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The Journal of Neuroscience : the... Dec 1983We have used antisera specifically directed against Met-enkephalin (met-ENK), somatostatin (SOM), substance P (SP), and vasoactive intestinal peptide (VIP), to study the... (Comparative Study)
Comparative Study
We have used antisera specifically directed against Met-enkephalin (met-ENK), somatostatin (SOM), substance P (SP), and vasoactive intestinal peptide (VIP), to study the development of neurons containing these peptides in the foregut of the chick. All four peptides were detected early in ontogeny, at 4 to 9 days of incubation (d.i.), and were localized primarily to cell bodies in the primitive myenteric plexus. There were differences in the times at which they were first detected and in the sequence of their appearance in the proventriculus, gizzard, and duodenum. The differentiation of these peptidergic neurons in the duodenum was examined in some detail. Cell bodies containing these peptides were first detected in the primitive myenteric plexus at 5 to 7 d.i. and increased in number from 7 to 11 d.i. Processes containing varicosities became prominent between 11 and 13 d.i. VIP was the first of the peptides to appear in the submucosal plexus and was found in more proximal regions of the duodenum at 5 d.i. Shortly thereafter, SOM- and SP-containing cell bodies were seen; met-ENK-containing cell bodies were never detected in the submucosal plexus. At 13 d.i., the circular smooth muscle contained a number of VIP-immunoreactive and a smaller number of SOM-immunoreactive processes. met-ENK- and SP-immunoreactive processes appeared in the circular smooth muscle between 17 and 21 d.i.; VIP- and SP-immunoreactive processes appeared in the mucosal plexus at 17 to 21 d.i. Our results suggest that neuropeptides appear very early in the ontogeny of enteric neurons, at the same time or even before cholinergic and serotonergic neurons express their phenotypes. These findings argue against a sequential developmental order in which peptidergic neurons appear after those containing acetylcholine and serotonin.
Topics: Animals; Cell Differentiation; Chick Embryo; Chickens; Enkephalin, Methionine; Fluorescent Antibody Technique; Intestines; Neurons; Somatostatin; Substance P; Vasoactive Intestinal Peptide
PubMed: 6197517
DOI: 10.1523/JNEUROSCI.03-12-02431.1983 -
The Journal of Clinical Investigation Sep 1991Human peripheral blood mononuclear cells are analyzed for preproenkephalin gene expression and peptide processing. Met-enkephalin immunoreactivity as detected with a...
Human peripheral blood mononuclear cells are analyzed for preproenkephalin gene expression and peptide processing. Met-enkephalin immunoreactivity as detected with a specific antiserum is found in the cytoplasm of monocytes but not in T lymphocytes. Secretion of met-enkephalin was analyzed with an RIA that is specific for the met-enkephalin pentapeptide. Unfractionated PBMC spontaneously released 40 pg/ml met-enkephalin and this increased two- to fourfold after stimulation with PHA. Lower levels (less than 100 pg/ml) of met-enkephalin were detected in supernatants from purified T cells that were activated with PHA and IL-2. In contrast, stimulation of purified monocytes with LPS or PMA resulted in the release of up to 600 pg/ml of the processed peptide. To examine whether T cells can produce met-enkephalin precursor peptides, T cell conditioned media were treated with trypsin and carboxypeptidase-B, which is known to release met-enkephalin from the propeptide. This increased levels of met-enkephalin to 400 pg/ml, indicating that lymphocytes secrete the propeptide but do not process it to met-enkephalin. The 1.4-kb preproenkephalin mRNA is detected in activated blood mononuclear cells and in purified monocytes and T cells. To determine whether monocytes or lymphocytes express met-enkephalin in vivo, lymphoid tissues were analyzed by immunohistochemistry. In human spleen tissue, positive cells were found in the red pulp but not in the follicles, which is also consistent with met-enkephalin expression in monocytes. In summary, these results show that human peripheral blood mononuclear cells express preproenkephalin mRNA and that monocytes, but not T cells, process the propeptide to metenkephalin.
Topics: Adult; Enkephalin, Methionine; Enkephalins; Gene Expression; Humans; Monocytes; Protein Precursors; RNA, Messenger; T-Lymphocytes
PubMed: 1885771
DOI: 10.1172/JCI115382 -
Proceedings of the National Academy of... Oct 1981Detailed competitive displacement curves of 3H-labeled [D-Ala2,Met5]enkephalinamide, [D-Ala2,D-Leu5]enkephalin, and dihydromorphine by a series of opiates and...
Detailed competitive displacement curves of 3H-labeled [D-Ala2,Met5]enkephalinamide, [D-Ala2,D-Leu5]enkephalin, and dihydromorphine by a series of opiates and enkephalins are biphasic, suggesting multiple sites. After treatment of tissue with naloxazone, the displacement of the three 3H-labeled ligands by all opiates and enkephalins tested becomes monophasic, losing the high-affinity displacement seen with low concentrations of both opiates and enkephalins. Coupled with Scatchard analysis of saturation experiments, these findings suggest a common site that binds both opiates and enkephalins equally well and with highest affinity (Kd values, less than 1 nM). Termed the mu 1 site, it corresponds to the previously described high-affinity site and appears to be the site responsible for analgesia under normal circumstances. The low-affinity binding of [3H]dihydromorphine (Kd, 3 nM) remaining after naloxazone treatment differs dramatically from low-affinity [D-Ala2,D-Leu5]-[3H]enkephalin binding (Kd, 5 nM). The mu 2 site, corresponding to the low-affinity [3H]dihydromorphine receptor sites, binds morphine (Ki, 10 nM) and dihydromorphine (Kd, 3 nM) far better than [D-Ala2,D-Leu5]enkephalin (Ki, 50 nM). Low-affinity [D-Ala2,D-Leu5]-[3H]enkephalin receptor sites bind [D-Ala2,D-Leu5]enkephalin (Ki, 5-8 nM) more potently than morphine (Ki, 71 nM) and correspond to the previously established delta receptor.
Topics: Animals; Binding, Competitive; Brain; Cell Membrane; Endorphins; Enkephalin, Methionine; Enkephalins; Kinetics; Morphine; Naloxone; Rats; Receptors, Opioid; Structure-Activity Relationship
PubMed: 6273857
DOI: 10.1073/pnas.78.10.6181 -
Journal of Anatomy Oct 1989The localisation of chromogranins A and B, met-enkephalin-arg6-gly7-leu8 (met-enk 8) and protein gene product 9.5 (PGP 9.5) in the adrenal medulla and extra-adrenal...
Localisation of chromogranin A and B, met-enkephalin-arg6-gly7-leu8 and PGP9.5-like immunoreactivity in the developing and adult rat adrenal medulla and extra-adrenal chromaffin tissue.
The localisation of chromogranins A and B, met-enkephalin-arg6-gly7-leu8 (met-enk 8) and protein gene product 9.5 (PGP 9.5) in the adrenal medulla and extra-adrenal chromaffin tissue has been studied in the developing rat by immunogold-silver staining. In the adult rat adrenal the cytoplasm of all medullary chromaffin cells showed a positive response with chromogranin A and B; in each case occasional groups of cells with a low reactivity that may have been NA cells were seen. Chromogranin A was first detected in adrenal medullary and extra-adrenal chromaffin cells at 18 days of gestation whilst chromogranin B was not detected in animals younger than 7 days. In 15 days old animals the adrenal medullary response to A and B was of the same intensity as that seen in the adult. Less than 1% of adult medullary chromaffin cells were responsive to met-enk 8 staining and medullary cells were unreactive in the fetus, with only extra-adrenal chromaffin tissue responding prenatally. During the first postnatal week immunoreactive cells appeared in the adrenal medulla in considerably greater proportions than in the adult gland. In contrast, positively stained nerve terminals associated with chromaffin cells and abundant in the adult adrenal were not detected during the first week of life. Immunoreactive nerve terminals were first seen early in the second week of life at a time when positive chromaffin cells were becoming less common. PGP 9.5 was located in all chromaffin cells of the adult adrenal and was readily detected in chromaffin cells in the adrenal and in extra-adrenal locations of the earliest stage examined (E16). Our findings suggest that the ontogenesis of the chromogranin-like immunostaining reflects the maturation of chromaffin granules and the PGP 9.5 immunostaining detected a protein common to cells of neuronal origin and expressed at an early stage of differentiation. The reciprocal relationship between the presence of enkephalins in chromaffin cells and in their presynaptic terminals merits further investigation.
Topics: Adrenal Medulla; Aging; Animals; Chromaffin System; Chromogranin A; Chromogranins; Enkephalin, Methionine; Fetus; Nerve Tissue Proteins; Neuropeptides; Rats; Rats, Inbred Strains; Ubiquitin Thiolesterase
PubMed: 2533591
DOI: No ID Found -
PloS One 2012Postoperative pain management is a critical aspect of patient care. The inflammatory state of the post-sternotomy surgical wound sensitizes nerve endings, causing pain....
BACKGROUND
Postoperative pain management is a critical aspect of patient care. The inflammatory state of the post-sternotomy surgical wound sensitizes nerve endings, causing pain. Unrelieved or improperly managed pain compromises wound healing. Peripheral opioid receptors play a major role in analgesia, particularly under inflammatory conditions where both opioid receptor expression and efficacy are increased. Leukocytic opioid peptides include β-endorphin (END), met-enkephalin (ENK), and dynorphin-A (DYN), with END and ENK being predominant.
METHODOLOGY/PRINCIPAL FINDINGS
This work represents the first study of inflammatory cells collected from post-sternotomy wounds of patients undergoing cardiac surgery including coronary artery bypass grafting (CABG). Wound fluid (WF) and cells were collected from sternal wounds using a JP Blake drain at 24, 48, and 72 hours post sternum closure. Anti-CD15 staining and flow cytometry revealed that polymorphonuclear neutrophils (PMN) are the predominant cells present in wound fluid collected post-surgery. Compared to peripheral blood (PB) derived PMN, significant increases in CD177+/CD66b+ PMN were observed suggesting activation of wound-site PMN. Such activation was associated with higher levels of opioid peptide expression in PMN derived from WF. Indeed, increased level of opioid peptides in sternal wound environment was noted 72 h post-surgery. We demonstrate that WF contains factors that can significantly induce POMC transcription in human PMNs. IL-10 and IL-4 were abundant in WF and both cytokines significantly induced POMC gene expression suggesting that WF factors such as IL-10 and IL-4 contribute towards increased opioid peptide expression in wound-site PMN.
CONCLUSIONS/SIGNIFICANCE
This approach provided a unique opportunity to study the cross-talk between inflammation and opioid peptides in PMN at a sternotomy wound-site. Wound-site PMN exhibited induction of END and ENK. In addition, sternal wound fluid significantly induced END expression in PMN. Taken together, these data constitute first clinical evidence that human wound-site PMNs are direct contributors of opioids at the sternal wound-site.
Topics: Aged; Dynorphins; Enkephalin, Methionine; Female; Flow Cytometry; Gene Expression Regulation; Humans; Inflammation; Interleukin-10; Interleukin-4; Male; Middle Aged; Neutrophils; Pain Management; Postoperative Period; Sternotomy; Wound Healing; beta-Endorphin
PubMed: 23118879
DOI: 10.1371/journal.pone.0047569 -
Scientific Reports Jul 2022With different countries facing multiple waves, with some SARS-CoV-2 variants more deadly and virulent, the COVID-19 pandemic is becoming more dangerous by the day and...
With different countries facing multiple waves, with some SARS-CoV-2 variants more deadly and virulent, the COVID-19 pandemic is becoming more dangerous by the day and the world is facing an even more dreadful extended pandemic with exponential positive cases and increasing death rates. There is an urgent need for more efficient and faster methods of vaccine development against SARS-CoV-2. Compared to experimental protocols, the opportunities to innovate are very high in immunoinformatics/in silico approaches, especially with the recent adoption of structural bioinformatics in peptide vaccine design. In recent times, multi-epitope-based peptide vaccine candidates (MEBPVCs) have shown extraordinarily high humoral and cellular responses to immunization. Most of the publications claim that respective reported MEBPVC(s) assembled using a set of in silico predicted epitopes, to be the computationally validated potent vaccine candidate(s) ready for experimental validation. However, in this article, for a given set of predicted epitopes, it is shown that the published MEBPVC is one among the many possible variants and there is high likelihood of finding more potent MEBPVCs than the published candidates. To test the same, a methodology is developed where novel MEBP variants are derived by changing the epitope order of the published MEBPVC. Further, to overcome the limitations of current qualitative methods of assessment of MEBPVC, to enable quantitative comparison and ranking for the discovery of more potent MEBPVCs, novel predictors, Percent Epitope Accessibility (PEA), Receptor specific MEBP vaccine potency (RMVP), MEBP vaccine potency (MVP) are introduced. The MEBP variants indeed showed varied MVP scores indicating varied immunogenicity. Further, the MEBP variants with IDs, SPVC_446 and SPVC_537, had the highest MVP scores indicating these variants to be more potent MEBPVCs than the published MEBPVC and hence should be preferred candidates for immediate experimental testing and validation. The method enables quicker selection and high throughput experimental validation of vaccine candidates. This study also opens the opportunity to develop new software tools for designing more potent MEBPVCs in less time.
Topics: COVID-19; Enkephalin, Methionine; Epitopes; Epitopes, B-Lymphocyte; Epitopes, T-Lymphocyte; Humans; Molecular Docking Simulation; Pandemics; Peptides; SARS-CoV-2; Vaccines, Subunit
PubMed: 35869117
DOI: 10.1038/s41598-022-16445-3 -
Gastroenterology Aug 2009Enteroendocrine cells, the largest and most diverse population of mammalian endocrine cells, comprise a number of different cell types in the gut mucosa that produce,... (Comparative Study)
Comparative Study
BACKGROUND & AIMS
Enteroendocrine cells, the largest and most diverse population of mammalian endocrine cells, comprise a number of different cell types in the gut mucosa that produce, store, and secrete small molecules, peptides, and/or larger proteins that regulate many aspects of gut physiology. Little is known about less typical endocrine cells in the intestinal mucosa that do not contain secretory granules, such as brush or caveolated cells. We studied a subset of these enteroendocrine cells in duodenum that produce several peptides, including endogenous opioids, and that also express the Trpm5 cation channel.
METHODS
We studied expression patterns of Trpm5 and other molecules by immunohistochemical and enzyme-linked immunosorbent assay analyses of intestinal tissues from transgenic mice that express green fluorescent protein from the Trpm5 promoter, as well as wild-type and Trpm5-null mice.
RESULTS
We describe a type of enteroendocrine cell in mouse duodenum that is defined by the presence of Trpm5 and that does not contain typical secretory granules yet expresses endogenous opioids (beta-endorphin and Met-enkephalin) and uroguanylin in apical compartments close to the lumen of the gut.
CONCLUSIONS
Solitary chemosensory cells that coexpress beta-endorphin, Met-enkephalin, uroguanylin, and Trpm5 exist in mouse duodenum. These cells are likely to secrete the bioactive peptides into the intestinal lumen in response to dietary factors; release of the opioid peptides requires the Trpm5 ion channel.
Topics: Animals; Biological Transport; Cells, Cultured; Duodenum; Enkephalin, Methionine; Enteroendocrine Cells; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation; Intestinal Mucosa; Mice; Mice, Transgenic; Models, Animal; Opioid Peptides; Sensitivity and Specificity; TRPM Cation Channels; beta-Endorphin
PubMed: 19272386
DOI: 10.1053/j.gastro.2009.02.070 -
International Journal of Molecular... Aug 2013We propose a protocol that provides a systematic definition of reaction coordinate and related free-energy profile as the function of temperature for the protein-folding...
We propose a protocol that provides a systematic definition of reaction coordinate and related free-energy profile as the function of temperature for the protein-folding simulation. First, using action-derived molecular dynamics (ADMD), we investigate the dynamic folding pathway model of a protein between a fixed extended conformation and a compact conformation. We choose the pathway model to be the reaction coordinate, and the folding and unfolding processes are characterized by the ADMD step index, in contrast to the common a priori reaction coordinate as used in conventional studies. Second, we calculate free-energy profile as the function of temperature, by employing the replica-exchange molecular dynamics (REMD) method. The current method provides efficient exploration of conformational space and proper characterization of protein folding/unfolding dynamics from/to an arbitrary extended conformation. We demonstrate that combination of the two simulation methods, ADMD and REMD, provides understanding on molecular conformational changes in proteins. The protocol is tested on a small protein, penta-peptide of met-enkephalin. For the neuropeptide met-enkephalin system, folded, extended, and intermediate sates are well-defined through the free-energy profile over the reaction coordinate. Results are consistent with those in the literature.
Topics: Enkephalin, Methionine; Models, Theoretical; Molecular Dynamics Simulation; Protein Conformation; Protein Folding
PubMed: 23917881
DOI: 10.3390/ijms140816058 -
American Journal of Hematology Sep 2001In the present study, beta-endorphin and met-enkephalin were tested for their antiplatelet activity in human platelet-rich plasma (PRP). Blood samples were obtained from... (Review)
Review
In the present study, beta-endorphin and met-enkephalin were tested for their antiplatelet activity in human platelet-rich plasma (PRP). Blood samples were obtained from 15 healthy subjects. The results of the study show that these two endogenous opioids (200 pg/ml final concentration) reduce platelet aggregation when it is induced by ADP at low dose (0.5 microM). It is likely due to conformational changes on the platelet membrane that cause a non-specific decreased susceptibility to platelet-aggregating agonists.
Topics: Adenosine Diphosphate; Blood Platelets; Dose-Response Relationship, Drug; Enkephalin, Methionine; Humans; Osmolar Concentration; Platelet Aggregation; beta-Endorphin
PubMed: 11559929
DOI: 10.1002/ajh.1140 -
PloS One 2018The objective of this study was to evaluate the effect of low-dose naltrexone (LDN) as a carboplatin chemotherapy-associated drug in female dogs with mammary carcinoma...
The objective of this study was to evaluate the effect of low-dose naltrexone (LDN) as a carboplatin chemotherapy-associated drug in female dogs with mammary carcinoma in benign mixed tumors (MC-BMT) after mastectomy and to assess its association with quality of life and survival rates. Sixty female dogs were included in this study, all of which had histopathological diagnosis of MC-BMT and were divided into three groups: G1 (control), consisting of animals submitted only to mastectomy with or without regional metastasis; G2, composed of treated animals that did not present with metastasis; and G3, treated dogs that presented with metastasis. G2 and G3 were also subdivided according to the treatment administered: chemotherapy alone (MC-BMT(-) C/MC-BMT(+) C) or LDN and chemotherapy (MC-BMT(-) C+LDN/MC-BMT(+) C+LDN). All animals were subjected to clinical evaluation, mastectomy, peripheral blood lymphocyte immunophenotyping, beta-endorphin and met-enkephalin quantification, and evaluation of survival rates and quality of life scores. The results showed higher serum concentrations of beta-endorphin and met-enkephalin, fewer chemotherapy-related side effects, and better quality of life and survival rates in the LDN-treated groups than in LDN-untreated groups (P < 0.05). Evaluation of clinical and pathological parameters indicated a significant association between the use of LDN and both prolonged survival and enhanced quality of life. These results indicate that LDN is a viable chemotherapy-associated treatment in female dogs with MC-BMT, maintaining their quality of life and prolonging survival rates.
Topics: Animals; Carboplatin; Dogs; Drug Synergism; Enkephalin, Methionine; Female; Immunophenotyping; Lymphocytes; Mammary Neoplasms, Animal; Mastectomy; Naltrexone; Neoplasm Metastasis; Quality of Life; Survival Analysis; Treatment Outcome; beta-Endorphin
PubMed: 30286124
DOI: 10.1371/journal.pone.0204830