RA
Epidemiology
♀/♂ 2-3:1, 40-50yrs, but also late-onset >60yrs. The overall prevalence of RA is 0.8% (0.3-
2.1%) and it steadily increases to 5% in women by the age of 70. Indigenous populations in
North America have increased risk. The lifetime risk of developing RA (USA) is 3.6% in
women and 1.7% in men. Having a family member with RA increases the chance to develop
RA by x3-9. A monozygotic twin of a patient with RA has 15% risk. Heritability is higher in
seropositive cases.
Causes
Genetics
•HLA-II (30%): DRB1*04:01 (*04:04, *01:01), shared epitope (MHC-II). HLA-DR4
refers to a group of alleles within the HLA-DRB1 gene (ex. HLA-DRB1*04). The disease in 1st-
degree relatives of a RA patient is 1.5x. DRB1*01:03, *13:01, *13:02 confer resistance to the
development of arthritis.
The shared epitope consists of a specific sequence of five amino acids
located in positions 70-74 of the third hypervariable region of certain HLA-DRB1 genes
(*04:01, *04:04, *01:01). The specific binding groove conferred by the SE preferentially
presents certain post-transnationally modified antigens (ex. citrullinated peptides).
•SNPs: PTPN22 (a phosphatase in lymphoid cells that dephosphorylates Zap70 and
Syk, negatively regulating TCR, BCR signaling; expression of the risk allele is associated with
ineffective negative selection of autoreactive lymphocytes), PADI4, CTLA4, STAT4 (plays a
critical role in differentiation of T cells).
•>100 loci outside the MHC have been linked to an increased risk of developing RA.
Environment
Smoking increases expression of PADI genes in the mucosal airways and enhances peptide
citrullination, generating ACPAs (arginine → citrulline), Porphyromonas gingivalis
(fibronectin citrullination), infectious agents (EBV, B19) (molecular mimicry → TLRs → NLRP3
→ inflammasome → IL-1β). Prevotella copri, Mycoplasma.
Citrullination: increased citrullination is not specific to RA and occurs regularly with
any environmental stress. What is unique to RA is the propensity for immune reactivity to
the neoepitopes created by protein citrullination with the production of ACPAs. It occurs
with higher frequency in the lungs of smokers and in gums of persons with periodontitis.
Carbamylation: unlike citrullination, carbamylation (homocitrullination) of proteins
is as a post-translational non-enzymatic modification of lysine → homocitrulline by stressed
cells. IgG antibodies against carbamylated proteins (anti-CarP) are present in ≈45% of RA
sera. IgG anti-CarP antibodies are present in 16% of ACPA(-) patients. Propably associated
with smoking. As carbamylation is an irreversible process, proteins tend to accumulate
homocitrulline residues over time. Stable extracellular matrix proteins in the joint, such as
collagens, are therefore attractive candidates for carbamylated autoantigens
The combination of carrying the HLA-SE and smoking led to a much higher risk of
ACPA(+) RA than simply adding the risk of smoking and carrying the HLA-SE alone, suggesting
a gene-environment interaction.
Experimental murine data shows that anti-PAD4 antibody binding to monocytes
triggers an inflammatory cascade that promotes immune cell recruitment to the joint and T
cell activation, culminating in synovial fibroblast activation and the development of more
severe arthritis.
Pathophysiology
,Synovium
2 layers, thin, consisted of mesenchymatic cells, primarily macrophages-like and fibroblasts-
like. The sublining consists largely of fibroblasts (FLS) and extracellular matrix (ex. collagen)
with scant blood vessels. In acute inflammation of RA, neutrophils are the most abundant
cells in the synovial fluid (they can make up 60-90% of the total leukocytes); in contrast, the
synovial membrane is rich in CD4⁺, plasma cells and macrophages - which drive the chronic
autoimmune inflammation. Chronic arthritis might have CD4+ in the synovium instead of
neutrophils (as does SLE).
Histopathology
•Hypertrophy of the lining layer: the histopathologic architecture of synovial lining
in RA is maintained by the adhesion molecule cadherin-11 expressed by FLS.
•Neoangiogenesis: as the synovium proliferates, relative synovial hypoxia is
associated with an increased production of the transcription factor HIF-1 that activates
transcription of genes that are fundamentally important for angiogenesis (VEGF, VEGF-R). As
the new vessels develop, cytokines produced in the synovium activate endothelial cells to
produce adhesion molecules (ICAM-1, VCAM-1, P-selectin, E-selectin). TNF may also
indirectly stimulate angiogenesis.
•Inflammatory infiltrate: consisting of macrophages, T-cells (CD4+>CD8+, Th1>Th2,
Th17>Tregs), B-cells, plasma cells and less DC and NK, found either randomly distributed
within the synovial sublining or grouped into follicular structures (20%).
•Pannus: a distinctive tissue located at the interface between synovium, cartilage
and bone at sites of joint destruction, where the bone isn’t covered by cartilage (bare area).
There are fibroblasts and macrophages, some lymphocytes and osteoclast precursors. Cells
within pannus produce proteinases, which degrade and destroy cartilage. Invasion of bone
by pannus is mediated by osteoclasts and results in the development of the characteristic
marginal erosions observed radiographically in RA.
Cells
Macrophages
i) cytokine production: macrophages are the main TNF and IL-1 producing cells in the
inflamed RA joint, but they also produce other inflammatory (and some anti-inflammatory)
cytokines. Although IL-6 is produced mainly by FLS, it seems that a rise in IL-6 is secondary to
macrophage activation.
ii) they reflect and amplify the proinflammatory factors present in their environment
iii) as APCs, they perpetuate the inflammatory response.
Fibroblasts
i) They produce proinflammatory cytokines (mostly IL-6 and chemokines) after being
stimulated by different cytokines (TNF, IL-17, IL-1).
ii) produce matrix degrading enzymes (MMPs) with which the pannus adheres and invades
cartilage. NOTCH3 signaling has been defined as a key factor that determines the location-
specific functions of sublining aggressive fibroblasts.
iii) express RANKL (promote osteoclastogenesis and bone erosion) (also T-cell produce it)
iv) express surface molecules such as VCAM-1 which act as chemoattractants for
macrophages retained in the intimal lining.
Of note, in inflamed RA synovium, fibroblasts express HLA-II molecules, but it is not
known whether they are able to act as effective APCs. Within RA synovium, FLS acquire
properties similar to locally invading tumors such as expression of proto-oncogenes, massive
proliferation and tissue invasion, typically observed in tumor cells. Their ability to circulate
, via lymphatics and blood stream is similar to the metastatic capacity of malignant cells and
provide a potential mechanism explaining the polyarticular nature of RA.
Neutrophils
They don’t express the integrin VLA-4, which may explain why they don’t remain in synovial
tissue where expression of its counter-receptor (VCAM-1) on FLS appears to be important in
the retention of other kinds of leukocytes. However, they’re abundant in RA synovial
effusions. They produce cytokines. Moreover, activated neutrophils may carbamylate
various proteins via the release of MPO and ROS, regardless of NETosis. They also have PAD,
responsible for citrullination. Experimental murine data shows that anti-PAD4 antibody
binding to monocytes triggers an inflammatory cascade that promotes immune cell
recruitment to the joint and T cell activation, culminating in synovial fibroblast activation
and the development of more severe arthritis.
NETosis: during NETosis, released citrullinated proteins may provide an
additional source of target antigens. PAD4 itself is also found in NETs, allowing it to continue
to citrullinate a host of other self-proteins after the NET is released. NETosis is known to
occur at high frequencies in lungs of smokers and gums affected by periodontitis, providing a
potential molecular mechanism for those these RA risk factors. Moreover, NETosis is
triggered by ACPA, RF, TNF and IL-17A, thereby setting up potential positive feedback loops.
Lymphocytes
-T cells: constitute >50% of the cells in most RA synovium, mostly CD4+ and memory T-cells,
which are present diffusely, in aggregates and in perivascular sites. There are also CD8+
which are present diffusely and are a major source of IFN-γ and TNF production in synovial
tissues. There appears to be a preponderance of Th1 and Th17 subset, with deficiency of Th2
and Tregs.
-B-cells/plasma cells (<5%): are present within and between aggregates and in foci; many
plasma cells produce RF as well as ACPAs. They’re stimulated by IL-10, BAFF and APRIL
present in the inflamed synovium.
Cytokine Perturbations
The RA synovium is characterized by the expression of cytokines and chemokines including
TNF, IL-1, IL-6, IL-10, IL-12, IL-18, (G)M-CSF, TGF-β. IFN-γ (Th1)and IL-17 (Th17) are
considered to play a central role in initiating and perpetuating synovitis as well as
contributing to osteoclast activation leading to erosive disease. The main TNF producers are
the macrophages, while the main IL-6 producers are FLS. IL-10 (which apart from being
immunosuppressive, it’s also a B-cell stimulant) producers are mainly macrophages.
•TNF: is not high in the sera, only in synovial biopsies, but IL-6 is. Although
macrophages are the main source of TNF production, signals from activated T-cells are likely
to be important in inducing TNF production within the inflamed joint. Inflammatory
cytokines such as TNF, IL-1 and IL-6 can substitute the RANK/RANKL signaling during
osteoclast activation.
-induces IL-1 (not reversely - maybe that’s why anti-IL-1 are not as effective)
-together with IL-1, induces fibroblast proliferation → IL-6, IL-8, GM-CSF.
•IL-17:
-induces RANKL production by fibroblasts → osteoclast activation
-together with IL-1, induces IL-6 production
•IL-6: in abundance in both synovium and serum. Upregulated IL-6 and soluble IL-6R
produced by lung macrophages enhance fibroblast proliferation and extracellular matrix
protein production (for ILD).
Epidemiology
♀/♂ 2-3:1, 40-50yrs, but also late-onset >60yrs. The overall prevalence of RA is 0.8% (0.3-
2.1%) and it steadily increases to 5% in women by the age of 70. Indigenous populations in
North America have increased risk. The lifetime risk of developing RA (USA) is 3.6% in
women and 1.7% in men. Having a family member with RA increases the chance to develop
RA by x3-9. A monozygotic twin of a patient with RA has 15% risk. Heritability is higher in
seropositive cases.
Causes
Genetics
•HLA-II (30%): DRB1*04:01 (*04:04, *01:01), shared epitope (MHC-II). HLA-DR4
refers to a group of alleles within the HLA-DRB1 gene (ex. HLA-DRB1*04). The disease in 1st-
degree relatives of a RA patient is 1.5x. DRB1*01:03, *13:01, *13:02 confer resistance to the
development of arthritis.
The shared epitope consists of a specific sequence of five amino acids
located in positions 70-74 of the third hypervariable region of certain HLA-DRB1 genes
(*04:01, *04:04, *01:01). The specific binding groove conferred by the SE preferentially
presents certain post-transnationally modified antigens (ex. citrullinated peptides).
•SNPs: PTPN22 (a phosphatase in lymphoid cells that dephosphorylates Zap70 and
Syk, negatively regulating TCR, BCR signaling; expression of the risk allele is associated with
ineffective negative selection of autoreactive lymphocytes), PADI4, CTLA4, STAT4 (plays a
critical role in differentiation of T cells).
•>100 loci outside the MHC have been linked to an increased risk of developing RA.
Environment
Smoking increases expression of PADI genes in the mucosal airways and enhances peptide
citrullination, generating ACPAs (arginine → citrulline), Porphyromonas gingivalis
(fibronectin citrullination), infectious agents (EBV, B19) (molecular mimicry → TLRs → NLRP3
→ inflammasome → IL-1β). Prevotella copri, Mycoplasma.
Citrullination: increased citrullination is not specific to RA and occurs regularly with
any environmental stress. What is unique to RA is the propensity for immune reactivity to
the neoepitopes created by protein citrullination with the production of ACPAs. It occurs
with higher frequency in the lungs of smokers and in gums of persons with periodontitis.
Carbamylation: unlike citrullination, carbamylation (homocitrullination) of proteins
is as a post-translational non-enzymatic modification of lysine → homocitrulline by stressed
cells. IgG antibodies against carbamylated proteins (anti-CarP) are present in ≈45% of RA
sera. IgG anti-CarP antibodies are present in 16% of ACPA(-) patients. Propably associated
with smoking. As carbamylation is an irreversible process, proteins tend to accumulate
homocitrulline residues over time. Stable extracellular matrix proteins in the joint, such as
collagens, are therefore attractive candidates for carbamylated autoantigens
The combination of carrying the HLA-SE and smoking led to a much higher risk of
ACPA(+) RA than simply adding the risk of smoking and carrying the HLA-SE alone, suggesting
a gene-environment interaction.
Experimental murine data shows that anti-PAD4 antibody binding to monocytes
triggers an inflammatory cascade that promotes immune cell recruitment to the joint and T
cell activation, culminating in synovial fibroblast activation and the development of more
severe arthritis.
Pathophysiology
,Synovium
2 layers, thin, consisted of mesenchymatic cells, primarily macrophages-like and fibroblasts-
like. The sublining consists largely of fibroblasts (FLS) and extracellular matrix (ex. collagen)
with scant blood vessels. In acute inflammation of RA, neutrophils are the most abundant
cells in the synovial fluid (they can make up 60-90% of the total leukocytes); in contrast, the
synovial membrane is rich in CD4⁺, plasma cells and macrophages - which drive the chronic
autoimmune inflammation. Chronic arthritis might have CD4+ in the synovium instead of
neutrophils (as does SLE).
Histopathology
•Hypertrophy of the lining layer: the histopathologic architecture of synovial lining
in RA is maintained by the adhesion molecule cadherin-11 expressed by FLS.
•Neoangiogenesis: as the synovium proliferates, relative synovial hypoxia is
associated with an increased production of the transcription factor HIF-1 that activates
transcription of genes that are fundamentally important for angiogenesis (VEGF, VEGF-R). As
the new vessels develop, cytokines produced in the synovium activate endothelial cells to
produce adhesion molecules (ICAM-1, VCAM-1, P-selectin, E-selectin). TNF may also
indirectly stimulate angiogenesis.
•Inflammatory infiltrate: consisting of macrophages, T-cells (CD4+>CD8+, Th1>Th2,
Th17>Tregs), B-cells, plasma cells and less DC and NK, found either randomly distributed
within the synovial sublining or grouped into follicular structures (20%).
•Pannus: a distinctive tissue located at the interface between synovium, cartilage
and bone at sites of joint destruction, where the bone isn’t covered by cartilage (bare area).
There are fibroblasts and macrophages, some lymphocytes and osteoclast precursors. Cells
within pannus produce proteinases, which degrade and destroy cartilage. Invasion of bone
by pannus is mediated by osteoclasts and results in the development of the characteristic
marginal erosions observed radiographically in RA.
Cells
Macrophages
i) cytokine production: macrophages are the main TNF and IL-1 producing cells in the
inflamed RA joint, but they also produce other inflammatory (and some anti-inflammatory)
cytokines. Although IL-6 is produced mainly by FLS, it seems that a rise in IL-6 is secondary to
macrophage activation.
ii) they reflect and amplify the proinflammatory factors present in their environment
iii) as APCs, they perpetuate the inflammatory response.
Fibroblasts
i) They produce proinflammatory cytokines (mostly IL-6 and chemokines) after being
stimulated by different cytokines (TNF, IL-17, IL-1).
ii) produce matrix degrading enzymes (MMPs) with which the pannus adheres and invades
cartilage. NOTCH3 signaling has been defined as a key factor that determines the location-
specific functions of sublining aggressive fibroblasts.
iii) express RANKL (promote osteoclastogenesis and bone erosion) (also T-cell produce it)
iv) express surface molecules such as VCAM-1 which act as chemoattractants for
macrophages retained in the intimal lining.
Of note, in inflamed RA synovium, fibroblasts express HLA-II molecules, but it is not
known whether they are able to act as effective APCs. Within RA synovium, FLS acquire
properties similar to locally invading tumors such as expression of proto-oncogenes, massive
proliferation and tissue invasion, typically observed in tumor cells. Their ability to circulate
, via lymphatics and blood stream is similar to the metastatic capacity of malignant cells and
provide a potential mechanism explaining the polyarticular nature of RA.
Neutrophils
They don’t express the integrin VLA-4, which may explain why they don’t remain in synovial
tissue where expression of its counter-receptor (VCAM-1) on FLS appears to be important in
the retention of other kinds of leukocytes. However, they’re abundant in RA synovial
effusions. They produce cytokines. Moreover, activated neutrophils may carbamylate
various proteins via the release of MPO and ROS, regardless of NETosis. They also have PAD,
responsible for citrullination. Experimental murine data shows that anti-PAD4 antibody
binding to monocytes triggers an inflammatory cascade that promotes immune cell
recruitment to the joint and T cell activation, culminating in synovial fibroblast activation
and the development of more severe arthritis.
NETosis: during NETosis, released citrullinated proteins may provide an
additional source of target antigens. PAD4 itself is also found in NETs, allowing it to continue
to citrullinate a host of other self-proteins after the NET is released. NETosis is known to
occur at high frequencies in lungs of smokers and gums affected by periodontitis, providing a
potential molecular mechanism for those these RA risk factors. Moreover, NETosis is
triggered by ACPA, RF, TNF and IL-17A, thereby setting up potential positive feedback loops.
Lymphocytes
-T cells: constitute >50% of the cells in most RA synovium, mostly CD4+ and memory T-cells,
which are present diffusely, in aggregates and in perivascular sites. There are also CD8+
which are present diffusely and are a major source of IFN-γ and TNF production in synovial
tissues. There appears to be a preponderance of Th1 and Th17 subset, with deficiency of Th2
and Tregs.
-B-cells/plasma cells (<5%): are present within and between aggregates and in foci; many
plasma cells produce RF as well as ACPAs. They’re stimulated by IL-10, BAFF and APRIL
present in the inflamed synovium.
Cytokine Perturbations
The RA synovium is characterized by the expression of cytokines and chemokines including
TNF, IL-1, IL-6, IL-10, IL-12, IL-18, (G)M-CSF, TGF-β. IFN-γ (Th1)and IL-17 (Th17) are
considered to play a central role in initiating and perpetuating synovitis as well as
contributing to osteoclast activation leading to erosive disease. The main TNF producers are
the macrophages, while the main IL-6 producers are FLS. IL-10 (which apart from being
immunosuppressive, it’s also a B-cell stimulant) producers are mainly macrophages.
•TNF: is not high in the sera, only in synovial biopsies, but IL-6 is. Although
macrophages are the main source of TNF production, signals from activated T-cells are likely
to be important in inducing TNF production within the inflamed joint. Inflammatory
cytokines such as TNF, IL-1 and IL-6 can substitute the RANK/RANKL signaling during
osteoclast activation.
-induces IL-1 (not reversely - maybe that’s why anti-IL-1 are not as effective)
-together with IL-1, induces fibroblast proliferation → IL-6, IL-8, GM-CSF.
•IL-17:
-induces RANKL production by fibroblasts → osteoclast activation
-together with IL-1, induces IL-6 production
•IL-6: in abundance in both synovium and serum. Upregulated IL-6 and soluble IL-6R
produced by lung macrophages enhance fibroblast proliferation and extracellular matrix
protein production (for ILD).
