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ORIGINAL ARTICLE
Ahead of print publication  

Fresh juice and extract of Drynaria quercifolia rhizome nutraceutical for the management of arthritis with special emphasis on enzyme levels


1 Organic and Bio Organic Chemistry Division, Central Leather Research Institute (Council of Scientific and Industrial Research), Chennai, Tamil Nadu, India; Department of Pharmacology, College of Pharmacy, National University of Science and Technology, Muscat, Sultanate of Oman
2 Organic and Bio Organic Chemistry Division, Central Leather Research Institute (Council of Scientific and Industrial Research), Chennai, Tamil Nadu, India

Date of Submission15-Mar-2020
Date of Decision01-May-2020
Date of Acceptance06-Jul-2020
Date of Web Publication17-Dec-2020

Correspondence Address:
Rajagopal Srinivasan,
Organic and Bio Organic Chemistry Division, Central Leather Research Institute (Council of Scientific and Industrial Research), Chennai 600020, Tamil Nadu.
India
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jpbs.JPBS_227_20

   Abstract 

Objective: The objective of this study was to screen fresh and concentrated juice extract of rhizome of Drynaria quercifolia (L.) for its anti-arthritic property using in vivo model. Materials and Methods: In vivo anti-arthritic effect was evaluated by complete Freund’s adjuvant (CFA)-induced arthritic model. Hematological and biochemical parameters were evaluated in the arthritic model with and without treatment. Assays of enzymatic and nonenzymatic antioxidants, lysosomal enzymes, and protein-bound carbohydrates were performed using the standard protocol. Histopathological examinations were carried out in different organ tissues. Results: Decrease in paw volume was noticed from 16th day and a significant reduction in paw volume was observed after 16th day in Drynaria Quercifolia Rhizome (DQR) (extract and fresh juice)-treated and methotrexate (MTx)-treated groups. Levels of hemoglobin, PCV (packed cell volume), and platelets were normal in all groups. Significant (P < 0.05) increase of TC (total count) and decrease of red blood cells were observed in the arthritic group (negative control) when compared with normal control. There was a significant change in liver enzyme levels. The activities of enzymatic antioxidants such as catalase, dismutase, and peroxidase in tissue homogenates also showed positive results. There was a significant (P < 0.05) increase in levels of lysosomal enzymes (alkaline phosphatase, acid phosphatase, and cathepsin D) in the arthritic control animals. These enzymes levels are being able to control by the treatment with DQR (extract and fresh juice) and standard drug (MTx). Histology reports confirm the presence of only a mild cartilage erosion in DQ fresh juice-treated group. Conclusion: Our findings pointed out the promising effect of fresh juice and extract of D. quercifolia Rhizome for the management of arthritis.

Keywords: Anti-arthritic, antioxidants, extract, fresh juice, in vivo



How to cite this URL:
Dhanalekshmi UM, Gowri T, Ramya B, Srinivasan R. Fresh juice and extract of Drynaria quercifolia rhizome nutraceutical for the management of arthritis with special emphasis on enzyme levels. J Pharm Bioall Sci [Epub ahead of print] [cited 2021 Jan 17]. Available from: https://www.jpbsonline.org/preprintarticle.asp?id=303526




   Introduction Top


Rheumatoid arthritis (RA) is caused by the disturbance of the inflammation process in the body especially in joints mainly by an autoimmune disorder and it leads to pain, swelling, and joint stiffness.[1] In the long stretch, patients may expose to symmetrical and bilateral disabilities in the hands, wrists, and knees.[1] In 2017, it has been estimated that more than 20 million people worldwide were living with RA, with over a million new cases diagnosed each year.[2] Globally, the age-standardized point prevalence and annual incidence rates of RA were increased by 7.4%.[3] Different mechanisms are involved in the pathophysiology of severe pain in RA and it mainly includes the inflammation process of the articular and periarticular structures through peripheral and central pain pathways. Arthritis pain also involves multimodality changes in nociceptive processing in the spinal cord and brain.[4] Central sensitization, in contrast to peripheral sensitization, represents an abnormal state of responsiveness or increased gain of the nociceptive system.[5] Studies show that there is a discordance between pain severity and the degree of joint damage.[5] In addition to the conventional therapy, the role of cannabinoid receptors 1 (CB-1) and gamma-aminobutyric acid (GABA) gene expression were studied as an anti-nociceptive pathways in the cartilage tissue of rats with arthritis.[6] In the case of RA, many of the commonly used drugs are becoming less acceptable due to serious adverse effects. Therapeutic treatment regimen with existing therapies produces harmful consequences which may counterbalance valuable results overtime. This necessitates the continued search for potent anti-arthritic agents with reduced or without any side effects. Studies based on the ethnobotanical use of plants have often proved to be a more efficient method of drug discovery. According to World Health Organization (WHO), still 80% of the world population rely on the plant-derived drugs and WHO has also recommended the evaluation of the effectiveness of plants in conditions where we lack safe modern drugs.[7]

Drynaria quercifolia (L.) J. Smith (Polypodiaceae) is an epiphytic medicinal pteridophyte, distributed widely in the evergreen forests of the Western Ghats of Kerala, locally called “Marappannakizhangu” or “Attukalkizhangu.” The rhizome is reported to be used by tribal communities of Tamil Nadu and Kerala to cure various diseases. Ethnomedicinal information shows that the rhizome of plant D. quercifolia (L.) J. Smith was traditionally used for the treatment of body pain, knee pain, joint pain, dyspepsia, diarrhea, typhoid, cholera, chronic jaundice, fever, and headache, and skin diseases and is believed to play a major role in the management of inflammatory processes.[8] However, no systematic investigations have been carried out to analyze the anti-arthritic activity of D. quercifolia Rhizome. In vitro anti-arthritic activity and in vivo toxicity study of Drynaria Quercifolia Rhizome (DQR) fresh juice and extract have been evaluated and reported by our group (IJPSR/RA-14019/01-20-in press). Hence, in this study an attempt has been made to screen the rhizome of D. quercifolia (extract and fresh juice) for its anti-arthritic property in in vivo CFA-induced arthritic rat model.


   Materials and Methods Top


Chemicals

Complete Freund’s adjuvant (CFA), epinephrine, 4,6 diphenyl 1,10 phenanthrolin, hemoglobin (Hb), DTNB, superoxide dismutase (SOD), acetylacetone, chloramine T were purchased from Sigma-Aldrich, St. Louis, Missouri. Ferric chloride and thiobarbituric acid (TBA) were purchased from Loba Chemicals, Mumbai, India. EDTA, p-nitrophenyl phosphate, sodium acetate trihydrate, reduced glutathione, copper sulfate, Folin–Coicalteau reagent, sodium potassium tartrate, 2,4 dinitro phenylhydrazine, ascorbic acid, pyridine, glycine, p-nitrophenol, Tris-Hcl buffer, p-dimethyl amino benzaldehyde, and trichloroacetic acid were purchased from Sisco Research Laboratory, Mumbai, India. All the other chemicals used were of analytical grade.

Sample collection and preparation

Fresh rhizome of D. quercifolia was collected from Kolli Hills, Namakkal District, Tamil Nadu, India. The fur portion was removed from the rhizome. The plant material was authenticated by Prof. V. Chelladurai, Research Officer (Retd), Govt. Siddha College, Tirunelveli, India. A voucher specimen (CLRICSIR/2011/09) has been preserved at our laboratory for future reference. The flesh part was isolated, sliced, and minced and press filtered. The resulting juice was centrifuged at 4000 rpm for 10 min. The supernatant liquid was concentrated under reduced pressure and lyophilized to obtain a brown colored free flow residue (crude). The residue was stored in a refrigerator at 4ºC–5ºC until use (2 kg flesh part gave 1000 mL of juice which gave 27.87 g of crude extract. The percentage yield was 2.8%). The crude extract was dissolved in water/phosphate buffer saline to required concentration and used for the experiments.

Experimental animals

This study was conducted after obtaining approval from the Institutional Animal Ethical Committee (IAEC. No: 15/01/AY18) and this protocol met the requirements of national guidelines of CPCSEA. Female albino Wistar rats with a bodyweight of 120–170 g approximately (average) used for this study were procured from King’s Institute, Guindy, Chennai, India and housed in the Institutional animal house under standard environmental conditions (23°C ± 1°C, 55% ± 5% humidity and 12h/12h light/dark cycle) and maintained with free access to standard diet (Hindustan Lever, Bangalore, India) and water ad libitum.

Anti-arthritic activity

Anti-arthritic activity was evaluated by the CFA-induced arthritic method.[9],[10] Arthritis was induced by a single intradermal injection of 0.1 mL of CFA containing 10mg/mL dry heat-killed Mycobacterium tuberculosis in sterile paraffin oil into the left hind paw of rats. After 14 days of CFA injection, rats were evaluated for arthritis development and then they were divided into five groups of six animals each. The details of experimental groups are mentioned in [Table 1]. The change in body weight and the swelling (using plethysmometer) in hind paws were examined for every 4 days alternatively during the course of experiment, that is, 0, 4, 8, 12, 16, 20, 24, and 28 days. The animals in Groups III, IV, and V were treated with their respective doses of drugs from day 15th to 28th after induction of arthritis, 14 days of treatment as mentioned in [Table 1].
Table 1: Details of experimental groups

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On the 29th day, blood was collected from the retro-orbital sinus for hematological and biochemical estimation. The animals were sacrificed by the euthanasia method. The liver, lungs, kidney, heart, and hind limbs were removed for biochemical assays and histopathological examination (preserved in 10% formalin).

Measurement of paw swelling

Changes in the edema of the left hind paw were measured from the ankle using a plethysmometer.

Percentage inhibition of paw volume was determined as follows:



where VC is the paw volume after induction, V0 is the paw volume before induction, and Vt is the paw volume after treatment;

Hematological and biochemical parameters

The hematological parameters such as red blood cell (RBC) count, differential count (lymphocytes, neutrophils, monocytes, and polymorphs), Hb, packed cell volume (PCV), and platelet count were analyzed by using MDC 4000 vet 3 part hematology analyzer. The biochemical parameters such as sugar, cholesterol, triglycerides (TGL), creatinine, serum glutamic pyruvic transaminase (SGPT), serum glutamic-oxaloacetic transaminase (SGOT), alkaline phosphatase (ALP), and bilirubin were analyzed by using Transasia Smart Batch Analyzer EM 200.

Biochemical assays using tissue homogenates

Tissue samples collected from all groups of animals were minced into fine pieces and homogenized by using IKAT 25 Teflon-homogenizer in 0.1M Tris-HCl buffer with a pH 7.4.

The samples were centrifuged in a centrifuge at 10,000 rpm for 15 min and the resultant supernatant was stored at –70ºC for following biochemical assays.

Assays of enzymatic and nonenzymatic antioxidants and lipid peroxide

The enzymatic antioxidants catalase (CAT), SOD, and glutathione peroxidase (Gpx) were measured in tissue homogenates. The nonenzymatic antioxidants such as reduced glutathione (GSH), vitamin C, and vitamin E were determined in tissue homogenates. The extent of lipid peroxidation was measured in tissue homogenates.[11],[12],[13],[14],[15],[16],[17]

Assays of lysosomal enzymes and protein-bound carbohydrates

The lysosomal enzymes such as ALP, acid phosphatase (ASP), and cathepsin D levels were measured in tissue homogenates. The protein-bound carbohydrates such as hexosamine and uronic acid levels were determined in liver, kidney, and lung tissues homogenates.[18],[19],[20]

Assay of hydroxyproline and myeloperoxidase

The hydroxyproline level was estimated in liver, lungs, kidneys, and bone homogenates. MPO activity was assayed as an index of neutrophil infiltration into the joints and tissues.[21],[22]

Histopathological examinations

At the end of the 28th day, all the animals were sacrificed to collect hind limbs (interphalangeal joint). The hind limbs were rinsed in ice-cold 0.9% saline and were fixed in 10% formalin for histopathological examinations.

Statistical analysis

The statistical parameters were analyzed by two-way analysis of variance (ANOVA) (Bonferroni test). The results are expressed as mean ± SD. Significant difference between the groups was assigned at P < 0.05.


   Results Top


Bodyweight

Bodyweight was significantly decreased from 4th to 28th day in the negative control group, that is, arthritic control group when compared with the normal control group. Such a reduction in body weight was not observed in animals treated with the standard drug and D. quercifolia (extract and juice). The results are depicted in [Table 2].
Table 2: Change in body weight

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Paw volume

In arthritis-induced rats, there was an appreciable increase in paw volume in the injected hind legs within 3–5 days. A significant increase (P < 0.05) of paw volume was continued up to 20th day and the increase in volume was stabilized after the 20th day. By this time the tail had become noticeably thickened. Decrease in paw volume was noticed from the 16th day and a significant reduction in paw volume was observed after the 16th day in DQ (fresh juice and extract) and standard treated groups. The paw volume changes in control and experimental groups of rats are shown in [Table 3].
Table 3: Paw volume measurement

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Percentage inhibition of paw volume

The results of percentage inhibition of paw volume are depicted in [Table 4]. Percentage inhibition of paw volume showed a gradual increase in positive (III), DQ extract (IV), and DQ fresh juice (V) groups after treatment. Maximum percentage inhibition was observed on day 28 in all the treatment groups. At the end of the study, the percentage inhibition of DQ extract group (IV) was similar to the positive group (III).
Table 4: Percentage inhibition of paw volume after treatment

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Hematological analysis

[Table 5] represents the hematological changes associated with the arthritic condition after treatment with DQ. Levels of Hb, PCV, platelet, and DC were normal in all the groups. Significant (P < 0.05) increase of TC and decrease of RBC was observed in arthritic group when compared with the normal group.
Table 5: Hematological parameters

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Biochemical analysis

The results of biochemical analysis are shown in [Table 6]. There was a significant increase in ALP and SGPT levels in the negative control group when compared to normal control; this was reversed to normal level by both standard treatment and D. quercifolia treatment. There was a significant increase in SGOT level in the negative group, extract-treated group, and fresh juice-treated group when compared to normal control group.
Table 6: Biochemical parameters

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Bio chemical assays using tissue homogenates

Assays of enzymatic and nonenzymatic antioxidants and lipid peroxide

[Figure 1]A, [B], and [C] shows the activities of enzymatic antioxidants CAT, SOD, and Gpx in tissue homogenates. In negative control, there was a significant decrease catalase level (except heart tissue sample) when compared to normal control. This was significantly (P < 0.05) increased by the treatment with standard drug, extract, and fresh juice of DQ. But in liver homogenate there was no significant improvement when compared to control, after treatment with standard drug. In negative control, there was a significant decrease SOD and Gpx levels when compared to normal control. This was significantly increased by standard drug, extract, and fresh juice of DQ treatment. By comparing these enzyme levels, the role of DQ in free radical scavenging is well understood.
Figure 1: (A) Amount of catalase enzyme present in tissue homogenates. (B) Amount of SOD enzyme present in tissue homogenates. (C) Amount of Gpx enzyme present in tissue homogenates. The values are expressed as mean ± SD, n = 6. a denotes significant at the level of P < 0.05 comparison with control, b denotes significant at the level of P < 0.05 comparison with negative control, and c denotes significant at the level of P < 0.05 comparison with positive control

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There was a significant decrease in reduced glutathione, vitamin E, and vitamin C levels in the negative control group when compared to normal control group. This was reversed to normal level by the treatment with standard drug (MTx), extract and fresh juice of DQ. Data are represented in [Figure 2]A, [B], and [C].
Figure 2: (A) Amount of reduced glutathione enzyme present in tissue homogenates. (B) Amount of vitamin E present in tissue homogenates. (C) Amount of vitamin C present in tissue homogenates. (D) Amount of MDA present in tissue homogenates. The values are expressed as mean ± SD, n = 6. a denotes significant at the level of P < 0.05 comparison with control, b denotes significant at the level of P < 0.05 comparison with negative control, and c denotes significant at the level of P < 0.05 comparison with positive control

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[Figure 2D] shows the extent of LPO levels in tissue homogenates. In negative control group (arthritic group), there was a significant (P < 0.05) increase in lipid peroxide level compared to normal control were noted. This was significantly decreased to normal level by the treatment of DQ (extract and fresh juice) and standard drug (MTx).

Assays of lysosomal enzymes and protein-bound carbohydrates

There was a significant (P < 0.05) increase in levels of lysosomal enzymes (ALP, ASP, and cathepsin D) in the arthritic control animals (negative control group) when compared to normal control group. This was reversed to normal by the treatment with DQ (extract and fresh juice) and standard drug (MTx). The results are shown in [Figure 3]A, [B], and [C].
Figure 3: (A) Amount of alkaline phosphatase present in tissue homogenates. (B) Amount of acid phosphatase present in tissue homogenates. (C) Amount of cathepsin D present in tissue homogenates. The values are expressed as mean ± SD, n = 6. a denotes significant at the level of P < 0.05 comparison with control and b denotes significant at the level of P < 0.05 comparison with negative control

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In negative control group, the significant (P < 0.05) increase in glycoproteins (hexosamine and uronic acid) was observed when compared to normal control group. This was decreased by the treatment with DQ (extract and fresh juice) and standard drug (MTx). The results are represented in [Figure 4]A and [B].
Figure 4: (A) Amount of hexosamine present in tissue homogenates. (B) Amount of uronic acid present in tissue homogenates. The values are expressed as mean ± SD, n = 6. a denotes significant at the level of P < 0.05 comparison with control and b denotes significant at the level of P < 0.05 comparison with negative control

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Assay of hydroxyproline and myeloperoxidase

In negative control group, there was a significant increase in hydroxyproline level (in lung and bone samples) was observed when compared to normal control group. This was decreased by the treatment with DQ (extract and fresh juice) and standard drug (MTx). The results are represented in [Figure 5]A. There was a significant increase in myeloperoxidase level in the negative control group (both in lung and bone samples) when compared to normal control group. This was reversed to normal by the treatment with the DQ (extract) and standard drug but not with DQ fresh juice treatment. The results were shown in [Figure 5B].
Figure 5: (A) Amount of hydroxyproline present in tissue homogenates. (B) Amount of myeloperoxidase present in tissue homogenates. The values are expressed as mean ± SD, n = 6. a denotes significant at the level of P < 0.05 comparison with control, b denotes significant at the level of P < 0.05 comparison with negative control, and d* denotes significant at the level of P < 0.05 comparison with extract-treated group

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Histopathological examination of interphalangeal joints

[Figure 6] shows the histopathological examination of bone. Arthritic control group (II) showed the erosion of cartilage and synovial hyperplasia (L), DQ fresh juice-treated group showed the mild cartilage erosion (O). There were no abnormalities in normal control, positive control, and DQ extract groups.
Figure 6: Histopathological slides (100×) of bones of arthritic Wistar albino rats treated with plant extract and standard drug. K: NC = no abnormality, L = negative group (erosion of cartilage and synovial hyperplasia), M = positive group (no abnormality), N = DQ extract-treated group (no abnormality), O = DQ juice-treated-mild cartilage erosion

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   Discussion Top


In this study, we have investigated the anti-arthritic effect of D. quercifolia rhizome in CFA-induced arthritic rats. Results showed that DQ significantly inhibited the paw swelling and this inhibition was accompanied by an increase in antioxidants levels, decrease in the levels of lysosomal enzymes, glycoproteins, hydroxyproline, and myeloperoxidase. Increased paw swelling observed in adjuvant-induced arthritic rats was found to be the result of edema of periarticular tissues. An increase in granulocytes and monocytes has been associated with inflammation process and changes in ankle diameter.[23] Changes in body weight are useful index to assess the course of the disease and the response to therapy of anti-arthritic agents in question. The loss of body weight observed in arthritic animals may be due to the reduced absorption of glucose and leucin through the rat intestine and also due to pathophysiological homeostatic imbalances.[24] Significant increase in body weight in DQR-treated group when compared with negative control is due to restoration of absorption capacity of intestine in arthritic animals. This increase in absorption capacity may be due to the presence of phytochemicals and antioxidant components in DQ. It has formerly been reported that phenolics and flavonoids possess anti-inflammatory, antioxidant activities and plays a major role in intestinal mucin cells physiology.[25] It has been evident that presence of flavonoids and phenol plays a major role in the management of arthritis inflammation process. Preliminary phytochemical screening of appropriate solvent extracts of DQR confirmed the presence of sterols, tannins, proteins, and amino acids, flavonoids, terpenoids, saponin, and carbohydrates.[26] Details of phytochemical analyses and in vitro arthritic activity of the DQR collected from the Kolli hills were evaluated and reported by our team (article in press).

The results also showed that the DQ and standard drug methotrexate (MTx) significantly suppressed the paw swelling. At the end of the study, there was no significant difference in paw volume of DQR-treated and positive control group. This shows the effectiveness of our plant in the management of inflammation in arthritis.

The hematological parameters did not show any significant difference between the DQ-treated groups, MTx-treated group, and normal control. In arthritic control group, there was a statistically significant difference (P < 0.05) in Hb and TC on comparing with normal control. This shows that the DQ does not affect the hematological parameters.

The biochemical parameters, glucose, cholesterol, creatinine, TGL, bilirubin did not show any statistically significant difference (P < 0.05) between the groups. The SGOT, SGPT, and ALP were elevated in arthritic control group compared to normal control. The SGPT and ALP showed a significant difference in DQ-treated groups and MTx-treated groups when compared to normal control, but the levels were within the normal physiological limits.

The enzymatic antioxidants such as catalase, SOD, and GPx activity were reduced in the arthritic control group compared to normal control. But in DQ-treated group and MTx-treated group, there was no significant decrease compared to normal control group. The increased production of oxygen free radicals in adjuvant-induced arthritic rats leads to decreased activities of both enzymatic and nonenzymatic antioxidants as a consequence of their increased consumption during the oxidative stress and cellular lysis.[27]

The nonenzymatic antioxidants such as GSH, vitamin C, and vitamin E activity were increased in DQ-treated groups whereas in arthritic control group there was a significant (P < 0.05) decrease. GSH is an important antioxidant that has been shown to destroy ROS and other free radicals both by nonenzymatic as well as enzymatic mechanisms, as it acts as a substrate for GPx and GST during the breakdown of H2O2 and lipid peroxides.[28] The decreased levels of GSH in adjuvant-induced arthritic rats might also be due to the decreased GR (glutathione reductase) activity.[29] Vitamin C is the effective antioxidant because of its ability to fight against oxygen free radicals and other molecules which trigger rheumatoid inflammation and also serve as a cofactor in collagen synthesis, the main protein in joint tissue and bone. Vitamin E is the major chain-breaking antioxidant, in spite of its antioxidant property, vitamin E also plays a vital role in anti-inflammation by decreasing the expression of pro-inflammatory cytokines, prevention of the NF-kB activation cascade and consequently arresting pro-inflammatory gene expression.[30] It has been reported that flavonoids are a good inhibitor of arachidonic acid peroxidation.[31] They interfere with the radical scavenging system and also increase the function of endogenous antioxidant systems. Administration of DQR fresh juice and extract decreased the oxidative stress by enhancing the activities of both enzymatic and nonenzymatic antioxidants which may be due to the presence of flavonoids and phenols.

Lipid peroxidation was significantly decreased in DQ- and MTx-treated groups when compared to arthritic control group. Lipid peroxidation is considered as a critical mechanism of the injury that occurs during RA. The malondialdehyde (MDA) concentration in adjuvant-induced arthritic animals suggests the role of free radicals during phagocytosis of immune complexes in this inflammatory arthropathy.[32] This shows that administration of DQR inhibited the lipid peroxidation.

The lysosomal enzymes such as ALP, ACP, and cathepsin D were significantly elevated in arthritic group. There was a significant decrease in DQR- and MTx-treated groups when compared to arthritic group. Lysosomal enzymes are the main factors playing a vital role in tissue injury and repair, inflammation, phagocytosis and they participate in pathological processes such as inflammation, degeneration, and RA.[33],[34] Activities of ALP, ACP, and Cathepsin D were significantly increased in arthritic rats, as these are good indices of liver and kidney impairment, which are also considered as the features of adjuvant arthritis.[35] Administration of DQR decreases the lysosomal enzymes release. The reduction on lysosomal enzymes proves the beneficial effect and indirectly confirms the protective role of the plant.

The levels of glycoproteins (hexosamine and uronic acid) were elevated in arthritic group when compared to control group. This elevation indicates the severity of the disease and this altered glycoproteins metabolism observed in arthritic animals is due to the increased release of acid hydrolases during arthritic condition, these enzymes are involved in the degradation of structural macromolecules in connective tissues and cartilage proteoglycans.[28] In DQR-treated groups, the glycoproteins levels were significantly decreased.

In the arthritic group, the hydroxyproline was significantly increased when compared to normal control group. Hydroxyproline is a biomarker for collagen hydrolysis. The hydroxyproline level was increased in arthritic animals, which indicates the increased hydrolysis of collagen in tissues and bone cartilage.[35],[36] In the DQR-treated groups there was a significant decrease in hydroxyproline level compared to arthritic control groups.

Myeloperoxidase activity was significantly increased in the arthritic group when compared to normal control. In the DQR-treated groups and MTx-treated group, there was a significant decrease in myeloperoxidase activity when compared to arthritic control group. The increased level of MPO in synovial tissues and hind paw of arthritic animals might be due to increased production of hypochlorous acid, which in turn has been reported to have a destructive effect on tissue components, especially those containing unsaturated lipids.[37],[38],[39] The result of this study shows that DQR has myeloperoxidase inhibition activity.

Histopathological examination of the joint showed mild cartilage erosion in DQ juice-treated group, but there was no significant changes in DQ extract-treated group and MTx-treated group. This shows that DQ extract suppressed the inflammatory changes associated with arthritis which was equivalent to MTx.


   Conclusion Top


This study showed the anti-arthritic potential of DQR. The quantitative enzymatic analysis of tissue homogenates supported the anti-arthritic activity of DQR. DQR extract had better anti-arthritic activity than DQR fresh juice.

Ethical policy and institutional review board statement

This study was conducted after obtaining approval from the Institutional Animal Ethical Committee (IAEC. No: 15/01/AY18) and this protocol met the requirements of national guidelines of CPCSEA.

Acknowledgement

RS, UMD, and TG are thankful to Mr. V. Elango, Animal House, CLRI for assistance in maintenance of experimental animals. UMD thanks the Council of Scientific and Industrial Research, India for research fellowship.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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    Figures

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