Introduction: the roots of Leptadenia hastata, ( L hastata) are remedies from the Senegalese pharmacopoeia and are as widely used as the leaves. However, few researchers have devoted themselves to their toxicity, unlike the leaves. However, in the traditional pharmacopoeia, the indications for use are very different. The aim of our study was to study the effect of the administration of methanolic extracts of roots of L hastata on renal tissue, using an animal model. Materials and Methods: a cohort of 18 rats was studied with a random distribution of the animals in 3 groups (n = 6). The first group was the control group. The treated groups (Group II and III) received the methanolic extract of L. hastata with an administration of 500 mg / kg / day and 1000 mg / kg / day respectively, for 28 days. Results: The dose of 1000 mg / kg / day was lethal in group III, from the first week in females. Serum creatinine was significantly higher in rats given the root extract. There was kidney damage with vacuolar degeneration starting at 500 mg / kg / day. The lesions were more severe in group III with glomerular involvement characterized by retraction in the renal corpuscles. Conclusion: If the leaves of L hastata seem to be safe for consumption according to the majority of authors in the literature, the roots of this same plant could be harmful. In addition to the lethality observed at the doses tested, the lesions in the renal parenchyma would be dose-dependent.
The leaves of Leptadenia hastata (L. hastata) are widely used in the Senegalese pharmacopoeia, and in the literature numerous studies have been reported concerning its medicinal properties [
The roots are also used as a remedy, but few toxicity studies have been reported in the literature regarding this part of the plant. The particular use of roots in sickle cell patients in traditional medicine motivated a study carried out in vitro [
Sickle cell anemia is a genetic disease prevalent in our geographic locality. There is no curative treatment and the disease is responsible for many complications, especially during pregnancy. The treatment is essentially preventive and the therapeutic properties of L. hastata in vitro on sickle cell anemia are promising [
The kidney plays a major role in eliminating metabolic waste, especially in the event of toxicity. In addition, renal failure is one of the common complications of hemoglobinopathies but can also be a secondary consequence of the toxicity induced by phytotherapy.
In order to explore in vivo the innocuity of the roots, before eventual clinical assays with patients with sickle cell anemia, we tested the plant using an animal model.
The aim of our study was therefore to study the effects on renal parenchyma, of the administration of methanolic extracts from the roots of L. hastata among a cohort of Wistar Albino rats.
This study was carried out thanks to a collaboration between the laboratory of Toxicology, the laboratory of Histology-Embryology-Cytogenetics and the laboratory of Pathology of Cheikh Anta Diop University. The experiment was approved by the Ethics Committee of Cheikh Anta Diop University of Dakar (Senegal).
Roots of L. Hastata were collected in the Bush, 18 km far from Dakar in the locality of Sangalcam (
Eighteen Wistar rats with eight to twelve weeks old and average weight of 180 - 220 g were studied. They were raised in plastic cages with wood shavings as bedding. The temperature of the room was maintained at 22˚C ± 4˚C with 12 hours of light and 12 hours of darkness. The rats had free access to food and water.
The animals were randomly divided into 3 groups (n = 6) with equal number of males and females. A stock solution of the 100 mg/mL concentration extract was prepared with 5% methanol in mineral water.
Group I the control group, received 2 ml of solution of mineral water containing methanol (10% of total volume).
Group II and III received the methanolic extract of L. hastata at doses of 500 mg/kg/j and 1000 mg/kg/j respectively for 28 days.
During this period, all the animals were observed daily for signs of toxicity and mortality. The changes in body weight, food and water intake and clinical signs were recorded according to the protocol described in the LD407 of the OECD.
After necropsy, samples from the kidneys were obtained and fixed in buffered formalin (10%) for at least 24 hours. The samples were dehydrated in graded series of ethanol, cleared in xylene, embedded in paraffin wax and sectioned at 4 - 5 μm. The slides were stained with the Hematoxylin and Eosin (H & E Stain). Changes were graded as follows: 1) showing no changes, 2) for mild changes (less than 30%), 3) moderate changes (more than 30% and less than 50%) and 4) for severe or marked changes (more than 50%), following guidelines from the INHAND Project (International Harmonization of Nomenclature and Diagnostic Criteria for Lesions in Rats and Mice) [
To analyze the slides, we used photonic microscope from Leica (Leica, microsystems, Wetzlar, Germany), DM 750 and Lasez software to capture and store the images.
On day 28, after anesthesia, blood samples were taken from the heart of the animal. Serum was obtained for biochemical analysis after centrifugation at 2500 rpm by min for 15 min. The biochemical indices such as aspartate aminotransferase (AST) and alanine aminotransferase (ALT) transaminases were analyzed. They were assessed in the laboratory of Biochemistry in Diamniadio Hospital using diagnostic kits purchased from Roche Diagnostics Ltd. (Germany).
Results of plasmatic creatinine were expressed as mean ± standard deviation (SD). The difference between the different groups was compared using test of variance (one-way ANOVA). Differences with values of p < 0.05 were considered statistically significant. Statistics were performed using GraphPad software.
The methanolic extract from the roots of L. hastata was lethal at the tested doses in male and female Wistar rats. At the dose of 1000 mg/kg/day, deaths were recorded during the first week for females and before the end of the test, all rats in group III were dead.
Creatinine
Creatinine levels were elevated in Group II. The biostatistical analysis of the biochemical parameters indicated statistically significant between the treated group and the control group. Results are presented in
Histology
Macroscopic examination
The kidneys were macroscopically normal with brownish color. After section, the parenchyma did not present any sign of particular abnormalities.
Microscopic examination
Microscopic examination of the kidney revealed vacuolar degeneration in the tubular epithelium in group II and group III and glomerulus lesions. The degree and the topography of these lesions are summarized in
| Group I Control | Group II 500 mg/kg/day | |
|---|---|---|
| Males | ||
| Creatinine (mg/L) | 7.27 ± 0.46 | 10.35 ± 2.47*** |
| Females | ||
| Creatinine (mg/L) | 7.38 ± 0.26 | 10.1 ± 2.32*** |
*** = p < 0.001 vs. control.
| Lesions | Group II 500 mg/kg/day | Group III 1000 mg/kg/day | Topography |
|---|---|---|---|
| Cytoplasmic vacuolar degeneration of tubules | 1 | 2, 3 | Cortical and medulla segment |
| Glomerulus retraction | 0 | 1, 2 | Cortical segment |
Traditional use of L Hastata roots
The roots of L. hastata have specific indications in the traditional pharmacopoeia. These medicinal indications are different from those of other parts of the plant such as the leaves or bark [
Therapeutic effect of roots on sickle cell anemia
Egunyomi [
The flavonoids which constitute the most concentrated component of the root [
The tissue damage caused by L. hastata roots
There was cytoplasmic vacuolization in the tubular epithelium with a retracted glomerulus and an enlargement of the filtration chamber. These damages were visible at the dose of 1000 mg/kg/day. At the dose of 500 mg/kg/day, vacuolization of the epithelium was present, but the rest of the parenchyma was relatively healthy.
L. hastata is reputed to be a harmless plant, according to most of studies [
An anti-androgenic effect has also been noted with a decrease in the level of androgens as well as a decrease in spermatogenesis in animal models according to Bayala [
Unlike the leaves, which have been widely studied and documented, few studies are devoted to the roots of L. hastata. The results of our study are additional information regarding the safety of this plant. The study also shows that the long-term use of L. hastata roots (from 500 m/kg/day) causes cellular damage to the kidney.
The methanolic extract of the roots of L. hastata is responsible for damage in the renal parenchyma with vacuolization of the tubular epithelium from a dose of 500 mg/kg/day. At the dose of 1000 mg/kg/day, the cytoplasmic vacuolization is more severe damage with glomerular damage.
However, the many medicinal properties of the roots of L. hastata and its use in herbal medicine for centuries in West Africa and also in animal husbandry traditions deserve further studies in toxicopathology. The potential promising pharmacological activities on sickle cell anemia are arguments justifying the determination of an effective and safe therapeutic dose for its users.
The authors declare that they have no competing interests.
This study received the approval of the ethics Committee from the faculty of Medicine of Cheikh Anta Diop University of Dakar.
Diallo, M.S., Bah, F., Dial, C.M., Diop, C., Faye, O., Seck, M. and Fall, M. (2020) Histopathological Study of the Kidney after Administration of Methanolic Extract of Roots of Leptadenia hastata in the Albino Wistar Rat. Open Journal of Pathology, 10, 67-75. https://doi.org/10.4236/ojpathology.2020.102007