ULCERATIVE COLITIS

 

 Ulcerative colitis (UC) and Crohn's disease (CD) are idiopathic, chronic, frequently disabling, inflammatory bowel diseases (IBD).

UC is characterized by mucosal inflammation limited to the colon, always involving the rectum and a variable extent of the more proximal colon in a continuous manner.

CD inflammation is transmural, most often discontinuous and may involve any portion of the gastrointestinal tract but most commonly involves the distal ileum. The prevalence of IBD in the United States is 200–300/100 000 with a similar prevalence for UC and CD.

IBD is considered a complex genetic disorder predicted to involve multiple genes of relatively low penetrance, since the familial patterns of inheritance do not conform to simple Mendelian models.

Overall, 10–20% of individuals with IBD report one or more additional relatives with IBD. Relatives of CD patients have a 10-fold risk of developing CD and relatives of UC patients have an 8-fold risk of developing UC. However, these diseases appear to be genetically related, as relatives of CD patients have a 4-fold risk of developing UC and relatives of UC patients have a 2-fold risk of developing CD.

 

An important candidate gene for IBD is the NFKB1 gene located at chromosome 4q24.

Nuclear Factor-B (NF-B) proteins are a family of transcription factors that regulate various biological defense processes, most notably innate and adaptive immune responses, acute phase reaction and apoptosis.

There are five members of the NF-B family in mammals: p50/p105, p65/RelA, c-Rel, RelB and p52/p100. Although many dimeric forms of NF-B have been detected, the major form of NF-B is a heterodimer of the p50 and p65/RelA subunits, encoded by the genes NFKB1 and NFKB2, respectively .

Human NFKB1 encodes two proteins, a 105 kDa, non DNA-binding, cytoplasmic molecule (p105) and a 50 kDa DNA-binding protein (p50) that corresponds to the N-terminus of p105. The NFKB1 gene spans 156 kb and has 24 exons with introns varying between 40 000 and 323 bp in length.

In most cells before stimulation, NF-B primarily resides in the cytoplasm in inactive complexes through association with a sequestering inhibitory protein, termed IB.

A wide range of stimuli, including bacterial and viral products, cytokines and oxidant-free radicals, activate NF-B. These stimuli promote NF-B nuclear translocation by a mechanism that involves IB phosphorylation and the ubiquitin-proteosome pathway. This phosphorylation appears to target IB for degradation and leads to its dissociation from the NF-B complex and subsequent translocation of NF-B to the nucleus. There, active NF-B binds to genomic DNA at promoter regions and thereby regulates gene transcription.

 

Inappropriate activation of NF-B has been implicated in inflammation associated with a variety of human diseases and pathologic conditions, among them asthma, inflammatory arthritis, septic shock, lung fibrosis, diabetes, cancer, AIDS, atherosclerosis, stroke and IBD. Furthermore, several anti-inflammatory and anti-cancer drugs work in part through inhibition of NF-B activation. For example, aspirin and glucocorticoids inhibit NF-B. Consistent with NF-B regulation of genes involved in the immune and inflammatory responses, mice null for several of the NF-B subunits show defects in clearing bacterial infection along with defects in B-cell and T-cell functions.

 

NF-B has a central pathogenic role in chronic intestinal inflammation. Using immunohistochemistry methods, in the inflamed intestinal mucosa of CD and UC patients, activated NF-B was increased and found localized to the macrophages and epithelial cells. Schreiber et al. similarly found CD and UC patients had increased NF-B activity in intestinal lamina propria cells.

Additionally, the therapeutic properties of mesalazine and sulfasalazine (the most common specific medical therapies for mild to moderate UC), rely in part on inhibition of NF-B activation Three CD associated mutations in the NOD2/CARD15 gene on chromosome 16 all have a defect in their ability to activate NF-B. Recent evidence suggests that this may result in a defect in the innate immune system's ability to protect the gut against invasive bacteria.

 

 

Laboratory investigations in UC

 

I will divide the testing into categories.

Genetic.

(1)            We are interested in HLA haplotypes as DR2 and DRB*0103 have some part to play in susceptibility.

(2)            NOD2 (also called CARD15) This gene on Cr16 codes for a nucleotide-binding oligomerization domain protein 2, located in the cytoplasm and possessing the ability to sense peptidoglycan fragments of bacterial origin. Following this action, comes activation of NFkappa beta. This is only one of the ways that NFkb can be activated.

(3)            ICAM R241 allele.

(4)            IL1 receptor antagonist allele.

(5)            Multi-drug resistance gene1a

(6)            Toll like receptor 5 (The TLR5 receptor recognizes and binds to flagellin of bacteria.

 

Immunology

 

We can test for

(1)            IgG levels and subsets and also IgA.

(2)            T cell subsets

(3)            Serum Cytokines, including IL1, IL6, TNF alpha, and Salivary IL6. IF gamma is largely located in tissue.

(4)            pANCA, ASCA antibodies, antigoblet cell antibodies, antipancreatics antibodies and anti-tropomyosin isoform 5 antibodies.

(5)            Complement and mannose binding lectin(this is now available through IMVS).

 

Histopathology

Careful examination of cell types as well as special and immunofluorescent stains.

 

Faecal studies, looking for

   (1) Distribution and numbers of normal flora

   (2) Pathogens

   (3) Bacterial toxins

   (4) PCRs for a range of these organisms

 

Blood factors and inflammatory indices

(1) Fibrinogen and von Willebrand factor.

(2) ESR, CRP and matrix metalloproteinase 9.

(3) Liver function.

 

Metabolic abnormalities,

 Vitamin 25 D3 and 1,25 dihydroxy vitamin D3.

Here we are looking for susceptibility because of Vitamin D3 deficiency, as this has been reported in UC and CD.

 

We will also look for overconversion of D3 to 12,25 D3 because of activated macrophages, which possess the 1 alpha hydroxylase needed for this conversion,

 

Nutritional deficiencies.

These may be primary or secondary.

Deficiencies of zinc may impair healing.

 

Therapy can also be divided into categories.

 

Anti-inflammatory agents.

(1)            Mesalazine. This inhibits NFkappa beta activation.

 has been taking suboptimal dosing.

(2)            Corticosteroids.These can be rectally or systemically administered..

(3)            Quercetin. This flavenoid inhibits Matrix Metalloproteinases, PLA2, Lipoxygenases and other inflammatory enzymes.

(4)            Lyprinol. This markedly decreases production of leukotrienes.

(5)            Other inhibitors of NFKB activation. Hops (isohumulones) and nettle contain such inhibitors.

 

 A product from Metagenics called Kaprex, contains hops (humulus lupulus), Rosemary and Olive leaf and has marked anti-inflammatory activity.

(6) Lactoferrin has been shown to have anti-microbial actions against pathogens and candida, but protection to normal flora as well as anti-inflammatory actions.

 

Since there is amazing similarity between angiotensin2 receptors and CCR2b receptors (on monocytes) some angiotensin 2 receptor blockers are anti-inflammatory and may also block the CCR2b receptors.

 

Bacterial products, monocyte chemotactic protein-1 and some cysteinyl leukotrienes are ligands for the CCDR2b recptors and evoke activation of IK B kinase thus allowing phosphorylation and activation of NFkappa beta. This then translocates to the cell nuclei where TNF alpha and IF gamma genes are activated to produce their products.

 

TNF alpha induces the genes for inducible nitric oxide synthase called inducible nitric oxide synthase (INOS).  The nitric oxide which forms reacts with superoxide in mitochondria to produce the peroxynitrite, a more reactive free radical. 

 

Superoxides are also a product of oxidative injury in cells in circumstances of chemical and microbiological damage.

Mitochondrial DNA is some ten times as susceptible to free radical injury as is nuclear DNA.                                        

   PEROXYNITRITE EFFECTS.

 

(1)            The peroxynitrite inactivates superoxide dismutase in mitochondria increasing the levels of superoxide.

 

(2)            But in addition the peroxynitrite activates a transcription factor called nuclear factor kappa beta (NFKb) which stimulates gene transcription for ILI, IL6, TNF alpha and IF gamma, and also gene transcription for inducible nitric oxide synthase.  (A self perpetuating cycle)

 

Thus both superoxide production (and decreased degradation) and nitric oxide production are kept high, perpetuating a higher level of peroxynitrite.

               

(3)            Peroxynitrite contributes to a decrease in ATP pools in the following fashion  (a further feed forward effect).

 

An enzyme called poly adenylate ribose synthase is activated by breaks in DNA.  Free radicals produce "nicks" in mitochondrial and nuclear DNA. 

 

The PARS promotes polyribosylation of histones, and the substrate NAD involved in all oxidative and energy metabolism conversion to NADH is depleted. 

 

 

Dietary modification

 

This includes exclusion of any suspect foods, and supplements with minerals. vitamins and antioxidants.

 

I tend to favour supplements of zinc, magnesium, selenium and molybdenum, along with reduction of omega 6 and trans fatty acids and addition of fish oils, co-enzyme Q10, vitamin E as mixed tocopherols, alpha lipoic acid, vitamin C in small frequent doses, and small amounts of extra B vitamins.

 

Probiotics such as Bioceuticals “Symbiotique” are important.

 

Possible future steps.

 

We need to know where either microbial products such as toxin (haemolysins, proteases etc) or components such as flagellin or cell wall lipopolysaccharides increase inflammatory activations and cell injuries.

 

It may be that staphylococci and mycoplasmas also act as super antigens greatly amplifying T cell activations.

 

The reading of microbial components by Toll-like receptor on macrophages can set going a macrophage production of specific cytokines.

 

An example is IL6, which on reaching the liver acts to induce the liver to produce mannose binding lectins and this in turn increases complement activations in the polymorph destruction of bacteria outside the cell, especially in the presence of specific antibodies.

 

Defects in the genes coding for mannose binding lectins may well allow impaired responses to some organisms.

 

Again local injury to colonic cells may activate irreversible damage cycles.

 

The future will include knowledge of each step in these activations in order to break these vicious cycles, at a number of points.

 

This will mean more specific and safer therapies.

 

Zinc is essential for health. It’s needed for the enzymes that regulate cell  division, growth, wound healing, and proper functioning of the immune  system. Zinc is an essential co-factor in a variety of cellular processes  including DNA synthesis, behavioral responses, reproduction, bone  formation, growth and wound healing. Zinc is a component of insulin and it  plays a major role in the efficiency of most of the functions of the body.  Zinc is necessary for the free-radical quenching activity of superoxide  dismutase (SOD), a powerful antioxidant enzyme which breaks down the  free-radical superoxide to form hydrogen peroxide. Zinc is required for  the proper function of T-lymphocytes. The mineral also plays a role in  acuity of taste and smell. And zinc is required for proper functioning of  genetics, immunity, formation of red blood cells, organ, muscle and bone  function, cell membrane stability, cell growth, division, differentiation  and genetics. Importantly, zinc is vital for the metabolism of Vitamin  A.