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ISSN (Print): 2320-5148

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ANTI-HYPERGLYCEMIC ACTIVITY OF SIMVASTATIN ALONE (THERAPEUTIC DOSE) AND COMBINATION OF SIMVASTATIN AND GLIPIZIDE (SUB THERAPEUTIC DOSES) ON ALLOXAN INDUCED HYPERGLYCEMIA IN ALBINO RATS

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ANTI-HYPERGLYCEMIC ACTIVITY OF SIMVASTATIN ALONE (THERAPEUTIC DOSE) AND COMBINATION OF SIMVASTATIN AND GLIPIZIDE (SUB THERAPEUTIC DOSES) ON ALLOXAN INDUCED HYPERGLYCEMIA IN ALBINO RATS

V.S. Harish Kumar*, N.R. Sindhu, Rajashri S. Patil and Umakant Patil

  1. S. Institute of Medical Sciences & Research Centre, Davangere, Karnataka, India

    ABSTRACTThe present study was designed to evaluate anti-hyperglycemic activity of simvastatin alone and the combination of sub therapeutic doses of simvastatin and glipizide. Hyperglycemia was induced experimentally in albino rats by subcutaneous injection of alloxan in a dose of 175 mg/kg body weight. After 72 hours of alloxan treatment, rats showing hyperglycemia (blood glucose level of 400 mg/dl and above) were included in the study. They were divided into four groups of 6 animals each (n=24). Oral administration of normal saline 0.5 ml, glipizide 2.5 mg/kg body weight, simvastatin 10 mg/kg body weight and sub therapeutic doses of both test (simvastatin 5 mg/kg body weight) and standard (glipizide 1.25 mg/kg body weight) drugs, was done respectively into each of the four groups for 30 consecutive days in order to assess the effect in terms of reduction in blood glucose level. Blood glucose was estimated on 0th, 10th, 20th, and 30th days of study in fixed time intervals. In the test group, there was a gradual fall in the blood glucose level which reached up to 308.3 mg/dl by 30th day of study (P < 0.001). In case of combination of sub therapeutic doses of simvastatin and glipizide, the fall in blood glucose level was gradual and sustained and it reached up to 201.5 mg/dl by the 30th day (P < 0.001).  These observations are comparable with the results obtained in case of glipizide treated rats, the standard group. Simvastatin appreciably lowered the blood glucose level, but the combination of sub therapeutic doses of simvastatin and glipizide, by virtue of their possible synergistic effect produced further reduction in the blood glucose level.  This study provides evidence in support of a potential anti-hyperglycemic effect of simvastatin and its combination with glipizide. Thus the combined treatment of simvastatin and glipzide may have added benefit for the diabetic patients associated with hyperlipidemia

    Keywords:

    Simvastatin,

    Anti-hyperglycemic,

    Glipizide,

    Alloxan

    INTRODUCTION: Diabetes mellitus (DM) is the world’s largest endocrinal disorder and a major threat to the global public health. It involves metabolic abnormalities of carbohydrates, fats and proteins. This metabolic disorder is characterized by hyperglycemia, associated with accelerated atherosclerosis leading to increased risk and incidence of myocardial infarction (MI), stroke and peripheral arterial disease.  Diabetes is also the leading cause of adult blindness and end-stage renal disease.

    If glycemic controls are achieved satisfactorily by therapeutic and non-therapeutic approaches, it is possible to reduce the progression of atherosclerosis, stabilize rupture-prone plaques and thus prevent arterial thromboembolism and eventually prevent cardiac death due to MI. The concomitant incidence of retinopathy and nephropathy can also be minimized. Overall therapeutic approaches to manage DM include usage of anti-diabetic drugs and anti-hyperlipidaemic drugs.

    Simvastatin, an anti-hyperlipidemic agent belongs to statins family, a class of drugs that competitively inhibit 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. This enzyme catalyzes the conversion of HMG-CoA to mevalonate, the rate-limiting step of the cholesterol synthesis pathway 1.  Recently, HMG-CoA reductase inhibitors have been reported to exert various other effects in addition to their lipid-lowering ability. Simvastatin attenuates leukocyte-endothelial cell interactions and subsequent blood-retinal barrier breakdown via suppression of vascular endothelial growth factor-induced intercellular adhesion molecule-1 (ICAM-1) expression in the diabetic retina. It may thus be useful in the prevention of diabetic retinopathy 2.

    Simvastatin is the mainstay in the management of hyperlipidemia. Clinical trials indicated that diabetics profit from this agent 3 as it is not uncommon to see both conditions co-existing.

    There have been very few studies on simvastatin as an anti-hyperglycemic agent and no study has investigated on its combination with glipizide for anti-hyperglycemic effect. Hence the present study was designed to evaluate the anti-hyperglycemic effect of simvastatin alone as well as its combination effect with glipizide in sub therapeutic doses, and to compare its anti-hyperglycemic effect with therapeutic dose of glipizide, a standard drug which is one of the sulfonylureas. This emphasis may widen the therapeutic horizon for the said agent.  

    MATERIALS AND METHODS:

    1. Drugs and chemicals: Simvastatin, Glipizide, Alloxan, Dextrose, Ether.
    2. Other requirements: Heparinized micro capillary tubes and Glucometer (ACCU-CHEK ACTIVE) with strips, Manufactured by- Roche Diagnostics GmbH, D-68298 Mannheim, Germany, Batch No- 91000247.
    3. Ethical clearance: The study was conducted in accordance with the National Institute of Health guidelines for the care and use of animals in research, and the protocol was approved by the Institutional Animal Ethical Committee, S. S. Institute of Medical Sciences and Research Centre, Davangere, Karnataka.
    4. Maintenance of rats: Healthy Albino rats (150-200gm), 12 weeks old of either sex, from the central animal house of S.S.I.M.S & R.C, Davangere, were used for the study. They were maintained under controlled conditions of temp 23±2◦C in 12hrs of light and 12 hrs of dark cycles per day. The animals were housed individually in polypropylene cages containing sterile paddy husk as bedding and were fed on normal diet & water ad libitium.
    5. Experimental Procedure:
      1. Induction of Diabetes- It is done by subcutaneous injection of alloxan in a dose of 175 mg/kg body weight 4, 5. Rats were treated with 10% dextrose orally to combat the early phase of hypoglycaemia 6. After 72 hours of alloxan treatment, the blood sugar was estimated and those with blood glucose level of 400mg/dl and above were included in this study.

    Groups- The rats were divided into four groups of six animals each and drugs were administered every day for 30 consecutive days.

    Group 1 (Control): received normal saline 0.5 ml by oral route daily for 30 days.

    Group 2 (Standard): received glipizide 2.5mg/kg body weight by oral route daily for 30 days 7.

    Group 3 (Test): received simvastatin10mg/kg body weight by oral route daily for 30 days 8.

    Group 4 (Sub therapeutic): received the sub therapeutic doses of both test (simvastatin 5 mg/kg body weight) and standard (glipizide 1.25 mg/kg body weight) drugs simultaneously by oral route daily for 30 days to assess their combined effect in terms of reduction in blood glucose level.

    1. Collection of blood sample and estimation of blood glucose level- Blood samples were collected from the retro-orbital plexus of each rat after induction of mild anesthesia with ether, 9  at the intervals of 0th min (before drug administration), 30th min, 1st hr, 2nd hr, 3rd hr, 6th hr, 12th hr and 24th hr (after the administration of drugs) on 0th day, 10th day, 20th day, 30th day and blood glucose levels were estimated by using glucometer and data were recorded for statistical analysis, which was done using repeated measures ANOVA test. The effect of simvastatin alone and the combination of it with glipizide was compared with the anti-hyperglycemic activity of glipizide, the standard drug.

    RESULTS: The rats blood glucose concentration of 400 mg/dl and above, obtained after 72 hrs of alloxan treatment were considered for the present study.

    Table 1 & chart 1: In the control group of rats receiving normal saline over a period of 30 days, there was no much variation in the blood glucose level. Initially on 0th day, the blood glucose level was 432.5 mg/dl and at the end of 30th day it was 451.2 mg/dl. Though there was a marginal increase in blood glucose compared to the initial level, it was not statistically significant.

    Table 2 & chart 2: In the second group treated with glipizide 2.5mg/kg body weight, hyperglycemic rats showed significant fall in blood glucose level from 402.7 mg/dl on 0th day to 166.8 mg/dl on 30th day. This group is considered as the standard.

    Table 3 & chart 3: In the test group of simvastatin (10mg/kg body weight) treated rats, there was gradual reduction in the blood glucose level up to 6th hr observation on the respective days of blood glucose estimation. There is a marked fall in the blood glucose level from 404.5 mg/dl on 0th day to 308.3 mg/dl on 30th day, which is statistically highly significant (P < 0.001). However the reduction in blood glucose level in this group is not as significant as that observed in the standard group (table 2 & chart 2).

    Table 4 & chart 4: In the fourth group of rats receiving the combination of sub therapeutic doses of simvastatin 5 mg/kg body weight and glipizide 1.25 mg/kg body weight over a period of 30 days, there was a gradual and sustained reduction in blood glucose level from 400.7 mg/dl on 0th day to 201.5 mg/dl on 30th day, which is statistically highly significant (P < 0.001). Blood glucose levels in this group showed a fall comparable to standard values obtained by treatment with glipizide (table 2 & chart 2).

    TABLE 1: CONTROL GROUP - EFFECT OF NORMAL SALINE (0.5 ml) ON BLOOD GLUCOSE (mg/dl) LEVEL OF ALLOXAN INDUCED HYPERGLYCEMIC RATS.

    Period 0thmin 30th min 1st hr 2nd hr 3rd hr 6th hr 12th hr 24th hr P*Value, sig
    0th Day 432.5±33.4 435.2± 33.7 438.5±33.0 436.0±35.5 437.8±33.5 439.7±31.1 439.2±33.8 439.0±34.0 0.12 NS
    10thDay 443.3±31.6 443.5±32.8 442.7±27.4 443.2±33.3 441.3±31.4 442.2±26.9 442.8±28.9 442.7±37.2 1.0 NS
    20thDay 442.3±29.2 443.7±30.7 438.3±30.4 443.3±33.2 442.2±31.9 439.3±28.8 443.0±28.1 448.0±32.0 0.12 NS
    30thDay 442.5±25.0 444.3±25.9 441.5±25.6 443.8±33.8 442.7±25.2 441.7±19.7 445.5±20.0 451.2±25.1 0.31 NS

    Values are means ± SD, * Repeated measures ANOVA test, NS= Non significant.

    TABLE 2: STANDARD GROUP - EFFECT OF GLIPIZIDE (2.5 mg/kg body weight) ON BLOOD GLUCOSE (mg/dl) LEVEL OF ALLOXAN INDUCED HYPERGLYCEMIC RATS

    Period 0th min 30 th min 1st hr 2nd hr 3rd hr 6th hr 12th hr 24th hr P* Value, sig
    0th Day 402.7±10.3 396.8±11.0 388.8±12.1 358.8±10.9 341.5±5.2 328.5±8.5 317.7±8.9 306.3±9.5 <0.001 HS
    10thDay 294.2±8.3 293.7±6.2 288.8±8.0 285.2±7.6 282.2±6.4 279.8±7.8 278.2±10.5 272.3±10.9 <0.001 HS
    20thDay 265.5±9.2 264.8±8.6 261.5±7.1 253.5±6.4 249.5±6.0 247.2±6.4 243.0±6.4 236.0±9.3 <0.001 HS
    30thDay 226.8±4.9 224.7±4.8 216.2±4.1 206.0±4.5 198.3±3.4 190.5±4.1 177.3±4.5 166.8±5.8 <0.001 HS

    Values are means ± SD, * Repeated measures ANOVA test, HS= Highly significant.

     TABLE 3: TEST GROUP - EFFECT OF SIMVASTATIN (10 mg/kg body weight) ON BLOOD GLUCOSE (mg/dl) LEVEL OF ALLOXAN INDUCED HYPERGLYCEMIC RATS

    Period 0th min 30 th min 1st hr 2nd hr 3rd hr 6th hr 12th hr 24th hr P* Value, sig
    0thDay 404.5±18.9 403.8±22.1 345.3±39.6 307.3±56.8 303.3±59.0 301.8±61.0 312.5±53.2 365.7±71.8 <0.001 HS
    10thDay 361.2±73.3 358.7±73.5 329.8±82.8 299.8±72.2 284.8±70.5 264.8±74.7 302.5±63.0 347.2±66.4 <0.001 HS
    20thDay 333.8±61.7 332.8±61.5 320.5±65.5 284.3±73.9 270.5±75.6 256.8±75.9 275.8±76.1 330.5±81.6 <0.001 HS
    30thDay 328.8±72.5 327.5±72.1 302.3±68.0 277.5±70.7 263.5±69.3 252.7±72.0 270.5±79.0 308.3±79.9 <0.001 HS

    Values are means ± SD, * Repeated measures ANOVA test, HS= Highly significant.

    TABLE 4: SUB THERAPEUTIC GROUP - COMBINATION EFFECT OF SIMVASTATIN (5mg/kg body weight) & GLIPIZIDE (1.25 mg/kg body weight) ON BLOOD GLUCOSE (mg/dl) LEVEL OF ALLOXAN INDUCED HYPERGLYCEMIC RATS

    Period 0th hr 30 th min 1st hr 2nd hr 3rd hr 6th hr 12th hr 24th hr P* Value, sig
    0thDay 400.7±19.4 397.2±19.6 389.7±17.9 369.7±21.1 356.5±16.6 345.0±10.5 310.0±10.1 321.3±8.7 <0.001 HS
    10thDay 314.7±6.4 312.2±6.0 305.0±6.0 291.5±8.7 287.5±5.2 284.5±4.6 281.3±10.3 279.7±10.7 <0.001 HS
    20thDay 275.8±5.4 274.5±6.5 269.8±4.7 264.7±4.2 260.0±3.6 254.2±6.9 250.3±5.7 246.2±5.3 <0.001 HS
    30thDay 240.8±3.8 239.7±3.7 235.5±3.5 231.8±3.3 229.5±5.9 227.2±8.4 213.2±9.2 201.5±5.5 <0.001 HS

    Values are means ± SD, * Repeated measures ANOVA test, HS= Highly significant.

     Fig 3

  2. The combination treatment with sub therapeutic doses of simvastatin and glipizide decreased the blood glucose concentration more than simvastatin treatment alone which is statistically highly significant.DISCUSSION: As already eluded, diabetes mellitus and hyperlipidemia often coexist and are among the high risk factors for cerebrovascular and cardiovascular morbidity and mortality. HMG-Co A reductase inhibitors have been shown to decrease the risk of cerebrovascular and cardiovascular events 10.  Statins are presently the main stay in the treatment of hyperlipidemia, which on regular use have shown to reduce the rate of morbidity and mortality in the above said disorders.In the present study, the results obtained by therapeutic dose of simvastatin alone and the combination of it with glipizide in sub therapeutic doses are quite appreciable. With reference to the standard drug, the effect of sub therapeutic dose combination of drugs has given nearly comparable results. Simvastatin reduced the blood glucose level appreciably until 6th hour after which a slight rise in blood glucose, lacking statistical significance was observed. However in case of combination, not even a slight rise in blood glucose level was observed on further readings after the 0th day, indicating persistent effect on the lowering of blood glucose on regular use.The results definitely indicate additive action of simvastatin on the pharmacodynamic response of glipizide even in doses below their normal therapeutic range. Simvastatin improves the prognosis of diabetic patients with coronary heart disease 11.    It exerts cardio protective effects and improves Insulin resistance, 12 which may be one of the contributing factors. Other studies have also shown multiple benefits of simvastatin like anti-inflammatory action, 13 suppression of oxidative stress  14   and its antioxidant effect 15 that may have role to play.CONCLUSION: Simvastatin reduces the blood glucose level, whereas the sub therapeutic doses of simvastatin and glipizide in combination will be more effective in lowering the blood glucose concentration, cost beneficial and possibly safer than the individual drug alone as the occurrence of adverse effects will decline with reduction in individual drug dose.

    The present observation requires confirmation with further extensive studies. If it is confirmed, it may further rationalize and widen therapeutic utility of simvastatin. Simvastatin is one of the important drugs used for hyperlipidemia. By this study we can emphasize upon the usage of simvastatin and its combination with glipizide, in sub therapeutic doses for the patients of diabetes associated with hyperlipidemia.

    ACKNOWLEDGEMENT: We would like to thank Micro Labs Ltd for providing pure drug samples.

    CONFLICT OF INTEREST: None declared.

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      How to cite this article:

      Kumar VSH, Sindhu NR, Patil RS and Patil U: Anti-Hyperglycemic Activity of Simvastatin alone (Therapeutic Dose) and Combination of Simvastatin and Glipizide (Sub Therapeutic Doses) on Alloxan induced Hyperglycemia in Albino Rats. Int J Pharm Sci Res. 3(11); 4398-4403.

       

Article Information

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4398-4403

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English

Ijpsr

V.S. Harish Kumar*, N.R. Sindhu, Rajashri S. Patil and Umakant Patil

S. S. Institute of Medical Sciences & Research Centre, Davangere, Karnataka, India

harishkvspharma@gmail.com

20 July, 2012

14 September, 2012

23 October, 2012

http://dx.doi.org/10.13040/IJPSR.0975-8232.3(11).4398-03

01 November,2012

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