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We have taken the liberty of highlighting what we feel are the most relevant points of this report on the study.
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Journal of Clinical Gastroenterology 2003; 37(4):336-33
Silymarin Retards the Progression of Alcohol-Induced Hepatic Fibrosis in Baboons
Charles S. Lieber, MD, MACP; Maria A. Leo, MD; Qi Cao, MD, PhD; Chaoling Ren, MD; Leonore M. DeCarli, BA
Hepatoprotective effects of silymarin in patients with alcoholic liver disease are controversial. For strict control, this was assessed in non-human primates.
As stated elsewhere, the possible reason for the controversy is from inconsistent results in other studies which may have been from lack of control over dose and compliance. The key words with regard to this study are “strict control”.
Twelve baboons were fed alcohol with or without silymarin for 3 years with a nutritionally adequate diet.
Silymarin opposed the alcohol-induced oxidative stress (assessed by plasma 4-hydroxynonenal) and the rise in liver lipids and circulating ALT. Alcohol also increased hepatic collagen type I by 50% over the 3 years with a significant rise in mRNA for [alpha]1 (I) procollagen, both prevented by silymarin. There were corresponding morphologic changes: at 36 months, 2 of 6 animals fed alcohol had cirrhosis and 2 septal fibrosis, with perivenular fibrosis in 2, whereas with alcohol + silymarin, there was only 1 cirrhosis and 1 septal fibrosis, with perivenular fibrosis in 2, and virtually no lesions in the remaining 2.
These results indicate a broad reaching beneficial effect of silymarin against the ravages of liver disease. It also states that the blood tests and biopsies were reliable predictors for the actual physical state of the livers after three years. The subjects with the worse blood test and biopsy results had the most damage.
Silymarin retards the development of alcohol-induced hepatic fibrosis in baboons, consistent with several positive clinical trials. The negative outcome observed in other trials possibly reflects poor compliance resulting in irregular or low silymarin intake. Thus, in view of the innocuity of silymarin, it might be advisable in future clinical studies to insure the controlled administration of sufficient amounts of silymarin.
This conclusion of this study is definite. Silymarin retards fibrosis.
Key Words: cirrhosis; collagen; oxidative stress
Received for publication January 7, 2003; accepted May 6, 2003.
From the Section of Liver Disease & Nutrition, Bronx VA Medical Center & Mount Sinai School of Medicine, Bronx, New York
Supported, in part, by NIH grant AA11115, the Department of Veterans Affairs, the Kingsbridge Research Foundation and MADAUS AG, Cologne, Germany.
Reprints: Dr. Charles S. Lieber, VA Medical Center (151-2), 130 West Kingsbridge Road, Bronx, NY 10468 (e-mail: email@example.com).
Silymarin, extracted from the milk thistle, is a mixture of the 3 isomeric compounds silibinin, silidianin, and silichristin; it has been used traditionally against a variety of liver ailments.1 Silibinin, the main isomer, was shown to prevent liver fibrosis induced by CCl4 in rats.2 It was also found to be a radical scavenger3 and, in patients with alcoholic liver disease,
Silibinin is another word for silybin, the main active ingredient in both Maximum Milk Thistle and UltraThistle. Actually, Maximum Milk Thistle and UltraThistle isolate silybin and then makes it 8 to 10 times more absorbable, bringing that much more to your liver.
some randomized controlled trials with silymarin showed beneficial effects such as improved survival.4 However, other controlled studies did not verify such an effect.5 Since clinical trials in alcoholics commonly suffer from significant variability in alcohol consumption and poor compliance with medications, we assessed the effectiveness of silymarin under controlled experimental conditions. Specifically, we addressed the question whether silymarin opposes alcohol-induced liver injury in terms of oxidative stress, abnormal liver tests, hepatic steatosis and fibrosis in non-human primates, which are much closer to man than the rodents used in all previous experimental studies.
Using baboons gave results which are more relevant to humans.
MATERIALS AND METHODS
Twelve baboons were studied in accordance with the institutional guidelines for animal research. They were fed for 36 months nutritionally adequate liquid diets containing alcohol (50% of calories),6 half of them supplemented with silymarin kindly donated by MADAUS AG (Cologne, Germany). The 2 groups were balanced with regard to sex, weight, and age. One baboon (#2) started 6 months late. Silymarin was given in the diet at a dose of 0.84 mg per calorie, an amount recommended by MADAUS AG as equivalent to the usual therapeutic dose in humans based on pharmacokinetic tests,
MADAUS AG makes Legalon, a prescription-strength standardize milk thistle from Germany. Maximum Milk Thistle has been compared to Legalon in a head to head clinical study and the absorption rate of silybin was 8 to 10 times greater with Maximum Milk Thistle. See study C10.
taking into account the higher metabolic rate in baboons compared with men. Daily alcohol intake was comparable in both groups: 2.84 1 0.33 and 3.34 1 0.32 g/kg body weight for the ethanol and the ethanol + silymarin groups, respectively. The corresponding data in g/animal/d were 49.8 1 4.6 versus 54.1 1 7.8. Intake of silymarin was 39.8 1 5.2 mg/kg body weight and 645 1 92 mg/animal/d.
Percutaneous liver biopsies were performed every 6 months with a Menghini needle. For light microscopy, specimens were coded for blind reading, prepared, and examined for the various stages of fibrosis as described before.7, 8 The tissue obtained was adequate for all morphologic and most chemical assessments except for a few of the collagen and triglyceride measurements because of variable yield of the percutaneous needle liver biopsies.
Measurements were carried out of plasma AST, ALT and total bilirubin (Sigma Kits; SIGMA Diagnostics), 4-hydroxynonenal by GC/MS,9 hepatic total triglycerides,10 collagen type I,11 and [alpha]1 (I) collagen mRNA.12
Plasma 4-hydroxynonenal increased strikingly with ethanol, and this was significantly prevented by silymarin at each of the 6 month time points (Fig. 1) during the first 2 years.
The fact that this increase was “significantly” prevented is very encouraging with regard to both Maximum Milk Thistle’s and UltraThistle’s effectiveness in protecting your liver.
There was a lesser increase of 4-hydroxynonenal in the 3rd year, possibly as a consequence of diminished metabolism secondary to liver disease.
Effect of alcohol (with or without silymarin) on plasma 4-hydroxynonenal. Alcohol increased 4-hydroxynonenal and this was opposed by silymarin, confirming its antioxidative properties. P values: alcohol versus alcohol + silymarin.(
Plasma ALT values (Fig. 2) were significantly lower after silymarin plus alcohol than with alcohol alone. There were no significant differences for AST and bilirubin.
ALT levels are often a better indicator of serious ongoing liver damage than AST. Keeping these low is a valuable effect of silymarin in this study.
Effect of alcohol (with or without silymarin) on plasma ALT. There was an overall significant attenuation by silymarin of the alcohol-induced rise in ALT. ns = not significant.
Attenuation is a lessening.
The overall histologic assessment showed lesser steatosis with silymarin, substantiated by triglyceride measurement which revealed an increase with ethanol and a significant reduction with silymarin after 1 year (161.6 1 37.7 versus 63.5 1 16.8 mg/g liver, P = 0.04) with a similar effect at 18 months (124.6 1 15.6 versus 69.6 1 12.3 mg/g liver, P = 0.02) and at 24 months (147.01 22.8 versus 77.2 1 10.1 mg/g liver, P = 0.02).
Both of these observations are strong testimony to the protective effects of milk thistle at the one year of treatment mark.
Concerning liver fibrosis, alcohol feeding resulted in an increase of collagen type I and silymarin prevented this rise (Figs. 3a, b). In addition, mRNA of [alpha] (I) procollagen was significantly lower after ethanol + silymarin than after ethanol alone (Figs. 4a, b). The histologic stages of fibrosis were consistent with these biochemical data. Indeed, in terms of the lesions achieved at 36 months, 4 of the 6 animals fed alcohol had an advanced stage of disease, reaching cirrhosis in 2 and septal fibrosis in another 2 (Fig. 5a). By contrast, with alcohol and silymarin, there was only 1 cirrhosis and 1 septal fibrosis (Fig. 5b). In the animals given ethanol + silymarin, 2 out of 6 were virtually fully protected after 3 years (Fig. 5b), but only 1 of the 6 animals fed alcohol alone had such an outcome (Fig. 5a). Histologic scores of the fibrosis were also decreased by silymarin (P = 0.03 by [chi]2 or Fisher exact test). Representative lesions observed at 36 months are shown in Figure 6A-F, illustrating the differences in steatosis and fibrosis between baboons fed alcohol with or without silymarin.
Inhibiting fibrosis is the key measure of effectiveness in slowing the progression of serious liver disease. Whether from alcohol or viral hepatitis, fibrosis leads to cirrhosis. As liver cells are killed, scar tissue is formed. This is fibrosis. At a certain point the scar tissue inhibits the function of the liver, this is cirrhosis. The more that functions are inhibited, the more advanced and serious the cirrhosis. The fact that silymarin dramatically inhibited fibrosis in this study is one of the strongest testaments to the daily use of either Maximum Milk Thistle or UltraThistle that we have yet encountered.
a, b. Effect of alcohol, with or without silymarin, on hepatic accumulation of collagen type I. Already after 18 months of alcohol, there was a significant rise in hepatic collagen type I (3a); this was prevented by silymarin which also opposed the subsequent increases (3a, 3b). ns = not significant.
This is more proof of a significant action against fibrosis.
a, b. Effect of alcohol, with or without silymarin, on hepatic mRNA of [alpha]1(I) procollagen. At 18 months, concomitant with the increase in collagen type I (see Figure 3a), there was a corresponding significant rise in mRNA (a). This was prevented by silymarin. This protective effect persisted for the remainder of the 3 years (a, b).
This shows that even after three years of daily use, milk thistle extract was still protecting the liver from fibrosis.
Effect of silymarin on alcohol-induced hepatic fibrosis. Whereas 4 out of 6 baboons fed alcohol for 36 months developed advanced stages of liver fibrosis, namely septal fibrosis (SF) in 2 and cirrhosis (C) in another 2 (5a), there was only 1 cirrhosis and 1 septal fibrosis after alcohol + silymarin (5b). PV = perivenular fibrosis. N = normal histology
This shows that the blood test indicators were correct in predicting levels of damage. The damage was commensurate with the markers from the blood tests.
Representative hepatic lesions in baboons fed alcohol (with or without silymarin) for 36 months. A. Baboon fed alcohol: Perivenular and pericellular fibrosis. Marked steatosis involving 75% of the hepatocytes. B. Baboon fed alcohol + silymarin: Perivenular fibrosis and minimal steatosis. C. Baboon fed alcohol: Septal fibrosis with incomplete nodular formation and marked steatosis. D. Baboon fed alcohol with silymarin. Septal fibrosis. E. Baboon fed alcohol: Cirrhotic liver with classic nodule surrounded by diffuse fibrosis and steatosis. F. Baboon fed alcohol + silymarin: Cirrhotic liver with sparse fibrosis and steatosis (Sirius Red, A-D W 120; E, F W 50).
These pictures show the actual comparative condition of the livers with or without silymarin. Note the dramatic difference in the ones protected by milk thistle. The structure is much, much closer to normal in all these examples.
Which of these would you want your liver to look like?
The effects of silymarin on alcoholic liver disease are controversial, with the presence4 or absence5 of positive effects. This issue had not been resolved by prior animal studies, carried out with rodents which did not duplicate the alcoholic liver fibrosis produced in humans. This difficulty, however, was overcome by the study of silymarin used, for the first time, in a model of alcoholic liver cirrhosis in the baboon.6 It allows for a full control of the 2 crucial variables, namely alcohol and silymarin consumption. Under these conditions, morphologic and biochemical parameters revealed a protective effect of silymarin against alcohol-induced liver injury.
This statement attests to the overwhelmingly positive results of the study with regard to the protective action of silymarin.
The results were significant in terms of oxidative stress, assessed by 4-hydroxynonenal, possibly the most reliable indicator of oxidative liver injury. Positive results were also obtained for ALT, lipids, and especially fibrosis and collagen metabolism. Thus, these favorable effects in non-human primates are consistent with some clinical trials,4 but not with others.5 Possible reasons for these discrepancies between the clinical trials may include variability in alcohol consumption as well as poor compliance with medication intake, not uncommon problems in alcoholic populations. Thus, the negative outcome observed in some of the clinical trials with silymarin may reflect the actual intake of a low amount of silymarin, whereas some of the positive trials may have been more successful in achieving the desirable dosage.
Another difference between the human trials and the present study is the fact that silymarin was used in the baboons to prevent alcoholic fibrosis, whereas in humans, it was given to treat patients with established alcoholic cirrhosis. Accordingly, it might be useful, in future clinical trials, to achieve and control the administration of sufficiently high doses of silymarin, with emphasis on careful assessment of medication compliance and of alcohol intake. In addition, it would be indicated to include patients at an early stage of disease in whom the effect of silymarin on the progression to cirrhosis could be assessed, since the baboons results reveal that silymarin slows the alcohol-induced disease process and can prevent the cirrhosis.
Silymarin slows the disease process for alcohol. And that process is not that different than the disease process of viral hepatitis. In either case, inflammation and fibrosis leads to cirrhosis, cirrhosis leads to death. This makes me all the more adamant about the importance of taking Maximum Milk Thistle or UltraThistle (because of its increased absorption) on a daily basis (because of the need for adequate levels on a consistent basis).
The authors thank L. Shoichet for skillful technical assistance, A. Ponomarenko for graphic and statistical support, Dr. U. Mengs (MADAUS AG) for his comments, and Y. Rodriguez for the diligent secretarial assistance.
2. Mourelle M, Muriel P, Favari L, et al. Prevention of CCl4-induced liver cirrhosis by silymarin. Fundam Clin Pharmacol. 1989; 3:183-191.
3. Comoglio A, Tomasi A, Malandrino S, et al. Scavenging effect of silipide, a new silybin-phospholipid complex, on ethanol-derived free radicals. Biochem Pharmacol. 1995; 50:1313-1316.
PubMed Abstract | CrossRef
4. Ferenci P, Dragosics B, Dittrich H, et al. Randomized controlled trial of silymarin-treatment in patients with cirrhosis of the liver. J Hepatology. 1989; 9:105-113.
5. Pares A, Planas R, Torres M, et al. Effects of silymarin in alcoholic patients with cirrhosis of the liver: results of a controlled, double-blind, randomized and multicenter trial. J Hepatology. 1998; 28:615-621.
PubMed Abstract | CrossRef
6. Lieber CS, DeCarli LM. Animal models of chronic ethanol toxicity.In: Packer L, ed. Methods in Enzymology: Oxygen Radicals in Biological Systems, Part C; Orlando, Florida: Academic Press Inc.; 1994: 585-594.
7. Lieber CS, Leo MA, Mak KM, et al. Choline fails to prevent liver fibrosis in ethanol-fed baboons but causes toxicity. Hepatology. 1985; 5:561-572.
8. Van Waes L, Lieber CS. Early perivenular sclerosis in alcoholic fatty liver: an index of progressive liver injury. Gastroenterology. 1977; 73:646-650.
9. van Kuijk FJGM, Siakotos AN, Fong LG, et al. Quantitative measurement of 4-hydroxyalkenals in oxidized low-density lipoprotein by gas chromatography-mass spectrometry. Anal Biochem. 1995; 224:420-424.
10. Homan R, Anderson MK. Rapid separation and quantitation of combined neutral and polar lipid classes by high-performance liquid chromatography and evaporative light-scattering mass detection. J Chromatogr. 1998; 708:21-26.
11. Moshage H, Casini A, Lieber CS. Acetaldehyde selectively stimulates collagen production in cultured rat liver fat-storing cells but not in hepatocytes. Hepatology. 1990; 12:511-518.
Journal of Clinical Gastroenterology 2003; 37(4):336-339
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