A novel di-functional fluorescent probe for ONOO- and Zn2+ imaging in cells.

Peroxynitrite (ONOO-) is one of the most significant reactive oxygen species (ROS) in living cells. Zn2+ in living cells plays an essential part in different physiological processes. The abnormal concentration of ONOO- and Zn2+ in living cells are related to many kinds of diseases, such as anemia, epilepsy, diarrhea, Alzheimer's disease, and so on. The relationship of ONOO- and Zn2+ in living cells when the relative disease occurs remains unknown. So we develop the first probe H-1 for detecting ONOO- and Zn2+ at the same time. The probe H-1 shows high selectivity, good anti-interference capability, low detection limit and short response time to ONOO- and Zn2+. When the probe was applied to detect ONOO- and Zn2+ in HeLa cells, we could observe the fluorescence changing in the green and blue channels separately without interference in real time. It has the potential to employ the relation of ONOO- and Zn2+ in some disease mechanism research.

Association between gut microbiota and Hirschsprung disease: a bidirectional two-sample Mendelian randomization study.

Background: Several studies have pointed to the critical role of gut microbiota (GM) and their metabolites in Hirschsprung disease (HSCR) pathogenesis. However, the detailed causal relationship between GM and HSCR remains unknown. Methods: In this study, we used two-sample Mendelian randomization (MR) analysis to investigate the causal relationship between GM and HSCR, based on the MiBioGen Consortium's genome-wide association study (GWAS) and the GWAS Catalog's HSCR data. Reverse MR analysis was performed subsequently, and the sensitivity analysis, Cochran's Q-test, MR pleiotropy residual sum, outlier (MR-PRESSO), and the MR-Egger intercept were used to analyze heterogeneity or horizontal pleiotropy. 16S rDNA sequencing and targeted mass spectrometry were developed for initial validation. Results: In the forward MR analysis, inverse-variance weighted (IVW) estimates suggested that Eggerthella (OR: 2.66, 95%CI: 1.23-5.74, p = 0.01) was a risk factor for HSCR, while Peptococcus (OR: 0.37, 95%CI: 0.18-0.73, p = 0.004), Ruminococcus2 (OR: 0.32, 95%CI: 0.11-0.91, p = 0.03), Clostridiaceae1 (OR: 0.22, 95%CI: 0.06-0.78, p = 0.02), Mollicutes RF9 (OR: 0.27, 95%CI: 0.09-0.8, p = 0.02), Ruminococcaceae (OR: 0.16, 95%CI: 0.04-0.66, p = 0.01), and Paraprevotella (OR: 0.45, 95%CI: 0.21-0.98, p = 0.04) were protective factors for HSCR, which had no heterogeneity or horizontal pleiotropy. However, reverse MR analysis showed that HSCR (OR: 1.02, 95%CI: 1-1.03, p = 0.049) is the risk factor for Eggerthella. Furthermore, some of the above microbiota and short-chain fatty acids (SCFAs) were altered in HSCR, showing a correlation. Conclusion: Our analysis established the relationship between specific GM and HSCR, identifying specific bacteria as protective or risk factors. Significant microbiota and SCFAs were altered in HSCR, underlining the importance of further study and providing new insights into the pathogenesis and treatment.

Case Report: Kaposiform hemangioendothelioma with PIK3CA mutation successfully treated with sirolimus.

Kaposiform hemangioendothelioma (KHE) is an extremely rare, locally aggressive vascular neoplasm. The etiopathogenesis of KHE is still poorly understood. In the present study, we found a new mutation in KHE (c.685delA, p.Thr229fs). The KHE patient with the PIK3CA mutation showed complete regression after sirolimus treatment. We propose that the presence of the PIK3CA mutation in KHE may correlate with good response to sirolimus.

Successful Treatment of Fibro-Adipose Vascular Anomaly with Sirolimus.

BACKGROUND: The purpose of this study was to present our initial experience in using sirolimus therapy to treat fibro-adipose vascular anomaly (FAVA). METHODS: We retrospectively reviewed the medical records of eight patients with FAVA who were treated with sirolimus at our hospital between July 2017 and October 2020. RESULTS: Six girls (75%) and two boys (25%) were included in the cohort; the average age was 8 years (range, 1-13 years). Vascular tumors developed mainly on the extremities, including the forearm (n = 2; 25.0%), calf (n = 4; 50.0%), and thigh (n = 2; 25.0%). The predominant symptoms included swelling of the lesion (n = 8; 100%), pain (n = 7; 87.5%), contracture (n = 3; 37.5%), and phlebectasia (n = 3; 37.5%). Magnetic resonance imaging was the primary method used for FAVA diagnosis, and all patients underwent enhanced MRI. All lesions were heterogeneous with hyperintense T1 signals. The fat-suppressed T2-weighted images also revealed heterogeneous hyperintense masses, thus indicating fibrofatty infiltration. All eight patients received a sirolimus treatment regimen after FAVA diagnosis. One patient underwent tumor resection but experienced recurrence, whereas the other six patients underwent biopsy. Histological examination revealed that the lesions consisted of fibrofatty tissue with abnormal venous channels and anomalous lymphatic vascular components. Sirolimus softened the masses and caused tumor shrinkage within 5.25 ± 2.6 weeks (range, 2-10 weeks) after treatment initiation. The tumors also involuted rapidly and became stable within 7.75 ± 2.25 months after treatment initiation (range, 6-12 months). All seven patients experiencing pain reported relief within 3.8 ± 1.8 weeks (range, 2-7 weeks) after initiation of sirolimus therapy. Sirolimus alleviated but did not fully resolve the contracture in three patients. Remarkably, five patients exhibited a complete response, and three patients exhibited a partial response. At the time of the last follow-up, three patients had begun to gradually taper off sirolimus after 24 months of treatment and maintained a low blood sirolimus concentration. No serious adverse effects were observed during treatment. CONCLUSION: FAVA is a complex vascular malformation that appears to respond well to sirolimus treatment. Thus, sirolimus may be an effective and safe treatment for FAVA. LEVEL OF EVIDENCE: LEVEL IV.

LINC01296 promotes neuroblastoma tumorigenesis via the NCL-SOX11 regulatory complex.

Neuroblastoma (NB) is the most common extracranial solid tumor in childhood. Long non-coding RNA LINC01296 has been shown to predict the invasiveness and poor outcomes of patients with NB. Our study validated its prognostic value and investigated the biological function and potential mechanism of LINC01296 regulating NB. Results illuminated that LINC01296 expression was significantly correlated with unfavorable prognosis and malignant clinical features according to the public NB database. We identified that silencing LINC01296 repressed NB cell proliferation and migration and promoted apoptosis. Moreover, LINC01296 knockdown inhibited tumor growth in vivo. The opposite results were observed through the dCas9 Synergistic Activation Mediator System (dCas9/SAM) activating LINC01296. Mechanistically, we revealed that LINC01296 could directly bind to nucleolin (NCL), forming a complex that activated SRY-box transcription factor 11 (SOX11) gene transcription and accelerated tumor progression. In conclusion, our findings uncover a crucial role of the LINC01296-NCL-SOX11 complex in NB tumorigenesis and may serve as a prognostic biomarker and effective therapeutic target for NB.

circRNA-TBC1D4, circRNA-NAALAD2 and circRNA-TGFBR3: Selected Key circRNAs in Neuroblastoma and Their Associations with Clinical Features.

OBJECTIVE: The roles of circRNAs in neuroblastoma (NB) are unclear. We used next-generation sequencing to detect the circRNA expression profiles in NB to identify the key circRNAs and analyzed the relationships between the circRNAs and clinical features. METHODS: Five paired neuroblastoma tumor and adjacent normal fetal adrenal medulla samples were collected for high-throughput RNA sequencing. Bioinformatics analysis was performed for functional annotation of the host genes of differentially expressed circRNAs. Validation of dysregulated circRNAs was performed by real-time quantitative reverse transcription polymerase chain reaction. The relationships between the key circRNAs and clinical features were analyzed. In addition, overexpression of key circRNAs in an NB cell line, as well as cell proliferation assays, colony formation assays and cell migration assays, was conducted to investigate the biological functions of key circRNAs. RESULTS: A total of 4704 differentially expressed circRNAs were found, including 2462 up-regulated and 2242 down-regulated circRNAs. According to our previous studies, the predicted target circRNAs of miR-21 involved in tumorigenic signaling pathways were selected, including circRNA-TBC1D4, circRNA-NAALAD2 and circRNA-TGFBR3. These circRNAs were associated with clinical features, and the circRNA expression was significantly lower (P < 0.05) in the NB tissues than in normal adrenal tissues. Overexpression of circRNA-TBC1D4 promotes NB cell migration, but not proliferation and colony-formation in vitro. CONCLUSION: We suggest that circRNA-TBC1D4, circRNA-NAALAD2 and circRNA-TGFBR3 may be cancer suppressor genes, which act by sponging miR-21 in NB. Further investigations are needed to elucidate the underlying mechanism.

Overexpression Prox1 in HemECs resembles Kaposiform hemangioendothelioma and cytotoxicity of sirolimus in vitro.

BACKGROUND: Kaposiform hemangioendothelioma (KHE) is a rare vascular tumor that occurs in children. Prox1 is a specific lymphatic marker for KHE. We intended to establish a Prox1 transgenic cell line resembling KHE and investigate the mechanism of sirolimus in treating KHE. METHODS: Prox1 was stably expressed in infantile hemangioma cell HemECs. RT-qPCR and Western blot were conducted to measure the expression of target genes. CCK-8, EdU assay, and cell cycle analysis were conducted to detect cell proliferation. Wound healing and transwell assay were used to evaluate cell migration and invasion. RESULTS: Both mRNA and protein levels of Prox1, LYVE-1, Podoplanin were upregulated in Prox1+ HemECs. An acceleration of cell growth and a rise in migration and invasion were observed with Prox1 overexpression. Sirolimus inhibited cell proliferation, promoted apoptosis and led to G1 phase arrest in Prox1+ HemECs. The expression of p-mTOR, p-4EBP1, and p-P70S6K decreased and the ratio of LC-3 II/LC-3 I elevated after treatment of sirolimus. CONCLUSIONS: Stable overexpression of Prox1 in HemECs induced a lymphatic endothelial reprogramming, and enhanced aggressive biological effects, partly resembled the invasion of KHE, and could serve as a novel model for KHE. Sirolimus may block mTOR-mediated pathways and induced autophagy in KHE.