Advanced glycation end product-modified low-density lipoprotein promotes pro-osteogenic reprogramming via RAGE/NF-κB pathway and exaggerates aortic valve calcification in hamsters.

BACKGROUND: Advanced glycation end product-modified low-density lipoprotein (AGE-LDL) is related to inflammation and the development of atherosclerosis. Additionally, it has been demonstrated that receptor for advanced glycation end products (RAGE) has a role in the condition known as calcific aortic valve disease (CAVD). Here, we hypothesized that the AGE-LDL/RAGE axis could also be involved in the pathophysiological mechanism of CAVD. METHODS: Human aortic valve interstitial cells (HAVICs) were stimulated with AGE-LDL following pre-treatment with or without interleukin 37 (IL-37). Low-density lipoprotein receptor deletion (Ldlr-/-) hamsters were randomly allocated to chow diet (CD) group and high carbohydrate and high fat diet (HCHFD) group. RESULTS: AGE-LDL levels were significantly elevated in patients with CAVD and in a hamster model of aortic valve calcification. Our in vitro data further demonstrated that AGE-LDL augmented the expression of intercellular cell adhesion molecule-1 (ICAM-1), interleukin-6 (IL-6) and alkaline phosphatase (ALP) in a dose-dependent manner through NF-κB activation, which was attenuated by nuclear factor kappa-B (NF-κB) inhibitor Bay11-7082. The expression of RAGE was augmented in calcified aortic valves, and knockdown of RAGE in HAVICs attenuated the AGE-LDL-induced inflammatory and osteogenic responses as well as NF-κB activation. IL-37 suppressed inflammatory and osteogenic responses and NF-κB activation in HAVICs. The vivo experiment also demonstrate that supplementation with IL-37 inhibited valvular inflammatory response and thereby suppressed valvular osteogenic activities. CONCLUSIONS: AGE-LDL promoted inflammatory responses and osteogenic differentiation through RAGE/NF-κB pathway in vitro and aortic valve lesions in vivo. IL-37 suppressed the AGE-LDL-induced inflammatory and osteogenic responses in vitro and attenuated aortic valve lesions in a hamster model of CAVD.

Abnormally elevated EZH2-mediated H3K27me3 enhances osteogenesis in aortic valve interstitial cells by inhibiting SOCS3 expression.

BACKGROUND AND AIMS: The osteogenic transition of aortic valve interstitial cells (AVICs) plays a critical role for the progression of calcific aortic valve disease (CAVD). Enhancer of zeste homolog 2 (EZH2) is an important methyltransferase for histone H3 Lys27 (H3K27) that has been found to be involved in osteogenesis. Here, we investigated the effect and mechanism of EZH2 in CAVD progression. METHODS: High throughout mRNA sequencing, qRT-PCR and immunoblot were performed to screen differentially expressed genes in non-CAVD and CAVD aortic valves. To investigate the role of EZH2 and SOCS3 in osteogenesis, AVICs were treated with siRNA, adenovirus and specific inhibitors, then osteogenic markers and mineralized deposits were examined. In vivo, the morphology and function of aortic valves were investigated by HE stain and echocardiography in ApoE-/- mice fed a long-term western diet (WD). RESULTS: We discovered that EZH2 was upregulated and SOCS3 was downregulated in calcified aortic valves. In AVICs, inhibition or silencing of EZH2 attenuated the osteogenic responses. On the other hand, demethylases inhibitor (GSK-J4) enhanced osteogenic transition of AVICs. Moreover, SOCS3 knockdown enhanced the expression of osteogenic markers, while SOCS3 overexpression suppressed osteogenesis and calcification. The chromatin immunoprecipitation and restored experiments indicated that EZH2 directly targeted SOCS3 to promote osteogenic responses of AVICs. In vivo, treatment with EZH2 inhibitor through intraperitoneal injection attenuated aortic valve thickening, calcification and dysfunction induced by WD. CONCLUSIONS: Collectively, we found that EZH2-mediated H3K27me3 enhanced osteogenesis and microcalcification of AVICs via inhibiting SOCS3 expression, which provides potential targets for future therapeutic interventions of CAVD.

Hesperetin, a Promising Dietary Supplement for Preventing the Development of Calcific Aortic Valve Disease.

BACKGROUND: No effective therapeutic agents for calcific aortic valve disease (CAVD) are available currently. Dietary supplementation has been proposed as a novel treatment modality for various diseases. As a flavanone, hesperetin is widely abundant in citrus fruits and has been proven to exert protective effects in multiple diseases. However, the role of hesperetin in CAVD remains unclear. METHODS: Human aortic valve interstitial cells (VICs) were isolated from aortic valve leaflets. A mouse model of aortic valve stenosis was constructed by direct wire injury (DWI). Immunoblotting, immunofluorescence staining, and flow cytometry were used to investigate the roles of sirtuin 7 (Sirt7) and nuclear factor erythroid 2-related factor 2 (Nrf2) in hesperetin-mediated protective effects in VICs. RESULTS: Hesperetin supplementation protected the mice from wire-injury-induced aortic valve stenosis; in vitro, hesperetin inhibited the lipopolysaccharide (LPS)-induced activation of NF-κB inflammatory cytokine secretion and osteogenic factors expression, reduced ROS production and apoptosis, and abrogated LPS-mediated injury to the mitochondrial membrane potential and the decline in the antioxidant levels in VICs. These benefits of hesperetin may have been obtained by activating Nrf2-ARE signaling, which corrected the dysfunctional mitochondria. Furthermore, we found that hesperetin could directly bind to Sirt7 and that the silencing of Sirt7 decreased the effects of hesperetin in VICs and potently abolished the ability of hesperetin to increase Nrf2 transcriptional activation. CONCLUSIONS: Our work demonstrates that hesperetin plays protective roles in the aortic valve through the Sirt7-Nrf2-ARE axis; thus, hesperetin might be a potential dietary supplement that could prevent the development of CAVD.

Marine-Derived Piericidin Diglycoside S18 Alleviates Inflammatory Responses in the Aortic Valve via Interaction with Interleukin 37.

Calcific aortic valve disease (CAVD) is a valvular disease frequently in the elderly individuals that can lead to the valve dysfunction. Osteoblastic differentiation of human aortic valve interstitial cells (HAVICs) induced by inflammation play a crucial role in CAVD pathophysiological processes. To date, no effective drugs for CAVD have been established, and new agents are urgently needed. Piericidin glycosides, obtained from a marine-derived Streptomyces strain, were revealed to have regulatory effects on mitochondria in previous studies. Here, we discovered that 13-hydroxypiericidin A 10-O-α-D-glucose (1→6)-ß-D-glucoside (S18), a specific piericidin diglycoside, suppresses lipopolysaccharide- (LPS) induced inflammatory responses of HAVICs by alleviating mitochondrial stress in an interleukin (IL)-37-dependent manner. Knockdown of IL-37 by siRNA not only exaggerated LPS-induced HAVIC inflammation and mitochondrial stress but also abrogated the anti-inflammatory effect of S18 on HAVICs. Moreover, S18 alleviated aortic valve lesions in IL-37 transgenic mice of CAVD model. Microscale thermophoresis (MST) and docking analysis of five piericidin analogues suggested that diglycosides, but not monoglycosides, exert obvious IL-37-binding activity. These results indicate that S18 directly binds to IL-37 to alleviate inflammatory responses in HAVICs and aortic valve lesions in mice. Piericidin diglycoside S18 is a potential therapeutic agent to prevent the development of CAVD.

4-Octyl itaconate suppresses the osteogenic response in aortic valvular interstitial cells via the Nrf2 pathway and alleviates aortic stenosis in mice with direct wire injury.

Calcific aortic valve disease (CAVD) is the most prevalent valvular heart disease in older individuals, but there is a lack of drug treatment. The cellular biological mechanisms of CAVD are still unclear. Oxidative stress and endoplasmic reticulum stress (ER stress) have been suggested to be involved in the progression of CAVD. Many studies have demonstrated that 4-octyl itaconate (OI) plays beneficial roles in limiting inflammation and oxidative injury. However, the potential role of OI in CAVD has not been thoroughly explored. Thus, we investigated OI-mediated modulation of ROS generation and endoplasmic reticulum stress to inhibit osteogenic differentiation in aortic valve interstitial cells (VICs). In our study, calcified aortic valves showed increased levels of ER stress and superoxide anion, as well as abnormal expression of Hmox1 and NQO1. In VICs, OI activated the Nrf2 signaling cascade and contributed to Nrf2 stabilization and nuclear translocation, thus augmenting the expression of genes downstream of Nrf2 (Hmox1 and NQO1). Moreover, OI ameliorated osteogenic medium (OM)-induced ROS production, mitochondrial ROS levels and the loss of mitochondrial membrane potential in VICs. Furthermore, OI attenuated the OM-induced upregulation of ER stress markers, osteogenic markers and calcium deposition, which were blocked by the Nrf2-specific inhibitor ML385. Interestingly, we found that OM-induced ER stress and osteogenic differentiation were ROS-dependent and that Hmox1 silencing triggered ROS production, ER stress and elevated osteogenic activity, which were inhibited by NAC. Overexpression of NQO1 mediated by adenovirus vectors significantly suppressed OM-induced ER stress and osteogenic markers. Collectively, these results showed the anti-osteogenic effects of OI on AVICs by regulating the generation of ROS and ER stress by activating the Nrf2 signaling pathway. Furthermore, OI alleviated aortic stenosis in a mouse model with direct wire injury. Due to its antioxidant properties, OI could be a potential drug for the prevention and/or treatment of CAVD.

Modulation effect of vaginal mucosal microflora and susceptibility to Neisseria gonorrhoeae infections: a systematic review and meta-analysis.

OBJECTIVE: The vaginal microbiota may modulate susceptibility to Neisseria gonorrhea (NG) infections. The objective of this meta-analysis was to evaluate the association between these NG infections and the vaginal microbiota. METHOD: A systematic review and meta-analysis was conducted to investigate the correlation of vaginal microbiota and NG risk. Primary sources of the reviewed studies were from inception through December 2018. Vaginal mucosa microflora were dichotomized into high-Lactobacillus vaginal microbiota and low-Lactobacillus vaginal microbiota (LL-VMB), using either Nugent score, Amsel's criteria, presence of clue cells or 16S rRNA gene sequencing. RESULTS: A total of 8 studies qualified for inclusion in this meta-analysis. LL-VMB could be regarded as worse prognostic factor, and the pooled OR was 1.33 (95% CI 1.02, 1.73; P = 0.04, I2 = 44%). LL-VMB was associated with a significantly higher susceptibility of NG. Trend for the sensitive analysis was consistence with the primary outcome. Significant publication bias was not detected by the funnel plot. CONCLUSION: In conclusion, the systematic review and meta-analysis has demonstrated that LL-VMB was significantly associated with a high NG susceptibility.

Interaction of MTHFR gene with smoking and alcohol use and haplotype combination susceptibility to psoriasis in Chinese population.

To investigate the association of MTHFR gene polymorphism and additional gene-gene interaction with psoriasis risk. GMDR model was used to screen the best gene-smoking and gene-drinking interaction combinations. Logistic regression was performed to investigate association between two SNPs and psoriasis. For psoriasis patient-control haplotype analyses, the SHEsis online haplotype analysis software ( http://analysis.bio-x.cn ) was employed. We found that carriers of homozygous mutant of rs1801133 polymorphism and heterozygous of rs1801131 are associated with increased psoriasis risk than those with wild-type homozygotes, OR (95%CI) were 2.01 (1.48-2.79), and 2.08 (1.56-2.86), respectively. We also found a significant gene-environment interaction between C677T and alcohol drinking. In all samples, the haplotype 1298A-677C was observed most frequently in two groups, with 49.43 and 55.71% for the patients and controls, respectively. The results also indicated that the haplotype containing 1298C and 677T alleles were associated with a statistically increased psoriasis risk, OR (95%CI) = 1.73 (1.12-2.46), P = 0.0002. Our study found that rs1801133, rs1801131 within MTHFR gene, and interaction between C677T and alcohol drinking and haplotype containing the 1298C and 677T alleles were all associated with increased psoriasis risk.

Silencing of miR­155 suppresses inflammatory responses in psoriasis through inflammasome NLRP3 regulation.

Psoriasis is a dermatosis with the major clinical symptoms of scale, erythema and itching, and it has a long disease course. In addition, it is easily recurrent and refractory, greatly affecting the physical and mental health of patients. In the present study, it was hypothesized that the function of miR­155 increases psoriasis­induced inflammation and that its expression may be dependent on inflammasome activation. miR­155 expression was examined by gene chip array and quantitative polymerase chain reaction analysis. miR­155 expression levels were significantly increased in an in vivo model of psoriasis compared with normal tissues, as was the expression of NLR family pyrin domain containing 3 (NLRP3). In vitro, using keratinocyte­induced HaCaT cells as a model for psoriasis, silencing of miR­155 was confirmed to significantly decrease inflammation and NLRP3/caspase­1 signaling. Activation of toll­like receptor 4 (TLR4) enhanced the miR­155­induced inflammatory response in the in vitro keratinocyte model. Treatment with a TLR4 inhibitor or an NLRP3 inhibitor reversed the miR­155­mediated inflammation in the same cell system. The present study demonstrated that miR­155 silencing suppressed psoriasis­associated inflammatory responses through inflammasome NLRP3 regulation.