Calcitonin Gene-related Peptide Protein Expression Research Articles

Calcitonin gene related peptide modified mesenchymal stem cells reduce restenosis after carotid balloon injury in rats

This work aimed to investigate the effects of calcitonin gene-related peptide (CGRP)-modified mesenchymal stem cells (MSCs) on vascular stenosis in carotid balloon-injured rats. The CGRP gene labeled with EGFP was transfected into bone marrow MSCs, and CGRP protein expression in MSCs was confirmed by immunofluorescence assays. A rat carotid balloon injury model was established using a surgical method. CGRP-modified MSCs were orthotopically transplanted into the injured area of the rats. At 28 days after cell transplantation, EGFP and CD31 expression was detected by immunofluorescence staining. Hematoxylin-eosin (H&E) staining was used to detect the intima/media area of the injured carotid artery stenosis site, and the expression of proliferating cell nuclear antigen (PCNA) was detected by immunohistochemistry. MSCs from rat bone marrow positively expressed CD29 and negatively expressed CD45. In vivo immunofluorescence staining showed that EGFP expression was significantly increased at the vascular injury site of rats transplanted with MSC_CGRP compared with the control group on the 28th day after cell transplantation and that the damaged vessels exhibited continuous CD31 expression. H&E staining showed that the vascular intimal proliferation area of rats transplanted with MSC_CGRP was significantly reduced compared with that of other groups. Furthermore, the immunohistochemistry results showed that PCNA expression in the endothelium of the MSC_CGRP group was lower than that of the other groups. Therefore, MSCs transfected with the CGRP gene can express the CGRP protein, and the implantation of CGRP-modified MSCs at the damaged site after carotid balloon-induced injury can reduce the neointimal area.

Calcitonin Gene-Related Peptide Is a Promising Marker in Ulcerative Colitis

Assessment of the severity and extent of disease activity continues to present challenges for physicians in the treatment of ulcerative colitis. Standard markers that can objectively reflect disease activity are useful for physicians to both evaluate the course of ulcerative colitis and monitor the effectiveness of therapy for any given patient. We hypothesize that calcitonin gene-related peptide (CGRP) can reflect the activity and severity of ulcerative colitis and be used as a marker to assess the effectiveness of various therapies. We examined the expression levels of CGRP by reverse transcription polymerase chain reaction (RT-PCR) and semi-quantitative immunohistochemisty in mucosal biopsies from 38 patients with UC and 18 controls. Levels of CGRP mRNA and protein expression were compared between patients and controls with the clinical activity index (CAI) and the endoscopic activity index (EAI) for various levels of UC severity. Our results showed that the levels of CGRP mRNA and protein expression were significantly reduced in UC patients compared to controls. This effect was more pronounced in patients with more severe cases of UC. There is a statistically significant negative correlation between levels of CGRP mRNA expression and CAI/EAI scores. A statistically significant negative correlation was also found between levels of CGRP protein expression and CAI/EAI scores. Overall, high CAI and EAI scores were accompanied by low CGRP mRNA and protein expression levels. Levels of CGRP protein and mRNA expression in the colonic mucosa of patients are closely associated with UC severity and corroborate traditional indices used to assess the disease.

P2X3 receptor mediates ectopic mechanical allodynia with inflamed lower lip in mice

Ectopic pain in other orofacial regions develops with local inflammation in separated orofacial structures. However, the basis for the spreading of pain to adjacent orofacial areas after local inflammation is still unknown. In the present study, we determined if the P2X3 receptor (P2X3R) was associated with altered mechanical sensitivity of the whisker pad skin following complete Freund's adjuvant (CFA) injection into the lower lip. Mice with local inflammation induced by CFA injection into the lower lip demonstrated significant mechanical allodynia of whisker pad skin. The mechanical allodynia was reversed by P2X3R antagonist, A-317491 administration into whisker pad skin. The number of P2X3R and calcitonin gene-related peptide (CGRP) positive trigeminal ganglion (TG) neurons that innervates the whisker pad skin and lower lip was increased after CFA injection into the lower lip. CGRP protein expression in TG ipsilateral to CFA injection was also significantly greater than that of the saline-injected mice. The present findings suggest that induced CGRP by local inflammation in the lower lip increases P2X3R in TG neurons, the increased P2X3Rs are involved in the sensitization of primary afferent neurons in the whisker pad skin. This P2X3R overexpression may underlie ectopic mechanical allodynia in the whisker pad skin after CFA injection into the lower lip.

The impact of short-lasting repeated vibrations on retrograde axonal transport, the expression of CGRP and parvalbumin in lower lumbar dorsal root ganglia

A prolonged exposure to vibration stimuli triggers pathological changes with many later manifested symptoms. Early vibration-induced changes are still not very well explored. Therefore, short 30 min vibration period per day with frequency 60 Hz repeated for 10 days was used, and the retrograde axonal transport from the sciatic nerve, the expression of calcitonin gene-related peptide (CGRP) and parvalbumin (PV) were studied in the dorsal root ganglia (DRGs) corresponding to lower lumbar spinal levels. Repeated vibration markedly decreased (25 and 34%) the accumulation of retrogradely transported Fluorogold to spinal motor neurons, whereas a significant increase (35 and 25%) was seen in the DRG primary sensory neurons corresponding to the L4 and L5 spinal level. Immunohistochemical studies showed a significant reduction of CGRP-positive small-sized neuronal cells in both DRGs. Fluoro-Jade labeling revealed that marked loss of CGRP-imunoreactive DRG sensory neurons is not due to neuronal degeneration. CGRP protein expression determined by Western blot analysis and optical density measurement, and NGF level measured by ELISA have been decreased, markedly only at the L4 DRG. PV protein expression was not affected by short repeated vibrations. Our results indicate that (a) short-lasting repeated vibrations affect the retrograde axonal transport in the DRG sensory neurons differently than in spinal motor neurons; (b) a decreased NGF-dependent CGRP production in the DRG primary sensory neurons plays an important role in early vibration-induced pathological mechanisms.

Characterization of CGRP protein expression in “satellite‐like” cells and dendritic arbours of the mouse olfactory bulb

The main olfactory bulb (OB) is made up of several concentric layers, forming circuitries often involving dendro-dendritic synapses. Important interactions between OB neurons occur in the external plexiform layer (EPL), where dendrites of tufted and Van Gehuchten cells form synapses with dendrites of deeper lying mitral, tufted, and granule cells. OB neurons display a variety of neurotransmitters. Here, the focus is on calcitonin gene-related peptide (CGRP), a 37-amino acid neuropeptide transmitter that is widely distributed in the central and peripheral nervous system. In the OB, CGRP-immunoreactive (ir) cell bodies were mostly observed in the mitral cell layer (MCL) of normal mice, and their number increased following colchicine treatment. Sparsely distributed CGRP-ir cell bodies were also found in the EPL and granular cell layer. Double-immunofluorescence experiments revealed a lack of co-localization between CGRP-like immunoreactivity (LI) and corticotropin-releasing factor- or galanin-LI, two markers for mitral cells, and no CGRP-LI was found in cholecystokinin-, parvalbumin-, or vasoactive intestinal polypeptide-ir tufted/Van Gehuchten cells. CGRP-ir cell bodies were not found to co-localize glutamic acid decarboxylase 67 (GAD67)-green fluorescence protein, gamma-aminobutyric acid (GABA)-, or calretinin-LI, although the possibility remains that CGRP-ir cells may contain low levels of GABA and/or GAD67 not detected by our methodology. Dendrites of CGRP-ir cells extensively ramified deep in the EPL and double-immunofluorescence revealed them to be adjacent with, often apparently contacting, dendrites of granule, mitral, tufted, and Van Gehuchten cells. We propose that these CGRP-ir cell bodies in the mouse OB are "satellite-like" cells within and, occasionally, close to the MCL.

Expression of calcitonin gene-related peptide, adrenomedullin, and receptor modifying proteins in human adipose tissue and alteration in their expression with menopause status

Calcitonin gene-related peptide (CGRP) is a vasoactive, proinflammatory neuropeptide implicated in the pathogenesis of cardiovascular disease. Elevated CGRP levels during hot flushes and pregnancy suggest that reproductive hormones may influence CGRP secretion. CGRP and the related protein adrenomedullin (ADM) may function through adipose tissue-mediated effects, since adipose tissue is an important site of cytokine production and the main site for estrogen production after menopause. This study examined mRNA and protein expression of CGRP, ADM, and the receptor activity-modifying proteins and the effects of menopausal status in human adipose tissue. Protein/mRNA levels were determined in adipose tissue biopsy samples collected from premenopausal (n = 22: follicle-stimulating hormone, <20 IU/L; estradiol [mean +/- SEM], 434.5 +/- 87.81 pmol/L) and postmenopausal (n = 25: follicle-stimulating hormone, >20 IU/L; estradiol, 43.4 +/- 6.95 pmol/L) women. Our studies determined that CGRP, ADM, and receptor activity-modifying proteins were expressed in abdominal fat, adipocytes, and preadipocytes. CGRP and ADM mRNA levels were increased in abdominal subcutaneous fat in postmenopausal women compared with premenopausal women (betaCGRP: premenopause Delta cycle threshold [Ct], 31.07 +/- 0.28 vs postmenopause DeltaCt, 30.35 +/- 0.17, P = 0.035; ADM: premenopause subcutaneous fat DeltaCt, 12.41 +/- 0.2 vs postmenopause subcutaneous fat DeltaCt, 11.55 +/- 0.14, P < 0.001) with CGRP differentially expressed in subcutaneous and omental depots. CGRP protein expression was higher in postmenopausal women (P < 0.05) in both fat depots. Our findings suggest that adipose tissue represents an important site for CGRP and ADM production and that menopause status alters their expression in abdominal fat. This offers a potential mechanism to explain the role of CGRP in menopausal vasomotor symptoms and the increased risk of cardiovascular disease in postmenopausal women.