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ORIGINAL ARTICLE |
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| Year : 2005 | Volume
: 59
| Issue : 2 | Page : 64-69 |
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Effect of fenugreek, onion and garlic on blood glucose and histopathology of pancreas of alloxan-induced diabetic rats
Gholamali A Jelodar1, M Maleki2, MH Motadayen1, S Sirus1
1 Department of Physiology, College of Veterinary Medicine, Shiraz University, Shiraz, 71345, Iran
2 Department of Pathology, College of Veterinary Medicine, Ferdowsi University, Mashhad, Iran
Correspondence Address:
Gholamali A Jelodar
Professor of Physiology, College of Veterinary Medicine, Shiraz University, Shiraz
Iran
doi:10.4103/0019-5359.13905
PMID: 15738612
BACKGROUND: Many traditional treatments have been recommended in the alternative system of medicine for treatment of diabetes mellitus; however, the mechanism of most of the herbals used has not been defined. AIMS: This study was carried out to clarify the effect of fenugreek, garlic and onion, recommended in Persian folklore medicine as beneficial in the treatment of diabetes, on blood glucose and their possible effect on pancreatic tissue. METHODS AND MATERIAL: Diabetes mellitus was induced in 20 out of 25 adult male albino rats, using intraperitoneal injection of 185 mg/kg BW alloxan. The diabetic rats were divided into four groups, three of which were fed a diet containing 12.5% BW Allium sativum (garlic), Allium cepa (onion) or Trigonella foenum-graecum (fenugreek) for 15 days. The fourth group (positive control) received an ordinary diet. The remaining non-diabetic rats (negative control group) received neither alloxan nor the mentioned plants. Following consumption of plants, blood glucose was measured every day and on the last day the pancreas were removed and stained with H&E and Gomeri aldehyde fuchsin (GAF). Morphology of the pancreatic sections and the following morphometric factors were studied: volume density of B cells, volume density of islets, percent of B cells, number of islets per square millimeter, average area of islets and average volume density of B cell in whole pancreas. STATISTICAL ANALYSIS USED: One-way Analysis of Variance (ANOVA) test and Duncan's multiple range tests were used to evaluate the data. RESULTS AND CONCLUSION: The results of this study indicate that only garlic was able to reduce blood glucose significantly compared with the control group (P<0.05). In the control positive group all the mentioned morphometric factors were significantly changed in comparison with the control negative (normal health) group, but the same did not show significant change between treated and untreated diabetics.
Keywords: Onion; garlic; fenugreek; alloxan; diabetes; histopathology; pancreas
How to cite this article:
Jelodar GA, Maleki M, Motadayen MH, Sirus S. Effect of fenugreek, onion and garlic on blood glucose and histopathology of pancreas of alloxan-induced diabetic rats. Indian J Med Sci 2005;59:64-9 |
How to cite this URL:
Jelodar GA, Maleki M, Motadayen MH, Sirus S. Effect of fenugreek, onion and garlic on blood glucose and histopathology of pancreas of alloxan-induced diabetic rats. Indian J Med Sci [serial online] 2005 [cited 2010 Sep 2];59:64-9. Available from: http://www.indianjmedsci.org/text.asp?2005/59/2/64/13905 |
| ¤ Introduction | |  |
Although herbal medicines have long been used effectively in treating many diseases in Asian communities and throughout the world, the mechanism of most of the herbals used have not been defined. Many traditional plant treatments for diabetes are also used, but most of the evidence for their beneficial effects is anecdotal.[1] Fenugreek, onion and garlic were reported to have a hypoglycemic effect.[2],[3],[4],[5] Although there are suggestions about the mechanism of action of plants, the exact mechanism is unclear. Plants may act on blood glucose through different mechanisms, some of them may have insulin-like substances,[6] some may inhibit insulinas activity,[7] and others may increase beta cells in the pancreas by activating regeneration of these cells.[8],[9] The fiber of plants may also interfere with carbohydrate absorption; thereby affecting blood glucose.[10] Our initial study showed a profound effect of garlic, onion and fenugreek on serum cholesterol, triglyceride, ALT, and AST of alloxan diabetic rats.[11] The aim of this study was to clarify the effect of above-mentioned plants, recommended in Persian folklore medicine as beneficial in the treatment of diabetes, on blood glucose and their possible effect on pancreatic tissue.
| ¤ Material and methods | | |
Animals
Adult male albino rats (230-260 g) were obtained from the animal house of a medical university and housed at 22±2 in an air-conditioned room and supplied with standard pellet food with tap water ad libitum. All rats received human care according to the criteria outlined in the "Guide for care and use of laboratory animals" prepared by the National Academy of Science and published by The National Institutes of Health.
Preparation of alloxan-induced diabetic rats
Alloxan tetra hydrate (Sigma) was dissolved in sterile distilled water.
Diabetes was induced in 20 rats by intraperitoneal injection of 185 mg/kg (5%). The rats were fasted 12h before and after the alloxan injection. The rats with blood glucose above 250 mg/dl, as well as with polydipsia, polyurea and polyphagia, which last for at least one week, were selected for the experiment.
The range of the diabetogenic dose of alloxan is quite narrow and even light overdosing may be generally toxic causing the loss of many animals;[12] to prevent toxic side effects, ranges of 95 to 195 mg/kg of alloxan (10 mg interval) were tested and 185 mg/kg was selected as the minimum and safest dose for induction of diabetes.
Preparation of food
The rats naturally were not interested in having the plants, so the plants were ground and mixed with standard pellet, and the diabetic experimental group were fed with 12.5% BW of plants, daily. Rats were fed with the prepared food first, and only when they finished that, additional ordinary food was given to them.
Experimental procedure
Following induction of diabetes in 20 rats, they were randomly divided into four groups; three of these were fed with plants for 15 days. While the fourth diabetic group (control positive) and a normal health (control negative), were fed with ordinary pellet food.
Blood glucose measurement
Blood glucose was tested every day. Blood was collected from the tail of fasting (10h) animals. The tail was embedded in 45 water bath and about one millimeter of its end was cut and a drop of blood was used for the blood glucose test with the help of glucometer GX (Ames, USA), further sampling did not need re-cutting of the tail. Accuracy of the glucometer was checked by the Orthotolidin method.
Histopatholgical study
On the last day of the experiment the rats were anesthetized and the tail part (Spelinic) of the pancreas was removed and kept in 10% formaldehyde. Dehydration and clearing of the tissues were formed automatically. The prepared 5-micron thickness sections were stained with Hematoxilin and Eosin (H/E) and Gomeri aldehyde-fuchsin (GAF), a beta cell specific staining.[13] Stained sections were quantitatively (morphometric) and qualitatively (morphological) evaluated. For quantitative analysis the following factors were evaluated:
- Volume density of islets, in 500 microscopic field percentage of islet tissue to total tissues was determination.[14]
- Volume density of B cells in islet tissue, obtained by counting approximately 1300 parts of islet and determination of density of B cells to total islet cells.[15]
- Percent of B cells, cells of approximately 4 islets on each tissue and 40 islets of each group were counted.
- Number of islets per square millimeter.
- The average area of the islets was determined by measuring the diameter of 4 islets in each section and totally of 40 islets in each group.
| ¤ Statistical analysis | | |
The results are expressed as mean± SD. The significance of the differences in the values was performed by one-way ANOVA test and Duncan's multiple range tests. P<0.05 was considered to be a significant difference.
| ¤ Results | | |
The effects of plants on fasting blood glucose (FBS). The effect of the plants on FBS is presented in [Table - 1]. Mean fasting blood glucose in the diabetic untreated group (control positive) was (258.7±7.7) during the whole period. In the normal health group this value was 67.2±4.1 mg/dl. In comparison with the positive control group, the group which consumed garlic showed significantly lower mean FBS (182.9±4.96) (P<0.05).
There was no significant difference of FBS between groups which received fenugreek or onion in comparison with the positive control group.
Effects of consumed plants on the histopathology of the pancreas
Histomorphologic change of pancreas
Pancreatic sections stained with HE showed that alloxan caused severe necrotic changes of pancreatic islets, especially in the center of the islets. Nuclear changes, karyolysis, disappearing of nucleus and in some places residue of destructed cells were visible. The relative reduction of the size and number of islets, especially around the large vessel, and also severe reduction of the number of B cells was obvious. Study of the pancreas of the treated groups did not show a significant difference as compared with the diabetic untreated group.
Results of the Histomorphometric study
The result of this section is summarized in [Table - 2]. In the diabetic untreated group all the 6 factors measured were significantly low in comparison with the normal health group (P<0.05). There was no significant difference between the histopathology of the treated and untreated diabetic rats.
| ¤ Discussion | | |
Consumption of garlic significantly decreased FBS; this finding agreed with others[2],[16],[17],[18]and seemed to support the theory that garlic acted as a hypoglycemic agent. Feeding of animals with other plants did not have a significant effect on FBS. The hypoglycemic effect of garlic may be due to the action of substances like allyle propyl disulphide or diallyle disulphide.[2]
In this study the pancreatic beta cells were destroyed with the help of alloxan. Alloxan and streptozotocin are the most usual substances to induce diabetes mellitus.[19] The rats were fasted for 12h before and after injection of alloxan; unfed animals are more susceptible to alloxan-induced diabetes.[20],[21] Following injection of alloxan, beta cells were selectively destroyed and the histopathogical study of specifically stained beta cells with GAF confirmed this.
The hypoglycemic effect of plants may be due to the presence of insulin-like substances in plants,[6] stimulation of B cells to produce more insulin,[16] high level of fiber which interferes with carbohydrate absorption[10] or the regenerative effect of plants on pancreatic tissue.[8],[9],[22] Specific staining along with ordinary HE staining has been used to clarify the effect of applied plants on pancreatic B cells. GAF staining which was used in this study, was first introduced by Gomeri,[13] and was applied by other scientists for staining of pancreatic B cells.[23],[24] Histopathological study of diabetic untreated rats showed almost complete destruction of B cells, which was due to the proper dose of alloxan used in this study. An inadequate dose will cause partial destruction of B cells in islets.[25] The histopathological study of the diabetic treated group did not show a significant difference with the diabetic untreated group. This finding reveals that the hypoglycemic effect of garlic is not through the action of garlic on the number of B cells, and will support the theory that garlic's hypoglycemic effect might be due to the action of substances like allyle propyl disulphide or diallyle disulphide[2] or due to an increase in the insulin response.[26]
Onion was reported to have a hypoglycemic effect[2],[17],[26] but our result didn't support these reports, which may be due to the volatile substances present in onion and expected to be escaped from onion during food processing.[2],[17] The finding of this study also may not support the report of Sharma, 1986, about the mild hypoglycemic effect of fenugreek in diabetic rats. In this study, the leaf of fenugreek was consumed but most of the available reports about the hypoglycemic effect of fenugreek are about fenugreek seed.[3],[5],[27] In conclusion, of the consumed plants, only garlic reduced FBS significantly but the histomorphometric study of the pancreas of the treated group didn't show a significant change of the pancreatic tissue, hence some other mechanism may be involved in its hypoglycemic action.
| ¤ References | | |
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| 10. | Nelson RW, Ihle SL, Lewis LD, Salisbury SK, Bottoms GD. Effects of dietary fiber supplementation on glycemic control in dogs with alloxan-induced diabetes mellitus. Am J Vet Res 1991;52:2060-6. |
| 11. | Jelodar GA, Nazifi S. Effect of onion, garlic and fenugreek on serum biochemical parameter of diabetic rats. Sci J Vet Med Ahwaz Univ 1998;1:77-85. |
| 12. | Lenzen S, Tiedge M, Jorns A, Munday R. Alloxan derivatives as a tool for elucidation of the mechanism of the diabetogonic action of alloxan. In: Lessons for animal diabetes, E Shafrir, editor, Boston: Birkhauser; 1996. p. 113-22. |
| 13. | Gomeri G. Aldehyde-fuchsin, A new staining for elastic tissue. Am J Path 1950;17:395-406. |
| 14. | Findlay JA, Thomas NW. Histology and cytology of the islets of Langerhans in the Mongolian gerbil Merion's Ungiculatus. Acta Anato 1980;108:446-62. [PUBMED] |
| 15. | O'Brien TD, Hayden DW, Johnson KH, FletcherTF. Immunohistochemical morphometry of pancreatic endocrine cells in diabetic, normoglycemic glucose intolerant and normal cats. J Comp Path 1986;96:357-69. |
| 16. | Chang MW, Johnson MA. Effect of garlic on carbohydrate metabolism and lipid synthesis in rats. J Nutr 1980;110:931-36. |
| 17. | Jain RC, Vyas CR. Hypoglycemic action of onion on rabbits. Br Med J 1974;2:730. |
| 18. | Jain RC, Vyas CR. Garlic in alloxan-induced diabetic rabbit. Am J Clin Nutr 1975;28:684-5. [PUBMED] |
| 19. | Szkudelski T. The mechanism of alloxan and streptozotocin action in B cells of the rat pancreas. Physiol Res 2001;50:536-46. |
| 20. | Szkudelski T, Kandulska K, Okulicz M. Alloxan in vivo does not only exert deleterious effects on pancreatic B cells. Physiol Res 1998;47:343-6. [PUBMED] [FULLTEXT] |
| 21. | Katsumata K, Katsumata K, Jr, Katsumata Y. Protective effect of diltiazem hydrochloride on the occurrence of alloxan-or streptozotocin-induced diabetes in rats. Horm Metab Res 1992;24:508-10. |
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| 23. | Mowry RW, Lowgley JB, Emmel VM. Only aldehyde fuchsin made from pararosanilin stains pancreatic beta cells granules and elastic fiber in anoxidized microsections: Problems caused by mislabeling of certain basic-fuchsin. Stain Technol 1980;55:91-104. |
| 24. | Poole MC, Brinn jE, Morgan CR. Cytomorphometric analysis of pancreatic islet tissue from alloxan diabetic rats. Anat Rec 1981;199:204A. |
| 25. | Lyman DG, Murray EGD. The pathology of the pancreas in experimental diabetes mellitus. Am J Med Sci 1945;210:381-96. |
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Tables
[Table - 1], [Table - 2]
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