Most of the concern about the safety of nitrates has arisen from epidemiological data on their potential to form N-Nitrosamines; these are thought to be cancer causing and also responsible for causing “blue-baby” syndrome. However, one must be careful in evaluating the data and when drawing any conclusions. Let us review the data for both these concerns. i) Blue-baby syndrome (methemoglobinemia) One case back in the 1950’s caused considerable sensation when a baby’s hemoglobin was thought to be compromised by supposedly excessive nitrates in well water and converting the hemoglobin to a form that was not able to transport oxygen easily.
More and more studies are showing the potentially damaging effects of chronic inflammation and its link to the development of various serious chronic diseases. Research has now identified chronic inflammation as a factor in the progression of cancer, diabetes, cardiovascular disease and Alzheimer’s disease, along with many others! The link between inflammation and these diseases is discussed in the following sections.
Inflammation and Cancer
In his 1971 State of the Union address President Richard Nixon pledged “I will ask for an appropriation of an extra $100 million to launch an extensive campaign to find a cure to cancer. Let us make a total national commitment to conquer this dread disease. America has long been the wealthiest nation in the world. Now it is time we become the healthiest nation in the world”. This appropriation led to the creation of the National Cancer Institute (NCI). Since then over 200 billion dollars has been spent, 1.56 million papers published, and 150,855 studies in mice reported. In spite of this, the US cancer death rate was the same in 2002 as it was in 1950 (193.9 per 100,000 versus 193.4 per 100,000)! Hopefully, however, research will continue, and we will move closer to finding the key to this deadly disease.
There is evidence that chronic inflammation is an important contributing factor. In general, cancers are a chronic disease caused by the prolonged exposure to a long list of damaging stimuli. These stimuli can include: infections or toxins like aflatoxin, hormonal insult like excess estrogen exposure post menopause, chronic irritation from tobacco smoke, carcinogens or asbestos exposure, a diet high in fats or sugar, alcohol, radiation and many more.
In the early 1860’s the renowned German pathologist Rudolf Virchow mentioned that certain cancers seemed to occur at sites of chronic inflammation. He based his hypothesis on the fact that certain irritants, together with long standing tissue injury invoked inflammation that ultimately led to cancer. In fact, all of the damaging stimuli listed above could contribute to chronic inflammation. There is a significant body of evidence demonstrating that chronic inflammation over a long period of time (20-30 years) can lead to cancer. For example, inflammation of the bronchus (bronchitis) due to the constant irritation from tobacco smoke can lead to cancer of the lung. Similarly, inflammation of the bladder (cystitis), colon (colitis), esophagus (esophagitis) and liver (hepatitis) leads to cancers of these tissues. Unfortunately, in the 1860’s this link between inflammation and cancer was considered too bold and largely went ignored. Recently, however, studies conducted in large groups of people have reinforced Virchow’s original observation suggesting a strong relationship between stimulators of inflammation and cancer. These epidemiological studies have identified stimulators of inflammation in the form of chronic irritants like tobacco smoke, alcohol, fried foods, UV light, infections, stress, red meat, grilled food, various heavy metals, solvents and trauma as major risk factors in various types of cancer. For example, the human papilloma virus has been linked to cervical cancer and chronic viral hepatitis B and C (HBV and HBC) infections have been identified as major risk factors for a specific liver cancer called hepatocellular carcinoma. Similarly, the bacteria Helicobacter pylori which is widely present in the stomach of most people world-wide and has a propensity to burrow its way into the lining of the stomach wall, is not only a primary contributor to the development of ulcers, but has also been strongly linked to the development of stomach cancer. Other chronic infections and/or inflammation like inflammatory bowel disease (IBD) also increase the risk of cancer of the colon. In fact it is estimated that approximately 15-20% of all human cancers are linked to infection and/or inflammation!Perhaps the best evidence for the significance of inflammation during cancerous growth comes from long term users of aspirin and non-steroidal anti-inflammatory drugs. Recent data indicates that these drug users reduce colon cancer risk by 40-50% and they also may be preventive for other cancers including lung, esophagus and stomach cancers. Additionally, other stronger NSAID’s have been shown to prevent the metastases or spread of certain forms of cancer. A summary of the cancers linked to chronic inflammation is listed in Table 3.
Inflammation is rightly regarded as a “secret killer” in diseases such as cancer. It is clear that there is a link between the two. However, this begs the question, how is inflammation linked to cancer? What is the mechanism of action? Which cellular processes are involved? If we can understand these things then perhaps we can use specific nutrients to target the key molecules or mechanisms that are involved in the development of cancer. Cancer is a multistage process defined by at least three main stages: initiation, promotion, and progression (Figure 2). These stages can then be further subdivided into various steps, including cellular transformation (where cells change their look and behaviour), promotion, survival, proliferation (multiplication of cells), invasion, angiogenesis (formation of blood vessels so that the tumour can be continually fed and grow), and metastasis (spreading of cancer to distant sites). These stages, while distinct, overlap considerably and involve various players like initiating compounds, signaling molecules, growth factors and numerous other mediators in a complex role play that links chronic inflammation to all of these stages. A list of these various signaling molecules, growth factors and other key players linking inflammation and cancer are described in Table 4, and their actions in the body and their contribution to inflammation and cancer initiation are shown in Figure 3.These molecular players are legitimate targets for the anti-cancer effects of various nutrients. For example, it has been shown that certain anti-inflammatory agents like the non-steroidal anti-inflammatory drugs (NSAID’s) reduce chronic inflammation and thus reduce the incidence of various cancers. Unfortunately, some of these NSAID’s are themselves harmful, and cause side-effects like gastric irritation and liver and kidney toxicity. However, it is possible that certain natural compounds could target such pathways without having the associated side-effects and thus could be used to help prevent or reduce the incidence of cancer.
Inflammation can work both ways with short term inflammation being protective and having anti-cancer effects, while long term inflammation can cause cancer. The intent of many natural products is to reduce chronic inflammation by targeting one or more of the above pathways that involve numerous inflammatory players including NF-κB, which plays an enormous role in inflammation and is discussed in greater detail in the next article, as well as various cytokines, chemokines, growth factors and hormones. For example, it has been noted that NF-κB is raised in every inflammatory condition, with especially strong links to cancer. Bharat Aggarwal of The University of Texas M.D. Anderson Cancer Center in Houston, Texas is one of the foremost researchers on NF-κB and its association with cancer. After thirty years of research, Aggarwal is convinced that NF-κB is the key culprit in cancer. According to Aggarwal all roads to cancer go through this single molecule!Several lines of evidence points to the key role NF-κB plays in inflammation and cancer. NF-κB has been associated with every known carcinogen and has also been linked with other inflammatory mediators like COX-2 and 5-LOX enzymes, tumour necrosis factor (TNF) and others. Furthermore, NF-κB levels are dramatically reduced by anti-cancer agents like natural polyphenols, flavonoids, vitamin E, lycopene, vitamin C and many others. All these findings provide compelling evidence that NF-κB is likely a major mediator of cancer.
Inflammation and Diabetes
The association between a reduced production of insulin and type-2 diabetes has been considered the hallmark of this condition. This reduction in insulin production causes an inability of insulin to transport sufficient quantities of glucose from the blood and into the tissues. The net result of this is that glucose is excreted into the urine. In fact, early physicians remarked that the high sugar content of the urine produced by diabetic patients could be used as a diagnostic marker of the disease. The insulin producing cells are called Beta cells and they are located in the islets of Langerhans in the pancreas. In diabetic patients, these Beta cells are either destroyed during the course of the disease and/or are unable to be regenerated quickly to produce sufficient quantities of insulin to handle the body’s requirements. However, type-2 diabetes is not only associated with a decreased production of insulin but also with a more recently discovered phenomenon called insulin resistance. Insulin resistance is a condition in which insulin becomes less effective at lowering glucose levels in the body even when it is present at normal levels.Type-2 diabetes is associated with a number of long term consequences including atherosclerosis or hardening of the blood vessels which leads to poor circulation and a loss of vessel elasticity as well as eventual heart disease and high blood pressure. Furthermore, diabetes can also affect smaller vessels, like those in the eyes, kidneys and the central nervous system, causing retinopathy, nephropathy and neuropathy which results in damage to the nerves, kidneys and blindness.Several theories have been proposed to help explain why some individuals develop inadequate insulin production and insulin resistance. One of the most credible of these theories is inflammation. The link between inflammation and type-2 diabetes has become increasingly strengthened by the results of numerous scientific studies in both animals and humans. There are several factors that may induce chronic inflammation in the body. Those most critical to the development of type-2 diabetes are shown in Table 5.
There are several lines of evidence that implicate inflammation in the development of diabetes. The strongest is data from both animal and human studies that shows raised levels of key inflammatory markers like C-reactive protein (CRP) or certain interleukins like IL-1B or IL-6 in patients or animals with Type-2 diabetes. Moreover, these studies have shown that the higher the levels of these inflammatory markers, the greater the chance the individual has diabetes. In fact, it has been argued that raised levels of these inflammatory biomarkers indicated that inflammation also results in the activation of the immune system, which may then actually attack various tissues of the body. Diabetes specialists call this auto-inflammatory disease which is similar to an auto-immune disease in that it results in the body attacking its own tissues; the difference is that the primary cause of an auto-inflammatory disease is inflammation. The second strong evidence of a link between inflammation and type-2 diabetes comes from biopsies of various tissues that show clear evidence of inflammatory cell involvement.
Despite the overwhelming evidence of the involvement of inflammation in diabetes, it remains uncertain whether inflammation is a cause of the disease or whether it is a resulting symptom. It is a classic case of a chicken or the egg scenario, which are so common in biological science. If inflammation is in fact a cause of diabetes, then we must examine the mechanisms by which this occurs. That is, how can inflammation cause type-2 diabetes? Research in this area has suggested that inflammation may lead to diabetes by causing cell death and by creating hypoxic conditions; this means that inflammation causes an environment in which the tissues are literally starved for oxygen, leading to cellular death.Multiple mechanisms may contribute to increased inflammation in type-2 diabetes, some of which are quite general and others that are highly specific. In the pancreas, inflammation may be initiated by excessive nutrients like glucose or free fatty acids that can activate the immune system (see Table 5). There are a large number of biochemical pathways and molecules that are implicated in the inflammatory process; however, as with cancer, the most extensively studied of these in diabetes is the NF-κB pathway.
Treatments to Reduce Inflammation in Type-2 Diabetes
Evidence for the role of inflammation in type-2 diabetes is quite strong and new treatments that block the activation of various inflammatory markers like NF-κB or interleukin are actively being developed. Research is beginning to show that an anti-inflammatory approach to treatment of the disease seems to lower blood glucose levels and improve insulin release as well as helping to limit the damaging effects that ensue. In particular, anti-inflammatory treatments appear to be very helpful for reducing the glycation of hemoglobin (also referred to as HbAC1) which is a commonly used marker for diagnosing diabetes. Glycation is a major factor Advances 15in the development of diabetic complications. Glycation is a chronic condition brought upon by consistently high levels of glucose in the blood. The resulting effect is that glucose molecules bind to various proteins in the body like hemoglobin or other proteins in the plasma, tissues or blood vessels. When a protein is glycated it becomes warped, which changes the structure and therefore the function of the protein. In the case of blood vessels, for example, glycation may result in a weakening of the blood vessel walls or a reduction in elasticity. These effects can lead to increased blood pressure and an increased incidence of vessel rupturing. The glycation of hemoglobin can be a major problem in diabetes. The use of anti-inflammatory compounds in the treatment of diabetes has also been shown to improve the release of insulin by the Beta cells. Many of these studies validate the potential of targeting inflammation as a therapeutic approach to treating type-2 diabetes and support a causative role of inflammation in this disease. Obviously more work is needed to test the effects of either single anti-inflammatory compounds or cocktails that will target the various pathways of inflammation involved in diabetes to achieve improved results. Nevertheless, there is ample evidence of a significant role of inflammation in type-2 diabetes as either the underlying cause or a resulting symptom that needs to be addressed.
Inflammation and Cardiovascular Disease
Cardiovascular disease includes a spectrum of closely related conditions associated with the heart and circulation and is the leading cause of death in the western world. Conditions included under the umbrella of Cardiovascular Disease include: heart failure, angina, heart attacks, high blood pressure and the formation thrombi, or large blood clots, which are the primary cause of stroke.Peter Libby of Harvard Medical School was among the first researchers to uncover a connection between inflammation and cardiovascular disease (CVD). Libby was studying atherosclerosis, a condition that results in hardening of the arteries due to the accumulation of plaque in the blood vessels.
This plaque can then lead to further damage to the vessels, and a large number of cardiovascular complications including high blood pressure and heart attacks. In the 1970’s, most researchers had accepted that there is a connection between high fat intake and atherosclerosis. Later, it was found that the type of fat intake is also important. For example, the consumption of saturated fats and especially trans fats, is largely responsible for the development of atherosclerosis. However, researchers still lacked a clear understanding of the sequence of events involved in the initiation of atherosclerosis. In piecing together this sequence of events, Libby noted that immune system cells associated with inflammation, like macrophages, were the first cells to arrive at the scene of blood vessel damage. Based on this initial observation, Libby then pieced together the events of atherosclerosis, much like a forensic scientist solving a crime. The first step of atherosclerosis involves cholesterol, which is a large molecule that in and of itself is not harmful, and has an important role in the healthy functioning of the body. For example, cholesterol plays an important role in the synthesis of various hormones (like estrogen, testosterone and stress hormones) Advances 17and is an important component the membranes lining the cells of the body. Unfortunately, the cholesterol molecule has a number of protruding arms called hydroxyl groups that are especially susceptible to damage by highly reactive molecules called free radicals. These free radicals include reactive oxygen and nitrogen species that commonly lurk in areas of the body that experience high levels of stress, like the blood vessels. When it is attacked by free radicals cholesterol becomes oxidized or nitrated, meaning that an oxygen or nitrogen atom is added to the molecule. It is this damaged cholesterol that is the “bad” cholesterol that gets deposited on the walls of the blood vessels as a fatty build-up. When the immune system detects this aberrant and “non-self ” form of cholesterol building up in the blood vessels it acts quickly to send the various immune cells into action. The damaged or oxidized cholesterol causes immune system cells, called monocytes to latch onto the walls of the blood vessels.
These cells then migrate into the walls of the vessel, and transform into macrophages which initiate the body’s inflammatory response and begin to devour cholesterol molecules. These cholesterol filled macrophages are called “foam cells” and form the fatty lipid core of the plaque. Immune cells also act to cordon off the foreign cholesterol molecules by forming a fibrous capsule around the lipid core to help prevent it from spreading to other areas. Unfortunately, there are usually many affected areas, which results in these capsules being formed in many areas along the walls of the blood vessels.At this point, the cholesterol filled foam cells begin to burrow deeper into the vessel wall and into the muscle layer around the blood vessel. The muscle cells in this layer respond to the aggressive expansionist behaviour by multiplying further. In the midst of this battle, other immune players send out signals to recruit more cells and immune system factors to the site of damage, and also release various inflammatory cytokines, resulting in a continuing loop of increasing inflammation. Eventually, the plaque can rupture, leading to the formation of a blood clot, which in turn can cause a heart attack and possible death. The details of plaque formation in the arteries are shown in Figure 5. This series of events suggests a connection between a chronic state of inflammation and the progression of atherosclerosis. Inflammation plays a role at every stage in the progression of atherosclerosis. In fact, very early atherosclerotic lesions are almost purely inflammatory in nature, consisting of a collection of fat laden inflammatory immune cells like macrophages. As atherosclerosis progresses, even more immune cells infiltrate the region of plaque formation, where they are a component of the cap covering the lipid core of the plaque. These immune cells exhibit signs of activation and release various inflammatory cytokines. When plaque rupture occurs, this happens at areas where the cap is thinnest. These areas tend to be highly abundant in activated immune cells which produce high levels of inflammatory molecules and various enzymes that can weaken the cap and activate cells in the core. This process transforms the stable plaque into a vulnerable, unstable structure that can rupture, leading to thrombosis or clot formation. Overall, Libby’s major contribution was to link high levels of various inflammatory markers like C-reactive protein (CRP) with the extent of inflammation and the extent of vascular damage. This connection between inflammatory markers and cardiovascular disease progression provides a series of predictive markers that may be able to be used to help identify patients with plaque build-up and the beginning stages of atherosclerosis earlier, allowing treatment to give at a an earlier stage of the disease. The value of these predictive, inflammatory markers has been shown in a variety of studies. Research has clearly shown that elevated inflammatory markers are associated with increased cardiovascular risk among healthy individuals as well as those at higher risk. For example, Lindahl and colleagues found that in patients with unstable coronary artery disease, levels of the inflammatory marker CRP was directly and strongly related to the long-term risk of death from cardiac causes (see Figure 7). Furthermore, various prescription drugs that are used to treat various aspects of cardiovascular disease, like statins, aspirin, fibrates and ACE inhibitors have also been shown to have the effect of reducing levels of various markers of inflammation, especially in patients with very high levels to start with.
Inflammation and Alzheimer’s Disease
Alzheimer’s Disease (AD) is a devastating and a progressive degenerative disease of the central nervous system that dramatically affects both the patient and their care-givers. Over two thirds of all cases of dementia (memory decay, diminished reasoning and personality changes affecting all areas of daily living) are associated with AD. It is estimated that by the age of 85, between a quarter and one third of all individuals will develop AD.
How does Alzheimer’s disease develop?
Amyloid-beta precursor protein (APP) is a protein molecule composed of around 700 amino acids and is located in within the membrane of nerve cells.
An enzyme called BACE1 breaks down APP into smaller subunits, one of which is called amyloid beta peptide (Aβ). This small, 42 amino acid subunit is thought to be one of the major contributors of to the development and progression of Alzheimer’s disease. Once formed, multiple Aβ join together to form large aggregates that are even more aggressive and damaging. These Aβ aggregates produce a plaque-like deposit around the nerve cells in the brain; much like a cholesterol laden plaque in the blood vessels forms and causes atherosclerosis or damage of blood vessels.The other possible contributing cause of AD are specific ‘tau’ proteins called neurofibrillary tangles (NFT) which also form around nerve cells. Proponents of the Aβ theory are jokingly referred to as “Baptists” while Advances 19those in the NFT camp are referred to as “Taoists”. Whether the main cause is Aβ or NFT, the net result is the destruction of the nerve cells. Both Aβ plaque formation and the formation of NFT are thought to stimulate localized and chronic inflammation around the affected nerve cells. Over many years, this chronic inflammation is likely to significantly exacerbate the pathogenesis of the disease. In the end, chronic inflammation combined with the generation of damaging free radical species and the destruction of synapses or “connections” between the nerve cells ultimately leads to the degeneration of the neural circuitry. The progression of Alzheimer’s disease is also accompanied by the eventual loss of brain volume or atrophy, and a loss of neurons and functional neural synapses. A theoretical progression of events involved in this process is shown in Figure 8.
The role on inflammation in Alzheimer’s may be even more nefarious than initially thought. New research shows that not only is inflammation a contributor to the progression of the disease, it may actually be one of the main causes! Two studies conducted at the Saint Louis University School of Medicine have suggested that AD occurs due to a malfunction in a transporter protein that is supposed to clear Aβ across the blood brain barrier and out of the brain. When this transporter malfunctions, the result is that Aβ accumulates in the brain, leading to AD. However, when the researchers looked further into what actually caused the transporter to malfunction, they found that the key was inflammation! When they induced inflammation in healthy mice they found that it turned off the transporter that lets Aβ exit the brain into the bloodstream. They also found that it revved up an entrance transporter that actually transported more Aβ into the brain! When the mice were given the NSAID indomethacin, the transporters went back to their normal functioning. This provides an explanation for a variety of epidemiological studies in humans that have shown that individuals using NSAID’s over a long period of time have a reduced risk of developing Alzheimer’s disease. However, as previously discussed, the chronic use of NSAID’s comes with its own risks and side-effects. The good news is that some natural products are also beginning to show considerable promise for reducing inflammation and other symptoms associated with AD. For example, one significant problem encountered in AD is that the Aβ is not cleared rapidly enough by the body’s phagocytes. These specialized white blood cells have the ability of engulf and eliminate foreign molecules, cells or other debris. A recent study in humans has shown that the combination of curcumin and vitamin D was able to enhance the clearance of the Aβ by phagocytes. An interesting aspect of the study is that the two nutrients were synergetic, meaning that the effect together was greater than the sum of their individual effects on Aβ clearance (Masoumi et al. 2009). The role and actions of curcumin and other natural anti-inflammatory agents are discussed in detail in the next article in this magazine.