Therapeutic effect of
1. Introduction
The liver plays an astonishing array of vital functions in the maintenance and performance of the body. Additionally, it is also handling the metabolism and excretion of drugs and other xenobiotics from the body thereby providing protection against foreign substances by detoxifying and eliminating them (saleem et al., 2010). Large numbers of xenobiotics are reported to be potentially hepatotoxic. Free radicals generated from the xenobiotic metabolism can induce lesions of the liver and react with the basic cellular constituents such as proteins, lipids, RNA and DNA (Ajith et al., 2007). A growing international health concern is Tuberculosis caused by
2. Materials and methods
2.1. Chemicals
Anti TB drug
2.2. Preparation of plant extract
Whole plant of
2.3. Animal maintenance and feeding
Female albino rats of
2.4. Experimental design
Animals were divided into various groups of six animals each.
Group I: Control (normal saline, 0.9%).
Group II:
Group III: Experimental control-RIF+INH+PZA+ETH at 52, 70,175,140 mg/kg, b.w., (p.o.), 3days/week.
Group VI- VII: INH+RIF+PZA (as in group III) +
Group VIII: Positive control-Silymerin -50mg/kg b.w. (p.o.), daily.
All the animals were euthanized 24hours after the last treatment to perform various biochemical and histological analysis.
2.4.1. Blood biochemical investigations
Blood was drawn from retro-orbital venous sinus and serum was isolated. Serum Urea, Uric acid and Creatinine determined by commercially prepared kit method (E-Merck, Germany). Serum AST and ALT activity were determined with Reitman and Frankel, 1957.
2.4.2. Tissue biochemical investigations
The tissues viz., liver and kidney were quickly excised, washed in ice cold, normal saline and blotted individually freed from extraneous material on ash-free filter paper. The tissues were then homogenized separately in hypotonic buffer (0.008% NaHCO3), pH 7.4, using a Potter-Elvejham homogenizer at 600-1000 rpm in ice cold conditions.
The crude tissue homogenate was centrifuged at 2000 rpm for 15 min (0-4°C). The supernatant was collected and stored at- 20°C until used for estimating tissue biochemical parameters. Lipid Peroxidation (LPO) was determined by measuring thiobarbituric acid reactive substances (TBARS) in tissues according to Sharma and Krishnamurthy, 1968.SOD and Catalase activity were determined according to Misra and Fridovich, 1972 and Aebi, 1974 respectively.
2.4.3. Histopathological investigations
Tissues were fixed with Bouin’s solution. They were later sectioned using a microtome, dehydrated in graded alcohol, embedded in paraffin section, and stained with hemotoxylin and Eosin (H & E).
2.5. Statistical analysis
Results are presented as mean ± S.E. of six animals used in each group. Data were subjected to statistical analysis through one-way analysis of variance (ANOVA) at 5% significance level followed by Student’s t-test at p≤ 0.05( Snedecor and Cochran,1989).
3. Results — Table 1-6 and histopathology
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69 ± 3.81 | 52 ± 2.87 |
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74 ± 4.09 | 53.6±2.96 |
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345±19.07# | 188 ± 10.39# |
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120.22 ± 6.65* | 81 ± 4.48* |
(81.44%) | (78.67%) | |
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112.34 ± 6.21* | 71 ± 3.92* |
(84.29%) | (86.02%) | |
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100.7 ± 5.57* | 63 ± 3.48* |
(88.51%) | (91.91%) | |
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94.4± 5.22* | 60 ± 3.32* |
(90.79%) | (94.11%) | |
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83 ± 4.59* | 59 ± 3.26* |
(94.92%) | (94.85%) | |
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193.3±10.68 | 4.1 ± 0.23 | 0.19 ± 0.010 |
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220±12.16 | 4.0 ± 0.22 | 0.2±0.011 |
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462±25.54# | 3.0±0.17# | 0.8± 0.044# |
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381.3±21.08* | 3.5 ± 0.19 | 0.65± 0.036* |
(30.03%) | (45.45%) | (24.59%) | |
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305.25±16.87* | 3.6 ± 0.199 | 0.58 ± 0.032* |
(58.33%) | (54.54%) | (36.06%) | |
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257±14.21* | 3.7 ± 0.20* | 0.34± 0.018* |
(76.29%) | (63.63%) | (75.4%) | |
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236±13.05* | 3.8± 0.21* | 0.32 ± 0.017* |
(84.10%) | (72.72%) | (78.68%) | |
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216±11.94* | 4.0± 0.22* | 0.3 ± 0.016* |
(91.55%) | (90.9%) | (81.96%) | |
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19.5±1.08 | 5.3± 0.292 | 0.4± 0.022 |
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24±1.33 | 5.6± 0.309 | 0.5±0.027 |
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52.6±2.91# | 10±0.552# | 0.8± 0.044# |
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47±2.59 | 7.6± 0.420* | 0.7± 0.038* |
(16.91%) | (51.06%) | (25%) | |
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44.1±2.44* | 6.22± 0.343* | 0.66± 0.036* |
(25.67%) | (80.42%) | (35%) | |
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35.7±1.97* | 6.1± 0.337* | 0.6± 0.033* |
(51.05%) | (82.97%) | (66%) | |
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30.7±1.70* | 6± 0.331* | 0.575± 0.032* |
(66.16%) | (88.90%) | (75%) | |
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27.8±1.544* | 5.8± 0.320* | 0.55± 0.030* |
(74.92%) | (89.36%) | (83%) | |
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0.31±0.017 | 0.3± 0.016 | 7.97± 0.441 | 7.5± 0.441 |
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0.341±0.018 | 0.33± 0.018 | 8.136±0.449 | 7.78± 0.449 |
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1.94±0.107# | 1.28±0.071# | 6.8± 0.376# | 6.17± 0.376# |
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0.69±0.038* | 1.096± 0.061* | 7.08± 0.391 | 6.69± 0.391 |
(76.68%) | (18.77%) | (23.93%) | (39.09%) | |
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0.57±0.034* | 1.096± 0.061* | 7.18± 0.396 | 7.0± 0.397 |
(84.04%) | (18.77%) | (32.47%) | (62.40%) | |
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0.44±0.0243* | 0.737± 0.041* | 7.4± 0.409 | 7.25± 0.409 |
(92.02%) | (55.47%) | (51.28%) | (81.20%) | |
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0.431±0.0238* | 0.721± 0.039* | 7.75± 0.428 | 7.33± 0.428 |
(92.57%) | (57.04%) | (81.19%) | (87.21%) | |
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0.335±0.018* | 0.665± 0.037* | 7.85± 0.434 | 7.34± 0.434 |
(98.46%) | (62.75%) | (89.74%) | (87.96%) | |
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70.7±3.908 | 48.78± 2.696 | 50.7± 2.802 | 70.1± 3.875 |
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76±4.201 | 51.2± 2.830 | 51.8±2.863 | 62.14± 3.435 |
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37.52±2.074# | 24.39±1.348# | 38.1± 2.106# | 50.8± 2.808# |
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44±2.432 | 33.3± 1.840* | 42.02± 2.322 | 59.67± 3.298 |
(19.52%) | (36.53%) | (31.11%) | (45.95%) | |
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45.6±2.520 | 35.9± 1.984* | 42.5± 2.349 | 59.97± 3.315 |
(24.35%) | (47.19%) | (34.92%) | (47.51%) | |
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55±3.040* | 41.15± 2.274* | 44.05± 2.435 | 61.96± 3.425* |
(52.68%) | (68.71%) | (47.22%) | (57.82%) | |
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60.9±3.366* | 43.13± 2.384* | 45.2± 2.498 | 62.1± 3.432* |
(70.46%) | (76.83%) | (56.34%) | (58.54%) | |
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68±3.759* | 41.83± 2.312* | 50.6± 2.775* | 62.57± 3.458* |
(91.86%) | (71.50%) | (96.03%) | (58.54%) | |
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1971±108.95 | 2449.99±135.44 |
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2114±116.86 | 2257.14±124.77 |
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1335.71±73.84# | 1785.7±98.72# |
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1721.4± 95.16* | 1814.28±100.29 |
(60.71%) | (4.30%) | |
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1864.28±103.06* | 1907.14±105.43 |
(83.02%) | (18.28%) | |
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1889±104.43* | 2042.85±112.93 |
(87.09%) | (38.71%) | |
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1966.06±108.69* | 2149.99±118.85* |
(99.22%) | (54.83%) | |
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1969±108.85* | 2107.14 ±116.48 |
(99.68%) | (48.38%) | |
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4. Discussion
The liver diseases remain one of the serious health problems as variedly exposed to xenobiotics. Modern medicines have little to offer for alleviation of hepatic diseases. Although
Effects of administration of Anti TB Drugs and
Significant rise above the normal upper limits in the measured serum transaminases in toxicant group on day 60 of the experiment was a biochemical indication of liver injury. Elevated levels of serum enzymes, AST and ALT are indicative of cellular leakage, and loss of functional integrity of cell membrane in liver (Ranawat et al., 2010). Oral administration of
The increased level of serum alkaline phosphatase is reliable marker of liver damage, which occurs due to the
As a measure of renal function status, serum urea, uric acid and creatinine are often regarded as reliable markers (Adebisi et al., 2000). Serum creatinine has been used to estimate glomerular filtration rate. Thus, elevations in the serum concentrations of these markers are indicative of renal injury (Adebisi et al., 2000; Adewole et al., 2007).The same was observed after toxicant administration. It may be due to dysfunctional and dystrophic changes in the liver and kidney. Experiment has shown that
Various biochemical parameters were measured in liver and kidney tissues. The levels of TBARS in liver and kidney tissues of ATD intoxicated rats were significantly elevated when compared to the level of TBARS in control animals. The increased lipid peroxidation results in changes in cellular metabolism of the hepatic and extra hepatic tissues, which ultimately leads to the whole cell deformity and cell death (Arun and Balasubramanian, 2011). The administration of herbal drug
Concomitant cellular oxidative stress was manifested by reduced GSH levels and increased lipid peroxidation. The inverse linear relationship between the ROS level and the GSH level indicated that free radical species were generated by exposure to anti TB drugs which reduced intracellular antioxidant levels. The results indicate that, the herbal drug
Superoxide dismutase and Catalase mutually function as important enzymes in elimination of Reactive oxygen species (ROS). SOD is the major attractive metalloprotein in the antioxidant family. The defensive antioxidant enzyme next to SOD is CAT. CAT is an enzymatic antioxidant widely distributed in all animal tissues, and the highest activity is found in the red cells and liver. Both are the key component of the antioxidant defense system. In the present study, the observed decrease in SOD and CAT activities were presumably associated with the increased oxidative stress caused by these toxicants that might be due to low level of zinc (a metal constituent of the enzyme SOD) in liver tissue (Arun and Balasubramanian, 2011) Therapy at 200-400 mg/kg b.w. reversed the SOD and CAT activity in the liver tissues and protected from free radical induced oxidative stress. These observations are substantiated by author (Gnanadesigan et al., 2011).(Table 5)
ATPase is a membrane bound enzyme. Since, phosphatase is a constituent of all the body tissues; it plays an important role in inorganic pyrophosphates activity. ATPase activity may be considered as a marker for assessing hepatocellular damage induced by hepatotoxic agents.(Table 6)
In our experiment, a concurrent fall was found in ATPase in liver after toxicants exposure. It might be due to dysfunctional and dystrophic changes in the mitochondria and cell membrane permeability. This damage was also very clearly visible in histopathological studies after toxicant administration. These observations are substantiated by other authors (Gao and Zhou, 2005; Krithika and Verma, 2009).The effect of the extracts on ATPase was as pronounced with 100mg/kg as with 200-400mg/kg b.w.
Liver damage induced by toxicant was associated with a variety of biochemical abnormalities following loss of integrity of the cell membrane or interference with normal hepatocytes metabolism and function.
The reason for hepatoprotective effect of the extracts may be due to presence of lignans and flavonoids which might have scavenged the free radical offering hepato protection.
5. Conclusion
The results of this study showed that common medicinal plant,
6. Histopathological studies
Acknowledgments
Authors are grateful to Jiwaji University for providing lab facility and UGC for financial assistance.
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