Insights into tissue accumulation, depletion, and health risk assessment of clopidol in poultry.

Clopidol is extensively used in livestock farming and residues of this antibiotic can persist in animal tissues, posing a risk to humans and the environment. In this study, we investigated the depletion of clopidol in various edible tissues of chickens (muscle, liver, kidney, fat, and eggs) using liquid chromatography-tandem mass spectrometry after the administration of a clopidol-contaminated diet (at 250 mg kg-1 for the high (1x) dose). After 14 d of exposure, the clopidol concentrations were highest in eggs (median: 9.83 mg/kg), followed by liver (3.56 mg/kg), kidney (3.01 mg/kg), muscle (1.56 mg/kg), and fat (0.727 mg/kg) at low exposure group, indicating that clopidol accumulated primarily in eggs rather than the other edible tissues. In addition, the maternal transfer ratios were estimated, and the transfer efficiencies of clopidol in muscle (egg-to-tissue ratio, ETR:1.81) and fat (2.06-58.2) were higher than those in liver (0.731-31.1) and kidney (0.832-38.9). Furthermore, we conducted a cumulative risk assessment for clopidol in edible chicken tissues using the hazard quotient (HQ) method. This assessment revealed that the exposure levels for Korean consumers pose an acceptable risk. However, for eggs from the 1x dose exposure group, the HQ values were greater than 1 for all age groups, particularly for young children (<18 y), suggesting that the higher daily consumption of eggs combined with the higher clopidol residues in eggs resulted in higher HQ values, which requires further attention. The findings of this study can assist in the management and monitoring of clopidol residues in chicken tissues and eggs.

Is fetal middle cerebral artery peak systolic velocity correlated with hemoglobin levels in cases of nonimmune fetal anemia?

OBJECTIVES: To establish whether a correlation exists between the fetal middle cerebral artery peak systolic velocity (MCA PSV) and fetal hemoglobin levels before intrauterine transfusion (IUT) in cases of severe fetal anemia. METHODS: This was a single-center, retrospective study of data from 49 fetuses treated with IUT for fetal anemia between 2003 and 2018. Severe fetal anemia was suspected when MCA PSV was or exceeded 1.55 multiples of the median. RESULTS: The causes of anemia were largely idiopathic, and the overall survival rate was 57%. MCA PSV and hemoglobin were correlated in all 34 fetuses with alloimmune fetal anemia, whereas the 15 fetuses with nonimmune causes showed no correlation. Of the 15 noncorrelated cases, twin pregnancy was most common, followed by idiopathic causes. All the twin pregnancies involved monochorionic twins. Fetal hydrops, especially ascites, was significantly associated with severe anemia. CONCLUSIONS: Fetal MCA PSV may not be a reliable independent factor for the diagnosis of severe fetal anemia in nonimmune cases, and the presence of associated hydrops implies that the fetus is more likely to have severe fetal anemia than in a fetus without hydrops.

Perinatal outcome of fetuses with congenital high airway obstruction syndrome: a single-center experience.

OBJECTIVE: To report our experience with management of fetuses with congenital high airway obstruction syndrome (CHAOS). METHODS: We retrospectively reviewed the cases of fetuses who were prenatally diagnosed and postnatally confirmed with CHAOS between 2010 and 2019 at Asan Medical Center, Seoul, Korea. RESULTS: Of 13 fetuses prenatally diagnosed with CHAOS, 7 were lost to follow-up and 6 were postnatally confirmed as having CHAOS. All fetuses, except one were delivered via cesarean section with an ex utero intrapartum treatment (EXIT) procedure. Two patients had coexisting congenital heart diseases requiring several cardiac surgeries following birth. Both of these patients demonstrated developmental delay; however, the remaining 4 had a normal development except for expressive language. Two infants died of respiratory complications, and the remaining 4 were alive at the end of the follow-up period. All 4 live patients underwent tracheostomy with planned reconstruction surgery. Three children are now able to phonate, and 1 can maintain a conservation. CONCLUSION: The proper management of CHAOS using the EXIT procedure results in high survival and low hypoxemia-induced complication rates. Therefore, an accurate prenatal diagnosis is necessary for an appropriate perinatal management.

Individual and regional factors associated with suicidal ideation among Korean elderly: a multilevel analysis of the Korea Community Health Survey.

OBJECTIVES: This study aimed to identify the individual and regional characteristics that influence suicidal ideation among the Korean elderly population. METHODS: Using data collected from the 2013 Korea Community Health Survey, a multilevel analysis was performed to establish an understanding of individual behavioral patterns and regional influences on suicidal ideation. RESULTS: Among the 77,407 individuals sampled, 11,236 (14.5%) elderly people over 60 years of age experienced suicidal ideation. Among individual factors, age, frequency of communication with friends, religious activity, social activity, leisure activity, trust in neighbors, subjective stress level, depressive symptoms, and subjective health status were significantly associated with suicidal ideation. The results showed that the lower the regional deprivation level, the higher the suicidal ideation odds ratio. In terms of regional size, the most significant effects were found in rural areas. CONCLUSIONS: This study suggested that suicidal ideation in the elderly is associated with community factors, such as the regional deprivation index, as well as personal factors.

Toxicological evaluation of 2-dodecylcyclobutanone, a unique radiolytic compound of palmitic acid.

This study was conducted to evaluate the toxic effects and potency of 2-dodecylcyclobutanone (2-dDCB), a unique compound derived from palmitic acid via irradiation. In a series of assays of bacterial reverse-mutation, in vitro chromosomal aberration, and in vivo micronucleus, negative responses were found by the treatment of 2-dDCB comparing vehicle control, dimethyl sulfoxide or corn oil. In the acute oral toxicity test, all of the mice administrated 2-dDCB survived, and there were no clinical and necropsy signs observed at any doses (0, 300, and 2000 mg/kg body weight) during the experimental period of 14 days. These results suggested that 2-dDCB is a relatively non-toxic substance with median lethality dose higher than 2000 mg/kg body weight. Moreover, there were no adverse effects noted in rats orally administrated 2-dDCB everyday via gavage for 28 days, even at the highest dose (2.0 mg/kg body weight/day) tested, which is 1000-times higher than the human daily intake of 2-dDCB estimated through an extreme exposure scenario. Overall, these results indicate that 2-dDCB is not likely to raise any human health concerns and irradiated foods containing palmitic acid can be recognized as safe for human consumption under the current international regulation systems for food irradiation.

Toxicological Evaluation of Flumequine in Pubertal Male Rats After Oral Administration for Six Weeks.

INTRODUCTION: Veterinarians use flumequine (FLU) widely but its toxicological effects are still unclear. MATERIAL AND METHODS: FLU doses of 53, 200, or 750 mg/kg were administered orally for six weeks to pubertal male rats for evaluation of their toxicity. RESULTS: Weight gain was poorer after seven days of exposure to FLU 750, but relative weights of the brain, adrenal and thyroid glands, and testes were notably higher. Haematological and lipid profile parameters, cardiac markers, and inorganic phosphate significantly increased in the FLU 750 group. Blood glucose, oestradiol and serum concentrations of immunoglobulins G (IgG) and E (IgE) significantly decreased after treatment. The levels of interleukins 10 (IL-10) and 6 (IL-6) fell significantly in the FLU 200 and FLU 750 groups. Cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) and cyclooxygenase-2 (Cox-2) expression amplified after treatment. Serum levels of free triiodothyronine (fT3) and free thyroxine (fT4) reduced in the FLU 200 and FLU 750 groups without changes in total T3 or T4 level. All doses of FLU significantly depressed concentrations of thyroid-stimulating hormone (TSH) and testosterone. Histopathology of thyroid glands from rats treated with FLU 750 showed degeneration and depletion of thyroid follicular epithelial cells. Expression of 8-hydroxydeoxyguanosine (8-OHdG) was increased in a dose-dependent manner in the brain, but decreased in the testes. Expression of CYP1A1 increased in the adrenal and pituitary glands. CONCLUSION: The results of this study suggest that the toxicity of FLU in rats is an effect of its disruptive influence on the pituitary-thyroid hormonal system and on the dysfunction of the immune system.

Trifloxystrobin induces tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis in HaCaT, human keratinocyte cells.

As the outermost layer of the body, the skin plays an important role in exposure to pesticides, which could have negative impacts on human health. Trifloxystrobin is a widely used fungicide of the strobilurin class, however, there is little information regarding the skin contact-associated toxic mechanism. Therefore, the present study was performed in order to identify the skin toxicity mechanism of trifloxystrobin using HaCaT (keratinocyte of human skin) cells. Following 24 or 48 h treatment, cell viability, and subsequent Annexin V-FITC/propidium iodide assay, TUNEL assay and Western blotting were performed to investigate the cell death mechanism of trifloxystrobin. Exposure to trifloxystrobin resulted in diminished viability of HaCaT cells in both a time- and concentration-dependent manner. The cell death was derived through apoptotic pathways in the HaCaT cells. Furthermore, we explored the effect of trifloxystrobin on TRAIL-mediated extrinsic apoptosis using siRNA transfection. Knockdown of death receptor 5 suppressed trifloxystrobin-provoked apoptosis. These results indicate that trifloxystrobin induces TRAIL-mediated apoptosis and has an inhibitory effect in keratinocytes that can interfere with the barrier function and integrity of the skin. This could be proposed as a mechanism of skin toxicity by trifloxystrobin and considered in the management of pesticide exposure.

Trifloxystrobin-induced mitophagy through mitochondrial damage in human skin keratinocytes.

Trifloxystrobin is a strobilurin class fungicide, the mode of action of which is to block the mitochondrial electron transport chain and inhibit energy production in fungi. Although adverse effects have been reported by occupational or environmental exposure of fungicides, the pathophysiological mechanism in human cells remains poorly understood. In the present study, we investigated the impact of trifloxystrobin on exposed skin at the cellular organelle level using HaCaT, the human skin keratinocyte cell line. Cells were treated with trifloxystrobin for 48 hr and trifloxystrobin showed detrimental effects on mitochondria evidenced by altered mitochondrial membrane potential and morphology. To identify autophagic degradation of the damaged mitochondria, confocal imaging and Western blotting were performed. Trifloxystrobin induced autophagy-related proteins in HaCaT cells. The mitochondrial reactive oxygen species scavenger mitoTEMPO was applied to further explore the mechanism of trifloxystrobin-mediated mitophagy in human skin cells. PINK1 and Parkin were overexpressed by trifloxystrobin, and mitoTEMPO alleviated the effects on mitophagy induction. Taken together, our findings indicated that mitochondrial damage and mitophagy may play a role in trifloxystrobin-induced toxicity in human keratinocytes and this could be suggested as a mechanism of cutaneous diseases developed by exposure.

Benomyl-induced effects of ORMDL3 overexpression via oxidative stress in human bronchial epithelial cells.

The respiratory system is a major site of exposure route during pesticide use. Although pesticide exposure is associated with chronic respiratory diseases including asthma, the underlying pathophysiological mechanism remains to be elucidated. In this study, we investigated the in vitro effects of benomyl-induced ORMDL3 overexpression on the toxicological mechanism using the human bronchial epithelial cell line 16HBE14o-. Benomyl increased reactive oxygen species and Ca2+ levels, and asthma-related ADAM33 and ORMDL3 expression in 16HBE14o- cells. Considering the change in Ca2+ level and protein expression, we focused on ORMDL3 to elucidate the mechanism of benomyl-induced asthma. Antioxidant treatment showed that benomyl-induced ORMDL3 and endoplasmic reticulum stress could be triggered by oxidative stress. Furthermore, ORMDL3 knockdown alleviated the effects of benomyl on intracellular Ca2+, and the expression of metalloproteinases, and proinflammatory cytokines involved in the pathogenesis of asthma. In conclusion, our results suggest that benomyl-induced ORMDL3 overexpression via oxidative stress might be a mechanism involved in asthma. Moreover, antioxidants and alleviating mechanisms that reduce ORMDL3 levels could serve as promising therapeutic targets for pesticide-induced asthma.

Chlorpyrifos induces NLRP3 inflammasome and pyroptosis/apoptosis via mitochondrial oxidative stress in human keratinocyte HaCaT cells.

Chlorpyrifos (CPF) has been widely used around the world as a pesticide for both agricultural and residential application. Although various studies have reported toxicity and health-related effects from CPF exposure, the molecular mechanism of CPF toxicity to skin has not been well-characterized. The present study determined the potential mechanism involved in skin toxicity of CPF using the HaCaT human skin keratinocyte cell line. After treating to HaCaT cells, CPF triggered reactive oxygen species (ROS) generation and mitochondrial oxidative stress. We focused on NLRP3 inflammasome, known to induce innate immune response. We used mitochondrial ROS (mROS) scavenger mitoTEMPO to demonstrate a role for mROS in NLRP3 inflammasome and programmed cell death induced by CPF. Our results showed that CPF provoked NLRP3 inflammasome and pyroptosis/apoptosis via an increase of mROS in HaCaT cells. This study proposes that CPF induces innate immune response and skin inflammation through activating the NLRP3 inflammasome in skin epithelial cells. CPF may lead to cutaneous disease conditions and antioxidants could be proposed for therapy against skin exposure to CPF.

Effect on the H19 gene methylation of sperm and organs of offspring after chlorpyrifos-methyl exposure during organogenesis period.

To elucidate the effect on the H19 gene methylation of sperm and organs in offspring by chlorpyrifos-methyl (CPM) exposure during organogenesis period, CPM was administered at doses of 4 (CPM4), 20 (CPM20), and 100 (CPM100) mg/kg bw/day from 7 days post coitum (d.p.c.) to 17 d.p.c. after mating CAST/Ei (♂) and B6 (♀). Anogenital distance (AGD) was measured at postnatal day (PND) 21. Clinical signs, body weights, feed and water consumption, organs weights, serum hormone values, and H19 methylation level of organ and sperm were measured at PND63. Body weights were significantly lower than control until PND6. AGD was significantly decreased in the CPM100 group in males and increased in the CPM20 group in females. The absolute weights of the thymus and epididymis were significantly increased for males in all of CPM treatment groups. In the CPM20 group, absolute weights of liver, kidney, heart, lung, spleen, prostate gland, and testes were significantly increased. Testosterone concentrations in serum were significantly increased by CPM treatment in males. H19 methylation level of liver and thymus showed decreased pattern in a dose-dependent manner in males. The levels of H19 methylation in sperm were 73.76 ± 7.16% (Control), 57.84 ± 12.94% (CPM4), 64.24 ± 3.79% (CPM20), and 64.24 ± 3.79% (CPM100). Conclusively, CPM exposure during organogenesis period can disrupt H19 methylation in sperm, liver, and thymus and disturb the early development of offspring.

Towards a strategic approaches in alternative tests for pesticide safety.

Pesticides have provided significant benefits including plant disease control and increased crop yields since people developed and utilized them. However, pesticide use is associated with many adverse effects, which necessitate precise toxicological tests and risk assessment. Most of these methods are based on animal studies, but considerations of animal welfare and ethics require the development of alternative methods for the evaluation of pesticide toxicity. Although the usage of laboratory animals is inevitable in scientific evaluation and alternative approaches have limitations in the whole coverage, continuous effort is necessary to minimize animal use and to develop reliable alternative tests for pesticide evaluation. This review discusses alternative approaches for pesticide toxicity tests and hazard evaluation that have been used in peer-reviewed reports and could be applied in future studies based on the critical animal research principles of reduction, replacement, and refinement.

Toxic effects of methylmercury, arsanilic acid and danofloxacin on the differentiation of mouse embryonic stem cells into neural cells.

This study was performed to assess the neurotoxic effects of methylmercury, arsanilic acid and danofloxacin by quantification of neural-specific proteins in vitro. Quantitation of the protein markers during 14 days of differentiation indicated that the mouse ESCs were completely differentiated into neural cells by Day 8. The cells were treated with non-cytotoxic concentrations of three chemicals during differentiation. Low levels of exposure to methylmercury decreased the expression of GABAA-R and Nestin during the differentiating stage, and Nestin during the differentiated stage. In contrast, GFAP, Tuj1, and MAP2 expression was affected only by relatively high doses during both stages. Arsanilic acid affected the levels of GABAA-R and GFAP during the differentiated stage while the changes of Nestin and Tuj1 were greater during the differentiating stage. For the neural markers (except Nestin) expressed during both stages, danofloxacin affected protein levels at lower concentrations in the differentiated stage than the differentiating stage. Acetylcholinesterase activity was inhibited by relatively low concentrations of methylmercury and arsanilic acid during the differentiating stage while this activity was inhibited only by more than 40 µM of danofloxacin in the differentiated stage. Our results provide useful information about the different toxicities of chemicals and the impact on neural development.

Exposure of pregnant mice to chlorpyrifos-methyl alters embryonic H19 gene methylation patterns.

The aim of this study was to identify whether chlorpyrifos methyl (CPM) exposure during pregnancy leads to changes in the methylation patterns of H19 gene. CPM 4, 20, 100 mg/kg bw/day was administered to 4 pregnant mice per group between 7 and 12 days post coitum (d.p.c.). Pregnant mice were killed at 13 d.p.c. The genomic methylation in primordial germ cells (PGCs) and fetal organs (the liver, intestine, and placenta) was examined. Four polymorphism sites in the H19 alleles of maternal (C57BL/6J) and paternal (CAST/Ei) alleles were identified at nucleotide position 1407, 1485, 1566, and 1654. The methylation patterns of 17 CpG sites were analyzed. The methylation level in male and female PGCs was not altered by CPM treatment in the maternal allele H19. The methylation level of the paternal H19 allele was altered in only male PGCs in response to the CPM treatment. The methylation level at a binding site for the transcriptional regulator CTCF2 was higher than that at the CTCF1 binding site in all CPM-treated groups. In the placenta, the aggregate methylation level of H19 was 56.89%in control group. But, those levels were ranged from 47.7% to 49.89% after treatment with increasing doses of CPM. H19 gene from the liver and intestine of 13 d.p.c. fetuses treated with CPM was hypomethylated as compared with controls, although H19 mRNA expression was unaltered. In the placenta, H19 expression was slightly increased in the CPM-treated group, although not significantly. IGF2 expression levels were not significantly changed in the placenta. In conclusion, CPM exposure during pregnancy alters the methylation status of the H19 gene in PGCs and embryonic tissues. We infer that these alterations are likely related to changes in DNA demethylase activity.

An indirect competitive assay-based aptasensor for detection of oxytetracycline in milk.

Oxytetracycline (OTC) is a common antibacterial agent used for the control of animal diseases. OTC abuse can seriously affect human health; therefore, we developed a biosensor using single-stranded DNA (ssDNA) aptamers for the detection of OTC. The binding probe aptamers for OTC were selected by a Systematic Evolution of Ligands by the exponential enrichment (SELEX) process and identified by the enzyme-linked aptamer assay (ELAA). Among the selected 5 aptamers, aptamer OTC3 showed the strongest affinity (Kd=4.7 nM) and highest specificity for OTC compared to structurally similar antibiotics, tetracycline and chlortetracycline. OTC was detected using indirect competitive ELAA. The limit of detection and quantitation with aptamer OTC3 were 12.3 and 49.8 µg/L, respectively, in milk and showed recovery rates of more than 90% in OTC-spiked milk. This biosensor method with high sensitivity and specificity based on indirect competitive ELAA can be applied to OTC detection in food products on-site because of the simplicity of detection.

Comparison of international guidelines of dermal absorption tests used in pesticides exposure assessment for operators.

The number of farmers who have suffered from non-fatal acute pesticide poisoning has been reported to vary from 5.7% to 86.7% in South Korea since 1975. Absorption through the skin is the main route of exposure to pesticides for farmers who operate with them. Several in vitro tests using the skins of humans or animal and in vivo tests using laboratory animals are introduced for the assessment of human dermal absorption level of pesticides. The objective of this study is to evaluate and compare international guidelines and strategies of dermal absorption assessments and to propose unique approaches for applications into pesticide registration process in our situation. Until present in our situation, pesticide exposure level to operator is determined just using default value of 10 as for skin absorption ratio because of data shortage. Dermal absorption tests are requested to get exposure level of pesticides and to ultimately know the safety of pesticides for operators through the comparison with the value of AOEL. When the exposure level is higher than AOEL, the pesticide cannot be approved. We reviewed the skin absorption test guidelines recommended by OECD, EFSA and EPA. The EPA recommends assessment of skin absorption of pesticides for humans through the TPA which includes all the results of in vitro human and animal and animal in vivo skin absorption studies. OECD and EFSA, employ a tiered approach, which the requirement of further study depends on the results of the former stage study. OECD guidelines accept the analysis of pesticide level absorbed through skin without radioisotope when the recovery using the non-labeled method is within 80~120%. Various factors are reviewed in this study, including the origin of skin (gender, animal species and sites of skin), thickness, temperature and, etc., which can influence the integrity of results.

Change in kidney damage biomarkers after 13 weeks of exposing rats to the complex of Paecilomyces sinclairii and its host Bombyx mori larvae.

Complex of Paecilomyces sinclairii and host larvae, Bombyx mori, is a well known health food; however, concerns about nephrotoxicity have been raised. Kidney toxicity was investigated after 13 weeks of administering the complex orally to rats with parameters including blood urea nitrogen (BUN), creatinine, and kidney damage biomarkers, beta-2-microglobulin (ß2m), glutathione S-transferase alpha (GST-α), kidney injury molecule 1 (KIM-1), tissue inhibitor of matrix metalloproteinase 1 (TIMP-1), vascular endothelial growth factor (VEGF), calbindin, clusterin, cystatin C, neutrophil gelatinase-associated lipocalin (NGAL), and osteopontin. Dose-dependent kidney cell karyomegaly and tubular hypertrophy were observed, with higher severity in males. There was a dose-dependent increase in KIM-1 and TIMP-1 levels in kidney and urinary KIM-1, cystatin C, ß2m, and osteopontin levels. KIM-1 and TIMP-1 increased in male kidneys had not recovered by 2 weeks after stopping exposure. Cystatin C in kidney was significantly lowered in all treatment groups at 13 weeks of administration. All the changes were more noticeable in males. These data indicate that the complex damage renal tubule cells with histopathological lesions and changes in biomarker levels. Kidney and urinary KIM-1 and cystatin C were the most markedly affected and early increased indicators among biomarkers tested, whereas BUN and creatinine were not affected.

Effects of oral deoxynivalenol exposure on immune-related parameters in lymphoid organs and serum of mice vaccinated with porcine parvovirus vaccine.

Mice were exposed to deoxynivalenol (DON) via drinking water at a concentration of 2 mg/L for 36 days. On day 8 of treatment, inactivated porcine parvovirus vaccine (PPV) was injected intraperitoneally. The relative and absolute weight of the spleen was significantly decreased in the DON-treated group (DON). Antibody titers to parvovirus in serum were 47.9 ± 2.4 in the vaccination group (Vac), but 15.2 ± 6.5 in the group treated with DON and vaccine (DON + Vac). The IgA and IgG was not different in the DON, Vac an,d DON + Vac groups. IgM was significantly lower only in the DON + Vac group. However IgE was significantly increased in the Vac and DON + Vac group, but no change was observed between the Vac and DON + Vac groups. The concentrations of IL-2, IL-4, GM-CSF, MCP-1 and Rantes in serum, and IL-1α in mesenteric lymph node and MIP-1ß in spleen were significantly increased by DON treatment compared to control. The concentrations of IL-2, IL-5, IL-6, IL-9, IL-12, IL-13 and Rantes in thymus, of IL-2 in spleen, and of IL-1α, IL-1ß, IL-3, IL-5, IL-10, IL-17, G-CSF, GM-CSF and MCP-1 in mesenteric lymph nodes were significantly decreased in mice compared to those in the Vac group, while concentrations of IL-1α, IL-2, IL-9, IL-13,G-CSF, GM-CSF, IFN-γ, MCP-1, MIP-1α and TNF-α were significantly increased in serum compared to the Vac group. In conclusion, the results presented here indicate that exposure to DON at 2.0 mg/L via drinking water can disrupt the immune response in vaccinated mice by modulating cytokines and chemokines involved in their immune response to infectious disease.

Evaluation of hepatotoxicity of chemicals using hepatic progenitor and hepatocyte-like cells derived from mouse embryonic stem cells: effect of chemicals on ESC-derived hepatocyte differentiation.

Embryonic stem cell testing is an alternative model system to assess drug and chemical toxicities because of its similar developmental characteristics with in vivo embryogenesis and organogenesis. This study evaluated the toxicity of chemicals at specific developmental stages of mouse embryonic stem cell (ESC)-derived hepatic differentiation; hepatic progenitor cells (HPCs), and hepatocyte-like cells (HCs). The toxic effects of carbon tetrachloride (CCl(4)), 5-fluorouracil (5-FU), and arsanilic acid (Ars) were evaluated by measuring the expressions of Cytokeratin (CK18) and GATA binding protein 4 (GATA-4) and the activities of aspartate transaminase (AST), lactate dehydrogenase (LDH), and alkaline phosphatase (ALP) during the hepatic differentiation process. Non-toxic doses of three chemicals at a range of 25 to 500 µM for CCl(4), 12.5 to 800 nM for 5-FU and 6.25 to 400 mM for Ars were treated. In the CCl(4)-treated group, significant decreases (P < 0.05) of the marker expression were observed by more than 300 µM from day 10 in CK18 and by more than 400 µM of CCl(4) from day 22 in GATA-4, respectively. However, both markers were decreased (P < 0.01) by treatments of all doses at day 40. In the 5-FU-treated group, the expressions of two proteins were not affected by any of the doses at day 10 and 22, whereas the GATA-4 expression was decreased (P < 0.05) by more than 400 nM of 5-FU at days 28 and 40. In the Ars-treated group, the CK18 expression was inhibited (P < 0.05) by more than 100 mM of Ars at day 22 but showed a tendency to recover. Although the GATA-4 was inhibited by all doses at day 22, the inhibition of GATA-4 recovered at days 28 and 40. ALP activities of three chemicals were significantly increased (P < 0.05) by a dose-dependent manner. The activities of AST and LDH were prone to be increased by more than 300 µM of CCl(4,) but not affected by all doses of 5-FU except for 800 nM of 5-FU in AST activities. In the Ars, the enzyme activities were significantly increased (P < 0.05) by more than 50 µM of Ars in AST and more than 6.25 µM of Ars in LDH. The present results indicate that CCl(4) has a more toxic effect on HCs, whereas Ars is more toxic to HPCs. Additionally, in vitro alternative testing using ESC-derived HPCs and HCs could provide useful information on chemical toxicity during the hepatic differentiation process and could be a useful model system for assessing chemical hepatotoxicity.

Kidney Toxicity Induced by 13 Weeks Exposure to the Fruiting Body of Paecilomyces sinclairii in Rats.

Paecilomyces sinclairiis (PS) is known as a functional food or human health supplement. However concerns have been raised about its kidney toxicity. This study was performed to investigate the kidney toxicity of PS by 13 week-oral administration to rats. Blood urea nitrogen (BUN), serum creatinine, and kidney damage biomarkers including beta-2-microglobulin (ß2m), glutathione S-transferase alpha (GST-α), kidney injury molecule 1 (KIM-1), tissue inhibitor of matrix metalloproteinase 1 (TIMP-1), vascular endothelial growth factor (VEGF), calbindin, clusterin, cystatin C, neutrophil gelatinase-associated lipocalin (NGAL) and osteopontin were measured during or after the treatment of PS. BUN, creatinine and kidney damage biomarkers in serum were not changed by PS. However, kidney cell karyomegaly and tubular hypertrophy were observed dose-dependently with higher severity in males. KIM-1, TIMP-1 and osteopontin in kidney and urine were increased dose dependently in male or at the highest dose in female rats. Increased urinary osteopontin by PS was not recovered at 2 weeks of post-exposure in both genders. Cystatin C in kidney was decreased at all treatment groups but inversely increased in urine. The changes in kidney damage biomarkers were more remarkable in male than female rats. These data indicate that the PS may provoke renal cell damage and glomerular filtration dysfunction in rats with histopathological lesions and change of kidney damage biomarkers in kidney or urine. Kidney and urinary KIM-1 and cystatin C were the most marked indicators, while kidney weight, BUN and creatinine and kidney damage biomarkers in serum were not influenced.