Pueraria lobate advances in molecular pharmacognosy / 中国中药杂志

Molecular pharmacognosy is a science of classification and identification, cultivation and protection, and production of active ingredients of graduated drugs at the molecular level. The proposal of molecular pharmacognosy allows the research of crude drugs to advance from the microscopic level to the genetic level. Pueraria lobata root, as a medicinal and edible plant, has high application value and economic value. There are many varieties that are easy to cause confusion, and it is not easy to distinguish and identify according to traditional identification methods. Moreover, the research of P. lobate root at the genetic level is still relatively shallow. the study received extensive attention of scholars. This article reviews recent research on molecular identification of P. lobate, transcriptome sequencing, cloning and synthesis of functional genes of P. lobate root in recent years in order to provide references for further promoting the development and utilization of P. lobate root and its active ingredients.

Construction and in vitro evaluation of targeted cisplatin-loaded nanoparticles for chemo-photothermal cancer therapy / 中国药科大学学报

@#To improve the therapeutic effect of cisplatin and reduce its side effects, a multifunctional drug delivery system with targeted and chemo-photothermal effect was constructed.Using polyethylene glycol polylactic acid block copolymer as a carrier, nanoparticles loaded with antitumor drug cisplatin and photosensitizer indocyanine green were prepared by ultrasonic emulsification, and the surface was then modified by cetuximab to prepare cetuximab-decorated and near-infrared (NIR)-activated nanoparticles (CPINPs).The physicochemical properties were characterized by mean particle size, Zeta potential, mAb conjugating rate and photothermal effect; the in vitro cell uptake was measured by laser confocal microscopy; and the in vitro antitumor activity was evaluated by CCK8 assay.The results indicated that CPINPs had mean particle diameter of (263.9 ± 3.73) nm, polydispersity index of 0.18 ± 0.03, Zeta potential of -(23.43 ± 0.42) mV, and cetuximab conjugating rate of (44.0 ± 1.72)%.The in vitro photothermal experiments showed that CPINPs upon NIR irradiation induced a photothermal effect, thus destroying the tumor cells. The in vitro cell uptake experiments demonstrated that NIR irradiation could promote cell uptake, and that more CPINPs were effectively internalized into A549 cells. The in vitro cytotoxicity test indicated that CPINPs treated with NIR irradiation had the effect of combined chemo-photothermal therapy, leading to higher cytotoxicity than that of free cisplatin or treatment without NIR, with IC50 values being (8.67 ± 0.04) μmol/L for 24 h incubation.To sumup the multifunctional drug delivery system constructed in the current work expected to be a more efficient targeted therapy strategy for lung cancer.

Difference Analysis of Chemical Constituents in Different Varieties of Puerariae Flos by UPLC-Q-TOF-MS / 中国实验方剂学杂志

Objective:To compare the chemical constituents of Puerariae Flos from three different varieties of <italic>Pueraria montana</italic> var. <italic>lobata</italic>， <italic>P. montana</italic> var. <italic>thomsonii</italic> and <italic>P</italic>. <italic>montana</italic> var<italic>. montana</italic>. Method:Ultra-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry （UPLC-Q-TOF-MS） was used with the mobile phase of 0.1% formic acid aqueous solution （A）-acetonitrile （B） for gradient elution （0-20 min， 10%-30%B； 20-30 min， 30%-55%B； 30-35 min， 55%-95%B； 35-37 min， 95%B； 37-40 min， 95%-10%B）， the flow rate was 0.25 mL·min^-1. Electrospray ionization （ESI） was used to scan and collect MS data in positive and negative ion modes with scanning range of <italic>m</italic>/<italic>z</italic> 50-1 500. The chemical components from different sources of Puerariae Flos were identified in combination with the chemical composition database and literature information. After the obtained data were normalized by MarkerView^TM 1.2.1， they were imported into SICMA-P 14.1 software for principal component analysis （PCA） and orthogonal partial least squares discriminant analysis （OPLS-DA） to select the main differentiated components among the three different varieties. Result:A total of 35 compounds were identified from three different varieties of Puerariae Flos， including 22 isoflavones， 6 flavonoids and 7 saponins. The flowers of <italic>P</italic>. <italic>lobata</italic>， <italic>P. montana</italic> var. <italic>thomsonii</italic> and <italic>P</italic>. <italic>montana</italic> var<italic>. montana</italic> contained 32， 35， 33 compounds， respectively. And 18 differential compounds were screened under the positive and negative ion modes， including kakkalide， tectoridin， 6″-<italic>O</italic>-xylosyl-tectoridin， 4'-methyltectorigenin-7-glucoside， glycitin， 6″-<italic>O</italic>-xylosyl-glycitin， irisolidone， kaikasaponin Ⅲ， 6″-<italic>O</italic>-malonylglycitin， kakkalidone， tectorigenin， rutin， soyasaponin BB， vitexin， biochanin A， genistin， kakkatin， azukisaponin Ⅱ. Conclusion:This research is the first to systematically study the chemical constituents of the flower of <italic>P</italic>. <italic>montana</italic> var<italic>. montana</italic>， although the flower of <italic>P</italic>. <italic>montana</italic> var<italic>. montana</italic> is used as adulterants， it has high contents of tectoridin and 6″-<italic>O</italic>-xylosyl-tectoridin， which has great potential for development. The efficacy components such as kakkalide and tectoridin in Puerariae Flos from the three sources of varieties are obviously different， and it is necessary to carefully consider the application of these three varieties as Puerariae Flos.

Analysis of edge morphology of partial veneers made by different processing techniques and materials / 北京大学学报(医学版)

OBJECTIVE@#To compare the edge morphology of partial veneers made of different materials by slurry molding, heat-pressed and computer aided design/computer aided manufacturing (CAD/CAM) techniques.@*METHODS@#Thirty premolars with smooth surface and intact enamel were selected and randomly divided into five groups, 6 specimens for each group. Group A were made from feldspathic porcelain (Noritake®) by slurry molding, while Group B were made from lithium disilicate glass ceramic (IPS E.max® Press) by heat-pressed. Group C/D/E were respectively made from feldspar porcelain block (VITA Mark II®), zirconia-reinforced glass ceramic (VITA Suprinity®) and hybrid ceramic with a ceramic-polymer network (VITA Enamic®) by CAD/CAM techniques. All the partial veneers luted with light-cured composite resin. Then the partial veneers were trimmed and polished to achieve the smooth finishing margin, clinical polishing sets were used according to the product descriptions. Scanning electron microscope (SEM) was used to observe the edge morphology of prostheses and the exposure of resin cements.@*RESULTS@#The smooth surface and knife-like edge of the partial veneers could be obtained after bonding, trimming and polishing. The edges of Group A were slightly rough and the width of the exposed adhesive was (106.00±9.17) μm. In Group B, the edges were smoother than Group A, and the exposed wide adhesive strip was visible, which was (138.33±20.59) μm. In Group E, the edges were smooth too, and the width of exposed adhesive strip was (186.00±5.66) μm. The edges of Group C and Group D were rough and uneven, and the adhesive was rarely exposed, they were (50.67±7.51) μm and (65.67±17.90) μm. There were all significant differences between two groups, except Group C and Group D.@*CONCLUSION@#After trimming and polishing in accordance with clinical procedures, the expected knife-like edge can be obtained in all groups. The width of the exposed resin adhesive of each group is different, the order: Mark II/Suprinity < Noritake < E.max Press < Enamic. The edge morphology of partial veneers in different processing technic and materials are different.

Establishment of a Huntington’s Disease in vitro Drug Screening Cell Model / 中国生物工程杂志

To develop a Huntington’s disease(HD) cell model in vitro to screen drugs targeting the aggregation of polyQ,different length of CAG repeat fragments were amplified by random primer PCR, identified by DNA sequencing and were fused to the N-terminus of CAT in the pCAR system respectively which had been constructed and identified before. Recombinant plasmids were transformed into and induced to express in the host E.coli. SDS-PAGE and chloramphenicol resistance test were done to determine the solubility of the polyQ and chloramphenicol resistance levels of the fusions. With different length of CAG repeat fragments cloned and expressed in the CAT-fusion protein reporting system, it is found that when the length of the fragments increased over 40, their encoding polyQ expressed as insoluble protein and chloramphenicol resistance levels are lower, while under 40, the polyQ expressed as soluble ones and chloramphenicol resistance levels are higher. A in vitro HD model that could minimize the pathological process of the HD thus has been developed. With which by measure the recombinant bacteria’s resistance to chloramphenicol, the polyQ’ solubility and folding state in vitro by quality and quantity could be determined. Thus this model can be used to screen drugs or bioactivity materials that can inhibit aggregation of the polyQ, which thereby shedding new light on the prevent, diagnosis and therapy of HD.

Effect of antisense oligonucleotides of ryanodine receptor on proliferation and Ca2+i of airway smooth muscle cells / 中国应用生理学杂志

<p><b>AIM</b>To investigate the effect of antisense oligonucleotides (ASON) of ryanodine receptor on proliferation and [Ca2+]i concentration of airway smooth muscle cells (ASMCs).</p><p><b>METHODS</b>ASMCs were cultivated with collagen enzyme digestion method. Different concentrations of ASON were added to the cultures with Lipofectamine 2000 to observe the ASMCs proliferation using MTS/PES method. The changes of ASMCs [Ca2+]i were also observed by flow cytometry. The expression of mRNA of subtypes of RyR was assayed by RT-PCR.</p><p><b>RESULTS</b>RyR ASON restrained the proliferation of ASMCs, decreased the expression of RyR and reduced the concentration of [Ca2+]i.</p><p><b>CONCLUSION</b>RyR ASON could inhibit the proliferation of ASMCs by influencing the concentration of [Ca2+]i after excited.</p>

Adaptation of myofibrilla, MHC and metabolic enzyme of rabbit diaphragm muscle to different frequency chronic electrical stimulation / 中国应用生理学杂志

<p><b>AIM</b>To detect effect of the different frequency of chronic electrical stimulation (CES) on myofibrillar isoform, myosin heavy chain (MHC) and metabolic enzyme activities.</p><p><b>METHODS</b>The histochemical method and SDS-polyacrylamide gel electrophoresis were respectively employed.</p><p><b>RESULTS</b>(1)There were a significant increase in I myo-fibrillar isoform and I MHC isoform and decrease in II B myofibrillar isoform and II B MHC isoforms in the chronic low frequency electrical stimulation (CLFES) 10 Hz and 20 Hz groups, but opposite results were found in the chronic high frequency electrical stimulation (CHFES) 50 Hz and 100 Hz groups. (2) There were a significant increase in the aerobic-oxidative enzyme activities and capacity, and a concomitant significant drop in glycolysis enzyme activities in CLFES groups, but opposite results were found in CHFES 50 Hz and 100 Hz groups.</p><p><b>CONCLUSION</b>It was suggested that there was a significant dependent relation between chronic electrical stimulation frequency and myofibrilla isoforms, myosin heavy chain (MHC) and metabolic enzyme activities.</p>

Expression of ryanodine receptor isoforms in airway smooth muscle cells of rats / 中国应用生理学杂志

<p><b>AIM</b>To detect the expression of ryanodine receptor (RyR) subtypes in normol rat airway smooth muscle cells(ASMCs) and changes during chronic asthma formation.</p><p><b>METHODS</b>ASMCs were cultured with collagen enzyme digestion method. The expression of subtypes of RyR were detected by RT-PCR. Purified PCR product linked with pGEM-T vector to make DNA sequence assay. Chronic asthma model was made with OVA, the changes of RyRs detected by RT-PCR.</p><p><b>RESULTS</b>All subtypes of RyR were expressed in airway smooth muscle cells of normol rat. The expression of RyR1 increased obviously compared with control group (P < 0.05) on chronic asthma.</p><p><b>CONCLUSION</b>Co-expression of three subtypes of RyR in ASMCs of normal rat, indicate that there are complicated intercellular Ca2+ regulation mechanism in ASM, moreover RyR1 might play a role during asthma development.</p>

Effect of chronic electrical stimulation of phrenic nerve at different frequencies on mRNA and protein expression of skeletal DHPR(alpha1) and RyRs in the diaphragm muscle of rabbits / 生理学报

The mRNA and protein expression of skeletal dihydropyridine receptor isoform alpha1 subunit (DHPR(alpha1)) and ryanodine receptor(1-3) (RyR(1-3)) during chronic electrical stimulation (CES) of phrenic nerve have rarely been explored. In the present study, we explored the signal translation mode of calcium release unit in diaphragm muscle of rabbits after CES. Thirty rabbits were used and randomly divided into the normal, 10, 20, 50 and 100 Hz groups. Phrenic nerve was continuously (5 weeks, 2x 2 h/d) stimulated at 10, 20, 50 and 100 Hz respectively (impulse width 0.2 ms, 3~6 waves/time, 45 times/min, 10~20 V). Reverse transcription PCR and immunohistochemical methods were employed. The results showed that mRNA and protein expressions of DHPR(alpha1) and RyR(1) in 10 and 20 Hz groups were more significantly lower than those in the control group (P<0.01), but mRNA and protein expressions of DHPR(alpha1) and RyR(1) were significantly higher in 50 and 100 Hz groups than those in the control group (P<0.01); a lower level of mRNA expression of RyR(2) was found in 10 and 20 Hz groups. It is suggested that the calcium release unit and the signal transduction mode between DHPR and RyRs were altered from conformational changes of linked proteins to Ca(2+)-induced Ca(2+) release (CICR) in the diaphragmatic muscle of rabbits after chronic low-frequency electrical stimulation of phrenic nerve for 5 weeks.