Diquat causes mouse testis injury through inducing heme oxygenase-1-mediated ferroptosis in spermatogonia.

Diquat dibromide (DQ) is a globally used herbicide in agriculture, and its overuse poses an important public health issue, including male reproductive toxicity in mammals. However, the effects and molecular mechanisms of DQ on testes are limited. In vivo experiments, mice were intraperitoneally injected with 8 or 10 mg/kg/ day of DQ for 28 days. It has been found that heme oxygenase-1 (HO-1) mediates DQ-induced ferroptosis in mouse spermatogonia, thereby damaging testicular development and spermatogenesis. Histopathologically, we found that DQ exposure caused seminiferous tubule disorders, reduced germ cells, and increased sperm malformation, in mice. Reactive oxygen species (ROS) staining of frozen section and transmission electron microscopy (TEM) displayed DQ promoted ROS generation and mitochondrial morphology alterations in mouse testes, suggesting that DQ treatment induced testicular oxidative stress. Subsequent RNA-sequencing further showed that DQ treatment might trigger ferroptosis pathway, attributed to disturbed glutathione metabolism and iron homeostasis in spermatogonia cells in vitro. Consistently, results of western blotting, measurements of MDA and ferrous iron, and ROS staining confirmed that DQ increased oxidative stress and lipid peroxidation, and accelerated ferrous iron accumulation both in vitro and in vivo. Moreover, inhibition of ferroptosis by deferoxamine (DFO) markedly ameliorated DQ-induced cell death and dysfunction. By RNA-sequencing, we found that the expression of HO-1 was significantly upregulated in DQ-treated spermatogonia, while ZnPP (a specific inhibitor of HO-1) blocked spermatogonia ferroptosis by balancing intracellular iron homeostasis. In mice, administration of the ferroptosis inhibitor ferrostatin-1 effectively restored the increase of HO-1 levels in the spermatogonia, prevented spermatogonia death, and alleviated the spermatogenesis disorders induced by DQ. Overall, these findings suggest that HO-1 mediates DQ-induced spermatogonia ferroptosis in mouse testes, and targeting HO-1 may be an effective protective strategy against male reproductive disorders induced by pesticides in agriculture.

Single-cell transcriptomic atlas of goat ovarian aging.

BACKGROUND: The ovaries are one of the first organs that undergo degenerative changes earlier in the aging process, and ovarian aging is shown by a decrease in the number and quality of oocytes. However, little is known about the molecular mechanisms of female age-related fertility decline in different types of ovarian cells during aging, especially in goats. Therefore, the aim of this study was to reveal the mechanisms driving ovarian aging in goats at single-cell resolution. RESULTS: For the first time, we surveyed the single-cell transcriptomic landscape of over 27,000 ovarian cells from newborn, young and aging goats, and identified nine ovarian cell types with distinct gene-expression signatures. Functional enrichment analysis showed that ovarian cell types were involved in their own unique biological processes, such as Wnt beta-catenin signalling was enriched in germ cells, whereas ovarian steroidogenesis was enriched in granulosa cells (GCs). Further analysis showed that ovarian aging was linked to GCs-specific changes in the antioxidant system, oxidative phosphorylation, and apoptosis. Subsequently, we identified a series of dynamic genes, such as AMH, CRABP2, THBS1 and TIMP1, which determined the fate of GCs. Additionally, FOXO1, SOX4, and HIF1A were identified as significant regulons that instructed the differentiation of GCs in a distinct manner during ovarian aging. CONCLUSIONS: This study revealed a comprehensive aging-associated transcriptomic atlas characterizing the cell type-specific mechanisms during ovarian aging at the single-cell level and offers new diagnostic biomarkers and potential therapeutic targets for age-related goat ovarian diseases.

The role of ferroptosis mediated by Bmal1/Nrf2 in nicotine -induce injury of BTB integrity.

Nicotine has shown the toxic effects on male reproductive system, and testicular damage is associated with ferroptosis, which is a non-apoptotic regulated cell death driven by iron-dependent lipid peroxidation. However, the role of nicotine on ferroptosis of testicular cells is largely elusive. In the present study, we showed that nicotine destroyed blood-testis barrier (BTB) by interfering with the circadian rhythm of BTB-related factors (ZO-1, N-Cad, Occludin and CX-43) and induced ferroptosis, as reflected via increased clock-control levels of lipid peroxide and decreased ferritin and GPX4, which involved in the circadian. Inhibition of ferroptosis with Fer-1 alleviated nicotine-induced injury of BTB and impaired sperm in vivo. Mechanically, we uncover that the core molecular clock protein, Bmal1, regulates the expression of Nrf2 via direct E-box binding to its promoter to regulate its activity, and nicotine decreases the transcription of Nrf2 through Bmal1 and inactivates Nrf2 pathway and its downstream antioxidant gene, which leads to the imbalance of redox state and ROS accumulation. Intriguingly, nicotine induced lipid peroxidation and subsequent ferroptosis by Bmal1-mediated Nrf2. In conclusion, our study reveals a clear role for the molecular clock in controlling Nrf2 in testis to mediate the ferroptosis induced by nicotine. These findings provide a potential mechanism to prevent smoking and/or cigarette smoke-induced male reproductive injury.

Melatonin Ameliorates Diquat-Induced Testicular Toxicity via Reducing Oxidative Stress, Inhibiting Apoptosis, and Maintaining the Integrity of Blood-Testis Barrier in Mice.

Diquat is a fast, potent, and widely used bipyridine herbicide in agriculture and it induces oxidative stress in several animal models. However, its genotoxic effects on the male reproductive system remain unclear. Melatonin is an effective free-radical scavenger, which has antioxidant and anti-apoptotic properties and can protect the testes against oxidative damage. This study aimed to investigate the therapeutic effects of melatonin on diquat-induced testicular injury in mice. The results showed melatonin treatment alleviated diquat-induced testicular injury, including inhibited spermatogenesis, increased sperm malformations, declined testosterone level and decreased fertility. Specifically, melatonin therapy countered diquat-induced oxidative stress by increasing production of the antioxidant enzymes GPX1 and SOD1. Melatonin treatment also attenuated diquat-induced spermatogonia apoptosis in vivo and in vitro by modulating the expression of apoptosis-related proteins, including P53, Cleaved-Caspase3, and Bax/Bcl2. Moreover, melatonin restored the blood-testicular barrier by promoting the expression of Sertoli cell junction proteins and maintaining the ordered distribution of ZO-1. These findings indicate that melatonin protects the testes against diquat-induced damage by reducing oxidative stress, inhibiting apoptosis, and maintaining the integrity of the blood-testis barrier in mice. This study provides a theoretical basis for further research to protect male reproductive health from agricultural pesticides.

BMP6 regulates AMH expression via SMAD1/5/8 in goat ovarian granulosa cells.

Anti-Müllerian hormone (AMH) is produced by ovarian granulosa cells (GCs)and plays a major role in inhibiting the recruitment of primordial follicles and reducing the sensitivity of growing follicles to follicle-stimulating hormone (FSH). Bone morphogenetic protein 6 (BMP6) has similar spatiotemporal expression to AMH during follicular development, suggesting that BMP6 may regulate AMH expression. However, the specific mechanism by which BMP6 regulates AMH expression remains unclear. The objectives of this study were to examine the molecular pathway by which BMP6 regulates AMH expression. The results showed that BMP6 promoted the secretion and expression of AMH in goat ovarian GCs. Mechanistically, BMP6 upregulated the expression of sex-determining region Y-box 9 (SOX9) and GATA-binding factor 4 (GATA4), which was associated with the transcriptional initiation of AMH. AMH expression was significantly decreased by GATA4 knockdown. Moreover, BMP6 treatment promoted the phosphorylation of SMAD1/5/8, whereas inhibiting the SMAD1/5/8 signaling pathway significantly abolished BMP6-induced upregulation of AMH and GATA4 expression. Interestingly, the activation of SMAD1/5/8 alone did not affect the expression of AMH or GATA4. The results suggested that BMP6 upregulated GATA4 through the SMAD1/5/8 signaling pathway, which in turn promoted AMH expression.

Solid-Liquid Phase Equilibrium of Ammonium Dihydrogen Phosphate and Agricultural Grade Ammonium Polyphosphate (Degree of Polymerization Ranging from 1 to 8) for Mixed Irrigation Strategy.

Water-soluble ammonium polyphosphate (APP) has the advantages of good solubility and slow-release characteristics and has the potential to be used in combination with monoammonium phosphate (MAP) as a high phosphorus content slow-release fertilizer to improve the utilization rate of phosphorus during irrigation. Herein, the effects of the APP1 concentration and temperature (278.2-313.2 K) on the solubility of MAP, solution density, and pH value in the ternary equilibrium system (APP1-MAP-water) were measured. The simplified Apelblat model, two empirical polynomials, and rational two-dimensional functions can describe the experimental solubility data, solution density, and pH value well, respectively, with reliable modeling parameters (R 2 > 0.99). In the OptiMax1001 reactor, the focused beam reflectance measurement (FBRM), the particle-view measurement (PVM), and the ReactIR 15 probes were used to observe and reverse verify that they can be synergistically codissolved to achieve economic efficiency. Basic thermodynamic data and models can guide their collaborative application in irrigation to improve the phosphorus utilization rate.

BMP6 Promotes the Secretion of 17 Beta-Estradiol and Progesterone in Goat Ovarian Granulosa Cells.

The purpose of this study was to investigate the effects of BMP6 on the function of goat ovarian granulosa cells (GCs). The results showed that the exogenous addition of BMP6 did not affect the EdU-positive ratio of ovarian GCs and had no significant effect on the mRNA and protein expression levels of the proliferation-related gene PCNA (p > 0.05). Meanwhile, BMP6 had no significant effect on the cycle phase distribution of GCs but increased the mRNA expression of CDK4 (p < 0.05) and CCND1 (p < 0.01) and decreased the mRNA expression of CCNE1 (p < 0.01). Moreover, BMP6 had no significant effect on the apoptosis rate of GCs and did not affect the mRNA expression levels of apoptosis-related genes BAX, BCL2, and Caspase3 (p > 0.05). Importantly, BMP6 upregulated the secretion of 17 beta-estradiol (E2) and progesterone (P4) in ovarian GCs (p < 0.01). Further studies found that BMP6 inhibited the mRNA expression of 3ß-HSD and steroid synthesis acute regulator (StAR) but significantly promoted the mRNA expression of the E2 synthesis rate-limiting enzyme CYP19A1 and the P4 synthesis rate-limiting enzyme CYP11A1 (p < 0.01). Taken together, these results showed that the exogenous addition of BMP6 did not affect the proliferation, cell cycle, and apoptosis of goat ovarian GCs but promoted the secretion of E2 and progesterone P4 in ovarian GCs by upregulating the mRNA expressions of CYP19A1 and CYP11A1.

Liver Receptor homolog-1 Regulates Apoptosis of Bovine Ovarian Granulosa Cells by Progestogen Receptor Signaling Pathway.

The purpose of the present investigation was to assess the function of LRH-1 on GCs and the mechanisms involved. Here, LRH- was highly expressed in the bovine GCs of atretic follicles. Treatment with 50 µM of LRH-1 agonist (DLPC) significantly induced the expression of LRH-1 (p < 0.05). In particular, LRH-1 activation blocked the progestogen receptor signaling pathway via downregulating progesterone production and progestogen receptor levels (p < 0.05), but had no effect on 17 beta-estradiol synthesis. Meanwhile, LRH-1 activation promoted the apoptosis of GCs and increased the activity of caspase 3 (p < 0.05). Importantly, upregulating the progestogen receptor signaling pathway with progestogen could attenuate the LRH-1-induced proapoptotic effect. Moreover, treatment with progestogen decreased the activity of the proapoptotic gene caspase 3 and increased the expression of antiapoptotic gene Bcl2 in LRH-1 activated GCs (p < 0.05). Taken together, these results demonstrate that LRH-1 might be dependent on the progestogen receptor signaling pathway to modulate bovine follicular atresia.

17ß-Estradiol Enhances Porcine Meiosis Resumption from Autophagy-Induced Gap Junction Intercellular Communications and Connexin 43 Phosphorylation via the MEK/ERK Signaling Pathway.

Estrogen and its analogues are ubiquitous in agricultural environments, with large biological functions of oocyte development. Gap junction intercellular communications (GJICs) are the structural basis in cumulus-oocyte complexes (COCs) and regulate oocyte maturation and developmental material transport through a number of pathways. This study mainly determines the effect and potential mechanism of estrogen (17ß-estradiol) in regulating GJICs in porcine COCs. In our study, 17ß-estradiol increased porcine nuclear maturation in a time-dependent manner. The analysis revealed that 17ß-estradiol upregulated the autophagy in COCs during in vitro maturation. In contrast with the control, 17ß-estradiol decreased GJICs in a time-dependent manner between cumulus cells and oocytes, while it was consistent with the control group at 24 h. Carbenoxolone (CBX) blocks GJICs as a negative control group used in our system. Autophagy inhibitor autophinib decreased oocyte maturation, and the reduced nuclear maturation treated with autophinib was abolished by 17ß-estradiol. Besides, the upregulation effect of autophinib on GJICs and transzonal projections (TZPs) was decreased by 17ß-estradiol. 17ß-Estradiol could reduce serine 368 phosphorylation of connexin 43 (Cx43) protein by autophinib in porcine COCs. These results were dependent upon the MEK/ERK signaling pathway. Furthermore, 17ß-estradiol-induced GJICs and Cx43 phosphorylation were inhibited by autophinib or the MEK/ERK pathway inhibitors (Trametinib and FR 180204), indicating that 17ß-estradiol regulated GJICs through the MEK/ERK signaling pathway. In conclusion, 17ß-estradiol improves the autophagy-mediated nuclear maturation with downregulating GJICs and TZPs in porcine COCs. Such an effect occurs by phosphorylation of Cx43, which was regulated via the MEK/ERK signaling pathway.

Isorhamnetin Promotes Estrogen Biosynthesis and Proliferation in Porcine Granulosa Cells via the PI3K/Akt Signaling Pathway.

Isorhamnetin is a natural flavonoid widely distributed in fruits and vegetables. However, the roles of isorhamnetin involved in steroidogenesis, proliferation, and apoptosis in ovarian granulosa cells (GCs) are poorly understood. We found that isorhamnetin promoted the secretion of estrogen and inhibited the secretion of progesterone and testosterone by modulating steroidogenesis-associated proteins and mRNA such as CYP19A1, StAR, and 3ß-HSD in ovarian GCs. Mechanistically, isorhamnetin stimulated the expression of the proliferating cell nuclear antigen and C-myc and promoted the proliferation of GCs via the PI3K/Akt signaling pathway. Furthermore, isorhamnetin increased the protein expression of CyclinB, CyclinD, CyclinE, and CyclinA, thereby raising the ratio of S-phase cells in response to GC proliferation. Changes in the expression of apoptosis-associated proteins (Bcl2, Bax, and cytochrome c) and intracellular reactive oxygen species levels showed that isorhamnetin inhibited GC apoptosis. Collectively, these findings indicate that isorhamnetin regulates steroidogenesis through the activation of PI3K/Akt, which promotes proliferation, inhibits apoptosis, and alleviates oxidative stress.

17ß-estradiol improves the developmental ability, inhibits reactive oxygen species levels and apoptosis of porcine oocytes by regulating autophagy events.

OBJECTIVES: Estrogen plays a critical role in the development and apoptosis of oocytes. Autophagy is an evolutionarily conserved and exquisitely regulated self-eating cellular process with important biological functions including the regulation of reproduction. This study aimed to determine the effect of autophagy regulated by the biologically active form of estrogen (17ß-estradiol) in porcine oocyte maturation in vitro. MATERIALS AND METHODS: We measured the effects of oocyte developmental competencies and autophagic activity in the porcine oocyte regulated by 17ß-estradiol using autophagic inhibitor (Autophinib). In addition, we studied the role of autophagy in reactive oxygen species (ROS) levels, mitochondrial distribution, Ca2+ production, mitochondrial membrane potential (ΔΨm), and early apoptosis by caspase-3, -8 activity in the mature oocytes. RESULTS: The results showed that the oocyte meiotic progression and early embryonic development were gradually decreased with Autophinib treatment, which was improved by 17ß-estradiol. Immunofluorescence experiments revealed that 17ß-estradiol primarily could promote the autophagy in the mature oocytes, and block the reduced-autophagic events by Autophinib. Moreover, 17ß-estradiol improved the Autophinib induced high ROS levels, abnormal mitochondrial distribution and low Ca2+ production in mature oocytes. Analyses of early apoptosis and ΔΨm showed that autophagy inhibition was accompanied by increased cellular apoptosis, and 17ß-estradiol reduced apoptosis rates of mature oocytes. Importantly, autophagy was downregulated by treatment with Autophinib, an activation of caspase-8 and cleaved caspase-3 increased. Those effects were abolished by 17ß-estradiol, which could upregulate autophagy. CONCLUSIONS: Our study have showed important implications that 17ß-estradiol could promote efficacy of the development of porcine oocytes, enhance the autophagy, reduce ROS levels and apoptosis activity in vitro maturation.

Development and characterization of in vitro self-assembled recombinant human follicle stimulating hormone originated from goat mammary epithelial cells.

Follicle stimulating hormone (FSH), composed of FSHα and FSHß subunits, is essential for female follicle development and male spermatogenesis. The recombinant human FSH (rhFSH) products on the market are mainly generated from mammalian cells and are expensive. Large animal mammary gland bioreactors are urgently needed to produce large amounts of rhFSH. However, there are currently no effective methods to prepare rhFSH by large animals mainly due to the fact that excessive accumulation of FSH might cause many adverse effects in animals. We herein report the development and characterization of functional self-assembled rhFSH produced in goat mammary epithelial cells (GMECs). FSHα and FSHß stably expressed in Chinese hamster ovary (CHO) cell lines were secreted into culture medium and well glycosylated. Importantly, FSHα and FSHß expressed apart were able to assemble into functional FSH. We next inserted human FSHα or FSHß gene separately into goat ß-Lactoglobulin locus in GMECs by CRISPR/Cas9. Inactive FSHα and FSHß subunits expressed from GMECs assembled into rhFSH as analyzed by His-tag pull down assay. Functional assessment of rhFSH by cAMP induction assay, mouse ovulation induction and rat ovarian weight gain experiments showed that the bioactivity of self-assembled rhFSH expressed by GMECs was comparable to that of Gonal-F both in vitro and in vivo. Our study demonstrated that FSHα and FSHß can be separately expressed and assembled into functional rhFSH, and provided the basis for future preparing FSH by goat mammary gland bioreactor with less health problems on the producing animals.

Electric field assisted benzene oxidation over Pt-Ce-Zr nano-catalysts at low temperature.

A novel catalytic system for benzene oxidation at low temperature is constructed by combining electric field with Pt-Ce-Zr nano-catalyst. The 1 wt% Pt/Ce0.75Zr0.25O2 catalyst assisted by electric field shows the best catalytic performance with 90% benzene conversion at 96.5 °C and excellent water resistance. The effect of electric field on catalysts and catalytic process is comprehensively investigated. The results of XRD, TEM, XPS and H2-TPR reveal that the electric field show negligible influence on the crystal structure and surface morphology of the catalyst, but it can lead to more oxygen vacancies. Therefore, more adsorbed oxygen with higher activity will be produced on the catalyst surface. The redox performance is improved due to the fact that valence distribution of Pt is changed in forms of more active sites composed of high valence oxides (PtO) generated in electric field. In situ DRIFTS is used to investigate the oxidation process of benzene and the results prove that electric field could accelerate the production and consumption of intermediate products, and produce new intermediate products such as carboxylic acid species, indicating that the introduction of electric field may open up a new rapid reaction path and promote the activation of benzene at low temperature.

WITHDRAWN: Transcriptional regulation of miR-483-3p mediated by IL-6/STAT3 axis promoted epithelial-mesenchymal transition and tumor stemness in glioma.

This article was originally published in Aging Advance Online Publications on November 4, 2020. Concerns were raised at that time about overlap between data in Figures 1D, 8A and 8B of this article and Figures in an unrelated retracted article (https://www.dovepress.com/cr_data/article_fulltext/s268000/268222/img/CMAR_A_268222_O_F0007g.jpg, (https://doi.org/10.2147/CMAR.S268222). The article was investigated, and the editorial decision was made to withdraw the article entirely in compliance with Aging's withdrawal policy. The institution affiliated with the published Aging article was notified about this action. This article will not appear internally in Aging or any external indexes or archives.

Melatonin protects mouse testes from palmitic acid-induced lipotoxicity by attenuating oxidative stress and DNA damage in a SIRT1-dependent manner.

Palmitic acid (PA), the main component of dietary saturated fat, has been known to increase in patients with obesity, and PA-induced lipotoxicity may contribute to obesity-related male infertility. Melatonin has beneficial effects on reproductive processes; however, the effect and the underlying molecular mechanism of melatonin's involvement in PA-induced cytotoxicity in the testes are poorly understood. Our findings showed that lipotoxicity was observed in mouse testes after long-term PA treatment and that melatonin therapy restored spermatogenesis and fertility in these males. Moreover, melatonin therapy suppressed PA-induced apoptosis by modulating apoptosis-associated proteins such as Bcl2, Bax, C-Caspase3, C-Caspase12, and CHOP in type B spermatogonial stem cells. Changes in the expression of endoplasmic reticulum (ER) stress markers (p-IRE1, p-PERK, ATF4) and intracellular Ca2+ levels showed that melatonin relieved PA-induced ER stress. Mechanistically, melatonin stimulated the expression and nuclear translocation of SIRT1 through its receptors and prevented PA-induced ROS production and mitochondrial dysfunction via SIRT1 signaling pathway. Furthermore, melatonin promoted SIRT1-mediated p53 deacetylation, thereby relieving G2/M arrest in response to PA-stimulated DNA damage. Collectively, these findings indicate that melatonin protects the testes from PA-induced lipotoxicity through the activation of SIRT1, which alleviates oxidative stress, ER stress, mitochondrial dysfunction, and DNA damage.

Rutin protects boar sperm from cryodamage via enhancing the antioxidative defense.

This study was aimed to investigate whether and how Rutin protects boar sperm against cryoinjury during cryopreservation. Five concentrations of Rutin with 0.2, 0.4, 0.6, 1.0, and 2.0 mM were added to the freezing extender of boar sperm, respectively, and the effects on quality and function of boar sperm after freezing-thawing were assessed. The results showed that the sperm motility, mitochondrial activity, plasma membrane integrity, and acrosomal integrity were significantly improved in 0.4 mM and 0.6 mM Rutin groups (p < .05). Compared with ganoderma lucidum polysaccharide (GLP) or Tanshinone IIA, Rutin exhibited higher rates of mitochondrial activity and acrosome integrity (p < .05). Mechanistically, the addition of Rutin at the concentration of 0.6, 0.8, and 1.0 mM significantly attenuated ROS accumulation and MDA production by improving antioxidant enzymatic activity, including SOD, CAT, and GSH-Px (p < .05). Functionally, a higher penetration rate and the increased total efficiency of fertilization were observed in the 0.4, 0.6, and 1.0 mM Rutin groups than in the control group (p < .05). Moreover, the addition of Rutin (0.6 mM) significantly induced an increase in both the cleavage and blastocyst rates (p < .05). In summary, supplementation with Rutin in cryopreservation medium protects boar sperm against ROS attack by enhancing the antioxidative defense.

A novel role of SIRT2 in regulating gap junction communications via connexin-43 in bovine cumulus-oocyte complexes.

SIRT2, the predominantly cytosolic sirtuin, plays important role in multiple biological processes, including metabolism, stress response, and aging. However, the function of SIRT2 in gap junction intercellular communications (GJICs) of cumulus-oocyte complexes (COCs) is not yet known. The purpose of the present study was to evaluate the effect and underlining mechanism of SIRT2 on GJICs in COCs. Here, we found that treatment with SIRT2 inhibitors (SirReal2 or TM) inhibited bovine oocyte nuclear maturation. Further analysis revealed that SIRT2 inactivation disturbed the GJICs of COCs during in vitro maturation. Correspondingly, both the Cx43 phosphorylation levels and MEK/MER signaling pathways were induced by SIRT2 inhibition. Importantly, SIRT2-mediated Cx43 phosphorylation was completely abolished by treatment with MEK1/2 inhibitor (Trametinib). Furthermore, treatment with SIRT2 inhibitors resulted in the high levels of MEK1/2 acetylation. Functionally, downregulating the MER/ERK pathways with inhibitors (Trametinib or SCH772984) could attenuate the closure of GJICs caused by SIRT2 inactivation in partly. In addition, inhibition of SIRT2 activity significantly decreased the membrane and zona pellucida localization of Cx43 by upregulating the levels of Cx43 acetylation. Taken together, these results demonstrated a novel role that SIRT2 regulates GJICs via modulating the phosphorylation and deacetylation of Cx43 in COCs.

LncRNA MEG3 inhibits HMEC-1 cells growth, migration and tube formation via sponging miR-147.

Long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) has been identified as a regulatory molecule in angiogenesis. The goal of this study was to illustrate how MEG3 affects the angiogenesis of vascular endothelial cells. Expression of MEG3, miR-147 and intracellular cell adhesion molecule-1 (ICAM-1) in human microvascular endothelial cell line (HMEC-1) was altered by transfection, then cell viability, apoptosis, migration, tube formation, as well as the correlation among MEG3, miR-147 and ICAM-1 were explored. MEG3 was down-regulated during tube formation of HMEC-1 cells. MEG3 expression suppressed cells viability, migration and tube formation, while it induced apoptosis. MEG3 could bind with miR-147 and repress miR-147 expression. MiR-147 induced ICAM-1 expression, and contained ICAM-1 target sequences. The anti-atherogenic actions of MEG3 were inhibited by miR-147, and the anti-atherogenic actions of miR-147 suppression were also inhibited when ICAM-1 was overexpressed. Further, ICAM-1 overexpression showed activated roles in Wnt/ß-catenin and Jak/Stat signaling pathways. In low-density lipoprotein receptor (Ldlr)-/- mice, MEG3 overexpression reduced CD68+, CD3+ and ICAM-1 areas in lesions and increased collagen content. MEG3 inhibited HMEC-1 cell growth, migration and tube formation. The anti-atherogenic actions of MEG3 might be mediated via sponging miR-147, and thereby repressing the expression of ICAM-1.

Fibrinogen, Neutrophil-to-Lymphocyte Rate and Platelet-to-Neutrophil Rate as Novel Acute Phase Indicators in Patients with Thromboangiitis Obliterans.

BACKGROUND/OBJECTIVES: The acute exacerbations and progressive deterioration seen in thromboangiitis obliterans (TAO) have been related to poor clinical outcomes. Here, we have studied the association of laboratory biomarkers with the acute phase of TAO (AP-TAO). METHODS/RESULTS: We conducted a retrospective case-control study on 112 patients with TAO and 98 healthy controls; comparing the neutrophil-to-lymphocyte rate (NLR), lymphocyte-to-monocyte rate (LMR), platelet-to-neutrophil rate (PNR), fibrinogen (FIB), and apolipoprotein A-I (ApoA-I). Significantly higher NLR level, as well as lower LMR, PNR, and ApoA-I levels were observed in patients with TAO, particularly the acute phase. Significantly increased FIB was only observed in AP-TAO. A positive correlation was found between NLR and with C-reactive protein (CRP) in the acute phase (r = 0.817, P < 0.001). Moreover, NLR, PNR, and FIB levels of 3.38, 45.12, and 3.69 were shown to be the predictive cut-off values for the AP-TAO (sensitivity 72.5, 82,4, and 66,7%, specificity 92.2, 78.4, and 96.1%; area under the curve [AUC] 0.875, 0.855, and 0.872), respectively. The FIB level was independently associated with the AP-TAO (OR = 11.420, P = 0.007). CONCLUSIONS: NLR, PNR, and FIB may be useful markers for the identification of inflammation and the AP-TAO. FIB may be an independent risk factor for the acute phase.