Loperamide-induced Constipation Activates Inflammatory Signaling Pathways in the Mid Colon of SD Rats Via Complement C3 and its Receptors.

BACKGROUND: Complement component 3 (C3) receptors play an important role as inflammatory mediators in the innate immune system, although their mechanisms were not well studied during constipation. OBJECTIVE: The aim of this study is to investigate the regulatory role of C3 and its receptors' downstream signaling during constipation. METHODS: Alterations in the C3, C3a receptor (C3aR), and C3b receptor (C3bR) expressions, PI3K/AKT pathway, RhoA/MLC pathway, MAP kinase pathway, and inflammatory cytokine expressions were measured in the mid colon of loperamide (Lop) treated SD rats. RESULTS: Lop treatment successfully induced constipation phenotypes, including decreased stool parameters and histological structure alterations. The expression levels of C3 were significantly increased, whereas expressions of C3aR and C3bR were decreased during Lop-induced constipation. Moreover, significant upregulation was observed in the phosphorylation levels of PI3K, AKT, and GSK3ß in mid colons of Lop treated SD rats. The expression of RhoA and phosphorylation of MLC were also enhanced in the Lop treated group. Furthermore, a similar pattern was detected in the MAP kinase pathway and inflammatory cytokine expressions. Subsequent to the Lop treatment, the phosphorylation of ERK and p38, as well as the mRNA levels of NF-κB, TNF-α, IL-6 and IL-1α were remarkably increased in the mid colon. CONCLUSION: These results indicate that Lop-induced constipation is tightly linked to the downregulation of C3aR and C3bR expressions, and upregulation of the C3 and C3Rs downstream signaling pathway, including PI3K/AKT, RhoA/MLC, and MAP kinase pathways as well as inflammatory cytokine expressions in the mid colon of SD rats.

Laxative effect and mechanism of Tiantian Capsule on loperamide-induced constipation in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Tiantian capsule (TTC), as a functional food, which consists of four herb medicines, including Aloe vera Burm.f. (25%), leaf juices, dried; Cucurbita moschata Duch. (25%), fructus, dried; Poria cocos (Schw.) Wolf. (12.5%), sclerotium, dried; Tremella fuciformis Berk. (12.5%), fruiting bodies, dried, and one extract xylooligosaccharides (25%) from Maize Cob by enzymolysis, has been commonly used in China to ameliorate constipation. AIM OF THE STUDY: The aim of the work is to elucidate the potential laxative mechanisms of TTC in loperamide-induced constipated rats. MATERIALS AND METHODS: LC-MS/MS was employed for analyzing the TTC extract. The gastrointestinal transit was evaluated by X-ray. The H&E and Alcian-Blue stain were applied to determine the changes of goblet cells and mucus layer, respectively. Meanwhile, levels of neurotransmitters were evaluated by enzyme-linked immunosorbent assay. The protein expressions were also measured by immunohistochemistry and Western blot. RESULTS: Our results showed that TTC administration attenuated constipation responses in aspects of fecal pellets number, water content of feces, stomach emptying and gastrointestinal transit. Further investigations revealed that TTC treatment not only induced the recovery of neurotransmitters, such as motilin, substance P, somatostatin, endothelin and vasoactive intestinal peptide, but also up-regulated the expressions of c-kit and stem cell factor (SCF). Additionally, the number of goblet cells and thickness of the mucus layer were elevated, and the guanylate cyclase C-cGMP signal pathway was also up-regulated after TTC treatment. CONCLUSION: Our findings demonstrated that the laxative effect of TTC in constipation rats is probably due to the regulation of bowel movement and intestinal fluid secretion.

Sacral nerve stimulation with appropriate parameters improves constipation in rats by enhancing colon motility mediated via the autonomic-cholinergic mechanisms.

Although sacral nerve stimulation (SNS) has been applied for treating constipation, its parameters were adopted from SNS for fecal incontinence, its effects are limited, and mechanisms are largely unknown. We investigated the effects and mechanism of SNS with appropriate parameters on constipation in rats treated with loperamide. First, using rectal compliance as an outcome measure, an experiment was performed to derive effective SNS parameters. Then, a 7-day SNS was performed in rats with constipation induced by loperamide. Autonomic functions were assessed by spectral analysis of heart rate variability (HRV) derived from an electrocardiogram. Serum levels of pancreatic polypeptide (PP), norepinephrine (NE), and acetylcholine (ACh) in colon were assessed. 1) Acute SNS at 5 Hz, 100 µs was found effective in enhancing rectal compliance and accelerating distal colon transit (P < 0.05 vs. sham SNS). 2) The 7-day SNS normalized loperamide-induced constipation, assessed by the number, weight, and water content of fecal pellets, and accelerated the distal colon transit (29.4 ± 3.7 min with sham SNS vs. 16.4 ± 5.3 min with SNS but not gastric emptying or intestinal transit. 3) SNS significantly increased vagal activity (P = 0.035) and decreased sympathetic activity (P = 0.012), assessed by spectral analysis of HRV as well as by the serum PP. 4) SNS increased ACh in the colon tissue; atropine blocked the accelerative effect of SNS on distal colon transit. We concluded that SNS with appropriate parameters improves constipation induced by loperamide by accelerating distal colon motility, mediated via the autonomic-cholinergic function.NEW & NOTEWORTHY Although sacral nerve stimulation (SNS) has been applied for treating constipation, its parameters were adopted from SNS for fecal incontinence, effects are limited, and mechanisms are largely unknown. This paper shows that SNS with appropriate parameters improves constipation induced by loperamide by accelerating distal colon motility mediated via the autonomic-cholinergic function.

Anti-Inflammatory Response and Muscarinic Cholinergic Regulation during the Laxative Effect of Asparagus cochinchinensis in Loperamide-Induced Constipation of SD Rats.

Several types of saponins and herbal plants containing saponins have been reported to have anti-inflammatory or laxative activities. To verify the therapeutic effects of saponin-enriched extracts of Asparagus cochinchinensis (SPA) on the anti-inflammatory response and on the cholinergic regulation in the gastrointestinal system, an alteration on the constipation phenotypes, on the inflammatory responses, and on the muscarinic cholinergic regulation were investigated in the transverse colons of Sprague Dawley (SD) rats with loperamide (Lop)-induced constipation after the treatment of SPA. Significant increases were observed on the total number of stools, the gastrointestinal transit, the thickness of the mucosal layer, the flat luminal surface, the number of paneth cells, and the lipid droplets in the Lop + SPA-treated group as compared to the Lop + Vehicle-treated group. SPA treatment induced the recovery of inflammatory cytokines (TNF-α, IL-1ß) and IL-6), inflammatory mediators (NF-κB and iNOS), the total number of infiltered mast cells, and mucin secretion. Also, some similar improvements were observed on the levels of acetylcholine esterase (AChE) activity and on the phosphorylation of myosin light chains (MLC) as well as the expression of muscarinic acetylcholine receptors M2/M3 (mAChR M2/M3) and their mediators. The results presented herein provide the first strong evidence that SPA stimulates anti-inflammatory responses and the muscarinic cholinergic regulation when exerting its laxative effects in the chronic constipation of Lop-induced models.

Enteromorpha and polysaccharides from enteromorpha ameliorate loperamide-induced constipation in mice.

Slow-transit constipation(STC)is a disease characterized by functional gastrointestinal disorder. Common laxatives used in clinical practice against constipation such as Senna have side effects. Enteromorpha(EP)is a common marine alga, and the polysaccharide extracted from EP has been reported possessing anti-cancer and anti-inflammation effects. The aim of this study is to investigate the effects of EP and Polysaccharides from Enteromorpha (PEP) on loperamide induced constipated mice model and illustrating mechanism of action. We investigated the effect of EP and PEP on fecal water content, defecation frequency and gastrointestinal transit (GI) time of loperamide-induced STC mice. In addition, serum Nitric Oxide (NO) content and vasoactive intestinal peptide receptor1 (VIPR1) as well as serotonin receptor (5-HT4) expression in the distal colon were analyzed. Furthermore, we determined the role of EP and PEP on microbiota distribution using stool genomic 16S rRNA sequencing. EP and PEP significantly enhanced intestinal motility function, and alleviated constipation associated intestinal inflammation. Moreover, EP and PEP significantly decreased serum NO concentration, down-regulated VIPR1 expression and up-regulated 5-HT4 expression in distal colon. Genomic stool DNA MiSeq Sequencing Analysis of microbiota community structures and distribution revealed that intestinal microecological changes caused by constipation recovered after both EP and PEP treatment. Our results indicate that EP and PEP are potent natural products which could be suggested in constipation therapy strategies.

Toxicological implications and laxative potential of ethanol root extract of Morella serrata in loperamide-induced constipated Wistar rats.

CONTEXT: Morella serrata L. (Myricaceae) is commonly used in South Africa to treat several diseases including constipation. OBJECTIVES: This study investigated toxicological implications and laxative potential of the ethanol root extract of the plant. MATERIALS AND METHODS: While normal control animals were placed on sterile placebo, the loperamide-constipated rats were treated with the extract at 75, 150 and 300 mg/kg doses for 7 days, and their feeding patterns and faecal properties were monitored. Gastrointestinal transit ratio and the toxicity profile of the tested doses were thereafter evaluated. RESULTS: The significantly increased faecal volume (192.08%), feed (63.63%), water intake (55.97%) and improved intestinal motility (95.05%) in the constipated rats following treatment with the extract (at 300 mg/kg) suggested laxative potential of the extract. The 1.5-2.0-fold normalization of the platelets, erythrocytes and leukocytes counts in the extract-treated constipated rats suggests its non-haematotoxic tendency. Furthermore, the extract (at the highest investigated dose) reversed the attenuation in the concentrations of the electrolytes (0.5-2-fold), total protein (62.12%) and albumin (55.88%) in the constipated animals as well as attenuated activities of hepatic enzymes (0.5-3.0-fold) and levels of urea (126.67%), creatinine (40.32%), cholesterol (3-fold) and triglycerides (9-fold). These further support its non-toxic and therapeutic attributes against constipation. CONCLUSION: Overall, the effect exhibited by M. serrata in this study competed well with Senokot (standard drug) and proved that it may be relatively safe and with excellent laxative potential, thus, supporting its pharmacological applications in South Africa.

Characterization of Changes in Global Genes Expression in the Distal Colon of Loperamide-Induced Constipation SD Rats in Response to the Laxative Effects of Liriope platyphylla.

To characterize the changes in global gene expression in the distal colon of constipated SD rats in response to the laxative effects of aqueous extracts of Liriope platyphylla (AEtLP), including isoflavone, saponin, oligosaccharide, succinic acid and hydroxyproline, the total RNA extracted from the distal colon of AEtLP-treated constipation rats was hybridized to oligonucleotide microarrays. The AEtLP treated rats showed an increase in the number of stools, mucosa thickness, flat luminal surface thickness, mucin secretion, and crypt number. Overall, compared to the controls, 581 genes were up-regulated and 216 genes were down-regulated by the constipation induced by loperamide in the constipated rats. After the AEtLP treatment, 67 genes were up-regulated and 421 genes were down-regulated. Among the transcripts up-regulated by constipation, 89 were significantly down-regulated and 22 were recovered to the normal levels by the AEtLP treatment. The major genes in the down-regulated categories included Slc9a5, klk10, Fgf15, and Alpi, whereas the major genes in the recovered categories were Cyp2b2, Ace, G6pc, and Setbp1. On the other hand, after the AEtLP treatment, ten of these genes down-regulated by constipation were up-regulated significantly and five were recovered to the normal levels. The major genes in the up-regulated categories included Serpina3n, Lcn2 and Slc5a8, whereas the major genes in the recovered categories were Tmem45a, Rerg and Rgc32. These results indicate that several gene functional groups and individual genes as constipation biomarkers respond to an AEtLP treatment in constipated model rats.

Synthesis and pharmacological activity of 2-(substituted)-3-{2-[(4-phenyl-4-cyano)piperidino]ethyl}-1,3-thiazolidin-4-ones.

Loperamide is a well-known peripherally acting opiate used for the treatment of diarrhoea. To gain more knowledge on the structure-activity relationships of antidiarrhoeal drugs and to develop new active molecules, a series of aryl-cyano-piperidinoalkyl-thiazolidinones related to Loperamide was synthesized and screened for antidiarrhoeal activity in mice by castor oil test. To characterize the potency and toxicity of the synthesized compounds ED50 and LD50 values were also determined. The thiazolidinones 2-6 displayed antidiarrhoeal activity at doses ranging between 15 and 82 mg/kg. Although the results show that the synthesized compounds are 15- to 80-fold less active respect to the reference compound, Loperamide, they are much less toxic (> or = 1000 mg/kg and 108.9 mg/kg, respectively). Besides, to evaluate the involvement of opioid receptors in antidiarrhoeal activity, Naloxone was administered prior to test the 2-phenyl-3-{2-[(4-phenyl-4-cyano)piperidino]ethyl}-1,3-thiazolidin-4-one (2), the more active compound of this series. The results obtained by this study, suggest that the antidiarrhoeal activity of this series of thiazolidinone derivatives could involve the opioid receptors.

Antidiarrhoeal activity of new thiazolidinones related to loperamide.

A series of thiazolidinones related to loperamide was synthesized and evaluated for antidiarrhoeal activity in mice, using the castor oil test. Of five compounds tested, antidiarrhoeal activity was found only for 2-(p-nitrophenyl)-3-¿3-[(4-(p-chlorophenyl)-4-hydroxy)piperidino]ethyl¿- 1,3-thiazolidin-4-one. The compound was less active than loperamide (ED50 values = 48.7 (24.8-95.6) and 0.91 (0.24-3.40) mg kg-1 respectively), but was also less toxic (LD50 values = 745.9 (545.2-929.8) and 108.9 (85.5-138.7) mg kg-1, respectively). Its antidiarrhoeal activity was counteracted by naloxone. Our results support the hypothesis that this compound, like loperamide, is an opiate-receptor agonist.

[Pharmacological studies of loperamide, an anti-diarrheal agent. I. Effects on diarrhea induced by castor oil and prostaglandin E. (author's transl)].

Effects of loperamide on diarrhea induced by castor oil and prostaglandin E1 were investigated in rats and mice and compared with those of narcotic analgesics, atropine, mecamylamine and local anesthetics. The following results were obtained. Loperamide markedly suppressed the appearance of diarrhea induced by oral administration of castor oil in rats and the ED50 values for 1 and 2 hr protection was 0.082 and 0.42 mg/kg p.o., respectively. Loperamide markedly suppressed the appearance of diarrhea induced by i.v. administration of prostaglandin E1 and the ED50 value for 2 hr protection was 0.24 mg/kg p.o. in rats. The ID120 min value of loperamide which was calculated on the basis of the dose producing a 20% or more inhibition of the charcoal transport in the small intestine for 120 min was 0.8 mg/kg p.o. in mice and this activity was 9.2 times more potent than that of morphine. The analgesic ED50 value (Haffner's method) and LD50 value of loperamide was 149 and 249 mg/kg p.o., respectively. These results suggest that loperamide has a potent anti-diarrheal activity and specificity to the gastrointestinal tract and inhibits the effect of prostaglandin E1 and ricinoleic acid on the intestinal tract in rats.