Detection and isolation of intestinal muscle relaxant substances from the root of Taverniera abyssinica A. Rich.

ETHNOPHARMACOLOGICAL RELEVANCE: In Ethiopian traditional medicine the root of Taverniera abyssinica A.Rich is known as a remedy for sudden gastrointestinal cramping and fever. In this study we have isolated and identified the bioactive principle of Taverniera abyssinica that exerts effects on isolated smooth muscle tissues of the rabbit duodenum and guinea-pig ileum. AIM OF THE STUDY: To isolate and purify the bioactive principle from the root of Taverniera abyssinica A.Rich by bioassay-guided fractionation, HPLC purification and masspectrometry, with further investigation of its bioactivity on isolated smooth muscle strips. MATERIALS AND METHODS: Roots of Taverniera abyssinica A.Rich extracted in 75% methanol/water were fractioned with a reverse phase column and then subjected to HPLC purification. Each fraction collected from the HPLC was tested for its bioactivity using electric field stimulation-evoked contractions of the rabbit duodenum and guinea-pig ileum. Finally, detailed structural analysis of the fraction displaying significant bioactivity was made by mass spectrometry. RESULTS: Through bioassay-guided fractionation and HPLC purification the bioactive fractions were identified. These were tested for bioactivity on isolated smooth muscle strips which showed about 80% inhibition of contractions evoked by electric field stimulation. These compounds were identified as formononetin, afrormosin and tectorigenin by using masspectrometry applying relevant standards for detection. CONCLUSION: The traditionally claimed smooth muscle-relaxing effect of the roots of Taverniera abyssinica A.Rich is essentially due the three isolated and purified the two isoflavones formononetin, afrormosin as well as the metoxyisoflavone tectorigenin, along with possibly other not yet purified bioactive substances, however with similar smooth muscle-relaxing properties.

Red blood cell folate level and associated factors of folate insufficiency among pregnant women attending antenatal care during their first trimester of pregnancy in Addis Ababa, Ethiopia.

Objective: Folate is an essential vitamin for de novo DNA synthesis and cell proliferation. Folate insufficiency at the time of conception and during the first trimester of pregnancy is associated with unintended pregnancy and birth outcomes, particularly neural tube defects. Hence, this study aimed to assess folate status and associated factors of folate insufficiency among pregnant women attending antenatal care during their first trimester of pregnancy in Addis Ababa, Ethiopia. Materials and methods: A cross-sectional study was conducted from 8 August 2017 to 3 January 2018 in Addis Ababa. In this study, 160 participants were enrolled via the convenience sampling method. Red blood cell folate was measured by the electrochemiluminescence binding assay method. Data were entered into Epi-Data version 3.1 and analyzed by SPSS version 22.0. Descriptive statistics were used to describe demographic characteristics and to determine the magnitude of folate deficiency. Logistic regression was used to identify the risk factors for folate deficiency. A p-value of less than 0.05 was considered statistically significant. Results: In this study, 44/160 (27%) participants had red blood cell folate level <400 ng/mL, insufficient to prevent neural tube defect. Multivariate regression showed that regular vegetable consumption was an independent determinant factor for red blood cell folate level (adjusted odds ratio: 0.41, confidence interval: 0.18-0.93). Conclusion: This study shows that a large magnitude of the first-trimester pregnant women had red blood cell folate concentrations below levels that are maximally protective against neural tube defects. Folic acid supplementation and supplemental nutrition containing green leafy vegetables should be promoted during the periconceptional period. In addition, the policymakers should set rules for mandatory folic acid fortification.

Folate, Vitamin B12, and Homocysteine Levels in Women With Neural Tube Defect-Affected Pregnancy in Addis Ababa, Ethiopia.

Background: Neural tube defects (NTDs) are prevalent congenital defects associated with pre-pregnancy diet with low levels of maternal folate. They are linked to severe morbidity, disability, and mortality, as well as psychological and economic burdens. Objective: The goal of this study was to determine the levels of folate, vitamin B12, and homocysteine in the blood of women who had a pregnancy impacted by NTDs. Subjects and Methods: A hospital-based case-control study was undertaken between September 2019 and August 2020. The study comprised a total of 100 cases and 167 controls. Enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of folate, vitamin B12, and homocysteine in the serum. Results: Only 39% of the cases and 54.5% of control mothers reported periconceptional use of folic acid/multivitamin, which indicated a statistically significant difference (p = 0.014). Logistic regression indicated that periconceptional use of folic acid/multivitamin was associated with NTDs (p = 0.015, OR = 1.873, 95% CI: 1.131-3.101). We found that 57% of the cases and 33.5% of controls, as well as 43% of cases and 20.4% of controls had serum folate and vitamin B12 levels below the cut-off value, respectively. Twenty-seven percent of the cases and 6.6% of controls had hyperhomocysteinemia (HHcy). The median concentrations of folate, vitamin B12, and homocysteine in cases and controls were 4.78 and 8.86 ng/ml; 266.23 and 455 pg/ml; 13.43 and 9.7 µmol/l, respectively. The median concentration of folate (p < 0.001) and vitamin B12 (p < 0.001) were significantly lower in the cases than controls, while the homocysteine concentration (p < 0.001) was significantly lower in the controls than cases. Folate [OR (95% CI) = 1.652 (1.226-2.225; p = 0.001)], vitamin B12 [OR (95% CI) = 1.890 (1.393-2.565; p < 0.001], and homocysteine [OR (95% CI) = 0.191 (0.09-0.405; p < 0.001)] levels were associated with NTDs. Conclusion: Folate and vitamin B12 are deficient in both cases and control mothers. The lower levels of folate and vitamin B12 with an elevated homocysteine level in NTD-affected pregnancy may be an indication that these biochemical variables were risk factors for NTDs. Folate/multivitamin supplementation and/or food fortification should be promoted.

Cytotoxic and other bioactivities of a novel and known sesquiterpene lactones isolated from Vernonia leopoldi (Sch. Bip. ex Walp.) Vatke in breast cancer cell lines.

Vernonia leopoldi (Sch. Bip. ex Walp.) Vatke (Asteraceae) is one of the widely used anti-cancer traditional medicinal plants in Ethiopia, despite the lack of data to support its therapeutic efficacy. Here we describe the isolation of compounds from the plant and the investigation of their cytotoxicity and other bioactivities. We identified the novel sesquiterpene lactone (SL) 11ß,13-dihydrovernodalol along with the three other SLs (vernomenin, vernolepin, and 11ß,13-dihydrovernodalin) and three flavonoids (apigenin, eriodyctiol, and luteolin) isolated from this plant for the first time. The structures of all the compounds were established based on extensive analysis of nuclear magnetic resonance spectroscopic data and confirmed by high-resolution electrospray ionization mass spectrometry. We then studied the biological activities of the SLs and found that all were cytotoxic at low µM ranges against MCF-7 and JIMT-1 breast cancer cells as well as against the normal-like MCF-10A breast epithelial cells evaluated in a spectrophotometric assay. All the SLs significantly reduced JIMT-1 cell migration after 72 h of treatment with 2 µM concentrations in a wound healing assay. Treatment with all SLs reduced the aldehyde dehydrogenase expressing cancer stem cell sub-population of the JIMT-1 cells significantly, evaluated by flow cytometry. Only 11ß,13-dihydrovernodalin resulted in a significant inhibition of tumor necrosis factor-α-induced translocation of nuclear factor κB to the cell nucleus. In addition, we show that the reporter fluorophore nitrobenzoxadiazole (NBD) can successfully be conjugated with an SL and that this SL-NBD conjugate is taken up efficiently in JIMT-1 cells. Therefore, the overall bioactivities of the SL compounds and specifically their effects against the stemness of breast cancer cells make them prime candidates for further in-depth investigation.

Effect of 70% Ethanol Extract and its Solvent Fractions of Artemisia afra (Jacq. Ex Willd.) against Pentylenetetrazole-Induced Seizure in Mice.

INTRODUCTION: Artemisia afra (Jacq. ex Willd.), commonly called African wormwood, is a highly aromatic perennial herb and a well-known medicinal plant, claimed to be effective and safe in the treatment of epilepsy. The whole-plant extract is traditionally used as an antiepileptic agent in Ethiopia. Aim of the Study. The aim of this study was, therefore, to evaluate the anticonvulsant effect of the hydroethanolic extract and solvent fractions of A. afra whole part in mice. MATERIALS AND METHODS: The effects of A. afra hydroethanolic extract and its solvent fractions were evaluated against pentylenetetrazole- (PTZ-) induced convulsions in mice. The onset and duration of PTZ-induced convulsions were determined with hydroethanolic A. afra extract and its solvent fractions. Data were analyzed using a one-way analysis of variance (ANOVA) followed by post hoc Tukey's multiple comparisons test. p < 0.05 was considered statistically significant. RESULTS: The hydroethanolic extract of A. afra, with all the three doses of 250, 500, and 1000 mg/kg, showed a significant delay (504.833 ± 62.835 ∗ s; p < 0.05 ∗ ; 551.833 ± 47.69 ∗∗ s; p < 0.01 ∗∗ ; and 808.333 ± 64.8 ∗∗∗ s; p < 0.001 ∗∗∗ , respectively) in the mean onset of convulsion and a decrease (17.000 ± 1.88 ∗∗∗ s, p < 0.05 ∗ ; 13.000 ± 1.8 ∗∗ s, p < 0.01 ∗∗ ; and 7.833 ± 1.07 ∗∗∗ s, p < 0.001, respectively) in the mean duration of convulsion against PTZ-induced convulsion in a dose-dependent manner compared to the control (92.833 ± 13.006 s; 34.167 ± 3.683 s), and its anticonvulsant activity was significantly less compared to that of diazepam (1001.167 ± 68.430 s; 4.500 ± 0.619 s). The solvent fractions, however, did not show anticonvulsant activity against PTZ-induced convulsion. CONCLUSION: Crude extract of A. afra has an anticonvulsant effect in mice. This might be attributed to the synergistic effects of two or more active ingredients present in the herb.

Solvent fractions of selected Ethiopian medicinal plants used in traditional breast cancer treatment inhibit cancer stem cells in a breast cancer cell line.

BACKGROUND: The incidence and mortality of breast cancer in women is increasing worldwide. Breast cancer contains a subpopulation of cells known as cancer stem cells (CSCs). The CSCs are believed to be responsible for chemotherapeutic resistance and are also involved in tumor initiation, progression, evolution, and metastasis to distant sites. The present study aimed to investigate the anti-CSC potential of selected Ethiopian medicinal plants traditionally used for breast cancer treatment. METHODS: The solvent fractions of three medicinal plants (the ethyl acetate fraction of Vernonia leopoldi, the aqueous fraction of Sideroxylon oxyacanthum, and the chloroform fraction of Clematis simensis) resulting from the methanolic crude extracts were selected based on their previously demonstrated cytotoxic effects on breast cancer cell lines. The effect of these solvent fractions on the status of the cancer stem cell subpopulation of the JIMT-1 cell line was assessed by flow cytometric evaluation of the proportion of aldehyde dehydrogenase positive cells and by measuring colony forming efficiency in a serum-free soft agar assay after treatment. Effects on cell migration using a wound healing assay and on tumor necrosis factor-α-induced translocation of nuclear factor-kappa B to the cell nucleus were also investigated. RESULTS: The solvent fractions showed a dose-dependent reduction in the aldehyde dehydrogenase positive subpopulation of JIMT-1 cells. The chloroform fraction of C. simensis (80 µg/mL) completely blocked colony formation of JIMT-1 cells. The wound healing assay showed that all fractions significantly reduced cell migration. The ethyl acetate fraction of V. leopoldi (0.87 µg/mL) significantly inhibited tumor necrosis factor-α-induced nuclear factor-kappa B translocation to the nucleus. CONCLUSION: The solvent fractions of the medicinal plants showed desirable activities against breast cancer stem cells in the JIMT-1 cell line, which warrants further studies.

Vitamin D Status and Association of VDR Genetic Polymorphism to Risk of Breast Cancer in Ethiopia.

Emerging evidence associates vitamin D deficiency and vitamin D receptor (VDR) genetic variations with risk for breast cancer. This study investigated the prevalence of vitamin D deficiency and its association with tumor characteristics and the implications of VDR genetic variations for risk of breast cancer in Ethiopia. This unmatched case⁻control study involved 392 female breast cancer patients and 193 controls. The plasma 25-hydroxyvitamin D (25(OH)D3) level was quantified in chemotherapy-naïve (N = 112) and tamoxifen-treated patients (N = 89). Genotyping for the VDR common variant alleles rs7975232 (ApaI), rs2228570 (FokI), and rs731236 (TaqI) was done. Eighty-six percent of the patients were vitamin D deficient (<50 nmol/L). Chemotherapy-naïve breast cancer patients had a higher prevalence of vitamin D deficiency (91.9% vs. 78.3%) compared to the tamoxifen-treated group (p < 0.001). The prevalence of severe vitamin D deficiency (<25 nmol/L) was significantly higher in chemotherapy-naïve (41.1%) than tamoxifen-treated (11.2%) patients. Vitamin D deficiency was not significantly associated with tumor characteristics or VDR genotype. The rs2228570 GG genotype was associated with increased risk of breast cancer (OR = 1.44, 95% confidence interval = 1.01-2.06). Our result indicates that rs2228570 might be a moderate risk factor for breast cancer development in the Ethiopian population. The high prevalence of severe vitamin D deficiency in treatment-naïve breast cancer patients indicates the need for nutritional supplementation of vitamin D at the time of chemotherapy initiation.

Antidiabetic and gastric emptying inhibitory effect of herbal Melia azedarach leaf extract in rodent models of diabetes type 2 mellitus.

Diabetes type 2 is associated with impaired insulin production and increased insulin resistance. Treatment with antidiabetic drugs and insulin strives for normalizing glucose homeostasis. In Ethiopian traditional medicine, plant extracts of Melia azedarach are used to control diabetes mellitus and various gastrointestinal disorders. The objective of this study was to clarify the antidiabetic effects of M. azedarach leaf extracts in diabetic type 2 experimental animals. In this study, mice were injected with Melia extract intraperitoneally. Plasma glucose was studied by using tail vein sampling in acute experiments over 4 h and chronic experiments over 21 days with concurrent insulin and body weight assessments. Glucose tolerance was studied by using intraperitoneal glucose (2 mg/g) tolerance test over 120 min. Gastric emptying of a metabolically inert meal was studied by the gastric retention of a radioactive marker over 20 min. Melia extracts displayed acute, dose-dependent antidiabetic effects in ob/ob mice similar to glibenclamide (p<0.05-0.001). Long-term administration of Melia extract reduced plasma glucose (p<0.001) and insulin (p<0.01-0.001) levels over 21 days, concurrent with body weight loss. Glucose tolerance test showed reduced basal glucose levels (p<0.05-0.01), but no difference was found in glucose disposal after long-term treatment with Melia extract. In addition, the Melia extract at 400 mg/kg slowed gastric emptying rate of normal Sprague-Dawley (p<0.001) and diabetic Goto-Kakizaki rats (p<0.001) compared with controls. It is concluded that the M. azedarach leaf extract elicits diabetic activity through a multitargeted action. Primarily an increased insulin-sensitizing effect is at hand, resulting in blood glucose reduction and improved peripheral glucose disposal, but also through reduced gastric emptying and decreased insulin demand.