Role and mechanism of miR-871-3p/Megf8 in regulating formaldehyde-induced cardiomyocyte inflammation and congenital heart disease.

OBJECTIVE AND DESIGN: We aimed to investigate the molecular mechanism underlying formaldehyde (FA)-induced congenital heart disease (CHD) using in vitro and in vivo models. MATERIALS AND SUBJECTS: Neonatal rat heart tissues and H9C2 cells were used for in vitro studies, while FA-exposed new-born rats were used for in vivo studies. TREATMENT: H9C2 cells were exposed to FA concentrations of 0, 50, 100 and 150 µM/mL for 24 h. METHODS: Whole transcriptome gene sequencing identified differentially expressed miRNAs in neonatal rat heart tissues, while Real-time quantitative PCR (RT-qPCR) assessed miR-871-3p and Megf8 expression. RNA pull-down and dual-luciferase reporter assays determined miR-871-3p and Megf8 relationships. Inflammatory cytokine expression was assessed by western blotting. A FA-induced CHD model was used to validate miR-871-3p regulatory effects in vivo. RESULTS: We identified 89 differentially expressed miRNAs, with 28 up-regulated and 61 down-regulated (fold change ≥ 2.0, P < 0.05). Inflammation (interleukin) and signalling pathways were found to control FA-induced cardiac dysplasia. miR-871-3p was upregulated in FA-exposed heart tissues, modulated inflammation, and directly targeted Megf8. In vivo experiments showed miR-871-3p knockdown inhibited FA-induced inflammation and CHD. CONCLUSION: We demonstrated miR-871-3p's role in FA-induced CHD by targeting Megf8, providing potential targets for CHD intervention and improved diagnosis and treatment strategies.

Identification and experimental validation of autophagy-related genes in abdominal aortic aneurysm.

AIM: Autophagy plays essential roles in abdominal aortic aneurysm (AAA) development and progression. The objective of this study was to verify the autophagy-related genes (ARGs) underlying AAA empirically and using bioinformatics analysis. METHODS: Two gene expression profile datasets GSE98278 and GSE57691 were downloaded from the Gene Expression Omnibus (GEO) database, and principal component analysis was performed. Following, the R software (version 4.0.0) was employed to analyze potentially differentially expressed genes related with AAA and autophagy. Subsequently, the candidate genes were screened using protein-protein interaction (PPI), gene ontology (GO) enrichment analysis, and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Finally, quantitative real-time polymerase chain reaction (RT-qPCR) was performed to detect the RNA expression levels of the top five selected abnormal ARGs in clinical samples obtained from the normal and AAA patients. RESULTS: According to the information contained (97 AAA patients and 10 healthy controls) in the two datasets, a total of 44 differentially expressed autophagy-related genes (6 up-regulated genes and 38 down-regulated genes) were screened. GO enrichment analysis of differentially expressed autophagy-related genes (DEARGs) demonstrated that some enrichment items were associated with inflammation, and PPI analysis indicated interaction between these genes. RT-qPCR results presented that the expression levels of IL6, PPARG, SOD1, and MAP1LC3B were in accordance with the bioinformatics prediction results acquired from the mRNA chip. CONCLUSION: Bioinformatics analysis identified 44 potential autophagy-related differentially expressed genes in AAA. Further verification by RT- qPCR presented that IL6, PPARG, SOD1, and MAP1LC3B may affect the development of AAA by regulating autophagy. These findings might help explain the pathogenesis of AAA and be helpful in its diagnosis and treatment.

Identification of long non-coding RNA in formaldehyde-induced cardiac dysplasia in rats.

Formaldehyde exposure during pregnancy can cause fetal congenital heart disease (CHD). However, the regulatory mechanism remains unclear. Studies on the biology of long non-coding RNAs (lncRNAs) show that lncRNAs can influence cardiac development and disease. However, expression patterns and regulatory mechanisms of action of lncRNAs in formaldehyde-induced CHD remain unclear. We used high-throughput sequencing strategies as a means of identifying lncRNA expression profiles in heart tissues of normal and formaldehyde-exposed newborn rats. Overall, 763 differentially expressed lncRNAs were identified, including 325 and 438 that were respectively up-regulated and down-regulated. GO and KEGG analyses indicated that the Ras and hedgehog signaling pathways may be important regulatory pathways in CHD caused by exposure to formaldehyde. A lncRNA-miRNA-mRNA co-expression network was constructed and a key miRNA, rno-miR-665, was identified. Furthermore, qRT-PCR analysis verified that the novel lncRNAs: MSTRG.27313.2, MSTRG.30629.2, MSTRG.36520.33, MSTRG.91234.1, and MSTRG.91233.9, were upregulated in the formaldehyde-exposed group. These differentially expressed lncRNAs identified during formaldehyde-induced CHD in newborn rats help explain CHD pathogenesis and provide an effective reference for diagnosing and treating CHD.

Comprehensive profile of circRNAs in formaldehyde induced heart development.

Circular RNAs (circRNAs) are a novel type of long non-coding RNAs that can regulate gene expression in heart development and heart disease. However, the expression pattern of circRNAs in congenital heart disease (CHD) induced by formaldehyde exposure is still unknown. We detected circRNAs expression profiles in heart tissue taken from six neonatal rat pups with formaldehyde exposure group and normal group using RNA-sequencing. Results revealed that a total of 54 circRNAs were dysregulated in the formaldehyde exposure group compared to the normal group. Among them, 31 were upregulated and 23 were downregulated (fold change = 2.0, p < 0.0 5). The qRT-qPCR results showed that expressions of 12:628708|632694, 18:77477060|77520779, 5:167486001|167526275 were significantly upregulated, while that of 7:41167312|4116775 and 20:50659751|5068786 were notably downregulated; the expression pattern was consistent with the RNA sequencing data. Bioinformatics analysis shows that the pathogenesis of formaldehyde exposure-induced CHD may involve Hippo-YAP pathway、Notch signaling pathway and other pathways. A key miRNA (rno-miR-665) was identified by constructing a circRNA-miRNA-mRNA co-expression network. In summary, the study illustrated that circRNAs differentially expressed in fetal heart tissues during formaldehyde exposure has potential biological functions and may be a biomarker or therapeutic target for CHD.

The cellular function and molecular mechanism of formaldehyde in cardiovascular disease and heart development.

As a common air pollutant, formaldehyde is widely present in nature, industrial production and consumer products. Endogenous formaldehyde is mainly produced through the oxidative deamination of methylamine catalysed by semicarbazide-sensitive amine oxidase (SSAO) and is ubiquitous in human body fluids, tissues and cells. Vascular endothelial cells and smooth muscle cells are rich in this formaldehyde-producing enzyme and are easily damaged owing to consequent cytotoxicity. Consistent with this, increasing evidence suggests that the cardiovascular system and stages of heart development are also susceptible to the harmful effects of formaldehyde. Exposure to formaldehyde from different sources can induce heart disease such as arrhythmia, myocardial infarction (MI), heart failure (HF) and atherosclerosis (AS). In particular, long-term exposure to high concentrations of formaldehyde in pregnant women is more likely to affect embryonic development and cause heart malformations than long-term exposure to low concentrations of formaldehyde. Specifically, the ability of mouse embryos to effect formaldehyde clearance is far lower than that of the rat embryos, more readily allowing its accumulation. Formaldehyde may also exert toxic effects on heart development by inducing oxidative stress and cardiomyocyte apoptosis. This review focuses on the current progress in understanding the influence and underlying mechanisms of formaldehyde on cardiovascular disease and heart development.

ßII spectrin (SPTBN1): biological function and clinical potential in cancer and other diseases.

ßII spectrin, the most common isoform of non-erythrocyte spectrin, is a cytoskeleton protein present in all nucleated cells. Interestingly, ßII spectrin is essential for the development of various organs such as nerve, epithelium, inner ear, liver and heart. The functions of ßII spectrin include not only establishing and maintaining the cell structure but also regulating a variety of cellular functions, such as cell apoptosis, cell adhesion, cell spreading and cell cycle regulation. Notably, ßII spectrin dysfunction is associated with embryonic lethality and the DNA damage response. More recently, the detection of altered ßII spectrin expression in tumors indicated that ßII spectrin might be involved in the development and progression of cancer. Its mutations and disorders could result in developmental disabilities and various diseases. The versatile roles of ßII spectrin in disease have been examined in an increasing number of studies; nonetheless, the exact mechanisms of ßII spectrin are still poorly understood. Thus, we summarize the structural features and biological roles of ßII spectrin and discuss its molecular mechanisms and functions in development, homeostasis, regeneration and differentiation. This review highlight the potential effects of ßII spectrin dysfunction in cancer and other diseases, outstanding questions for the future investigation of therapeutic targets. The investigation of the regulatory mechanism of ßII spectrin signal inactivation and recovery may bring hope for future therapy of related diseases.

miR-153-3p Targets ßII Spectrin to Regulate Formaldehyde-Induced Cardiomyocyte Apoptosis.

Background: Formaldehyde (FA) is ubiquitous in the environment and can be transferred to the fetus through placental circulation, causing miscarriage and congenital heart disease (CHD). Studies have shown that ßII spectrin is necessary for cardiomyocyte survival and differentiation, and its loss leads to heart development defects and cardiomyocyte apoptosis. Additionally, previous studies have demonstrated that miRNA is essential in heart development and remodeling. However, whether miRNA regulates FA-induced CHD and cardiomyocyte apoptosis remains unclear. Methods: Using commercially available rat embryonic cardiomyocytes and a rat model of fetal cardiomyocyte apoptosis. Real-time quantitative PCR (RT-qPCR) and Western blot were performed to examine the level of miR-153-3p, ßII spectrin, caspase 7, cleaved caspase7, Bax, Bcl-2 expression in embryonic cardiomyocytes and a rat model of fetal cardiomyocyte apoptosis. Apoptotic cell populations were evaluated by flow cytometry and Tunel. Luciferase activity assay and RNA pull-down assay were used to detect the interaction between miR-153-3p and ßII spectrin. Masson's trichrome staining detects the degree of tissue fibrosis. Fluorescence in situ hybridization (FISH) and Immunohistochemistry were used to detect the expression of miR-153-3p and ßII spectrin in tissues. Results: Using commercially available rat embryonic cardiomyocytes and a rat model of fetal cardiomyocyte apoptosis, our studies indicate that miR-153-3p plays a regulatory role by directly targeting ßII spectrin to promote cardiomyocyte apoptosis. miR-153-3p mainly regulates cardiomyocyte apoptosis by regulating the expression of caspase7, further elucidating the importance of apoptosis in heart development. Finally, the results with our animal model revealed that targeting the miR-153-3p/ßII spectrin pathway effectively regulated FA-induced damage during heart development. Recovery experiments with miR-153-3p antagomir resulted in the reversal of FA-induced cardiomyocyte apoptosis and fetal cardiac fibrosis. Conclusion: This study investigated the molecular mechanism underpinning the role of ßII spectrin in FA-induced CHD and the associated upstream miRNA pathway. The study findings suggest that miR-153-3p may provide a potential target for the clinical diagnosis and treatment of CHD.

Hierarchical micro/nanofibrous membranes of sustained releasing VEGF for periosteal regeneration.

The periosteum plays a vital role in both development and injury healing process of bone. However, few researches have focused on artificial periosteum, which was also limited by the complexity on its construction and biological risks for clinical practice. In order to tackle this issue, inspired by the structural development of periosteum, we put forward a hierarchical micro/nanofibrous bionic periosteum with sustained releasing of VEGF as exogenous vascularized fibrous layer of periosteum to induce endogenous cambium layer in vivo for complete regeneration of periosteal and bone tissue, through collagen self-assembly and micro-sol electrospinning technologies. The VEGF encapsulated in hyaluronan-PLLA core-shell structure was demonstrated to be released in a durable way for angiogenesis in fibrous layer and bone defect area. Meanwhile, the self-assembly of collagen together with electrospun fibers contributed to a hierarchical micro/nanostructure which greatly mimicked the microenvironment of extracellular matrix to provide structural and biochemical cues for cell adhesion, proliferation and differentiation, and lead to the formation of cambium layer which mimicked the in-situ ossification manner as intramembranous ossification. As the motif of this study, the periosteal regeneration was characterized both by osteoblasts and periostin, which represented structural and molecular mechanisms respectively. Furthermore, the periosteal biomaterial proposed here possessed the superior abilities of scar inhibition, angiogenesis, osteogenesis to repair the bone defect in a uniform and rapid manner by inherent periosteal ossific mechanism involved in both intramembranous and endochondral ossification. Thus, the endogenous-exogenous combined bionic periosteum proved to be efficient and versatile in triggering periosteal and bone regeneration and hopefully supply a promising strategy for solving clinical issue.

Vitamin E Reversed Apoptosis of Cardiomyocytes Induced by Exposure to High Dose Formaldehyde During Mice Pregnancy.

In this study, we investigated the protection effect of Vitamin E (Vit E) on formaldehyde (FA) exposure during pregnancy induced apoptosis of cardiomyocytes, and used an HL-1 cell line to confirmed the findings in vivo.Pregnant mice received different doses of FA (0.5 mg/kg, 1.0 mg/kg, 1.5 mg/kg, 0.1 µg Vit E, or 1.5 mg/kg + 0.1 µg Vit E). TUNEL staining was used to reveal the apoptosis in cardiomyocytes, and SOD, MDA, GSH, Livin, and Caspase-3 in cardiomyocytes were detected by ELISA, RT-PCR, and Western blot. For in vitro study, HL-1 cells were treated with vehicle, 5 µmol/L FA, 25 µmol/L FA, 50 µmol/L FA, 10 mg/L Vit. E, and 50 µmol/L FA+ 10 mg/L Vit E, respectively. CCK-8 assay and flow cytometry were used to evaluate cell vitality and apoptosis. A high dose of FA exposure led to cytotoxicity in both pregnant mice and offspring, as TUNEL staining revealed a significant apoptosis of cardiomyocytes, and the alternation in SOD, GSH, MDA, Livin, and Caspase-3 was found in cardiomyocytes. 0.1 µg Vit. E could reverse high doses of FA exposure induced apoptosis of cardiomyocytes in both pregnant mice and offspring. The in vitro study revealed that FA exposure induced a decrease of cell viability and increased cell apoptosis, as well as oxidative stress in HL-1 cells with alternation in SOD, GSH, MDA, Livin, and Caspase-3.This study revealed a high dose of FA induced oxidative stress and apoptosis of cardiomyocytes in both pregnant mice and offspring, and Vit E supplement during pregnancy reversed the systemic and myocardial toxicity of FA.

Loss of Cystic Fibrosis Transmembrane Regulator Impairs Intestinal Oxalate Secretion.

Patients with cystic fibrosis have an increased incidence of hyperoxaluria and calcium oxalate nephrolithiasis. Net intestinal absorption of dietary oxalate results from passive paracellular oxalate absorption as modified by oxalate back secretion mediated by the SLC26A6 oxalate transporter. We used mice deficient in the cystic fibrosis transmembrane conductance regulator gene (Cftr) to test the hypothesis that SLC26A6-mediated oxalate secretion is defective in cystic fibrosis. We mounted isolated intestinal tissue from C57BL/6 (wild-type) and Cftr-/- mice in Ussing chambers and measured transcellular secretion of [14C]oxalate. Intestinal tissue isolated from Cftr-/- mice exhibited significantly less transcellular oxalate secretion than intestinal tissue of wild-type mice. However, glucose absorption, another representative intestinal transport process, did not differ in Cftr-/- tissue. Compared with wild-type mice, Cftr-/- mice showed reduced expression of SLC26A6 in duodenum by immunofluorescence and Western blot analysis. Furthermore, coexpression of CFTR stimulated SLC26A6-mediated Cl--oxalate exchange in Xenopus oocytes. In association with the profound defect in intestinal oxalate secretion, Cftr-/- mice had serum and urine oxalate levels 2.5-fold greater than those of wild-type mice. We conclude that defective intestinal oxalate secretion mediated by SLC26A6 may contribute to the hyperoxaluria observed in this mouse model of cystic fibrosis. Future studies are needed to address whether similar mechanisms contribute to the increased risk for calcium oxalate stone formation observed in patients with cystic fibrosis.

Usefulness of thyroglobulin in cervical lymph node fine-needle aspirations at initial thyroidectomy for the diagnosis of metastases in papillary thyroid cancer.

OBJECTIVE: To study the role of fine-needle aspiration biopsy and thyroglobulin measurement (FNAB-Tg) in diagnosing cervical lymph node (CLN) metastases in patients with papillary thyroid cancer (PTC) before thyroidectomy. METHODS: Ultrasonography (US)-guided FNAB was performed on 92 patients with PTCs with 105 CLNs before surgery. The wash-out of the FNAB-Tg level was detected. RESULTS: Based on the final pathology, 67 lymph nodes (LNs) were positive for metastasis, and 38 LNs were negative for metastasis. The sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of FNAB-Tg in thyroidectomized patients was 100%, 86.8%, 95.2%, 97.3%, and 95.2%, respectively. CONCLUSIONS: FNAB-Tg is a useful technique for diagnosing CLN metastasis of PTC before thyroidectomy.
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Synergism between phospholipase D2 and sorbitol accumulation in diabetic cataract formation through modulation of Na,K-ATPase activity and osmotic stress.

Phospholipase D (PLD), a highly regulated enzyme that generates the second messenger phosphatidic acid, functions in signal transduction, membrane trafficking and cytoskeletal reorganization. PLD is thought to be involved in the pathogenesis of diabetic complications by activating PKC. Since PKC and PLD are present in the lens we sought to determine if PLD plays a role in diabetic cataract development. We developed transgenic mice that overexpress PLD2, one of the two mammalian isoforms of PLD. These mice developed congenital nuclear cataracts, but not diabetic cataracts. Histological analysis revealed vacuole formation in the fiber cells, mediated potentially by the substantially increased Na,K-ATPase activity. In the presence of the aldose reductase overexpressing transgene that increases lens osmotic pressure, these double transgenic mice developed more severe congenital cataract and became susceptible to develop diabetic cataract. Together, these data suggest that increased PLD2 activity in the lens under hyperglycemic condition might impair its osmoregulatory mechanism and reduce its ability to cope with the osmotic stress triggered by sorbitol accumulation.