Anti-tumor Effect of Draconis Sanguis Petroleum Ether Fraction on Human Gastric Cancer HGC-27 and MGC-803 Cells / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Draconis Sanguis petroleum ether fraction （DSPEF） on the proliferation， apoptosis， migration， and autophagy of human gastric cancer HGC-27 and MGC-803 cells， and preliminarily elucidate its molecular mechanism. MethodCell counting kit-8 （CCK-8） assay was used to detect the effect of DSPEF at different concentrations （0， 20， 40， 60， 80 mg·L-1） on the proliferation of HGC-27 and MGC-803 cells after 24， 48， 72 h. Hoechst staining and flow cytometry were used to explore the effects of DSPEF at different concentrations on the apoptosis and apoptosis rate of HGC-27 and MGC-803 cells after 48 h treatment， respectively. The wound healing assay and acridine orange staining were used to investigate the effects of DSPEF on the migration and autophagy of HGC-27 and MGC-803 cells， respectively. Western blot was used to detect the expression levels of signaling pathway-related proteins in HGC-27 and MGC-803 cells treated with DSPEF for 48 h. ResultCompared with the control group， DSPEF（30 mg·L-1） inhibited the proliferation and migration of HGC-27 and MGC-803 cells in a concentration- and time-dependent manner （P<0.05）， and induced the apoptosis （P<0.01） and autophagy of HGC-27 and MGC-803 cells. DSPEF （60 mg·L-1） down-regulated the protein levels of phosphorylated mammalian target of rapamycin （p-mTOR） （P<0.05， P<0.01） and down-regulated phospho-signal transducer and activator of transcription 3 （p-STAT3） in HGC-27 and MGC-803 cells （P<0.01）， suggesting that DSPEF presumedly inhibited the proliferation and migration of human gastric cancer HGC-27 and MGC-803 cells and induced their apoptosis and autophagy by inhibiting the mTOR/STAT3 signaling pathway. ConclusionThe down-regulation of the mTOR/STAT3 signaling pathway may be involved in the anti-gastric cancer effect of DSPEF. This study is expected to provide a reference for the investigation of the anti-tumor effect of Draconis Sanguis.

Effect of Xuebijing injection on endothelial microparticles and renal cortical microcirculation in septic rats / 中华危重病急救医学

Objective:To clarify the characteristics of renal cortical microcirculation and its relationship with the expression of plasma endothelial microparticle (EMP) in septic rats, and to evaluate the effect of Xuebijing injection as an adjuvant therapy of antibiotics on septic AKI.Methods:The 8-10 weeks old specific pathogen free (SPF) male Sprague-Dawley (SD) rats were divided into sham operation group (Sham group), positive drug control group and Xuebijing group by the random number table method, with 10 rats in each group. The cecal ligation and puncture (CLP) with large ligation (ligated 75% of the cecum) was used to prepare a rat high-grade sepsis model; in the Sham group, the cecum was stretched without ligation or puncture. Due to the high mortality of CLP with large ligation, Xuebijing injection (4 mL/kg, 12 hours per time) and imipenem/cilastatin injection (90 mg/kg, 6 hours per time) were administered to the rats in the Xuebijing group via the tail vein immediately after the model was produced. Normal saline and imipenem/cilastatin were administered to the rats by the same methods in the positive drug control group. The rats in the Sham group were treated with the same volume of normal saline as any of the other two groups at the same frequency. At 48 hours after model reproduction, the mean arterial pressure (MAP) and blood lactic acid (Lac) of the rats were measured. The renal cortical microcirculation was monitored by using side stream dark-field imaging. Renal hypoxia signals were assessed by pimonidazole chloride immunohistochemistry. Plasma EMP levels were determined by using flow cytometry, and then the correlation between EMP and microcirculation parameters of renal cortex was analyzed. At the same time, the serum creatinine (SCr) was measured, and the renal injury score (Paller score) was used to evaluate the severity of renal tissue pathological damage.Results:Compared with the Sham group, perfused vessel density (PVD), microvascular flow index (MFI) and MAP in the positive drug control group and the Xuebijing group decreased significantly, the positive expression of hypoxia probe (pimonidazole) increased, Lac, EMP, Paller score and SCr increased significantly. However, compared with the positive drug control group, the renal cortical microcirculation in the Xuebijing group was improved significantly, PVD and MFI were increased significantly [PVD (mm/mm 2): 16.20±1.20 vs. 9.77±1.12, MFI: 2.46±0.05 vs. 1.85±0.15, both P < 0.05], Lac was reduced significantly (mmol/L: 4.81±1.23 vs. 6.08±1.09, P < 0.05), MAP increased slightly [mmHg (1 mmHg = 0.133 kPa): 84.00±2.00 vs. 80.00±2.00, P > 0.05], suggested that Xuebijing injection improved renal microcirculation perfusion in septic rats, and this effect did not depend on the change of MAP. The positive expression of pemonidazole in renal cortex of the Xuebijing group was significantly lower than that of the positive drug control group [(35.89±1.13)% vs. (44.93±1.37) %, P < 0.05], suggested that Xuebijing injection alleviated renal hypoxia. The plasma EMP levels of rats in the Xuebijing group were significantly lower than those in the positive drug control group (×10 6/L: 3.49±0.17 vs. 5.78±0.22, P < 0.05), and the EMP levels were significantly negatively correlated with PVD and MFI ( r values were -0.94 and -0.95, respectively, both P < 0.05), indicated that the increase of plasma EMP was highly correlated with renal microcirculation disorder, and Xuebijing injection inhibited the increase of plasma EMP levels. The Paller score in the Xuebijing group was significantly lower than that in the positive drug control group (46.90±3.84 vs. 62.70±3.05, P < 0.05), and the level of SCr was also significantly lower than that in the positive drug control group (μmol/L: 121.1±12.4 vs. 192.7±23.9, P < 0.05), which suggested that Xuebijing injection relieved kidney injury and improved renal function in septic rats. Conclusion:As an adjuvant therapy of antibiotics, Xuebijing injection could inhibit the expression of plasma EMP in rats with sepsis, improve renal cortex microcirculation, and reduce kidney injury.

Preliminary audiological evaluation of the SoundBite bone conduction devices in adults with single-sided deafness / 中华耳鼻咽喉头颈外科杂志

Objective: The auditory deficits of single-sided deafness (SSD) can be treated with a novel intra-oral device, SoundBite, which delivers sound by applying vibratory signal to the teeth. The purpose of this study was to evaluate the efficacy and benefit of the bone conduction device for Chinese adults with SSD. Methods: Eighteen patients aged 19-66 yrs with acquired, permanent sensorineural SSD and no current treatment by any other devices for SSD, were recruited in a prospective controlled, nonrandomized, unblinded study. They were requested the continually daily wear of the new device over a 30-day free trial period. The intra-oral hearing device was placed around two maxillary teeth and was similar to a small partial denture or retainer. The audiological tests included pure tone air conduction thresholds, monosyllable word recognition score (WRS) in quiet and sentence reception thresholds in noise (via CMNmatrix test). The benefit was determined with the Abbreviated Profile of Hearing Aid Benefit (APHAB) and the Speech, Spatial and Qualities of Hearing Scale (SSQ) questionnaire. Results: The monosyllable WRS and the 50% threshold of signal-to-noise ratio (SNR50) were significantly better in all aided conditions. The head shadow effect, assessed by the SNR50 via CMNmatrix test improved an average of 2.6 dB after 30 days' wearing compared with unaided condition (P<0.001). The APHAB scores improved (P<0.05) for all subjects for the Global and Ease of Communication, Reverberation, Background Noise subscales. The SSQ scores improved (P<0.05) for all subjects for Speech, Spatial and Qualities of Hearing subscales. Conclusion: The SoundBite is a good alternative to the well-established implantable bone conduction devices in patients with SSD. An improvement in listening ability in noise and quiet as well as a decrease of the head shadow effect is validated as the expected.

LncRNA AFAP1-AS1/miR-27b-3p/VEGF-C axis modulates stemness characteristics in cervical cancer cells / 中华医学杂志(英文版)

BACKGROUND@#Long non-coding RNA (lncRNA) actin filament-associated protein 1 antisense RNA 1 (AFAP1-AS1) functions as a competing endogenous RNA to regulate target genes expression by sponging microRNAs (miRs) to play cancer-promoting roles in cancer stem cells. However, the regulatory mechanism of AFAP1-AS1 in cervical cancer (CC) stem cells is unknown. The present study aimed to provide a new therapeutic target for the clinical treatment of CC.@*METHODS@#Hyaluronic acid receptor cluster of differentiation 44 variant exon 6 (CD44v6)(+) CC cells were isolated by flow cytometry (FCM). Small interfering RNAs of AFAP1-AS1 (siAFAP1-AS1) were transfected into the (CD44v6)(+) cells. The levels of AFAP1-AS1 were measured by quantitative real-time PCR (qRT-PCR). Sphere formation assay, cell cycle analysis, and Western blotting were used to detect the effect of siAFAP1-AS1. RNA pull-down and luciferase reporter assay were used to verify the relationship between miR-27b-3p and AFAP1-AS1 or vascular endothelial growth factor (VEGF)-C.@*RESULTS@#CD44v6(+) CC cells had remarkable stemness and a high level of AFAP1-AS1. However, AFAP1-AS1 knockdown with siAFAP1-AS1 suppressed the cell cycle transition of G(1)/S phase and inhibited self-renewal of CD44v6(+) CC cells, the levels of the stemness markers octamer-binding transcription factor 4 (OCT4), osteopontin (OPN), and cluster of differentiation 133 (CD133), and the epithelial-mesenchymal transition (EMT)-related proteins Twist1, matrix metalloprotease (MMP)-9, and VEGF-C. In the mechanism study, miR-27b-3p/VEGF-C signaling was demonstrated to be a key downstream of AFAP1-AS1 in the CD44v6(+) CC cells.@*CONCLUSIONS@#LncRNA AFAP1-AS1 knockdown inhibits the CC cell stemness by upregulating miR-27b-3p to suppress VEGF-C.

Hypoglycemic effects of Tibetan medicine Huidouba in STZ-induced diabetic mice and db/db mice / 中草药·英文版

Objective: Huidouba (HDB) is a Chinese folk medicine used to treat diabetes in Sichuan Province, China. Therefore, we investigated the anti-diabetic effects of HDB and its underlying mechanisms. We hypothesized that HDB treatment could enhance glucose tolerance and insulin sensitivity, and thus prevent a hyperglycemia state. Methods: To test the hypothesis, streptozotocin (STZ)-induced diabetic mice and db/db mice, widely used models of hyperglycemia and insulin-resistant diabetes, were either treated with HDB, metformin, or acarbose. Blood glucose, oral glucose tolerance test, insulin tolerance test, pancreatic histopathology and serum biochemistry were detected to assess the hypoglycemic effect of HDB. Results: HDB treatments were found to show the effect in reducing glucose levels. HDB also resulted in a significant reduction in body weight and food intake in the STZ-induced diabetic mouse model. Furthermore, it significantly improved glucose and insulin tolerance in the two diabetic mouse models. Importantly, insulin, glucagon, pancreatic polypeptide, and somatostatin immunohistochemistry revealed that HDB treatment improved the function and the location of the cells in the islets compared with the other two treatments. HDB treatment resulted in significant restoration of islet function. Our results illustrated the underlying mechanism of HDB in the progression of diabetes, and HDB can be an effective agent for the treatment of diabetes. Conclusion: The results of this study suggested that HDB can reduce blood glucose levels in STZ-induced hyperglycemic mice and db/db mice.

Huidouba Ameliorates Kidney Oxidative Stress Injury by Down-regulating Nox4 Expression in Rats with Diabetic Nephropathy / 中国实验方剂学杂志

Objective:From a new perspective，to explore therapeutic effect of Huidouba (HDB) on alleviating kidney oxidative damage in rats with diabetic nephropathy (DN) and provide a scientific basis for developing HDB as a potential Tibetan medicine for treatment of DN. Method:Rats were fed with high-fat diet (HFD) and injected with streptozocin (STZ, 65 mg·kg-1) intraperitoneally to induce DN model, while rats in Blank group were injected with an equal volume of vehicle and fed with normal chow. The successfully modeling DN rats were randomly divided into three groups, 8 rats per group, DN model group (10 mL·kg-1·d-1), Metformin group (0.045 g·kg-1·d-1) and HDB group (0.18 g·kg-1·d-1). Monitor body weight (BW) and fasting blood glucose (FBG) weekly, and collect 24 hours urine before and after medication to examine microalbuminuria (mAlb). Calculate kidney index (KI) after sacrificing, analyze mAlb, serum creatinine (SCr) and blood urea nitrogen (BUN) with a fully automatic biochemical analyzer. Histopathology of kidney was observed by Masson staining. Lipid peroxidation malondialdehyde (MDA) assay kit was used to examine MDA content in kidney tissue. Nox4, as a subtype of triphosphopyridine nucleotide (NADPH) oxidase family was determined by Western blot and immunofluorescence assay of kidney tissue. Result:Compared with blank group, levels of FBG, 24 h mAlb, SCr, BUN and MDA in DN model group were increased (P<0.01), tissue damage was obvious and Nox4 expression in glumeruli was increased significantly (P<0.01). Compared with DN model group, levels of FBG, 24 h mAlb, SCr, BUN and MDA in drug administration groups were decreased (P<0.01), kidney injury was alleviated and Nox4 expression was down-regulated（P<0.01）. Conclusion:HDB as a Yiqiyangyin Tibetan medicine, could ease oxidative stress injury of kidney and reduce proteinuria in DN rats, thus prevent the development of DN. Its mechanism is closely related to down-regulating Nox4 expression of kidney tissue in DN rats.

Hepatoprotective Effect of Portulacae Herba on Carbon Tetrachloride Induced Acute Liver Injury in Mice / 中国实验方剂学杂志

Objective::To explore the effect and mechanism of Portulacae Herba protecting carbon tetrachloride (CCl4)-induced acute liver injury. Method::Sixty Kunming mice were randomly divided into normal group, model group, silybin group (200 mg·kg-1) and Portulacae Herba high, medium, low (2, 1, 0.5 g·kg-1) dose groups. After continuous intragastric administration for 5 days, mice in each group were intraperitoneally injected with 0.2% CCl4 peanut oil solution to establish acute liver injury model, except normal mice. After 23 hours of modeling, serum and liver tissue were collected. Fully automatic analysis of serum serum liver function indicators in mice. Liver tissues were taken for hematoxylin-eosin staining (HE) staining to observe liver pathological changes. RNA Sequencing (RNA-seq) was used to analyze differential genes and functional enrichment, real-time fluorescence quantification PCR(Real-time PCR) was used to verify the mRNA expression of cytochrome P450 family members(CYP)26A1, CYP2C37, CYP2C44, CYP2C50, CYP2C54. Result::Compared with normal group, the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), total bilirubin (TBIL), malondialdehyde (MDA) in model group were significantly increased (P<0.05), and the activities of triglyceride (TG) and superoxide dismutase (SOD) were significantly decreased (P<0.05). Compared with model group, Portulaca Herba significantly reduced ALT, AST, TBIL and MDA levels in mice with acute liver injury (P<0.05), significantly increased SOD activity (P<0.01), and decreased the degree of liver tissue damage in mice. Compared with normal group, the mRNA expressions of CYP2C44, CYP2C50 in mice with acute liver injury were significantly decreased (P<0.05). Compared with model group, the mRNA expressions of CYP26A1, CYP2C37, CYP2C44, CYP2C50 and CYP2C54 were significantly increased in all dose groups of Portulaca Herba (P<0.05, P<0.01). Conclusion::Portulacae Herba has significant protective effects on acute liver injury caused by CCl4, and its mechanism may be related to the regulation of cytochrome P450 related genes.

Screening combination ratio and exploring mechanism of Momordicae Semen and Epimedii Folium / 中国中药杂志

The aim of this paper was to obtain low toxicity and high efficiency anti-tumor Chinese medicine through screening the combination ratios of Momordicae Semen and Epimedii Folium, and to explore the anti-tumor mechanism of the combination of two drugs by observing their effect on apoptosis-related proteins in cancer cells. Methyl thiazolyl tetrazolium(MTT) assay was used to observe the effect of drug combination on the proliferation of tumor cells from different tissue sources. The effects of the combination of the two drugs on tumor cells were analyzed by Compusyn software. Plate cloning assay was used to observe the effect of combination of these two drugs on the proliferation of A549 cells in vitro. The expression of reactive oxygen species(ROS) and apoptotic proteins p53, Bcl-2 and Bax were compared by using ROS kit and Western blot. Lewis lung cancer model was used to observe the anti-tumor effect of drugs in vivo. The results showed that the anti-tumor effect of their ethanol extract was more significant than that of water extract, and the anti-proliferation effect was strongest when the ratio was 1∶1(P<0.05). Compusyn analysis showed that the combination of the two drugs had synergistic effect. Further studies showed that after combined use, the number of clonogen formation in A549 cells was significantly reduced(P<0.01); ROS production was increased; the expression of apoptosis-related protein p53 was up-regulated, and the ratio of Bcl-2/Bax was decreased. In vivo animal study showed that the tumor inhibition rate was 53.06%(P<0.05) in the high dose group. As compared with the single use of the two drugs, the combination of the two drugs had more significant anti-proliferative effect on tumors, and the optimum ratio was 1∶1. The combination of the two drugs at a ratio of 1∶1 inhibited the proliferation of various tumor cells, and had no significant effect on normal liver cells LO2 when compared with other ratios. Therefore, it can be preliminarily inferred that the combination of the two drugs may have the effect of synergism and detoxification. Further studies showed that the combination of the two drugs can significantly inhibit the proliferation of A549 cells, and its mechanism may be related to the activation of endogenous apoptotic pathway. In vivo experiments also showed that the tumor inhibition rate increased with the increase of drug concentration.

LC-MS guided isolation of two anti-inflammatory cyclic dihydrochalcane trimers from Chinese dragon's blood / 中国中药杂志

As an important integral part of traditional Chinese medicine chemical biology( TCMCB),it is of great importance to rapid isolate,and reliably identify the chemical components in herbal medicines. Phytochemical studies on the anti-inflammatory active part of Chinese dragon's blood,the red resin of Dracaena cochinchinensis,resulted in the isolation of two compounds,nordracophane( 1) and dracophane( 2),using LC-MS and chromatographic techniques( Silica gel,ODS and preparative HPLC). The structures,cyclic dihydrochalcane trimers,were elucidated on the basis of 1 D and 2 D NMR,MS,IR and UV spectral analysis. Compound 1 is a new compound,and 2 is isolated from D. cochinchinensis for the first time. Both compounds exhibited significant inhibition of nitric oxide production in lipopolysaccharides( LPS)-stimulated RAW264. 7 cells with IC50 values of( 14. 9±4. 50) and( 9. 0±0. 7) μmol·L-1.

Identification of active components in Longxue Tongluo Capsules against ischemic brain injury based on component-activity relationship / 中国中药杂志

Ten fractions(A-J) were prepared by separation of Longxue Tongluo Capsules(LTC) by using silica gel column chromatography and orthogonal experimental design,showing similar chemical profiles with different abundances of peaks.These ten samples were assessed with UHPLC-QE OrbitrapHRMS for 97 common peaks.For the pharmacological activity experiment,three kinds of in vitro cell models including lipopolysaccharide(LPS)-induced BV-2 microglial cells NO release model,oxygen-glucose deprivation/reoxygenation(OGD/R)-treated HUVEC vascular endothelial cells injury model,and OGD/R-treated PC-12 nerve cells injury model were employed to evaluated the bioactivity of each fraction.Based on the contribution of each identified component,grey relation analysis and partial least squares(PLS) analysis were performed to establish component-activity relationship of LTC,identify the potential active components.After that,validation of the potential active components in LTC was carried out by using the same models.The results indicated that 4 phenolic compounds including 7,4'-dihydroxyhomoisoflavanone,loureirin C,4,4'-dihydroxy-2,6-dimethoxydihydrochalcone,and homoisosocotrin-4'-ol,might be the active components for anti-neuroinflammation effect;five phenolic compounds such as 3,5,7,4'-tetrahydroxyhomoisoflavanone,loureirin D,7,4'-dihydroxyhomoisoflavane,and 5,7-dihydroxy-4'-methoxy-8-methyflavane,might have positive effects on the vascular endothelial injury;three phenolic compounds including 5,7,4'-trihydroxyflavanone,7,4'-dihydroxy-5-methoxyhomoisoflavane,and loureirin D,might be the active components in LTC against neuronal injury.