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. 2019 Dec 10:2019:4921086.
doi: 10.1155/2019/4921086. eCollection 2019.

In Vivo, Proteomic, and In Silico Investigation of Sapodilla for Therapeutic Potential in Gastrointestinal Disorders

Sameen Fatima Ansari  1 , Arif-Ullah Khan  1 , Neelum Gul Qazi  1 , Fawad Ali Shah  1 , Komal Naeem  1
Affiliations

In Vivo, Proteomic, and In Silico Investigation of Sapodilla for Therapeutic Potential in Gastrointestinal Disorders

Sameen Fatima Ansari et al. Biomed Res Int. .

Abstract

This study aims to delineate the effects of Manilkara zapota Linn. (Sapodilla) fruit chloroform (Mz.CHCl3) and aqueous (Mz.Aq) extracts tested through different techniques. Antidiarrheal activity and intestinal fluid accumulation were examined by using castor oil-induced diarrhea and castor oil fluid accumulation models. Isolated rabbit jejunum tissues were employed for in vitro experiments. Antimotility and antiulcer were performed through charcoal meal transient time and ethanol-induced ulcer assay, molecular studies were conducted through proteomic analysis, and virtual screening was performed by using a discovery studio visualizer (DSV). Mz.CHCl3 and Mz.Aq extracts attributed dose-dependent (50-300 mg/kg) protection (20-100%) against castor oil-induced diarrhea and dose-dependently (50-300 mg/kg) inhibited intestinal fluid secretions in mice. Mz.CHCl3 and Mz.Aq extracts produce relaxation of spontaneous and K+ (80 Mm) induced contractions in isolated tissue preparations and decreased the distance moved by charcoal in the gastrointestinal transit model in rats. It showed gastroprotective effect in ulcerative stomach of rats and decreased levels of IL-18 quantified by proteomic analysis. Histopathological results showed ethanol-induced significant gastric injury, leading to cloudy swelling, hydropic degeneration, apoptosis, and focal necrosis in all gastric zones using hematoxylin and eosin (H&E) staining. Moreover, ethanol increased the activation and the expression of tumor necrotic factor (TNF-α), cyclooxygenase (COX-2), and nuclear factor kappa-light-chain-enhancer of activated B cells (p-NFκB). In silico results were comparative to in vitro results evaluated through virtual screening. Moreover, ethanol increased the activation and expression of tumor necrotic factor, cyclooxygenase, and nuclear factor kappa-light-chain-enhancer of activated B cells. This study exhibits the gastroprotective effect of Manilkara zapota extracts in the peritoneal cavity using a proteomic and in silico approach which reveals different energy values against target proteins, which mediate the gastrointestinal functions.

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Conflict of interest statement

The authors declare that they have no conflicts of interest.

Figures

Figure 1
Figure 1
2D chemical structure of Manilkara zapota constituent (methyl chlorogenate) drawn through GaussView 5.0 Software and saved in PDB format.
Figure 2
Figure 2
Inhibitory effect of Manilkara zapota extracts: chloroform (Mz.CHCl3), aqueous (Mz.Aq), and atropine on castor oil-induced fluid accumulation in mice. Results are expressed as mean ± SEM, n = 5. Antisecretory effect is expressed as Pi/Pm × 1000 g, where Pi is the weight of the small intestine and Pm is the weight of mouse; #p < 0.001 versus saline group, ∗∗∗p < 0.001 versus castor oil group, one-way analysis of variance with Tukey's post hoc test.
Figure 3
Figure 3
Concentration-dependent inhibitory effect on spontaneous and K+ (80 mM) induced contractions of Manilkara zapota extracts: (a) chloroform (Mz.CHCl3), (b) aqueous (Mz.Aq), (c) papaverine, and (d) verapamil in isolated tissue preparations. Results are expressed as mean ± SEM (n = 3–5).
Figure 4
Figure 4
Gross-appearance of gastric mucosa in rats: (a) pretreated with saline, 10 mL/kg, severe injuries are seen, as ethanol (1 mL/100 g) produced excessive hemorrhagic necrosis of gastric mucosa pretreated with Manilkara zapota extracts: (b–d) chloroform (Mz.CHCl3), (e–g) aqueous (Mz.Aq) at doses of 50, 100, and 300 mg/kg, respectively, and (h) pretreated with omeprazole (20 mg/kg). The injuries were reduced with the increase of Mz.CHCl3 and Mz.Aq doses and omeprazole compared with ulcer control. At 300 mg/kg, Mz.CHCl3 and Mz.Aq showed most efficacious gastroprotective action.
Figure 5
Figure 5
Effect of Manilkara zapota extracts: chloroform (Mz.CHCl3) and aqueous (Mz.Aq) in comparison with saline, ethanol, and omeprazole groups on IL-18 in ethanol-induced ulcer model. IL-18 levels were measured by ELISA assay. ∗p < 0.05, ∗∗p < 0.01, and #p < 0.001. Analyzed by one-way ANOVA followed by Tukey's post hoc test.
Figure 6
Figure 6
Effects of Mz.Aq extract on ethanol-mediated stomach histopathologic changes. Stomach tissues (n = 5) from each experimental group were processed for histological evaluation at 1 hour after ethanol challenge. (a) Representative histological changes of apoptotic markers (TNF-α, P-NFκB, and COX-2) in stomach, scale bar = 20 μm, magnification 40x: control group, ethanol group, Mz.Aq extract group, and omeprazole group. (b, c) Severity scores of stomach injury in different groups (n = 5) calculated via relative density of value (AU) and relative integrated density. Data were analyzed by two-way ANOVA followed by Tukey's post hoc test using GraphPad Prism software. Mean ± SEM. ∗∗∗p < 0.001 versus ethanol group; ∗p < 0.01 versus ethanol group. (d) Histopathological examination of saline group, ethanol group, test group, and omeprazole group.
Figure 7
Figure 7
(a, b) The interactions of methyl chlorogenate and phenylephrine against the target adrenergic α1 receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 8
Figure 8
(a, b) The interactions of methyl chlorogenate and pirenzepine against the target muscarinic M1 receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 9
Figure 9
(a, b) The interactions of methyl chlorogenate and atropine against the target muscarinic M3 receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 10
Figure 10
(a, b) The interactions of methyl chlorogenate and domperidone against the target dopaminergic D2 receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 11
Figure 11
(a, b) The interactions of methyl chlorogenate and calmidazolium against the target calmodulin receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 12
Figure 12
(a, b) The interactions of methyl chlorogenate and verapamil against the target voltage gated L-type calcium channels, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 13
Figure 13
(a, b) The interactions of methyl chlorogenate and omeprazole against the target H+/K+ ATPase receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 14
Figure 14
(a, b) The interactions of methyl chlorogenate and ranitidine against the target histaminergic H2 receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 15
Figure 15
(a, b) The interactions of methyl chlorogenate and loperamide against the target mu-opioid receptor, respectively, evaluated through Biovia Discovery Studio 2016.
Figure 16
Figure 16
(a, b) The interactions of methyl chlorogenate and papaverine against the target phosphodiesterase enzyme, respectively, evaluated through Biovia Discovery Studio 2016.
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