Cytotoxic activity of Hypericum perforatum L. on K562 erythroleukemic cells: differential effects between methanolic extract and hypericin

The influence of a methanolic extract of Hypericum perforatum L. and of purified hypericin has been comparatively tested on the growth of a human erythroleukemic cell line (K562). After 1 h exposure to increasing concentrations (as hypericin content) of both agents in the dark, leukemic cells were grown for 24 h and 48 h. The effects on cell growth were determined by viable cell count, flow cytometry analysis and fluorescence microscopy. Our data show that purified hypericin has only a weak inhibitory effect on cell growth and no effect in inducing apoptotic cell death. In contrast, the Hypericum flower extract shows a significant concentration-dependent and long-lasting inhibition of cell growth, and induces apoptotic cell death. This work con fi rms the interesting role of Hypericum perforatum L. in cancer therapy and strongly supports the hypothesis that agents, other than hypericin, present in the total extract can impair tumor cell growth acting separately or in a combined manner.     

Anthocyanins induce cell cycle perturbations and apoptosis in different human cell lines

To investigate the mechanistic basis for the biological properties of anthocyanins, two aglycone anthocyanins [delphinidin (DY) and cyanidin (CY)] were used to examine their effects on cell cycle progression and on induction of apoptosis in human cancer cells (uterine carcinoma and colon adenocarcinoma cells) and in normal human fibroblasts. These compounds differ in the number and position of hydroxyl groups on the beta ring in the molecular structure. Cellular uptake of anthocyanins was confirmed by HPLC analysis and no metabolites were detected. The clonogenic assay showed that CY induces a dose-dependent growth inhibitory effect only in fibroblasts. This effect was confirmed by flow cytometric analysis, showing a significant reduction of cells in S phase. In contrast, DP inhibited cell growth in normal and tumour cell lines. This event is accompanied in fibroblasts by an accumulation of cells in the S phase suggesting a block in the transition from S to G2 phase. On the other hand, in tumour cell lines we observed a reduction of cells in G1 phase, paralleled by the appearance of a fraction of cells with a hypodiploid DNA content, thus demonstrating an apoptotic effect by DP. The occurrence of apoptosis induced by DP was confirmed by morphological and biochemical features, including nuclear condensation and fragmentation, annexin V staining, DNA laddering and poly(ADP-ribose) polymerase-1-proteolysis. Furthermore, the mitochondrial membrane potential of apoptotic cells after treatment with DP was significantly lost. The different effects exerted by DP as compared with CY suggest that the presence of the three hydroxyl groups on the beta ring in the molecular structure of DP may be important for its greater biological activity.     

Ursodeoxycholic acid does not affect ethinylestradiol bioavailability in women taking oral contraceptives

Objective Contraception is recommended for female patients during ursodeoxycholic acid (UDCA) treatment for the potential teratogenic effect of this bile acid, and the aim of our study was to determine whether this treatment affects the bioavailability of ethinylestradiol (EE₂).Methods In this double-blind, randomised study, we measured EE₂ pharmacokinetics in eight healthy volunteers randomly allocated to receive oral contraceptive (30 g EE₂ and 75 g gestodene) plus either UDCA (8–10 mg/kg per day) or placebo for 21 days during the first of three consecutive menstrual cycles. After a washout period during the second cycle, the subjects received the alternative treatment during the third menstrual cycle. Serum EE₂ and UDCA were measured using radioimmunoassay and gas chromatography–mass spectrometry, respectively. Results The profile for serum EE₂ concentration was similar during UDCA (mean maximum serum concentration 177 pg/ml, SEM 59) and during placebo treatment (153 pg/ml, SEM 62), and mean area under the curve (AUC) was 1374 pg/h per ml (SEM 580) and 1320 pg/h per ml (SEM 551) during the two regimens, respectively. The point estimates and 90% confidence intervals of UDCA/placebo ratios for EE₂ AUC and for maximum serum concentration were 1.1 (0.8–1.5) and 1.2 (1.0–1.4), respectively. Mean serum triglycerides concentration increased from 58.3 mg/dl (SEM 6.8) at enrolment to 91.4 mg/dl (SEM 10.7) during placebo (P<0.01) and to 88.6 mg/dl (SEM 13.7) during UDCA treatment (P<0.05). During UDCA treatment, serum enrichment with this bile acid and with the metabolite iso-UDCA was 29% (16%) and 3% (2%), respectively.Conclusion Co-administration with UDCA does not affect the bioavailability of EE₂ in healthy volunteers, indicating that contraceptive efficacy is not affected.     

Acupuncture in the prophylactic treatment of migraine without aura: a comparison with flunarizine

OBJECTIVES: In a randomized controlled trial extending over 6 months, we evaluated the effectiveness of acupuncture versus flunarizine in the prophylactic treatment of migraine without aura. METHODS: One hundred sixty women with migraines were randomly assigned to acupuncture treatment (group A, n = 80) or to an oral therapy with flunarizine (group F, n = 80). In group A, acupuncture was carried out in weekly sessions for the first 2 months and then once a month for the next 4 months. The same acupoints were used at each treatment: LR3 Taichong, SP6 Sanyinjiao, ST36 Zusanli, CV12 Zhongwan, LI4 Hegu, PC6 Neiguan, GB20 Fengchi, GB14 Yangbai, EX-HN5 Taiyang, GV20 Baihui. In group F, 10 mg flunarizine were given daily for the first 2 months and then for 20 days per month for the next 4 months. RESULTS: The frequency of attacks and use of symptomatic drugs significantly decreased during treatment in both groups. The number of attacks after 2 and 4 months of therapy was significantly lower in group A than in group F, and analgesic consumption was significantly lower in group A at 2 months of treatment. At 6 months no such differences existed between the two treatment groups. Pain intensity was significantly reduced only by acupuncture treatment. Side effects were significantly less frequent in group A. CONCLUSIONS: Acupuncture proved to be adequate for migraine prophylaxis. Relative to flunarizine, acupuncture treatment exhibited greater effectiveness in the first months of therapy and superior tolerability.     

An inter- and intra-laboratory comparison of breath ¹³CO₂analysis

BACKGROUND: 13C breath test analysis requires accurate 13CO2 measurements. AIM:: To perform a multicentre study to evaluate the repeatability and reproducibility of breath 13CO2 analysis. METHODS: Two series of 25 paired randomly coded tubes (each consisting of 23 13CO2-enriched breath samples and two samples of standard reference pure CO2 with certified delta 13CPDB) were sent to participating centres for 13CO2 measurement. Each series of tubes was analysed 10 days apart. The repeatability and reproducibility of 13C measurements was assessed by Mandel's k and h statistics. RESULTS: Twenty-two centres participated in the study: 18 showed good inter- and intra-laboratory variability, whilst four showed abnormally high inter- or intra-laboratory variability. Breath test results were also significantly affected by the accuracy of the 13C analytical procedures. CONCLUSIONS: A low accuracy of 13C measurements may significantly affect the results of breath tests, leading to inappropriate clinical decisions. Standardization of 13C analysis is required to guarantee optimal 13C measurements and accurate 13C breath test results.     

Combined diagnostic imaging of adenomyomatosis of the gallbladder

The consistency of a request for liver MRI and MR-cholangiopancreatography in a patient with an occasional US-finding of mural thickening of gallbladder fundus > or = 3 mm in size was assessed. The differential US diagnosis in case of focal mural thickening was analyzed in combination with the other findings to justify the radiologicoclinical reliability of the request. Overall, these findings suggested the diagnostic hypothesis of adenomyomatosis, then confirmed by a second level in-depth examination, namely liver MRI and MR-cholangiopancreatography.