|Year : 2019 | Volume
| Issue : 1 | Page : 63-68
Evaluation of the wound healing effects of Jatropha curcas latex on intact and wounded skin in wistar rats
Hilary Oguekhian Elimian, Gerald Ikechi Eze
Department of Anatomy, University of Benin, Benin City, Edo State, Nigeria
|Date of Submission||02-Oct-2018|
|Date of Decision||09-Mar-2019|
|Date of Acceptance||12-Mar-2019|
|Date of Web Publication||28-Nov-2019|
Dr. Hilary Oguekhian Elimian
Department of Anatomy, University of Benin, Benin City, Edo State
Source of Support: None, Conflict of Interest: None
BACKGROUND: Wounds have posed a major challenge to human. It has been a major cause of morbidity and mortality. One of the herbs commonly used among the Edo people to treat wound is Jatropha curcas. It is used for the treatment of superficial wound, to secure hemostasis on wounded skin, as purgative, soap, and for making hedge around farm. It is a species of flowering plant in the genus Jatropha in the spurge family Euphorbiaceae.
AIM: This study aimed at evaluating the effects of J. curcas sap on normal and wounded skin of rats. The objectives were to investigate the wound-healing effects of J. curcas in Wistar rats as well as its effects on intact skin.
MATERIALS AND METHODS: This study involved the use of 24 Wistar rats which were placed in four Groups of six Wistar rats each (labeled A, B, C, and D). Group A had 3-cm long incision inflicted on the back of the Wistar rats and was treated with the latex of J. curcas. This was compared with Group B that had the same size of wound and was treated with 0.9% normal saline. Group C had the crude sap of J. curcas applied on intact skin. Group D had no wound and nothing applied to the skin. The animals were sacrificed after 28 days, and the histology of the normal and wounded skin was examined.
RESULTS: The J. curcas formed blood clots within 5 s of application unlike that treated with normal saline. The histological examination showed that the healing with J. curcas was greater with the re-establishment of the skin layer. The unwounded skin treated with J. curcas latex showed exfoliation of the stratum corneum.
CONCLUSION: This study has shown that the latex of Jatropha curcas has haemostatic properties. There was greater wound healing effects when compared to normal saline.
Keywords: Incision, Jatropha curcas, wound, wound healing
|How to cite this article:|
Elimian HO, Eze GI. Evaluation of the wound healing effects of Jatropha curcas latex on intact and wounded skin in wistar rats. J Exp Clin Anat 2019;18:63-8
|How to cite this URL:|
Elimian HO, Eze GI. Evaluation of the wound healing effects of Jatropha curcas latex on intact and wounded skin in wistar rats. J Exp Clin Anat [serial online] 2019 [cited 2020 Feb 20];18:63-8. Available from: http://www.jecajournal.org/text.asp?2019/18/1/63/271865
| Introduction|| |
Wound healing refers to the process by which damaged tissues of the body are repaired by living tissues. The earliest account of wound healing dated back to about 2000 BC, when the Sumerians employed two modes of treatment which consisted of spiritual method of using incantation and physical method (Barbul et al., 2015). Over the year, researches on wound healing have revolutionized modes of wound care. Various herb and natural products have been used for wound care (Oliver-Bever, 1986). One of the plants used in the traditional setting for various medicinal purposes and wound care is Jatropha curcas (Hikwa, 1995; Makkar et al., 1997).
J. curcas is a species of flowering plant in the genus Jatropha in the spurge family Euphorbiaceae (Janick and Paull, 2008). J. curcas is a source of phytochemicals with varying biological activities both in vitro and in vivo. The plant contains alkaloids, phorbol, tannins, phytates, saponins, esters, lignans, cyclic peptides, rhamnose, fucose, arabinose, xylose, mannose, galactose, glucose, glucuronic acid, galacturonic acid, and terpenes (Devappa et al., 2011). Among the terpenes, diterpenes have shown a wide range of biological activities, for example, phorbol esters which produce multiple biological effects. The phorbol esters has has skin irritant effect as well as tumour promoting effects. it also has anti-inflammatory, molluscicidal, insecticidal, and fungicidal activities.(Makkar and Becker, 2009; Devappa et al., 2010 a, b; Devappa et al., 2011).
From experiments done, the seed oil of J. curcas was shown not to be effective in experimental wound healing when compared to control, but microcurrent application alone or combined with the oil extract of J. curcas exerts significant difference in the wound healing of Wistar rats. These positive results were due to microcurrent stimulation. A significant decrease in the epithelization period was observed (Shetty et al., 2006). A mixture of extract from the leaf and back of J. curcas was shown to accelerate wound healing in rats by increasing the skin breaking strength, wound contraction dry granulation tissue weight, and hydroxyproline levels (Shetty et al., 2006). J. curcas seed oil was alone shown not to be effective on experimental wound healing.
Topical application of methanol extract of J. curcas incorporated into petroleum jelly ointment base, on excised wound in rats, was shown to significantly (P < 0.05) cause a higher rate of wound healing and reduced epithelization period (Esimone et al., 2009). The histopathological examination of granulation tissue showed much advanced phase of healing, with more collagen, which has organized to form bundles (Shetty et al. 2006).
The seed oil of J. curcas was found to contain skin tumor promoters in a two-stage mouse carcinogenesis experiment. The “irritant fraction” present in the methanol extract of the seed oil when partially purified induced ornithine decarboxylase in mouse skin and inhibited the specific binding of 3H-12-O-tetradecanoylphorbol-13-acetate (TPA) to a particulate fraction of mouse skin. After initiation with 7,12-dimethylbenz[a] anthracene, this “irritant fraction” induced tumor in the skin of 36% of the mice tested in 30 weeks (Horiuchi et al., 1987).
Roots of J. curcas were undertaken for pharmacognostic studies and evaluation of antidiarrheal activity in albino mice. The methanol fraction after successive extraction showed activity against castor oil induced diarrhea and intraluminal accumulation of fluid. It also reduced gastrointestinal motility after charcoal meal administration in albino mice. The results indicated that the action of J. curcas root methanol extract could be through a combination of inhibition of elevated prostaglandin biosynthesis and reduced propulsive movement of the small intestine (Mujumdar et al., 2000).
Bacteriological and parasitologic tests were carried out on laboratory bench surfaces using the sap and crushed leaves of J. curcas. Observation showed that the sap exhibited germicidal actions on the growth of common bacteria of Staphylococcus, Bacillus, and Micrococcus species on contact and retained such effects on treated laboratory bench surface for close to 6 h after initial application (Fagbenro-Beyioku and Oyibo, 1998; Ahmed and Adam, 1974). Hydroalcoholic extract of the leaves, stem, and root of J. curcas had showed significant antioxidant activity using in vitro antioxidant models such as nitric oxide radical scavenging activity, hydroxyl radical scavenging activity, reducing power method, and hydrogen peroxide radical scavenging activity (Diwani et al., 2009).
The latex of this plant is used by Edo people (especially farmers) to secure hemostasis in superficial wound. It is believed to stop bleeding and aid wound healing; hence, the need for this study is to establish the wound-healing effects of the latex of J. curcas and its possible effects on intact skin.
| Materials and Methods|| |
Surgical blade, dissecting set, formalin, alcohol, xylene, hematoxylin, slide, microscope, and measuring rule were used in the study.
This was a prospective study which involved the use of 24 Wistar rats. These rats were placed in four groups of six rats each. The groups were labeled A, B, C, and D. The rats in Groups A and B had a 3-cm wound inflicted on their back and was treated with J. curcas latex and normal saline, respectively. The wounds were examined every 72 h to check for the physical changes in the wounds. This included the nature of the ulcer, the floor, edge, base, and the surrounding skin and the dimensions of the ulcer in two axes – longitudinal and transverse using a calibrated plastic rule. Group C had the skin of the back shaven and the latex of J. curcas. Group D had no wound on them. The study spanned through a period of 4 weeks. The portion of the skin treated was harvested and examined histologically.
This study was done in the animal house of the Anatomy Department of the University of Benin, Benin City. Consent was obtained from the Ethics and Research Committee of the College of Medicine, University of Benin, Benin City. The reference number of the ethical approval is CMS/PO/109/Vol.I/205. J. curcas was identified at the Botany Department of the University of Benin.
Twenty-four Wistar rats were procured from the animal farm of the University of Benin. The animals were placed in four Groups, with each group consisting of six Wister rats (one rat/cage). These groups were labeled A, B, C, and D.
The sap of J. curcas was obtained by depilation of the leaves, and the latex was allowed to flow freely into a sterile universal bottle. The sterile bottle with the latex in situ was kept in an ice pack and transported to the animal house for use. The latex was allowed to warm to normal room temperature before use.
Group A – The group consisted of six Wistar rats with wound inflicted on their back. This involved the creation of a surgical wound at the back of the rat. The rats were anesthetized with chloroform. The skin at the back of the rats was prepared by first shaving the hair. The skin was then cleaned with chlorhexidine and then 70% alcohol. After which the rat was placed on a clean board which was also cleaned with chlorhexidine and 70% alcohol, an incision was made at the back of the rat which was 3-cm long by cutting through the full depth of the skin and subcutaneous tissues. This group had their wound treated with the latex of J. curcas, every 48 h by applying it topically on the wound.
Group B – This group consisted of six Wistar rats. This involved the creation of a surgical wound at the back of the rat. The rat was anesthetized with chloroform. The skin at the back of the rats was prepared by first shaving the hair. The skin was then cleaned with chlorhexidine and then 70% alcohol. After which the rat was placed on a clean board which was also cleaned with chlorhexidine and 70% alcohol, an incision was made at the back of the rat which was 3-cm long by cutting through the full depth of the skin and subcutaneous tissues. These wounds were treated with 0.9% normal saline, every 48 h by applying it topically on the wound.
Group C – This group consisted of six Wistar rats with a portion of their skin over the back shaven and delineated with indelible ink. The latex of J. curcas was applied on the skin of the rat once every 48 h for 4 weeks, to the shaven portion of the back.
Group D – this group is the control consisting of six Wistar rats. This group has no wound on their skin.
These rats were kept in separate cages (one rat/cage). They were fed with growers mash (top feeds).
The wounds were examined 72 hourly, and the dimensions were measured using a tape rule. The length and breadth of the wounds were measured in millimeters.
The experimental animals were all anesthetized and sacrificed after 28 days using vaporized chloroform. The portion of the skin treated with J. curcas and normal saline was excised. A portion of normal skin from the back of the control group was excised.
The tissues were fixed and stained using the routine hematoxylin and eosin method of staining.
| Results|| |
Following the application of J. curcas latex to the wounds, there was immediate formation of clot over the wounds within 5 s, and hemostasis was secured [Plate 1]. Three days after, the ulcer treated with J. curcas was observed to have a brownish dry scab on the floor of the wound [Plate 2]. The wounds had sloppy edges. The wound edges were contracting progressively.
The wounds treated with 0.9% normal saline clotted after 2 min [Plate 3]. On the 3rd day, the wound was noticed to have a pinkish scab on the floor [Plate 4]. The edges were sloppy and the wounds were contracting. The dimension of the wound decreased periodically until the wounds were fully healed macroscopically.
The sizes of the wounds for both Groups A and B decreased progressively. Complete healing was first noticed on the 22nd day from both the groups. On the 23rd day, the wounds were fully healed [Figure 1].
There was no gross abnormality observed on the part of the intact skin treated with J. curcas (Group C). No anomaly was observed in the skin of the control group (Group D).
[Plate 5] and [Plate 6] showed the skin of the control group. The epidermis and dermis were normal. The epidermis composed of stratified squamous epithelium keratinized. The hair follicles and the sebaceous glands were normal.
[Plate 7] and [Plate 8] showed wounded rat skin treated with J. curcas latex. The epidermis and dermis were normal with exfoliation of the stratum corneum.
[Plate 9] showed wounded rat skin treated with 0.9% normal saline solution. There was a distortion in the architecture of the layers of the skin. There are patchy interruptions in the epidermis extending to the dermis.
[Plate 10] showed unwounded skin treated with J. curcas with normal dermis. There is exfoliation of the stratum corneum layer of the epidermis.
| Discussion|| |
Wound healing is the process by which damaged parts of the body are repaired by living tissues. Attempts have been made by human over the years to use several modalities and plant extract to improve and facilitate the process of wound healing. One of the commonly used plants is J. curcas. The whole latex of J. curcas reduced the clotting time of human blood (Osoniyi and Onajobi, 2003). This shows that the latex of J. curcas has coagulant properties when the whole latex is used. This corroborated the results of this study, in which there was clot formation over the wounds treated with J. curcas within 5 s of application of the latex, as shown in [Plate 1]. This effect may be due to the acceleration of the clotting process by some of the constituents of the latex.
The bark extract of J. curcas accelerates the wound-healing process by increasing the skin breaking strength, granulation tissue breaking strength, wound contraction, dry granulation tissue weight, and hydroxyproline levels (Shetty et al., 2006). It also increases the rate of epithelization, and the collagens were better arranged in bundles (Shetty et al., 2006). This supported the results of the experimental group treated with J. curcas. There was a significant contraction of the wound edges, as shown on [Plate 2]. The histological examination of the skin treated with J. curcas showed re-epithelialization of the skin of the rats [Plate 7] and [Plate 8]. The layers of the skin were re-established unlike in the control group that had patchy interruptions of the skin layers [Plate 9]. This suggested that the latex aided the process of wound healing in these rats.
The seed oil of J. curcas has been found to induce tumor in the skin of 36% of mice after 30 weeks of application of the seed oil, in a two-stage mouse carcinogenesis experiment. The irritant fraction was the methanol extract (Horiuchi et al., 1987). This shows that prolonged use of this seed oil has adverse effects. This supports the histological findings of the experimental Group C (shaven skin treated with J. curcas), in which there was exfoliation of the stratum corneum. This suggests that prolonged use of the latex can act as a skin irritant.
| Conclusion|| |
This study has shown that the latex of J. curcas has hemostatic properties. It forms clot over wound within 5 s of application. It causes significant wound contraction. Histologically, greater wound healing effects were observed. Topical application of the latex on intact skin resulted in exfoliation of the stratum corneum in Wistar rats. The experiment has shown that J. curcas latex has a good hemostatic function, and its effect on wound healing following topical application is excellent.
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