Let-7c-5p associate with inhibition of phenobarbital-induced cell proliferation in human palate cells.

Cleft palate (CP) is one of the most common congenital diseases, and is accompanied by a complicated etiology. Medical exposure in women is among one of the reasons leading to CP. Recently, it has been reported that microRNA (miRNA) plays a crucial role in palate formation and the disruption of miRNA that influence the development of CP. Although association with pharmaceuticals and miRNAs were suggested, it has remained largely unknow. The aim of the current investigation is to elucidate upon the miRNA associated with the inhibition of phenobarbital (PB)-induced cell proliferation in human embryonic palatal mesenchymal (HEPM) cells. We showed that PB inhibited HEPM cell viability in a dose-dependent manner. We demonstrated that PB treatment suppressed cyclin-D1 expression in HEPM cells. Furthermore, PB upregulated let-7c-5p expression and downregulated the expression of two downstream genes (BACH1 and PAX3). Finally, we demonstrated that the let-7c-5p inhibitor alleviated PB-induced inhibition of cell proliferation and altered BACH1 and PAX3 expression levels. These results suggest that PB suppresses cell viability by modulating let-7c-5p expression.

A novel alternative method for long-term evaluation of male reproductive toxicity and its recovery using a pre-pubertal mouse testis organ culture system.

Successful treatment of pediatric cancers often results in long-term health complications, including potential effects on fertility. Therefore, assessing the male reproductive toxicity of anti-cancer drug treatments and the potential for recovery is of paramount importance. However, in vivo evaluations are time-intensive and require large numbers of animals. To overcome these constraints, we utilized an innovative organ culture system that supports long-term spermatogenesis by placing the testis tissue between a base agarose gel and a polydimethylsiloxane ceiling, effectively mirroring the in vivo testicular environment. The present study aimed to determine the efficacy of this organ culture system for accurately assessing testicular toxicity induced by cisplatin, using acrosin-green fluorescent protein (GFP) transgenic neonatal mouse testes. The testis fragments were treated with different concentrations of cisplatin-containing medium for 24 h and incubated in fresh medium for up to 70 days. The changes in tissue volume and GFP fluorescence over time were evaluated to monitor the progression of spermatogenesis, in addition to the corresponding histopathology. Cisplatin treatment caused tissue volume shrinkage and reduced GFP fluorescence in a concentration-dependent manner. Recovery from testicular toxicity was also dependent on the concentration of cisplatin received. The results demonstrated that this novel in vitro system can be a faithful replacement for animal experiments to assess the testicular toxicity of anti-cancer drugs and their reversibility, providing a useful method for drug development.

Quantitative BONCAT Allows Identification of Newly Synthesized Proteins after Optic Nerve Injury.

Retinal ganglion cells (RGCs) die after optic nerve trauma or in degenerative disease. However, acute changes in protein expression that may regulate RGC response to injury are not fully understood, and detailed methods to quantify new protein synthesis have not been tested. Here, we develop and apply a new in vivo quantitative measure of newly synthesized proteins to examine changes occurring in the retina after optic nerve injury. Azidohomoalanine, a noncanonical amino acid, was injected intravitreally into the eyes of rodents of either sex with or without optic nerve injury. Isotope variants of biotin-alkyne were used for quantitative BONCAT (QBONCAT) mass spectrometry, allowing identification of protein synthesis and transport rate changes in more than 1000 proteins at 1 or 5 d after optic nerve injury. In vitro screening showed several newly synthesized proteins regulate axon outgrowth in primary neurons in vitro This novel approach to targeted quantification of newly synthesized proteins after injury uncovers a dynamic translational response within broader proteostasis regulation and enhances our understanding of the cellular response to injury.SIGNIFICANCE STATEMENT Optic nerve injury results in death and degeneration of retinal ganglion cells and their axons. The specific cellular response to injury, including changes in new protein synthesis, is obscured by existing proteins and protein degradation. In this study, we introduce QBONCAT to isolate and quantify acute protein synthesis and subsequent transport between cellular compartments. We identify novel candidate protein effectors of the regenerative response and uncover their regulation of axon growth in vitro, validating the utility of QBONCAT for the discovery of novel regulatory and therapeutic candidates after optic nerve injury.

Involvement of Bmal1 and Clock in Bromobenzene Metabolite-Induced Diurnal Renal Toxicity.

Circadian rhythms are endogenous oscillators that regulate 24 h behavioral and physiological processes. Our previous investigation demonstrated that bromobenzene metabolite (4-bromocatechol: 4-BrCA) exhibited chronotoxicity (i.e., the nephrotoxicity induced by 4-BrCA was observed during the dark phase, while not observed at light phase in mice). However, the molecular mechanism is still unknown. The aim of the present study is to investigate the cellular molecule(s) involved in the 4-BrCA-induced nephrotoxicity using mouse renal cortex tubular cell lines (MuRTE61 cells). We found that 4-BrCA showed dose dependent (0.01-1 mM) cell proliferation defect in MuRTE61 cells. By treating with 0.03 mM 4-BrCA, we demonstrated that major clock genes (Bmal1, Clock, Cry1, Cry2, Per1, and Per2) were significantly downregulated. Interestingly, the expression levels of two genes, Bmal1 and Clock, continued to decrease after 3 h of treatment with 4-BrCA, while Cry1, Per1, and Per2 were unchanged until 24 h of treatment. Moreover, BMAL1 and CLOCK levels are higher at light phase. We speculated that BMAL1 and CLOCK might function defensively against 4-BrCA-induced nephrotoxicity since the expression levels of Bmal1 and Clock were rapidly decreased. Finally, overexpression of Bmal1 and Clock restored 4-BrCA-induced cell proliferation defect in MuRTE61 cells. Taken together, our results suggest that Bmal1 and Clock have protective roles against 4-BrCA-induced nephrotoxicity.

Copper-induced diurnal hepatic toxicity is associated with Cry2 and Per1 in mice.

BACKGROUND: This study aimed to investigate diurnal variations in copper-induced hepatic toxicity and the molecular mechanisms underlying this chronotoxicity. METHODS: Male C57BL/6J mice were intraperitoneally injected with copper chloride (CuCl2) at zeitgeber time 2 (ZT2) or 14 (ZT14), twice per week for 5 or 8 weeks. Seventy-two hours after the final CuCl2 injection, the mice were euthanized, and plasma samples were collected. The livers and kidneys were collected and weighed. In vitro experiments were performed to assess cell viability and fluctuations in clock gene expression levels in Hepa1-6 cells after CuCl2 treatment. We examined copper homeostasis- and apoptosis-related genes under clock genes overexpression. RESULTS: Repeated CuCl2 administration for 8 weeks resulted in more severe toxicity at ZT14 compared to ZT2. CuCl2 administration at ZT14 elevated plasma aspartate aminotransferase, hepatic tumor necrosis factor-α, and interleukin-6 for 5 weeks, whereas the toxic effects of CuCl2 administration at ZT2 were weaker. Moreover, CuCl2 treatment inhibited Hepa1-6 cell viability in a dose-dependent manner. We observed increased expression of three clock genes (Ciart, Cry2, and Per1) after CuCl2 treatment. Among them, overexpression of Cry2 and Per1 accelerated CuCl2-induced inhibition of Hepa1-6 cell viability. Moreover, we found that the overexpression of Cry2 and Per1 regulates cleaved caspase-3 by modulating the copper transporter genes ATP7B and CTR1. CONCLUSION: These results suggest that CuCl2-induced diurnal toxicity is associated with Cry2 and Per1 expression through the regulation of copper transporter genes in mice.

Quantitative evaluation of spermatogenesis by fluorescent histochemistry.

Identifying the types of spermatogenic cells that compose seminiferous tubules, as well as qualitative confirmation of the presence or absence of disorders, has been regarded as crucial in spermatogenesis. Sperm count and fertilizing capacity, both of which depend on the quality as well as quantity of spermatogenesis, are factors critical to fertilization. However, the quantitative assessment of spermatogenesis is not commonly practiced. Spermatogenesis has species-specific stages; when the specific stage in the seminiferous tubules is precisely determined, the types of spermatogenic cells in each stage can be spontaneously identified. Thereafter, a unique marker is used to classify the cells observed in each stage. Quantitative assessment of spermatogenesis has the potential to detect inapparent spermatogenesis disorders or numerically indicate the degree of the disorder. To this end, a histochemical approach using unique markers is indispensable for the quantitative assessment of spermatogenesis. Future developments in techniques to measure cell populations using computer software will further facilitate the establishment of quantitative assessment of spermatogenesis as a standard analysis method that can contribute significantly to advance our understanding of spermatogenesis.

Two-year intermittent exposure of a multiwalled carbon nanotube by intratracheal instillation induces lung tumors and pleural mesotheliomas in F344 rats.

BACKGROUND: A mounting number of studies have been documenting the carcinogenic potential of multiwalled carbon nanotubes (MWCNTs); however, only a few studies have evaluated the pulmonary carcinogenicity of MWCNTs in vivo. A 2-year inhalation study demonstrated that MWNT-7, a widely used MWCNT, was a pulmonary carcinogen in rats. In another 2-year study, rats administered MWNT-7 by intratracheal instillation at the beginning of the experimental period developed pleural mesotheliomas but not lung tumors. To obtain data more comparable with rats exposed to MWNT-7 by inhalation, we administered MWNT-7 to F344 rats by intratracheal instillation once every 4-weeks over the course of 2 years at 0, 0.125, and 0.5 mg/kg body weight, allowing lung burdens of MWNT-7 to increase over the entire experimental period, similar to the inhalation study. RESULTS: Absolute and relative lung weights were significantly elevated in both MWNT-7-treated groups. Dose- and time-dependent toxic effects in the lung and pleura, such as inflammatory, fibrotic, and hyperplastic lesions, were found in both treated groups. The incidences of lung carcinomas, lung adenomas, and pleural mesotheliomas were significantly increased in the high-dose group compared with the control group. The pleural mesotheliomas developed mainly at the mediastinum. No MWNT-7-related neoplastic lesions were noted in the other organs. Cytological and biochemical parameters of the bronchoalveolar lavage fluid (BALF) were elevated in both treated groups. The lung burden of MWNT-7 was dose- and time-dependent, and at the terminal necropsy, the average value was 0.9 and 3.6 mg/lung in the low-dose and high-dose groups, respectively. The number of fibers in the pleural cavity was also dose- and time-dependent. CONCLUSIONS: Repeated administration of MWNT-7 by intratracheal instillation over the 2 years indicates that MWNT-7 is carcinogenic to both the lung and pleura of rats, which differs from the results of the 2 carcinogenicity tests by inhalation or intratracheal instillation.

Ocular biometry with swept-source optical coherence tomography-based optical biometer in Japanese patients with EYS-related retinitis pigmentosa: a retrospective study.

BACKGROUND: This study aimed to identify the features of ocular biometry in patients with EYS-related retinitis pigmentosa using IOLMaster 700. METHODS: We retrospectively reviewed the medical records of patients with retinitis pigmentosa. Patients with records of the following were included: (1) ocular biometry measurements using the IOLMaster 700 and (2) genetic diagnostic tests. Axial length, keratometry, anterior chamber depth, aqueous depth, lens thickness, central corneal thickness (CCT), and corneal diameter (white to white) measurements were extracted. Based on keratometry measurements, (1) standard keratometric astigmatism, (2) posterior corneal astigmatism, and (3) total corneal astigmatism were obtained. Demographics and biometric parameters were compared between patients with EYS-related retinitis pigmentosa and other patients with retinitis pigmentosa. RESULTS: A total of 86 eyes of 44 patients (23 females and 21 males; mean age: 47.7 years) with retinitis pigmentosa were included. Of these, 18 were identified as having EYS variants. CCT was significantly thinner (P < 0.001) and the posterior corneal curvature at the steepest meridian was significantly smaller (P = 0.024) in patients with EYS-related retinitis pigmentosa than in other patients with retinitis pigmentosa. The magnitudes of all corneal astigmatism measurements was higher in patients with EYS-related RP, although these differences were not statistically significant. CONCLUSION: Patients with EYS-related retinitis pigmentosa had unique features in ocular biometry, such as thinner central corneal thickness and smaller posterior corneal curvature radius at the steepest meridian compared with other patients with retinitis pigmentosa. The findings suggest that patients with retinitis pigmentosa have different ocular dimension features among the different causative genes.

Association of Bruch's membrane opening and optic disc morphology to axial length and visual field defects in eyes with primary open-angle glaucoma.

PURPOSE: To examine the morphology of Bruch's membrane opening (BMO), optic disc, and peripapillary atrophy (PPA) by scanning laser ophthalmoscopy (SLO) and spectral-domain optical coherence tomography (SD-OCT), and to determine their association with the axial length and visual field defects. METHODS: This was a cross-sectional study of 94 eyes of 56 subjects; 77 eyes were diagnosed with primary open-angle glaucoma and 17 eyes as normal. The margins of the optic disc were determined in the SLO images, and that of the BMO in the SD-OCT images. The ovality and area of the BMO and the optic disc were measured. The beta and gamma-PPA areas were also measured. The association of each parameter with the axial length and the mean deviation (MD) of the visual field tests was determined by generalized estimating equations (GEEs). RESULTS: The optic disc ovality was associated with the axial length and the MD (ß = -0.47, P = 7.6 × 10-4 and ß = 0.12, P = 0.040). The BMO ovality was not significantly associated with the axial length and the MD. The BMO area was associated with the axial length (ß = 0.30, P = 0.029). A larger BMO area was associated with a thinner BMO-based neuroretinal rim width (BMO-MRW) after adjustments for the MD (ß = -0.30, P = 2.1 × 10-4). The beta- and gamma-PPA areas were associated with the axial length (ß = 0.50, P = 7.4 × 10-5 and ß = 0.62, P = 4.2 × 10-6). CONCLUSIONS: The optic disc ovality was associated with both the axial length and MD, whereas BMO ovality was not. Attention should be paid to the influence of the axial length-related enlargement of the BMO.

Circumpapillary retinal nerve fiber layer thickness, anterior lamina cribrosa depth, and lamina cribrosa thickness in neovascular glaucoma secondary to proliferative diabetic retinopathy: a cross-sectional study.

BACKGROUND: To compare the lamina cribrosa between eyes with and without neovascular glaucoma (NVG) using enhanced depth imaging spectral-domain optical coherence tomography. METHODS: Forty-six patients with proliferative diabetic retinopathy were enrolled in this cross-sectional study. The patients were divided into two groups based on the absence or presence of NVG (the non-NVG group and the NVG group, respectively). The intraocular pressure (IOP), circumpapillary retinal nerve fiber layer (cpRNFL) thickness, anterior lamina cribrosa depth (ALD), and laminar thickness (LT) were compared between the groups. RESULTS: In the non-NVG group, the mean age was 66.2 ± 2.4 (mean ± standard error) years, mean maximum IOP was 18.8 ± 1.8 mmHg, mean cpRNFL thickness was 91.2 ± 3.9 µm, mean ALD was 407.0 ± 22.9 µm, and mean LT was 155.0 ± 4.7 µm. In the NVG group, the mean age was 61.4 ± 2.1 years, mean maximum IOP was 33.1 ± 1.6 mmHg, mean cpRNFL thickness was 73.6 ± 3.4 µm, mean ALD was 403.9 ± 20.1 µm, and mean LT was 156.9 ± 4.2 µm. The IOP was significantly higher and the cpRNFL was significantly thinner in the NVG group (P < 0.001 and P = 0.002, respectively). However, the age, ALD, and LT were not statistically different between the groups (P = 0.151, 0.919, and 0.757, respectively). CONCLUSIONS: Although the cpRNFL was thinner, the structure of the lamina cribrosa was unchanged in the NVG eyes. Axonal loss of the retinal ganglion cells in NVG patients was prior to lamina cribrosa deformation.

Structural and Functional Analyses in Nonarteritic Anterior Ischemic Optic Neuropathy: Optical Coherence Tomography Angiography Study.

BACKGROUND: Retinal and optic disc perfusion in nonarteritic anterior ischemic optic neuropathy (NAION) is incompletely understood. Our aim was to investigate the characteristics of the microvascular structures at the peripapillary area and optic disc, and their associations with retinal structure and function in patients with NAION. METHODS: We conducted a prospective, observational case series study. Thirty-four eyes, consisting of 15 NAION eyes and 19 normal eyes, were included. Optical coherence tomography (OCT) angiography was used to measure the vessel densities in the peripapillary superficial retina and whole-depth mode inside the optic disc. Measurement of circumpapillary retinal nerve fiber layer (cpRNFL) thickness was performed using OCT. Sectorial division analysis of cpRNFL was performed by eliminating the influences of the difference in disc rotation between OCT images and OCT angiography images. RESULTS: The vessel densities of peripapillary retina and inside the optic disc were significantly reduced in the NAION compared to the normal (both P < 0.001). Both the severity of visual field defect and cpRNFL thinning were significantly associated with the peripapillary vessel density (P = 0.006, P = 0.046), but not with the optic disc vessel density (P = 0.981, P = 0.856). cpRNFL and peripapillary vessel density showed reduction predominantly in the superior sectors, corresponding to the visual field defect. However, the correlations showed discrepancy of the sectors. CONCLUSIONS: The microvascular structures in the peripapillary retina and optic disc were reduced, but the cpRNFL thinning was associated with vessel density only in the peripapillary retina, indicating that the vessel densities in the peripapillary retina and optic disc may be differently affected in the pathological process of NAION.

The relationship between self-reported habitual exercise and visual field defect progression: a retrospective cohort study.

BACKGROUND: Exercise reduces intraocular pressure (IOP) in the short term. However, it is not known whether exercise contributes to slower glaucomatous visual field defect progression. METHODS: Twenty-four primary open-angle glaucoma or exfoliation glaucoma patients who were evaluated by the Humphrey Field Analyzer (HFA) 24-2 program ≥ four times in 3 years were enrolled. Patients with a history of intraocular surgery in past 3 years or other eye diseases threatening visual fields were excluded. Patients were classified into two groups whether they had exercise habits or not. RESULTS: Eleven patients had exercise habits. The mean ± standard error of IOP and MD slope were 14.8 ± 0.9 mmHg and +0.20 ± 0.20 dB/year in the exercise group and 13.3 ± 0.8 mmHg and -0.53 ± 0.18 dB/year in the non-exercise group (P = 0.24 and P = 0.01, respectively). Higher IOP [odds ratio (OR) = 0.44/1 mmHg increase; P = 0.02] and habitual exercise (OR = 0.04; P = 0.02) reduced the visual field defect progression risk in logistic regression analyses. CONCLUSIONS: Patients with self-reported exercise habits had slower glaucoma progression.

Persephin: A potential key component in human oral cancer progression through the RET receptor tyrosine kinase-mitogen-activated protein kinase signaling pathway.

Persephin (PSPN) is a neurotrophic factor of the glial cell line-derived neurotrophic factor (GDNF) family that promotes survival of multiple populations of neurons. Little is known about the relevance of PSPN in human malignancy including oral squamous cell carcinoma (OSCC). This study was undertaken to evaluate PSPN mRNA and protein expression by analyzing cellular proliferation and the cell cycle in PSPN knockdown cells in vitro. PSPN mRNA and protein were significantly (P < 0.05) up-regulated in OSCC-derived cells compared with human normal oral keratinocytes (n = 7). Cellular proliferation decreased significantly (P < 0.05) in PSPN knockdown cells with reduced receptor tyrosine kinase (RTK) signaling, and cell-cycle arrest at the G1 phase resulted from up-regulation of the cyclin-dependent kinase inhibitors (p21(Cip1) , p27(Kip1) , p15(INK4B) , and p16(INK4A) ). Furthermore, the PSPN protein expression in 101 primary OSCCs was significantly (P < 0.05) higher than in normal counterparts. Among the clinical variables analyzed, overexpression of PSPN also was related closely (P < 0.05) to tumoral size. Our results suggested that PSPN is a possible key regulator of OSCC progression via PSPN-RET-mitogen-activated protein kinase activation and that PSPN overexpression may have diagnostic potential for OSCC.

Discovery and genetic analysis of non-bitter Tartary buckwheat (Fagopyrum tataricum Gaertn.) with trace-rutinosidase activity.

In a screening of about 500 lines of Tartary buckwheat, we identified lines that contained no detectable rutinosidase isozymes using an in-gel detection assay. We confirmed that seeds of these individuals had only a trace level of in-vitro rutinosidase activity. To investigate the heritability of the trace-rutinosidase characteristic, we analyzed the progeny of crosses between rutinosidase trace-lines, 'f3g-162', and the 'Hokkai T8'. The F2 progeny clearly divided into two groups: those with rutinosidase activity under 1.5 nkat/g seed (trace-rutinosidase) and those with activity over 400 nkat/g seed (normal rutinosidase). The segregation pattern of this trait in F2 progeny exhibited 1 : 3 ratio (trace-rutinosidase : normal rutinosidase), suggesting that the trace-rutinosidase trait is conferred by a single recessive gene; rutinosidase-trace A (rutA). In addition, sensory panelists evaluated the bitterness of flour from trace-rutinosidase individuals and did not detect bitterness, whereas flour from normal rutinosidase individuals was found to have strong bitterness. Although at least three bitter compounds have been reported in Tartary buckwheat seeds, our present findings indicate that rutin hydrolysis is the major contributing factor to bitterness. In addition, the trace-rutinosidase line identified here, 'f3g-162', is a promising material for generating a non-bitter Tartary buckwheat variety.

Breeding of 'Manten-Kirari', a non-bitter and trace-rutinosidase variety of Tartary buckwheat (Fagopyrum tataricum Gaertn.).

Here, we developed a new Tartary buckwheat cultivar 'Manten-Kirari', whose flour contains only trace amounts of rutinosidase and lacked bitterness. The trace-rutinosidase breeding line 'f3g-162' (seed parent), which was obtained from a Nepalese genetic resource, was crossed with 'Hokkai T8' (pollen parent), the leading variety in Japan, to improve its agronomic characteristics. The obtained progeny were subjected to performance test. 'Manten-Kirari' had no detectable rutinosidase isozymes in an in-gel detection assay and only 1/266 of the rutinosidase activity of 'Hokkai T8'. Dough prepared from 'Manten-Kirari' flour contained almost no hydrolyzed rutin, even 6 h after the addition of water, whereas the rutin in 'Hokkai T8' dough was completely hydrolyzed within 10 min. In a sensory evaluation of the flour from the two varieties, nearly all panelists detected strong bitterness in 'Hokkai T8', whereas no panelists reported bitterness in 'Manten-Kirari'. This is the first report to describe the breeding of a Tartary buckwheat cultivar with reduced rutin hydrolysis and no bitterness in the prepared flour. Notably, the agronomic characteristics of 'Manten-Kirari' were similar to those of 'Hokkai T8', which is the leading variety in Japan. Based on these characteristics, 'Manten-Kirari' is a promising for preparing non-bitter, rutin-rich foods.

Histological differences between the central and peripheral areas of the testes of busulfan-administered mice.

Busulfan is an anticancer drug known to cause serious damage to seminiferous tubules in the testes and deplete germ cells in human and animal models. The testicular artery is anastomosed with deferential and cremasteric arteries and is divided into capsular arteries, which give rise to the centripetal arteries and then recurrent arteries. The arterial blood in the testicular tissue is supplied by such a consequent system of arterial vessels, in order from the peripheral to the central area. As anticancer drugs are generally distributed throughout the whole body via the bloodstream and the running and distribution of arteries differ among the testicular areas, we hypothesized that the efficacy of busulfan differs in different testicular areas, particularly between the central and peripheral areas. In this study, busulfan was intraperitoneally injected at 40 mg/kg body weight into C57BL/6J male mice. After 28 days, in busulfan-treated mice, the diameters of seminiferous tubules were significantly higher in the central than in the peripheral area of the testes. The seminiferous tubular areas also significantly decreased in the peripheral areas compared with the central areas. The number of germ cells per seminiferous tubule was significantly higher in the central than in the peripheral area. Sertoli cell nuclei were detached into the lumen in the peripheral area. The number of Leydig cells was significantly lower in the peripheral areas. These data suggest that the effects of busulfan differ between the central and peripheral areas of the testis at 4 weeks after busulfan administration.

Protective effect of Sasa veitchii extract against all-trans-retinoic acid-induced inhibition of proliferation of cultured human palate cells.

Cleft palate is the most common facial birth defect worldwide. It is caused by environmental factors or genetic mutations. Environmental factors such as pharmaceutical exposure in women are known to induce cleft palate. The aim of the present study was to investigate the protective effect of Sasa veitchii extract against medicine-induced inhibition of proliferation of human embryonic palatal mesenchymal cells. We demonstrated that all-trans-retinoic acid inhibited human embryonic palatal mesenchymal cell proliferation in a dose-dependent manner, whereas dexamethasone treatment had no effect on cell proliferation. Cotreatment with Sasa veitchii extract repressed all-trans-retinoic acid-induced toxicity in human embryonic palatal mesenchymal cells. We found that cotreatment with Sasa veitchii extract protected all-trans-retinoic acid-induced cyclin D1 downregulation in human embryonic palatal mesenchymal cells. Furthermore, Sasa veitchii extract suppressed all-trans-retinoic acid-induced miR-4680-3p expression. Additionally, the expression levels of the genes that function downstream of the target genes ( ERBB2 and JADE1 ) of miR-4680-3p in signaling pathways were enhanced by cotreatment with Sasa veitchii extract and all-trans-retinoic acid compared to all-trans-retinoic acid treatment. These results suggest that Sasa veitchii extract suppresses all-trans-retinoic acid-induced inhibition of cell proliferation via modulation of miR-4680-3p expression.

Involvement of microRNA-4680-3p against phenytoin-induced cell proliferation inhibition in human palate cells.

Cleft palate (CP) is one of the most common birth defects and is caused by a combination of genetic and/or environmental factors. Environmental factors such as pharmaceutical exposure in pregnant women are known to induce CP. Recently, microRNA (miRNA) was found to be affected by environmental factors. The aim of the present study was to investigate the involvement of miRNA against phenytoin (PHE)-induced inhibition of proliferation in human embryonic palatal mesenchymal (HEPM) cells. We demonstrated that PHE inhibited HEPM cell proliferation in a dose-dependent manner. We found that treatment with PHE downregulated cyclin-D1 and cyclin-E expressions in HEPM cells. Furthermore, PHE increased miR-4680-3p expression and decreased two downstream genes (ERBB2 and JADE1). Importantly, an miR-4680-3p-specific inhibitor restored HEPM cell proliferation and altered expression of ERBB2 and JADE1 in cells treated with PHE. These results suggest that PHE suppresses cell proliferation via modulation of miR-4680-3p expression.