Enhancing aortic valve drug delivery with PAR2-targeting magnetic nano-cargoes for calcification alleviation.

Calcific aortic valve disease is a prevalent cardiovascular disease with no available drugs capable of effectively preventing its progression. Hence, an efficient drug delivery system could serve as a valuable tool in drug screening and potentially enhance therapeutic efficacy. However, due to the rapid blood flow rate associated with aortic valve stenosis and the lack of specific markers, achieving targeted drug delivery for calcific aortic valve disease has proved to be challenging. Here we find that protease-activated-receptor 2 (PAR2) expression is up-regulated on the plasma membrane of osteogenically differentiated valvular interstitial cells. Accordingly, we develop a magnetic nanocarrier functionalized with PAR2-targeting hexapeptide for dual-active targeting drug delivery. We show that the nanocarriers effectively deliver XCT790-an anti-calcification drug-to the calcified aortic valve under extra magnetic field navigation. We demonstrate that the nano-cargoes consequently inhibit the osteogenic differentiation of valvular interstitial cells, and alleviate aortic valve calcification and stenosis in a high-fat diet-fed low-density lipoprotein receptor-deficient (Ldlr-/-) mouse model. This work combining PAR2- and magnetic-targeting presents an effective targeted drug delivery system for treating calcific aortic valve disease in a murine model, promising future clinical translation.

FAIM Enhances the Efficacy of Mesenchymal Stem Cell Transplantation by Inhibiting JNK-Induced c-FLIP Ubiquitination and Degradation.

Background: The poor survival rates of transplanted mesenchymal stem cells (MSCs) in harsh microenvironments impair the efficacy of MSCs transplantation in myocardial infarction (MI). Extrinsic apoptosis pathways play an important role in the apoptosis of transplanted MSCs, and Fas apoptosis inhibitory molecule (FAIM) is involved in regulation of the extrinsic apoptosis pathway. Thus, we aimed to explore whether FAIM augmentation protects MSCs against stress-induced apoptosis and thereby improves the therapeutic efficacy of MSCs. Methods: We ligated the left anterior descending coronary artery (LAD) in the mouse heart to generate an MI model and then injected FAIM-overexpressing MSCs (MSCsFAIM) into the peri-infarction area in vivo. Moreover, FAIM-overexpressing MSCs were challenged with oxygen, serum, and glucose deprivation (OGD) in vitro, which mimicked the harsh microenvironment that occurs in cardiac infarction. Results: FAIM was markedly downregulated under OGD conditions, and FAIM overexpression protected MSCs against OGD-induced apoptosis. MSCsFAIM transplantation improved cell retention, strengthened angiogenesis, and ameliorated heart function. The antiapoptotic effect of FAIM was mediated by cellular-FLICE inhibitory protein (c-FLIP), and FAIM augmentation improved the protein expression of c-FLIP by reducing ubiquitin-proteasome-dependent c-FLIP degradation. Furthermore, FAIM inhibited the activation of JNK, and treatment with the JNK inhibitor SP600125 abrogated the reduction in c-FLIP protein expression caused by FAIM silencing. Conclusions: Overall, these results indicated that FAIM curbed the JNK-mediated, ubiquitination-proteasome-dependent degradation of c-FLIP, thereby improving the survival of transplanted MSCs and enhancing their efficacy in MI. This study may provide a novel approach to strengthen the therapeutic effect of MSC-based therapy.

SRT1720 Pretreatment Promotes Mitochondrial Biogenesis of Aged Human Mesenchymal Stem Cells and Improves Their Engraftment in Postinfarct Nonhuman Primate Hearts.

Declined function of aged mesenchymal stem cells (MSCs) diminishes the benefits of cell therapy for myocardial infarction (MI). Our previous study has demonstrated that SRT1720, a specific SIRT1 activator, could protect aged human MSCs (hMSCs) against apoptosis. The purpose of the present study was to investigate the role of mitochondria in the antiapoptotic effects of SRT1720. In addition, we established a nonhuman primate MI model to evaluate cell engraftment of SRT1720-pretreated aged hMSCs (SRT1720-OMSCs). A hydrogen peroxide (H2O2)-induced apoptosis model was established in vitro to mimic MI microenvironment. Compared with vehicle-treated aged hMSCs (Vehicle-OMSCs), SRT1720-OMSCs showed alleviated apoptosis level, significantly decreased caspase-3 and caspase-9 activation, and reduced release of cytochrome c when subjected to H2O2 treatment. Mitochondrial contents were compared between young and aged hMSCs and our data showed that aged hMSCs had lower mitochondrial DNA (mtDNA) copy numbers and protein expression levels of components of the mitochondrial electron transport chain (ETC) than young hMSCs. Also, treatment with SRT1720 resulted in enhanced MitoTracker staining, increased mtDNA levels and expression of mitochondrial ETC components in aged hMSCs. Furthermore, SRT1720-OMSCs exhibited elevated mitochondrial respiratory capacity and higher mitochondrial membrane potential. In vivo study demonstrated that SRT1720-OMSCs had higher engraftment rates than Vehicle-OMSCs at 3 days after transplantation into the infarcted nonhuman primate hearts. Taken together, these results suggest that SRT1720 promotes mitochondrial biogenesis and function of aged hMSCs, which is involved in its protective effects against H2O2-induced apoptosis. These findings encourage further exploration of the optimization of aged stem cells function via regulating mitochondrial function.

Aldo-keto reductase family 1 member B induces aortic valve calcification by activating hippo signaling in valvular interstitial cells.

AIMS: Calcific aortic valve disease (CAVD) is a primary cause of cardiovascular mortality; however, its mechanisms are unknown. Currently, no effective pharmacotherapy is available for CAVD. Aldo-keto reductase family 1 member B (Akr1B1) has been identified as a potential therapeutic target for valve interstitial cell calcification. Herein, we hypothesized that inhibition of Akr1B1 can attenuate aortic valve calcification. METHODS AND RESULTS: Normal and degenerative tricuspid calcific valves from human samples were analyzed by immunoblotting and immunohistochemistry. The results showed significant upregulation of Akr1B1 in CAVD leaflets. Akr1B1 inhibition attenuated calcification of aortic valve interstitial cells in osteogenic medium. In contrast, overexpression of Akr1B1 aggravated calcification in osteogenic medium. Mechanistically, using RNA sequencing (RNAseq), we revealed that Hippo-YAP signaling functions downstream of Akr1B1. Furthermore, we established that the protein level of the Hippo-YAP signaling effector active-YAP had a positive correlation with Akr1B1. Suppression of YAP reversed Akr1B1 overexpression-induced Runx2 upregulation. Moreover, YAP activated the Runx2 promoter through TEAD1 in a manner mediated by ChIP and luciferase reporter systems. Animal experiments showed that the Akr1B1 inhibitor epalrestat attenuated aortic valve calcification induced by a Western diet in LDLR-/- mice. CONCLUSION: This study demonstrates that inhibition of Akr1B1 can attenuate the degree of calcification both in vitro and in vivo. The Akr1B1 inhibitor epalrestat may be a potential treatment option for CAVD.

Knockdown of estrogen-related receptor α inhibits valve interstitial cell calcification in vitro by regulating heme oxygenase 1.

Calcific aortic valve disease (CAVD) is the most common valvular heart disease in adults. The cellular mechanisms of CAVD are still unknown, but accumulating evidence has revealed that osteogenic differentiation of human valve interstitial cells (hVICs) plays an important role in CAVD. Thus, we aimed to investigate the function of estrogen-related receptor α (ERRα) in the osteogenic differentiation of hVICs. We found that the level of ERRα was significantly increased in CAVD samples compared to normal controls. In addition, ERRα was significantly upregulated during hVIC osteogenic differentiation in vitro. Gain- and loss-of-function experiments were performed to identify the function of ERRα in hVIC calcification in vitro. Inhibition of endogenous ERRα attenuated hVIC calcification, whereas overexpression of ERRα in hVICs promoted this process. RNA sequencing results suggested that heme oxygenase-1 (Hmox1) was a downstream target of ERRα, which was further confirmed by western blotting. Additionally, we also found that downregulation of Hmox1 by shHmox1 efficiently reversed the inhibition of calcification induced by ERRα shRNA in hVICs. ChIP-qPCR and luciferase assays indicated that Hmox1 was negatively regulated by ERRα. We found that overexpression of Hmox1 or its substrates significantly inhibited hVIC calcification in vitro. In conclusion, we found that knockdown of ERRα can inhibit hVIC calcification through upregulating Hmox1 and that ERRα and Hmox1 are potential targets for the treatment of CAVD.

TPP1 Enhances the Therapeutic Effects of Transplanted Aged Mesenchymal Stem Cells in Infarcted Hearts via the MRE11/AKT Pathway.

BACKGROUND: Poor cell survival after transplantation restricts the therapeutic potential of mesenchymal stem cell (MSC) transplantation into infarcted hearts, particularly in older individuals. TPP1, a component of the shelterin complex that is involved in telomere protection, is highly expressed in young MSCs but declines in aged ones. Here, we explore whether TPP1 overexpression in aged mouse MSCs improves cell viability in vivo and in vitro. METHODS: Aged mouse MSCs overexpressing TPP1 were injected into the peri-infarct area of the mouse heart after left anterior descending coronary artery ligation. In parallel, to evaluate cellular-level effects, H2O2 was applied to MSCs in vitro to mimic the microenvironment of myocardial injury. RESULTS: In vivo, the transplantation of aged MSCs overexpressing TPP1 resulted in improved cell survival, enhanced cardiac function, and reduced fibrosis compared to unmodified aged MSCs. In vitro, TPP1 overexpression protected aged MSCs from H2O2-induced apoptosis and enhanced DNA double-strand break (DSB) repair. In addition, the phosphorylation of AKT and the key DSB repair protein MRE11 were both significantly upregulated in aged MSCs that overexpressed TPP1. CONCLUSIONS: Our results reveal that TPP1 can enhance DNA repair through the AKT/MRE11 pathway, thereby improving the therapeutic effects of aged MSC transplantation and offering significant potential for the clinical application of autologous transplantation in aged patients.

Uptake and transformation of decabromodiphenyl ether in different rice cultivars: Evidence from a carbon-14 study.

The differences of PBDE absorption, accumulation, and metabolism in different cultivars of the same crop are rarely explored. This study used 14C tracing to fully demonstrate the uptake and transformation of soil-borne BDE209 in three rice cultivars, including two indica (HHZ and YD1) and one japonica cultivars (NJ3). Results showed that about 6.9, 17.2, and 17.4% of the applied 14C-BDE209 were transformed to 14C-metabolites in soils planted with HHZ, YD1, and NJ3, respectively. The 14C-BDE209 and its 14C-metabolites in soil could be absorbed by the rice and gradually transported to its root, stem, leaf, and grain, with the total whole-plant uptake of 8.52, 4.55 and 3.43 nmol for HHZ, YD1, and NJ3, respectively. The cultivar of HHZ had the greatest whole-plant 14C absorption but the lowest ΣPBDEs residues in its grain, with the ΣPBDEs of 421.8, 454.2 and 967.0 ng g-1 for HHZ, YD1, and NJ3, respectively. BDE-209 accounted for 90%, 31% and 50% of the ΣPBDEs in the grain from HHZ, YD1, and NJ3, respectively. The estimated daily intake (EDI) amounts of ΣPBDEs were 928, 1056, and 2675 ng kg-1 bw d-1 via consuming rice grains from HHZ, YD1, and NJ3, respectively, which were below the safe threshold limits for human consumption. This study proved the different BDE-209 absorption, accumulation and transformation in different rice cultivars, which potentially suggests the need of considering cultivar differences in assessing the dietary risks of PBDEs.