Investigation of the effect of curcumin on oxidative stress, local inflammatory response, COX-2 expression, and microvessel density in  induced enteritis, myositis and myocarditis in mice

A. M. R. Hamed; I. R. Abdel-Shafi; M. D. A. Elsayed; A. M. Mahfoz; S. E. Tawfeek; M. S. A. Abdeltawab

Open Access

Investigation of the effect of curcumin on oxidative stress, local inflammatory response, COX-2 expression, and microvessel density in Trichinella spiralis induced enteritis, myositis and myocarditis in mice

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| May 04, 2022

Volume 59 (2022): Issue 1 (March 2022)

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Published Online: May 04, 2022

Page range: 18 - 36

Received: Jun 01, 2021

Accepted: Nov 30, 2021

DOI: https://doi.org/10.2478/helm-2022-0002

Keywords
curcumin, oxidative stress, cyclooxygenase-2, CD34

© 2022 A. M. R. Hamed, I. R. Abdel-Shafi, M. D. A. Elsayed, A. M. Mahfoz, S. E. Tawfeek, M. S. A. Abdeltawab, published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Schematic presentation of the research procedure

Bar chart showing means and standard deviations of the adult T. spiralis worm count per 100 mL of intestinal fluid in the different study groups. (Alb/Cur150mg= combined albendazole and curcumin 150mg/Kg regimen group); *significant difference between groups at P<0.05

Bar chart showing means and standard deviations of the T. spiralis larval count per gram muscle tissue in the different study groups. (Alb/Cur150mg= combined albendazole and curcumin 150mg/Kg regimen group); *significant at difference between groups P<0.05

Bar chart showing means and standard deviations of serum MDA levels in nmol/mL during the intestinal phase (I) and muscular phase (M) of T.spiralis infection in the different study groups. (Alb/Cur150mg= combined albendazole and curcumin 150mg regimen group);*significant difference between groups at P<0.05

H&E-stained sections of the intestinal phase of (a,b) Group 1 (Control non-infected group) showing the normal intestinal villous structure. (c,d) Group 2 (Infected non-treated group) showing adult worms in the intestine (ad) encroaching on and distorting the intestinal villi, dense mononuclear inflammatory infiltration (I), cut section in the adult embedded within the intestinal villi (arrow heads) and goblet cell proliferation (G) (e,f) Group 3 (Albendazole-treated group) showing improvement in the intestinal villous architecture with moderate infiltration with inflammatory cells (I), (g,h) Group 4 (Curcumin 150mg-treated group) showing moderate sub-epithelial inflammatory infiltration and few embedded larvae (arrow head), (I,j) Group 5 (Curcumin 300mg-treated group) showing intact intestinal villi and mild cellular infiltrate, (k,l) Group 6 (Combined Curcumin 150 mg + Albendazole group) showing healthy intestinal villi with a core of connective tissue, normal epithelial covering, intact brush border, and few goblet cells (G) (Scale bar 50μm).

H&E-stained sections of the muscle phase of (a) Group 1 (Control non-infected group) showing normal architecture of the gastrocnemius muscle, diaphragm and myocardium. (b) Group 2 (Infected non-treated group) showing multiple depositions of T. spiralis encysted larvae (L) embedded in the gastrocnemius and diaphragm musculature. The cysts appear surrounded by a thick wall and granuloma formation. The myocardium is infiltrated by mononuclear inflammatory cells (I), (c) Group 3 (Albendazole-treated group) showing degenerated larvae (L) surrounded by dense inflammatory cells (I). Scattered inflammatory foci (f) are noticed in the myocardium. (d) Group 4 (Curcumin 150mg-treated group) showing moderate inflammatory infiltration (I) in the muscle and diaphragm in addition to mild separation of the myocardial fibrils (arrow heads), (e) Group 5 (Curcumin 300mg-treated group) showing marked improvement with completely degenerated larvae in the gastrocnemius muscle, limited inflammatory cells in the diaphragm and minimal separation of myofibrils in the heart (arrow heads). Group 6 (Combined Curcumin 150 mg + Albendazole group) showing degenerated larvae (L) invaded by inflammatory infiltration(Scale bar 50μm).

COX-2 immunohistochemistry in the intestinal phase of (a) Group 1 (Control non-infected group), (b) Group 2 (Infected non-treated group), (c) Group 3 (Albendazole-treated group), (d) Group 4 (Curcumin 150mg-treated group), (e) Group 5 (Curcumin 300mg-treated group), (f) Group 6 (Combined Curcumin 150 mg + Albendazole group) showing strong positive COX-2 immuno-reactivity in the intestinal mucosa and in the inflammatory cells of the T. spiralis-infected group and moderate cytoplasmic immunohistochemical expression in the curcumin-treated groups. Weak cytoplasmic COX-2 expression was noticed in the lamina propria and intestinal epithelial cells of the combined curcumin and albendazole group (Scale bar 50μm).

COX-2 immunohistochemistry in the muscle phase of (a) Group 1 (Control non-infected group) showing no immunostaining, (b) Group 2 (Infected non-treated group) showing strong positive immunoreactivity around encapsulated T. spiralis larvae and in the cellular infiltrates, (c) Group 3 (Albendazole-treated group) showing mild COX-2 immunostaining, (d) Group 4 (Curcumin 150mg-treated group) showing moderate COX-2 immunostaining in the muscle fibers around degenerated larvae, (e) Group 5 (Curcumin 300mg-treated group) showing distinct positive COX-2 immunostained cells in the muscle fibers surrounding the degenerated larvae, (f) Group 6 (Combined Curcumin 150 mg + Albendazole group) showing very minimal COX-2immunohistochemical expression (Scale bar 50μm).

COX-2 immunohistochemistry in the myocardium of (a) Group 1 (Control non-infected group), (b) Group 2 (Infected non-treated group), (c) Group 3 (Albendazole-treated group), (d) Group 4 (Curcumin 150mg-treated group), (e) Group 5 (Curcumin 300mg-treated group), (f) Group 6 (Combined Curcumin 150 mg + Albendazole group) showing strong positive Cox-2 immuno-reactivity in the myocardial fibers of the T. spiralis-infected group and moderate cytoplasmic immunohistochemical expression in the curcumin-treated groups. Weak cytoplasmic COX-2 expression is noticed in the myofibrils of combined curcumin and albendazole group (Scale bar 50μm).

CD34 immunohistochemistry in the intestinal phase of (a) Group 1 (Control non-infected group), (b) Group 2 (Infected non-treated group), (c) Group 3 (Albendazole-treated group), (d) Group 4 (Curcumin 150mg-treated group), (e) Group 5 (Curcumin 300mg-treated group), (f) Group 6 (Combined Curcumin 150 mg + Albendazole group) showing marked increase in CD34 immuno-expression in the inflammatory cells and capillaries of the infected group. Albendazole and curcumin treatment showed a reduction in the CD34 immuno-reactive capillaries and inflammatory cells (Scale bar 50μm).

CD34 immunohistochemistry in the muscle phase of (a) Group 1 (Control non-infected group), (b) Group 2 (Infected non-treated group), (c) Group 3 (Albendazole-treated group), (d) Group 4 (Curcumin 150mg/kg-treated group), (e) Group 5 (Curcumin 300mg/Kg-treated group), (f) Group 6 (Combined Curcumin 150 mg/kg + Albendazole group) showing marked increase in CD34 immunohistochemistry in the dense inflammatory cellular infiltration and capillary endothelial surrounding the encysted larvae (L) in the infected group. The albendazole-treated group shows moderate CD34 expression in the epimysium surrounding a degenerated larva (L). The curcumin 150 mg/Kg group and the curcumin 300 mg/Kg group show moderate CD34 immunohistochemical expression in the capillary network and inflammatory cells invading the disintegrated larvae. The combined curcumin 150 mg/kg + albendazole group shows few CD34 positive capillaries around homogenized larvae (L) with degenerated capsules (Scale bar 50μm).

CD34 immunohistochemistry in the myocardium of (a) Group 1 (Control non-infected group), (b) Group 2 (Infected non-treated group), (c) Group 3 (Albendazole-treated group), (d) Group 4 (Curcumin 150mg/kg-treated group), (e) Group 5 (Curcumin 300mg-treated group), (f) Group 6 (Combined Curcumin 150 mg/kg + Albendazole group) showing numerous CD34 positive immuno-reactive capillaries in the T. spiralis-infected group. Moderate CD34 expression is noticed in the curcumin 150 mg/kg and curcumin 300 mg/kg-treated groups, while minimal CD34 immunoreactivity is noticed in the combined curcumin 150 mg/kg and albendazole-treated groups (Scale bar 50μm).

Mean expression levels of inflammatory changes during the muscle phase of infection; (0) signifies no change, (1) mild, (2) moderate changes and (3) severe changes.

Muscle phase

	Study groups	Fibrosis	Capsular inflammation	Interstitial inflammation	Muscle degeneration	Larval degeneration
Mean	Negative control(NC)	0.00	0.00	0.00	0.00	0.00
	Positive control(1)	3.00	3.00	3.00	1.00	0.00
	Curcumin150mg(2)	2.67	2.83	2.83	0.83	0.17
	Curcumin300mg(3)	0.67	0.83	0.83	0.00	0.83
	Albendazole(4)	2.83	2.33	2.67	0.83	1.00
	Alb/Cur150mg(5)	2.00	2.17	2.33	0.50	0.50

Pearson r correlation test between COX-2 and CD34 expression detected by IHC.

Tissue sections	Pearson r correlation test between COX-2 and CD34 expression detected by IHC
	Drug groups	Pearson r value	P value
Intestinal tissue sections	Positive control	NaN	NaN
	Curcumin 150 mg/kg	-0.250	0.633
	Curcumin 300 mg/kg	0.500	0.312
	Albendazole 50 mg/kg	-0.500	0.312
	Combined curcumin 150 mg/kg and albendazole	NaN	NaN

Skeletal muscle tissue sections	Positive control	0.944	<0.001^*
	Curcumin 150 mg/kg	NaN	NaN
	Curcumin 300 mg/kg	NaN	NaN
	Albendazole 50 mg/kg	NaN	NaN
	Combined curcumin 150 mg/kg and albendazole	-0.500	0.312

Cardiac muscle tissue sections	Positive control	0.944	<0.001
	Curcumin 150 mg/kg	NaN	NaN
	Curcumin 300 mg/kg	NaN	NaN
	Albendazole 50 mg/kg	NaN	NaN
	Combined curcumin 150 mg/kg and albendazole	-0.500	0.312

Means and standard deviations of CD34 expression levels in sections from the small intestine, skeletal muscle and heart of the different study groups.

CD34expression (Mean ± SD)

Tissue samples	Small intestine	Skeletal muscle	Cardiac muscle
Study groups
Negative control group	0.7±0.5	0.0±0.0	0.0±0.0
Positive control group	2.7±0.5	2.7±0.4	2.7±0.4
Curcumin 150 mg/kg	1.3±0.5	1.7±0.5	1.7±0.5
Curcumin 300 mg/kg	1.7±0.5	1.7±0.5	1.7±0.5
Albendazole 50 mg/kg	0.7±0.5	1.0±0.0	1.0±0.0
Albendazole 50 mg/kg and curcumin 150 mg/kg	1.0±0.0	0.7±0.5	0.7±0.5

Mean expression levels of inflammatory changes during the intestinal phase of infection; (0) signifies no change, (1) mild, (2) moderate changes and (3) severe changes.

Intestinal phase

	Study groups	Inflammation	Goblet cells	Apoptosis	Fibrosis	Oedema
Mean	Negative control(NC)	1.00	0.17	0.00	0.17	0.00
	Positive control(1)	3.00	1.00	1.00	2.00	1.00
	Curcumin150mg(2)	2.83	1.00	0.00	1.67	1.00
	Curcumin300mg(3)	0.83	0.17	0.00	0.17	0.17
	Albendazole(4)	1.33	0.67	0.33	1.00	0.50
	ALb/Cur150mg(5)	1.17	0.00	0.33	0.33	0.67

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Investigation of the effect of curcumin on oxidative stress, local inflammatory response, COX-2 expression, and microvessel density in Trichinella spiralis induced enteritis, myositis and myocarditis in mice

Article Category: Research Article

Published Online: May 04, 2022

Page range: 18 - 36

Received: Jun 01, 2021

Accepted: Nov 30, 2021

DOI: https://doi.org/10.2478/helm-2022-0002

Keywords
curcumin, oxidative stress, cyclooxygenase-2, CD34

© 2022 A. M. R. Hamed, I. R. Abdel-Shafi, M. D. A. Elsayed, A. M. Mahfoz, S. E. Tawfeek, M. S. A. Abdeltawab, published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

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Mean expression levels of inflammatory changes during the muscle phase of infection; (0) signifies no change, (1) mild, (2) moderate changes and (3) severe changes.

Pearson r correlation test between COX-2 and CD34 expression detected by IHC.

Means and standard deviations of CD34 expression levels in sections from the small intestine, skeletal muscle and heart of the different study groups.

Mean expression levels of inflammatory changes during the intestinal phase of infection; (0) signifies no change, (1) mild, (2) moderate changes and (3) severe changes.

Investigation of the effect of curcumin on oxidative stress, local inflammatory response, COX-2 expression, and microvessel density in Trichinella spiralis induced enteritis, myositis and myocarditis in mice

Article Category: Research Article

Published Online: May 04, 2022

Page range: 18 - 36

Received: Jun 01, 2021

Accepted: Nov 30, 2021

DOI: https://doi.org/10.2478/helm-2022-0002

Keywordscurcumin, oxidative stress, cyclooxygenase-2, CD34

© 2022 A. M. R. Hamed, I. R. Abdel-Shafi, M. D. A. Elsayed, A. M. Mahfoz, S. E. Tawfeek, M. S. A. Abdeltawab, published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Fig 1

Fig 2

Fig 3

Fig 4

Fig 5

Fig 6

Fig 7

Fig 8

Fig 9

Fig 10

Fig 11

Fig 12

Mean expression levels of inflammatory changes during the muscle phase of infection; (0) signifies no change, (1) mild, (2) moderate changes and (3) severe changes.

Pearson r correlation test between COX-2 and CD34 expression detected by IHC.

Means and standard deviations of CD34 expression levels in sections from the small intestine, skeletal muscle and heart of the different study groups.

Mean expression levels of inflammatory changes during the intestinal phase of infection; (0) signifies no change, (1) mild, (2) moderate changes and (3) severe changes.

Keywords
curcumin, oxidative stress, cyclooxygenase-2, CD34