Lessons on harmonization of scoring criteria for dicentric chromosome assay in South Korea.

PURPOSE: Networking with other biodosimetry laboratories is necessary to assess the radiation exposure of many individuals in large-scale radiological accidents. The Korea biodosimetry network, K-BioDos, prepared harmonized scoring guidelines for dicentric chromosome assay to obtain homogeneous results within the network and investigated the efficiency of the guidelines. MATERIALS AND METHODS: Three laboratories in K-BioDos harmonized the scoring guidelines for dicentric chromosome assay. The results of scoring dicentric chromosomes using the harmonized scoring guidelines were compared with the laboratories' results using their own methods. Feedback was collected from the scorers following the three intercomparison exercises in 3 consecutive years. RESULTS: K-BioDos members showed comparable capacity to score dicentrics in the three exercises. However, the results of the K-BioDos guidelines showed no significant improvement over those of the scorers' own methods. According to the scorers, our harmonized guidelines led to more rejected metaphases and ultimately decreased the number of scorable metaphases compared with their own methods. Moreover, the scoring time was sometimes longer with the K-BioDos protocol because some scorers were not yet familiar with the guidelines, though most scorers reported that the time decreased or was unchanged. These challenges may cause low adherence to the guidelines. Most scorers expressed willingness to use the guidelines to select scorable metaphases or identify dicentrics for other biodosimetry works, whereas one did not want to use it due to the difference from their calibration curves. CONCLUSIONS: We identified potential resistance to following the harmonized guidelines and received requests for more detailed methods. Our findings suggest that the harmonized criteria should be continually updated, and education and training should be provided for all scorers. These changes could allow members within the biodosimetry network to successfully collaborate and support each other in large-scale radiological accidents.

Effects of Siegesbeckia herba extract against particulate matter 10 (PM10 ) in skin barrier-disrupted mouse models.

OBJECTIVES: Skin barrier disruption is a significant problem of the older population in an aging society. It is characterized by increased transepidermal water loss and decreased skin water content, and particulate matter (PM) is a social issue that can contribute to the exacerbation of skin inflammation. Thus, addressing this problem is urgent. METHODS: Skin barrier-disrupted mouse models were induced by two methods using acetone application or tape-stripping. This study investigated the antioxidative and anti-inflammatory properties of the Siegesbeckia herba extract (SHE) on PM-induced changes in skin barrier-disrupted mouse models. To examine changes in skin water content, inflammatory cytokines, and keratinocyte differentiation markers, mouse models were treated with vehicle 100 µL, PM10 100 µL (100 µg/mL), SHE 100 µL, or PM10 100 µL (100 µg/mL) plus SHE 100 µL. RESULTS: SHE preserved skin hydration in the skin barrier-disrupted mouse models regardless of the presence of PM10 . SHE also inhibited the upregulation of inflammatory cytokines such as interleukin (IL)-1ß, IL-4, IL-6, IL-8, and tumor necrosis factor-α and normalized the downregulation of keratinocyte differentiation markers against PM10 in skin barrier-disrupted mouse models. CONCLUSIONS: This study elucidated the therapeutic effects of SHE against PM10 in skin barrier-disrupted mouse models.

O2 variant chip to simulate site-specific skeletogenesis from hypoxic bone marrow.

The stemness of bone marrow mesenchymal stem cells (BMSCs) is maintained by hypoxia. The oxygen level increases from vessel-free cartilage to hypoxic bone marrow and, furthermore, to vascularized bone, which might direct the chondrogenesis to osteogenesis and regenerate the skeletal system. Hence, oxygen was diffused from relatively low to high levels throughout a three-dimensional chip. When we cultured BMSCs in the chip and implanted them into the rabbit defect models of low-oxygen cartilage and high-oxygen calvaria bone, (i) the low oxygen level (base) promoted stemness and chondrogenesis of BMSCs with robust antioxidative potential; (ii) the middle level (two times ≥ low) pushed BMSCs to quiescence; and (iii) the high level (four times ≥ low) promoted osteogenesis by disturbing the redox balance and stemness. Last, endochondral or intramembranous osteogenesis upon transition from low to high oxygen in vivo suggests a developmental mechanism-driven solution to promote chondrogenesis to osteogenesis in the skeletal system by regulating the oxygen environment.

Application of a semi-automated dicentric scoring system in triage and monitoring occupational radiation exposure.

The dicentric chromosome assay (DCA) is considered the gold standard for radiation biodosimetry, but it is limited by its long dicentric scoring time and need for skilled scorers. The automation of scoring dicentrics has been considered a strategy to overcome the constraints of DCA. However, the studies on automated scoring methods are limited compared to those on conventional manual DCA. Our study aims to assess the performance of a semi-automated scoring method for DCA using ex vivo and in vivo irradiated samples. Dose estimations of 39 blind samples irradiated ex vivo and 35 industrial radiographers occupationally exposed in vivo were estimated using the manual and semi-automated scoring methods and subsequently compared. The semi-automated scoring method, which removed the false positives of automated scoring using the dicentric chromosome (DC) scoring algorithm, had an accuracy of 94.9% in the ex vivo irradiated samples. It also had more than 90% accuracy, sensitivity, and specificity to distinguish binary dose categories reflecting clinical, diagnostic, and epidemiological significance. These data were comparable to those of manual DCA. Moreover, Cohen's kappa statistic and McNemar's test showed a substantial agreement between the two methods for categorizing in vivo samples into never and ever radiation exposure. There was also a significant correlation between the two methods. Despite of comparable results with two methods, lower sensitivity of semi-automated scoring method could be limited to assess various radiation exposures. Taken together, our findings show the semi-automated scoring method can provide accurate dose estimation rapidly, and can be useful as an alternative to manual DCA for biodosimetry in large-scale accidents or cases to monitor radiation exposure of radiation workers.

Sprayable nanomicelle hydrogels and inflammatory bowel disease patient cell chips for development of intestinal lesion-specific therapy.

All-in-one treatments represent a paradigm shift in future medicine. For example, inflammatory bowel disease (IBD) is mainly diagnosed by endoscopy, which could be applied for not only on-site monitoring but also the intestinal lesion-targeted spray of injectable hydrogels. Furthermore, molecular conjugation to the hydrogels would program both lesion-specific adhesion and drug-free therapy. This study validated this concept of all-in-one treatment by first utilizing a well-known injectable hydrogel that underwent efficient solution-to-gel transition and nanomicelle formation as a translatable component. These properties enabled spraying of the hydrogel onto the intestinal walls during endoscopy. Next, peptide conjugation to the hydrogel guided endoscopic monitoring of IBD progress upon adhesive gelation with subsequent moisturization of inflammatory lesions, specifically by nanomicelles. The peptide was designed to mimic the major component that mediates intestinal interaction with Bacillus subtilis flagellin during IBD initiation. Hence, the peptide-guided efficient adhesion of the hydrogel nanomicelles onto Toll-like receptor 5 (TLR5) as the main target of flagellin binding and Notch-1. The peptide binding potently suppressed inflammatory signaling without drug loading, where TLR5 and Notch-1 operated collaboratively through downstream actions of tumor necrosis factor-alpha. The results were produced using a human colorectal cell line, clinical IBD patient cells, gut-on-a-chip, a mouse IBD model, and pig experiments to validate the translational utility.

Chromosome aberration dynamics in breast cancer patients treated with radiotherapy: Implications for radiation biodosimetry.

Although radiological accidents often result in partial-body radiation exposure, most biodosimetry studies focus on estimating whole-body exposure doses. We have evaluated time-dependent changes in chromosomal aberrations before, during, and after localized fractionated radiotherapy. Twelve patients with carcinoma in situ of the breast who underwent identical adjuvant radiation therapy (50 Gy in 25 fractions) were included in the study. Lymphocytes were collected from patients before, during, and after radiotherapy, to measure chromosome aberrations, such as dicentric chromosomes and translocations. Chromosome aberrations were then used to calculate whole- and partial-body biological absorbed doses of radiation. Dicentric chromosome frequencies in all study participants increased during radiotherapy (p < 0.05 in Kruskal-Wallis test). Increases of translocation frequencies during radiotherapy were observed in seven of the twelve patients. The increased levels of dicentric chromosomes and translocations persisted throughout our 1-year follow-up, and evidence of partial-body exposure (such as Papworth's U-value > 1.96) was observed more than 1 year after radiotherapy. We found that cytogenetic biomarkers reflected partial-body fractionated radiation exposure more than 1 year post-exposure. Our findings suggest that chromosome aberrations can be used to estimate biological absorbed radiation doses and can inform medical intervention for individuals suspected of fractionated or partial-body radiation exposure.

Cancer Patient Tissueoid with Self-Homing Nano-Targeting of Metabolic Inhibitor.

The current paradigm of cancer medicine focuses on patient- and/or cancer-specific treatments, which has led to continuous progress in the development of patient representatives (e.g., organoids) and cancer-targeting carriers for drug screening. As breakthrough concepts, i) living cancer tissues convey intact profiles of patient-specific microenvironmental signatures. ii) The growth mechanisms of cancer mass with intense cell-cell interactions can be harnessed to develop self-homing nano-targeting by using cancer cell-derived nanovesicles (CaNVs). Hence, a tissueoid model of ovarian cancer (OC) is developed by culturing OC patient tissues in a 3D gel chip, whose microchannel networks enable perfusion to maintain tissue viability. A novel model of systemic cancer responses is approached by xenografting OC tissueoids into ischaemic hindlimbs in nude mice. CaNVs are produced to carry general chemotherapeutics or new drugs under pre/clinical studies that target the BRCA mutation or energy metabolism, thereby increasing the test scope. This pioneer study cross-validates drug responses from the OC clinic, tissueoid, and animal model by demonstrating the alignment of results in drug type-specific efficiency, BRCA mutation-dependent drug efficiency, and metabolism inhibition-based anti-cancer effects. Hence, this study provides a directional foundation to accelerate the discovery of patient-specific drugs with CaNV application towards future precision medicine.

An intercomparison exercise to compare scoring criteria and develop image databank for biodosimetry in South Korea.

PURPOSE: Mutual cooperation of biodosimetry laboratories is required for dose assessments of large numbers of people with potential radiation exposure, as in mass casualty accidents. We launched an intercomparison exercise to validate the performance of biodosimetry laboratories in South Korea. MATERIALS AND METHODS: Participating laboratories shared metaphase images from dicentric chromosome assays (DCAs) and fluorescence in situ hybridization (FISH)-based translocation assays, which were evaluated based on their own scoring protocols. RESULTS: Overall, the coefficient of variation among three laboratories was less than 10% for counting scorable metaphases and chromosomal aberrations. However, there was variation in the interpretation of the International Atomic Energy Agency guidelines for selecting scorable metaphases and identifying chromosomal aberrations. In a technical workshop, scoring discrepancies were extensively discussed in order to harmonize biodosimetry protocols in Korea. In addition, metaphase images with agreement among all participating laboratories were compiled into an image databank, which can be used for education and training of scorers. CONCLUSIONS: These findings and exercises may improve the accuracy of dose assessment, as well as increase the capacity for biodosimetry in South Korea.

Hormone autocrination by vascularized hydrogel delivery of ovary spheroids to rescue ovarian dysfunctions.

The regeneration potential of implantable organ model hydrogels is applied to treat a loss of ovarian endocrine function in women experiencing menopause and/or cancer therapy. A rat ovariectomy model is used to harvest autologous ovary cells while subsequently producing a layer-by-layer form of follicle spheroids. Implantation of a microchannel network hydrogel with cell spheroids [vascularized hydrogel with ovarian spheroids (VHOS)] into an ischemic hindlimb of ovariectomized rats significantly aids the recovery of endocrine function with hormone release, leading to full endometrium regeneration. The VHOS implantation effectively suppresses the side effects observed with synthetic hormone treatment (i.e., tissue overgrowth, hyperplasia, cancer progression, deep vein thrombosis) to the normal levels, while effectively preventing the representative aftereffects of menopause (i.e., gaining fatty weight, inducing osteoporosis). These results highlight the unprecedented therapeutic potential of an implantable VHOS against menopause and suggest that it may be used as an alternative approach to standard hormone therapy.

Modulation of Cancer Cell Growth and Progression by Caveolin-1 in the Tumor Microenvironment.

Caveolin-1 (Cav-1), a major structural component of cell membrane caveolae, is involved in a variety of intracellular signaling pathways as well as transmembrane transport. Cav-1, as a scaffolding protein, modulates signal transduction associated with cell cycle progression, cellular senescence, cell proliferation and death, lipid homeostasis, etc. Cav-1 is also thought to regulate the expression or activity of oncoproteins, such as Src family kinases, H-Ras, protein kinase C, epidermal growth factor, extracellular signal-regulated kinase, and endothelial nitric oxide synthase. Because of its frequent overexpression or mutation in various tumor tissues and cancer cell lines, Cav-1 has been speculated to play a role as an oncoprotein in cancer development and progression. In contrast, Cav-1 may also function as a tumor suppressor, depending on the type of cancer cells and/or surrounding stromal cells in the tumor microenvironment as well as the stage of tumors.

The standardized Korean Red Ginseng extract and its ingredient ginsenoside Rg3 inhibit manifestation of breast cancer stem cell-like properties through modulation of self-renewal signaling.

BACKGROUND: The ginsenoside Rg3, one of active components of red ginseng, has chemopreventive and anticancer potential. Cancer stem cells retain self-renewal properties which account for cancer recurrence and resistance to anticancer therapy. In our present study, we investigated whether the standardized Korean Red Ginseng extract (RGE) and Rg3 could modulate the manifestation of breast cancer stem cell-like features through regulation of self-renewal activity. METHODS: The effects of RGE and Rg3 on the proportion of CD44high/CD24low cells, as representative characteristics of stem-like breast cancer cells, were determined by flow cytometry. The mammosphere formation assay was performed to assess self-renewal capacities of breast cancer cells. Aldehyde dehydrogenase activity of MCF-7 mammospheres was measured by the ALDEFLUOR assay. The expression levels of Sox-2, Bmi-1, and P-Akt and the nuclear localization of hypoxia inducible factor-1α in MCF-7 mammospheres were verified by immunoblot analysis. RESULTS: Both RGE and Rg3 decreased the viability of breast cancer cells and significantly reduced the populations of CD44high/CD24low in MDA-MB-231 cells. RGE and Rg3 treatment attenuated the expression of Sox-2 and Bmi-1 by inhibiting the nuclear localization of hypoxia inducible factor-1α in MCF-7 mammospheres. Suppression of the manifestation of breast cancer stem cell-like properties by Rg3 was mediated through the blockade of Akt-mediated self-renewal signaling. CONCLUSION: This study suggests that Rg3 has a therapeutic potential targeting breast cancer stem cells.

15-Keto prostaglandin E2 suppresses STAT3 signaling and inhibits breast cancer cell growth and progression.

Overproduction of prostaglandin E2 (PGE2) has been linked to enhanced tumor cell proliferation, invasiveness and metastasis as well as resistance to apoptosis. 15-Keto prostaglandin E2 (15-keto PGE2), a product formed from 15-hydroxyprostaglandin dehydrogenase-catalyzed oxidation of PGE2, has recently been shown to have anti-inflammatory and anticarcinogenic activities. In this study, we observed that 15-keto PGE2 suppressed the phosphorylation, dimerization and nuclear translocation of signal transducer and activator of transcription 3 (STAT3) in human mammary epithelial cells transfected with H-ras (MCF10A-ras). 15-Keto PGE2 inhibited the migration and clonogenicity of MCF10A-ras cells. In addition, subcutaneous injection of 15-keto PGE2 attenuated xenograft tumor growth and phosphorylation of STAT3 induced by breast cancer MDA-MB-231 cells. However, a non-electrophilic analogue, 13,14-dihydro-15-keto PGE2 failed to inhibit STAT3 signaling and was unable to suppress the growth and transformation of MCF10A-ras cells. These findings suggest that the α,ß-unsaturated carbonyl moiety of 15-keto PGE2 is essential for its suppression of STAT3 signaling. We observed that the thiol reducing agent, dithiothreitol abrogated 15-keto PGE2-induced STAT3 inactivation and disrupted the direct interaction between 15-keto PGE2 and STAT3. Furthermore, a molecular docking analysis suggested that Cys251 and Cys259 residues of STAT3 could be preferential binding sites for this lipid mediator. Mass spectral analysis revealed the covalent modification of recombinant STAT3 by 15-keto PGE2 at Cys259. Taken together, thiol modification of STAT3 by 15-keto PGE2 inactivates STAT3 which may account for its suppression of breast cancer cell proliferation and progression.

Src-mediated phosphorylation, ubiquitination and degradation of Caveolin-1 promotes breast cancer cell stemness.

Cancer stem cells (CSCs) are responsible for tumor initiation, metastasis and recurrence. Caveolin-1 (Cav-1) is a major protein of caveolae, which participates in various cellular functions, such as vesicle trafficking, cholesterol homeostasis, tumor progression, etc. In the present study, we investigated a role for Cav-1 in regulating the stemness of human breast cancer (MDA-MB-231) cells. Cav-1 expression was significantly lower in tumorspheres than in adherent cells. The silencing of Cav-1 enhanced stemness of MDA-MB-231 cells. Mechanistically, Cav-1 silencing was accompanied by enhanced expression of Bmi-1, which is a representative self-renewal regulator, and promoted epithelial-mesenchymal transition. In a CSC-like state, reduced Cav-1 depends on its destabilization through ubiquitin-proteasome degradation. We further found that Src-mediated phosphorylation of Cav-1 at the Tyr 14 residue is essential for its degradation. Taken together, these findings suggest that Cav-1 destabilization by Src may play a pivotal role in manifestation and maintenance of stemness in breast cancer cells.

Role of heme oxygenase-1 and its reaction product, carbon monoxide, in manifestation of breast cancer stem cell-like properties: Notch-1 as a putative target.

Cancer stem cells (CSCs) constitute a subpopulation of transformed cells that possess intrinsic ability to undergo selfrenewal and differentiation, which drive tumour resistance and cancer recurrence. It has been reported that CSCs possess enhanced protection against oxidative stress induced by reactive oxygen species compared with nonstem-like cancer cells. In the present work, we investigated the role of heme oxygenase-1 (HO-1), a representative antioxidant enzyme, on the stemness and selfrenewal of human breast CSCs. We found that pharmacologic or genetic inhibition of HO-1 attenuated the sphere formation, whereas HO-1 inducers enhanced the number and the size of tumourspheres in breast CSCs. Carbon monoxide (CO) is endogenously generated as a consequence of degradation of heme by HO-1. The proportion of populations of CD44+/CD24- cells retaining CSC properties was increased in MDA-MB-231 cells treated with a CO-releasing molecule (CORM-2). Following CORM-2 treatment, the expression of Notch-1 and related genes Jagged-1 and Hes1 was increased, which was accompanied by the mammosphere formation. Taken together, these findings suggest that HO-1-derived CO production stimulates the formation of mammospheres in breast cancer cells through activation of Notch-1 signalling.

Erratum to: HIFU: Effects and Clinical Effectiveness of Non-surgical Therapy for Uterine Fibroids.

[This corrects the article on p. 59 in vol. 22, PMID: 27617237.].

Bead affinity chromatography in a temperature-controllable microsystem for biomarker detection.

This paper describes a temperature-controllable bead affinity chromatography (BAC) in a microsystem for biomarker detection, and preparing samples for matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) analysis. Cancer marker proteins were captured in the microsystem by BAC with RNA aptamer-immobilized microbeads. The captured proteins were then denatured and released from the microbeads by controlling temperature. The microsystem consists of a microreactor for trapping microbeads and a temperature control unit for thermal treatment of the trapped beads. We used polymethylsilxoane or single crystalline silicon in fabricating two different types of reaction chamber to compare the differences in performance originated from the materials. Carcinoembryonic antigen was concentrated and purified from human serum using the microsystem and detected by MALDI-TOF MS to demonstrate the usefulness of the microsystem. The microsystem simplifies a sample preparation process required for protein analysis and cancer biomarker detection, which will accelerate the process of cancer research.

Postpartum changes in body composition.

Parity is associated with weight retention and has long-lasting and detrimental effects on the health of women. Previous studies have shown that increasing parity was independently associated with an increased prevalence of metabolic syndrome. Postpartum weight is made up of several components including uterine and mammary tissues, body water (intracellular (ICW) and extracellular water (ECW)), and fat. These components change in variable amounts postpartum, thereby distinctly affecting the interpretation of individual weight retention; however, it is unclear which components contribute to weight retention. The aims of this longitudinal study were to evaluate changes in body composition during the postpartum period and to investigate their effects on weight retention. This prospective study examined 41 healthy, pregnant women who gave birth at Korea University Guro Hospital. We measured body composition at 2 days, 2 weeks, and 6 weeks postpartum using bioelectrical impedance analysis. Weight decreased during this postpartum period (P < 0.001); the postpartum weight retention from prepregnancy to 6 weeks postpartum was 4.43 ± 4.0 kg. Among various body composition components, ECW, ICW, total body water, and fat-free mass (FFM) decreased postpartum. However, fat mass (FM) and visceral fat area, the components that experienced the greatest changes, increased postpartum. Our results demonstrate that the postpartum period is associated with a preferential accumulation of adipose tissue in the visceral compartment, even though overall body weight is decreased. Further studies are needed to evaluate the changes in body composition over longer time periods and their long-term effects on health.