Alcohol Intake (0.5 Ml And 1.0 Ml) Once Per Day And The Functional Status Of Hepatic Tissues Of Wistar Rats

ABSTRACT

This study was aimed at the evaluation of the effect of alcohol intake (0.5 ml and 1.0 ml once per day for 21 days) and the functional status of the hepatic tissues of wistar rats. The branded alcohol (Seaman’s Schnapps 40% alcohol by volume) was obtained from a grocery store located at Relief market Owerri, Imo State, Nigeria. The alcohol was prepared by further diluting it with equal volume of water. This preparation was repeated every day for the period of three (3) weeks of administration of alcohol. Twelve (12) male albino rats were divided into three (3) groups of four (4) rats each. The rat groups were grouped according to the treatments received by oral gavage on daily basis for 21 days. Group one received 0.5 ml/kg body weight of alcohol once daily, Group two received 1.0 ml/kg body weight of alcohol once daily, Group three received feed and water only and served as the normal control group. At the end of the treatment period of 21 days, blood volumes of 3.0 mL were drawn from orbital sinus of the rat groups after being deprived of feed and water for 12 h. The blood samples were allowed to clot, after which serum from corresponding rat groups were measured for hepatic tissue biomarkers, namely, AST, ALT, ALP activities and bilirubin concentrations. The AST and ALP levels after the administration of 0.5 ml and 1.0 ml of alcohol once per day were higher and significantly different from the control group, while there was no significant difference between ALT, total bilirubin and conjugated bilirubin levels and the control. This is an indication that the intake of alcohol even at low doses has damaging effects on the liver.

CHAPTER ONE

1.0   INTRODUCTION AND LITERATURE REVIEW

1.1 INTRODUCTION

Alcohol are organic compounds identified by one or more hydroxyl (−OH) groups attached to a carbon atom of an alkyl group (Leroy, 2018). Alcohol is often considered as organic derivatives of water in which one of the hydrogen atoms has been replaced by an alkyl group, typically represented by R in organic structures. There are different types of alcohol: propyl, methyl, ethyl, and butyl alcohol, but for the production of alcoholic beverages, ethyl alcohol (ethanol) is the type used, while death or blindness could result if the other three are consumed at all (Leroy, 2018). Ethanol is normally produced from the fermentation of yeast, sugar, and starches (Tarek, 2009). It can also be consumed mixed in beverages or in its raw form, but the end product after digestion by the body is formaldehyde, which is deleterious to the body and also the cause of alcohol poisoning. Liver cirrhosis, gastric ulcers, gastritis, fatty liver, alcoholic hepatitis, and pancreatitis are examples of some disorders resulting from alcohol intoxication. Moderate consumption of alcohol may have some health benefits that could prolong life, but when taken in excess or as an escape route from problems, then abuse or alcoholism is inevitable. Abuse of alcohol usually leads to alcohol use disorder also known as alcoholism or alcohol addiction which can lead to a violent behavior. Addiction to alcohol refersto physical and psychological dependency on alcohol to the point that tolerance is built up to alcohol (Rarek, 2009). Furthermore, once the body becomes adapted to excessive consumption of alcohol, an abrupt discontinuation leads to withdrawal symptoms which can be a threat to life and the symptoms normally includes hallucination, tremors, convulsions, anxiety, etc. Alcohol addiction may also result in the following: restlessness, depression, erratic behavior, decreased involvement in extracurricular activities, loss of interest in work or school, lack of interest in relationships, preoccupation with drinking, inability to control drinking, and violent behavior (Vaillant and Keller, 2018).

The level of the blood alcohol is increased when the quantity of alcohol consumed exceeds the normal dose which the body could metabolize, and then intoxication sets it. The higher the blood alcohol concentration, the higher the risk of diseases associated with the liver, kidney, and pancreas and the abundance of free radicals in the body system.

In this present research, I investigated the effect of daily alcohol intake on the functional status of hepatic tissues of albino rats.

1.2 Literature Review

1.2.1. Blood

Blood is a specialized body fluid. The blood that runs through the veins, arteries, and capillaries is known as whole blood, a mixture of about 55 percent plasma and 45 percent blood cells. About 7 to 8 percent of your total body weight is blood. An average-sized man has about 12 pints of blood in his body, and an average-sized woman has about nine pints (Atterbury et al., 2007).

1.2.1.1.  Components of Blood

Many people have undergone blood tests or donated blood, but hematology – the study of blood – encompasses much more than this. Doctors who specialize in hematology (hematologists) are leading the many advances being made in the treatment and prevention of blood diseases (Migliacci, 2013).

The liquid component of blood is called plasma, a mixture of water, sugar, fat, protein, and salts. The main job of the plasma is to transport blood cells throughout the body along with nutrients, waste products, antibodies, clotting proteins, chemical messengers such as hormones, and proteins that help maintain the body’s fluid balance (Heuft et al., 2008). 

Known for their bright red color, red cells are the most abundant cell in the blood, accounting for about 40 to 45 percent of its volume. The shape of a red blood cell is a biconcave disk with a flattened center – in other words, both faces of the disc have shallow bowl-like indentations (a red blood cell looks like a donut) (Matthes et al., 2014).

Production of red blood cells is controlled by erythropoietin, a hormone produced primarily by the kidneys. Red blood cells start as immature cells in the bone marrow and after approximately seven days of maturation are released into the bloodstream. Unlike many other cells, red blood cells have no nucleus and can easily change shape, helping them fit through the various blood vessels in your body. However, while the lack of a nucleus makes a red blood cell more flexible, it also limits the life of the cell as it travels through the smallest blood vessels, damaging the cell’s membranes and depleting its energy supplies. The red blood cell survives on average only 120 days (Burnouf et al., 2009).

Red cells contain a special protein called hemoglobin, which helps carry oxygen from the lungs to the rest of the body and then returns carbon dioxide from the body to the lungs so it can be exhaled. Blood appears red because of the large number of red blood cells, which get their color from the hemoglobin. The percentage of whole blood volume that is made up of red blood cells is called the hematocrit and is a common measure of red blood cell levels (Tsantes et al., 2014).

White blood cells protect the body from infection. They are much fewer in number than red blood cells, accounting for about 1 percent of your blood (Treleaven et al., 2011).

The most common type of white blood cell is the neutrophil, which is the “immediate response” cell and accounts for 55 to 70 percent of the total white blood cell count. Each neutrophil lives less than a day, so your bone marrow must constantly make new neutrophils to maintain protection against infection. Transfusion of neutrophils is generally not effective since they do not remain in the body for very long (Hardwick, 2008).

The other major type of white blood cell is a lymphocyte. There are two main populations of these cells. T lymphocytes help regulate the function of other immune cells and directly attack various infected cells and tumors. B lymphocytes make antibodies, which are proteins that specifically target bacteria, viruses, and other foreign materials (Tsantes et al., 2014).

Unlike red and white blood cells, platelets are not actually cells but rather small fragments of cells. Platelets help the blood clotting process (or coagulation) by gathering at the site of an injury, sticking to the lining of the injured blood vessel, and forming a platform on which blood coagulation can occur. This results in the formation of a fibrin clot, which covers the wound and prevents blood from leaking out. Fibrin also forms the initial scaffolding upon which new tissue forms, thus promoting healing (Migliacci, 2013).

A higher than normal number of platelets can cause unnecessary clotting, which can lead to strokes and heart attacks; however, thanks to advances made in antiplatelet therapies, there are treatments available to help prevent these potentially fatal events. Conversely, lower than normal counts can lead to extensive bleeding (Matthes et al., 2014).

1.2.1. 2    Serum

Serum, the portion of plasma remaining after coagulation of blood, during which process the plasma protein fibrinogen is converted to fibrin and remains behind in the clot. Antiserum, which is prepared from the blood of animals or humans that have been exposed to a disease and have developed specific antibodies, is used to protect persons against disease to which they have been exposed (Heuft et al., 2008).

Serum is the fluid and solute component of blood which does not play a role in clotting. It may be defined as blood plasma without the clotting factors, or as blood with all cells and clotting factors removed. Serum includes all proteins not used in blood clotting; all electrolytes, antibodies, antigens, hormones; and any exogenous substances (e.g., drugs or microorganisms). Serum does not contain white blood cells (leukocytes), red blood cells (erythrocytes), platelets, or clotting factors (Atterbury et al., 2007).

The study of serum is serology. Serum is used in numerous diagnostic tests as well as blood typing. Measuring the concentration of various molecules can be useful for many applications, such as determining the therapeutic index of a drug candidate in a clinical trial (Treleaven et al., 2011).

To obtain serum, a blood sample is allowed to clot (coagulation). The sample is then centrifuged to remove the clot and blood cells, and the resulting liquid supernatant is serum (Migliacci, 2013).

 1.2.2.  Alcohol and Alcoholic Beverages

There are different types of alcohol based on their uses; some are used as solvents, antifreeze solutions, e.g., isopropyl and methyl alcohol, paint removers in chemistry laboratories and chemical factories, and as nail polish remover and cleansing solvent, for example, methylated spirit for domestic use (Nordmann et al., 2002). There are different types of alcohol depending on the number of carbon atoms present and the position of the OH bond in the formula, but the most common alcohol is ethanol (CH₃CH₂OH or C₂H₅OH). During the process called fermentation, alcohol is produced. Fermentation of yeast usually results into sugar breaking down into carbon dioxide and alcohol. But carbon dioxide is taking off the process through gas bubbles leaving a mixture of water and ethanol. Sugar fermentation to alcohol can be used in different applications especially in the production of alcoholic beverages, for example, the extracts of grapes and barley are fermented in the wine and beer industry to produce alcoholic beverages. Any drink or beverage that contains ethanol is normally referred to as alcoholic beverage and are either produced by fermentation or distillation (Naimiet al., 2003). There are different types of alcoholic and nonalcoholic beverages. Alcoholic drinks can be classified into five categories: (i) Wine, i.e., still, sparkling, fortified, or aromatized; (ii) beer, i.e., ales, lagers, and stouts; (iii) cider and perry; (iv) distilled spirits, i.e., vodka, gin, rum, whiskey, brandy, and others; and (v) liqueurs, i.e., flavored with fruit, citrus, herb, kernel, flower, cream, and berry (Buckman et al., 2015).

Frequent alcohol consumption can be deleterious to human’s health by affecting the following: (i) Brain: the communicative pathways of the brain are disrupted which may result in mood swing, lack of coordination, and coherence. (ii) Heart: it may also affect the heart by causing cardiomyopathy, arrhythmias, stroke, and high blood pressure. (iii) Liver: different types of problems may occur in the liver including liver inflammations, cirrhosis, alcoholic hepatitis, fibrosis and steatosis, or fatty liver (Naimiet al., 2003). (iv) Pancreas: heavy consumption of alcohol may cause the blood vessels in the pancreas to be swollen and inflamed thereby preventing proper digestion (Buchman et al., 2015). (v) Skin: abuse and consumption of alcohol can cause a variety of skin disorders (Smith and Fenske, 2000). Thermoregulation of the body also results in skin vasodilation and sweating (Yoda et al., 2005). (vi) Cancer: heavy drinkers stand the chance of developing cancers like cancer of the mouth, esophagus, throat, liver, and breast (Horowitz et al., 2009). Alcohol meddles with folate assimilation and function in the body, which may be one way alcohol can build danger of causing specific cancers. (vii) Immune system: alcoholism results in immune system being compromised, and the body is easily prone to diseases like pneumonia and tuberculosis (Kelvin et al., 2014).

1.2.3  Alcoholism

Alcoholism is alcohol abuse and a prevalent problem nowadays. A person is considered addicted if he/she continues to drink alcohol without interruption with the knowledge that this person knows about the dangers of alcohol addiction. Alcohols are of the worst psychotropic substance (Barclay and Stewart, 2008). Also, alcoholism leads to serious social, economic, and health problems. Thus, excessive alcohol consumption has consequences that may sometimes result in death. Alcoholism is a chronic disease that is often fatal. Alcohol inhibits people’s ability to think properly, and most addicts deny the existence of the problem (Smith and Robinson, 2013). However, alcoholism is considered one of the most widespread problems in the world. Alcohol abuse is a global problem causing the lack of development of the individual and society alike. There are many factors that cause alcohol addiction and genetics is one of the most important causes of alcohol addiction. studies conducted on the families of alcoholics found that the children of alcoholics have a greater predisposition to alcohol addiction, and children born into families of addicts are at risk of addiction.

1.2.4  History of the use of alcohol

The origin of alcoholic beverages is lost in the mists of prehistory. Fermentation can occur in any mashed sugar-containing food (such as grapes, grains, berries, or honey) left exposed in warm air. Yeasts from the air act on the sugar, converting it to alcohol and carbon dioxide. Alcoholic beverages were thus probably discovered accidentally by preagricultural cultures (Lucia, 2003). Early peoples presumably liked the effects, if not the taste, and proceeded to purposeful production. From merely gathering the wild-growing raw materials, they went on to regular cultivation of the vine and other suitable crops. Few preliterate groups did not learn to convert some of the fruit of the earth into alcohol. In the case of starchy vegetation, quite primitive agriculturists learned how to convert the starch to fermentable sugar through preliminary chewing (Saliva contains an enzyme that carries out this conversion) (Babor, 2006).

Alcohol is the oldest and still one of the most widely used drugs. The making of wines and beers has been reported from several hundred preliterate societies. The importance of these alcoholic beverages is evident in the multiplicity of customs and regulations that developed around their production and uses (Marciniak et al., 2002). They often became central in the most valued personal and social ceremonies, especially rites of passage, and were ubiquitous in such activities as births, initiations, marriages, compacts, feasts, conclaves, crownings, magic rites, medicine, worship, hospitality, war making, peace making, and funerals (Lucia, 2003).

The manufacture and sale of alcoholic beverages was already common in the earliest civilizations, and it was commercialized and regulated by government (Rouche, 2003). The oldest known code of laws, the Code of Hammurabi of Babylonia (c. 1770 BCE), regulated drinking houses. Sumerian physician-pharmacists prescribed beer (c. 2100 BCE) in relatively sophisticated pharmacopoeias found on clay tablets. Later Egyptian doctors, in their medical papyri (c. 1500 BCE) (Babor, 2006), included beer or wine in many of their prescriptions. Semitic cuneiform literature of the northern Canaanites, in prebiblical Ugarit, contains abundant references to the ubiquitous religious and household uses of alcohol.

Water, a precious commodity in the earliest agriculturally dependent civilizations, was probably the original fluid used as offering in worship rites. In time, other fluids (milk, honey, and later wine) were substituted. That alcoholic beverages should have displaced other fluids in early religions, both as offering and drink, is not surprising. The capacity of alcohol to help the shaman or priest and other participants reach a desired state of ecstasy or frenzy could not long have escaped observation, and its powers were naturally attributed to supernatural spirits and gods (Babor, 2006). The red wine in religious uses was eventually perceived as symbolizing the blood of life and, in this spiritual sense, ultimately passed into the Christian Eucharist. The records of the ancient Egyptian as well as of the Mesopotamian civilizations attest that drinking and drunkenness had passed from the state of religious rite to common practice, often troublesome to government and accompanied by acute and chronic illnesses (Raymond, 2007). There are ample indications that some people so loved drink and were so abandoned to drunkenness that they must be presumed to have been alcoholics (Samuelson, 2008).

1.2.5  Alcohol Consumption Worldwide

The worldwide per capita consumption of distilled spirits, beer, and wine has generally increased since 1950, with beer consumption increasing more than the consumption of either spirits or wine (Kuhn et al., 2008). Although the consumption of all alcoholic beverages generally rose during the second half of the 20th century, deviations from these general tendencies were evident in many countries. Countries with traditionally high levels of total alcohol consumption (e.g., France, Italy, Portugal, Spain, and Switzerland) exhibited stable or only slightly increased per capita consumption (Vonghiaet al., 2008). Beer consumption in traditionally heavy beer-drinking countries (e.g., Germany, Belgium, Australia, New Zealand, Ireland, the United Kingdom, and Denmark) did not increase as much as did the worldwide pattern. Wine consumption on a per capita basis actually declined in some countries with traditionally high levels of consumption (e.g., France and Italy), while it rose in some countries with relatively low wine consumption, especially the United States and Australia. In the 1990s alcohol consumption declined in most of the developed countries but increased in many developing countries (Roehrs and Ruth, 2001).

Many factors are believed to have affected these patterns in total alcohol consumption. One factor is the increased use of alcohol (primarily beer and wine) with meals, in part a result of long-term increases in per capita income in many countries and in part a result of fermentation technology that has kept the price of some alcoholic beverages relatively low, facilitating the purchase of beer and wine. Another factor is improved marketing and advertising (Zakhari, 2006). For example, in North America, particularly in the United States, the introduction of low-calorie beer and wine in the early 1970s was instrumental in the increased per capita consumption of alcohol in the late 20th century.

The countries leading in total alcohol consumption, based on data from 2016, werethe Czech Republic, Australia, Portugal, Slovakia, and Hungary. There were significant disparities in the level of consumption across countries among different types of alcoholic beverages. For example, although most of the leading consumers of alcoholic beverages drank significant quantities of wine, many drank relatively low quantities of distilled spirits (Lohr, 2005). The leading beer-drinking countries were the Czech Republic, Namibia, Austria, Germany, and Poland. South Korea, Seychelles, Russia, Estonia, and Lithuania were among the leading consumers of distilled spirits. Rates of heavy alcohol consumption were highest in developed and high-income countries (Rehm et al., 2010).

1.2.6  Alcohol Metabolism

The by-product of fermentation and/or distillation of natural juices, fruit and vegetable mashes is pure ethyl alcohol, a volatile liquid, which is subsequently modified by either adding additional alcohol or through distillation, vaporization and even condensation. Such beverages as wine and beer are obtained from fermentation and contain five percent and 11 percent alcohol, respectively (Day et al., 2009). Fortified wines, for example, sherry and port, contain both wine and pure alcohol. Vodka, gin, rum and whisky are distilled products and have a much higher alcohol content (Day et al., 2009).

Once ingested orally, alcohol is rapidly absorbed from the stomach, small intestine and colon, and subsequently evenly distributed. It is metabolized by the

enzyme, alcohol dehydrogenase, which is located in the gastric mucosa and the liver. Ninety-five percent of alcohol is metabolized to carbon dioxide and water, and the remainder is partly excreted by the kidney and partly expelled in the air. In an hour, the body can metabolize approximately 30 mls of alcohol which is equivalent to approximately one standard drink (Mills et al., 2004). The rate of metabolism of warfarin, diazepam and propranolol are increased after long-term alcohol use. Cimetidine and ranitidine inhibit gastric alcohol dehydrogenase leading to increased blood alcohol levels following a given exposure (Taylor, 2003).

1.2.7  Excretion of Alcohol

Most of the ethanol a person drinks (95–98%) is removed from the body by oxidative metabolism and less than 10% is eliminated unchanged by excretion via the lungs, the kidneys, and skin. After moderate drinking, 2–5% of the dose of ethanol can be recovered by analysis of urine, exhaled air, and sweat. These body fluids are often analyzed in forensic toxicology as proof a person has consumed alcohol in situations when, for some reason, they are required to remain abstinent (Marques and McKnight, 2009). When subjects drank 0.54, 0.68, or 0.85 g/kg as neat whisky, urine samples collected during 7 hr contained

0.70, 0.80, and 1.55% of the dose administered (Jones, 2000). In these experiments, the maximum BAC reached was 120 mg %, so these percentages of the dose excreted are expected to be higher on reaching higher BAC as expected for first-order urinary excretion of ethanol.

1.2.8   Nonoxidative metabolites

A small fraction (<0.2%) of the dose of ethanol a person drinks undergoes conjugation reactions in the liver to produce ethyl glucuronide (EtG) and ethyl sulfate (EtS), which are the major nonoxidative metabolites (Palmer, 2009). The pharmacokinetic profiles of EtG and ethanol in blood are different in several respects (Halter et al., 2008). For example, the EtG curve in blood rises more slowly than the BAC and reaches a peak concentration in blood 1–2 hr later than the ethanol peak. The EtG and ethanol curves are shifted in time and the EtG concentration in blood is about 1,000 times lower than the concentration of ethanol. EtG is also eliminated from the body more slowly and is detectable in blood and urine for several hours longer than ethanol, which means that analysis of EtG is useful to disclose recent drinking in different clinical and forensic situations (Hoisethet al., 2007).

After gas chromatography-mass spectrometry methods were introduced for analysis of EtG and EtS in blood, urine, and hair strands, a number of forensic applications have emerged (Schmitt et al., 2005). These metabolites are used as biomarkers for recent drinking and to control abstinence in people expected to refrain from drinking, because of their employment (safety-sensitive work) or participation in treatment programs for alcohol abuse (Walsham and Sherwood, 2012).

In postmortem (PM) toxicology, the analysis of EtG in blood and urine can help in differentiation between ethanol that might be produced after death from antemortem ingestion (Hoisethet al., 2007). In this connection, there appears to be some advantages of analyzingEtS metabolite as a biomarker of recent drinking and whether postmortem synthesis of ethanol has occurred (Helander and Beck, 2005; Krabsethet al., 2014). Another method to test the origin of PM ethanol is

to determine the ratio of two metabolites of serotonin, the 5-HTOL/5-HIAA ratio as described elsewhere (Helanderet al., 2005).

Other nonoxidative metabolites of ethanol include various fatty acid ethyl esters (FAEE) and phosphatidylethanol (PEth), both of which have found applications in clinical medicine as biomarkers of heavy drinking (Staufer and Yegles, 2016)

1.2.9   Socioeconomic Impact of Alcohol Consumption

Statistic in the Great Britain comparing alcoholic insanity with other kinds of insanities shows that 2249 alcoholic patients with insanity were detected against only 53 with other reasons than alcohol. In US 85 percent of psychological patients in bedlam were alcoholic patients. If statistic gathers about the number of drunken who committed a crime or destroyed houses and lives, it would be actually high. In France 440 people die because of alcohol daily (4). From the perspective of statistic which the philosopher Huger distributed in twentieth annual of Science journal, 60 percents of intentional murdering, 75 percent of maims, 30 percent of immoral crimes (like committing adultery with incest), 20 percent of thievery were related to alcohol and alcoholic drinks and in otherstatistic from the same professor 40 percent of convict children had some history of alcohol consumption (4). In an article entitled “is alcohol more dangerous than heroin?” it came that; today alcohol is considered as emergency, because its detrimental effects are very wide. These effects consist of rape, injuries following falling, mental disorders and lack of true upbringing of the children. Wide effects of these encompass all the societies. Today alcoholic drinks need to be less available and advertised (20). Room indicated that solving the problem of alcoholic drinks should be the priority in all part of the world (21).

1.2.10  Alcohol and Health

1.2.10.1.  Mental Damages in Alcohol Consumption

Alcohol alone disturbs inhibitory mechanisms and activator of behavior control (Sher, 2006). Memory dysfunction: it is seen in individuals who consume high amounts continuously or high amounts in one time. Depression: depression can cause alcohol consumption and reversely. Anxiety: it is common in giving up period of alcohol. Some of the patients with anxiety consume alcohol again, alcohol consumption and anxiety has a two way relationship. Suicide: behaviors for suicide intention or decision to hurt themselves is more frequent in alcohol consumer compared to others. 6 to 8 percents of high amounts consumers suicide at last. Personality disorders: personality changes in the form of egotism, not paying attention to others, no to obey behavioral and moral rules, dishonesty and lack of accountability and to be jealous which is a rare complication but serious following consumption of high amounts of alcohol are seen. Suspicion to wife is a lot seen among them (Brust, 2005).

1.2.10.2.   Effects on Central and Peripheral Nervous Systems

Researches believe that individuals who consume alcohol become shy and talkative, it is not due to stimulatory effects of alcohol on brain, whereas it is owing to loss of control of high centers on moderations (Brust, 2006). The most important effect of alcohol is the weakness of central nervous system. Following its effects on nervous system it causes reactions in vessels which drain blood to the surface of the body and cause skin flushing, at first causes warmth in the body but after a short while body loses its warmth two times more. Drunken first becomes hyperactive and its body temperature goes up as mentioned above and tachypnea occurs. Then speech centers become affected and with the weakness of controlling mechanisms talkativeness occurs. Then hearing center is affected and individual hear some sounds. After that visual center is affected and individual sees fictitious images. Finally equilibrium center is affected, physical activities balance disrupts. Therefore with drinking alcohol continence is destroyed mostly which is the root of timidity and modesty, and leads to abnormal tasks (Brust, 2006). Alcohol peripheral neuropathy (ALN) is a complication in alcoholics which causes sensory, motor and autonomic disorders. Alcohol per se is a significant predisposing factor of developing this kind of neuropathy (Su, 2005). Alcohol induced seizure is seen in individuals who consume high amounts of alcohol which is present with several attacks without any previous history of seizure (Schuckit, 2005). Several articles indicate the association between alcohol consumption and increased risk of brain attacks (Aminoff and Parent, 2008). Alcohol has antiplatelet effects which cause subarachnoid hemorrhage which can cause hemorrhagic brain attack (Sher, 2006).

1.2.10.3.  Cardiovascular Effects

Alcohol cause many changes in cardiovascular system. Long term consumption of high amounts of alcohol contributes to dilative cardiomyopathy and fibrosis and ventricular hypertrophy (Ronksleyet al., 2011). Many researches have confirmed the most harmful effect of alcohol on cardiovascular system as increasing the risk of atrial fibrillation (Kloner and Rezkalla, 2007). In a study Danish researchers followed 47949 in 6 years founded that alcohol consumption causes increased risk of atrial fibrillation in men. Atrial fibrillation is a dysrhythmia in heartbeat, which may cause death because of the lack of control on atrium beat (Ronksleyet al., 2011). In amounts higher than 30 grams per day is associated with increased risk of ischemic and hemorrhagic heart attack (Bhatt et al., 2007). Obvious relationship has been approved between long term alcohol consumption and functional and structural heart muscle dysfunction (Noth and Swislocki, 2001). Ectopic hyperactivity of ventricles is detected after alcohol consumption (Bhatt et al., 2007). Also high amount of alcohol consumption leads to increased risk of heart failure (38-40). From 80 cross sectional investigations seeking the relationship of alcohol consumption and hypertension, approximately all of them indicated that increased consumption of alcohol leads to higher blood pressure and increased risk of hypertension. Moreover higher consumption causes interactions with antihypertensive drugs (Mukamai, 2010).

1.2.10.4.  Sexual Dysfunctions

Erectile dysfunctions or delayed ejaculation are other consequences of alcohol consumption. Alcohol effects on vascular and nervous systems cause complications in relationships with sexual partner in high amount alcohol consumer (Sher, 2006). Alcohol induced sexual dysfunction causes changes in sexual hormones balance like testosterone, estrogen and prolactin in both genders and also decreases number of viable sperm production. Delayed ejaculation, decreased libido, sexual cycle disorders and delayed orgasm in women is the effects of these hormonal changes (Fingerhood, 2007). Alcohol consumption and opioids decrease sexual tendency (Molina et al., 2010).

1.2.10.5.   Effects on Liver

Alcoholic liver disease is still a reason of death. Human liver is able to oxidize 8 grams of alcohol per hour, but higher amounts appear in blood. Concentrations below 0.05 percent does not cause any symptom in human, but higher than this amount leads to some symptoms like a decrease in physical activities balance and visual disorders. Lethal dose is different among individuals (CDCP, 2010). However, alcohol comprises high energy, but it must be first changed into lipids to be applicable and then changed into energy in a long pathway. So it is believed that alcohol affect liver health and it is also called death calorie. Hepatic disease is the most frequent medical side effects of alcohol misuse. It is estimated that about 15 to 30 percents goes toward complicated hepatic diseases finally. The disease starts serenely. Alcoholic fatty liver may progress to alcoholic hepatitis and consequently cirrhosis and hepatic failure. In the United States, long term alcohol consumption is the main reason of hepatic cirrhosis and the need for liver transplant. Long term alcohol consumption has been the main reason of chronic pancreatitis in the west (Dasarathy and McCullough, 2007). There is a relationship with vitamin C and B depletion with alcohol consumption. Alcohol is a diuretic which can cause to loose water (Sher, 2006). Long term alcohol consumption leads to increased risk of hepatocellular adenoma with stimulation of cells called H-RAS-MUTANT (Molina et al., 2010). Alcohol metabolites are able to enter processes which lead to hepatic autoimmune diseases similar to ALD (Fingerhood, 2007). Alcohol consumption may also lead to disease progression and acceleration in patients with hepatitis C (Rehm et al., 2010).

1.2.10.6.   Effects on Bone

Alcohol consumption decreases not only the density and strength of bone, but also bone repair after fractures. Researches designated that alcohol consumption damages bone cells called osteoclasts, and with division of these damaged cells to osteoblasts, which are considered as the main bone cells, bone strength decreases (Realm et al., 2003). In a study performed in Copenhagen, consumption of high amounts of alcohol was found to be a main risk factor for pelvic fracture (Lee and Snape, 2008).

Effects on Pancreas Chronic pancreatitis is a progressive inflammatory disease which involves pancreatic ducts. This disease results from a combination of environmental factors like alcohol and smoking (Dawson, 2008). Alcohol induced injury can be also acute. The most common causes of acute pancreatitis are cholelithiasis and alcohol consumption, which alcohol is the main reason of chronic pancreatitis as well (Maria et al., 2001). Alcohol injury on pancreas is based on fibrosis necrosis theory initiate with an acute process. Following there would be repetitive attacks leading to atrophic changes of acinar cells and fibrosis. At last leads to complicationsin exocrine and endocrine activities of pancreas (Derk and Horatius, 2005). Ethanol metabolism in acinar cells and other pancreatic cells and production of toxic metabolites play pivotal roles in acute and chronic injury of pancreas. Pancreas metabolizes ethanol in oxidative pathway using ADH enzyme and cytochrome P4502E1 and in non oxidative pathway using FAEE enzyme. It has been shown that PSC cells contain ADH enzyme too which leads to injury initiation (Lieber, 2006). Nowadays it has been publicized apparently that activation of PSC cells has a key role in production of cancerous stroma. Production of cancerous stroma has a close relationship with cancerous cells and provides a good environment for the local growth and metastasis. PSC cells activates under the effects of ethanol and acetaldehyde (alcohol toxic product) (Maria et al., 2001). Acinar cells are able to metabolize alcohol. Direct alcohol toxicity and its metabolites on these cells leads to autodigestive processes of the pancreas tissue due to stimulant factors (Rehm et al., 2010).

1.2.10.7.  Fetal Alcoholic Syndrome

Adverse effects of alcohol on intrauterine growth comprises a wide spectrum of anomalies, behavioral and neurocognitive disorders, which the most complete word to indicate it seems to be FASD (fetal alcohol spectrum disorders). After developing diagnostic tools to diagnose more applicable and precise it broke down into several fields like PFAS, AFS and ARND (Molina et al., 2010). In a study performed on children affected with FASD, approximately all of them showed a significant clinical delay in fine and gross motor skills (Fingerhood, 2007). In another study all children with FASD diagnosis had ADHD (a kind of hyperactivity disorder) (Lee and Snape, 2008). In a large study conducted in the United States on 1400 patients with FASD, 28% had encephalopathy and 52% neurobehavioral disorders (Rehm et al., 2010). Frequent complications in patients with FASD comprises 11% preterm labor, 70% intrauterine growth retardness, 45% microcephaly, 55% digits disorders (congenital digits flexion; camptodactyly), 51% visual disorders (disorder in focusing; refractive error), 43% dental complications (Dental Crowding), 38% nails hypoplasia, 38% strabismus, 22% urogenital anomalies and 18% congenital cardiac defects (Realm, 2003). Eye complications is considered as one of the most frequent and important disorders in fetal alcoholic syndrome which has been reported in about 90% of children with fetal alcohol syndrome, including microphthalmia, loss of neurons in the retina, optic nerve hypoplasia, and dysmyelination (Maria et al., 2001). Complications which are known as fetal alcoholic syndrome include; intrauterine growth retardness, microcephaly, incomplete development of maxillofacial and joint abnormalities. In more severe forms congenital heart defects and mental retardness is occurred (Sher, 2006).

1.2.10.8.   Skin Complications

Psoriasis is a dermatologic disorder which causes swelling and exfoliation. In this disturbing disease, skin cellsgrow from the depth of skin thereafter separate promptly from the skin. Based on researches performed, an obvious relationship has been proven between environmental factors and psoriasis. From these factors smoking and alcohol have significant roles (Noth and Swislocki, 2001). In an investigation performed, 15% of patients with psoriasis reported consumption of high amounts of alcohol (Vongiaet al., 2008). Moreover, involved skin surface in affected patients was related to the amount of alcohol consumption significantly (Asarathy and McCullough, 2007).

1.2.10.9.  Alcohol and Cancer

Alcohol consumption increases the risk of cancer in mouth, pharynx, larynx, esophagus and liver (Rehm et al., 2010). Based on investigations performed, moderate alcohol consumption increases the risk of endometrial cancer in young women (Kersaw and Guidot, 2008). In other studies conducted on the effect of CYP2E1 and ALDH2 genes pleomorphism, breast cancer progression risk was 1.4 higher in alcoholic women compared to others (Kopans, 2007).

1.2.11  The Liver

Liver is an organ in the upper abdomen that aids indigestion and removes waste products and worn-out cellsfrom the blood. It is a vital organ present in vertebrate andsome other animals, which has a wide range of functionsincludingdetoxification and protein synthesis. The liveris our greatest chemical factory, it builds complexmolecules from simple substances absorbed from thedigestive tract, it neutralises toxins, it manufactures bilewhich aids fat digestion and removes toxins through thebowels (Matonet al., 1993). But the ability of the liver toperform these functions is often compromised by numerous substances we are exposed to on a daily basis;these substances include certain medicinal agents which when taken in over doses and sometimes when introduced within therapeutic ranges injures the organ (Gagliano et al., 2007). Liver disease is worldwide problem. Conventionaldrugs used in the treatment of liver diseases are sometimes inadequate and can have serious adverse effects. Therefore, it is necessary to search for alternative drugs for the treatment of liver disease in order to replace currently used drugs of doubtful efficacy and safety (Ozbeket al., 2004). In the absence of reliable liver protective drugs in allopathic medical practices, herbsplay a role in the management of various liver disorders.

Figure 1: Intrahepatic vascular and biliary anatomy (Decker;2007).

 1.2.11.1   Livermarker Enzymes

Liver function tests (LFTs or LFs), are groups of clinical biochemistry laboratory blood assays designed to give information about the state of a patient’s liver (Lee, 2009). According to some, liver transaminases (AST/ALT (SGOT/SGPT) are not liver function testsbut are biomarkers of liver injury in a patient with some degree of intact liver function. Other sources include transaminase (McClatchey, 2002).

Aspartate aminotransferase (AST)

This is present in many tissues and is useful in evaluating muscle and liver damage in small and large animals. AST is not liver specific in any domestic animal species and the reference range in horses is rather broad. Skeletal muscle is the second largest source of AST in animals.

Alanine aminotransferase is the most frequently relied biomarker of hepatotoxicity (Dufour, et al., 2000). It is a liver enzyme that plays an important role in amino acid metabolism and gluconeogenesis. It catalyzes the reductive transfer of an amino group from alanine to α-ketoglutarate to yield glutamate and pyruvate. Normal levels are in the range of 5-50 U/L. Elevated level of this enzyme is released during liver damage. The estimation of this enzyme is a more specific test for detecting liver abnormalities since it is primarily found in the liver (Nathwani, et al., 2005)

In combinations with the physical examination and history, the evaluation of other serum enzymes should aid in differentiating the source of increased AST levels. AST is present in both the cytoplasm and mitochondria of hepatocytes (and many other cells) and will elevate in states of altered membrane permeability. In such cases, levels are expected to be less than in states of frank necrosis, when both cytoplasmic and mitochondrial enzymes are released.

Alanine aminotransferase (ALT)

ALT is considered to be liver specific in small animals. This enzyme is present in high concentrations in the cytoplasm of hepatocytes. Plasma concentrations increase with hepatocellular, damage/necrosis, hepatocyte proliferation, or hepatocellular degeneration. ALT is a cytoplasmic enzyme and is considered to be liver specific in primates and some other small animal species (Nathwani, et al., 2005)

Alkaline phosphatase (ALP)

Alkaline phosphatase is a hydrolase enzyme that is eliminated in the bile. It hydrolyzes monophosphates at an alkaline pH. It is particularly present in the cells which line the biliary ducts of the liver. It is also found in other organs including bone, placenta, kidney and intestine. Several isozymes have been identified in humans and preclinical species. Normal levels are in the range of 20-120U/L. It may be elevated if bile excretion is inhibited by liver damage. Hepatotoxicity leads to elevation of the normal values due to the body’s inability to excrete it through bile due to the congestion or obstruction of the biliary tract, which may occur within the liver, the ducts leading from the liver to the gallbladder, or the duct leading from the gallbladder through the pancreas that empty into the duodenum [small intestine]. Increase in alkaline phosphatase and/or bilirubin with little or no increase in ALT is primarily a biomarker of hepatobiliary effects and cholestasis (Ramaiah, et al., 2007). In humans, increased ALP levels have been associated with drug induced cholestasis (Wright et al., 2007).

1.2.10.2 Bilirubin

Bilirubin is the main bile pigment that is formed from the breakdown of heme in red blood cells. The broken down heme travels to the liver, where it is secreted into the bile by the liver. Normally, a small amount of bilirubin circulates in the blood. Serum bilirubin is considered a true test of liver function, as it reflects the liver’s ability to take up, process, and secrete bilirubin into the bile.

Bilirubin production and excretion follows a specific pathway. When the reticuloendothelial system breaks down old red blood cells, bilirubin is one of the waste products. This “free bilirubin”, is in a lipid-soluble form that must be made water-soluble to be excreted. The free, or unconjugated, bilirubin is carried by albumin to the liver, where it is converted or conjugated and made water soluble. Once it is conjugated into a water-soluble form, bilirubin can be excreted in the urine. An enzyme, glucuronyltransferase, is necessary for the conjugation of bilirubin. Either a lack of this enzyme, or the presence of drugs that interfere with glucuronyltransferase, impairs the liver’s ability to conjugate bilirubin. Because the bilirubin is chemically different after it goes through the conjugation process in the liver, lab tests can differentiate between the unconjugated or indirect bilirubin and conjugated or direct bilirubin. The terms “direct” and “indirect” reflect the way the two types of bilirubin react to certain dyes. Conjugated bilirubin is water-soluble and reacts directly when dyes are added to the blood specimen. The non-water soluble, free bilirubin does not react to the reagents until alcohol is added to the solution. Therefore, the measurement of this type of bilirubin is indirect. Test results may be listed as “BU” for unconjugated bilirubin and “BC” for conjugated bilirubin. Total bilirubin measures both BU and BC.

Conjugated and Unconjugated Bilirubin

Bilirubin is a yellowish breakdown product of the heme. It is a part of the hemoglobin molecule that is in the red blood cells. It is thrown out of our body by means of bile or urine. Hence an increase in the level of bilirubin indicates the person could be suffering from certain diseases like jaundice. It is lipid soluble as it is a four ring structure known as tetrapyrrole. Bilirubin is split into two unconjugated versus conjugated bilirubin. Bilirubin when high is brown. However when the level of bilirubin is slightly higher than normal it is yellowish. In some cases depending on the level of bilirubin, when it is elevated it may show even on our skin and sclera.

There are differences between unconjugated and conjugated bilirubin. Unconjugated bilirubin is not soluble with water and conjugated bilirubin is soluble with water. In order to explain this, tetrapyrrole is digested by reticuloendothelial cells that then result in unconjugated bilirubin. The Unconjugated bilirubin combines with albumin and is carried to the liver. From the liver it then joined or “conjugated” to glucuronide by an enzyme called UDP-glucuronyltransferase.

Conjugated bilirubin reacts quickly as compared to unconjugated bilirubin. In order to produce Azobilirubin, which is a red-violet compound; dyes are added to blood sample. Conjugated bilirubin is known as direct bilirubin whereas unconjugated bilirubin is known as indirect bilirubin. Unconjugated bilirubin still produces Azobilirubin whereas conjugated bilirubin only produces it if dye is added.

Unconjugated bilirubin is fat soluble however conjugated bilirubin is water-soluble and hence can be excreted through kidneys. An increase in the level of conjugated bilirubin means an indication towards hepatobiliary disease. Unconjugated bilirubin generally reacts slowly in the absence of an accelerator namely ethanol. In order to calculate indirect bilirubin the formula, Total bilirubin -Direct bilirubin, is used.

Direct bilirubin is measured without an accelerator. Unlike unconjugated bilirubin which binds with neural tissue and also leads to kernicterus (a serious disorder damages the basal ganglia and other parts of the central nervous system) if left untreated or other forms of toxicity, conjugated bilirubin does not bind significantly to neural tissue neither does it lead to any form of toxicity or kernicterus. A very popular or well-known disease that is caused by elevated level of bilirubin in the body is Gilbert’s syndrome. This is mainly hereditary and is caused by elevated level of unconjugated bilirubin but is not serious and may lead to mild jaundice if exerted.

1.3.    Justification of the Study

A large proportion of heavy drinkers develop serious alcoholic liver disease. Susceptibility to alcoholic hepatitis and cirrhosis appears to be influenced by heredity, gender, diet, and co-occurring liver illness. Most alcoholic liver damage is attributed to alcohol metabolism. Liver injury may be caused by direct toxicity of metabolic by-products of alcohol as well as by inflammation induced by these byproducts. Exposure of liver cells to bacterial toxins may contribute to liver disease. Escalating liver injury can lead to fibrosis and, ultimately, to cirrhosis (Penny, 2013; Lombardi et al., 2015).

 1.4 Aim and Objective of Study

The aim of this study is to evaluate the functional status of hepatic tissues of albino rats place on alcohol (0.5 ml and 1.0 ml) once daily.

1.4.1  Objectives of study

To determine:

• The effect of daily alcohol intake (once daily) on Aspartate aminotransferase activity of albino rats.
• The effect of daily alcohol intake (once daily) on alanine aminotransferase activity of albino rats.
• The effect of daily alcohol intake (once daily) on alkaline phosphatase activity of albino rats.
• The effect of daily alcohol intake (once daily) on total and conjugated bilirubin concentration of albino rats.
  

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  Pages:  71

  Category: Project

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  Chapters: 1-5                                 

  Material contains Table of Content, Abstract and References.