EVALUATION OF WOUND HEALING ACTIVITY OF Clerodendrum Infortunatum Linn. IN DIABETIC RATS.
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Date
2015
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Birsa Agricultural University, Kanke, Ranchi, Jharkhand
Abstract
The present study was undertaken to evaluate the wound healing activity
of Clerodendrum infortunatum LINN. in normal and diabetic rats. Open excision
wounds of 2×2 cm2 were experimentally created on the back region of the
normal and diabetic rats to evaluate the time dependent wound healing effects
(days 0, 3, 7, and 15) of C. infortunatum ointment (4%). This study was
conducted in two phases i.e. Phase I and Phase II. In phase I study, we evaluated
the time dependent effects of C. infortunatum ointment (4%) on cutaneous wound
healing in normal rats. In phase II study, we evaluated the time dependent
effects of C. infortunatum ointment (4%) on cutaneous wound healing in diabetic
rats.
In the present study, phytochemical tests were carried out in ethanolic
extract of C. infortunatum leaves in order to determine the presence of chemical
constituents. The tests were aimed to identify the presence of alkaloids,
carbohydrates, flavonoids, glycosides, reducing sugars, saponins, sterols,
terpenes, tannins, and proteins. Phytochemical screening showed the presence
of alkaloids, glycosides, reducing sugars, sterol, terpene in higher extent,
flavanoids were found moderately and saponin and tannin were found tracer
amount where as the carbohydrates, and proteins were absent in the extract.
The C. infortunatum (4% ointment) was applied topically on the wound
twice daily for 15 days to evaluate the time dependent wound healing effects of
C. infortunatum , as compared to ointment base applied on normal control rats.
The six animals (each time point) from each group were sacrificed on days 3, 7,
and 15 to collect the granulation tissue.
The tissue was then used for the estimation of various oxidative stress
related parameters, estimation of hydroxyproline content and for the studies of
gross histopathological changes by H & E staining, of wound sections on days 3,
7 and 15.
The salient findings of phase I trial are given below:
1. The reduction in wound area was significant on day 3,7 and 15 in C.
infortunatum ointment (4%) treated group, as compared with control in
normal rats .
2. C. infortunatum ointment (4%) treated wounds revealed significantly
higher percent wound contraction as compared to control group. It was
also revealed that the percentage of wound contraction was significantly
higher (p<0.001) in treated group on day 7, day 15 as compared to control.
However percentage of wound contraction was not significantly (p>0.05)
vary on day 3 in C. infortunatum treated in compared to control group.
3. SOD activity in the normal treated group was significantly (p<0.05)
increased on days 3, 7 post wounding in a time dependent manner as
compared to control group. C. infortunatum ointment treatment caused a
significant increased of SOD level on day 3, on day 7. However, on day
15, the level of SOD was increased in treated group but not significantly
vary as compared to control group
4. Similarly, the activity of catalase in normal treated group was
significantly (p<0.05) increased on day 7 and on day 15 as compared to
control. However, the levels of catalase were increased but not
significantly vary in normal treated group on day 3 of post wounding as
compared to control group.
5. GSH contents were found to be significantly (p<0.001) increased on day 3,
day 7 and day 15 as compared to control in a time dependent manner. C.
infortunatum ointment in treated group revealed a significantly higher
level of GSH in granulation tissue on day 3,7,15 as compared to control
group.
6. C. infortunatum ointment (4%) treatment caused significantly (p<0.001)
decreased the level Lipid peroxidation in normal treated group as
compared to control group on days 7 and 15. However, on day 3 the level
of LPO was not significantly (p>0.05) varied in treated group as
compared to control group.
7. The level of hydroxyproline were significantly (p<0.05) increased on day
3 in C. infortunatum ointment treated wounds, as compared to control rats.
After day3, the level of hydroxyproline in C. infortunatum treated group
significantly (p<0.01) increased on day 7 and day 15 as compared to
control group.
8. H & E stained sections showed that treated wounds had marked
proliferation of fibroblasts with collagen deposition, new and wellformed
capillaries in well-formed granulation tissue covered by newly
formed epithelial layer. Whereas, control groups had presence of marked
inflammatory cells until day 15.
The salient findings of phase II trial are given below:
1. There was significantly increased on wound contraction (absolute area) in
C. infortunatum ointment treated diabetic group on day 3, 7 and on day 15
as compared to diabetic control.
2. C. infortunatum ointment (4%) treated diabetic wounds revealed
significantly higher percent wound contraction as compared to diabetic
control group. It was also revealed that the percentage of wound
contraction was significantly higher (p<0.05) in diabetic treated group on
day7and day 15 as compared to diabetic control. However percentage of
wound contraction was not significantly (p>0.05) vary on day 3 in C.
infortunatum diabetic treated versus diabetic control group.
3. SOD activity in the diabetic treated group was significantly (p<0.05)
increased on days 3, 7 post wounding in a time dependent manner as
compared to diabetic control group. However, on day 3, the level of SOD
was increased in treated group but not significantly vary as compared to
control group.
4. The activity of catalase in diabetic treated group was significantly
(p<0.001) highly increased on day 3, 7 and 15 as compared to diabetic
control. The level of catalase was increased progressively on day 3, 7 and
day 15 in diabetic treated group as compared to control group.
5. GSH contents were found to be significantly (p<0.001) increased in
diabetic treated group on day 7and day 15.On day 3 GSH level were
increased non significantly in diabetic treated group as compared to
diabetic control. C. infortunatum ointment treated group revealed a
significantly higher level of GSH in granulation tissue on day 7 and day
15, however it was not significantly increased on day 3 as compared to
diabetic control group.
6. The level of LPO was decreased significantly in diabetic treated group in
comparison to diabetic control on day 3, 7 and on day 15.
7. The level of hydroxyproline were significantly (p<0.05) increased on day
3 in C. infortunatum ointment treated diabetic wounds, as compared to
diabetic control rats. After day3, the level of hydroxyproline in C.
infortunatum treated group significantly (p<0.01) increased on day 7and
day 15as compared to diabetic control group.
8. Diabetic treated group exhibited uniform and thick granulation tissue
formation with marked proliferation of fibroblasts, well-formed new
blood vessels and few inflammatory cells. However, wound area of
control group was dominated by more cells that are inflammatory and
loosely formed less granulation tissue.
CONCLUSION
C. infortunatum ointment (4%) treatment showed faster and organized
healing of cutaneous wounds in normal as well as in diabetic rats. Percentage
contraction of wound was significantly higher in treated groups as compared to
control group whereas the percentage contraction was much higher in normal
treated groups as compared to diabetic groups. Up-regulation of various antioxidative
enzymes as well as hydroxyproline content and histopathological
results revealed that there was improved healing by C. infortunatum ointment
application in treated groups. The ethanolic extract ointment (4%) of C.
infortunatum has shown great potential in treating diabetic wounds as well as
normal wound on topical application.
Description
EVALUATION OF WOUND HEALING ACTIVITY OF Clerodendrum Infortunatum Linn. IN DIABETIC RATS.
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