Warning: strftime() [function.strftime]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 5186

Warning: strftime() [function.strftime]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 5186

Warning: strftime() [function.strftime]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 5186

Warning: strftime() [function.strftime]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 5186

Warning: mktime() [function.mktime]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 6277

Warning: mktime() [function.mktime]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 6277

Warning: date() [function.date]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 1086

Warning: date() [function.date]: It is not safe to rely on the system's timezone settings. You are *required* to use the date.timezone setting or the date_default_timezone_set() function. In case you used any of those methods and you are still getting this warning, you most likely misspelled the timezone identifier. We selected 'America/New_York' for 'EDT/-4.0/DST' instead in /home/treesfor/domains/tfljournal/public_html/lib-common.php on line 1086
Trees for Life Journal - Role of Rosemary Leaf Extract Against Various Doses of Gamma Radiation
 

Research Articles

Role of Rosemary Leaf Extract Against Various Doses of Gamma Radiation

Garima Sancheti and P. K. Goyal

PEER REVIEWED
There may be tremendous practical value to understanding how to neutralize or reduce the damaging (and lethal) effects of radiation on the human body. Treatment of mice with rosemary extract prior to irradiation is reported herein to prolong life and reduce the symptoms of radiation sickness.
PEER REVIEWED


Garima Sancheti and P. K. Goyal*

Radiation & Cancer Biology Laboratory, Department of Zoology, University of Rajasthan, Jaipur – 302 004 (India) *Corresponding author

Email: pkgoyal2002@rediffmail.com

Trees for Life Journal 2007, Vol 2:2

The electronic version of this article is the complete one and can be found online at: http://www.tfljournal.org/article.php/2007042416194363

Received: July 29, 2006; Accepted: January 28, 2007; Published: August 1, 2007

Copyright: ©2007 Garima Sancheti et al.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.




In light of the various nuclear power plant accident, military and terrorism scenarios that have entered the world consciousness over the past two generations, there is tremendous practical value to understanding how to neutralize or reduce the damaging (and lethal) effects of radiation on the body. Despite substantial research over the past three decades, no chemical compounds have been found to be perfectly safe and effective for this purpose. Research on the use of plant extracts to protect against radiation exposure is not widely known, however the low toxicity and minimal side effects of many plant products are well known in both western science and traditional medicine. Examining the benefits of plant-based therapy with rigorous quantitative science is beginning to gain a toehold in the field of radioprotection. There are now a number of mechanism-based reports of substantial in vitro and in vivo activity from plant preparations [reviewed in March 2007: Jagetia (2007) Journal of Clinical Biochemistry and Nutrition. 40: 74-81]. By presenting the following peer-reviewed study, we hope to add to that body of evidence and to stimulate further research in this neglected arena.




Abstract

The present investigation reports the radiomodulatory effect of Rosmarinus officinalis (rosemary) leaf extract against radiation-induced hematological alterations in Swiss albino mice at various post-autopsy intervals (i.e., between 24 hours to day 30). Treatment of animals with rosemary extract (1000 mg/ kg body wt) prior to irradiation was found to delay the onset of mortality and reduced the symptoms of radiation sickness such as ruffled hairs, lethargy, anorexia and diarrhea in comparison to radiation alone treated animals. Rosemary treated experimental groups exhibited a dose dependent rise (9 < 6 < 3 Gy) in the number of leucocytes (i.e., lymphocytes, monocytes, basophils, eosinophils and neutrophils) by the 30th day post autopsy interval in comparison to the control. Irradiation resulted in a significant increase in lipid peroxidation levels (p< 0.01, p< 0.001) and a reduction in glutathione levels (p<0.05, p<0.001) in blood as observed in radiation alone treated animals. Conversely, treatment of mice with rosemary extract exhibited a significant decrease (p< 0.01, p< 0.001) in lipid peroxidation level and an increase (p< 0.05, p<0.001) in glutathione content.

Introduction

Radiation protection is at a cross-road after radiation incidents and unacceptable tragedies such as those at Chernobyl and Three Mile Island. Radiation induced damage to the normal tissues can be partially reduced by the use of radioprotectors that reduce the damaging effects of radiation, including radiation-induced lethality (4,22,38). Various workers have investigated the potential application of radioprotective chemicals in the event of planned and unplanned exposure (i.e., clinical oncology, radiation site cleanup, military scenarios, radiological terrorism, radiation accidents, etc.) (18,25,40).

A substantial amount of research has been carried out in the field of chemical radioprotection during the last few decades; however, no safe and ideal synthetic radioprotectors are available to date. Recently, interest has generated in developing the potential drugs of plant origin for the amelioration of radiation effects. Plants and their products are well known to have an advantage over the synthetic compounds in terms of their potential low/no toxicity at the effective dose with minimum or no side effects (3,9,33,34,41). However, the use of medicinal plants suffers from lack of robust scientific evidence to support their use. Therefore, studies supporting or rejecting their role in the treatment of various health disorders is of great need (3).

Rosemary (Rosmarinus officinalis), belonging to the family Lamiaceae, is a common medicinal and aromatic plant grown in many parts of the world. It is indigenous to Southern Europe, particularly on the dry rocky hills of the Mediterranean region. Rosemary is used as a culinary herb, a beverage drink, as well as in cosmetics; in folk medicine it is used as a tonic and stimulant, analgesic, antirheumatic, carminative, diuretic, expectorant, anti-epileptic, anti-spasmodic in renal colic, dysmenorrhoea, for relief of respiratory disorders, effects on human fertility, and the stimulation of hair growth (1). Rosemary has been shown to be safe in toxicity studies in animal models when added as an antioxidant to food (35).

Since time immemorial, the plant has been used traditionally by people for curing various health disorders around the world. Caribs of Guatemala use rosemary to cure various human diseases (12). Rosemary has been described as a wonder-drug in literature and in various medieval drug monographs as well (36,44). Thus, wide acceptability and diverse pharmacological and anti-oxidative properties of the plant stimulated us to evaluate the radio modulatory effect of Rosmarinus officinalis in Swiss albino mice exposed to various doses of gamma radiation.

Materials and Methods

Animal care and handling

Male Swiss albino mice (Mus musculus), 6-8 weeks old, weighing 20-24 g., from an inbred colony were used for the present study. The animals were provided standard mice feed (procured from Hindustan Lever Ltd., India) and water ad libitum and were maintained under controlled conditions of temperature and light (Light: dark, 10 hrs: 14 hrs.). Four animals were housed in a polypropylene cage with locally procured paddy husk (Oryza sativa) as bedding throughout the experiment. Tetracycline-containing water (0.13 mg/ml) was provided once a fortnight and was given as a preventive measure against infections. Animal care and handling were performed according to the guidelines set by the World Health Organization (WHO), Geneva, Switzerland and the INSA (Indian National Science Academy), New Delhi, India. The Departmental Animal Ethical Committee approved the present study.

Irradiation

The cobalt teletherapy unit (Co-60) at the Cancer Treatment Centre, Radiotherapy Department, SMS Medical College & Hospital, Jaipur, India, was used for irradiation. Unanaesthetized animals were restrained in well-ventilated Perspex boxes and exposed to various doses of gamma radiation (i.e., 3, 6 and 9 Gy) at a distance (SSD) of 80 cm from the source at a dose rate of 0.85 Gy/min.

Taxonomic description of the plant

Rosemary is an evergreen shrub growing to 1.5 m by 1.5 m at a medium rate. The leaves of rosemary are about 1 inch long, linear, revolute, dark green above and paler and glandular beneath, with camphoraceous aromatic odour. The scented hermaphrodite flowers are small and pale blue. Much of the active volatile principle resides in their calyces. There are various other varieties of the plant, but the green-leaved variety is the kind used medicinally (13).

Preparation of plant extract

The identification of the plant Rosmarinus officinalis (family: Lamiaceae) was done by a botanist, Dr Deepak Acharya, (Voucher Specimen No: DDC/2001/DEPTBT/ACHARYA2430) of the Department of Botany, Danielson College, Chhindwara, Madhya Pradesh (India). The non-infected leaves of the plant were collected, carefully cleaned, shade dried and powdered in a grinder. The plant material was prepared by extracting 200 gm of leaf powder with double distilled water by refluxing for 36 hrs (12 hrs. x 3) at 55 ± 5°C. Pellets of the extract were obtained by evaporation of its liquid contents in the incubator. An approximate yield of 22 % extract (w/w) was obtained.

The required dose for treatment was prepared by dissolving the drug pellets in double distilled water and administered by oral gavage with a micropipette (100 µl/ animal) at a dose of 1000 mg/ kg body wt./animal (1000 mg of 22% of original plant weight). Henceforth, rosemary leaf extract will be called RE.

Experimental Design

Optimum dose determination

A dose selection of Rosmarinus officinalis (RE) was done on the basis of a drug tolerance study. For this purpose, various doses of RE extract (100, 200, 400, 800, 1000, 1500 and 2000 mg/kg body wt.) were tested for their tolerance (once in a day for 5 consecutive days) in Swiss albino mice. One hour after the last administration of RE, mice were exposed to 8 Gy gamma irradiation. All these animals were then observed for 30 days for scoring signs of radiation sickness or mortality. Thus, the optimum tolerated dose of RE (1000 mg/ kg b. wt.) was determined and used for further detailed experimentation (Fig 1).

The LD50/30 and Dose Reduction Factor

The efficacy of any protective agent is evaluated by the determination of its dose reduction factor (DRF). The DRF of R. officinalis extract (RE) based on LD50/30 survival experiment was calculated after irradiating a large number of Swiss albino mice to different doses of gamma rays in the presence (experimental) or absence (control) of RE. The percentage of mice surviving at each radiation dose till 30 days following such exposures was used to construct survival dose response curves. Regression analysis was done to obtain LD50/30, and dose reduction factor was computed as:

DRF =

LD50/30 of Experimental Animals

LD50/30 of Control Animals

The LD50/30 values for control and experimental animals obtained from the survival data are 6.85 and 10.47 respectively. The dose reduction factor of R. officinalis against radiation treatment was calculated on the basis of the survival experiment and was measured as 1.53.

Modification of radiation response

A total of 48 animals used for the experiment were assorted into 4 groups. Mice of group 1 (sham irradiated) were orally administered double distilled water (DDW) at a dose of 1000 mg/ kg body weight, volume equal to RE. Animals belonging to group 2 (RE alone) were given daily rosemary extract at a dose of 1000 mg/ kg/ animal for 5 consecutive days, one hour before irradiation. Animals of group 3 (radiation control) were exposed to various doses of gamma rays alone (i.e., 3, 6 and 9 Gy) one hour after DDW treatment on day 5. Group 4 (RE experimental) received RE (1000 mg/ kg body wt./ animal) as in group 2. One hour after last administration of RE, mice were exposed to various doses of gamma rays, (i.e., 3, 6 and 9 Gy), respectively. These animals were observed daily for any sign of sickness, morbidity, behavioral toxicity and mortality. A minimum of 6 animals from each group were necropsied on days 1, 3, 5, 10, 20 and 30 post-treatment intervals to study hematological and biochemical parameters.

Hematological study

For the study, blood was collected from the orbital sinus of animals from each group in a vial containing 0.5 M EDTA. Differential leucocyte counts (lymphocytes, monocytes, basophils, eosinophils and neutrophils) were determined by adopting standard procedures. The number of each type of leucocytes (lymphocytes, monocytes, basophils, eosinophils and neutrophils) and the total number of leucocytes counted were recorded. The percentage of each type of leucocyte was calculated by the formula:

Number of type of leucocyte

× 100

Total number of leucocytes counted

Biochemical determinants

Biochemical alterations were studied in animals of all the groups at one hour post- exposure to gamma radiation. The level of glutathione (GSH) was determined in blood by the method of Beutler et al. (5). The lipid peroxidation (LPx) level in the serum was measured by the assay of thiobarbituric acid reactive substances (TBARS) according to the method of Okhawa et al. (30).

Statistical analysis

The result for all the groups at various necropsy intervals were expressed as mean ± standard error of the mean (S. E. M.) to evaluate whether the mean of the sample drawn from experimental (RE experimental) deviated significantly from respective control (Irradiation control). Student’s ‘t’ test was used by the method of Bourke et al. (7). The significance level was set at different levels as p<0.05, p<0.01 and p<0.001.

Results

Data are presented in Tables 1-3 and Figures 1-4. The radioprotective effect of rosemary leaf extract (RE) was studied in mice treated with 1000 mg/ kg body wt. RE before exposure to 3, 6 and 9 Gy of gamma radiation. No noticeable signs of behavioral change, sickness or mortality were observed in Sham irradiated/ RE-treated group. Animals exposed to 3 and 6 Gy gamma rays alone did not show mortality throughout the experimental period, but slight laziness was observed in some animals. Animals exposed to 9 Gy gamma rays exhibited epilation, ruffled hair, watering of eyes, weight loss and became lethargic. No animal could survive in the 9 Gy irradiated alone group beyond day 10. Animals pretreated with RE did not exhibit mortality or any symptoms of radiation sickness. General health, activeness, food and water intake were found to be normal in RE pretreated irradiated animals.

After whole body exposure to different doses of gamma radiation (i.e., 3, 6 and 9 Gy), lymphocyte percentages remained significantly lower than normal, and could not regain a normal value even by the last day of autopsy interval (day 30). No significant changes in monocytes, eosinophils and basophil counts were registered in any of the groups. However, monocytes followed a pattern similar to lymphocytes. Following irradiation, a significant increase above normal was observed in neutrophil counts. A normal value could not be restored in any of the irradiated groups till day 30 post-exposure.

Daily administration of 1000 mg/ kg of RE for 5 consecutive days rendered recovery in the different types of leucocytes (i.e., lymphocytes, monocytes, eosinophils, basophils and neutrophils) in comparison to irradiated alone groups, and values close to normal were registered in a dose dependent manner (3> 6> 9 Gy) by post-treatment day 30.

Biochemical determinants

There was no significant difference observed in the levels of glutathione (GSH) and lipid peroxidation (LPx) in the blood content of sham irradiated (group 1) or RE alone treated animals (group 2). In concomitant treatment of RE and radiation (group 4), GSH was found to be further lowered than the radiation treated group. A significant elevation in the values of blood GSH as compared to group 3 was estimated in RE experimental animals. An increase in LPx levels above normal was evident in serum of irradiated mice, while a significant decrease in such values was evident in the RE pretreated irradiated group.

Discussion

Radiation injuries are manifested as a result of enhanced production of free radicals due to oxidative stress. Exposure to radiation causes ionization of molecules in the cells, which sets off potentially damaging reactions via free radical production (23). Free radical mediated processes and oxidative stress have been implicated in the pathogenesis of the aging process and various diseases such as atherosclerosis, liver damage, arthritis, cancer, and neurodegenerative disorders (2). The prevailing view is that intake of antioxidant nutrients can reduce the risk of free radical-related health problems and may prove to be protective against ionizing irradiation.

The present study revealed that the number of lymphocytes declined in a dose dependent manner after exposure to 3, 6 and 9 Gy gamma irradiation. A rapid depression was observed at early intervals which may be attributed to direct destruction of such cells in peripheral blood of mice (31). No significant changes in monocytes, eosinophils and basophils were observed after whole body exposure to different doses of gamma radiation. It may be attributed to the fact that mature granulocytes are radioresistant whereas lymphocytes are extremely sensitive to radiation (8). Furthermore, neutrophil granules altered inversely as compared to lymphocytes. These cells exhibited an early rise while lymphocytes and monocytes declined soon after exposure thus showing an opposite behavior. This can be explained by an abortive rise phenomenon as described earlier by workers (26,39).

According to Hall (14), by the time the number of circulating cells in the blood reaches minimum value as the mature circulating cells begin to die off, the supply of new cells from the depleted precursor population becomes inadequate to replace these, thereby making radiation effects become apparent. Also, this abrupt increase may have appeared due to an abortive rise phenomenon (6,26) or can be interpreted as stimulation effect (11). Hastening the maturation of granulocyte precursors in bone marrow and their release into general circulation can be attributed to a rise in neutrophil counts (28,42).

It is evident from the present study that administration of RE reduced radiation sickness and mortality, and provided protection to differential leucocytes counts (i.e., lymphocytes, monocytes, basophils, eosinophils and neutrophils) in the peripheral blood of mice from the damaging gamma radiation. It has been observed that rosmarinic acid (found in rosemary) is effective in relation to blood circulation and to improve hemodynamics in occlusive arterial diseases (1). Rosemary has been found to contain certain antioxidative (32) and free radical scavenging activity (45) in its active compounds like caffeic acid, carnosolic acid, chlorogenic acid, rosmanol, rosmarinic acid, carnosol, different diterpenes (16,43), rosmari- diphenol, rosmariquinone (17) and other natural antioxidants such as ursolic acid, alkaloid rosmaricine and glucocolic acid (21). In a recent study, carnosic acid was found to render protection to UVA irradiated human skin fibroblasts (29).

The basic effect of radiation on cellular membranes is believed to induce lipid peroxidation (LPx) by the production of free radicals that have the potential to damage DNA and cause cell death (24,27). The level of radiation-induced LPx increased considerably in a dose dependent manner in the entire group 3 irradiated animals, whereas a decrease in the values was observed in RE-treated group 4. The inhibition observed in the LPx level in blood of RE administered animals may have been responsible for the observed radioprotection by plant extract. This view is supported by the investigation of an anti-lipoperoxidant activity of young sprouts of Rosmarinus officinalis that significantly reduced the formation of malondialdehyde in rat hepatocytes1 (9). Sotelo-Felix et al. proposed that carnosol could scavenge free radicals induced by carbon tetrachloride, consequently avoiding the propagation of lipid peroxides in the liver of mice (37).

Studies conducted by Haraguchi et al. report an inhibition of superoxide and lipid peroxidation by 4 diterpenoids from rosemary, i.e. carnosic acid, carnosol, rosmanol and epirosmanol (15). Del Bano et al. investigated the efficacy of carnosic acid, carnosol and rosmarinic acid (active constituents of rosemary) and found these to be radioprotective against chromosomal damage induced by γ-rays (10). The exact mechanism of action of rosemary is yet to be elucidated; however, it may act as a protective by scavenging free radicals triggered by radiation.

Glutathione (GSH) is one of the antioxidant enzymes that act as the first line of defense against pro-oxidant stress, thus performing as a free radical scavenger. Oral administration of DDW or RE did not significantly influence the endogenous GSH level in blood. In the present study, GSH levels were found to be lower in the blood of irradiated alone animals than that observed in the RE pre-treated mice. The levels of GSH were found to be elevated in the blood of mice after RE administration.

One of the mechanisms of RE protection against radiation can be an elevation in the glutathione level that is mediated through the modulation of cellular antioxidant level. Rosmarinic acid has been experimentally found to have a significant antioxidant role through free radical scavenging activity (1). Kilic et al. observed that lipid peroxidation starts as soon as the endogenous GSH gets exhausted, and the addition of GSH stops further peroxidation promptly (20). Increase in the GSH concentration, towards normal, could have resulted in reduced levels of LPx, thereby protecting against damage caused by radiation in the RE pre-treated irradiated group.

The mechanism of the radioprotective action of Rosmarinus officinalis leaf extract in this animal model may thus be its free radical scavenging activity and its ability to thus protect cellular molecules from oxidative damage. Furthermore, it inhibited lipid peroxidation and modulated GSH levels in blood of these Swiss albino mice. The activity of rosemary may also be attributed to stimulating or protecting hematopoiesis in bone marrow and a subsequent increase of hematological constituents in the peripheral blood. Since significant protection was obtained at a non-toxic low dose, RE may have an advantage over the known radioprotectors. Further investigations are in progress to study the exact mechanism of action and clinical applicability of R. officinalis in radioprotection.

 

Table 1: Variation in differential leucocyte counts (DLC) of 3 Gy irradiated Swiss albino mice at various autopsy intervals

Post-Irradiation Interval

Treatment Group

Lymphocytes

(%)

Monocytes

(%)

Eosinophils

(%)

Basophils

(%)

Neutrophils

(%)

 

Day 1

Irradiation Control

48.4±0.10c

2.2±0.50b

1.4±0.66

0.8±0.33

47.2±0.79c

RE Experimental

54.8±3.16c

2.6±0.82

2.0±0.21

0.6±0.35

40.0±1.50c

 

Day 3

Irradiation Control

52.0±1.56c

2.0±0.44c

1.8±0.52

0.6±0.21

43.6±0.91c

RE Experimental

57.8±1.28b

2.4±0.35

1.6±0.59

0.4±0.17

37.8±1.88c

 

Day 5

Irradiation Control

58.2±1.38b

1.6±0.92b

2.4±0.21

0.4±0.21

37.4±1.65c

RE Experimental

59.6±2.16

1.8±1.01

2.6±0.40

0.8±0.17

35.2±1.80

 

Day 10

Irradiation Control

58.6±1.50b

1.8±0.40b

1.8±0.53

0.4±0.33

36.4±1.30c

RE Experimental

61.6±1077

1.4±0.21

2.2±0.48

0.6±0.35

34.2±1.86c

 

Day 20

Irradiation Control

56.8±3.28c

1.6±0.45c

2.0±0.33

0.8±0.28

38.8±1.95

RE Experimental

59.2±1.92

2.4±0.77

1.6±0.17

0.6±0.33

36.2±1.62

Day 30

Irradiation Control

58.8±2.42a

1.8±0.87c

2.0±0.48

0.6±0.21

36.0±1.08c

RE Experimental

64.8±1.34

2.8±0.63

2.2±0.77

0.4±0.17

30.2±0.60

Sham-irradiated

67.2±2.16

3.2±0.40

2.8±0.17

0.8±0.21

25.4±0.52

RE alone

67.4±1.34

3.4±0.22

3.2±0.54

0.8±0.12

25.2±0.45

a’ indicates significant (p< 0.05) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

b’ indicates significant (p<0.01) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

c’ indicates significant (p<0.001) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

 

 

 

Table 2: Variation in differential leucocyte counts (DLC) of 6 Gy irradiated Swiss albino mice at various autopsy intervals

Post-Irradiation Interval

Treatment Group

Lymphocytes

(%)

Monocytes

(%)

Eosinophils

(%)

Basophils

(%)

Neutrophils

(%)

 

Day 1

Irradiation Control

51.8±1.68c

2.4±0.66

2.2±0.35a

0.2±0.33

43.4±1.03c

RE Experimental

54.4±2.18

2.2±0.35

2.6±0.66

0.6±0.17

40.2±1.80b

 

Day 3

Irradiation Control

53.8±2.77b

2.4±0.17a

1.8±0.63

0.4±0.21a

41.6±1.63c

RE Experimental

51.6±1.46

2.2±0.44

1.4±0.95

0.2±0.35

44.6±1.85

 

Day 5

Irradiation Control

58.2±0.92

1.2±0.21c

2.6±0.96

0.4±0.33

39.6±1.10c

RE Experimental

56.4±1.94

1.6±0.33

2.2±1.06

0.4±0.28

39.4±1.79

 

Day 10

Irradiation Control

50.0±3.35c

1.4±0.78a

2.2±0.33a

0.6±0.21

45.8±1.51c

RE Experimental

57.2±2.60

1.0±0.25

1.2±0.56a

0.4±0.17

40.2±0.93b

 

Day 20

Irradiation Control

54.6±2.14c

1.2±0.48b

1.8±0.45a

0.2±0.35

42.2±0.40c

RE Experimental

57.8±2.60

2.2±0.46a

2.0±0.48

0.8±0.28

37.2±0.79c

Day 30

Irradiation Control

52.2±1.66c

1.6±0.60a

2.0±0.53

0.6±0.10

43.6±0.80c

RE Experimental

63.8±1.37

1.8±0.43

2.0±1.16

0.4±0.28

31.8±1.03

Sham-irradiated

67.2±2.16

3.2±0.40

2.8±0.17

0.8±0.21

25.4±0.52

RE alone

67.4±1.34

3.4±0.22

3.2±0.54

0.8±0.12

25.2±0.45

a’ indicates significant (p< 0.05) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

b’ indicates significant (p<0.01) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

c’ indicates significant (p<0.001) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

 

 

Table 3: Variation in differential leucocyte counts (DLC) of 9 Gy irradiated Swiss albino mice at various autopsy intervals

Post-Irradiation Interval

Treatment Group

Lymphocytes

(%)

Monocytes

(%)

Eosinophils

(%)

Basophils

(%)

Neutrophils

(%)

 

Day 1

Irradiation Control

52.0±2.38

3.0±0.44

2.8±0.21

0.2±0.21

42.0±0.92

RE Experimental

54.0±1.32

2.0±0.52

2.6±0.40

0.2±0.17

41.2±1.61

 

Day 3

Irradiation Control

51.8±1.44

2.8±0.21

2.6±0.17

0.2±0.21

42.6±2.95

RE Experimental

53.8±0.96

2.0±0.63

2.0±0.25

0.4±0.43

41.8±1.13

 

Day 5

Irradiation Control

56.0±2.24

2.4±0.48

3.0±0.35

0.4±0.17

38.2±2.51

RE Experimental

58.2±1.86

2.6±0.95

1.8±0.84

0.2±0.35

37.2±1.08

 

Day 10

Irradiation Control

55.8±1.45

2.4±0.71

2.8±0.65

0.8±0.28

38.2±2.57

RE Experimental

66.4±2.56

2.2±0.45

1.4±1.12

0.6±0.33

29.4±1.68

 

Day 20

Irradiation Control

N.S.

N.S.

N.S.

N.S.

N.S.

RE Experimental

65.6±1.73

2.2±0.77

1.4±1.08

0.2±0.21

30.4±3.68

Day 30

Irradiation Control

N.S.

N.S.

N.S.

N.S.

N.S.

RE Experimental

64.2±1.16

2.0±0.71

1.2±0.82

0.4±0.17

31.4±3.83

Sham-irradiated

67.2±2.16

3.2±0.40

2.8±0.17

0.8±0.21

25.4±0.52

RE alone

67.4±1.34

3.4±0.22

3.2±0.54

0.8±0.12

25.2±0.45

N.S. = No Survival

a’ indicates significant (p< 0.05) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

b’ indicates significant (p<0.01) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

c’ indicates significant (p<0.001) difference with respect to Sham-irradiated v/s Irradiation control, as well as Irradiation control v/s RE experimental

 

Figure 2(Click to enlarge)

Fig 1: Days of survival of mice pretreated with different doses of RE and exposed to 8 Gy gamma radiation


Figure 3(Click to enlarge)

Fig 2: Variations in the body weight of mice at post-irradiation intervals treated with (experimental) or without Rosemary extract (RE) treatment


Figure 4(Click to enlarge)

Fig 3: Variations in the glutathione (GSH) and lipid peroxidation (LPx) level in peripheral blood of mice with / without Rosemary extract (RE) treatment


Figure 5(Click to enlarge)

Fig 4: Survival dose-response curve for determination of LD50/30 (Survival data collected for treatment of radiation doses and calculated by regression analysis)

 

REFERENCES

 

  1. Al-sereiti MR, KM Abu-amer and P Sen. (1999) Pharmacology of rosemary (Rosmarinus officinalis Linn.) and its therapeutic potentials. Indian Journal of Experimental Biology 37: 124-130.
  2. Ames BN, MK Shigenaga and TM Hagen. (1993) Oxidants, antioxidants, and the degenerative diseases of aging. Proceedings of the National Academy of Sciences of the United States of America 90: 7915-7922.
  3. Ammon HP and MA Wahl. (1991) Pharmacology of Curcuma longa. Planta Medica 57: 1-7.
  4. Baliga MS, GC Jagetia, P Venkatesh and JN Ullor. (2004) Radio protective effect of abana, a polyherbal drug following total body irradiation. British Journal of Radiology 77: 1027-1035.
  5. Beutler E, O Duron and BM Kellin. (1963) Improved method for the determination of blood glutathione. Journal of Laboratory and Clinical Medicine 61: 882-888.
  6. Bloom W and LO Jacobson. (1948) Some hematological effects of irradiation. Blood 3: 586.
  7. Bourke GJ, LE Daly and JC McGilvary. (1985) In Interpretation and Uses of Medical Statistics. 3rd edition. Blackwell Scientific Publication. Oxford.
  8. Casarett AP. (1968) In Radiation Biology. Prentice-Hall, Inc. Englewood Cliffs, New Jersey pp-178.
  9. Coleman CN, HB Stone, JE Moulder and TC Pellmar. (2004) Modulation of radiation injury. Science 304: 693-694.
  10. Del Baño MJ, J Castillo, O Benavente-García, J Lorente, R Martín-Gil, C Acevedo and M Alcaraz. (2006) Radioprotective-antimutagenic effects of rosemary phenolics against chromosomal damage induced in human lymphocytes by γ-rays. Journal of Agricultural and Food Chemistry 54(6): 2064 -2068
  11. Errera M and A Forssberg. (1960) In Mechanisms in Radiology. Vol. II. Academic Press. New York.
  12. Giron LM, V Freire, A Alonzo and A Caceres. (1991) Ethnobotanical survey of the medicinal flora used by the Caribs of Guatemala. Journal of Ethnopharmacology 34: (2/3): 173-187.
  13. Grieve M. (1971) A Modern Herbal: The Medicinal, Culinary, Cosmetic and Economic Properties, Cultivation and Folk-Lore of Herbs, Grasses, Fungi, Shrubs & Trees with their Modern Scientific Uses. Dover Publications, Inc. New York ISBN: 0-486-22798-7.
  14. Hall EJ. (2000) Radiobiology for the Radiologist. 5th Edition. Lippincot, Williams and Wilkins. Philadelphia, USA.
  15. Haraguchi H, T Saito, N Okamura and A Yagi. (1995) Superoxide and lipid peroxidation were inhibited by 4 diterpenoids from rosemary: carnosic acid, carnosol, rosmanol and epirosmanol. Planta Medica 61: 333-336.
  16. Hoefler C, J Fleurentin, F Mortier, JM Pelt and J Guillemain. (1987) Journal of Ethnopharmacology 19: 133.
  17. Houlihan C, C Ho and S Chang. (1985) Journal of the American Oil Chemists’ Society 62: 96.
  18. Johnson RH. (2004) Dealing with the terror of nuclear terrorism. Health Physics 87: S3-7.
  19. Joyeux M, A Rolland, J Fleurentin, F Mortier and P Dorfman. (1990) Tert-butyl hydroperoxide-induced injury in isolated rat hepatocytes: A model for studying anti-hepatotoxic crude drugs. Planta Medica 56: 171-174.
  20. Kilic D, H Sayan, B Gonul and I Egeham. (2000) The effect of granulocyte macrophase colony stimulating factor on glutathione and lipid peroxidation in a rat model. European Journal of Surgical Oncology 26: 701-704.
  21. Kotb DF. (1985) Medicinal plants in Libya. Arab Encyclopedia House. Tripoli 720.
  22. Kumar A, J Ram, RM Samarth and M Kumar. (2003) Modulatory influence of Adhatoda Vasica Nees leaf extract against gamma irradiation in Swiss albino mice. Phytomedicine 12: 285-293.
  23. LaVerne JA. (2000) OH radicals and oxidizing products in the gamma radiolysis of water. Radiation Research 153: 196-200.
  24. Leyko W and G Bartosz. (1986) Membrane effects of ionizing radiation and hyperthermia. International Journal of Radiation Biology and Related Studies in Physics Chemistry and Medicine 49: 743-770.
  25. Mettler FAJ and GL Volez. (2002) Major radiation exposure- What to expect and how to respond. New England Journal of Medicine 346: 1554 -1561.
  26. Nachtwey DS, EJ Ainsworth and GF Leong. (1967) Recovery from radiation injury in swine as evaluated by split dose technique. Radiation Research 31: 353.
  27. Noda Y, PL McGeer, EG McGeer and M Comporti. (1993) Lipid peroxidation: Biopathological significance. Molecular Aspects of Medicine 14: 199-207.
  28. Nunia V and PK Goyal. (2004) Prevention of gamma radiation induced anaemia in mice by diltiazem. Journal of Radiation Research 45: 11-17.
  29. Offert EA, JC Gauthier, O Avanti, C Scaletta, F Runge, K Kramer and LA Applegate. (2002) Photoprotective potential of lycopene, beta-carotene, vitamin E, vitamin C and carnosic acid in UVA-irradiated human skin fibroblasts. Free Radical Biology and Medicine 32(12): 1293-1303.
  30. Ohkhawa H, N Ohishi and K Yogi. (1979) Assay for lipid peroxidation in animal tissue by thiobarbituric acid reaction. Analytical Biochemistry 95: 351.
  31. Patt HM, EB Tyree, RL Straube and DE Smith. (1949) Cysteine protection against X-irradiation. Science 110: 213-214.
  32. Saito Y, A Shiga, Y Yoshida, T Furuhashi, Y Fujita and E Niki. (2004) Effects of a novel gaseous antioxidative system containing rosemary extract on the oxidation induced by nitrogen dioxide and ultraviolet radiation. Bioscience, Biotechnology and Biochemistry 68(4): 781-786.
  33. Sarkar FH and Y Li. (2004) The role of isoflavones in cancer chemoprevention. Frontiers in Bioscience 9: 2714-2724.
  34. Scarterzzimi P and E Speroni. (2000) Review of some plants of Indian traditional medicine with antioxidant activity. Journal of Ethnopharmacology 71: 23- 43.
  35. Schuler P. (1990) Natural antioxidants exploited commercially. In Food Antioxidants. Ed. BJF Hudson. Elsevier Applied Science. London. Chapter-4.
  36. Selmi G. (1967) Therapeutic use of rosemary through the centuries. Policlinico (Prat) 74: 439-441.
  37. Sotelo-Felix JI, D Martinez-Fong and P Muriel De la Torre. (2002) Protective effect of carnosol on CCl (4)-induced acute liver damage in rats. European Journal of Gastroenterology and Hepatology 14(9): 1001-1006.
  38. Stone HB, JE Moulder, Coleman, et al. (2003) Models for evaluating agents intended for the prophylaxis, mitigation and treatment of radiation injuries. Report of an NCI workshop, Dec 3-4. Radiation Research 162: 711-728.
  39. Valentine WN and ML Pearse. (1952) Studies on the radiosensitivity of bone marrow. I. The relative sensitivity of erythroid and myeloid elements. Blood 7: 1.
  40. Waselenko JK, MacVittie, WF Blakely, et al. (2004) Medical management of the acute radiation syndrome: Recommendations of the strategic National Stockpile Radiation Working group. Annals of Internal Medicine 140: 1037- 1051.
  41. Weiss JF and MR Landauer. (2003) Protection against ionizing radiation by antioxidant nutrients and phytochemicals. Toxicology 189: 1-20.
  42. Wuensche, HW. (1938) Fortlaujende untersuchungen uner dis Eingllusses der Roentgenstrahlen auf das knochenmark. Archive of Experimental Pathology and Pharmacology 189: 581.
  43. Wu JW, M Lee, C Ho and S Chang. (1982) Journal of the American Oil Chemists’ Society 59: 339.
  44. Zimmermann V. (1980) Rosemary as a medicinal plant and wonder-drug. A report on the medieval drug monographs. Sudhoffs Arch Z Wissenschaftsgesch 64: 351-370.
  45. Zhao BL, XJ Li, RG He, SJ Cheng and WJ Xin. (1989) Cell Biophysics 14: 175.

 


View this Article by:
Print PDF | Discussion (0 comments)

Add Comment

 Copyright © 2014 Trees for Life Journal
 All trademarks and copyrights on this page are owned by their respective owners.

Powered By Geeklog