Effects of the combination of Epimedii Folium and Ligustri Lucidi Fructus on apoptosis and autophagy in SOP rats and osteoblasts via PI3K/AKT/mTOR pathway.

BACKGROUND: This study aimed to investigate the effects of the combination of Epimedii Folium (EF) and Ligustri Lucidi Fructus (LLF) on regulating apoptosis and autophagy in senile osteoporosis (SOP) rats. METHODS: Firstly, we identified the components in the decoction and drug-containing serum of EL (EF&LLF) by Ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS). Secondly, SOP rats were treated with EF, LLF, EL and caltrate to evaluate the advantages of EL. Finally, H2O2-, chloroquine-, and MHY1485-induced osteoblasts were treated with different doses of EL to reveal the molecular mechanism of EL. We detected bone microstructure, oxidative stress levels, ALP activity and the expressions of Bax, Bcl-2, caspase3, P53, Beclin-1, p-PI3K, PI3K, p-Akt, Akt, p-mTOR, mTOR, and LC3 in vivo and in vitro. RESULTS: 36 compounds in EL decoction and 23 in EL-containing serum were identified, including flavonoids, iridoid terpenoids, phenylethanoid glycosides, polyols and triterpenoids. EL could inhibit apoptosis activity and increase ALP activity. In SOP rats and chloroquine-inhibited osteoblasts, EL could improve bone tissue microstructure and osteoblasts functions by upregulating Bcl-2, Beclin1, and LC3-II/LC3-I, while downregulating p53 in all treatment groups. In H2O2-induced osteoblasts, EL could upregulate the protein and mRNA expressions of Bcl-2 while downregulate LC3-II/LC3-I, p53 and Beclin1. Besides, EL was able to down-regulate PI3K/AKT/mTOR pathway which activated in SOP rats and MHY1485-induced osteoblasts. CONCLUSIONS: These findings demonstrate that EL with bone protective effects on SOP rats by regulating autophagy and apoptosis via PI3K/Akt/mTOR signaling pathway, which might be an alternative medicine for the treatment of SOP.

Ultrasound-targeted microbubble destruction promotes PDGF-primed bone mesenchymal stem cell transplantation for myocardial protection in acute Myocardial Infarction in rats.

BACKGROUND: Ultrasound-targeted microbubble destruction (UTMD) has emerged as a promising strategy for the targeted delivery of bone marrow mesenchymal stem cells (MSCs) to the ischemic myocardium. However, the limited migration capacity and poor survival of MSCs remains a major therapeutic barrier. The present study was performed to investigate the synergistic effect of UTMD with platelet-derived growth factor BB (PDGF-BB) on the homing of MSCs for acute myocardial infarction (AMI). METHODS: MSCs from male donor rats were treated with PDGF-BB, and a novel microbubble formulation was prepared using a thin-film hydration method. In vivo, MSCs with or without PDGF-BB pretreatment were transplanted by UTMD after inducing AMI in experimental rats. The therapeutic efficacy of PDGF-BB-primed MSCs on myocardial apoptosis, angiogenesis, cardiac function and scar repair was estimated. The effects and molecular mechanisms of PDGF-BB on MSC migration and survival were explored in vitro. RESULTS: The results showed that the biological effects of UTMD increased the local levels of stromal-derived factor-1 (SDF-1), which promoted the migration of transplanted MSCs to the ischemic region. Compared with UTMD alone, UTMD combined with PDGF-BB pretreatment significantly increased the cardiac homing of MSCs, which subsequently reduced myocardial apoptosis, promoted neovascularization and tissue repair, and increased cardiac function 30 days after MI. The vitro results demonstrated that PDGF-BB enhanced MSC migration and protected these cells from H2O2-induced apoptosis. Mechanistically, PDGF-BB pretreatment promoted MSC migration and inhibited H2O2-induced MSC apoptosis via activation of the phosphatidylinositol 3-kinase/serine-threonine kinase (PI3K/Akt) pathway. Furthermore, crosstalk between PDGF-BB and stromal-derived factor-1/chemokine receptor 4 (SDF-1/CXCR4) is involved in the PI3K/AKT signaling pathway. CONCLUSION: The present study demonstrated that UTMD combined with PDGF-BB treatment could enhance the homing ability of MSCs, thus alleviating AMI in rats. Therefore, UTMD combined with PDGF-BB pretreatment may offer exciting therapeutic opportunities for strengthening MSC therapy in ischemic diseases.

Noninvasive left ventricular pressure-strain myocardial work in patients with well-functioning bicuspid aortic valves and aortic dilation: a preliminary study.

Background: Noninvasive left ventricular pressure-strain myocardial work (MW) is a novel method for evaluating left ventricular function that integrates myocardial deformation and afterload and has certain advantages over global longitudinal strain (GLS). The study aimed to analyze MW in patients with well-functioning bicuspid aortic valve (BAV) and explore the influences of aortic dilation and arterial stiffness on left ventricular function. Methods: A total of 104 patients with well-functioning BAVs and 50 controls were enrolled in our study. Global work index (GWI), global constructive work (GCW), global wasted work (GWW), global work efficiency (GWE), GLS, and aortic stiffness index were measured. Based on the ascending aortic diameter, patients with BAV were divided into 3 subgroups (nondilated, mildly dilated, and moderately dilated). Results: GWI, GCW, GWW, and aortic stiffness index were significantly increased (P<0.001, P=0.023, P<0.001, and P<0.001, respectively), while GWE and GLS were significantly decreased among patients with BAV compared with controls (all P values <0.001). Patients with BAV and mildly and moderately dilated aortas had an increased GWW and aortic stiffness index but a decreased GWE compared with patients with BAV and nondilated aortas (all P values <0.05); meanwhile, GCW and GLS did not differ among the BAV subgroups (all P values >0.05). GWI was elevated in patients with BAV and moderately dilated aortas compared with patients with BAV and nondilated aortas (P<0.05). On multivariable analysis, the aortic stiffness index was an independent influencer of GWI, GCW, GWW, and GWE (P=0.025, P=0.049, P<0.001, and P=0.001, respectively). The aortic diameter was highly correlated with the aortic stiffness index (r=0.863; P<0.001). Conclusions: MW could assess early myocardial impairment in patients with well-functioning BAV. MW may help to differentiate the detrimental effect of aortic dilation on left ventricular function, whereas GLS may not.

Combined Epimedii Folium and Ligustri Lucidi Fructus with dexamethasone alleviate the proliferation of airway smooth muscle cells by regulating apoptosis/autophagy.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine (TCM) theory believes kidney deficiency is the root cause of chronic refractory asthma with pathological changes of airway remodeling. Our previous experiments confirmed that the combination of Epimedii Folium and Ligustri Lucidi Fructus (ELL) with the effect of nourishing Yin and Yang of the kidney could improve the pathological changes of airway remodeling in asthmatic rats, but the specific mechanism remains unclear. AIM OF THE STUDY: This research was designed to reveal the synergy of ELL and dexamethasone (Dex) in the proliferation, apoptosis, and autophagy of airway smooth muscle cells (ASMCs). MATERIALS AND METHODS: Primary cultures of ASMCs from rats were prepared and induced with histamine (Hist), Z-DEVD-FMK (ZDF), rapamycin (Rap), or 3-Methyladenine (3-MA) at generation 3-7 for 24 or 48 h. Subsequently, the cells were treated with Dex, ELL, and ELL&Dex for 24 or 48 h. The effect of various concentrations of inducers and drugs on cell viability was detected by Methyl Thiazolyl Tetrazolium (MTT) assay, cell proliferation was tested using immunocytochemistry (ICC) by detecting Ki67 protein, cell apoptosis was measured by Annexin V-FITC/PI assay and Hoechst nuclear staining, cell ultrastructure was observed by transmission electron microscopy (TEM), and immunofluorescence (IF), Western blot (WB) combined with quantitative real-time PCR (qPCR) were used for measuring autophagy and apoptosis-related genes including protein 53 (P53), cysteinyl aspartate-specific proteinase (Caspase)-3, microtubule-associated protein 1 light chain 3 (LC3), Beclin-1, mammalian target of rapamycin (mTOR) and p-mTOR. RESULTS: In ASMCs, Hist and ZDF promoted cell proliferation, significantly decreased Caspase-3 protein expression, and up-regulated Beclin-1 levels; Dex alone and in combination with ELL promoted Beclin-1, Caspase-3, and P53 expression, enhancing autophagy activity and apoptosis in Hist and ZDF-induced AMSCs. In contrast, Rap inhibited cell viability, increased Caspase-3, P53, Beclin-1, and LC3-II/I and decreased the levels of mTOR and p-mTOR with promoting apoptosis and autophagy; ELL or ELL&Dex reduced P53, Beclin-1, and LC3-II/I to down-regulate apoptosis and the excessive autophagic state of ASMCs induced by Rap. In the 3-MA model, cell viability and autophagy were reduced; ELL&Dex significantly upgraded the expression of Beclin-1, P53, and Caspase-3 and promoted apoptosis and autophagy of ASMCs. CONCLUSIONS: These results suggest that ELL combined with Dex may regulate the proliferation of ASMCs by promoting apoptosis and autophagy and be a potential medicine for the treatment of asthma.

Dynamic changes in marker components during the stir-frying of Pharbitidis Semen, and network analysis of its potential effects on nephritis.

Introduction: Pharbitidis Semen (PS) has been widely used in traditional Chinese medicine to treat several diseases such as nephritis. PS is usually stir-fried to enhance its therapeutic efficacy before use in clinical practice. However, the changes in phenolic acids during stir-frying and the mechanisms of their therapeutic effects on nephritis are still unclear. Methods: Here, we studied the processing-induced chemical changes and elucidated the mechanism of PS in the treatment of nephritis. We determined the levels of the 7 phenolic acids in raw PS (RPS) and stir-fried PS (SPS) using high-performance liquid chromatography, analyzed the dynamic compositional changes during stir-frying, and used network analysis and molecular docking to predict and verify compound targets and pathways corresponding to nephritis. Results: The dynamic changes in the 7 phenolic acids in PS during stir-frying are suggestive of a transesterification reaction. Pathway analysis revealed that the targets of nephritis were mainly enriched in the AGE-RAGE, hypoxia-inducible factor-1, interleukin-17, and tumor necrosis factor signaling pathways among others. Molecular docking results showed that the 7 phenolic acids had good binding ability with the key nephritic targets. Discussion: The potential pharmaceutical basis, targets, and mechanisms of PS in treating nephritis were explored. Our findings provide a scientific basis for the clinical use of PS in treating nephritis.

Apoptotic tumor cell-derived microparticles loading Napabucasin inhibit CSCs and synergistic immune therapy.

BACKGROUND: Cancer stem cells (CSCs) are crucial for the growth, metastasis, drug resistance, recurrence, and spread of tumors. Napabucasin (NAP) could effectively inhibit CSC, but its mechanism has not been fully explained. Additionally, NAP also has the drawbacks of poor water solubility and low utilization. Therefore, this study not only elaborated the new mechanism of NAP inhibiting CSCs, but also built NAP-loaded nanoprobes using apoptotic tumor-derived microparticles (TMPs) as carriers to combine diagnose and treat of colon cancer and lessen the adverse effects of NAP. RESULTS: The study discovered a new mechanism for NAP inhibiting tumors. NAP, in addition to inhibiting STAT3, may also inhibit STAT1, thereby inhibiting the expression of CD44, and the stemness of colon cancer. N3-TMPs@NAP was successfully synthesized, and it possessed a lipid bilayer with a particle size of 220.13 ± 4.52 nm, as well as strong tumor binding ability and anti-tumor effect in vitro. In static PET/CT imaging studies, the tumor was clearly visible and showed higher uptake after N3-TMPs@NAP injection than after oral administration. The average tumor volume and weight of the N3-TMPs@NAP group on day 14 of the treatment studies were computed to be 270.55 ± 107.59 mm3 and 0.30 ± 0.12 g, respectively. These values were significantly lower than those of the other groups. Additionally, N3-TMPs@NAP might prevent colon cancer from spreading to the liver. Furthermore, due to TMPs' stimulation of innate immunity, N3-TMPs@NAP might stimulate anti-tumor. CONCLUSIONS: As a combined diagnostic and therapeutic nanoprobe, N3-TMPs@NAP could successfully conduct PET/CT imaging, suppress CSCs, and synergistically stimulate anticancer immune responses. Additionally, this nanoprobe might someday be employed in clinical situations because TMPs for it can be produced from human tissue and NAP has FDA approval.

A rare combination of Ebstein's anomaly with left ventricular outflow tract obstruction.

We describe a rare case of Ebstein's anomaly (EA) combined with left ventricular outflow tract obstruction in a 54-year-old man that was accurately identified by echocardiography, cardiac magnetic resonance imaging (CMR). The imaging result was ultimately validated by surgery. We emphasize the clinical importance of using echocardiography and CMR together to provide a thorough, noninvasive explanation of these results.

Epimedii Folium and Ligustri Lucidi Fructus Promote Osteoblastogenesis and Inhibit Osteoclastogenesis against Osteoporosis via Acting on Osteoblast-Osteoclast Communication.

Osteoblast (OB) and osteoclast (OC) play important roles in bone formation and bone resorption, which can communicate with each other through cytokine paracrine. Previous studies have confirmed that Epimedii Folium (EF) and Ligustri Lucidi Fructus (LLF) used alone or in combination can treat osteoporosis (OP) through regulating bone remodeling, but the effects of EF and LLF on osteoblastogenesis, osteoclastogenesis, and OB-OC communication are unclear. In this study, we investigated the direct and indirect effects of EF and LLF on OBs and OCs via monoculture and coculture (transwell) models of OBs and OCs. We found that the combination of EF and LLF (EF&LLF) could promote osteoblastogenesis and inhibit osteoclastogenesis directly and indirectly. In order to study the mechanisms of EF&LLF on indirectly regulating osteoblastogenesis and osteoclastogenesis, we detected the expression of cytokines by which OBs and OCs could communicate with each other. We found that EF&LLF could downregulate the expression of RANKL and M-CSF and the protein ratio of RANKL/OPG of OBs and Atp6v0d2 expression of OCs and upregulate the expression of OPG and TGF-ß1 of OBs and the expression of TGF-ß1, BMP-2, and IGF-1 of OCs, indicating that EF&LLF could regulate cytokine expressions of OBs/OCs to affect OB-OC communication. In addition, EF&LLF had a better effect on regulating cytokines of OBs and OCs than EF or LLF in single use. This study suggested that EF&LLF exhibited the effects of promoting osteoblastogenesis and inhibiting osteoclastogenesis via acting on OB-OC communication and provided some scientific evidences for EF&LLF against OP.

Engineering goat milk-derived extracellular vesicles for multiple bioimaging-guided and photothermal-enhanced therapy of colon cancer.

Multimodal image-guided photothermal therapy (PTT) has great application potential in cancer treatment due to its advantages of low side effects and good efficacy. There is an urgent need for PTT nanocarriers with high loading efficiency and modified surfaces. Goat milk-derived extracellular vesicles (GMVs) an ideal PTT nanoplatforms due to their anti-inflammatory ability, tumor retention ability, high yield, and high biosafety. This study used GMVs to design a theranostic nanoprobe for positron emission tomography/computer tomography/near-infrared fluorescence (PET/CT/NIRF) imaging and image-guided PTT for colon cancer. The key genes, important biological processes, and important signaling pathways of indocyanine green (ICG)-mediated PTT and N3-GMV@ICG-mediated PTT were analyzed. The nanoprobe triggered anti-tumor immune and inflammation responses to enhance PTT. In addition, the nanoprobe could attenuate PTT-induced inflammation benefiting from the anti-inflammatory efficacy of GMVs. Therefore, our findings conceptually advanced the diagnosis and treatment of colon cancer. We believed that the nanoprobe had broad clinical transformation prospects, and GMVs might be ideal nanocarriers for constructing integrated diagnostic and PTT probes.

Novel strategy for primary epithelial cell isolation: Combination of hyaluronidase and collagenase I.

OBJECTIVE: Different strategies for epithelial cell isolation significantly affect the viability and physiological properties of primary cells. Trypsin digestion, a conventional method, causes collateral damage owing to its strong digestive potential. To better preserve the physiological properties of epithelial tissues, we aimed to develop a modified method (hyaluronidase and collagenase I combination) for primary cell isolation. METHOD: We used conventional and modified methods to compare cell viability, morphology and stemness. Additionally, we investigated the passaging stability of epithelial cells and their capacity for organoid formation. Finally, we compared the two methods for isolating urothelial, oesophageal, lingual, and epidermal epithelial cells. RESULT: Gingival epithelial cells obtained using the modified method had higher viability, better morphology and stronger stemness than those obtained using the conventional method. Additionally, primary cells obtained using the modified method were stably passaged. Regarding organoid culture, adopting the modified method led to a significant increase in the growth rate and expression of the stem cell markers cytokeratin (CK)-19 and Ki-67. Furthermore, the modified method outperformed the conventional method for isolating urothelial, epidermal, oesophageal and lingual epithelial cells. CONCLUSION: We demonstrated that the combination of hyaluronidase and collagenase I outperformed trypsin in preserving the physiological properties of primary cells and organoid formation. The modified method could be broadly applied to isolate different types of epithelial cells and facilitate studies on organoids and tissue engineering.

Application of Speckle Tracking Echocardiography for Evaluating Ventricular Function after Transcatheter Pulmonary Valve Replacement.

Pulmonary regurgitation usually leads to right heart dilatation and eventually right heart dysfunction, which is associated with a poor prognosis. Transcatheter pulmonary valve replacement is a developing treatment for pulmonary valve dysfunction that can take the place of traditional surgery and make up for the shortcomings of a large injury. Echocardiography plays a significant role in assessing ventricular function; however, conventional echocardiographic parameters have several limitations. Speckle tracking echocardiography has been regarded as a more accurate tool for quantifying cardiac function than conventional echocardiography. Therefore, the aim of this review was to summarize the application of speckle tracking echocardiography for evaluating right and left ventricular functions in patients after transcatheter pulmonary valve replacement.