GUJHS. 2007 March; Vol. 4, No. 1
Daliha Aqbal, Assya Abdallah, Estafani Bolloso, Allan Angerio, PhD
Department of Human Science
School of Nursing and Health Studies
C-reactive protein (CRP), a prominent acute-phase protein present in inflammatory responses, performs a major role in inflammatory bowel disease (IBD). Characterized by chronic gastrointestinal irritation, IBD results in the stimulation of three specific cytokines: interleukin-1β, interleukin-6, and tumor-necrosis factor-α. These cytokines trigger the elevated production of CRP in the hepatocytes, increasing its plasma concentration from 0.8mg/L to approximately 45mg/L, almost a 500-fold increase. As a result of its palpable prevalence and proliferation in IBD, this research explores the role of CRP as a marker, and as a pro and anti-inflammatory agent. As a clinical aid, CRP levels serve as a marker for detecting IBD, differentiating it from functional bowel disorders, categorizing the disease severity and risk, and monitoring the effective treatments based on disease activity. CRP, as a pro-inflammatory agent operates by associating itself with the main cytokinal release of interleukin-6 and engaging in pseudo-antibody reactions with extrinsic or autologous ligands, which initiate the complement system. Finally, as an inhibitor of inflammation, CRP prevents neutrophil-endothelial adhesion through the shedding of L-selectin molecule expression in neutrophil cells possibly by binding to CD-32. While CRP functions as a pro and anti-inflammatory agent in inflammatory responses, its net effect in IBD patients requires further investigation.
Keywords: C-Reactive Protein, Inflammatory Bowel Disease, Crohn’s Disease, Ulcerative Collitis, Functional Bowel Disorder, Acute Phase Proteins, Cytokines, Hepatocytes, Tumor-Necrosis Factor-Alpha, Interleukin-6, Interleukin-1β, modified C-Reactive Protein, Inflammation, L-selectin
Inflammatory bowel disease (IBD) characterizes a range of chronic ailments affecting the digestive tract. It has been known to be a serious and life-long condition affecting many young people between the ages of 15 and 25 years old (Vermiere, 2005). Over the years, reports of IBD have gradually increased, affecting 150 to 250 people per 100,000 population. IBD diverges into two main forms: Ulcerative colitis (UC), a disease that has become relatively stabilized among people, and a second form, Crohn’s disease (CD) that researchers have determined to be steadily escalating (Isaacs, 2005). Each of these chronic intestinal inflammatory diseases is differentiated by its physical appearance and by the quantitative magnitude of their spread in the body. UC inflames the mucosal lining of the colon, affecting it in a continuous way, while CD distresses the entire gastrointestinal tract from the mouth to the anus in a discontinuous, transmural manner described as “cobblestone” (Vermiere, 2005) (Angerio, 2005). The distinctions between them can further be determined by the presence of HLA-G, a class I molecule of the major histocompatibility complex (MHC) in the intestinal tissue of UC and not in CD. Genes of the MHC influence disease susceptibility and disease behavior for patients with IBD (Torres, 2004). CD and UC often present themselves with bloody diarrhea, abdominal pain, anorexia and weight loss (Cabrera-Abreu, 2004). Patients suffering from IBD encounter severe physical pain and emotional trauma.
The ambiguity of the disease lies in the fact that there have been no determined causes or cures for IBD, but rather an accumulation of predictions and drugs that have been known to affect the disease in a specific manner. Current research indicates that IBD is an autoimmune process, in which the gut flora or intestinal bacteria of the small intestine and upper stomach trigger sustained inflammation as the body initiates the production of antibodies against its own tissue (Vermiere, 2005). Genetic disposition and environmental contributions serve as the most probable cause in triggering the autoimmune process. According to one study, between 6 percent and 32 percent of patients with IBD have a first or second relative affected by it, siblings being at a higher risk than parents, offspring, or second degree relatives. This research categorized IBD as a polygenic disorder in which multiple loci on chromosomes 16 and 12 are responsible for its behavior (Watts, 2002). Environmental factors, such as smoking, the use of oral contraceptives, appendectomy, and various infections also lead to IBD. Further research indicates that an over-proliferation of T cells may also contribute to uncontrolled inflammation due to its ubiquitous presence in patients with CD (Isaacs, 2005). Until further research determines the definite cause of IBD, the general theory hypothesizes that any factor interfering with the mucosal lining of the intestines or capable of initiating an inflammatory response can lead to IBD in genetically susceptible hosts (Angerio, 2005).
With no knowledge of the exact cause of IBD, there has also been no cure for it, but a variety of treatments ranging from immuno-suppression with azathioprine or 6-mercaptopurine to inducing remission in active IBD patients with sulfasalazine (Barrett, 2004). Other medications target the specific cytokine releases that occur with IBD which result in the intense inflammation. Drugs such as infliximab, operate against the tumor necrosis factor-α cytokine, while others, such as MRA suppress the release of interleukin-6, the most prominent lymphokine associated with IBD (Sandborn, 2004). Increased cytokine release could possibly denote that the over abundance of T cells within IBD patients does contribute to the cause of IBD since T cells synthesize and secrete these polypeptides. Nevertheless, the ambiguity of IBD continues to stimulate scientists to research this perplexing disease that affects many people around the world.
Recent research has associated the presence of IBD with the elevation of C-reactive proteins (CRP), an acute-phase protein discovered in 1930 in patients with pneumococcal pneumonia. These acute-phase proteins are responsible for initiating responses to inflammation by increasing or decreasing their plasma concentrations by at least 25 percent during inflammatory distresses. IBD patients may have a 1000-fold increase in their CRP levels in relation to the intensity and proliferation of the disease (Gabay, 1999). As a result of its active presence in this gastrointestinal disease, this research primarily focuses on its simultaneous role as an effective inflammatory marker, and a possible pro and anti-inflammatory agent in IBD.
The efficacy of a disease marker lies in its ability to aid clinicians in their attempt at diagnosis. Ideally, a disease marker should be able to determine an individual’s risk for a given disorder, monitor the progress of the disease activity, and regulate the effectiveness of a treatment (Nielson, 2000) (Vermiere, 2004). CRP possesses all of the aforementioned characteristics allowing it to act as an effective disease marker for IBD due to its inherent sensitivity towards inflammation when its plasma concentration levels increase dramatically (Vermiere 2004) (Pouilles, 2002). As an active marker for IBD, CRP differentiates and diagnosis patients with bowel disorders, determines disease severity, and measures the success of various treatments and drugs (Mazlam, 1994).
During an inflammatory response, interleukin-6 (IL-6), interleukin-1β (I-1β), and tumor necrosis factor-α (TNF-α), cytokines forming the immune system, stimulate the liver, the primary site of CRP production, to exponentially increase the circulating presence of CRP at the site of injury. Under normal circumstances plasma levels of CRP are approximated to be at 0.8 mg/L (Vermiere, 2004). In IBD patients, this level may increase to greater than 45 mg/L, a life-threatening stage predicting the need for a coloctemy (Vermiere, 2004). Increasing hepatocytes in the liver, allowing for a greater rate of CRP synthesis, cause such a life threatening stage. An increased CRP level measurement is direct evidence that the body has started to mobilize its defenses (Mazlam, 1994). Although an accurate marker for IBD, CRP is considered a less reliable marker of inflammation and disease activity in patients with UC, unless dealing with a very severe case, due to its overall lower CRP concentration (Vermiere, 2005). Recent research indicates that CRP serves as a marker not only for IBD, but to differentiate between IBD and functional bowel disorders (FBD). FBD, also known as Irritable Bowel Syndrome (IBS) is characterized by the presence of abdominal pain and discomfort (Evans, 2006). The key factor differentiating between FBD and IBD is the absence of structural abnormalities, especially inflammation. Rather, patients affected by FBD exhibit only functional disruptions altering their bowel habits (Bradesi, 2002). Furthermore, FBD does not have oncogenic potential, wherein lies the need for distinguishing between the bowel diseases.
A study from St. Mark’s Hospital in London, UK, found that CRP levels increased in all patients diagnosed with CD, and 50 percent of those diagnosed with UC after undergoing a clinical examination and a rectal biopsy. The diagnosis was based on the patient’s erythrocyte sedimentation rate (ESR), CRP, and al-glycoprotein levels. Taking a similar approach, Beattie el al., found that 100 percent of patients with CD and 60 percent of patients with UC demonstrated increased CRP levels (Vermiere, 2005). In a study performed by Poullis, it was found that CRP was found to have higher plasma concentration in IBD than FBD (Pouilles, 2002). All the aforementioned studies suggest that using a lower CRP cut-off value would increase the sensitivity therefore increasing the accuracy in IBD diagnosis. As a result of the direct correlation between IBD and CRP, CRP was found to be the best laboratory marker for the differentiation of patients with IBD from all other bowel disorders (Vermiere, 2005).
Disease activity, measured by the concentration of CRP present at the affected site, is greatly aided by its short half-life. The CRP half life of nineteen hours, unaffected by any physiological or pathophysiological circumstances, allows for the rapid decrease in CRP concentration during periods of reduced inflammatory activity in IBD (Nielson, 2000) (Vermiere, 2004). A linear relationship has been found to exist between the amount of CRP present and the clinical activity but more apparent for CD than UC. Although this linear relationship exists, absolute cut-off levels to distinguish between mild, moderate, and severe cases of the disease have yet to be determined. Consequently, it is believed that comparing CRP levels over a period of time gives more accurate results than absolute cut-off levels when determining IBD’s degree of severity in an individual. In a study by Fagan et al, it was found that although both CRP and ESR levels correlated with disease activity, CRP demonstrated a more accurate correlation, thus being a more accurate monitor of disease activity (Vermiere, 2005).
CRP, as a marker, contributes greatly in the determination of a drug’s effectiveness on IBD. The effectiveness of a drug on IBD is assessed through CRP changes after therapy. Anti-inflammatory and immunosuppressive drugs have no affect on CRP response. Consequently, any change in CRP response during treatment reflects the effect of the drug on IBD. The beneficial effect of a drug on intestinal inflammation presents a decrease in CRP, while persistently raised CRP levels indicate the impotence of a drug towards controlling inflammation (Vermiere, 2004). Louis et al’s study comparing patients with a baseline level CRP greater than 5mg/L and less than 5mg/L concluded that those individuals who started with the greater CRP concentration had a greater response to therapy. This raises the question of whether it is beneficial for research to include only those individuals with greater concentrations of CRP. Although the changes are more evident in such individuals, those with a CRP concentration less than 5 mg/L were still affected. Therefore, regardless of the severity of IBD, all patients show response when treated with beneficial drugs.
CRP serves as a pro-inflammatory agent through its interplay with the aforementioned cytokines: interleukin-6 (IL-6), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α). The following process describes the possible development of IBD through its associated interaction with CRP, a possible inducer of the inflammatory response. Some unknown agent initiates the course of the disease by stimulating the release of the three prominent cytokines, all of which have an effect on the extravasation of CRP. This protein, which is composed of five identical nonglycosylated polypeptide subunits, or protomers, is mainly produced by the hepatocytes within the parenchymal cells of the liver. (Pepys, 2003). The interplay of these cytokines: TNF-α, IL-6, and IL-1β, serve as an important consideration in determining the efficacy of each on the secretion of CRP. Evidence suggests that IL-6, which is produced mainly by macrophages and T cells, influences CRP levels significantly due to its direct ability to induce the hepatic release of acute phase proteins (Gabay 1999) (Nielson, 2000). Furthermore, IL-6 enhances the effect of IL-1β while simultaneously inhibiting the expression of TNF-α as shown in research conducted in mice (Gabay, 1999). While most current drugs, such as infliximab seem to target the TNF-α in the treatment of IBD, this research suggests that the focus on the therapeutics of IBD should be shifted due to the greater contribution of IL-6 as the main cytokine. Infliximab is not effective in patients with elevated
CRP since it cannot induce remission in substantial percentages of patients with IBD, more specifically with CD patients. (Sandborn, 2004) (Bamias, 2005). Further research indicates that endothelin-1, a potent vasoconstrictor also contributes to IBD due to its interaction with IL-6, which further enhances its role in the inflammation process (Angerio, 2005). Hence, IL-6 stimulates the greatest secretion of this cyclic pentameric protein, which begins to circulate and enter into the inflamed site of the large intestine or the small intestine depending on which of the two forms the disease assumes (CD being more apparent in the small intestine, while UC predominantly occurring in the colon and large intestines) (Angerio, 2005). Inflammatory cytokines thus stimulate CRP, which acts as a pro-inflammatory agent due its association with them.
Its pro-inflammatory role becomes further identifiable when CRP binds to phosphocholine residues, a constituent of the phospholipids of cell membranes and plasma lipoproteins (Vermiere, 2005). Extrinsic or autologous ligands, which are mainly modified plasma lipoproteins, damaged cell membranes, and other constituents of microorganisms may also form other CRP-ligand complexes. This chemical bond permits CRP to exemplify the key properties of antibodies, which induce inflammation through its activity with pathogens (Pepys, 2003). Hence, CRP has the ability to aggregate and precipitate the cellular structures bearing these ligands, by activating the complement system when it is recognizes C1q, a subunit of the C1 enzyme within this system (Pepys, 2003). Triggering this system results in opsonization and phagocytosis, both of which magnify the CRP-ligand complex, in an effort to make it more susceptible to the action of phagocytes (Vermiere, 2005). As a pro-inflammatory mediator, CRP can initiate the elimination of targeted cells by interacting with both the humoral and cellular immune responses of inflammation (Gabay, 1999). In its ability to behave as similarly as an antibody, CRP thus serves as an effective pro-inflammatory agent as observed in Figure A and B.
nhibition of lymphocyte extravasation and influx at the site of injury has been found to be an effective strategy in the treatment of IBD (Sandborn, 2004). Among its various roles, CRP functions as an anti-inflammatory agent by preventing the adhesion of neutrophils to endothelial cells and the inhibition of neutrophil chemotaxis (Gabay, 1999).
Neutrophil migration to sites of inflammation involves a complex interaction of leukocytes with endothelial cells via regulated expression of surface adhesion molecules. The initial attachment of neutrophils to endothelium is mediated by L-selectin, a cell adhesion molecule constitutively expressed by neutrophils (Zouki, 1997). In an experiment analyzing the reaction between L-selectin saturated endothelial cells and neutrophils incubated with CRP or CRP peptides, Zouki et al demonstrated that CRP and CRP peptides affect the L-selectin mediated interaction of neutrophils with endothelial cells, the first step of neutrophil extravasation. CRP and CRP peptides 174-185 and 201-206 bind to receptors on neutrophils and inhibit the neutrophil-endothelial adhesion by the cleavage and shedding of L-selectin, while CRP peptides 77-82 had no effect. Neutrophils treated with CRP and CRP peptides were unable to adhere to endothelial cells due to the loss of L-selectin expression caused by the CRP treatment (Zouki, 1997).
Modified forms of CRP are also characterized by this anti-inflammatory feature. In the absence of calcium, a chemical necessary for CRP’s pentameter structure, CRP dissociates into individual subunits that cannot assume the original pentameter structure but instead form an aggregation of monomers. The general term used to describe these isoforms of CRP is modified-CRP (mCRP) (Vermiere, 2004) (Heuertz, 2005). Despite the differences in structure, mCRP is also able to inhibit neutrophil adhesion to endothelial cells. As mCRP binds to the IgG receptor FcγRIII (CD16) found on neutrophils, the shedding of L-selectin occurs and adhesion is blocked (Vermiere, 2004).
Furthermore, there is growing evidence that CRP naturally exists in multiple polymorphic forms. One such form, a CRP-mCRP complex, increases the inhibition of neutrophil-endothelial adhesion. Studies show that the binding of CRP to neutrophils is increased in the presence of mCRP concentrations greater than 10 µg/mL (Zouki, 1997). Heuertz et al. suggest that native CRP binds to mCRP which then binds to CD16, an indirect method for halting phagocytic action and migration. As the neutrophil-endothelial adhesion fails due to binding with CRP, CRP peptides, and mCRP, neutrophilic migration to injured areas diminishes, thereby suggesting the possibility of CRP as a major mechanism to limiting the inflammatory response in IBD.
Various experiments explain how CRP functions as both an inflammatory and anti-inflammatory agent at once. Experiments involving transgenic mice concluded that CRP functions as both an inflammatory and anti-inflammatory agent; however its net effect on each individual test mouse was anti-inflammatory (Gabay, 1999). It is also suggested that CRP starts out as an inflammatory agent and reverses its role over time. In its role as an opsonin, CRP results in an influx of white blood cells into the injured area further exaggerating the inflammatory response. Eventually, phagocytosis of bacteria will result in the decreased production of pro-inflammatory cytokines and the increased production of anti-inflammatory cytokines (Vermiere, 2004). While this explanation sufficiently describes CRP in any inflammatory site, it can not be directly applied CRP’s role in IBD as it is yet unknown if bacteria are the cause of inflammation in IBD. Further research into the causes of IBD will reveal more clearly CRP’s role as an inflammatory agent, an anti-inflammatory agent, or both.
Further study of the CRP-neutrophil binding site explains how CRP’s anti-inflammatory effect is not observed in all instances of high CRP presence. For CRP to function as an anti-inflammatory agent and inhibit neutrophil-endothelial adhesion, it must bind to a specific receptor on neutrophils. Recent reports suggest that FcγIIa (CD 32) is the receptor for CRP on neutrophils (Heuertz, 2005). However, not all individuals are able to bind CRP, only those with FcγIIa of the R/R131 allotype can. Only 25% of the American population carries this phenotype (Heuertz, 2005). As a result, only individuals with CD32 R/R131 bind CRP, inhibit neutrophil chemotaxis, and exhibit CRP’s anti-inflammatory role. This might explain why high levels of CRP in IBD and other inflammatory diseases do not always correspond with anti-inflammatory reactions. It might also explain why IBD is so prevalent in the United States and Western countries as opposed to other areas of the world.
IBD coexists significantly with the presence of CRP due to the various roles this protein can assume in effected patients. As an inflammatory marker, it has the ability to predict, monitor, and evaluate IBD in terms of its presence, severity, and therapeutics. Its role as an indicator of the disease corresponds directly with its presence in simultaneously inducing and inhibiting inflammation. This contradictory coexistence as a pro and anti-inflammatory agent in IBD, however, seems to create a biological dichotomy, which requires further research into the causes of IBD to determine its particular role. While it prominently seems to inhibit inflammation, without further research and experimentation, there is no accurate conclusion and therefore no appropriate therapeutic measures. If the latter does prove to be true, however, CRP could be utilized as an artificial drug in the treatment of IBD, a novel discovery which could significantly impact society and the scientific community. Yet, if it is an inflammation inducer, research could be aimed at using drugs that inhibit CRP, itself, instead of the current therapeutic measures, which target the specific interleukin associated with CRP. As continual research with CRP progresses, IBD seems to have a more optimistic future, one in which involves the leading participation of this acute phase protein.
Angerio, Allan D, Dominick Bufalino, Melissa Bresnick, Christine, Sarah Brill.
“Inflammatory Bowel Disease and Endothelin-1.” Critical Care Nursing
Quarterly. 2005; 28: 208-213.
Bamias, Giorgos, Mark R. Nyce, Sarah A De La Rue, and Fabio Cominelli. “New Concepts in the Pathophysiology of Inflammatory Bowel Disease.” Annals of Internal Medicine. 2005; 143: 895-904.
Barrett H. Barnes. Stephan M. Borowitz, Frank T. Saulsbury. Martha Hellem, and
James L. Sutphen. “Discordant Erythrocyte Sedimentation Rate and C-reactive
Protein in Children With Inflammatory Bowel Disease Taking Azathioprine or 6-
Mercaptopurine.” Journal of Pediatric Gastroenterology and Nutrition. 2004; 38:
Bradesi, Sylvie, James A. McRoberts, Peter A. Anton, Emeran A. Mayer. “Inflammatory Bowel Disease and Irritable Bowel Syndrome: Separate or Unified.” Current Opinion Gastroenterology. 2002; 19: 336-342.
Cabrera-Abreu, J C, P Davies, Z Matek, M S Murphy. “Performance of blood tests in diagnosis of inflammatory bowel disease in a specialist clinic.” Archives of Disease in Childhood. 2004; 89: 69-71.
Evans, BW, WK Clark, DJ Moore, PJ Whorwell. “Tegaserod for the Treatment of Irritable Bowel Syndrowm.” The Cochrane Database of Systematic Reviews. 2006; 1.
Gabay, Cem, and Irving Kushner. “Acute-Phase Proteins and Other Systematic
Responses to Inflammation,” The New England Journal of Medicine, 1999; 340:
Heuertz, Rita M, Gregory P. Schneider, Lawrence A. Potempa, and Robert O. Webster. “Native and modified C-reactive protein bind different receptors on human neutrophils.” The International Journal of Biochemistry & Cell Biology. 2005; 37: 320-335.
Isaacs, Kim L, James D. Lewis, William J. Sandborn, Bruce E. Sands, and Stephan R.
Targon. “State of the Art: IBD Therapy and Clinical Trials in IBD.”
Inflammatory Bowel Disease, 2005; 11: S3-S12.
Mazlam, M.Z. and HJF Hodgson. “Why measure C reactive protein?” Gut.1994;
Nielson, Ole H, Ben Vainer, Soren M. Madsen, Jacob B. Seidelin, and Neils H.H. Heegaard. “Established and Emerging Biological Activity Markers of Inflammatory Bowel Disease.” The American Journal of Gastroentrerology. 2000; 95:359-360.
Pepys, Mark B. and Gideon M. Hirshfield. “C-reactive protein: a critical update.” Journal of Clinical Investigation. 2003; 111: 1805-1812.
Pouilles, Andrew P. Sameer Zar, Krishna K. Sundaram, Simon J. Moodie, Paul Risley, Andrew Theodossi, and Michael A. Mendall. “A new, highly sensitive assay for C-reactive protein can aid the differentiation of inflammatory bowel disorders from constipation-and diarrhea-predominant functional bowel.” European Journal of Gastroenterology and Hepatology. 2002; 14:409-410.
Sandborn, W J, and W A Faubion. “Biologics in inflammatory bowel disease: how
much progress have we made?” Gut. 2004; 53: 1366-1373.
Torres, M.I., M. Le Discorde, P. Lorites, A. Rios, M.A. Gassull, A. Gil, J.
Maldonado, J. Dausset, and E.D. Carosella. “Expression of HLA-G in
inflammatory bowel disease provides a way to distinguish between ulcerative colitis and Crohn’s disease.” International Immunology. 2004;16: 579-583.
Vermeire, Séverine, Gert Van Assche, and Paul Rutgeerts. “C-reactive Protein as a Marker for Inflammatory Bowel Disease.” Inflammatory Bowel Disease. 2004; 10:661-665.
Vermiere, Séverine, Gert Van Assche, and Paul Rutgeerts. “The role of C-
reactive protein as an inflammatory marker in gastrointestinal diseases.”
Gastroenterology & Hepatology. 2005; 2: 580-586.
Watts, D A, and J Satsangi. “The genetic jigsaw of inflammatory bowel disease.”
Gut 2002; 50: iii31-iii36.
Zouki, Christine, Micheline Beauchamp, Chantal Baron, János G. Filep. “Prevention of in Vitro Neutrophil Adhesion to Endothelial Cells through Shedding of L-Selectin by C-reactive Protein and Peptides Derived from c-reactive protein.” Journal of Clinical Investigation. 1997; 100 522-529.