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Cardiovascular diseases are fatal threats to human health and also important fields in drug discovery. Organoid is a miniature with the structure and function similar to the organ, which is formed by the self-updating and specific differentiation of stem cells during the in vitro culture. Considering its characteristics of human origin, physical features, self-assembling and genetic stability, heart organoid has attracted much attention in the study of cardiogenesis, cardiovascular diseases modeling and related drug research. Hence, this article will review the development of heart organoids and its construction strategies, highlighting its application and prospects in drug discovery.
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This study investigated the effect of puerarin on human umbilical vein endothelial cells (HUVEC) injured with hydrogen peroxide (H2O2). HUVEC were divided into three groups: a control group, a model group (H2O2 400 μmol·L-1) and a puerarin-treated group (3, 10, 30 and 100 μmol·L-1). HUVEC were cultured with varied concentration of puerarin for 2 h and treated with H2O2 for another 24 h. Cell proliferation was detected by a CCK-8 assay. The mitochondrial membrane potential was measured by a JC-1 fluorescent probe. A transwell chamber assay was adopted to observe cell migration ability. Mitochondrial respiratory function was measured in a two-chamber titration injection respirometer (Oxygraph-2k). The expression of interleukin-1β (IL-1β), interleukin-18 (IL-18) and tumor necrosis factor-α (TNF-α) was detected by quantitative real-time PCR. The expression of pyroptosis-mediated proteins, including cleaved-cysteinyl aspartate-specific proteinase-1 (caspase-1), N-gasdermin D (N-GSDMD), NOD-like receptor protein 3 (NLRP3) and purinergic ligand-gated ion channel 7 receptor (P2X7R) was detected by Western blot. The results show that 400 μmol·L-1 H2O2 treatment for 24 h causes obvious damage to HUVEC. Compared with the model group, puerarin protected against cellular injury in a dose-dependent manner, with the greatest effect at a dose of 30 and 100 μmol·L-1. Puerarin significantly decreased the mitochondrial membrane potential and improved mitochondrial function. Puerarin inhibited cell migration induced by H2O2, suppressed the expression of IL-1β, IL-18 and TNF-α, and down-regulated the pyroptosis-mediated protein. These changes are statistically significant (P < 0.05). These findings demonstrate that puerarin has a protective effect against H2O2-induced oxidative damage of HUVEC by inhibiting the migration of HUVEC cells. The mechanism may be related to improved mitochondrial respiratory function and inhibition of pyroptosis.
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Aim Timely re establishment of coronary blood How in patients with myocardial infarction is the cornerstone of their treatment; however, substantial amount of damage can oecur as a consequence of reperfusion.In recent years it has been found that receptor interacting protein kinase 3 ( RIPK3 ) contributes remarkably to myocardial ischemia-reperfusion injury (MIRI).RIPK3 can regulate necroptosis through RIPK1/RIPK3/MLKL and CaMKII, respectively, and participate in the MIRI process.This artiele reviews the researeh progress of RIPK3-mediated ne¬ croptosis involved in MIRI from endoplasmic reticulum stress, mitochondrial fragmentation disturbanee, cardiac microvascular dysfunction and inflammation, and focuses on whether RIPK3 can be used as a new target for anti-MIRI, so as to provide a new strategy and choice for improving the clinical treatment effect and prognosis of ischemic heart disease.
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Compound reserpine and triamterene tablets (CRTT), a compound antihypertensive drug developed by Chinese scientists, is still widely used in clinical practice. However, the mechanisms by which CRTT treats hypertension remain to be fully understood. This study used network pharmacology to analyze CRTT's antihypertensive mechanisms with in vitro experiments. The targets of the four chemical components of CRTT were collected from the Swiss Target Prediction database; 1 828 protein targets related to hypertension were collected from the Therapeutic Target Database (TTD) and Online Mendelian Inheritance in Man (OMIM) database. The CRTT-hypertension network model was constructed using a search tool for recurring instances of neighbouring genes (STRING). Gene ontology (GO) and pathway enrichment analysis of targets of interest was conducted with the Metascape database. In the in vitro study, human umbilical vein endothelial cells (HUVEC) and vascular smooth muscle cells (VSMC) were treated with 1 μmol·L-1 angiotensin Ⅱ (AngⅡ) and CRTT was administered at concentrations of 0.01, 0.1, and 1 μmol·L-1. Changes in the phosphatidylinositol-3-kinase/protein serine threonine kinase/endothelial nitric oxide synthase (PI3K/Akt/eNOS) pathway in HUVEC and the cyclic guanosine monophosphate/cGMP-dependent protein kinase (cGMP/PKG) pathway in VSMC were determined by Western blot. Network pharmacological analysis revealed that the antihypertensive effect of CRTT is closely associated with biological pathways such as vascular tone regulation, adrenergic receptor activation, protein kinase activity and signaling pathways such as the cGMP/PKG signaling pathway, vascular smooth muscle contraction, neuroactive ligand-receptor interaction, adrenergic signaling in cardiomyocytes and calcium signaling pathways. The in vitro study confirmed that CRTT increased the levels of phosphorylated phosphatidylinositol-3-kinase (p-PI3K), phosphorylated protein serine threonine kinase (p-Akt), phosphorylated endothelial nitric oxide synthase (p-eNOS) in HUVEC and the levels of eNOS, phosphorylated vasodilator-stimulated phosphoprotein (p-VASP), and PKG in VSMC through multiple targets and pathways. These results suggest that the activation of PI3K/Akt/eNOS pathway and endothelial-dependent NO/cGMP signaling may be involved in the CRTT-mediated hypotensive effect.
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This study was to investigate the protective effects of puerarin on myocardial ischemia/reperfusion (MI/R) injury and the underlying mechanism. The MI/R-model was established by ligating the left anterior descending artery (LAD) for 60 min followed by 24 h reperfusion, puerarin (10, 30, and 100 mg·kg-1) was orally administered 20 min before reperfusion. Cardiac function, myocardial infarct index, cardiac damage markers, inflammatory cytokines, and apoptosis index were measured to evaluate the protective effects of puerarin on MI/R injury. The activation of Nod-like receptor protein 3 (NLRP3) inflammasome and Toll like receptor 4 (TLR4)/myeloid differentiation factor 88 (Myd88)/nuclear factor kappa B (NF-κB) pathway were determined by Western blot. All animal experimental procedures were approved by the ethics committee of the Institute of Materia Medica, Peking Union Medical College, Chinese Academy of Medical Sciences. The results showed that puerarin could significantly improve cardiac function, reduce myocardial infarct size, decease the levels of lactic dehydrogenase (LDH), aspartate transaminase (AST), creatine kinase-MB (CK-MB), and cardiac troponin T (cTnT) and suppress cardiomyocyte apoptosis. Meanwhile, puerarin could notably decrease the levels of inflammatory cytokines such as interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α). Western blot analysis revealed that puerarin could downregulate the expression of TLR4, Myd88, NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), cleaved-caspase 1, cleaved-gasdermin-D (GSDMD), IL-1β, and IL-18, as well as the phosphorylation levels of inhibitor of NF-κB α (IκBα), IκB kinase β (IKKβ), and NF-κB. These findings demonstrated that puerarin could alleviate MI/R injury by suppressing NLRP3 inflammasome activation, possibly via TLR4/Myd88/NF-κB pathway.
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Mitochondria play a key role in cell metabolism. In addition to synthesizing ATP, they also participate in many physiological and pathological processes, including apoptosis, inflammation, oxidative stress, neuronal disease, tumor development, and aging. Most gene transcription of mitochondrial proteins occurs in the nucleus, so the biogenesis of mitochondria and the maintenance of mitochondrial homeostasis mainly depend on the expression of nuclear genes (nDNA) and mitochondria-nucleus interactions. Conversely, mitochondria can affect the expression of nuclear genes through nuclear transcription factors, a process called mitochondrial retrograde signaling. This review summarizes the research progress on mitochondria-nucleus retrograde signaling and its regulation, including the ways by which mitochondria regulate nuclear genes and affect biological processes, and discusses new strategies for the treatment of diseases that involve mitochondrial retrograde signaling in disease pathology.
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Objective To observe the clinical efficacy of Tangzhiping Prescription on non-proliferative diabetic retinopathy (NPDR) and the effects on serum vascular epithelial growth factor (VEGF) and pigment epithelium derived factor (PEDF). Methods Totally 86 NPDR patients were randomly divided into treatment group (43 cases) and control group (43 cases). Both groups were given hypoglycemic, anti hypertensive and lipid-regulatory basic therapy. The control group was given calcium distillate capsules, 0.5 g per time, 3 times a day, orally; the treatment group was treated with Tangzhiping Prescription based on the control group, 1 dosage per day, twice a day, orally. Four weeks were set as one treatment course. Treatment for both groups lasted for three courses. Clinical efficacy and fundus efficacy of both groups were evaluated. TCM symptom scores, fundus scores, and visual condition were observed; FPG, 2 h PG, Hb A1 C, TC, TG, HDL-C and LDL-C and changes in the contents of VEGF and PEDF were detected. Results The control group and the treatment group lost 2 and 3 cases respectively. The total effective rate of clinical efficacy and total fundus efficiency of the control group were 65.00％ and 68.35％, respectively, and the treatment group were 87.50％ and 84.62％ respectively, with statistical significance (P＜0.05). Compared with before treatment, TCM symptom scores and fundus scores decreased significantly after treatment in both groups (P＜0.01); After treatment, the TCM symptom scores and fundus scores in the treatment group were lower than those in the control group (P＜0.05, P＜0.01). Compared with before treatment, visual acuity improved significantly after treatment in both groups (P＜0.05, P＜0.01). Compared with before treatment, levels of FPG, 2 h PG, Hb A1 c, TC, TG, and LDL-C decreased significantly in both groups after treatment (P＜0.01). After treatment, the levels of 2 h PG, Hb A1 c, TC, TG, and LDL-C in the treatment group were significantly lower than the control group (P＜0.05, P＜0.01). Compared with before treatment, the levels of VEGF decreased and PEDF levels increased in both groups, with statistical significance (P＜0.01). After treatment, the improvement of VEGF and PEDF in the treatment group was better than that in the control group (P＜0.05, P＜0.01); negative correlation was found between VEGF and PEDF (r=-0.320, P＜0.01). Conclusion Tangzhiping Prescription can effectively improve the clinical symptoms of patients with NPDR and slow down the progress of NPDR via reducing the blood glucose and blood lipids, and regulating the contents of VEGF and PEDF.
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OBJECTIVE To investigate the effects of total flavonoids of bugloss(TFB)on left ventricular (LV)remodeling after myocardial infarction(MI),LV size and function was compared in mice subjected to left anterior descending coronary artery ligation. METHODS 28 d after MI, the infarcted fraction of the LV and LV mass, systolic and diastolic function were measured. Capillary density and myocyte width in the nonischemic portion of the LV were also determined.RESULTS 28 d after MI,both groups had dilated LVs with decreased fractional shortening and lower ejection fractions. Although the infarcted size of the LV was similar in both groups,LV end-diastolic internal diameter,end-diastolic volume,and mass were lower, but fractional shortening, ejection fraction, and the maximum rate of developed LV pressure(dp/dtmax)were greater in TFB treated mice than in control mice.Impairment of diastolic func-tion, as measured by the time constant of isovolumic relaxation (t) and the maximum rate of LV pres-sure decay(dp/dtmin),was more marked in control mice than in TFB treated mice.Mortality after MI was greater in control mice than in TFB treated mice.In control mice,capillary density and myocyte width in the nonischemic portion of the LV did not differ before and 28 days after MI, whereas in TFB treated mice, capillary density increased and myocyte width declined after MI. CONCLUSION These results suggest that the presence of TFB limits LV dysfunction and remodeling in a murine model of MI in part by decreasing myocyte hypertrophy in the remote myocardium.
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This study is to investigate the effect of total flavonoids of Uygur medicine bugloss (BTF) on rats with myocardial ischemia/reperfusion injury, and to explore the mechanisms by which it acts. Left anterior descending (LAD) coronary artery in rats was occluded for 30 min followed by 4 h reperfusion. Meanwhile, BTF dissolved in saline was administered intraperitoneally at dosage of 10, 30 and 50 mg x kg(-1). Electrocardiograph, infarction index, serum myocardial enzymes and heart function were determined to evaluate the effect of BTF. Some other observations were carried out to explore whether inhibiting inflammation and apoptosis is involved in the mechanisms underlying BTF. Our results showed that in ischemia/reperfusion injured rats BTF could dose-dependently reduce myocardial infarction index and myocardial enzyme leakage, and enhance heart function, indicating that it possesses significant cardio protection. ELISA analysis showed that BTF could decrease the content of myocardial inflammatory cytokines such as IL-1beta, IL-6 and TNF-alpha. Western-blotting confirmed that BTF could increase the expression of anti-apoptotic protein Bcl-2 and reduce the expression of proapoptosis protein Bax. Further more, the phosphorylation level of PI3K and Akt was upregulated by BTF treatment. BTF can protect rat against myocardial ischemia/reperfusion injury. Anti-inflammation and inhibition of apoptosis through upregulating PI3K/Akt signal pathway may contribute to the protective effect of BTF.
Subject(s)
Animals , Rats , Apoptosis , Apoptosis Regulatory Proteins , Boraginaceae , Chemistry , Flavonoids , Pharmacology , Heart , Interleukin-6 , Myocardial Infarction , Myocardial Reperfusion Injury , Drug Therapy , Myocardium , Phosphatidylinositol 3-Kinases , Phosphorylation , Protective Agents , Proto-Oncogene Proteins c-akt , Signal Transduction , Tumor Necrosis Factor-alpha , bcl-2-Associated X ProteinABSTRACT
Nine compounds were isolated and purified by column chromatographic techniques including macroporous resin, silica gel, ODS, Sephadex LH-20, and preparative reversed-phase HPLC. Their structures were elucidated as taxifolin (1), naringenin (2), chalconaringenin (3), acacetin (4), quercetin 3-O-beta-D-galactopyranoside (5), 6-prenylnaringenin (6) xanthohumol (7), desmethylxanthohumol (8), xanthohumol B (9) on the basis of MS and NMR spectroscopic data analysis. Compounds 1-5 were isolated from Humulus lupulus for the first time.
Subject(s)
Dextrans , Chemistry , Drugs, Chinese Herbal , Chemistry , Flavanones , Chemistry , Flavonoids , Chemistry , Humulus , Chemistry , Magnetic Resonance Spectroscopy , Mass Spectrometry , Molecular Structure , Quercetin , ChemistryABSTRACT
AIM@#To investigate the relationship between cerebroprotection of pinocembrin and epoxyeicosatrienoic acids (EETs) and their regulating enzyme soluble epoxide hydrolase (sEH).@*METHODS@#Rats underwent middle cerebral artery occlusion (MCAO) to mimic permanent focal ischemia, and pinocembrin was administrated via tail vein injection at 10 min, 4 h, 8 h and 23 h after MCAO. After 24 MCAO, rats were re-anesthetized, and the blood and brain were harvested and analyzed.@*RESULTS@#Pinocembrin displayed significant protective effects on MCAO rats indicated by reduced neurological deficits and infarct volume. Importantly, co-administration of 0.2 mg·kg(-1) 14, 15-EEZE, a putative selective EET antagonist, weakened the beneficial effects of pinocembrin. 14, 15-EET levels in the blood and brain of rats after 24 h MCAO were elevated in the presence of pinocembrin. In an assay for hydrolase activity, pinocembrin significantly lowered brain sEH activity of MCAO rats and inhibited recombinant human sEH activity in a concentration-dependent manner (IC50, 2.58 μmol·L(-1)). In addition, Western blot and immunohistochemistry analysis showed that pinocembrin at doses of 10 mg·kg(-1) and 30 mg·kg(-1) significantly down-regulated sEH protein in rat brain, especially the hippocampus CA1 region of MCAO rats.@*CONCLUSION@#Inhibiting sEH and then increasing the potency of EETs may be one of the mechanisms through which pinocembrin provides cerebral protection.
Subject(s)
Animals , Humans , Male , Rats , Arachidonic Acids , Metabolism , Brain , Metabolism , Brain Ischemia , Drug Therapy , Genetics , Metabolism , Disease Models, Animal , Epoxide Hydrolases , Genetics , Metabolism , Flavanones , Protective Agents , Rats, Sprague-DawleyABSTRACT
To screen potential hamster chymase 2 inhibitors, a high-throughput screening (HTS) model was established. Recombinant hamster chymase 2 with active form was cloned and expressed in E. coli. The HTS model with total volume of 50 microL in 384-well microplate was based on fluorescence analysis and was proved sensitive as well as specific (Z' = 0.84). A total of 40 080 samples (including 28 060 compounds and 12 020 natural products) were screened, and 613 samples with inhibition greater than 90% were selected for further rescreening. Finally, compounds J16647 and J16648 were identified with high inhibitory activity on chymase 2, and whose IC50 values were 0.823 and 0.690 micromol x L(-1), respectively.
Subject(s)
Animals , Cricetinae , Rats , Chymases , Enzyme Inhibitors , Pharmacology , Escherichia coli , Metabolism , High-Throughput Screening Assays , Methods , Inhibitory Concentration 50 , Structure-Activity RelationshipABSTRACT
To screen potential human soluble protein tyrosine phosphatase 1B (PTP1B) inhibitors, a high-throughput screening (HTS) model in 384-well microplate with total volume of 50 microL was established. Recombinant PTP1B was cloned and expressed in E. coli. with its specific substrate 4-nitrophenyl phosphate disodium salt hexahydrate (PNPP). The HTS model was based on enzyme reaction rate with enhanced sensitivity and specificity (Z' = 0.78). A total of 24,240 samples were screened, among them 80 samples with inhibition greater than 70% were selected for further rescreening. Finally, six compounds with high inhibitory activity were identified, whose IC50 values were 21.58, 18.39, 15.37, 11.92, 37.27, and 36.61 microg x mL(-1), separately. The results indicated that the method was stable, sensitive, reproducible and also suitable for high-throughput screening.
Subject(s)
Humans , Drug Evaluation, Preclinical , Methods , Enzyme Inhibitors , Pharmacology , Escherichia coli , Metabolism , High-Throughput Screening Assays , Methods , Inhibitory Concentration 50 , Protein Tyrosine Phosphatase, Non-Receptor Type 1 , Metabolism , Sensitivity and Specificity , Vanadates , PharmacologyABSTRACT
To screen potential human soluble epoxide hydrolase (hsEH) inhibitors, a high-throughput screening model in 384-well microplate with total volume of 50 microL was established. Recombinant hsEH was cloned and expressed in E. coli. and its specific substrate PHOME was synthesized. The HTS model was based on fluorescence analysis with enhanced sensitivity and specificity (Z' = 0.65). A total of 47 360 samples (including 25 040 compounds and 22 320 natural products) were screened, of which 950 samples with inhibition greater than 80% were selected for further rescreening. Finally, two compounds with high inhibitory activity were identified, whose IC50 value were 8.56 and 4.31 micromol x L(-1), separately. The results indicated that the method was stable, sensitive, reproducible and also suitable for high-throughput screening.