Effect of Black Seed Thymoquinone on Rheumatoid Arthritis

Thymoquinone (2-isoprpyl-5-methyl-1,4-benzoquinone) is the most active component of Black cumin (Nigella sativa) seed oil. It is widely used in traditional medicine to treat a wide range of illnesses.

According to various research findings, thymoquinone exert important health-beneficial effects including antioxidant, anti-inflammatory and anti-cancer effects. As an antioxidant agent, thymoquinone normalizes glutathione levels and increases the activity of antioxidant enzymes such as glutathione peroxidase, catalase, and superoxide dismutase.


Black Cumin Tea Contains Thymoquinone

Black cumin seed is made into a tea and served as an antioxidant agent. Since it contains thymoquinone and other biologically active phytochemicals, black cumin seed tea should be recommended for patients who suffer rheumatoid arthritis. It can also be served to cancer patients who are undergoing chemotherapy.

How To Prepare Black Cumin Seed Tea

  • To prepare black cumin seed tea, assemble 2 cups of water, 2 tea spoonful of raw black cumin seeds and honey. Then follow the steps as follows:
  • Add the 2 cups of water and black cumin seeds in a pot and heat till it boils.
  • Remove from heat once it boiled.
  • Cover the pot and allow it to steep for about 10 minutes.
  • Strain the water into a cup using a mesh. You may add the seed to enjoy the whole benefits if you so wish.
  • Add two spoonful of honey and stir
  • Serve and enjoy your tea


Benefits of Thymoquinone and Black Cumin Seed Tea in Treatment of Rheumatoid Arthritis

The uncontrolled inflammation associated with rheumatoid arthritis arises as a result of the uncontrolled production and activity of various inflammatory cytokines, TNFα, IL-1, IL-17, IL-6 and the resulting inflammation is responsible for the pain, tenderness, swelling, redness and stiffness of joints.

Black cumin seeds and especially thymoquinone has been shown to be beneficial against rheumatoid arthritis and autoimmune diseases.


Thymoquinone Inhibits NF-kB Signaling In Rheumatoid Arthritis

NF-kB regulates the expression of many genes, enzymes, cytokines, cell cycle regulatory molecules as well as angiogenic factors. It induces inflammation by influencing the expression several pro-inflammatory cytokines, chemokines , acute phase proteins and growth factors.

Thymoquinone inhibits NF-kB induced inflammatory response in rheumatoid arthritis patients by inhibiting its translocation into the nucleus. Thymoquinone also inhibits NF-kB activities by suppressing TNF-induced-NF-kB activation. This is done by inhibiting TNF-induced IKBα phosphorylation and degradation as well as p65 phosphorylation and nuclear translocation.


Thymoquinone Inhibits Prostaglandins and COX-2 in Rheumatoid Arthritis

Prostaglandins (PGs) are arachidonic acid metabolites. They are found at elevated levels in synovial fluid and also in synovial membrane, where they function in the development of vasodilation, fluid extravasation and pain in synovial tissues. Aside these functions, Prostaglandin E2 (PGE2) and COX are upregulated in synovial tissues in rheumatoid arthritis patients, where PGE2 synergizes with IL-23 to stimulate Th17 cell proliferation.

Th17 in turn stimulates the release of pro-inflammatory cytokines and promotes bone resorption. PGE2 also mediates complex interactions that lead to the development of articular cartilage erosions and juxta-articular bone.

Cyclooxygenase enzymes (COX), especially COX-2 are involved in inflammatory responses. COX-2 is induced by pro-inflammatory cytokines, mainly IL-1.

Thymoquinone suppresses the expression of COX-2 protein by inhibiting NF-kB signaling pathway activation and induces the expression of cytoprotective enzymes.

Effect of Thymoquinone on PI3k/Akt Signaling Pathway

Phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway is an intracellular regulatory signal transduction pathway that is activated by toxic substances or cellular stimuli that regulate many cellular processes including cell growth, survival and apoptosis.

When abnormally activated, PI3K/Akt signaling pathway is involved in the pathogenesis of many diseases including diabetes mellitus, cancer, and rheumatoid arthritis. In RA, PI3K/Akt signaling pathway play important role through the expression of different types of pro-inflammatory mediators that degrade IKβ and activate NF-kB signaling pathway.

Thymoquinone induces apoptosis by blocking PI3K/Akt signaling pathway in DU-145 cell line. It also deactivate PI3K/Akt and NF-kB signaling pathway and regulate various gene products such as p65 and COX-2.



J.K. Kundu, L. Liu, J.-W. Shin, Y.-J. Surh., Thymoquinone inhibits phorbol ester-induced activation of NF-κB and expression of COX-2, and induces expression of cytoprotective enzymes in mouse skin in vivo Biochem. Biophys. Res. Commun., 438 (4) (2013), pp. 721-727


Umar, J. Zargan, K. Umar, S. Ahmad, C.K. Katiyar, H.A. Khan Modulation of the oxidative stress and inflammatory cytokine response by thymoquinone in the collagen induced arthritis in Wistar rats Chem. Biol. Interact., 197 (1) (2012), pp. 40-46


Vaillancourt, P. Silva, Q. Shi, H. Fahmi, J.C. Fernandes, M. Benderdour

Elucidation of molecular mechanisms underlying the protective effects of thymoquinone against rheumatoid arthritis J. Cell. Biochem., 112 (1) (2011), pp. 107-117


Thymoquinone inhibits IL-1β-induced inflammation in human osteoarthritis chondrocytes by suppressing NF-κB and MAPKs signaling pathway Inflammation, 38 (6) (2015), pp. 2235-2241


Khan, A. Sureda, T. Belwal, S. Çetinkaya, İ. Süntar, S. Tejada, H.P. Devkota, H. Ullah, M. Aschner. Polyphenols in the treatment of autoimmune diseases. Autoimmun. Rev., 18 (7) (2019), pp. 647-657


Stańczyk J, Kowalski ML. Rola cyklooksygenaz oraz prostaglandyn w patogenezie reumatoidalnego zapalenia stawów [The role of cyclooxygenase and prostaglandins in the pathogenesis of rheumatoid arthritis]. Pol Merkur Lekarski. 2001 Nov;11(65):438-43.


Molecular Mechanisms of Osteoarthritis

Osteoarthritis (OA) is a chronic degenerative disorder which occurs when the cartilage that cushions the ends of bones in joints wear down over a period of time. The molecular mechanisms of osteoarthritis have not received much attention among the public, except the few who are in research and development of drugs. OA is the most prevalent chronic joint disease, which increases in prevalence with age and affects majority of persons over 65.
Unlike Rheumatoid arthritis (RA), OA is caused by mechanical wear and tear at the joints. Also, while RA can occur at any stage, OA occurs later in life of the individual. Both however, share some similarities in their symptoms. But, while RA affects both sides of the affected joints, OA can affect one side of the joint.

Symptoms of Osteoarthritis

Symptoms of Osteoarthritis may develop slowly and worsen over a period of time. Some of the signs and symptoms include:
• Loss of flexibility at the affected joints.
• Pain at the affected joint.
• Stiffness of joints especially upon awaking.
• Swelling.
• Tenderness of tissues around the affected joints.

Pathological Progression of Osteoarthritis

The pathological process of osteoarthritis begins by an alteration in the composition and organization of the extracellular matrix. When this happens, the articular chondrocytes exhibit a transient proliferative response and increased matrix synthesis (Col2, aggrecan etc) in an attempt to initiate repair of damage caused by pathological stimulation.
Changes in the composition and structure of the cartilage further stimulate chondrocytes to produce more catabolic factors involved in cartilage degradation. As a result, the proteoglycans and collagen network break down, leading to the disruption of the cartilage integrity. Later on, the chondrocytes will undergo apoptosis and the articular cartilage will be completely lost. The molecular mechanisms of osteoarthritis involve several pathways that network with each other to induce inflammatory response in osteoarthritis patients and include growth factors and cytokines.


Growth Factors and Osteoarthritis

Transforming growth factor B (TGF-B) plays important role in skeletogenesis and inhibits chondrocyte hypertrophy and maturation. The inhibition of TGF-B represents a potential mechanism in the development of OA, and the loss of TGF-B is associated with cartilage damage. Some other growth factors that play important roles in cartilage homeostasis are fibroblast growth factor-2 and 18 (FGF-2 and FGF-18). FGF-2 has a potent catabolic and anti-anabolic role in human cartilage homeostasis. It is released in supraphysiological amounts during loading/ injury of the cartilage matrix. It activates multiple transduction signal pathways (MAPKs) such as ERK, p38 and JNK. FGF-2 can stimulate MMP-13 expression, which is the major degrading enzyme to type II collagen. Its stimulation also mediates the up-regulation of matrix-dependent enzyme expression (ADAMTS-5 and MMP-13) and the down-regulation aggrecan expression. But PKCგ inhibition significantly impairs the detrimental effects mediated by FGF-2.
FGF-18 which is a secreted heparin-binding polypeptide growth factor is involved in cartilage growth and maturation and is implicated in the development of functional cartilage and bone tissue. It has roles in enhancing regeneration and repression of damaged cartilage by stimulating chondrogenesis.

Molecular Mechanism of Wnt/β-catenin Signaling in Osteoarthritis

Wnts are a family of extracellularly secreted glycoproteins that are in numerous biological activities, including cell proliferation, differentiation, polarization and fate determination. They have been implicated in the development of some diseases through canonical β-catenin-dependent and noncanonical β catenin-dependent signaling pathways.
During cartilage development, Wnt/β-catenin signaling activity is strictly regulated in chondrogenesis and chondrocyte maturation. Excessive Wnt pathway activation in the adult articular cartilage under IL-1β stimulation is thought to be an OA progression susceptibility factor. An increase in expression of Wnt1 pathway activator, Wnt inducible-signaling pathway protein 1 (WISP-1) induces articular cartilage degradation through up-regulation of the expression MMPs and aggrecanases in chondrocytes and macrophages. But the expression levels of some wnt pathway antagonists, such as sclerostin, dickkopf WNT signaling pathway inhibitor 1 (DKK1), and secreted frizzled-related protein 3 (sFRP3) usually decreases in parallel with OA progression. Therefore, an up-regulation of the expression of these antagonists alleviates OA cartilage destruction. A mutant in sFRP3 causes increased levels of active β-catenin, thereby promoting abnormal articular chondrocyte hypertrophy and results in hip and knee in OA patients.

Molecular Mechanism of the PI3K/AkT/mTOR Pathway in OA Pathogenesis

The PI3/AkT/mTOR pathway plays significant roles in the molecular mechanisms of osteoarthritis in three aspects of OA which are cartilage, synovial inflammation and subchondral bone sclerosis.

Cartilage Homeostasis

Cartilage homeostasis is important to articular health and is defined as the state in which synthesis of extracellular matrix (ECM) is balanced by its degradation. The destruction of cartilage homeostasis initiates and boosts OA pathogenesis. It is marked by elevated MMPs, a disintegrin and metalloproteinases with thrombospondrin motifs (ADAMTSs) and a reduced collagen II and aggrecan levels.
The PI3K/AkT pathway plays a role in both ECM anabolism and catabolism. When stimulated, AkT promotes the synthesis of collagen II and an over-expression of AkT enhances proteoglycan synthesis in human chondrocytes under the stimulation of tert-butylhydroperoxide (tBHP). When PI3k is activated, Akt is phosphorylated, which promotes the activation of mTORC1. The activated mTORC1 inhibits autophagy by associating with ULK1- Atg13-FIR200 complex, thereby inhibiting autophagosome formation. Inhibiting of mTOR and the subsequent increase in autophagic activity may restore homeostasis in articular cartilage chondrocytes.

NF-kB in OA Pathogenesis

When excessively activated, NF-kB signaling up-regulates hypoxia-inducible factor 2α (HIF-2α) transcription. This results in enhanced OA progression through further induction of catabolic factors such as MMPs, VEGF and IHH. But a moderate activity of NF-kB is required to maintain healthy articular cartilage.
The NF-kB-HIF-2α axis may be a potential therapeutic target. This is because a moderate dose of an IKK inhibitor can suppress HIF-2α expression without significant effect on cell survival.


Almonte-Becerril M, Navarro-Garcia F, Gonzalez-Robles A, Vega-Lopez MA, Lavalle C, Kouri JB. 2010. Cell death of chondrocytes is a combination between apoptosis and autophagy during the pathogenesis of osteoarthritis within an experimental model. Apoptosis 15(5): 631– 638.
Anandacoomarasamy A, Leibman S, Smith G, Caterson I, Giuffre B, Fransen M, Sambrook PN, March L. 2012. Weight loss in obese people has structure-modifying effects on medial but not on lateral knee articular cartilage. Ann Rheum Dis 71(1): 26– 32.
Benderdour M, Tardif G. Pelletier JP, Di Battista JA, Reboul P, Ranger P, Martel-Pelletier J. 2002. Interleukin 17 (IL-17) induces collagenase-3 production in human osteoarthritic chondrocytes via AP-1 dependent activation: Differential activation of AP-1 members by IL-17 and IL-1beta. J Rheumatol 29(6): 1262– 1272.
Billinghurst RC, Dahlberg L, Ionescu M, Reiner A, Bourne R, Rorabeck C, Mitchell P, Hambor J, Diekmann O, Tschesche H, Chen J, Van Wart H, Poole AR. 1997. Enhanced cleavage of type II collagen by collagenases in osteoarthritic articular cartilage. J Clin Invest 99(7): 1534– 1545.
Blanco FJ, Guitian R, Vazquez-Martul E, de Toro FJ, Galdo F. 1998. Osteoarthritis chondrocytes die by apoptosis. A possible pathway for osteoarthritis pathology. Arthritis Rheum 41(2): 284– 289.
Bohensky J, Terkhorn SP, Freeman TA, Adams CS, Garcia JA, Shapiro IM, Srinivas V. 2009. Regulation of autophagy in human and murine cartilage: Hypoxia-inducible factor 2 suppresses chondrocyte autophagy. Arthritis Rheum 60(5): 1406– 1415.
Little CB, Barai A, Burkhardt D, et al. Matrix metalloproteinase 13-deficient mice are resistant to osteoarthritic cartilage erosion but not chondrocyte hypertrophy or osteophyte development. Arthritis Rheum
Glasson SS, Askew R, Sheppard B, et al. Deletion of active ADAMTS5 prevents cartilage degradation in a murine model of osteoarthritis. Nature. 2005; 434:644–648.
Majumdar MK, Askew R, Schelling S, et al. Double-knockout of ADAMTS-4 and ADAMTS-5 in mice results in physiologically normal animals and prevents the progression of osteoarthritis. Arthritis Rheum. 2007; 56:3670–3674139.
Kobayashi H, Chang SH, Mori D, Itoh S, Hirata M, Hosaka Y, Taniguchi Y, Okada K, Mori Y, Yano F, et al. Biphasic regulation of chondrocytes by Rela through induction of anti-apoptotic and catabolic target genes. Nat Commun. 2016;7:13336.
Goldring MB, Otero M. Inflammation in osteoarthritis. Curr Opin Rheumatol. 2011;23(5):471–8.

Mendez MG, Janmey PA. Transcription factor regulation by mechanical stress. Int J Biochem Cell Biol. 2012;44(5):728–32.
Nam J, Aguda BD, Rath B, Agarwal S. Biomechanical thresholds regulate inflammation through the NF-kappaB pathway: experiments and modeling. PLoS One. 2009;4(4), e5262.


Nuclear Factor-Kappa B (NF-kB)

Nuclear factor-kappa B (NF-kB) is a transcription factor that plays critical functional roles in inflammation, immunity, cell proliferation, differentiation, and survival. It exists in an inactive state in the cytosol and can be stimulated by molecules such as TNFα, and other cell stressors.

NF-kB is found in almost all cellular cell types and is identified as a regulator of kB light chain expression in mature B and plasma cells.

Because NF-kB has the ability to influence the expression of numerous genes, its activity is tightly regulated at multiple levels. The primary mechanism for regulating NF-kB is through inhibitory IkB proteins and the IKK complex, which phosphorylates IkBs.

Nuclear Factor-kappa B (NF-kB) Activation

NF-kB activation is initiated by TNFα. When TNFα binds to TNF receptors and activate them, IkB kinase (IKK) is ultimately triggered, which leads to the phosphorylation of IkB. The phosphorylation of IkB results in ubiquitination and degradation of IkB. When this happens, the remaining NF-kB dimmer (e.g., p65/p50 or p50/p50 subunits) translocates to the nucleus, where it then binds to a DNA consensus sequence of target genes.


Nuclear Factor-kappa B (NF-kB) Structure-Function relationship

NF-kB is a multiple-gene family of proteins that can form stable homo- and heterodimeric complexes, which vary in their DNA binding specificity and transcriptional activation potential.

There are five proteins of the NF-kB family in mammalian cells; they are RelA (P65), C-Rel, RelB, and NF-kB1 (p50 and its precursor p105), and NF-kB2 (p52 and its precursor p100). NF-kB and Rel proteins share a highly conserved 300 amino acid N-terminal Rel homology domain (RHD), which is responsible for DNA binding, dimerization, and association with IkB inhibitory proteins.

The p50/p65 complex shows strong transcriptional activation, whereas p50/p50 and p52/p52 homodimers suppress transcription of NF-kB target genes.


Nuclear Factor-kappa B (NF-kB) Is Inhibited By IkB Family Proteins

NF-kB is inhibited by IkB family proteins through NF-kB/IkB complex formation. There are seven inhibitory protein members of IkB family. They are IkBα, IkBβ, IkBε, IkBϒ, Bcl3, NF-kB1 precursor and NFkB2 precursor.

The IkB family members have a common ankyrin repeat domain. They regulate the subcellular localization, and hence, the DNA binding and transcriptional activity of NF-kB proteins.


IkB Regulates NF-kB Translocation to the Nucleus.

NF-kB is localized in the cytoplasm in an NF-kB/IkB complex, which is inactive. The inactive NF-kB/IkB co2mplex is a result of masking of the nuclear localization signals (NLS) on the NF-kB subunits by the IkB proteins. Hence, the degradation of IkB would lead to unmasking of the NLS, allowing NF-kB to undergo translocation to the nucleus.

The IkB proteins show a preference for specific NF-kB/Rel complexes, which provides a means to regulate the activation of distinct Rel/NF-kB complexes.

After the binding and transcriptional activity of NF-kB on DNA, it induces the expression of IkBα, which enters the nucleus and remove NF-kB  from DNA by forming NF-kB/IkB complex with the released NF-kB. The complex is then expelled from the nucleus back to the cytoplasm as a result of potent nuclear export signals on IkB and p65.


IkB Degradation Is Mediated By Ubiquiti/Proteasome System

The activation of NF-kB is achieved through the signal induced proteolytic degradation of IkB. This degradation is initiated by the stimuli dependent phosphorylation of IkB at specific N-terminal residues (S32/S36 for IkBα, S19/S23 for IkBβ), and is mediated by the ubiquitin/proteasome system.

Phosphorylation of IkB however, is not enough to initiate degradation of IkB. Ubiquitination and subsequent degradation depends on the recognition of phosphorylated IkB by the β-TrCp, an F-box/WD containing component of the Skp1-cullin-F-box (SCF) class of E3 ubiquitin ligases.


IKKs Mediates The Phosphorylation Of IkB On Serine 32 and 36

Phosphorylation of IkBa on the Serine 32 and 36 is mediated by IkB kinases (IKKs), whose activity is induced by activators of the NF-kB pathway. IKK contains two subunits, IKK1 (IKKα) and IKK2 (IKKβ). It also contains a regulatory subunit, NEMO ( IKKy, IKKAPI, FIP3).

IKK1 and IKK2 are homologous kinases, and both contain an N-terminal kinase domain and a C-terminal region with two protein interaction motifs, a leucine zipper (LZ), and ahelix-loop-helix (HLH) motif.

The LZ domain is responsible for demerization of IKK1 and IKK2. It is also essential for IKK complex activity. The IKK1/2 complex associates with NEMO through a short interaction motif located at the C-terminus of either catalytic subunit. Short peptides derived from the interaction motif can be used to disrupt the IKK complex and prevent its activation. NEMO connects the IKK complex to upstream activators through its C-terminus, which contains a zinc finger motif. NEMO also undergoes stimulus dependent interaction with components of TNF receptor complex.





Hacker H, Karin M. 2006. Regulation and function of IKK and IKK-related kinases. Sci STKE 2006: re13.

Harhaj EW, Maggirwar SB, Sun SC. 1996. Inhibition of p105 processing by NF-kB proteins in transiently transfected cells. Oncogene 12: 2385–2392.

Hatakeyama S, Kitagawa M, Nakayama K, Shirane M, Matsumoto M, Hattori K, Higashi H, Nakano H, Okumura K, Onoe K, et al. 1999. Ubiquitin-dependent degradation of IkB is mediated by a ubiquitin ligase Skp1/Cul 1/F-box protein FWD1. Proc Natl Acad Sci 96: 3859–3863.

Chen G , Cao P , Goeddel DV . TNF-induced recruitment and activation of the IKK complex require Cdc37 and Hsp90 . Mol Cell 2002 ; 9 : 401 – 10.

Hu Y , Baud V , Delhase M , et al . Abnormal morphogenesis but intact IKK activation in mice lacking the IKKalpha subunit of IkappaB kinase . Science 1999 ; 284 : 316 – 20 .

Li Q , Lu Q , Hwang JY , et al . IKK1-deficient mice exhibit abnormal development of skin and skeleton . Genes Dev 1999 ; 13 : 1322 – 8.

Li Q , Van Antwerp D , Mercurio F , et al . Severe liver degeneration in mice lacking the IkappaB kinase 2 gene . Science 1999 ; 284 : 321 – 5.

Takeda K , Takeuchi O , Tsujimura T , et al . Limb and skin abnormalities in mice lacking IKKalpha . Science 1999 ; 284 : 313 – 16.

Ghosh S , Karin M . Missing pieces in the NFkappaB puzzle . Cell 2002 ; 109 ( Suppl. ) : S81 – 96.

Yang J , Lin Y , Guo Z , et al . The essential role of MEKK3 in TNFinduced NFkappaB activation . Nat Immunol 2001 ; 2 : 620 – 4.

Li ZW , Chu W , Hu Y , et al . The IKKbeta subunit of IkappaB kinase (IKK) is essential for nuclear factor kappaB activation and prevention of apoptosis . J Exp Med 1999 ; 189 : 1839 – 45.