Objective To explore the scanning techniques of piriformis and different ultrasonographic characteristics of normal and abnormal piriformis.Methods A total of 40 cases diagnosed with unilateral piriformis syndrome underwent ultrasonic examination.Then ultrasonic scanning techniques of piriformis were summarized.Contours,thickness and smoothness of epimysium and ultrasonic echo of internal muscle texture of piriformis were compared between the normal and abnormal piriformis.The study was approved by the Third Affiliated Hospital Ethics Committee of Zhejiang Chinese Medical University (Approval no.ZSLL-JS-2016-18).Results Interruption of ultrasonic echo of ilium could be considered as ultrasonographic signs for locating piriformis quickly and accurately.Abnormal piriformis in suffering side of patients with piriformis syndrome showed obscure contours and being thicker than the other side [x2 =9.899,P =0.002;(25.81 ± 0.30)mm vs (22.29 ± 0.27)mm,t =13.604,P =0.000].Moreover,there were significant differences in comparing smoothness of epimysium and ultrasonic echo of internal muscle texture of piriformis between the two sides(x2 =23.226,P =0.000;x2 =54.848,P =0.000).Conclusions Interruption of ultrasonic echo of ilium may be an important sign for locating piriformis.Ultrasound can display piriformis clearly and distinguish ultrasonographic images of normal piriformis accurately from abnormal piriformis,which can be taken as an basis imaging for clinical diagnosis of piriformis syndrome.

Objective To evaluate the feasibility of Mycobacteria growth indicator tube ( MGIT ) liquid culture combined with rifampin resistance test real-time PCR ( Xpert MTB/RIF test) in the diagnosis of tuberculosis.Methods 652 cases of suspected pulmonary tuberculosis from October 2014 to March 2015 in Quzhou People′s Hospital were enrolled. The morning sputum samples were collected for acid-fast staining, L?wenstein-Jensen ( L-J) culture, BACTEC MGIT culture and Xpert MTB/RIF test.Samples with positive results of MGIT liquid culture were subjected to strain identification and drug sensitivity test with fluid method.Methodological comparison between four methods was made and the performance of MGIT liquid culture combined with Xpert MTB/RIF test in the rapid diagnosis and drug resistance detection in Mycobacterium tuberculosis was evaluated by chi-square test. Results Among the samples from 399 confirmed tuberculosis patients, the diagnostic sensitivity of the 4 methods ( acid-fast staining, L-J culture, MGIT culture and Xpert MTB/RIF test) were 17.0%(68/399), 23.8%(95/399), 37.8% (151/399) and 37.3%(149/399) respectively.The sensitivity and specificity of MGIT culture combined with Xpert MTB/RIF test was 39.8%(159/399) and 94.8%(240/253).The sensitivity of MGIT culture combined with Xpert MTB/RIF test was significantly higher than acid-fast staining (χ2 =50.9, P<0.01 ) and L-J culture (χ2 =23.7,P<0.01).The average detection time of MGIT culture was 7.5 days ( smear positive and Xpert MTB/RIF positive), 13.4 days (smear negative and Xpert MTB/RIF positive) and 16.9 days ( smear negative and Xpert MTB/RIF negative) .The sensitivity and specificity of Xpert MTB/RIF test in rifampin resistance detection were 9/9 and 97.3% ( 129/132 ) respectively.The average detection time of fluid method for drug sensitivity test was 8.3 days.Conclusions MGIT liquid culture combined with Xpert MTB/RIF test can detect Mycobacterium tuberculosis and the drug sensitivity rapidly.The method is highly sensitive and specific.It is important for clinical diagnosis and treatment of tuberculosis.

Objective To determine the relationship between visual signal intensity and quantitative signal intensity of HCC assessed with DWI and histopathological differentiation of HCC.Methods The MR examinations including MRI plain scan,LAVA dynamic enhanced scan and DWI (1.5T,b value:0 and 600 s/mm2) of 224 patients who had surgically resected HCCs were retrospectively reviewed.Histopathological examinations revealed that there were 31 well-,169 moderately-,and 24 poorly-differentiated HCCs.The incidence of each visually evaluated signal intensity and quantitative signal intensity of HCC assessed with DWI signal intensity and the relationship between signal intensity and histopathological differentiation were assessed for each sequence.Results (1) On DWI,56.7％ of HCCs appeared as obviously hyperintense,24.1％ tumors appeared as moderate hyperintense,and 19.2％ tumors appeared as isotense or slight hyperintense to the surrounding hepatic parenchyma.There was a significant difference between isotense/slight hyperintense and obvious hyperintense and histopathological differentiation (P ＜ 0.05).There was no significant difference between isotense/slight hyperintense and moderate hyperintense and histopathological differentiation (P ＜ 0.05).There was no significant difference between moderate hyperintense and obvious hyperintense and histopathological differentiation (P ＞ 0.05).Visually evaluated signal Intensity of HCC showed an inverse correlation with histopathological differentiation (r =-0.324,P ＜ 0.05).On DWI,the tumors tended to show a brighter signal with decreasing histopathological differentiation.(2) There was a significant difference in DWI signal intensity value among the well,moderately and poorly differentiated HCCs (P ＜ 0.05).The SI value of well differentiated HCCs was lower than that of moderately differentiated HCCs and poorly differentiated HCCs (P ＜ 0.05).The SI value of moderately differentiated HCCs was lower than that of poorly differentiated HCCs.However,there was no significant difference between the SI value of the moderately and poorly differentiated HCCs (P ＞ 0.05).ROC analysis showed that the optimal cutoff point of SI value in diagnosing well differentiated HCCs was 66.5.A cutoff SI value equal to or less than 66.5 was used to differentiate well-differentiated HCC from moderately-and poorly-differentiated HCC with a sensitivity of 90.1％ and a specificity of 71.9％.Conclusions On DWI,the tumors tended to show a brighter,visually evaluated signal intensity and higher quantitative signal intensity with decreasing histopathological differentiation (P ＜ 0.05).The quantitative signal intensity of HCC assessed with DWI signal intensity could only predict well differentiated HCC.It was limited in predicting histopathological differentiation of HCC using evaluating signal intensity and quantitative signal intensity of HCC assessed with DWI.

Traditional herbal medicines, Panax ginseng, Panax quinquefolium and Panax notoginseng, attract our attention for their extensive and powerful pharmacological activities. Ginsenosides are the main active constituents of these medicinal herbs. The related glycosyltransferases involved in ginsenoside biosynthesis are the key enzymes which catalyze the last important step. Modification of ginsenoside aglycones by glycosyltransferases produces the complexity and diversity of ginsenosides, which have more extensive pharmacological activity. At present, ginsenoside aglycones and compound K have been obtained by synthetic biology. As the last step of ginsenoside biosynthesis, glycosylation of ginsenoside aglycones has been studied intensively in recent years. This review summarizes the basic strategies and research advances in studies on glycosyltransferases involved in ginsenoside biosynthesis, which is expected to lay the theoretical foundation for the in-depth research of biosynthetic pathway of ginsenosides and their production by synthetic biology.
Biosynthetic Pathways ; Ginsenosides ; biosynthesis ; Glycosyltransferases ; metabolism ; Panax ; chemistry ; Plants, Medicinal ; chemistry ; Synthetic Biology

Pichia pastoris is one of the most important systems used in the field of molecular biology for the expression of recombinant proteins. The system has advantages of high expression, high stability, high secretion, easy high-density fermentation and low cost. Many factors affect the expression of recombinant protein, such as gene copy number, codon usage preference, type of promoter, molecular chaperones, glycosylation, signal peptide and fermentation process. In this review, research advances of the above aspects are summarized, which lay a foundation for improving the expression of recombinant proteins in P. pastoris.
Fermentation ; Gene Dosage ; Glycosylation ; Molecular Chaperones ; Pichia ; metabolism ; Promoter Regions, Genetic ; Protein Sorting Signals ; Recombinant Proteins ; biosynthesis

Lanosterol synthase is encoded by the erg7 gene and catalyzes the cyclization of 2, 3-oxidosqualene, which is a rate-limiting step of the inherent mevalonate (MVA)/ergosterol metabolic pathway in Saccharomyces cerevisiae. The intermediate 2, 3-oxidosqualene is also the precursor of triterpenoids. Therefore, the cyclization of 2, 3-oxidosqualene is the key branch point of ergosterol and triterpenoids biosynthesis. Down-regulation of 2, 3-oxidosqualene metabolic flux to ergosterol in S. cerevisiae may redirect the metabolic flux toward the triterpenoid synthetic pathway reconstructed by the synthetic biology approach. To construct erg7 knockout cassette harboring the loxP-Marker-loxP element, long primers were designed, which were homologous to the sequences of both erg7 ORF and plasmid pUG66. The cassette was transformed into diploid wild strain INVSc1 by LiAc/SS Carrier DNA/PEG method and then erg7 gene haploid deficient mutant was obtained by homologous recombination. The results of semiquantitative PCR and real-time quantitative PCR revealed that erg7 expression level in erg7 gene haploid deficient mutant is one time lower than that in wild strain. The results of TLC and HPLC showed that the ergosterol content in deficient mutant decreased to 42% of that in wild strain.
Chromatography, High Pressure Liquid ; DNA Primers ; Down-Regulation ; Ergosterol ; metabolism ; Haploidy ; Intramolecular Transferases ; genetics ; Polymerase Chain Reaction ; Saccharomyces cerevisiae ; genetics ; Squalene ; analogs & derivatives ; metabolism

Tetradrine-tashionone II(A)-PLGA composite microspheres were prepared by the SPG membrane emulsification method, and the characterization of tetradrine-tashionone II(A) -PLGA composite microspheres were studied in this experiment. The results of IR, DSC and XRD showed that teradrine and tashionone II(A) in composite microspheres were highly dispersed in the PLGA with amorphous form. The results of tetradrine-tashionone II(A) -PLGA composite microspheres in vitro release experiment showed that the cumulative release amounts of tetradrine and tashionone II(A) were 6.44% and 3.60% in 24 h, and the cumulative release amounts of tetradrine and tashionone II(A) were 89.02% and 21.24% in 17 d. The process of drug in vitro release accorded with the model of Riger-Peppas. Tetradrine-tashionone II(A) -PLGA composite microspheres had slow-release effect, and it could significantly reduce the burst release, prolong the therapeutic time, decrease the dosage of drugs and provide a new idea and method to prepare traditional Chinese medicine compound.
Benzofurans ; chemistry ; Benzylisoquinolines ; chemistry ; Drug Carriers ; chemistry ; Drug Compounding ; instrumentation ; methods ; Drugs, Chinese Herbal ; chemistry ; Kinetics ; Lactic Acid ; chemistry ; Microspheres ; Particle Size ; Polyglycolic Acid ; chemistry

In recent years, polysaccharides have received much attention because of their high safety and good immunological activity. The study of polysaccharide in vivo process is a key scientific problem that needs to be solved for polysaccharide drug development. Some progress has been made in the field of polysaccharide pharmacokinetics and immunomodulation. However, due to the lack of both chromogenic and light-absorbing groups and the complex molecular structure of polysaccharides, the in vivo processes and immunomodulatory mechanisms of polysaccharides have been slow to be investigated. The effective combination of multiple techniques can break the bottleneck of difficult tracing and unknown immunomodulatory mechanism of polysaccharides in vivo, and promote the development and utilization of polysaccharides. In this paper, we systematically summarize the key techniques in the study of polysaccharide in vivo processes and immunomodulatory mechanisms in order to provide technical references and research ideas for the study of polysaccharide in vivo processes and immunomodulatory mechanisms.

Natural products are important sources of drug discovery. However, the traditional methods of extraction and isolation, as well as chemical synthesis for obtaining natural products are associated with issues such as operational complexity, high costs, low efficiency, and environmental pollution. Constructing microbial cell factories through synthetic biology methods to produce medicinal natural products has the advantages of high efficiency, low cost, and environmental protection. Nevertheless, the scope and yield improvement of the products are limited by the limitations of enzymes in microbial cell factories. Protein engineering is considered one of the most effective approaches to overcome these limitations. This article introduces commonly used methods of protein engineering technology and summarizes its specific applications in improving enzyme performance, modifying the enzymatic environment, and promoting the development of synthetic biology tools in the field of pharmaceutical natural product synthesis. Furthermore, it analyzes the current bottlenecks and challenges in protein engineering and looks forward to its future application prospects, offering insights for the development and practical use of protein engineering technology.

Nicotinic acetylcholine receptors (nAChRs) belong to ligand-gated ion channel receptors, of which α7 nAChR subtype is widely distributed in the cerebral cortex, thalamus, hippocampus, and also identified in microglia, macrophages, bone marrow cells, etc. Previous studies revealed that α7 nAChR is closely related to the function of the cholinergic anti-inflammatory pathway, and is a vital target for drug development of Alzheimer's disease and schizophrenia. The establishment of a stable α7 nAChR in vitro drug screening system is crucial for the efficient screening of novel drugs targeting this target. Recombinant expression of different subtypes of nAChRs on Xenopus laevis oocyte membranes and current detected by two-electrode voltage clamp (TEVC) is an advanced and complex model for novel drug screening. Molecular chaperones can assist the assembly of some nAChR subunits to form functional receptors, providing a stable expression model for the screening of compounds targeting this receptor. In this study, a molecular chaperone gene of α7 nAChR, transmembrane protein 35A (Tmem35a), was isolated and cloned from rats. We constructed the recombinant expression vector and obtained the cRNA of Tmem35a by in vitro transcription technique. Two cRNAs (Tmem35a and α7) were mixed and injected into X. laevis oocytes for expression. Then, the effects of this molecular chaperone on the current expression and pharmacological properties of α7 nAChR were evaluated by the TEVC. The results revealed that TMEM35A, also known as novel acetylcholine receptor chaperone (NACHO) could effectively increase the expression of α7 nAChR protein on oocyte membranes, and the amount of α7 nAChR protein was increased about 1-fold. The peak current induced by agonist acetylcholine (ACh) was increased about 10-fold. After injection of Tmem35a cRNA, the median effect concentration (EC₅₀) value of α7 nAChR to agonist ACh is 228.5 μmol·L^-1, which shows almost no difference from native α7 nAChR (EC₅₀: 223.3 μmol·L^-1), indicating the preservation of the normal properties of α7 nAChR. The results of this investigation indicate that the molecular chaperone NACHO effectively assists the heterologous expression of α7 nAChR in X. laevis oocytes, which provides a model for screening the potency of lead compounds targeting α7 nAChR. All animal experiments in this study were reviewed and approved by the Ethics Committee of Guangxi University (approval number: GXU-2023-0249).