Application progress on data-driven technologies in intelligent manufacturing of traditional Chinese medicine extraction / 中国中药杂志

The application of new-generation information technologies such as big data, the internet of things(IoT), and cloud computing in the traditional Chinese medicine(TCM)manufacturing industry is gradually deepening, driving the intelligent transformation and upgrading of the TCM industry. At the current stage, there are challenges in understanding the extraction process and its mechanisms in TCM. Online detection technology faces difficulties in making breakthroughs, and data throughout the entire production process is scattered, lacking valuable mining and utilization, which significantly hinders the intelligent upgrading of the TCM industry. Applying data-driven technologies in the process of TCM extraction can enhance the understanding of the extraction process, achieve precise control, and effectively improve the quality of TCM products. This article analyzed the technological bottlenecks in the production process of TCM extraction, summarized commonly used data-driven algorithms in the research and production control of extraction processes, and reviewed the progress in the application of data-driven technologies in the following five aspects: mechanism analysis of the extraction process, process development and optimization, online detection, process control, and production management. This article is expected to provide references for optimizing the extraction process and intelligent production of TCM.

Application of RUNX2 gene over expression vector modified exosomes from BMSC combined with calcium carbonate scaffold system in bone defect / 中国骨伤

OBJECTIVE@#To investigate the effect of RUNX2 gene overexpression vector modified exosomes derived from bone marrow mesenchymal stem cells (BMSCs) combined with calcium carbonate scaffold system in bone defect.@*METHODS@#Rabbit BMSCs were used as the research object, and BMSCs were identified by flow cytometry. Construct RUNX2 gene overexpression vector, transfect BMSCs with lentivirus, and collect exosomes by ultracentrifugation. The morphology of exosomes was observed by transmission electron microscope, the expression of exosome marker CD63 was detected by Western blot, and the calcium carbonate scaffold was constructed by three chamber parallel automatic temperature control reaction system. According to whether the RUNX2 gene overexpression vector was transfected or not, the complex of BMSCs and calcium carbonate scaffold was divided into three groups, namely BMSCs group, RUNX2 overexpression group and exosome group. The osteogenic differentiation of BMSCs was detected by oil red O staining and RT-PCR. There were 9 clean adult healthy male New Zealand white rabbits, aged (12.97±1.21) months, with a body weight of (19.3±3.6) kg, with 3 rabbits in each group. The animal model of skull defect was constructed by surgical method, and the repair of bone defect was evaluated by imaging, he staining and Masson staining.@*RESULTS@#The results of flow cytometry showed that the expression of CD29 protein, CD44 protein, CD11b protein and CD45 protein on the surface of BMSCs were 99.5%, 100%, 0.1% and 0.1%, respectively. Transmission electron microscopy showed that the exosomes were bilayer vesicles with a diameter of 50 to 150 nm. Western blot showed that the molecular marker CD63 of exosomes was positive. Oil red O staining showed that the osteogenic differentiation of BMSCs in exosome group was significantly higher than that in RUNX2 overexpression group and BMSCs group. The results of RT-PCR showed that the relative expressions of RUNX2, BMP-2 and ALP mRNA in BMSCs in exosome group were significantly higher than those in RUNX2 overexpression group and BMSCs group (P<0.05). The imaging results showed that the repair effect of skull defect in exosome group was better than that in RUNX2 overexpression group. HE staining and Masson staining showed that the repair effect of skull defect in exosome group was better than that in RUNX2 overexpression group (P<0.05). MSCs in exosome group was significantly higher than that in RUNX2 overexpression group and BMSCs group. The results of RT-PCR showed that the relative expressions of RUNX2, BMP-2 and ALP mRNA in BMSCs in exosome group were significantly higher than those in RUNX2 overexpression group and BMSCs group(P<0.05). The imaging results showed that the repair effect of skull defect in exosome group was better than that in RUNX2 overexpression group. HE staining and Masson staining showed that the repair effect of skull defect in exosome group was better than that in RUNX2 overexpression group(P<0.05).@*CONCLUSION@#Compared with RUNX2 gene overexpression vector transfection, extraction of exosomes directly can promote the differentiation of BMSCs into osteoblasts more efficiently, and the combination with calcium carbonate scaffold can better promote the healing of bone defects. So as to provide new ideas and methods for the clinical treatment of bone defects.

Research on batch-to-batch quality control of compound Danshen extract based on physical characterization and multivariate statistical analysis / 中国中药杂志

Physical attributes of Chinese herbal extracts are determined by their chemical components, and the physical and chemical attributes jointly affect the preparation process performance and the final product quality. Therefore, in order to improve the quality control of Chinese herbal extracts, we should comprehensively study the batch-to-batch consistency of physical and chemical attributes as well as the correlations between them. This paper first explored the physical attributes affecting the preparation process performance of the compound Danshen extract and developed a method for characterizing the texture attributes. With such main chemical components as water, phenolic acids, saponins, and saccharides and texture, rheology, and other physical attributes taken into consideration, the batch-to-batch quality fluctuation of products from different production lines and time was analyzed by principal components analysis(PCA). Finally, the correlation and partial least squares(PLS) analysis was conducted, and the regression equation was established. The fitting result of the PLS model for dynamic viscosity was satisfying(R~2Y=0.857, Q~2=0.793), suggesting that the chemical components could be adjusted by the component transfer rate in the extraction process, the impurity removal rate in the alcohol precipitation process, and the water retention rate of the concentration process to meet the control of the extract dynamic viscosity. This study clarified the correlations between physical and chemical attributes of the compound Danshen extract and established a method for controlling its physical attributes based on process regulation, which would provide reference for improving the quality control of Chinese herbal extracts.

Research on feedforward control method of mixing process for compound Danshen Dripping Pills based on quality by design(QbD) / 中国中药杂志

The mixing process is one of the key operation units for solid preparation of traditional Chinese medicine. The physical properties such as particle size, density and viscosity of the mixture are key factors that need to be controlled, which will directly affect the performance of the preparation molding process and product quality. Subsequent dripping process performance and appearance qua-lity of dripping pills will be affected by dynamic viscosity of materials in the mixing process. Based on this, with mixing process of compound Danshen dripping pills as the object, a feedforward control method for the dripping pill mixing process was established based on the concept of quality by design(QbD). Firstly, critical quality attribute(CQA)-dynamic viscosity, critical material attributes(CMAs)-the moisture content of compound Danshen extract, average molecular weight of polyethylene glycol 6000 and critical process parameter(CPP)-mixing temperature were identified through the analysis of properties for multiple batches of the raw materials and excipients as well as technological mechanism. Then the Box-Behnken experimental design was used to establish the regression model among CMA, CPP and CMA(R■=0.972 0, RMSE =16.24) to obtain the design space. Finally, through the verification of three batches within the design space, the mixing process temperature was adjusted according to the properties of the raw materials and exci-pients to achieve accurate control of the dynamic viscosity attribute. The relative deviation between the actual dynamic viscosity value and the target value was less than 3.0 %. The feedforward control of the mixing process of compound Danshen dripping pills was rea-lized in this study, which can contribute to improving quality consistency of the mixing process intermediates, simultaneously provide a reference for the research on the process quality control of other Chinese medicine dripping pills.

Lentivirus mediated siRNA hsa-circ-0000885 transfection of BMSCs and osteoclast co-culture system on cell differentiation, proliferation and apoptosis / 中国骨伤

OBJECTIVE@#To explore the effects of siRNA hsa-circ-0000885 modified bone marrow mesenchymal stem cells (BMSCs) on osteogenic differentiation, cell proliferation and apoptosis in order to provide new ideas and methods for the clinical treatment of osteoporosis (OP).@*METHODS@#From September 2018 to February 2020, 13 patients with osteoporosis admitted to our hospital were selected as the research objects, including 11 females and 2 males, with an age of (65.45±10.77) years old. After obtaining the informed consent of patients, peripheral blood tissues were extracted. Then the expression level of cir-cRNA in peripheral blood mononuclear cells(PBMC) was detected by circ RNA chip. The expression of circ RNA was silenced by siRNA technology. The BMSCs were transfected with lentivirus. According to the siRNA interference plasmid hsa-circ-0000885, the cells were divided into the blank group, the empty vector group and the siRNA interference group. After 72 hours of treatment, the cell cycle was detected by flow cytometry, the apoptosis level was detected by AV-PI kit, and the osteogenic differentiation ability of BMSCs was detected by ALP staining.@*RESULTS@#The expression of hsa-circ-0000885 in PBMC of patients with osteoporosis was significantly higher than that of healthy controls (@*CONCLUSION@#The lentivirus mediated siRNA hsa-circ-0000885 plasmid transfected into BMSCs and osteoclast co culture system can promote cell proliferation, inhibit apoptosis and promote osteogenic differentiation of BMSCs, which can be used as a potential therapeutic target for OP patients.

Characteristics of collateral circulation in adult moyamoya disease based on modified Suzuki staging / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the characteristics of collateral circulation in adult moyamoya disease (MMD).</p><p><b>METHODS</b>The clinical data were collected from all adult patients with MMD undergoing digital subtractive angiography (DSA) in our department from 2006 to 2016. Based on the imaging findings, the patients were divided into ischemia group and bleeding group. A double-blind analysis was conducted of the CT or magnetic resonance imaging findings and the severity of the disease was graded using the modified Suzuki score (mSS). We classified the anastomotic networks in MMD into the superficial meningeal type and deep parenchymal type. The superficial meningeal type was further classified into the leptomeningeal and the durocortical networks, and the deep parenchymal networks into subependymal networks and the inner striatal and inner thalamic networks.</p><p><b>RESULTS</b>No significant difference was found in the distribution of mSS scores between the hemorrhage group and the ischemic group (Χ=5.812, v=5, P=0.325), but the posterior communicating artery and internal carotid artery diameter ratio (Pcom/ICA ratio) was significantly greater in the hemorrhage group (t=2.119, v=108, P=0.036). The Pcom/ICA ratio differed significantly among the groups with different mSS scores (f=8.924, P=0.00), higher in groups with mSS scores of 3, 4 and 5. The incidence of anterior choroidal artery dilation differed significantly between hemorrhage and ischemic groups (Χ=11.79, P=0.001). The incidences of durocortical networks (Χ=0.327, P=0.567) and subependymal networks (Χ=0.011, P=0.917) were comparable between hemorrhage group and ischemic groups, but the incidence of leptomeningeal networks (P=0.018) and inner striatal and inner thalamic networks (Χ=7.551, P=0.006) differed significantly between the two groups.</p><p><b>CONCLUSION</b>The collateral circulation vascular system is an important component of cerebral blood flow in MMD patients and varies from patient to patient. Patients with MMD exhibit increased Pcom/ICA ratio with abnormal expansion of the anterior choroidal artery, and the leptomeningeal networks and the inner striatal and inner thalamic networks are independent risk factors for cerebral hemorrhage.</p>

Study on chemical components distribution in Paeoniae Radix Alba and its processing methods / 中国中药杂志

After studying the discipline of chemical components distribution in peony root, it was found that the content distribution of albiflorin, paeoniflorin, benzoylpaeoniflorin and paeonol was inhomogeneous in different parts of peony root, as same as in different roots belonging to a same plant. Therefore, in order to minimize the effects of different qualities of materials on experimental result, the content changing percentages of four chemical components was chosen as indicators to study processing method of Paeoniae Radix Alba. The processing technique of Paeoniae Radix Alba was put peony roots in the boiled water for at most ten minutes, and cut them into 2-3 mm slices after peeling the root bark, finally dry them in a oven under 55 degrees C, which was similar to the traditional processing method used in Bozhou. And the peeling, boiling and drying methods would affect the contents of albiflorin, paeoniflorin, benzoylpaeoniflorin and paeonol on several levels.

Determination of chemical components of Paeoniae Radix Alba decoction pieces and its quality evaluation / 中国中药杂志

<p><b>OBJECTIVE</b>An UPLC method was developed to evaluate the quality of Paeoniae Radix by simultaneously determining four components, paeoniflorin, albiflorin, benzoylpaeoniflorin and paeonol in Paeoniae Radix Alba decoction pieces.</p><p><b>METHOD</b>The UPLC chromatographic column was ACQUITY UPLC® HSS T3. The mobile phase was acetonitrile-0.05% phosphoric acid water with gradient elution. The column temperature was 30 °C and detection wavelength was 230 nm with a flow rate of 0.4 mL · min(-1). A linear model was obtained through principal component analysis (PCA), and PCA scores were used to evaluate the quality of Radix Paeoniae Alba decoction pieces comprehensively.</p><p><b>RESULT</b>Paeoniflorin, albiflorin, benzoylpaeoniflorin and paeonol could be well separated from other components, and the results of specificity, precision, repeatability, linearity, recovery rate and stability reached the standards, respectively. The content of paeoniflorin in 9 batches of Paeoniae Radix Alba decoction pieces was below the standard given by Chinese Pharmacopoeia (2010 edition). Using the comprehensive scoring method with principal component analysis, the results showed that the samples from Zhejiang province have better quality than those from Anhui and Shandong provinces.</p><p><b>CONCLUSION</b>The method established in this study can effectively determine the content of paeoniflorin, albiflorin, benzoylpaeoniflorin and paeonol, which could be used for quality control of Paeoniae Radix Alba.</p>

Estabishment of Identification Method for Longdan Xiegan Pills(Honey Pills) by TLC / 中国药师

Objective:To research the TLC identification method for Longdan Xiegan pills( honey pills) . Methods:TLC was used in the identification. The samples were extracted by 70% methanol with a heating reflux method, and then extracted by the agents with dif-ferent polarity, including petroleum ether, ethyl acetate and butanol. The petroleum ether part was detected by fluorescence at 365nm for Angelica sinensis, and 1% vanillin-sulfuric acid color reaction was used to detect Alisma orientale. The ethyl acetate part was determined by fluorescence at 365nm for Scutellaria baicalensis, and the butanol part was detected with chloroform-methanol-water (30∶12∶3) as the developing solvent for Bupleurum and Glycyrrhiza, and with acetone-ethyl acetate-water (6∶6∶1) as the developing solvent for gentiopi-croside, geniposide and liquiritin. Results:The developed TLC spots were clear with good separation, high specificity and promising re-producibility. Conclusion:The method can be exactly used in the qualitative identification and quality control of Longdan Xiegan pills ( honey pills) .

Optimization of extraction technology from Paeoniae Radix Alba using response surface methodology / 中国中药杂志

To ensure the stability of chemistry components and the convenience of operation, ultrasound method was chosen to study in this investigation. As the total common peaks area in chromatograms was set to be evaluation index, the influence on the technology caused by extraction time, ethanol concentration and liquid-to-solid ratio was studied by using single factor methodology, and the extraction technology of Paeoniae Radix Alba was optimized by using response surface methodology. The results showed that the extracting results were most affected by ethanol concentration; liquid-to-solid ratio came the second and extraction time thirdly. The optimum ultrasonic-assisted extraction conditions were as follow: the ultrasonic extraction time was 20.06 min, the ethanol concentration in solvent was 72.04%, and the liquid-to-solid ratio was 53.38 mL · g(-1), the predicted value of total common peaks area was 2.1608 x 10(8). Under the extraction conditions after optimization, the total common peaks area was 2.1422 x 10(8), and the relative deviation between the measured and predicted value was 0.86%, so the optimized extraction technology for Paeoniae Radix Alba is suitable and feasible. Besides, for the purpose of extracting more sufficiently and completely, the optimized extraction technology had more advantages than the extraction method recorded in the monogragh of Paeoniae Radix Alba in Chinese Pharmacopoeia, which will come true the assessment and utilization comprehensively.

Determination of 23-Acetate Alisol B in Longdan Xiegan Honey Pills by HPLC / 中国药师

Objective:To establish a method for the determination of 23-acetate alisol B in Longdan Xiegan honey pills by HPLC. Methods:The analysis was performed on a Waters Symmetry C18 (250 mm × 4. 6 mm,5μm) column with the mobile phase of acetonitrile-0. 1% phosphoric acid (62 ∶ 38). The flow rate was 1. 0 ml·min-1, the column temperature was 35℃ and the detection wavelength was set at 208nm. Results: The linear range of 23-acetyl alisol B was 19. 999 5- 1 999. 9500 ng(r =0. 999 9), and the average recovery was 95. 56%(RSD = 0. 7%, n = 6). Conclusion: The method is simple, rapid and accurate, and can be used to control the quality of Longdan Xiegan honey pills with good repeatability and recovery.

Practice and exploration of the health reform in Wuhu city / 中华医院管理杂志

Wuhu City has been exploring,since 2007,an innovative health and medical system featuring both the reform of public hospitals and that of the primary care system.This reform aims at strengthening the public health service system,improving the basic medical service system,speeding up the construction of the medical security system,and improving the drug supply system.It is characteristic of a core,two wings,and four-wheel drive,which tells the essence of Wuhu' s reform practices and outcomes.

Cloning and expression analysis of leucoanthocyanidin reductase gene in Fagopyrum dibotrys / 药学学报

The leucoanthocyantin reducase (LAR) gene, an important functional gene of catechins biosynthesis pathway, was cloned from Fagopyrum dibotrys (D.Don) Hara by degenerate PCR and rapid amplification of cDNA ends (RACE). The full-length cDNA of FdLAR is 1 581 bp (GenBank accession: JN793953), containing a 1 176 bp ORF encoding a 391 amino acids protein, and its 3'-untranslated region has an obvious polyadenylation signal. The recombinant plasmid containing FdLAR completed ORF was transformed into E. coli BL21 (DE3). The target fusion peptide with molecular weight of 66 kD was expressed under the condition of 16 degrees C and induced by IPTG at final concentration of 1.0 mmol x L(-1). Bioinformation analysis indicated that the amino acid sequence of FdLAR showed great homology to other LAR with the NADB-Rossmann conversed domain in the N-terminus. Real-time quantitative PCR was used to detect the expression levels of FdLAR gene during different development periods. The determination of flavonoids contents in appropriate rhizomes showed that the relationship between FdLAR gene expression and the accumulation of flavonoids displayed different trends during vegetative growth and reproductive growth stages, suggesting that the FdLAR gene may be involved in the pathway of flavonoid metabolisms in Fagopyrum dibotrys.

Advances in the quantitative analytical methods of drug polymorphism / 药学学报

Polymorphism of drug is known to influence the stability, dissolution, bioavailability and other performance characteristics of the products. Therefore, the crystal form of the drug must be identified and determined in order to ensure consistent product performance. Even if the identification and characterization of crystal forms are performed thoroughly and the effective crystal form is selected for preparation, it is important to ensure that the effective crystal form in the final product remains unchanged. Therefore, it is essential to quantitate the content of the effective crystal form in the product to control the quality and performance of them. X-ray powder diffraction, FT-Raman, mid-IR, near-IR, terahertz pulsed spectroscopy, solid-state NMR spectroscopy, and DSC are the quantitative methods of crystal form used in the recent 10 years. This review briefly highlights the basic principles and the progress of these methods and discusses the perspective as they apply to pharmaceutical research and development.

Inhibitory effect of docetaxel on proliferation of human lens epithelial cells / 中华实验眼科杂志

Background Some drugs with inhibitory effect on the proliferation of lens epithelial cells have a limiting application in clinic because of their adverse response.To screen the effective and less side-effect drug for supressing LECs growth is very inportant for the prevention and treatment of after cataract.Objective This study was to explore the effects of docetaxel on LECs growth and compare its role with epirubicin hydrochloride,pirarubicin hydrochloTide and rahitrexed.Methotis Immortalized human LECs line (SRA01/04) were cultured and passaged.Different concentrations of docetaxel,epirubicin hydrochloride,pirarubicin hydrochloride and rahitrexed were added into the medium respectively for 24.48 and 72 hours.The proliferation of LECs was detect by M1Yr.Flow cytometry analysis Was used to analyze the influence of different concentrations of docetaxel on cellular cycle at 48 hours after addition of docetaxel,and Annexin V-FITC/PI marking method was used to assesse the apoptosis of LECs under the action of docetaxel.Expression of bcl-2 protein in LECs Was evaluated by Westeru blot. Result The growth rate of LECs Wag 100%in 8-519 pmol/L doeetaxel groups with the normal cell shape.Majority of abnormal cells and low growth rate were found in 66 nmoVL docetaxel group at 48 and 72 hours.The IC50 of docetaxel was lowest in 48 and 72 hours in docetaxel group in comparison to epirubicin hydrochloride and pirarubicin hydrochloride. However,no evident inhibition on LECs growth in 23.22-523.56 μmol/L of raltitrexed.At 48 hours,the percentage of LECs in G2/M phase increased as the asccnte of concentration of docetaxel,showing a significant difference among 4 groups(F=2633.05,P＜0.01).The percentage of early apoptotic cells increased to 22.4%(χ2=20.00,P＜0.01) and 27.9%(χ2=42.68,P＜0.01)from normal control 3.1% at 48 hours after LECs exposed to 8.3 nmol/L and 266 nmol/L docetaxe.The expression of bcl-2 protein in LECs was obviously weakened after addition of docetaxel,especially 8.3 nmol/L docetaxel group. Conclusion Docetaxel,epirubicin hydrochloride and pirarubicin hydrochloride can inhibit the proliferation of human LECs in vitro.But there is no supression on LECs growth inraltitrexed.Docetaxel is proved to have a strongly arrested effect on the proliferation of LECs in comparison with epirubicin hydrochloride and pirarubicin hydrochloride and play its role at concentration-and time-dependent manner.

Molecular cloning and characterization of a novel isoflavone reductase-like gene (FcIRL) from high flavonoids-producing callus of Fagopyrum cymosum / 药学学报

Lignans are important defensive compounds in plants and have good biological activities protecting human health. In order to study the medicinal secondary metabolism of Fagopyrum cymosum (Trev.) Meisn, a traditional Chinese medicine with anti-tumor effect, a novel isoflavone reductase-like gene, FcIRL, was cloned using RACE strategy from a cDNA library of high flavonoids-producing callus. The full-length cDNA of the FcIRL was 1 217 bp (accession no. EU116032), which contained a 942 bp open reading frame (ORF) encoding a 313 amino acid protein. Two stop codons (TAG) and a putative polyadenylation signal ATAAA at 24 bp upstream from the polyadenylation site was found in 5' and 3' UTR, separately. And no intron was found in the genomic sequence yet. FcIRL contained a predicted N-terminal acetylation site (M1-K5) and a NADPH-binding motif (G10-G-T-G13-Y-I-G16) in the N-terminal region, a conserved NmrA (nitrogen metabolite repression regulator) domain (V6-N244), multi-phosphorylation sites and one conserved N-glycosylation site (N214). Sequence homology comparison, phylogenetic analysis and advanced structures prediction all suggested that FcIRL belonged to the class of pinoresinol-lariciresinol reductase (PLR), which is a key enzyme in synthetic pathway of 8-8'-linked lignans, with function in catalyzing reduction of pinoresinol and lariciresinol into secoisolariciresinol, and medicinal secondary metabolism and resistance in F. cymosum.

Pathology of enterovirus 71 infection: an autopsy study of 5 cases / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the clinicopathologic features of fatal enterovirus 71 (EV71) infection.</p><p><b>METHODS</b>Autopsy was performed in 5 neonates died of EV71 infection. Tissue samples from major organs were collected, formalin-fixed and examined under light microscopy. Immunohistochemical study was carried out in selected examples.</p><p><b>RESULTS</b>Four of the 5 cases showed predominant changes in central nervous system, with encephalitis and encephalomyelitis identified mainly in brainstem and upper cervical spinal cord. Histologic findings included neuronal degeneration and necrosis, neuronophagia, perivascular cuffing and diffuse or nodular hyperplasia of macrophages/microglia. Cerebral edema, brain herniation and aseptic meningitis were also noted. The lungs showed mainly pulmonary congestion, neurogenic pulmonary edema and focal hemorrhage. There were minimal changes in the intestinal epithelium. The intestinal lymphoid tissue however was hyperplastic and associated with apoptosis of follicular center cells. The remaining case had cerebral edema and mild meningitis. The lung alveolar septa were thickened with lymphocytic infiltrates. Some alveolar cells were hyperplastic and associated with diffuse hyaline membrane formation. No specific abnormalities were identified in gastrointestinal tract. In all the 5 cases studied, there was enlargement of lung hilar and mesenteric lymph nodes, coupled with apoptosis of follicular center cells. In general, no significant pathologic changes were demonstrated in heart, liver and kidneys.</p><p><b>CONCLUSIONS</b>In fatal EV71 infection, the major pathologic changes lie in the central nervous system. The pulmonary lesions are mainly secondary in nature. The usual cause of death is cerebral edema complicated by brain herniation and pulmonary edema. It is also noteworthy that some cases show only lung damages, without classic neurologic changes.</p>

Molecular confirmation of enterovirus type 71 infection: a post-mortem study of two cases / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the diagnostic application of molecular detection of enterovirus type 71 (EV71) infection using post-mortem paraffin-embedded tissue.</p><p><b>METHODS</b>Two autopsy cases of EV71 infection were studied by histopathological and immunohistochemical methods. Reverse transcription polymerase chain reaction (RT-PCR) was performed to detect the viral RNA in paraffin-embedded tissue samples.</p><p><b>RESULTS</b>Characteristic features of acute encephalitis were seen in the brain, with most prominent lesions found in the brain stem in both cases. Inflammatory cells were largely CD68-positive microglia with a few CD15-positive neutrophils in the areas of neuronal necrosis. The 5'-untranslated region of EV71 was detected in the medulla by RT-PCR using paraffin-embedded tissues of both cases. Sequencing analysis of the RT-PCR products showed 100% homology to the EV71 strain, recently submitted to the GenBank database from Fuyang, Anhui province.</p><p><b>CONCLUSIONS</b>Molecular detection of EV71 can be performed on formalin-fixed, paraffin-embedded tissue samples from fatally infected patients. Timely and accurate diagnosis of the infection by such molecular approach is crucial for the proper clinical and public health intervention.</p>

Preparation of in situ gel systems for the oral delivery of ibuprofen and its pharmacokinetics study in beagle dogs / 药学学报

The in situ gel systems can form gel in situ after administration to achieve sustained release, thus provides a promising strategy for drug delivery systems. The aim of this study was to design and prepare in situ gel systems for the oral delivery of ibuprofen (IBU-ISG) and study its pharmacokinetics in Beagle dogs. The characteristics of the basic material of gellan gum (Kelcogel, Kel) and sodium alginate (Manugel, M) were studied through investigating the complex viscosity of the Kel or M solution with or without different concentrations of calcium ion or sodium citrate to ascertain the amount range of the excipients. The measurement of complex viscosity of the solution (0. 5% Kel and 1% M) with different concentrations of sodium citrate and calcium ion was carried out to select the suitable proportion of calcium ion and sodium citrate. The formulation of binary IBU-ISG was optimized by monitoring the complex viscosity before gelling in vitro release property. The optimized formulation contains 1.0% sodium alginate, 0.5% gellan gum, 0. 21% sodium citrate and 0.056% calcium chloride. A single oral dose of IBU-ISG and reference formulation (IBU suspension) were given to each of the 6 healthy Beagle dogs, ibuprofen in plasma at different sampling times was determined by RP-HPLC. The pharmacokinetics parameters in 6 Beagle dogs were calculated. The Tmax of IBU-ISG and reference formulation were (1.8 +/- 0.6) and (0.4 +/- 0. 1) h. The Cmax values were (29.2 +/- 7.6) and (37.8 +/- 2.2) microg x mL(-1). The T(1/2) were (2.3 +/- 0.5) and (2.0 +/- 0.9) h, and the AUC(0-t) were (131.0 +/- 38.6) and (117.3 +/- 23.1) microg x mL(-1) x h, respectively. The binary IBU-ISG was successfully prepared.

siRNA silences mdr1 gene expression and reverses apoptosis resistance of K562/ADM cells line / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the effect of small interfering RNA(siRNA) on silence of mdr1 gene and reversal of apoptosis resistance in multidrug-resistant (MDR) human leukemia K562/ADM cell.</p><p><b>METHODS</b>Human MDR leukemia cell line K562/ADM was used as the target cells. Two siRNAs (mdr1 siRNA-1 and mdr1 siRNA-2) targeted mdr1 gene were chemically synthesized and transfected into K562/ADM cells with liposome. Expression of mdr1 mRNA was determined by real-time PCR, P-glycoprotein (P-gp) expression and caspase-3 activity were measured with flow cytometry (FCM), and the cell apoptosis was observed by optical and electronic microscopy for morphology and Annexin V/PI staining.</p><p><b>RESULTS</b>The mdr1 siRNA-1 and mdr1 siRNA-2 could markedly down-regulate the expression of mdr1 gene in K562/ADM cells, the expression of mdr1 mRNA decreased by 91.2% and 82.0% , and the P-gp by 74.1% and 84.4%, respectively. The caspase-3 activity was markedly enhanced, and the active caspase-3 in K562/ADM cells increased by about 40% compared to liposome alone and non-silencing controls. the sensitivity of K562/ADM cells to adriamycin-induced apoptosis was significantly augmented, the apoptotic rate of the cells treated with siRNA plus adriamycin increased by about 60% compared to adriamycin alone.</p><p><b>CONCLUSION</b>siRNAs silence the expression of mdr1/P-gp to overcome the P-gp-mediated apoptosis resistance in drug-resistant K562/ADM cells.</p>