Nesfatin-1 mitigates calcific aortic valve disease via suppressing ferroptosis mediated by GSH/GPX4 and ZIP8/SOD2 axes.

OBJECTIVE: Calcific aortic valve disease (CAVD) predominantly affects the elderly and currently lacks effective medical treatments. Nesfatin-1, a peptide derived from the cleavage of Nucleobindin 2, has been implicated in various calcification processes, both physiological and pathological. This study explores the impact of Nesfatin-1 on the transformation of aortic valve interstitial cells (AVICs) in CAVD. METHODS AND RESULTS: In vitro experiments showed that Nesfatin-1 treatment mitigated the osteogenic differentiation of AVICs. Corresponding in vivo studies demonstrated a deceleration in the progression of CAVD. RNA-sequencing of AVICs treated with and without Nesfatin-1 highlighted an enrichment of the Ferroptosis pathway among the top pathways identified by the Kyoto Encyclopedia of Genes and Genomes analysis. Further examination confirmed increased ferroptosis in both calcified valves and osteoblast-like AVICs, with a reduction in ferroptosis following Nesfatin-1 treatment. Within the Ferroptosis pathway, ZIP8 showed the most notable modulation by Nesfatin-1. Silencing ZIP8 in AVICs increased ferroptosis and osteogenic differentiation, decreased intracellular Mn2+ concentration, and reduced the expression and activity of superoxide dismutase (SOD2). Furthermore, the silencing of SOD2 exacerbated ferroptosis and osteogenic differentiation. Nesfatin-1 treatment was found to elevate the expression of glutathione peroxidase 4 (GPX4) and levels of glutathione (GSH), as confirmed by Western blotting and GSH concentration assays. CONCLUSION: In summary, Nesfatin-1 effectively inhibits the osteogenic differentiation of AVICs by attenuating ferroptosis, primarily through the GSH/GPX4 and ZIP8/SOD2 pathways.

SIRT6-mediated Runx2 downregulation inhibits osteogenic differentiation of human aortic valve interstitial cells in calcific aortic valve disease.

Calcific aortic valve disease (CAVD) is a progressive cardiovascular disorder involving multiple pathogenesis. Effective pharmacological therapies are currently unavailable. Sirtuin6 (SIRT6) has been shown to protect against aortic valve calcification in CAVD. The exact regulatory mechanism of SIRT6 in osteoblastic differentiation remains to be determined, although it inhibits osteogenic differentiation of aortic valve interstitial cells. We demonstrated that SIRT6 was markedly downregulated in calcific human aortic valves. Mechanistically, SIRT6 suppressed osteogenic differentiation in human aortic valve interstitial cells (HAVICs), as confirmed by loss- and gain-of-function experiments. SIRT6 directly interacted with Runx2, decreased Runx2 acetylation levels, and facilitated Runx2 nuclear export to inhibit the osteoblastic phenotype transition of HAVICs. In addition, the AKT signaling pathway acted upstream of SIRT6. Together, these findings elucidate that SIRT6-mediated Runx2 downregulation inhibits aortic valve calcification and provide novel insights into therapeutic strategies for CAVD.

Fibrillin-1 mutation contributes to Marfan syndrome by inhibiting Cav1.2-mediated cell proliferation in vascular smooth muscle cells.

Marfan syndrome (MFS) is an autosomal dominant connective tissue disorder caused by mutation in fibrillin-1 (FBN1). However, the molecular mechanism underlying MFS remains poorly understood. The study aimed to explore how the L-type calcium channel (CaV1.2) modulates disease progression of MFS and to identify a potential effective target for attenuating MFS. KEGG enrichment analysis showed that the calcium signaling pathway gene set was significantly enriched. We demonstrated that FBN1 deficiency exhibited inhibition on both the expression of Cav1.2 and proliferation of vascular smooth muscle cells (VSMCs). Then, we examined whether FBN1 mediates Cav1.2 via regulating TGF-ß1. Higher levels of TGF-ß1 were observed in the serum and aortic tissues from patients with MFS. TGF-ß1 modulated Cav1.2 expression in a concentration-dependent manner. We evaluated the role of Cav1.2 in MFS by small interfering RNA and Cav1.2 agonist Bay K8644. The effect of Cav1.2 on cell proliferation was dependent on c-Fos activity. These results demonstrated FBN1 deficiency decreased the expression levels of Cav1.2 via regulation of TGF-ß1, and downregulation of Cav1.2 inhibited cell proliferation of human aortic smooth muscle cells (HASMCs) in MFS patients. These findings suggest that Cav1.2 may be an appealing therapeutic target for MFS.

Celastrol confers ferroptosis resistance via AKT/GSK3ß signaling in high-fat diet-induced cardiac injury.

Obesity-induced cardiac dysfunction is a severe global disease associated with high dietary fat intake, and its pathogenesis includes inflammation, oxidative stress, and ferroptosis. Celastrol (Cel) is a bioactive compound isolated from the herb Tripterygium wilfordii, which has a protective influence on cardiovascular diseases. In this study, the role of Cel in obesity-induced ferroptosis and cardiac injury was investigated. We found that Cel alleviated ferroptosis induced by Palmitic acid (PA), exhibiting a decrease in the LDH, CK-MB, Ptgs2, and Lipid Peroxidation levels. After cardiomyocytes were treated with additional LY294002 and LiCl, Cel exerted its protective effect through increased AKT/GSK3ß phosphorylation and decreased level of lipid peroxidation and Mitochondrial ROS. The systolic left ventricle (LV) dysfunction of obese mice was alleviated via ferroptosis inhibition by elevated p-GSK3ß and decreased Mitochondrial ROS under Cel treatment. Moreover, mitochondrial anomalies included swelling and distortion in the myocardium which was relieved with Cel. In conclusion, our results demonstrate that ferroptosis resistance with Cel under HFD conditions targets AKT/GSK3ß signaling, which provides novel therapeutic strategies in obesity-induced cardiac injury.

Corrigendum: Clinical outcomes following surgical mitral valve plasty or replacement in patients with infectious endocarditis: A meta-analysis.

[This corrects the article DOI: 10.3389/fsurg.2022.1048036.].

Mechanical versus Bioprosthetic Aortic Valve Replacement in Middle-Aged Adults: A Systematic Review and Meta-Analysis.

BACKGROUND: Mechanical prostheses and bioprosthetic prostheses have their own advantages and disadvantages. Mechanical ones are recommended for younger patients (<50 years old), and bioprosthetic ones are recommended for older patients (>70 years old). There is still debate regarding which kind of prosthesis is better for middle-aged patients (50 to 70 years old) receiving aortic valve replacement (AVR). To solve this problem, we conducted this meta-analysis. Given that only one randomized controlled trial (RCT) study was included, we conducted a subgroup analysis of RCT and propensity score matching (PSM) retrospective studies to reduce the bias. METHODS: We systematically searched articles related to clinical outcomes of mechanical and bioprosthetic prostheses in middle-aged patients receiving AVR in the PubMed, Cochrane Library, and China National Knowledge Infrastructure (CNKI) databases. The published date was up to 1 October 2022. Studies were excluded if not only middle-aged patients were included, or if they lacked direct comparisons between mechanical and bioprosthetic prostheses. RESULTS: In total, 22 studies with 32,298 patients were included in the final analysis. The results show that patients aged between 50 and 70 receiving AVR with mechanical prostheses achieved better long-term survival and fewer reoperations and valve-related events but suffered more with bleeding events. No significant difference could be found in terms of early mortality and long-term cardiac death. The same results could be observed in the subgroup analysis of RCT and PSM retrospective studies. CONCLUSION: Both mechanical and bioprosthetic prostheses are beneficial to middle-aged patients undertaking AVR procedures. However, mechanical prostheses show better clinical outcomes in long-term survival and comorbidities. Individual recommendation is still necessary.

Adipose tissue macrophage-derived exosomes induce ferroptosis via glutathione synthesis inhibition by targeting SLC7A11 in obesity-induced cardiac injury.

Ferroptosis is an iron-dependent programmed cell death characterized by the accumulation of reactive oxygen species (ROS). Long-term high fat diet (HFD) was found to be associated with ferroptosis and cardiac injury. HFD-induced obesity is characterized by sustained, low-grade inflammation in adipose tissue, while macrophage infiltration plays a crucial role in inflammation. Exosomes (Exos) derived from adipose tissue macrophages (ATMs) participate in the physiological processes of recipient cells. In this study, we investigated the role of ATM-Exos in obesity-induced ferroptosis and cardiac injury. We found that HFD-induced obesity resulted in higher mRNA expression levels of specific markers, e.g., prostaglandin endoperoxide synthase 2 (PTGS2), and increased the levels of lipid peroxides, including malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE). Macrophages infiltrated the adipose tissues, as examined by flow cytometry. Exosomes derived from ATM-Exos were analyzed using transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). Obese ATM-Exos administration induced higher levels of PTGS2, MDA, 4-HNE, lipid ROS, and mitochondrial injury. Obese ATM-Exos further provoked obvious cardiac injury, demonstrated by abnormal levels of cardiac enzymes and inflammatory factors. Systolic left ventricle (LV) function anomalies were induced by ATM-Exos in obese mice. miR-140-5p is abundant in obese ATM-Exos and promotes ferroptosis in cardiomyocytes. Solute carrier family 7 member 11 (SLC7A11) is a downstream target of miR-140-5p, which induces ferroptosis via inhibition of GSH synthesis by targeting SLC7A11. Attenuating exosomal-miR-140-5p expression alleviates ferroptosis and cardiac injury induced by obese ATM exosomes by alleviating GSH inhibition. In summary, the current study provides evidence that obese ATM-exosomal miR-140-5p promotes ferroptosis by regulating GSH synthesis and provides a novel therapeutic strategy for targeting obese ATM-Exos in obesity-induced cardiac injury.

Transcription factor Sp2 promotes TGFB-mediated interstitial cell osteogenic differentiation in bicuspid aortic valves through a SMAD-dependent pathway.

Calcification of the bicuspid aortic valve (BAV) involves differential expression of various RNA genes, which is achieved through complex regulatory networks that are controlled in part by transcription factors and microRNAs. We previously found that miR-195-5p regulates the osteogenic differentiation of valvular interstitial cells (VICs) by targeting the TGF-ß pathway. However, the transcriptional regulation of miR-195-5p in calcified BAV patients is not yet clear. In this study, stenotic aortic valve tissues from patients with BAVs and tricuspid aortic valves (TAVs) were collected. Candidate transcription factors of miR-195-5p were predicted by bioinformatics analysis and tested in diseased valves and in male porcine VICs. SP2 gene expression and the corresponding protein levels in BAV were significantly lower than those in TAV, and a low SP2 expression level environment in VICs resulted in remarkable increases in RNA expression levels of RUNX2, BMP2, collagen 1, MMP2, and MMP9 and the corresponding proteins. ChIP assays revealed that SP2 directly bound to the transcription promoter region of miR-195-5p. Cotransfection of SP2 shRNA and a miR-195-5p mimic in porcine VICs demonstrated that SP2 repressed SMAD7 expression via miR-195-5p, while knockdown of SP2 increased the mRNA expression of SMAD7 and the corresponding protein and attenuated Smad 2/3 expression. Immunofluorescence staining of diseased valves confirmed that the functional proteins of osteogenesis differentiation, including RUNX2, BMP2, collagen 1, and osteocalcin, were overexpressed in BAVs. In Conclusion, the transcription factor Sp2 is expressed at low levels in VICs from BAV patients, which has a negative impact on miR-195-5p expression by binding its promoter region and partially promotes calcification through a SMAD-dependent pathway.

Clinical outcomes following surgical mitral valve plasty or replacement in patients with infectious endocarditis: A meta-analysis.

Background: For degenerative mitral disease, more and more evidences support that mitral valve plasty (MVP) has much better clincial outcomes than mitral valve replacement (MVR). However, the advantages of MVP in patients suffering from infectious endocarditis (IE) are unclear. To evaluate the appropriateness of MVP in IE patients, we conducted this meta-analysis. Based on the difference between active and healed phase, we not only compared the result of patients with IE, but also identified the subgroup with active IE. Methods: We systematically searched the clinical trials comparing clinical outcomes of MVP and MVR in patients suffering from IE. Relevant articles were searched from January 1, 2000 to March 18, 2021 in Pubmed and Cochrane Library. Studies were excluded if they were with Newcastle-Ottawa Scale (NOS) score less than 6 or lacking of direct comparisons between MVP and MVR. Results: 23 studies were involved and 25,615 patients were included. Pooled analysis showed fewer adverse events and early or long-term death in the MVP group. However, more reoperations existed in this patient group. And the reinfection rate was close between two groups. Similar results were observed after identifying active IE subgroup, but there is no difference in the freedom from reoperation due to all-events. Conclusions: Although limitimations exited in this study, patients suffering from IE can benefit from both MVP and MVR. For surgeons with consummate skills, MVP can be the preferred choice for suitable IE patients.

Ultrafast identification of Pinelliae Rhizoma using colorimetric direct-VPCR.

With the increasing growth of the herbal market, a rapid and easy-to-use system is highly desirable in the high-throughput identification of massive herbal medicine samples. Here, an ultrafast and colorimetric detection system was devised based on simplifying template preparation and a newly developed amplification technique, named colorimetric direct-VPCR. The system was successfully applied to the identification of Pinelliae Rhizoma. Compared to the traditional method, the whole test can be finished within 30 min from the sample treatment to the testing results. The method was evaluated by correctly identifying 72 samples obtained from 9 different habitats, demonstrating its high reliability. In summary, we present an ultrafast (less than 30 min) and colorimetric detection platform (under ultraviolet lamp) based on direct-VPCR for the identification of Pinelliae Rhizoma. The high practicability (100% accuracy) of this pipeline enables it to be a promising method in the routine detection of other herbal materials. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-03035-9.

Hinokinin alleviates high fat diet/streptozotocin-induced cardiac injury in mice through modulation in oxidative stress, inflammation and apoptosis.

Type 2 diabetes, a global health concern has been considered as major risk factor for cardiovascular diseases. Hinokinin, an emerging bioactive lignin, is reported to show wide range of pharmacological activities. However, the protective role and mechanisms of Hinokinin against type 2 diabetes-mediated cardiotoxicity are still remains unknown. An experimental type 2 diabetic mice model was created by treating animals with high fat diet for four weeks and intraperitoneal injection of streptozotocin (35 mg/kg body weight). Post-type 2 diabetic induction, animals orally treated with Hinokinin (20 or 40 mg/kg body weight) for six weeks. The type 2 diabetic mice exhibited a rise in blood glucose level as well as glycated hemoglobin (HbA1c %), decrease in weekly body weights, decrease in food intake, reduction in absolute heart weight, fall in serum insulin level with altered lipid profile and cardiac functional damage. Diabetic mice treated with Hinokinin attenuated hyperglycemia, dyslipidemia and cardiac dysfunction. In addition, Hinokinin ameliorated histological alterations, fibrosis and glycated proteins in HFD/STZ-induced mice. Type 2 diabetic condition in mice exacerbated oxidative stress, inflammatory status and apoptosis. Hinokinin treatment significantly assuaged oxidative stress, inflammation and apoptosis and elevated antioxidant defenses in diabetic heart. The underlying mechanisms for such mitigation involved the modulation of Nrf2/Keap1/ARE pathway, MAPKs (JNK, p38 and ERK 1/2) and TLR4/MyD88/NF-κB mediated inflammatory pathways and mitochondrial-dependent (intrinsic) apoptosis pathway. In conclusion, the results of this study provided clear evidence that Hinokinin protects against HFD/STZ (type 2 diabetes)-induced cardiac injury by alleviating oxidative stress, inflammation and apoptosis.

[Development of FTIR fingerprint for identification of armand clematis stem (Chuanmutong) and related herbs].

Armand clematis stem (Clematidis Armandii Caulis, Chuanmutong) is a widely used Chinese herb to disinhibit urine and relieve stranguria. It is difficult to be identified owing to its various macroscopic feature and unknown characteristic compounds. Thus, total of 24 Chuanmutong samples and 7 related herbs including four manshurian aristolochia stem (Aristolochiae Manshuriensis Caulis, Guanmutong) and three akebia stem (Akebiae Caulis, Mutong) samples were collected and analyzed in the range of 4 000 - 400 cm⁻¹ by Fourier Transform Infrared (FTIR) and two-dimensional infrared correlation spectroscopy (2D-FTIR) techniques. The FTIR spectra of 24 Chuanmutong samples are consistent in the spectrum profiles, position and intensity of characteristic peaks. 20 of the 24 Chuanmutong samples were randomly selected as calibration samples to calculate and simulate mean spectrum. This mean spectrum is named as FTIR fingerprint of Chuanmutong with characteristic peaks at 3 412, 2 932, 1 739, 1 639, 1 509, 1 456, 1 426, 1 376, 1 332, 1 261, 1 159, 1 035, 897 ,609 cm⁻¹. Meanwhile, the limited level (Mean-3σ=0.992 6) to identify true or false Chuanmutong by correlation coefficient of FTIR spectra was calculated based on the 20 Chuanmutong calibration samples. Then, the rest 4 Chuanmutong, 4 Guanmutong and 3 Mutong samples were used as validation samples to evaluate the identification efficacy. The result shows that the FTIR spectra of 4 Chuanmutong validation samples were similar to the fingerprint. Their correlation coefficients of FTIR spectra were over the limited level and accepted as Chuanmutong. However, the spectra of Guanmutong and Mutong were significantly different from Chuanmutong fingerprint. The correlation coefficients of Guanmutong (0.902 1-0.940 4, n=4) and Mutong (0.954 9-0.978 9, n=3) FTIR spectra were less than the limited level and rejected from Chuanmutong. Furthermore, the number, position and intensity of auto-peaks on the 2D-FTIR were drastically different among the three herbs. It is concluded that the developed FTIR fingerprinting can be rapidly and accurately identify Chuanmutong and differentiate from related herbs.