Functional Analysis of an Intronic FBN1 Pathogenic Gene Variant in a Family With Marfan Syndrome.

Marfan syndrome (MFS) is an autosomal dominant connective tissue disorder that canonically affects the ocular, skeletal, and cardiovascular system, in which aortic tear and rupture is the leading cause of death for MFS patients. Genetically, MFS is primarily associated with fibrillin-1 (FBN1) pathogenic variants. However, the disease-causing variant in approximately 10% of patients cannot be identified, partly due to some cryptic mutations that may be missed using routine exonic sequencing, such as non-coding intronic variants that affects the RNA splicing process. We present a 32-year female with typical MFS systemic presentation that reached to a clinical diagnosis according to the revised Ghent nosology. We performed whole-exome sequencing (WES) but the report failed to identify known causal variants when analyzing the exonic sequence. However, further investigation on the exon/intron boundaries of the WES report revealed a candidate intronic variant of the fibrillin 1 (FBN1) gene (c.248-3 C>G) that predicted to affect the RNA splicing process. We conducted minigene splicing analyses and demonstrated that the c.248-3 C>G variant abolished the canonical splicing site of intron 3, leading to activation of two cryptic splicing sites and causing insertion (c.248-1_248-2insAG and c.248-1_248-282ins). Our study not only characterizes an intronic variant to the mutational spectrum of the FBN1 gene in MFS and its aberrant effect on splicing, but highlights the importance to not neglect the exon/intron boundaries when reporting and assessing WES results. We point out the need of conducting functional analysis to verify the pathogenicity of intronic mutation, and the opportunity to re-consider the standard diagnostic approaches in cases of clinically diagnosed MFS with normal or variant of unknown significance genetic results.

LncRNA HOXA-AS3 Promotes the Progression of Pulmonary Arterial Hypertension through Mediation of miR-675-3p/PDE5A Axis.

Pulmonary arterial hypertension (PAH) seriously threatens the elder people. Long non-coding RNAs (lncRNAs) are involved in multiple diseases. However, the study of the lncRNAs in the occurrence of PAH is just beginning. For this, we sought to explore the biological function of lncRNA HOXA cluster antisense RNA 3 (HOXA-AS3) in PAH. Hypoxia (HYP) was used to mimic in vitro model of PAH. Gene and protein expressions in cells were detected by q-PCR and Western blotting, respectively. In addition, cell proliferation and viability were tested by CCK-8 and MTT assay. Cell apoptosis was measured by flow cytometry. Wound healing was used to detect cell migration. Furthermore, the connection of HOXA-AS3, miR-675-3p, and phosphodiesterase 5A (PDE5A) was verified by dual-luciferase report assay. HOXA-AS3 and PDE5A were upregulated in human pulmonary artery smooth muscle cells (HPASMCs) in the presence of HYP, while miR-675-3p was downregulated. Moreover, knockdown of HOXA-AS3 suppressed the growth and migration of HPASMCs, but induced the apoptosis. Overexpression of miR-675-3p achieved the same effect. MiR-675-3p inhibitor or overexpression of PDE5A notably reversed the inhibitory effect of HOXA-AS3 knockdown on PAH. Finally, HOXA-AS3 could sponge miR-675-3p, and PDE5A was directly targeted by miR-675-3p. HOXA-AS3 increased the development of PAH via regulation of miR-675-3p/PDE5 axis, which could be the potential biomarker for treatment of PAH.

Resection of a Giant Bronchogenic Cyst in the Left Atrium.

A bronchogenic cyst in the left atrium is rare. We report the case of a 17-year-old male patient who was admitted to the emergency department because of severe chest pain and dyspnea. He was diagnosed using echocardiography and computed tomography, which revealed a huge cardiac tumour in the dome of the left atrium. He was surgically treated with tumour enucleation. The resultant atrial dome defect was reconstructed with a bovine pericardial patch. Pathologic investigation revealed that the tumour was a bronchogenic benign cyst.

Effects of Isolated Impaired Fasting Glucose on Brain Injury During Cardiac Surgery Under Cardiopulmonary Bypass.

Objective: To evaluate the effects of isolated impaired fasting glucose (IIFG) on brain injury in patients undergoing cardiopulmonary bypass (CPB) surgery. Methods: Patients with rheumatic heart valve disease who underwent elective mitral valve replacement were included and divided into control and IIFG groups. Pre-, intra-, and postoperative blood glucose levels, serum insulin levels, insulin resistance index (HOMA-IR), lactic acid levels, and neuron-specific enolase (NSE) and S100B levels were measured. The cerebral oxygen extraction ratio (OER) was calculated. Cognitive function was assessed via the Mini-Mental State Examination (MMSE). Results: HOMA-IR levels were higher in the IIFG group than the control group 30 min after the beginning of CPB, at the termination of CPB, and 2 h after the termination of CPB. Cerebral OER and lactic acid increased intraoperatively in both groups, especially in the IIFG group. NSE and S100B levels were higher in the IIFG group than in the control group at the termination of CPB, 2 h after the termination of CPB, and at 24 h postoperatively. The MMSE scores did not significantly differ between the two groups. Delirium occurred in two patients in the IIFG group, and one in the control group. No other signs and symptoms of brain injuries were detected in either group. Conclusions: The increased postoperative NSE and S100B levels in the IIFG group compared with controls may be associated with severe insulin resistance and stress hyperglycemia. However, the IIFG group did not have clinical manifestations of brain injuries, including cognitive impairment.

MicroRNA-495 Ameliorates Cardiac Microvascular Endothelial Cell Injury and Inflammatory Reaction by Suppressing the NLRP3 Inflammasome Signaling Pathway.

BACKGROUND/AIMS: In recent years, microRNA-495 (miR-495) has been reported to be a tumor-suppressor miR that is down-modulated in cancers. However, its potential mechanism remains unknown. Therefore, this study aimed to demonstrate the role of miR-495 in cardiac microvascular endothelial cell (CMEC) injury and inflammatory reaction by mediating the pyrin domain-containing 3 (NLRP3) inflammasome signaling pathway. METHODS: Overall, 40 mice were assigned into myocardial ischemia/reperfusion injury (MIR) and sham groups. After model establishment, the levels of troponin T (TnT), troponin I (TnI), N-terminal pro-B-type natriuretic peptide (NT-proBNP), creatine kinase isoenzyme MB (CK-MB), myoglobin (MYO), tumor necrosis factor-alpha (TNF-α), and interleukin 1beta (IL-1ß) were detected by Enzyme-Linked Immunosorbent Assay (ELISA). Apoptosis was evaluated using Terminal deoxy (d)-UTP nick end labeling (TUNEL) staining, the level of NLRP3 protein was determined by immunohistochemical assay, and miR-495 was detected by in situ hybridization (ISH). The infarct size was determined using 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. The expression of miR-495 and the mRNA and protein levels of NLRP3, TNF-α, IL-1ß, IL-18 and caspase-1 were evaluated by RT-qPCR and western blot analysis. After transfection, the cells were treated with a miR-495 mimic, a miR-495 inhibitor, or siNLRP3. Cell proliferation was measured by the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and cell cycle and apoptosis by flow cytometry. RESULTS: Mice with myocardial I/R injury had elevated levels of TnT, TnI, NT-proBNP, CK-MB, MYO, TNF-α and IL-1ß; enhanced cell apoptosis; increased expression of NLRP3, TNF-α, IL-1ß, IL-18 and caspase-1; and decreased miR-495 expression. MiR-495 was confirmed to target NLRP3. Moreover, miR-495 reduced the mRNA and protein levels of NLRP3, TNF-α, IL-1ß, IL-18 and caspase-1, inhibited cell apoptosis and decreased cells at the G0/G1 phase while improving cell proliferation and increasing cells at the S phase. However, the effects of NLRP4 were proved to be reciprocal. CONCLUSION: In conclusion, the current study indicated that miR-495 improved CMEC injury and inflammation by suppressing the NLRP3 inflammasome signaling pathway.

The protective effect of umbelliferone ameliorates myocardial injury following ischemia­reperfusion in the rat through suppression NLRP3 inflammasome and upregulating the PPAR-γ.

The present study investigated whether the protective effect of umbelliferone could regulate myocardial injury following ischemia­reperfusion and improve mitochondrial respiratory function, thereby relieving myocardial injury following ischemia­reperfusion in rats. In the present study, the extent of inflammation and oxidative stress were analyzed using ELISA. Western blot analysis was employed to investigate the protein expression levels of the PYD domains­containing protein 3 (NLRP3) inflammasome and peroxisome proliferator­activated receptor-γ (PPAR­Î³). Compared with the myocardial injury following ischemia­reperfusion group, umbelliferone significantly prevented myocardial injury, inhibited oxidative stress markers (superoxide dismutase and malondialdehyde), reduced inflammation (tumor necrosis factor­α and interleukin­6) and myocardial apoptosis levels (caspase­3/9 and apoptosis regular B­cell lymphoma­2­associated X protein) in the myocardial injury following ischemia­reperfusion group of rats. Umbelliferone treatment also suppressed NACHT, LRR and NLRP3 inflammasome activation and induced PPAR­Î³ expression. The results of the present study suggested that the protective effect of umbelliferone may ameliorate myocardial injury following ischemia­reperfusion in the rat through the suppression of the NLRP3 inflammasome and upregulating PPAR­Î³ expression.

Comparative evaluation of topical pranoprofen and fluorometholone in cases with chronic allergic conjunctivitis.

PURPOSE: This study was conducted to compare the efficacy of 0.1% fluorometholone and 0.1% pranoprofen in cases with chronic allergic conjunctivitis. METHODS: In an investigator-masked trial, patients with chronic allergic conjunctivitis were randomized to treatment with 4 times daily 0.1% pranoprofen (PN) or 0.1% fluorometholone (FL) eye drops for 4 weeks. A 4-point rating scale assessing the severity of 5 symptoms and 4 signs (0 = none, 1 = mild, 2 = moderate, and 3 = severe) was used. A linear mixed model was used to explore the rate of score changes. Regression analysis was used to evaluate the relation between clinical outcome and age. RESULTS: A total of 75 patients were enrolled at the baseline. There were no significant differences in the demographics and baseline skin prick scores between both groups. Mean baseline scores in PN and FL group were 6.71 ± 2.28 and 6.41 ± 2.06, respectively. The scores rapidly decreased to 3.35 ± 1.58 and 2.91 ± 1.71 on day 7, respectively. Fluorometholone showed a more rapid effect compared with pranoprofen during the first week of treatment (P < 0.05) but not later. Regression analysis showed that age was negatively associated with response to fluorometholone (younger than 29 years). The intraocular pressure increased by 0.7 mm Hg in the FL group and decreased by 0.5 mm Hg in the PN group on day 28 (P > 0.05). CONCLUSIONS: Both fluorometholone and pranoprofen were effective for management of cases with chronic allergic conjunctivitis. Fluorometholone provided more rapid relief as compared with pranoprofen. The effect of fluorometholone was more pronounced in younger patients.