Lnc-HZ01 with m6A RNA methylation inhibits human trophoblast cell proliferation and induces miscarriage by up-regulating BPDE-activated lnc-HZ01/MXD1 positive feedback loop

Novel lncRNA‐HZ04 promotes BPDE‐induced human trophoblast cell apoptosis and miscarriage by upregulating IP3R1/CaMKII/SGCB pathway by competitively binding with miR‐hz04

“…Fifteen women who had undergone artificial miscarriages to terminate unwanted pregnancies (the HC group) and 15 patients with two or more consecutive unexplained miscarriage (the RM group) in the age between 25 and 30 were recruited, as described previously. 21,37,38 All HC groups have previous pregnancies. Any woman with one of the following features was excluded, including autoimmune abnormality, luteal phase defects, hyperprolactinemia, antiphospholipid antibody syndrome, or hyperandrogenemia; uterine abnormalities or cervical incompetence; abnormal karyotype of the parents or abortus; polycystic ovarian syndrome; the symptoms of endocrine or metabolic diseases; viral infectious diseases; tuberculosis, HBV, HCV, HIV, or with positive results from γ-interferon release tests; and eclampsia or preeclampsia.…”

“…2.9 | Quantitative real-time polymerase chain reaction RT-qPCR analysis was performed as described previously. 21,37,38 Total RNAs were extracted from cells or tissues. Subsequently, mRNA and lncRNA were reversely transcribed into cDNA using the Prime Script RT Reagent Kit with gDNA Eraser (Takara).…”

“…The methods were similar to those described previously. 21,37,38 The primary antibodies contained anti-IP 3 R 1 (dilution 1:1000, ab264281, Abcam, Cambridge, UK), anti-CaMKII (dilution 1:1000, 4436S, Cell Signaling Technology, Boston, USA), anti-p-CaMKII (dilution 1:1000, 3356S, Cell Signaling Technology), anti-SGCB (dilution 1:100, sc-5306, SANTA Cruz, Texas, USA), anti-p53 (ab1101, Abcam), and anti-GAPDH (dilution 1:10 000, ab8245, Abcam). The secondary antibodies included goat anti-rabbit IgG (dilution 1:5000, ab205718, Abcam) and goat anti-mouse IgG (dilution 1:5000, ab205719, Abcam).…”

“…EZ-Magna Chromatin immunoprecipitation (ChIP) Kit (Millipore) was used in ChIP assays, as described previously. 37,38 After sonication of the chromatin, the anti-p53 monoclonal antibody (ab1101, Abcam) was used for immunoprecipitation at 4°C overnight on an inverse rotator, with IgG (ab172730, Abcam) as negative control. The complex was washed thrice and DNA was separated.…”

“…In our recent work, we have identified that a novel lnc-HZ01 and MXD1 (MAX dimerization protein 1) comprise a positive self-feedback loop, which is activated by BaP or BPDE exposure. 37 This loop is upregulated in both RM tissues and BPDE-exposed trophoblast cells, which further promotes EIF4E transcription, inhibits trophoblast cell proliferation, and induces miscarriage. We also found that, in BPDE-exposed human trophoblast cells and in RM tissues, lnc-HZ03 and miR-hz03 upregulates p53/SAT1 axis and promote spermine metabolism, which further promote BPDE-induced human trophoblastic cell apoptosis and the occurrence of miscarriage.…”