Assessment of genotoxicity of Ssanghwa-tang, an herbal formula, by using bacterial reverse mutation, chromosome aberration, and in vivo micronucleus tests

Objectives: Ssanghwa-tang (SHT) is a traditional herbal formula comprising nine medicinal herbs, and it is used for reducing fatigue in Korea. SHT exerts various effects such as anti-inflammatory, antioxidant, and anti-aging activities, and protection against acute hepatotoxicity. However, the genotoxicity of SHT has not yet been established.Methods: Ten components were identified in SHT water extract by using high-performance liquid chromatography analysis. We assessed the genotoxicity of SHT by using bacterial reverse mutation (Ames test), chromosome aberration, and in vivo micronucleus tests.Results: The contents of paeoniflorin, glycyrrhizin, and liquiritin apioside in SHT were 15.57, 6.94, and 3.48 mg/g extract, respectively. SHT did not increase the revertant colonies of Salmonella typhimurium and Escherichia coli strains in the presence or absence of metabolic activity. Although SHT did not induce structurally abnormal chromosomes in Chinese hamster lung (CHL) cells in the presence of metabolic activity, the number of structurally aberrated chromosomes increased dose-dependently in the absence of metabolic activity. In the in vivo micronucleus test, SHT did not affect the formation of micronuclei compared with the vehicle control.Conclusions: Genotoxicity of SHT was not observed in the Ames test and in vivo micronucleus test. However, based on the results of chromosome aberration test, it can be presumed that SHT has the potential to induce genotoxicity because it induced structurally abnormal chromosomes in the absence of metabolic activity.

Cytotoxic and Genotoxic Effects of Methanol Extracts of Some Gypsophila L. Species

In this study, the cytotoxic and genotoxic effects of Gypsophila perfoliata L. var. perfoliata, Gypsophila perfoliata L. var. araratica Kit Tan, Gypsophila pilosa Hudson and Gypsophila osmangaziensis Ataşlar & Ocak plant extracts have been examined by using Allium Test method. Methanol extracts of plants have been prepared in 4 different concentrations (0.625 mg/ml, 1.250 mg/ml, 2.500 mg/ml and 5.000 mg/ml). After the onion roots were treated at these concentrations of plant extracts for 24 hours and 48 hours, mitosis preparations were prepared from these root tips. With the data being obtained by evaluating the preparations, mitotic index (%) and chromosome aberration (%) values have been calculated. Distilled water has been used as control group. It was found out that mitotic index and chromosome aberration values of all species showed significant differences compared to control group in the extract concentration range of 1.250–5.000 mg/ml. It has been determined that the most widely observed chromosome aberrations were disturbed metaphase, sticky metaphase, c-metaphase, disturbed anaphase and anaphase bridge.

P–572 Purifying selection for aneuploidy cells in mosaicism embryo at post-implantation stage

Study question Why low ratio mosaicism embryos develop to normal karyotype babies? Summary answer Our in vitro implantation assay clarified purifying selection for aneuploid cells in post implantation embryos. What is known already There are some reports about healthy live birth after transfer of mosaic embryos, which was reported for the first time from Italy in 2015. It is also reported that the abnormal cell is screened with the mouse in the embryo development, and only a normal cell contributes to the development. But it has not been examined in human. Study design, size, duration To clarify the change of aneuploid cells and mitochondrial activity in human embryo, we biopsied several parts from one blastocyst and examined karyotype. After in vitro implantation assay for biopsied embryos, we compared the karyotype of biopsy sample with that of cultured cell mass. Participants/materials, setting, methods Under the ethical review of Yokohama City University and informed consent with patients, we collected human surplus blastocysts those are donated after successful clinical treatment or discarded because of poor development grade. We biopsied multiple parts from one blastocyst and cultured the biopsied embryos, and extracted whole DNA from the biopsy samples and cultured embryos. Karyotyping by next generation sequencing were performed. Main results and the role of chance We analyzed 34 samples from 11 embryos, including 25 biopsy sample from 11 embryos and 9 cell mass from 7 cultured embryos. In the karyotype tracking results, even though biopsy sample analysis before the culture were uniformed aneuploid or chromosome mosaic, the developing embryo cell mass had normal karyotype. In one embryo as an example, among the three biopsied extra trophectoderm samples from that, two of them were mosaic, and one of them had uniformed chromosome 21 trisomy and chromosome 16 mosaic monosomy. But the embryo formed multiple cell mass in implantation assay. We examined karyotype of three cell mass, and the result from all were normal karyotype. We suggested that the chromosome aberration cells were screened in the human embryo development, and when the function was not carried out the embryo stopped the development. Limitations, reasons for caution Because of small number of samples available, we need more samples for a more accurate evaluation. Furthermore, we cannot evaluate the absolute mechanism that cells with chromosome aberration decreases. Wider implications of the findings: Conventional PGT-A techniques are based on uniformed embryos developing hypothesized past time. As showed in some clinical reports, PGT-A can reduce of spontaneous abortion and chance of embryo transfer. Thinking about aneuploid cell purifying system in embryo development, effectiveness of PGT-A should be more questionable for infertility treatment. Trial registration number A200326004

Case Report: Occupation Radiation Disease, Skin Injury, and Leukemia After Accidental Radiation Exposure

Objective: Follow-up observation of radiation accident in which a worker developed acute radiation disease and eventually died of leukemia. The case provided key practical information for the study on clinical effects of radiation on the health of workers.Case Presentation: We observed and followed-up the progression and effect of radiation exposure at various stages in a 28-year-old male patient. We examined the chromosomal morphology, white blood cell count, and sperm count. Laboratory tests for leukemia diagnosis and other clinical parameters were performed.Results: After the patient was irradiated, the white blood cell level decreased, the sperm count dropped to 0, and the libido completely disappeared. The patient's chromosome aberration cell rate and total chromosome aberration cell rate were 7.33 and 7.66%, respectively. Examination of leukemia diagnostic experiments revealed that abnormal cells accounted for 60%; bone marrow examination showed that prolymphocytes abnormally proliferated, accounting for 89%, and had positive extracellular iron staining. After the initial treatment, the patient's white blood cell level increased and was finally maintained at a normal level, the sperm count returned to normal levels, and libido was restored. The patient died of acute lymphoblastic leukemia 34 years after the exposure.Conclusion: More attention has been paid to the long-term effects of ionizing radiation-induced malignant tumors. The occupational protection of radiographic inspection workers should be strengthened to reduce and avoid occupational injuries to protect the health and safety of workers.

Comparison of Radiation-Induced DNA Damage between Conventional and Computed Tomography Coronary Angiography

Purpose: Conventional coronary angiography (CCA) and coronary computed tomography angiography (CCTA) are the most frequently used imaging modalities to diagnose coronary artery disease (CAD). The amount of radiation and genotoxic damages of these imaging methods showed variation with the improved technology. Thus we sought to compare the ionizing radiation doses and radiation-induced DNA damage in patients who were performed CCA and CCTA.Methods: 76 patients (39 in CCA group, 37 in CCTA group) were enrolled. Patients undergoing CCTA were grouped according to the use of flash technique (22 patients with CCTA-flash, 15 patients with CCTA-other). The effective radiation dose was recorded. Genotoxicity was compared with chromosome aberration test before and after imaging methods.Results: There was a significant difference between the groups in effective radiation doses given to patients. Radiation was lowest in the CCTA-flash group, followed by CCA, and non-flash CCTA group. There was no change in chromosome aberration rate after CCTA-flash group (p = 0.479). There was a significant increase in chromosome aberration rates after CCA and CCTA-other groups (CCA: p = 0.001; CCTA-other: p = 0.01). Conclusions: CTA which was taken with flash technique in dual-energy CT devices delivers lower dose radiation compared to other groups. Due to this significant difference, radiation-induced genetic damage was significantly less in patients with CCTA undergoing flash technique.

Biological features of blood lymphocytes of the primary patients with endometrial cancer

The aim: to examine the radiosensitivity of chromosomes of T-lymphocytes in the blood of primary patients with endometrial cancer depending on the radiation dose. It was expected that the investigations would reveal a cytogenetic parameter as a predictor of radiosensitivity in non-malignant cells of patients exposed to curative irradiation. Materials and methods. Blood samples from 20 primary patients and 30 conditionally healthy donors were examined. Peripheral blood T-lymphocytes culture test system with metaphase chromosome aberration analysis was used. X-ray test-irradiation was performed at G0-stage of the cell cycle in the dose range of 0.5–3.0 Gy. Results. It was shown that the spontaneous level of chromosome aberrations in lymphocytes of primary patients before anti-tumour therapy is 7,82±0,33 aberrations/100 metaphases. This is more than 2-fold higher than the upper limit of average population index and approximately 6-fold higher than the data of own control. In our study during X-ray irradiation of cells cultures of patients, it was found for the first time that the total frequency of radiation-induced chromosome aberrations obeys the classical linear quadratic dose dependence with a predominance of linear component values; the frequency of radiation markers – also linear quadratic dose dependence, but with a predominance of quadratic component. Conclusions. High specificity of T-lymphocyte chromosomes to exposure to ionizing radiation as well as strict dependence of chromosome aberration yield on exposure dose justify their use as predictors of radiosensitivity of healthy cells from the tumour environment. The revealed dependences of induction of chromosomal damage in T-lymphocytes of patients with endometrial cancer prove the need for a personalized approach to plan the course of radiation therapy