Resistant Starch And Colorectal Cancer Biology Essay

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Starch is a polysaccharide made up of glucose monomers joined together with α-1,4 glycosidic bonds to form a linear polymer, known as amylose; or with α-1,4 and α-1,6 glycosidic bonds to form a larger branched polymer, known as amylopectin. Starch is formed in the leaves of all green photosynthesising plants and is an important carbohydrate in the human diet, providing the major source of dietary calories and found in many staple foods such as rice, potatoes, wheat and maize.**

There are two crystalline forms of starch, A and B, which contain different relative proportions of amylose and amylopectin. Type A starches are 23-29 glucose units in chain length and are found in cereals. Type B have chain lengths 30- 44 glucose units and are found in tubers and amylose-rich starches. A third type, C, found in legumes, appears to be a mixture of A and B forms, which resists digestion as do type B (Topping and Clifton, 2001).

Starch is hydrolyzed by the enzyme α-amylasev which is found, in saliva, in the mouth; and in the duodenum, after being released by the pancreas. Firstly α-amylases release maltodextrins that are then hydrolysed to free glucose, by membrane-bound maltases, which is absorbed (Annison and Topping, 1994). Until research by Englyst et al (1982), showing that starches could be fermented in the large intestine, it was believed that starch was fully digested in the small intestines. This starch called Resistant Starch that by definition is "the sum of starch and products of starch digestion not absorbed in the small intestine of healthy individuals" (Asp, 1996). It can therefore be thought of as a dietary fibre.

Dietary fibre, as a term to describe all indigestible plant polysaccharides, was first used by Trowell et al. (1976). Although the definition has been refined over the decades it was generally accepted that plant materials including; non starch polysaccharides, oligosaccharides and lignin, belonged to either soluble or insoluble dietary fibres. However, Mcleary (2003) provided a definition that included Resistant Starch, which is both soluble and insoluble and can therefore be thought of as a third type of dietary fibre (Phillips et al. 2008).

Resistant starch is not found in large amounts in nature but is produced in the commercial manufacture or domestic preparation of household foods, in particular by the heating and subsequent cooling of starchy foods (Jenkins et al., 2000). In Western diets, the intakes of RS have been recorded as 5 to 10g per day (Brighenti et al., 1998).

There are four types of Resistant Starch (RS) that are classed on how they are resistant to enzyme digestion and structural consideration, RS1- RS4. RS1 is the class of starch that, due to the presence of intact cell walls, is physically inaccessible to digestion. RS2 forms have native starch granules that resist digestion by the composition of ungelatinised granules; this includes high-amylose maize starch (HAMS), which retains its structure during the processing of foods. RS3 is generally formed by the retrogradation of starch granules. RS4 are those starches that have had their digestibility decreased through chemical modification (Lorraine, 2002; Nugent, 2005).

Table 1: Classification of types of resistant starch, food sources, and factors affecting their resistance to digestion in the colon

From Nugent (2005)

Even though all starchy foods naturally contain RS, each different category has diverse physiological effects due to the varying structural compositions and the ways of processing and cooking they underwent. The digestibility of RS also varies within individuals and therefore these several factors show why degradation by bacterial fermentation is not uniform, for example 96% of RS2 in green banana was digested but only 89% of RS2 from raw potato was digested (Cummings et al., 1996).

Starch digestion and/or absorption varies because α-amylase activity can be directly affected by factors intrinsic to starchy foods including NSP, α-amylase inhibitors and the formation of amylose-lipid complexes (Englyst et al., 1992); as well as extrinsic factors such as additives (e.g. phosphorous) (Niba, 2003). Smaller food particles have a larger surface area to volume ratio and are therefore digested more rapidly than larger ones (Annison and Topping, 1994). Encapsulation of the starch granule results in inefficient digestion due to cell wall components and therefore disruption of the cell wall is necessary for unhindered starch digestion (Tovar et al., 1990).

Colonic microflora metabolise the remaining, undigested carbohydrates (i.e. RS and NSP) in the large intestine, providing the majority of enzymatic substrate for saccharolytic colonic bacteria, which use it for characteristic anaerobic fermentation producing short chain fatty acids (SCFA), including acetate, propionate and butyrate (Bauer-Marinovic et al., 2006). RS is therefore digested by bacterial amylases and the glucose produced is then further metabolised, via the formation of pyruvate, into SCFA and gases, including CO2, H2 and CH4. Absorption of SCFA in the large intestine results in the eventual digestion and absorption of RS energy in the colon (Sharma et al., 2008). There are a variety of reactions and metabolic process involved in fermentation, in which anaerobic microbial breakdown of organic matter yields metabolisable energy. This is used for microbial growth and maintenance and also other metabolic end products used by the host (Macfarlane and Gibson, 1995).

There has been much hope of the health benefits of RS (Jenkins et al., 2000) and its human consumption has been shown to improve colonic health by having a mild laxative effect, softening stools, decreasing faecal pH, increasing luminal SCFA concentrations, increasing faecal bulk and reducing the accumulation of harmful by-products of protein fermentation (i.e. ammonia (NH3) and phenols) (Birkett el al., 1996; Young and Le Leu, 2004). The increasing of digesta mass results in the dilution of carcinogens; while the decrease in digesta pH inhibit the formation and absorption of carcinogens (e.g. secondary bile acids). RS also has other broader protective mechanisms, associated with dietary fibre, such as alterations of gut microbiota to a more beneficial state and decreasing transit time, which contribute to protective effects against colorectal cancer (McGarr et al., 2005). There have also been animal studies that have shown that RS prevented colonic DNA damage in rats fed high protein diets (Toden et al., 2006).

The three principle SCFA produced as a result of colonic fermentation of RS; acetate, propionate and butyrate, have important roles in the maintaining a healthy bowel, which includes increasing colonic blood flow, maintaining a low colonic pH and improving mineral and water absorption (Topping and Clifton, 2001).

The cecum and the ascending colon are the regions in which there is the greatest fermentation rate of RS, mirroring the supply of carbohydrate in the diet (Topping and Clifton, 2001). SCFA levels fall during passage through the colon, due to their uptake and utilisation by colonocytes and bacteria (Nugent, 2005), with higher levels in the cecum and proximal colon and lower levels in the sigmoid colon, which correlate with the pattern of pH during fermentation (Nardgaard, 1998). Within the large intestine there are variations in the pH levels during fermentation, with a pH range of 5.4 - 5.9 in the cecum and the ascending colon, which increases to pH 6.2 in the transverse colon and finally ranging from pH 6.6 - 6.9 in the descending colon (Nardgaard, 1998). This pH trend is due to the decreasing rate of SCFA production and the uptake and utilisation of SCFA by colonocytes.

RS increases the production of total SCFA and also the individual concentrations of acetate, propionate and butyrate (Ferguson et al., 2000). The various types of RS also show discrepancies in the increase in production of three SCFA, as shown by Cummings et al. (1996) in which RS2 is reported to have increased the concentration of butyrate in humans, whilst RS3 did not. Even within the same type of RS there have been marked differences; potato starch, unlike HAMS, was found to enhance the proportion of butyrate, with both sources of substrate classified as RS2 (Ferguson et al., 2000).

Various data shows that colonic SCFA production is in the order of acetate > propionate ≥ butyrate, with a respective molar ratio of approximately 60:20:20 (Cummings, 1981). However various polysaccharides produce different SCFA, for example starch fermentation primarily yield acetate, propionate and butyrate, whilst pectin and xylan fermentation yields acetate only as the main product (Englyst et al., 1987).

It has also been shown, both in vitro and in animal studies, that RS produces more butyrate than NSP (Noakes et al., 1996, Ferguson et al., 2000). However, not all published studies on the effects of RS on human have shown the same effect on butyrate production (Hylla et al., 1998). This could be due to the type and quantity of RS used and the background dietary fibre, which may affect the location of the fermentation of RS.

A study in which 53 Australians consumed a mean total starch intake of 131g (within a typical Western diet), of which 5g was RS, determined that those with the highest RS intakes had the highest fecal butyrate concentration and therefore potentially reduced risk of colorectal cancer (Birkett et al. 1997).

There have been acylated starches designed that provide rapid and sustained delivery of SCFA to the colon, which in animal studies have shown to be twice as effective as HAMS in raising colonic SCFA levels of the acid that has been esterified; butyrylated SCFA result in a greater increase in butyrate than HAMS (Clarke et al. 2008).

Out of the three main SCFA, butyrate is the one that is the most interesting due to its protective effects against colorectal cancer (Whitehead et al., 1986). It is the primary energy source for colonic epithelium and is preferred to glutamine and glucose as respiratory fuels for the colonocyte (Roediger, 1982). The production of butyrate by fermentation is also associated with reduced rate of aberrant crypt foci (ACF) formation (Coleman et al., 2002) and tumour mass, provided that there is fermentation within the distal colon (Le Leu et al., 2007)

One of the mechanisms in which butyrate may be thought of as being protective against colorectal cancer is by inhibiting histone deacetylase and therefore enhancing the apoptosis of genetically damaged cells, through a histone hyperacetylation- mediated pathway (Hinnebusch et al., 2002).At physiological concentration, it has also been shown to inhibit proliferation and also induce differentiation of colon cancer cells in vitro, which may, in part, explain the correlation between a high fibre diet and the low incidence of colorectal cancer (Young et al., 2005).

Bajka et al. (2008) found that the in their animal model, colonic exposure to SCFA resulted in a strong negative correlation with DNA damage and therefore higher colonocyte apoptosis with HAMS, which would elevate levels of butyrate. Conversely, in a study conducted by Le Leu et al. (2002), there was no change in the acute apoptotic response to a genotoxic carcinogen between a RS (HAMS) and RS- free diet, suggesting that HAMS did not offer any protection against colorectal cancer via the regulation of acute apoptotic response. However, as was stated, only a modest concentration of RS was used, and that to see a positive result would require higher concentrations of RS.

Apoptosis is an important regulatory process that protects against the development of tumours and is suggested to be a better predictor of cancer outcome than cell proliferation in carcinogen- induced models (Bedi et al., 1995).

It has been shown that a lower, and hence acidic, pH is linked with protection from colorectal cancer (Walker et al., 1986). A lower luminal pH can influence many beneficial processes, including preventing the conversion of primary bile acids into secondary bile acids and of the ionisation of NH3, the balance of bacterial species (Macfarlane and Cummings, 1991). There is also a decrease in the activity of bacterial enzymes such as b- glucuronidase, which causes the deconjugation and reactivation of potential mutagens (Muir et al. 1998).

The decrease in colonic pH is caused by the accumulation of SCFA and this decreases the solubility of free bile acids, which may decrease the potential tumour promoter activity that secondary bile acids can cause (Grubben et al., 2001). An increased colonic acidification, of which is below a pH 6.5, inhibits colonic bacterial enzyme 7 α-dehydroxylase, which converts primary bile acids into secondary bile acids (Thornton, 1981). 7 α-dehydroxylase converts primary into secondary bile acids deoxycholic and iithochoic acids (Van Munster et al. 1994). In the presence of deoxycholic acid there is an increased production of phosphatidylcholine diacylglycerol (DAG) in the human femermentation systems (Morotomi et al. 1990). DAG increases the affinity of protein kinase C (PKC) for calcium, rendering it active at physiological levels and phosphorylating various target organs. Phorbol esters, which mimic DAG (in that they also activate PKC) are known to be tumour promoters but are not degraded (Bingham, 1997). Furthermore, decreasing colonic pH increases calcium availability for it to bind to free bile acids and fatty acids (Wargovich et al., 1984).

It has been noted that secondary bile acids show involvement, as a promoting agent, in the adenoma-carcinoma sequence of colorectal cancer. Christl et al. (1997) conducted a study, carried out in vitro, that found that at pH 6, there was significantly more primary bile acids that remained with a lesser production of secondary bile acids, and that at pH 7 total bile acid concentration was lower. This indicated that there is an inhibition of bacterial breakdown of primary to secondary bile acids, when starch is simultaneously fermented. Showing one of the mechanisms in which resistant starch has the potential protective factor against colorectal disease, by the fact that the reduction of pH is explained by the SCFA production.

Secondary bile acids have a cytotoxic effect on colonic mucosa that leads to a compensatory increase in their proliferation. The hyper- proliferative mucosa has an enhanced sensitivity to mutagenic substances and is associated with an increased risk of colorectal cancer (Van Munster and Nagengast, 1993).

In humans consuming a low RS diet, the colon has a neutral pH 7.2 ± 0.2. With an acidic environment there is healthy bacterial proliferation and inhibition of pathogenic bacteria (Le Leu et al., 2002).

In a low pH, alkaline toxic compounds degrade, which inhibits their absorption into the body (Bird et al., 2000).

A common effect of the major SCFA is their ability to stimulate and enhance colonic muscular contraction, which raises muscular tone and large bowel oxygenation and nutrient transport (Bird et al., 2000)

There are further ways in which butyrate could be administered to achieve protective health benefits against colorectal cancer. In animal models, butyrate was given as gastroresistant, slow release pellets and was recorded to increase apoptosis in the colonic epithelium, providing additional evidence of its beneficial effects (Caderni et al., 1998). It has also been suggested that butyrate enemas, in long- term therapy, are useful to reduce the risk of colorectal cancer in patients with ulcerative colitis (D'Argenio et al., 1996). A study by Bonnotte et al. (1998), showed that butyrate enhances the sensitivity of colon carcinoma cell lines to Fas- mediated apoptosis, which gives implications that it could be used as an adjuvant with chemotherapy and immunotherapy in colorectal cancer.