Salvianolic acid C attenuates cerebral ischemic injury through inhibiting neuroinflammation via the TLR4-TREM1-NF-κB pathway.

BACKGROUND: Stroke is a leading cause of mortality and disability with ischemic stroke being the most common type of stroke. Salvianolic acid C (SalC), a polyphenolic compound found in Salviae Miltiorrhizae Radix et Rhizoma, has demonstrated therapeutic potential in the recovery phase of ischemic stroke. However, its pharmacological effects and underlying mechanisms during the early stages of ischemic stroke remain unclear. This study aimed to examine the potential mechanism of action of SalC during the early phase of ischemic stroke using network pharmacology strategies and RNA sequencing analysis. METHODS: SalC effects on infarct volume, neurological deficits, and histopathological changes were assessed in a mouse model of transient middle cerebral artery occlusion (tMCAO). By integrating RNA sequencing data with a cerebral vascular disease (CVD)-related gene database, a cerebral ischemic disease (CID) network containing dysregulated genes from the tMCAO model was constructed. Network analysis algorithms were applied to evaluate the key nodes within the CID network. In vivo and in vitro validation of crucial targets within the identified pathways was conducted. RESULTS: SalC treatment significantly reduced infarct volume, improved neurological deficits, and reversed pathological changes in the tMCAO mouse model. The integration of RNA sequencing data revealed an 80% gene reversion rate induced by SalC within the CID network. Among the reverted genes, 53.1% exhibited reversion rates exceeding 50%, emphasizing the comprehensive rebalancing effect of SalC within the CID network. Neuroinflammatory-related pathways regulated by SalC, including the toll-like-receptor 4 (TLR4)- triggering receptor expressed on myeloid cells 1 (TREM1)-nuclear factor kappa B (NF-κB) pathway, were identified. Further in vivo and in vitro experiments confirmed that TLR4-TREM1-NF-κB pathway was down-regulated by SalC in microglia, which was essential for its anti-inflammatory effect on ischemic stroke. CONCLUSIONS: SalC attenuated cerebral ischemic injury by inhibiting neuroinflammation mediated by microglia, primarily through the TLR4-TREM1-NF-κB pathway. These findings provide valuable insights into the potential therapeutic benefits of SalC in ischemic stroke.

Identification of bioactive anti-angiogenic components targeting tumor endothelial cells in Shenmai injection using multidimensional pharmacokinetics.

Shenmai injection (SMI) is a well-defined herbal preparation that is widely and clinically used as an adjuvant therapy for cancer. Previously, we found that SMI synergistically enhanced the activity of chemotherapy on colorectal cancer by promoting the distribution of drugs in xenograft tumors. However, the underlying mechanisms and bioactive constituents remained unknown. In the present work, the regulatory effects of SMI on tumor vasculature were determined, and the potential anti-angiogenic components targeting tumor endothelial cells (TECs) were identified. Multidimensional pharmacokinetic profiles of ginsenosides in plasma, subcutaneous tumors, and TECs were investigated. The results showed that the concentrations of protopanaxadiol-type (PPD) ginsenosides (Rb1, Rb2/Rb3, Rc, and Rd) in both plasma and tumors, were higher than those of protopanaxatriol-type (Rg1 and Re) and oleanane-type (Ro) ginsenosides. Among PPD-type ginsenosides, Rd exhibited the greatest concentrations in tumors and TECs after repeated injection. In vivo bioactivity results showed that Rd suppressed neovascularization in tumors, normalized the structure of tumor vessels, and improved the anti-tumor effect of 5-fluorouracil (5FU) in xenograft mice. Furthermore, Rd inhibited the migration and tube formation capacity of endothelial cells in vitro. In conclusion, Rd may be an important active form to exert the anti-angiogenic effect on tumor after SMI treatment.

Protective effects of Danshen injection against erectile dysfunction via suppression of endoplasmic reticulum stress activation in a streptozotocin-induced diabetic rat model.

BACKGROUND: Erectile dysfunction (ED) is a common complication of diabetes. This study aimed to explore the beneficial effect of Danshen injection on ED in a streptozotocin (STZ)-induced diabetic rat model and the underlying mechanism. METHODS: The diabetic rat model was established by an intraperitoneal injection of 60 mg/kg STZ in male Sprague-Dawley rats. The diabetic rats were intraperitoneally injected with Danshen solution (0.5 or 1 mL/kg/day) or the same volume of saline for 6 weeks. Age-matched rats served as controls. After 6 weeks, erectile function and histological morphology of the corpora cavernosum were assessed. Oxidative stress indicators, including superoxide dismutase (SOD) activity, malondialdehyde (MDA) content, and reactive oxygen species (ROS) levels, were measured in penile tissues. The expression levels of glucose-regulated protein 78 (Grp78), growth arrest and DNA damage-inducible gene 153 (GADD153/CHOP) were determined by immunohistochemistry, immunoblotting, and RT-PCR. Apoptosis was detected by a TUNEL assay. RESULTS: The erection times of diabetic rats were significantly less than those of control rats. Danshen injection could improve erectile function via increased erection times. Danshen injection was also found to ameliorate the morphological abnormalities of the corpora cavernosum, to reduce the number of apoptotic cells, and to suppress caspase-3 activation in penile tissue, accompanied by downregulation of the endoplasmic reticulum stress biomarkers Grp78 and CHOP. Danshen injection could increase SOD activity as well as reduce ROS and MDA levels in diabetic rats, indicating suppression of oxidative stress. CONCLUSION: Danshen injection could rescue diabetes-associated ED, possibly via suppressing the oxidative stress and endoplasmic reticulum (ER) stress-induced apoptosis pathways.

iTRAQ-Based Proteomic Analysis Reveals Recovery of Impaired Mitochondrial Function in Ischemic Myocardium by Shenmai Formula.

Shenmai formula (SM) has been a traditional medicinal remedy for treating cardiovascular diseases in China for 800 years; however, its mechanism of action remains unclear. To explore the mechanism underlying cardioprotective effects of SM, iTRAQ-based proteomic approach was applied to analyze protein of myocardium in rats with myocardial ischemic injury. Upon treatment with SM and its two major components Red ginseng (RG) and Radix Ophiopogonis (OP), 101 differentially expressed proteins were filtered from a total of 712 detected and annotated proteins. They can be classified according to their locations and functions, while most of them are located in intracellular organelle, participating in cellular metabolic process. The functions of them are mostly associated with mitochondrial oxidative phosphorylation/respiration. The differentially expressed proteins were validated by liquid chromatography-tandem mass spectrometry and Western blotting (ATP5D, NDUFB10, TNNC1). Further in vitro experiments found that SM could attenuate hypoxia induced impairment of mitochondrial membrane potential and cellular ATP concentration in neonatal rat ventricular myocytes. Interestingly, the result of quantitative mitochondrial biogenesis assays revealed that SM had dominant positive effects on the maximum respiration, ATP-coupled respiration, and spare capacity of mitochondria in response to hypoxia. Hence, our findings suggest that SM promotes mitochondrial function to protect cardiomyocytes against hypoxia, which provides a possible illustration for conventional botanical therapy on a molecular level.

Comparison of bioactive components and pharmacological activities of ophiopogon japonicas extracts from different geographical origins.

Ophiopogon japonicus (Linn. f.) Ker-Gawl (O. japonicas), mainly cultivated in Sichuan and Zhejiang province in China, has different bioactive components and therefore their pharmacological activities. To explain the different clinical efficacy of O. japonicas derived preparations, herein we report differences of pharmacological activities between Sichuan and Zhejiang O. japonicas and behind them the exact differences of bioactive components. Based on a LC/MS-IT-TOF method, the differences of bioactive components between Sichuan and Zhejiang O. japonicas extracts were analyzed and respective characteristic components were picked out. We determined 39 ophiopogonones and 71 ophiopogonins compounds in Sichuan and Zhejiang O. japonicas extracts and found the contents of these compositions have several times difference. Evidenced by experimental data of pharmacological activities in inhibiting cardiomyocyte damage induced by H2O2, mouse macrophage cell inflammation induced by lipopolysaccharide and cytotoxicity in vitro, Zhejiang O. japonicas extract had a stronger antioxidant and anti-inflammatory capacity than Sichuan O. japonicas extract, and the two O. japonicas extracts exhibited selective cytotoxicity on different cancer cell lines in vitro. These data shed light on the links between bioactive components and pharmacological activities of O. japonicas derived preparations. Thus, geographical origin of O. japonicas should be considered to be a key factor in efficacy studies and further clinical application.

Shenmai injection enhances the cytotoxicity of chemotherapeutic drugs against colorectal cancers via improving their subcellular distribution.

Shenmai injection (SMI) is a Chinese patent-protected injection, which was mainly made of Red Ginseng and Radix Ophiopogonis and widely used for treating coronary heart disease and tumors by boosting Qi and nourishing Yin. In this study we examined whether SMI could produce direct synergetic effects on the cytoxicity of adriamycin (ADR) and paclitaxel (PTX) in colorectal cancers in vivo and in vitro, and explored the underlying pharmacokinetic mechanisms. BALB/c nude mice with LoVo colon cancer xenografts were intraperitoneally injected with ADR (2 mg·kg-1·3d-1) or PTX (7.5 mg·kg-1·3d-1) with or without SMI (0.01 mL·g-1·d-1) for 13 d. Co-administration of SMI significantly enhanced the chemotherapeutic efficacy of ADR and PTX, whereas administration of SMI alone at the given dosage did not produce visible anti-cancer effects, The chemosensitizing action of SMI was associated with increased concentrations of ADR and PTX in the plasma and tumors. In Caco-2 and LoVo cells in vitro, co-treatment with SMI (2 µL/mL) significantly enhanced the cytotoxicity of ADR and PTX, and resulted in some favorable pharmacokinetic changes in the subcellular distribution of ADR and PTX. In addition, SMI-induced intracellular accumulation of ADR was closely correlated with the increased expression levels of P-glycoprotein in 4 colon cancer cell lines (r2=+0.8558). SMI enhances the anti-cancer effects of ADR and PTX in colon cancers in vivo and in vitro by improving the subcellular distributions of ADR and PTX.

Danshen injection ameliorates STZ-induced diabetic nephropathy in association with suppression of oxidative stress, pro-inflammatory factors and fibrosis.

Diabetic nephropathy (DN) is one of the most frequent complications in diabetes mellitus. This study aimed to explore whether Danshen injection is protective to renal tissue in diabetes. Intraperitoneal injection of streptozotocin (STZ) (60mg/kg) was used to induce diabetes in rats. Some STZ-induced diabetic rats were also intraperitoneally injected with Danshen solution at two different dosages (0.5 or 1ml/kg/day) for 6weeks. Our results showed that serum creatinine (sCr) and blood urea nitrogen were significantly increased in STZ-induced diabetic rats, which was alleviated upon Danshen injection. Danshen injection was also found to ameliorate hypertrophy and dilatation of renal tubule and glomeruli possibly by decreasing the expression of collagen and fibronectin in association with suppression of TGF-ß1/Smad pathway. Further investigation revealed that Danshen injection could increase the activity of superoxide dismutase (SOD), and reduce reactive oxygen species (ROS) and malondialdehyde (MDA) levels in STZ-induced diabetic rats, indicating suppression of oxidative stress. In addition, we also found that Danshen injection could suppress IκB/NF-κB signaling pathway and reduce the level of a number of pro-inflammatory factors, including tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß) and interleukin-6 (IL-6) in the diabetic renal tissue, indicating suppression of inflammation. In conclusion, our results demonstrated that Danshen injection may rescue STZ-induced diabetic nephropathy, possibly via suppressing the oxidative stress, inflammatory responses and fibrosis progression.

Dihydroartemisinin inhibits the Bcr/Abl oncogene at the mRNA level in chronic myeloid leukemia sensitive or resistant to imatinib.

Due to the mutations of the Bcr/Abl oncogene that obstacle the binding of the protein with imatinib, the resistance to imatinib has developed in a significant portion of chronic myeloid leukemia (CML) patients. It stimulated the search for novel molecules for treatment of imatinib-resistance CML. Inhibiting the amplification of Bcr/Abl oncogene is believed to be a new effective strategy to override the imatinib resistance on CML cells. In present research, we demonstrated that dihydroartemisinin (DHA), a safe and effective antimalarial analog of artemisinin, could significantly inhibit the Bcr/Abl fusion gene at the mRNA level in CML cells sensitive or resistant to imatinib (including the primary CML cells with T315I mutation) and induce cell death. Moreover, dihydroartemisinin could also lead to the inhibition of the Bcr/Abl protein expression and tyrosine kinase activity, and strongly suppress on the downstream signals of Bcr/Abl, which included inhibition of tyrosine kinase activity of AKT and ERK, promotion of cytochrome c release from the mitochondria and the consequential activation of caspase-9/3 in imatinib-resistant CML cells. These results suggest for the first time that Dihydroartemisinin might be a potential novel drug candidate for treatment of imatinib-resistant CML and worthy of further study.

[Chemical constituents from Rehmannia glutinosa].

OBJECTIVE: To study the chemical constituents from the roots of Rehmannia glutinosa. METHOD: The compounds were isolated by various chromatographic methods and identified by spectroscopic analysis. RESULT: Twelve compounds were isolated and their structures were identified as 5-hydroxymethyl-pyrrole-2-carbaldehyde (1), 5-hydroxymethyl furfural (2), tyrosol (3), 5,6-dihydroxy-beta-ionone (4), 6-O-E-feruloyl ajugol (5), acteoside (6), leucosceptoside A (7), martynoside (8), isomartynoside (9), purpureaside C (10), jionoside A1 (11), and jionoside B1 (12). CONCLUSION: Compounds 1, 3 and 9 were isolated from the genus Rehmannia for the first time.

[Relationship of angiopoietins expression with microvessel density (MVD) in human colorectal tumors].

OBJECTIVE: To investigate the expression of angiopoietin (Ang)-1 and Ang-2 in colorectal tumors and its relations to microvessel density (MVD) in tumor tissue. METHODS: Ang-1, Ang-2 and factor VIII-related antigen were stained immunohistochemically in 91 cases of primary colorectal adenocarcinoma, 20 cases of colorectal adenoma and 24 cases of normal colorectal mucosal tissue, and MVD was also assayed in above tissue specimens. RESULT: (1) A significantly higher Ang-1 (7.07+/-2.00) was observed in normal tissue compared with 1.75 +/-1.98 in the adenoma and 1.40 +/- 1.22 in the adenocarcinoma (P<0.01). (2) Ang-2 protein positive rate in adenocarcinoma was significantly higher than that in normal tissue and adenoma (P<0.01). The expression of Ang-2 in adenocarcinoma was closely associated with poor differentiation and vessel invasion. (3) There were significant correlations between Ang-1 and Ang-2 (r=-0.338, P<0.01), Ang-1 and MVD (r=-0.388, P<0.01), Ang-2 and MVD (r=0.594, P<0.01) in the 135 cases. CONCLUSION: The overexpression of Ang-2 may play an important role in angiogenesis of colorectal adenocarcinoma. It can be regarded as an index for malignancy and prognosis in colorectal adenocarcinoma.