Objective: The aim of this study was to explore the ototoxicity of toluene in the early development of zebrafish embryos/larvae. Methods: Zebrafish were utilized to explore the ototoxicity of toluene. Locomotion analysis, immunofluorescence, and qPCR were used to understand the phenotypes and molecular mechanisms of toluene ototoxicity. Results: The results demonstrated that at 2 mmol/L, toluene induced zebrafish larvae death at 120 hours post fertilization (hpf) at a rate of 25.79% and inhibited the rate of hatching at 72 hpf. Furthermore, toluene exposure inhibited the distance travelled and average swimming velocity of zebrafish larvae while increasing the frequency of movements. As shown by fluorescence staining of hair cells, toluene inhibited the formation of lateral line neuromasts and middle line 1 (Ml Conclusion This study indicated that toluene may affect the development of both the inner ear and lateral line systems in zebrafish, while the lateral line system may be more sensitive to toluene than the inner ear.
Animals ; Ear, Inner/growth & development* ; Embryo, Nonmammalian/drug effects* ; Gene Expression Regulation, Developmental/drug effects* ; Hair Cells, Auditory/metabolism* ; Lateral Line System/growth & development* ; Locomotion/drug effects* ; Ototoxicity/physiopathology* ; Toluene/toxicity* ; Zebrafish

OBJECTIVE: To investigate the molecular mechanism of trichloroethylene (TCE) cardiac developmental toxicity on zebrafish embryos and to try to provide experimental data for related intervention. METHODS: Zebrafish embryos were purchased from the National Zebrafish Resource Center. The embryos were divided into DMSO(control group), DMSO+CHIR, DMSO+XAV, TCE, TCE+CHIR and TCE+XAV groups(TCE at the concentration of 1, 10 and 100 ppb, with the DMSO as control; DMSO: Dimethyl suldoxide; CHIR: CHIR-99021, Wnt agonist; XAV: XAV-939, Wnt antagonist), 60 embryos per group. Zebrafish embryos were fed in systematic aquaculture water, 28℃. The water was replaced every 24 h and drugs were added according to the grouping scheme. The cardiac tissues were dissected and analyzed by transcriptome microarray after RNA extraction. The expressions of Wnt signaling pathway related genes were verified by q-PCR. Wnt atagonist XAV and activator CHIR were used alone or in combination to further evaluate the possibility of the Wnt signaling participating in the cardiac developmental toxicity induced by TCE. RESULTS: Compared with control, Zebra fish embryos exposed to TCE showed a significant increase in heart defects, and the main phenotypes were abnormal atrioventricular ratio, looping defects and pericardial edema. The results of microarray profiling showed that the expressions of genes related to Wnt signaling pathway were affected significantly. The results of qPCR further confirmed that TCE inhibited the expressions of Wnt pathway target genes Axin2, Sox9b and Nkx2.5(P＜0.05). Wnt agonist CHIR reduced the TCE-induced cardiac malformation rate significantly, while the addition of Wnt antagonist XAV markedly enhanced the cardiac developmental toxicity of TCE. CONCLUSION Exposure to TCE leads to heart malformation in zebrafish embryos. Wnt signaling pathway may be involved in the cardiac developmental toxicity induced by TCE.
Animals ; Gene Expression Regulation, Developmental ; drug effects ; Heart ; drug effects ; embryology ; Transcriptome ; Trichloroethylene ; adverse effects ; Wnt Signaling Pathway ; drug effects ; Zebrafish

Adiposity ; drug effects ; Animals ; Diethylhexyl Phthalate ; toxicity ; Female ; Gene Expression Regulation, Developmental ; drug effects ; Male ; Maternal Exposure ; Metabolic Diseases ; chemically induced ; PPAR gamma ; genetics ; metabolism ; Pregnancy ; Prenatal Exposure Delayed Effects ; Rats ; Rats, Sprague-Dawley

Oligodendrocytes (OLs) are myelinating glial cells that form myelin sheaths around axons to ensure rapid and focal conduction of action potentials. Here, we found that an axonal outgrowth regulatory molecule, AATYK (apoptosis-associated tyrosine kinase), was up-regulated with OL differentiation and remyelination. We therefore studied its role in OL differentiation. The results showed that AATYK knockdown inhibited OL differentiation and the expression of myelin genes in vitro. Moreover, AATYK-deficiency maintained the proliferation status of OLs but did not affect their survival. Thus, AATYK is essential for the differentiation of OLs.
Animals ; Animals, Newborn ; Apoptosis Regulatory Proteins ; genetics ; metabolism ; Cell Differentiation ; drug effects ; physiology ; Cell Proliferation ; drug effects ; genetics ; Cells, Cultured ; Cuprizone ; toxicity ; Demyelinating Diseases ; chemically induced ; metabolism ; pathology ; Embryo, Mammalian ; Gene Expression Regulation, Developmental ; genetics ; Ki-67 Antigen ; metabolism ; Mice ; Mice, Inbred C57BL ; Myelin Basic Protein ; metabolism ; Myelin Proteolipid Protein ; metabolism ; Myelin Sheath ; drug effects ; metabolism ; Oligodendroglia ; drug effects ; metabolism ; Protein-Tyrosine Kinases ; genetics ; metabolism ; RNA, Small Interfering ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley

This study investigated the role of long non-coding RNAs (lncRNAs) in the development of the palatal tissues. Cleft palates in mice were induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Expression levels of long non-coding RNA H19 (lncRNA H19) and insulin-like growth factor 2 (IGF2) gene were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The rate of occurrence of cleft palate was found to be 100% by TCDD exposure, and TCDD could cause short upper limb, cerebral fissure, webbed neck, and short neck. The expression levels of lncRNA H19 and IGF2 gene specifically showed embryo age-related differences on E13, E14, and E15 in the palatal tissues. The expression levels of lncRNA H19 and IGF2 gene showed an inverse relationship on E13, E14, and E15. These findings demonstrated that lncRNA H19 and IGF2 can mediate the development of mouse cleft palate.
Animals ; Cleft Palate ; chemically induced ; genetics ; pathology ; Female ; Gene Expression Regulation ; Gene Expression Regulation, Developmental ; drug effects ; Male ; Mice ; Mice, Inbred C57BL ; Palate ; metabolism ; Polychlorinated Dibenzodioxins ; toxicity ; RNA, Long Noncoding ; genetics ; Real-Time Polymerase Chain Reaction

Lead exposure is a known potential risk factor for neurodegenerative diseases such as Alzheimer's disease (AD). Exposure to lead during the critical phase of brain development has been linked with mental retardation and hypophrenia in later life. This study was aimed to investigate the effects of lead exposure of pregnant mice on the expressions of insulin-degrading enzyme (IDE) and nerve growth factor (NGF) in the hippocampus of their offspring. Blood samples were collected from the tail vein, and after anesthetizing the pups, the brain was excised on postnatal day 21. Lead concentrations were determined by graphite furnace atomic absorption spectrophotometry, and the expressions of IDE and NGF were determined by immunohistochemistry and Western blotting. Results showed that the reduction in IDE and NGF expression in the hippocampus of pups might be associated with impairment of learning and memory and dementia induced by maternal lead exposure during pregnancy and lactation.
Animals ; Down-Regulation ; Female ; Gene Expression Regulation, Developmental ; drug effects ; Gene Expression Regulation, Enzymologic ; drug effects ; Hippocampus ; drug effects ; growth & development ; metabolism ; Insulysin ; genetics ; metabolism ; Lead ; toxicity ; Mice ; Pregnancy ; Prenatal Exposure Delayed Effects ; chemically induced

OBJECTIVEThis study aimed to investigate the expression pattern and function of Nuclear receptor subfamily 2 group E member 1 (Nr2e1) in retinoic acid (RA)-induced brain abnormality.

METHODSThe mouse model of brain abnormality was established by administering 28 mg/kg RA, and neural stem cells (NSCs) were isolated from the mouse embryo and cultured in vitro. Nr2e1 expression was detected by whole mount in situ hybridization, RT-PCR, and Western blotting. Nr2e1 function was determined by transducing Nr2e1 shRNA into NSCs, and the effect on the sonic hedgehog (Shh) signaling pathway was assessed in the cells. In addition, the regulation of Nr2e1 expression by RA was also determined in vitro.

RESULTSNr2e1 expression was significantly downregulated in the brain and NSCs of RA-treated mouse embryos, and knockdown of Nr2e1 affected the proliferation of NSCs in vitro. In addition, a similar expression pattern of Nr2e1 and RA receptor (RAR) α was observed after treatment of NSCs with different concentrations of RA.

CONCLUSIONOur study demonstrated that Nr2e1 could be regulated by RA, which would aid a better understanding of the mechanism underlying RA-induced brain abnormality.

Animals ; Brain ; cytology ; embryology ; Cell Proliferation ; Down-Regulation ; Gene Expression Regulation ; Gene Expression Regulation, Developmental ; drug effects ; Mice ; Mice, Inbred C57BL ; Neural Stem Cells ; drug effects ; physiology ; Receptors, Cytoplasmic and Nuclear ; genetics ; metabolism ; Tretinoin ; pharmacology

Objective: To compare the effects of 50 Hz 1.8 mT sinusoidal magnetic field (SEMF) and 50 Hz 0.6 mT pulsed electromagnetic field(PEMF) on the maturation and mineralization of rat calvaria osteoblasts. Methods: Primary cultured rat calvarial osteoblasts were divided into 3 groups:blank control group, SEMF group and PEMF group. The rats in SEMT and PEMT groups were treated with 50 Hz 1.8 mT SEMF or 50 Hz 0.6 mT PEMF for 90 min/d, respectively. Western blotting and Real-time RT-PCR were used to detect the protein and mRNA expressions of Collagen-1, bone morphogenetic protein 2 (BMP-2), osterix (OSX) and Runt-associated transcription factor 2(Runx-2). The alkaline phosphatase(ALP) activity was detected by ALP test kits at d6 and d9 after treatment, and by ALP staining using azo coupling at d10 after treatment. The formation of calcium nodules was observed by alizarin red staining. Results: Compared with blank control group, the protein and mRNA expressions of Collagen-1, BMP-2, OSX and Runx-2 in SEMT and PEMT groups were significantly increased (P <0.01 or P <0.05); while the mRNA expressions of Collagen-1 and BMP-2 in PEMF group were significantly higher than those in SEMF group. After 6 days treatment, the activity of ALP in PEMF group was significantly higher than that in blank control group (P<0.05), while such difference was not observed in SEMF group (P0.05); after 9 days treatment, the activities of ALP in both PEMF and SEMP groups were significantly higher than that in blank control group (all P<0.05), but the difference between PEMF and SEMF groups was not significant (P0.05). After 10 days treatment, ALP staining was increased in both PEMF and SEMF groups compared with that in blank control group (all P<0.01), and the stained area was bigger in PEMF group than that in SEMF group (P<0.05). After 12 days treatment, calcium nodules were increased in PEMF and SEMF groups compared with that in blank control group (all P<0.01), and more calcium nodules were observed in PEMF group than SEMF group (P<0.05). Conclusion: Both 50 Hz 1.8 mT that in SEMF and 50 Hz 0.6 mT PEMF can promote the maturation and mineralization of osteoblasts, and the effect of PEMF is more marked.
Animals ; Calcification, Physiologic ; drug effects ; Cell Differentiation ; Cells, Cultured ; Electromagnetic Fields ; Gene Expression Regulation, Developmental ; radiation effects ; Magnetic Fields ; Osteoblasts ; cytology ; radiation effects ; Rats ; Skull ; drug effects

Objective: To investigate the effect of icaritin on maturation and mineralization of mouse osteoblast MC3T3-E1 cells and its mechanism. Methods: The cultured MC3T3-E1 cells were divided into blank control group, CXC chemokine receptor type 4 (CXCR4) inhibitor (AMD3100) group, icaritin group, and icaritin plus AMD3100 group. The expression of CXCR4, stromal cell-derived factor 1 (SDF-1) and osteogenesis-related genes and proteins were detected by real-time RT-PCR and Western blotting after drug treatment for 24 h. The alkaline phosphatase (ALP) activity was determined with ALP kit on d3 and d6; calcium nodules were detected by alizarin red staining after drug treatment for 14 d. Results: Real time RT-PCR showed that compared with the blank control group, relative expressions of CXCR4, SDF-1 and osteogenesis-related genes in icaritin group were significantly increased (P<0.05 or P<0.01); After AMD3100 treatment, the relative expression of CXCR4 gene was decreased (P<0.05). Western blot showed that compared with the blank control group, relative expressions of CXCR4, SDF-1 and osteogenesis-related proteins in the icaritin group were significantly increased (all P<0.01), but were decreased after AMD3100 was added (all P<0.01). The ALP activity of icaritin group was significantly higher than that of blank control group (all P<0.01) on d3 and d6 after drug treatment, while the activity of ALP was significantly decreased after AMD3100 treatment (all P<0.01). At d14 after drug treatment, compared with the blank control group, the area of alizarin red staining was increased in the icaritin group, while it was significantly reduced after the addition of AMD3100. Conclusion: Icaritin may promote maturation and mineralization of mouse osteoblast MC3T3-E1 cells through CXCR4/SDF-1 signaling pathway.
3T3 Cells ; Animals ; Calcification, Physiologic ; drug effects ; Chemokine CXCL12 ; metabolism ; Flavonoids ; pharmacology ; Gene Expression Regulation, Developmental ; drug effects ; Mice ; Osteoblasts ; cytology ; drug effects ; Receptors, CXCR4 ; metabolism ; Signal Transduction ; drug effects

The effect of high concentrations of testosterone on ovarian follicle development was investigated. Primary follicles and granulosa cells were cultured in vitro in media supplemented with a testosterone concentration gradient. The combined effects of testosterone and follicle-stimulating hormone (FSH) on follicular growth and granulosa cell gonadotropin receptor mRNA expression were also investigated. Follicle growth in the presence of high testosterone concentrations was promoted at early stages (days 1-7), but inhibited at later stage (days 7-14) of in vitro culture. Interestingly, testosterone-induced follicle development arrest was rescued by treatment with high concentrations of FSH (400 mIU/mL). In addition, in cultured granulosa cells, high testosterone concentrations induced cell proliferation, and increased the mRNA expression level of FSH receptor (FSHR), and luteinized hormone/choriogonadotropin receptor. It was concluded that high concentrations of testosterone inhibited follicle development, most likely through regulation of the FSH signaling pathway, although independently from FSHR downregulation. These findings are an important step in further understanding the pathogenesis of polycystic ovary syndrome.
Androgens ; pharmacology ; Animals ; Cell Proliferation ; drug effects ; Female ; Follicle Stimulating Hormone ; genetics ; metabolism ; pharmacology ; Gene Expression Regulation, Developmental ; Granulosa Cells ; cytology ; drug effects ; metabolism ; Mice ; Ovarian Follicle ; cytology ; drug effects ; growth & development ; metabolism ; Primary Cell Culture ; RNA, Messenger ; genetics ; metabolism ; Receptors, FSH ; genetics ; metabolism ; Receptors, Gonadotropin ; genetics ; metabolism ; Receptors, LH ; genetics ; metabolism ; Signal Transduction ; drug effects ; genetics ; Testosterone ; antagonists & inhibitors ; pharmacology