effects other than those mediated by nitric oxide, prostaglandins and adrenergic compounds. Investigation of the mechanisms underlying the vasoactive effect of neuropeptides may be important for understanding and treating retinal diseases where disturbances in retinal flow regulation are involved in the disease pathogenesis.

Intradermal substance P as a challenge agent in healthy individuals.

Pharmacological challenge models are deployed to evaluate drug effects during clinical development. Intradermal injection of Substance P (SP) neuropeptide, a potential challenge agent for investigating local mediators, is associated with wheal and flare response mediated by the MRGPRX2 receptor. Although dose-dependent data on SP effects exist, full characterization and information on potential carryover effect after repeated challenge are lacking. This open-label, two-part, prospective enabling study of SP intradermal challenge in healthy participants aimed to understand and distinguish between wheal and flare responses following various SP doses. Part 1 included one challenge visit to determine optimum SP dose range for evaluation in part 2, which determined variability in 20 participants and used intradermal microdialysis (IDM) for SP-challenged skin sampling. At 5, 15, 50, and 150 pmol doses, respectively, posterior median area under the curve (AUC; AUC0-2h ) was 4090.4, 5881.2, 8846.8, and 9212.8 mm2 /min, for wheal response, and 12020.9, 38154.3, 65470.6, and 67404.4 mm2 /min for flare response (SP-challenge visit 2). When the challenge was repeated ~2 weeks later, no carryover effect was observed. IDM histamine levels were relatively low, resulting in low confidence in the data to define temporal characteristics for histamine release following SP challenge. No safety concerns were identified using SP. Wheal and flare responses following intradermal SP challenge were dose-dependent and different. The results indicate that this challenge model is fit-for-purpose in future first-in-human studies and further assessment of novel drugs targeting dermal inflammatory disease responses, such as chronic spontaneous urticaria, chronic inducible urticaria, and pseudo-allergic reactions.

COMPARATIVE CHARACTERISTICS OF SUBSTANCE P CONCENTRATION AND NEUROPATHIC PAIN COMPONENT IN PATIENTS WITH ACUTE AND CHRONIC COURSES OF VERTEBROGENIC LUMBAR-SACRAL PAIN SYNDROMES

The aim. The primary aim of this study is to assess the variance in substance P concentration, taking into account pain intensity and the presence of a neuropathic component, among patients experiencing acute and chronic vertebral lumbosacral pain syndromes.&#x0D; Materials. Measurement of substance P concentration in serum blood, evaluation of pain syndrome using the VAS scale, and assessment of pain according to the DN4 questionnaire were performed in patients receiving comprehensive treatment. A total of 125 patients were included in the study, comprising 65 patients with acute pain and 60 patients with chronic pain syndrome.&#x0D; Results. Pain ratings according to the Visual Analog Scale (VAS) among patients from the acute pain group was 7.15±1.07 on the VAS scale, while patients in the chronic pain group rated it at 6.09±0.96. On the 7th day of treatment, patients in the acute pain group reported a pain rating of 4.46±0.73, whereas those in the chronic pain group reported a rating of 4.22±0.97. The level of substance p(SP) in serum blood on the first day of treatment was 13.96±1.93 ng/mL in the acute pain group and 10.34±2.92 ng/mL in the chronic pain group. By the 7th day of treatment, the level decreased to 7.72±1.90 ng/mL in the acute pain group and 8.03±2.14 ng/mL in the chronic pain group. According to the DN4 questionnaire, neuropathic component was ascertained in 38% of patients with acute pain syndrome and 56% of participants with chronic pain. The average DN4 score in the acute pain group was 2.26±1.01, while in the chronic pain group, it was 4.53±1.02. In the group of patients with chronic pain syndrome and the presence of a neuropathic component, the level of SP was 10.89±1.78 ng/mL. In contrast, without the presence of a neuropathic component, it was 6.51±1.94 ng/mL.&#x0D; Conclusions. The trajectory of acute and chronic pain syndromes in patients with lumbosacral vertebral pain syndromes exhibits dissimilarities. Patients experiencing acute pain displayed a more favorable trend characterized by a reduction in pain intensity and a decrease in pain-related neurotransmitter concentration in the bloodstream.

Preparation of co-electrospinning membrane loaded with simvastatin and substance P to accelerate bone regeneration by promoting cell homing, angiogenesis and osteogenesis.

Bone regeneration is a complex process that requires the coordination of various biological events. Developing a tissue regeneration membrane that can regulate this cascade of events is challenging. In this study, we aimed to fabricate a membrane that can enrich the damaged area with mesenchymal stem cells, improve angiogenesis, and continuously induce osteogenesis. Our approach involved creating a hierarchical polycaprolactone/gelatin (PCL/GEL) co-electrospinning membrane that incorporated substance P (SP)-loaded GEL fibers and simvastatin (SIM)-loaded PCL fibers. The membrane could initiate a burst release of SP and a slow/sustained release of SIM for over a month. In vitro experiments, including those related to angiogenesis and osteogenesis (e.g., migration, endothelial network formation, alkaline phosphatase activity, mineralization, and gene expression), clearly demonstrated the membrane's superior ability to improve cell homing, revascularization, and osteogenic differentiation. Furthermore, a series of in vivo studies, including immunofluorescence of CD29+/CD90+ double-positive cells and immunohistochemical staining for CD34 and vWF, confirmed the co-electrospinning membrane's ability to enhance MSC migration and revascularization response after five days of implantation. After one month, the Micro-CT and histological (Masson and H&E) results showed accelerated bone regeneration. Our findings suggest that a co-electrospinning membrane with time-tunable drug delivery could advance the development of tissue engineering therapeutic strategies and potentially improve patient outcomes.

AbobotulinumtoxinA improves skin properties and sebum quality in the rhino mouse

Besides neuronal cells, botulinum neurotoxins (BoNTs) can also affect other cell types such as fibroblasts or keratinocytes. These cells play a key role in skin conditions. Maintaining a high-quality sebum secretion is essential to avoid premature aging. This study explored the effect of abobotulinumtoxinA (aboBoNT-A) in the rhino mouse. Briefly, anaesthetized animals were injected via the intra-dermal route (ID; four sites of injection) by either vehicle or 0.1, 0.3 and 1 Unit aboBoNT-A per mouse. A reference group was administered with adapalene gel 0.1% (daily local application) for 15 days. Adapalene is a third-generation retinoid and is used as first-line treatment of moderate acne. The body weight and the thickness of the dorsal skin were measured on days 1, 5, 10 and 15; erythema and scaling were recorded at the same time. On day 15, animals were ethically euthanized and skin samples were collected for histology, ELISA and lipidomic assays. AboBoNT-A administered ID at the doses 0.1 U and 0.3 U per mouse was well tolerated. 1 U aboBoNT-A (per mouse) induced a transient loss of muscle tone associated with a slight body weight loss after which mice recovered a good health status. AboBoNT-A did not show any significant effect on utricles surface area but induced a significant anti-inflammatory effect on dermis at the two highest doses. Moreover, aboBoNT-A showed neither side effects commonly observed with local retinoids, nor hyperplasia or dermis inflammation. No change in skin Interleukin-1alpha (IL-1α) cytokine levels was evidenced with aboBoNT-A, whereas a dose-dependent increase of substance P (SP) concentration in the skin was recorded, suggesting that aboBoNT-A induces neuropeptide accumulation in tissue by inhibiting exocytosis mechanisms. Lipidomic analysis showed that aboBoNT-A significantly increased the sebum concentration of several lipid species, presenting skin protecting properties. Overall, these data suggest that ID aboBoNT-A has skin rejuvenation, anti-inflammatory and moisture-boosting properties.

[Effect of deep needling "Tianshu" (ST25) on colonic motility in rats with slow transit constipation].

To observe the effect of acupuncture of "Tianshu"(ST25) at different depths on colonic transportation function, expressions of colonic substance P (SP) and interstitial cells of Cajal (ICC) in rats with slow transit constipation (STC), so as to explore its mechanisms underlying improvement of STC．. Fifty male Wistar rats were selected and randomly divided into control，STC model，conventional acupuncture，deep needling group 1 and deep needling group 2 groups，with 10 rats in each group．The STC model was established by gavage of 1 mg/mL compound diphenoxylate suspension (10 mg/kg), once every other day for 21 days, and rats of the control group were given the same dose of distilled water by gavage．EA (2 Hz, 2 mA) was applied to "Tianshu"(ST25), with the acupuncture needle inserted to a depth of 3 mm for rats of the conventional acupuncture group, 4.5 mm for those of deep needling group 1, and 10 mm for those of the deep needling group 2. The acupuncture needle was twirled for 1 min, then retained for 15 min each time, once a day for 15 consecutive days．Following modeling, rats of the 4 groups and the control group received gavage of active carbon 2 mL (100 g/L) for observing the excretion time of the first black stool grain to assess the intestinal transit function. The colonic myoelectric activities (frequency and amplitude) were recorded by using BIOPAC multichannel physiograph. The immunoactivity of SP and c-kit (a transmembrane protein kinase for identification of ICC) of colonic musculature was detected by immunohistochemistry. Compared with the control group，the time of excretion of the first black stool grain, and the amplitude of colonic electromyogram (EMG) were significantly increased (P<0.01), while the frequency of EMG, expressions of SP and c-kit (ICC) were significantly decreased in the model group (P<0.01). In contrast to the model group, both deep needling group 1 and 2 had a decrease in the time of excretion of the first black stool grain, and amplitude of intestinal EMG, and an increase of frequency of intestinal EMG, and immunoactivity of SP and c-kit (P<0.01). The effect of deep needling 2 is superior to that of deep needling 1 in reducing the time of excretion of the first black stool grain (P<0.05), lowering the amplitude of EMG of the gut smooth muscle (P<0.05) and in increasing the frequency of EMG (P<0.05) and the expressions of SP and c-kit (P<0.05). No significant changes were found in the levels of frequency and amplitude of EMG, and expressions of SP and c-kit after routine needling in comparison with the model group (P>0.05), except the excretion time of the first black stool grain (P<0.05). Deep needling at ST25 at depth of 4.5 mm and 10 mm，especially at depth of 10 mm，has a significant effect in promoting gut motility to ameliorate constipation in rats with STC, which may be related to its function in up-regulating the expressions of SP and ICC activity.

Distribution and synaptic organization of substance P-like immunoreactive neurons in the mouse retina.

Substance P (SP), a neuroprotective peptidergic neurotransmitter, is known to have immunoreactivity (IR) localized to amacrine and/or ganglion cells in a variety of species' retinas, but it has not yet been studied in the mouse retina. Thus, we investigated the distribution and synaptic organization of SP-IR by confocal and electron microscopy immunocytochemistry in the mouse retina. SP-IR was distributed in the inner nuclear layer (INL), inner plexiform layer (IPL), and ganglion cell layer (GCL). Most of the SP-IR somas belonged to amacrine cells (2.5% of all) in the INL and their processes stratified into the S1, S3, and S5 layers of the IPL, with the most intense band in the S5 layer. Some SP-IR somas can also be observed in the GCL, which were identified as displaced amacrine cells (82%, 1269/1550) and ganglion cells (18%, 281/1550) by antibodies against AP2α and RBPMS, respectively. Such SP-IR ganglion cells (1.2% of all RGCs) can be further divided into 3 subgroups expressing SP/α-Synuclein (α-Syn), SP/GAD67, and/or SP/GAD67/α-Syn. Possible physiological and pathological roles of these ganglion cells are discussed. Further, electron microscopy evidence demonstrates that SP-IR amacrine cells receive major inputs from other SP-IR amacrine cell processes (146/242 inputs) and output mostly to SP-negative amacrine cell processes (291/673 outputs), suggesting series inhibition among amacrine cells. These results reveal for the first time an explicit distribution, novel ganglion cell features, and synaptic organization of SP-IR in the mouse retina, which is important for the future use of mouse models to study the roles of SP in healthy and diseased (including Parkinson's disease) retinal states.

Electroacupuncture on Baihui (DU20) and Xuehai (SP10) acupoints alleviates psoriatic inflammation by regulating neurotransmitter substance P- Neurokinin-1 receptor signaling

BackgroundAt present, acupuncture-related practices have been widely used to treat psoriasis. In our study, we investigated the effect and explored the mechanism of electroacupuncture (EA) on acupoints Baihui (DU20) and Xuehai (SP10) for the treatment of psoriasis. MethodsImiquimod-induced psoriasis-like mouse model was used in this study. Mice were treated with electroacupuncture at DU20 and SP10 (depth of 2–3 mm, frequency of 2/15 Hz, intensity of 0.5–1.0 mA, 10 min/day). The severity of psoriasis-like lesions for each group was assessed. In addition, histological analysis of the lesions were performed. The levels of inflammatory cytokines were determined using Elisa. The expression levels of Substance P (SP) and NK1R were measured using Western blotting. In addition, NK1R inhibitor was administrated to evaluate the target of electroacupuncture in our mouse model. ResultsElectroacupuncture significantly alleviated IMQ-induced skin lesions and epidermal thickness, accompanied with reduced keratinocyte proliferation, CD3+, CD4+, and CD8+ T cells infiltration. The reduced levels of inflammatory cytokines was observed after electroacupuncture treatment. In addition, electroacupuncture inhibited the expression levels of SP and NK1R. NK1R inhibitor could ameliorate lesional symptoms and suppress epidermal thickening and CD3+, CD4+, and CD8 + T cell infiltration. ConclusionsElectroacupuncture relieved psoriasis-like inflammation and T cell infiltration. This therapeutic action was likely mediated by the modulation of Substance P and its receptor NK1R.