Effects of Curcuma longa Rhizoma on MIA-induced Osteoarthritis in Rat Model

Article information

J Korean Med. 2019;40(3):35-58

Publication date (electronic) : 2019 September 30

doi : https://doi.org/10.13048/jkm.19026

Department of Rehabilitation Medicine of Korean Medicine, College of Korean Medicine, Daegu Haany University

Correspondence to: Young Jun Kim, Department of Rehabilitation Medicine of Korean Medicine, Daegu Haany University Pohang Korean Medicine Hospital, 411, Saecheonnyeon-daero, Nam-gu, Pohang-si, Gyeongsangbuk-do, 37685, Korea, Tel: +82-54-281-0055, Fax: +82-54-281-7463, E-mail: kimyj@dhu.ac.kr

Received 2019 July 2; Revised 2019 August 7; Accepted 2019 August 13.

Abstract

Objectives

The aim of this study was to investigate the anti-inflammatory effects of Curcuma longa rhizoma extract in an experimental rat model of osteoarthritis.

Methods

Osteoarthritis was induced in rats by injecting monosodium iodoacetate (MIA) into the knee joint cavity of rats. The rats were divided into 5 groups (Normal, Control, positive comparison, low (CL) and high (CH) concentration groups). Rats in the low concentration (CL) group had MIA-induced osteoarthritis; they were treated with Curcuma longa rhizoma extract at a dose of 50mg/kg body weight. Rats in the high concentration (CH) group had MIA-induced osteoarthritis; they were treated with Curcuma longa rhizoma extract at a dose of 100mg /kg body weight. Hind paw weight distribution and ROS levels were measured. At the end of all treatments, changes in alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), and creatinine levels were analyzed. In addition, inflammatory protein levels were evaluated by western blot analysis.

Results

In this study, hind paw weight distribution significantly improved in the CL and CH groups, while. Reactive oxygen species (ROS) production significantly decreased in both. The levels of ALT, AST, BUN, and creatinine did not significantly change in either group. The production of nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4), p47^phox, and Ras-related C3 botulinum toxin substrate 1 (RAC1) decreased in both. Catalase, heme oxygenase-1 (HO-1) and superoxide dismutase (SOD) significantly increased in the CL and CH groups, respectively. Nuclear factor erythroid 2 (Nrf2) increased, but there were no significant differences between the experimental and control groups. Inflammatory cytokines, including nuclear factor-kappa Bp65 (NF-κBp65), interleukin-1beta (IL-1β), and tumor necrosis factor-alpha (TNF-α), decreased significantly in both the CL and CH groups.

Conclusions

Our results showed that Curcuma longa rhizoma extract has anti-inflammatory effects. Anti-inflammatory activity is regulated by the inhibition of inflammatory cytokines and mediators, such as NF-κB, therefore, it suppresses cartilage damage as well.

Keywords: Curcuma longa rhizoma extract; Osteoarthritis; Anti-inflammation; Monosodium iodoacetate (MIA)

Fig. 1

HPLC profile of combined extract of Curcuma longa rhizoma 30% EtOH extract.

(a) Chemical structure of Curcumin (C₂₁H₂₀O₆), (b) Linearity curve of curcumin, (c) HPLC profile of Curcumin and Curcuma longa rhizoma 30% EtOH extract.

Fig. 2

Alanine transaminase (ALT) in serum.

Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group. There is no significant difference in the experimental groups.

Table 2

Body Weight and Food Intake

Fig. 3

Aspartate transaminase (AST) in serum.

Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group. #, Significantly different from the normal (##, p<0.01)

Fig. 4

Blood urea nitrogen (BUN) in serum.

Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group. There is no significant difference in the experimental groups.

Fig. 5

Creatinine in serum.

Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group.There is no significant difference in the experimental groups.

Fig. 6

Scavenging activity of Curcuma longa rhizoma 30% EtOH extract on DPPH radical.

(A) Scavenging activity of L-ascorbic acid against DPPH radical, (B) Scavenging activity of Curcuma longa rhizoma 30% EtOH extract against DPPH radical.

Fig. 7

Scavenging activity of Curcuma longa rhizoma 30% EtOH extract on ABTS radical.

(A) Scavenging activity of L-ascorbic acid against ABTS radical, (B) Scavenging activity of Curcuma longa rhizoma 30% EtOH extract against ABTS radical.

Fig. 8

Oxidative stress biomarker (ROS) in serum.

Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group. #, Significantly different from the normal (###, p<0.001). *, Significantly different from the control (***, p<0.001).

Fig. 9

Western blot analysis of NADPH oxidase 4 (NOX4) expression.

Expression levels in the MIA-induced osteoarthritis rats. Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group. #, Significantly different from the normal (##, p<0.01).

Fig. 10

Western blot analysis of p47^phox expression.

Expression levels in the MIA-induced osteoarthritis rats. Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group. There is no significant difference in the experimental groups.

Fig. 11

Western blot analysis of Ras-related C3 botulinum toxin substrate 1 (RAC1) expression.

Fig. 12

Western blot analysis of nuclear factor erythroid 2 (Nrf2) expression.

Fig. 13

Western blot analysis of glutathione heme oxygenase-1 (HO-1) expression.

Expression levels in the MIA-induced osteoarthritis rats. Refer to Table 2 for groups. All data are expressed as the means ± SEM, n=6 rats per group. *, Significantly different from the control (*, p<0.05).

Fig. 14

Western blot analysis of superoxide dismutase (SOD) expression.

Fig. 15

Western blot analysis of catalase expression.

Fig. 16

Western blot analysis of glutathione peroxidase (GPx) expression.

Fig. 17

Western blot analysis of nuclear factor-kappa B p65 (NF-κBp65) expression.

Fig. 18

Western blot analysis of inducible nitric oxide synthase (iNOS) expression.

Fig. 19

Western blot analysis of tumor necrosis factor-alpha (TNF-α) expression.

Fig. 20

Western blot analysis of interleukin-1beta (IL-1β) expression.

Fig. 21

Western blot analysis of matrix metalloproteinase-2 (MMP-2) expression.

Fig. 22

Western blot analysis of matrix metalloproteinase-9 (MMP-9) expression.

Fig. 23

Western blot analysis of tissue inhibitor of metalloproteinases 1 (TIMP1) expression.

Fig. 24

Western blot analysis of tissue inhibitor of metalloproteinases 2 (TIMP2) expression.

Fig. 25

The histological analysis of the knee joint tissues after treatment of CL and CH in MIA-induced osteoarthritis rats (H&E staining, ×200).

In the normal, synovial tissue and cartilage were well observed. In the control, most of aricular cartilage and synovial tissue were destructed. In the CL & CH, articular cartilage and synovial tissue were preserved, compared with the control. (A) Nor, (B) Con, (C) Indo, (D) CL, (E) CH. Refer to Table 2 for groups.

Fig. 26

The histological analysis of the knee joint tissues after treatment of CL and CH in MIA-induced osteoarthritis rats (Safranin O staining, × 200).

In the normal, there are much of proteoglycan around the cartilage. In the control, most of proteoglycan was broken down. In the CL & CH, proteoglycan was well preserved compared with the control. Especially, CH group showed histologically similar to normal group. (A) Nor, (B) Con, (C) Indo, (D) CL, (E) CH. Refer to Table 2 for groups.

Table 1

The Operating Condition of HPLC for Analysis of Curcumin

Table 3

The Changes in Relative Hind Paw Weight Distribution in MIA-induced Osteoarthritis Rats

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Group	Body weight			Food intake (g/day)

	Initial (g)	Final (g)	Gain (g)
Nor	329.91 ± 6.53	373.82 ± 9.04	43.91 ± 5.63	22.76 ± 1.84
Con	310.73 ± 12.47^###	355.64 ± 16.45#	44.91 ± 6.88	20.74 ± 3.03
Indo	312.00 ± 14.61	356.00 ± 27.56	44.00 ± 17.26	20.31 ± 1.00
CL	305.78 ± 8.54	339.56 ± 19.49	33.78 ± 20.33	21.85 ± 1.14
CH	315.33 ± 12.94	360.22 ± 15.87	44.89 ± 6.15	22.18 ± 0.97

Group	0 weeks	1 weeks	2 weeks	3 weeks
Nor	100.18 ± 14.97	98.66 ± 12.03	97.45 ± 10.89	102.94 ± 21.87
Con	106.90 ± 15.84	180.14 ± 49.22^##	279.32 ± 55.82^###	644.25 ± 239.77^###
Indo	103.25 ± 13.10	204.26 ± 36.02	191.39 ± 77.62*	127.89 ± 54.03^***
CL	109.11 ± 14.74	194.94 ± 72.15	258.16 ± 103.42	257.79 ± 62.75^***
CH	97.47 ± 13.11	193.96 ± 69.76	230.36 ± 108.48	160.98 ± 65.47^***