Diabetic cardiomyopathy is characterized by diabetes‐induced myocardial abnormalities, accompanied by inflammatory response and alterations in inflammation‐related signalling pathways. Kirenol, isolated from Herba Siegesbeckiae, has potent anti‐inflammatory properties. In this study, we aimed to investigate the cardioprotective effect of kirenol against DCM and underlying the potential mechanisms in a type 2 diabetes mellitus model. Kirenol treatment significantly decreased high glucose‐induced cardiofibroblasts proliferation and increased the cardiomyocytes viability, prevented the loss of mitochondrial membrane potential and further attenuated cardiomyocytes apoptosis, accompanied by a reduction in apoptosis‐related protein expression. Kirenol gavage could affect the expression of pro‐inflammatory cytokines in a dose‐dependent manner but not lower lipid profiles, and only decrease fasting plasma glucose, fasting plasma insulin and mean HbA1c levels in high‐dose kirenol‐treated group at some time‐points. Left ventricular dysfunction, hypertrophy, fibrosis and cell apoptosis, as structural and functional abnormalities, were ameliorated by kirenol administration. Moreover, in diabetic hearts, oral kirenol significantly attenuated activation of mitogen‐activated protein kinase subfamily and nuclear translocation of NF‐κB and Smad2/3 and decreased phosphorylation of IκBα and both fibrosis‐related and apoptosis‐related proteins. In an Electrophoretic mobility shift assay, the binding activities of NF‐κB, Smad3/4, SP1 and AP‐1 in the nucleus of diabetic myocardium were significantly down‐regulated by kirenol treatment. Additionally, high dose significantly enhanced myocardial Akt phosphorylation without intraperitoneal injection of insulin. Kirenol may have potent cardioprotective effects on treating for the established diabetic cardiomyopathy, which involves the inhibition of inflammation and fibrosis‐related signalling pathways and is independent of lowering hyperglycaemia, hyperinsulinemia and lipid profiles.

Background: The mevalonate pathway generates endogenous cholesterol and intermediates including geranylgeranyl pyrophosphate (GGPP). By reducing GGPP production, statins exert pleiotropic or cholesterol-independent effects. The potential regulation of GGPP homeostasis through dietary intake and the interaction with concomitant statin therapy is unknown. Methods: We developed a sensitive HPLC technique to quantify dietary GGPP and conducted proteomics, qRT-PCR screening and western blot to determine signaling cascades, gene expression, protein-protein interaction and protein membrane trafficking in wild type and transgenic rats. Results: GGPP contents were highly variable depending on food source that differentially regulated blood GGPP levels in rats. Diets containing intermediate and high GGPP reduced or abolished the effects of statins in rats with hypoxia- and monocrotaline-induced pulmonary hypertension: this was rescuable by methyl-allylthiosulfinate and methyl-allylthiosulfinate-rich garlic extracts. In human pulmonary artery smooth muscle cells (HPASMCs) treated with statins, hypoxia activated RhoA in an extracellular GGPP-dependent manner. Hypoxia-induced ROCK2/Rab10 signaling was prevented by statin and recovered by exogenous GGPP. The hypoxia-activated RhoA/ROCK2 pathway in rat and HPASMCs upregulated the expression of Ca 2+ -sensing receptor (CaSR) and hypoxia-induced mitogenic factor/FIZZ1 (HIMF), a mechanism attenuated by statin treatment and regained with exogenous GGPP. Rab10-knockdown almost abrogated hypoxia-promoted CaSR membrane-trafficking, a process diminished by statin and resumed by exogenous GGPP. Hypoxia-induced pulmonary hypertension was reduced in rats with CaSR mutated at the binding motif of HIMF and the interaction between dietary GGPP and statin efficiency was abolished. In humans fed with a high GGPP diet, blood GGPP levels were increased, and this abolished statin-lowering effects on plasma GGPP and hypoxia-enhanced RhoA activity of blood monocytes that were both also rescued by garlic extracts. Conclusions: There is important dietary regulation of GGPP levels that interferes with the effects of statin therapy in experimental pulmonary hypertension. These observations rely on a key and central role of i) RhoA-ROCK2 cascade activation and ii) Rab10-faciliated CaSR membrane trafficking with iii) subsequent overexpression and binding of HIMF to CaSR. These findings warrant clinical investigation for the treatment of pulmonary hypertension and perhaps other diseases by combining statin together with garlic-derived methyl-allylthiosulfinate or garlic extracts and thus circumventing dietary GGPP variations.

Experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS), is characterized by CNS demyelination mediated by autoreactive T cells. Kirenol, a biologically active substance isolated from Herba Siegesbeckiae, has potent anti-inflammatory activities. Here we investigated effects of kirenol on EAE. Kirenol treatment markedly delayed onset of disease and reduced clinical scores in EAE mice. Kirenol treatment reduced expression of IFN-γ and IL-17A in the serum and proportion of Th1 and Th17 cells in draining lymph nodes. Priming of lymphocytes was reduced and apoptosis of MOG-activated CD4+ T cells was increased in kirenol treated EAE mice. Kirenol treatment of healthy animals did not affect the lymphocytes in these non-immunized mice. Further in vitro studies showed that kirenol inhibited viability of MOG-specific lymphocytes and induced apoptosis of MOG-specific CD4+ T cells in a dose- and time-dependent manner. Kirenol treatment upregulated Bax,downregulated Bcl-2,and increased activation of caspase-3 and release of cytochrome c, indicating that a mitochondrial pathway was involved in kirenol induced apoptosis. Moreover, pretreatment with either a pan-caspase inhibitor z-VAD-fmk or a more specific caspase 3 inhibitor Ac-DEVD-CHO in lymphocytes reduced kirenol induced apoptosis. Our findings implicate kirenol as a useful agent for the treatment of MS.

Objective: The cardiotoxicity of doxorubicin (DOX) reduces the quality of life and prognosis of cancer patients, and therefore its clinical application has been largely restricted. This study aimed to assess the effects of cryptotanshione (CPT) on DOX-induced rat cardiac insufficiency. Results: CPT treatment significantly suppressed apoptosis in vitro. The oral administration of CPT significantly improved cardiac function in the rat model, reduced collagen production and suppressed apoptosis and the production of reactive oxygen species in the heart tissue. Transcriptomic profiling and its relevant bioinformatics analysis showed that CPT suppressed doxorubicin-induced cardiotoxicity by inhibiting p53 signaling pathway. Conclusion: Transcriptomic profiling and bioinformatics analysis can be used to evaluate the cardio-protective effect of CPT through inactivating p53 signaling pathway in the doxorubicin-mediated myocardial damage model. Methods: F-actin staining and flow cytometry were used to assess the effects of CPT on cardiomyocytes. In vivo, echocardiography and hemodynamic evaluation were used to assess the effects of CPT on the cardiac dysfunction in rats. Furthermore, transcriptomic profiling and bioinformatics analysis, as well as western blot analysis, were used to determine that CPT induced changes in the signaling pathways in the model.

Background: Pulmonary arterial hypertension is an incurable disease, in which the extracellular CaSR (calcium sensing receptor) is mechanistically important. This study was aimed to genetically link the CaSR gene and function to the disease severity. Methods: Sanger sequencing, Sugen/hypoxia pulmonary arterial hypertension rat model, CaSR mutated rat, transcriptional reporter assay and measurement of CaSR activity were used. Results: Sanger sequencing identified a significant association between the variant rs1042636(A>G), located in CaSR exon 7, and idiopathic pulmonary arterial hypertension (IPAH) formation in patients. The frequency of 2968G homozygotes was higher in patients with IPAH compared with healthy individuals (23.6% versus 17.5%; P =0.001, OR=1.864), and the minor alleles of rs6776158, rs1048213, and rs9883099, located in CaSR promoter, raised the IPAH odds ratio to 2.173. Patients with IPAH carrying heterozygotes or homozygotes genotype of rs1042636 showed markedly higher pulmonary artery pressure and reduced survival compared with individuals carrying the wild-type allele. The minor alleles of rs6776158, rs1048213, and rs9883099 increased CaSR expression in reporter assay. In Sugen/hypoxia pulmonary arterial hypertension rats, the point mutation replicating rs1042636 found in IPAH exacerbated pulmonary arterial hypertension severity by promoting the overexpression and the enhanced activity of CaSR. Conclusions: Our functional genomic analysis thus indicates that the CaSR minor alleles of rs1042636, rs6776158, rs1048213, and rs9883099 contribute to the development and severity of IPAH. These findings may benefit clinical prognosis and treatment for IPAH.