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Abnormal expression of long non-coding RNAs (lncRNAs) is associated with the dysfunctions of human trophoblast cells and the occurrence of miscarriage (abnormal early embryo loss). BBC3/PUMA (BCL2 binding component 3) plays significant roles in regulation of cell apoptosis. However, whether specific lncRNAs might regulate BBC3 in trophoblast cells and further induce apoptosis and miscarriage remains completely unclear. Through screening, we identified a novel lnc-HZ12, which was significantly highly expressed in villous tissues of recurrent miscarriage (RM) patients relative to their healthy control (HC) group. Lnc-HZ12 suppressed chaperone-mediated autophagy (CMA) degradation of BBC3, promoted trophoblast cell apoptosis, and was associated with miscarriage. In mechanism, lnc-HZ12 downregulated the expression levels of chaperone molecules HSPA8 and LAMP2A in trophoblast cells. Meanwhile, lnc-HZ12 (mainly lnc-HZ12-SO2 region in F2 fragment) and HSPA8 competitively bound with the 169RVLYNL174 patch on BBC3, which prevented BBC3 from interactions with HSPA8 and impaired the formation of BBC3-HSPA8-LAMP2A complex for CMA degradation of BBC3. Thus, lnc-HZ12 upregulated the BBC3-CASP9-CASP3 pathway and induced trophoblast cell apoptosis. In villous tissues, lnc-HZ12 was highly expressed, CMA degradation of BBC3 was suppressed, and the apoptosis levels were higher in RM vs HC villous tissues, all of which were associated with miscarriage. Interestingly, knockdown of murine Bbc3 could efficiently suppress placental apoptosis and alleviate miscarriage in a mouse miscarriage model. Taken together, our results indicated that lnc-HZ12 and BBC3 played important roles in trophoblast cell apoptosis and miscarriage and might act as attractive targets for miscarriage treatment. Abbreviation: 7-AAD: 7-aminoactinomycin D; BaP: benzopyrene; BBC3/PUMA: BCL2 binding component 3; ChIP: chromatin immunoprecipitation; CHX: cycloheximide; CMA: chaperone-mediated autophagy; CQ: chloroquine; DMSO: dimethyl sulfoxide; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HC: healthy control; HSPA8: heat shock protein family A (Hsp70) member 8; IP: immunoprecipitation; LAMP2A: lysosomal associated membrane protein 2; LncRNA: long non-coding RNA; mRNA: messenger RNA; MT: mutant-type; NC: negative control; NSO: nonspecific oligonucleotide; PARP1: poly(ADP-ribose) polymerase 1; RIP: RNA immunoprecipitation; RM: recurrent miscarriage; TBP: TATA-box binding protein; WT: wild-type. [ABSTRACT FROM AUTHOR]

Human trophoblast cells are crucial for healthy pregnancy. However, whether the defective homologous recombination (HR) repair of dsDNA break (DSB) in trophoblast cells may induce miscarriage is completely unknown. Moreover, the abundance of BRCA1 (a crucial protein for HR repair), its recruitment to DSB foci, and its epigenetic regulatory mechanisms, are also fully unexplored. In this work, it is identified that a novel lnc‐HZ10, which is highly experssed in villous tissues of recurrent miscarriage (RM) vs their healthy control group, suppresses HR repair of DSB in trophoblast cell. Lnc‐HZ10 and AhR (aryl hydrocarbon receptor) form a positive feedback loop. AhR acts as a transcription factor to promote lnc‐HZ10 transcription. Meanwhile, lnc‐HZ10 also increases AhR levels by suppressing its CUL4B‐mediated ubiquitination degradation. Subsequently, AhR suppresses BRCA1 transcription; and lnc‐HZ10 (mainly 1‐447 nt) interacts with γ‐H2AX; and thus, impairs its interactions with BRCA1. BPDE exposure may trigger this loop to suppress HR repair in trophoblast cells, possibly inducing miscarriage. Knockdown of murine Ahr efficiently recovers HR repair in placental tissues and alleviates miscarriage in a mouse miscarriage model. Therefore, it is suggested that AhR/lnc‐HZ10/BRCA1 axis may be a promising target for alleviation of unexplained miscarriage. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) derived from parental cells could communicate with neighboring or distant recipient cells. The components in EVs, especially non-coding RNAs, including microRNAs, long non-coding RNAs, and circular RNAs, could regulate the functions of the recipient cells. Meanwhile, EVs could also be used as valuable biomarkers and drug delivery carriers. Moreover, environmental toxicants may alter EVs components and regulate EVs-mediated pathogenesis of various diseases. In this review, we mainly summarized the roles of EV-derived non-coding RNAs in the regulation of cell dysfunctions in various adverse pregnancy outcomes, such as preeclampsia (PE), gestational diabetes mellitus (GDM), and miscarriage. Moreover, the effects of environmental toxicants on the components and functions of EVs, as well as on their regulatory roles in these diseases, were also discussed. [ABSTRACT FROM AUTHOR]

Strand displacement DNA synthesis is essential for DNA replication. Gp90, the sole DNA polymerase of Pseudomonas aeruginosa phage 1, can bypass multiply DNA lesions. However, whether it can perform strand displacement synthesis is still unknown. In this work, we found that gp90 exo― could perform strand displacement synthesis, albeit its activity and processivity were lower than those of primer extension. Gp90 exo― itself could not unwind Y-shaped or fork DNA. Tail and gap at DNA fork were necessary for efficient synthesis. High GC content obviously inhibited strand displacement synthesis. Consecutive GC sequence at the entrance of fork showed more inhibition effect on DNA synthesis than that in the downstream DNA fork. The fraction of productive polymerase and DNA complex (A values) was higher for fork than gap; while their average extension rates (k p values) were similar. However, both A and k p values were lower than those for the primer/template (P/T) substrate. The binding of gp90 exo― to fork was tighter than P/T or gap in the absence of dATP. In the presence of dATP to form ternary complex, the binding affinity of gp90 exo― to P/T or gap was increased compared with that in the binary complex. Abasic site, 8-oxoG, and O 6-MeG inhibited and even blocked strand displacement synthesis. This work shows that gp90 exo― could perform strand displacement DNA synthesis at DNA fork, discovering the presence of new functions of PaP1 DNA polymerase in DNA replication and propagation of PaP1. • Gp90 exo― could perform strand displacement DNA synthesis. • Tail and gap at DNA fork were necessary for efficient synthesis. • High GC content and DNA lesions obviously inhibited strand displacement synthesis. • E. coli Pol I KF exo― and Pol T7 ― could perform this synthesis, but Dpo4 and hPol ι could nearly not. • This work discovers new functions of polymerase in DNA replication and propagation of PaP1. [ABSTRACT FROM AUTHOR]

Long non‐coding RNA (lncRNA) is one of the important regulators of many malignancies. However, the biological function and clinical significance of a large number of lncRNAs in gastric cancer remain unclear. Therefore, we analysed the TCGA data to find that LINC01303 is significantly up‐regulated in gastric cancer tissues. However, the biological function of LINC01303 in GC remains unknown. In our study, we found that the expression of LINC01303 was significantly higher in GC tissues than in adjacent tissues by real‐time quantitative PCR. We can significantly inhibit the malignant proliferation, migration and invasion of GC cells by silencing LINC01303 expression. In addition, LINC01303 knockdown can also inhibit GC growth in vivo. After the bioinformatics analysis, we found that LINC01303 can be used as a miR‐101‐3p sponge to competitively adsorb miR‐101‐3p with EZH2. Therefore, our results indicate that LINC01303 promotes the expression of EZH2 by inhibiting miR‐101‐3p activity and promotes GC progression. In summary, in this study, we demonstrated for the first time that the LINC01303/miR‐101‐3p/EZH2 axis promotes GC progression. [ABSTRACT FROM AUTHOR]

Proper migration and invasion of trophoblast cells into endometrium is vital for successful embryo implantation during early pregnancy. Benzo[a]pyrene-7, 8-diol-9, 10-epoxide (BPDE) is an ultimate carcinogenic product of benzo[a]pyrene (BaP), which causes multiple trophoblast-related diseases. However, the mechanism of BPDE-inhibited migration/invasion of trophoblast cells is still unclear. In this work, we found that BPDE significantly inhibited the filopodia formation and migration/invasion of human trophoblast Swan 71 cells. BPDE up-regulated the level of miR-194-3p, which further inhibited the phosphoinositide 3-kinase (PI3K)/AKT/ cell division cycle 42/ p21 (RAC1) activated kinase 1 signaling pathway and depressed the filophdia formation of Swan71 cells. Addition of 740 Y-P, the activator of phosphoinositide 3-kinase, could stimulate cell migration/invasion, confirming the involvement of this pathway. Knock-down of miR-194-3p up-regulated this pathway and promoted filopodia formation and migration/invasion. Conversely, overexpression of miR-194-3p down-regulated this pathway and inhibited cell migration/invasion. Therefore, miR-194-3p takes important roles in the BPDE-inhibited filopodia formation and cell migration/invasion, providing valuable information in the BPDE-induced dysfunctions of human extravillous trophoblast cells. [ABSTRACT FROM AUTHOR]

Human trophoblast cells play important role in embryo-fetal development. However, trophoblast cells are sensitive to environmental carcinogens. Recently, we have discovered that BPDE inhibits trophoblast cell migration, invasion and proliferation, and also increases trophoblast cell apoptosis. Ferroptosis is a newly identified iron-dependent non-apoptotic programmed cell death. Whether BPDE might induce trophoblast cell ferroptosis is completely unknown. In this work, we have discovered that ferroptosis does occur in BPDE-treated human trophoblast cells. Iron metabolism up-regulates intracellular free Fe2+ level, produces excessive ROS (reactive oxygen species), and accelerates lipid peroxidation. GPX4 efficiently eliminates ROS and inhibits lipid peroxidation. Thus, ferroptosis is well suppressed due to the balance of these two pathways in healthy trophoblast cells. However, BPDE exposure could promote iron metabolism, increase intracellular free Fe2+ level, produce excessive ROS, and result in lipid peroxidation. Meanwhile, BPDE also down-regulates GPX4 expression level by promoting its proteasomal degradation, which eventually produces excessive ROS and induces lipid peroxidation. Thus, BPDE exposure induces trophoblast cell ferroptosis by unbalancing these two pathways, providing new insight in understanding BPDE-induced dysfunctions of human trophoblast cells. • BPDE exposure induces human trophoblast cell ferroptosis. • BPDE exposure increases intracellular free Fe2+ level and ROS level. • BPDE promotes GPX4 proteasomal degradation and reduces its protein level. • BPDE produces excessive ROS and induces lipid peroxidation. [ABSTRACT FROM AUTHOR]

rNTPs are structurally similar to dNTPs, but their concentrations are much higher than those of dNTPs in cells. rNTPs in solutions or rNMP at the primer terminus or embedded in template always inhibit or block DNA replication, due to the reduced Mg2+ apparent concentration, competition of rNTPs with dNTPs, and the extra repulsive interaction of rNTP or rNMP with polymerase active site. In this work, unexpectedly, we found rNTPs can promote T7 DNA replication with the maximal promotion at rNTPs/dNTPs concentration ratio of 20. This promotion was not due to the optimized Mg2+ apparent concentration or the direct incorporation of extra rNMPs into DNA. This promotion was dependent on the concentrations and types of rNTPs. Kinetic analysis showed that this promotion was originated from the increased fraction of polymerase-DNA productive complex and the accelerated DNA polymerization. Further evidence showed that more polymerase-DNA complex was formed and their binding affinity was also enhanced in the presence of extra rNTPs. Moreover, this promotion in T7 DNA replication also accelerated the lysis of T7-infected host Escherichia coli. This work discovered that rNTPs could promote DNA replication, completely different from the traditional concept that rNTPs always inhibit DNA replication. • Appropriate concentration ratios of rNTPs/dNTPs can promote T7 DNA replication. • rNTPs increased the amount of polymerase-DNA productive complex and the accelerated DNA polymerization. • More polymerases-DNA complex was formed and their binding affinity was enhanced in the presence of extra rNTPs. • This promotion for T7 DNA replication accelerated the lysis of its host E. coli infected with T7. [ABSTRACT FROM AUTHOR]

Background: Recurrent miscarriage (RM) affects 1%-3% of pregnancies. However, in almost 50% of cases, the cause is unknown. Increasing evidence have shown that benzo(a)pyrene [B(a)P], a representative of polycyclic aromatic hydrocarbons (PAHs), is correlated with miscarriage. However, the underlying mechanisms of B(a)P/benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE)-induced trophoblast cell dysfunctions and miscarriage remain largely unknown., Objective: The objective was to discover the role(s) of a novel lncRNA, lnc-HZ09 , in the regulation of BPDE-inhibited migration and invasion of trophoblast cells and the occurrence of miscarriage., Method: Human trophoblast cells were treated with 0, 0.25, 0.5, 1.0 ,  or  1.5 μ M BPDE with or without corresponding lnc-HZ09 silencing or overexpression. Using these cells, we evaluated cell migration and invasion, the mRNA and protein levels of members of the PLD1/RAC1/CDC42 pathway, the regulatory roles of lnc-HZ09 in PLD1 transcription and mRNA stability, and lnc-HZ09 transcription and stability. Human villous tissues were collected from RM ( n = 15 ) group and their matched healthy control (HC, n = 15 ) group. We evaluated the levels of BPDE-DNA adducts, lnc-HZ09, and the mRNA and protein expression of members of the PLD1/RAC1/CDC42 pathway, and correlated their relative expression levels. We further constructed 0, 0.05 or 0.2 mg / kg B(a)P-induced mouse miscarriage model (each n = 6 ), in which the mRNA and protein expression of members of the Pld1/Rac1/Cdc42 pathway were measured., Results: We identified a novel lnc-HZ09 . Human trophoblast cells treated with lnc-HZ09 exhibited less cell migration and invasion. In addition, the levels of this lncRNA were higher in villous tissues from women with recurrent miscarriage than those from healthy individuals. SP1-mediated PLD1 mRNA levels were lower, and HuR-mediated PLD1 mRNA stability was less in trophoblast cells overexpressing lnc-HZ09 . However, trophoblast cells treated with MSX1 had higher levels of lnc-HZ09 , and METTL3-mediated m6A methylation on lnc-HZ09 resulted in greater lnc-HZ09 RNA stability. In BPDE-treated human trophoblast cells and in RM villous tissues, MSX1-mediated lnc-HZ09 transcription and METTL3-mediated lnc-HZ09 stability were both greater. In our mouse miscarriage model, B(a)P-treated mice had lower mRNA and protein levels of members of the Pld1/Rac1/Cdc42 pathway., Discussion: These results suggest that in human trophoblast cells, BPDE exposure up-regulated lnc-HZ09 level, suppressed PLD1/RAC1/CDC42 pathway, and inhibited migration and invasion, providing new insights in understanding the causes and mechanisms of unexplained miscarriage. https://doi.org/10.1289/EHP10477.

Normal pregnancy is essential for human reproduction. However, BaP (benzo(a)pyrene) and its metabolite BPDE (benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide) could cause dysfunctions of human trophoblast cells and might further induce miscarriage. Yet, the underlying mechanisms remain largely unknown. Herein, we identified a novel upregulated lnc-HZ04 and a novel downregulated miR-hz04 in villous tissues of unexplained recurrent miscarriage (RM) relative to those in healthy control tissues and also in BPDE-treated human trophoblast cells. Lnc-HZ04 directly and specifically bound with miR-hz04, diminished the reduction effects of miR-hz04 on IP 3 R 1 mRNA expression level and on IP 3 R 1 mRNA stability, and then activated the Ca 2+ -mediated IP 3 R 1 /p-CaMKII/SGCB pathway, which further promoted trophoblast cell apoptosis. The miR-hz04 target site on lnc-HZ04 played crucial roles in these regulations. In normal trophoblast, relatively less lnc-HZ04 and more miR-hz04 suppressed this apoptosis pathway and gave normal pregnancy. After exposure to BPDE or in RM tissues, p53 was upregulated, which might promote p53-mediated lnc-HZ04 transcription. Relatively more lnc-HZ04 and less miR-hz04 activated this apoptosis pathway and might further induce miscarriage. BaP could also induce mice miscarriage by upregulating its corresponding murine apoptosis pathway. Therefore, BPDE-induced apoptosis of human trophoblast cells was associated with the occurrence of miscarriage. This work discovered the regulation roles of lnc-HZ04 and miR-hz04 and provided scientific and clinical understanding of the occurrence of unexplained miscarriage., (© 2021 Federation of American Societies for Experimental Biology.)

Egg yolk immunoglobulins (IgY), as nutraceutical supplement for therapeutic or prophylactic intervention, have been extensively studied. The effects of IgY on small molecular toxin-induced toxicity in animals are unclear. In the present study, the protection of highly purified and specific anti-AFB 1 IgY against AFB 1 -induced genotoxicity and oxidative damage on the rat liver model were investigated. Our results revealed that AFB 1 induced significant oxidative damage markers, as well as AFB 1 -induced protein expression in antioxidant, pro- and antiapoptosis processes in rat liver. These effects could be significantly inhibited by cogavage with anti-AFB 1 IgY in a dose-dependent manner. However, anti-AFB 1 IgY did not significantly induce hepatic CAT and SOD1. To explore mechanisms, metabolite experiments were established to evaluate the influence of anti-AFB 1 IgY on the absorption of AFB 1 in rats. Middle and high doses of anti-AFB 1 IgY reduced hepatic AFB 1 -DNA adducts by 43.3% and 52.9%, AFB 1 - N 7 -guanine urinary adducts by 19.6% and 34.4%, and AFB 1 -albumin adducts by 10.5% and 21.1%, respectively. The feces of high dose anti-AFB 1 IgY cogavaged rats contained approximately 2-fold higher AFB 1 equivalents at 3-6 h after ingestion than AFB 1 group feces, indicating IgY inhibited AFB 1 uptake. These results had provided insight that anti-AFB 1 IgY could prevent animal organs from damage caused by AFB 1 and will be beneficial for the application of detoxification antibody as a supplement in food.