Animals ; Epithelial-Mesenchymal Transition ; Fibrosis ; Kidney ; pathology ; Kidney Tubules ; cytology ; Male ; Rats ; Rats, Sprague-Dawley ; Ureteral Obstruction ; pathology

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Non-Small-Cell Lung ; genetics ; Female ; Genes, erbB-1 ; genetics ; Humans ; Lung Neoplasms ; genetics ; Male ; Middle Aged ; Mutation ; Nucleic Acid Amplification Techniques ; methods ; Receptor, Epidermal Growth Factor ; genetics

The oral administration of bioactive macromolecular drugs such as proteins, peptides and nucleic acids represents unprecedented challenges from the drug delivery point of view. One key consideration is how to overcome the gastrointestinal tract absorption barrier. Recent studies suggest that microfold cell (M cell), a kind of specialized antigen-sampling epithelial cell which is characterized by a high endocytic rate and low degradation ability, may play an important role in macromolecule oral absorption. The development of an in vitro M cell coculture system and its modified models greatly advanced the study of M cells and the development of oral delivery system for macromolecular drugs. The special structure, function and formation characteristics, and biomarkers of M cell are summarized in this review. The applications of in vitro M cell models in developing oral delivery system ofbioactive macromolecular drugs are discussed.
Administration, Oral ; Animals ; Drug Delivery Systems ; methods ; Humans ; Intestinal Mucosa ; cytology ; Macromolecular Substances ; administration & dosage ; pharmacokinetics ; Models, Biological ; Peptides ; administration & dosage ; pharmacokinetics ; Peyer's Patches ; cytology ; Proteins ; administration & dosage ; pharmacokinetics ; Vaccines ; pharmacokinetics

To establish a fluorescent quantitative PCR method (FQ-PCR) with TaqMan probe for simultaneous detection of polyomavirus (BKV) and cytomegalovirus (CMV) and to evaluate its clinical application in the renal transplantation recipients. The conservative sequences of BKV and CMV were targeted and amplified by nested PCR technique. The PCR products were cloned into the plasmids pcDNA3. 1(+). The recombinant plasmid containing target sequences of BKV and CMV were constructed as external standards. The TaqMan-based assay was optimized. For evaluating the assay, the sensitivity was determinated by diluted standard (5 X 103-10icopies/mL), and the specificity was verified by negative control and positive control, and the precision was assessed by intra-assay coefficient of variation (ICV) through detecting standard repeatedly (20 times). A total of 480 blood samples of renal transplantation recipients were used to detect BKV and CMV DNA simultaneously with FQ-PCR, and the concentrations of FK506 were measured by ELISA. The association of DNA copy and concentrations of FK506 was analyzed. The cloned target BKV and CMV DNA was confirmed by sequencing and analysis. The sensitivity of the FQ-PCR assay reached 5 X 103 copies/ml in detecting BKV or CMV DNA. Control DNA verified the assay specifically detecting target DNA. The precision of the assay to quantif target DNA copies was acceptable (Intra-assay CV was 3.44% for BKV and 2.23% for CMV; Inter-assay CV was 4. 98% for BKV and 3.76% for CMV;). Of 480 samples, 130 samples (27. 08%) were CMV DNA positive, significantly higher than the BKV DNA positive (13.33%, 64/480, P<0.05). The positive BKV or CMV DNA was found to be associated with high concentrations of FK506 (P<0. 05). In conclusion, the developed real-time PCR assay for detecting both CMV and BKV DNA simultaneously was s high sensitive, precise and time-effectiveand could be applied in the monitoring of the CMV and BKV infection in the renal transplantation recipients.
Adolescent ; Adult ; Aged ; Conserved Sequence ; Cytomegalovirus ; genetics ; isolation & purification ; Cytomegalovirus Infections ; diagnosis ; virology ; DNA, Viral ; blood ; Female ; Humans ; Immunosuppressive Agents ; blood ; Kidney Transplantation ; adverse effects ; Male ; Middle Aged ; Polyomavirus ; genetics ; isolation & purification ; Polyomavirus Infections ; diagnosis ; virology ; Real-Time Polymerase Chain Reaction ; methods ; Reproducibility of Results ; Sensitivity and Specificity ; Species Specificity ; Tacrolimus ; blood ; Time Factors ; Tumor Virus Infections ; diagnosis ; virology ; Viral Load ; Young Adult

OBJECTIVETo explore the primary experimental methods to construct tissue engineering blood vessel.

METHODSUsing the collagen-chitosan to prefabricate the scaffolds with 3-dimensional structure, the proliferated human endothelial cells (ECs), smooth muscle cells (SMCs) and fibroblasts act as the seed cells. The cells were seeded to scaffolds in two-step method, and engineering tissue were matured by static culture or bioreactor culture. Extracellular matrix contents and the platelet aggregation were examined in engineering tissue, tissue engineering blood vessels were taken as patches to repair the man-made defaults on the rats aorta.

RESULTSThe proliferated human ECs, SMCs and fibroblast can hold activity and act as seed cells. The prefabricated scaffolds, with excellently cell and tissue biocompatibility, can facilitated cells adherence and upgrowth, the cells quantities and extracellular matrix contents in engineered tissue are time dependent increase (P < 0.05). Platelet aggregation tests confirm the tissue engineering blood vessel have some anti-coagulability. Using the engineering tissue patch to repair the default, 6 aortas in 8 animal were patency till 10 days post-operation.

CONCLUSIONSThe seeding cells can be seeded on the 3-dimensional collagen-chitosan scaffolds and matured, the tissue engineering blood vessel can be constructed primarily.

Animals ; Bioartificial Organs ; Biocompatible Materials ; Blood Vessel Prosthesis ; Cell Culture Techniques ; methods ; Cell Division ; Chitosan ; Collagen ; Endothelial Cells ; cytology ; Fibroblasts ; cytology ; Humans ; Myocytes, Smooth Muscle ; cytology ; Rats ; Rats, Sprague-Dawley ; Tissue Engineering ; methods

AIM:To investigate the distribution and mechanism of M 1/M2 macrophages in inflammatory injury and repair process of renal tissues .METHODS: SD male rats ( n=45 ) were randomly divided into 2 parts: ischemia-reperfusion injury (IRI) and unilateral ureteral obstruction (UUO) renal injury.The rats with IRI were divided into sham operation group and operation groups (0, 6, 24, and 72 h after operation), and the rats with UUO were divided into sham operation group and operation groups (3, 7 and 14 d after operation).Automatic biochemical analyzer was used to detect serum levels of creatinine and urea nitrogen .The degree of renal injury in IRI group and UUO group were detected by HE staining.The expression of CD68 was examined by immunohistochemical staining .The levels of inducible nitric oxide syn-thase (iNOS), arginase-1 (Arg-1), transforming growth factor-β1 (TGF-β1) and tumor necrosis factor-α(TNF-α) were measured by ELISA.The polarizations of M1 ( CD68 +, F4/80 + and CD16/32 +) and M2 ( CD68 +, F4/80 +and CD206 +) macrophages were analyzed by flow cytometry .RESULTS:In IRI group, the infiltration of CD68 +macrophages and the degree of injury were increased with the prolongation of time in the renal tissues .At 24 h, the tissue injury and macrophage infiltration were the most serious , but then decreased .At 72 h, the tissue damage and CD68 +macrophage in-filtration were significantly reduced .In UUO group, obstructive injury was increased with the prolongation of time , and at 14 d, marked fibrous hyperplasia occurred .The infiltration of CD68 +macrophages at 7 d was the most serious , but then reduced at 14 d.Flow cytometry analysis showed that M 1 macrophages were the majority in the early stages of UUO and IRI, and the result of ELISA identified the higher level of iNOS .At the late stage of injury , the M1 macrophages were de-creased, while the M2 macrophages were increased with higher level of Arg-1.M1 macrophage-mediated early injury was due to the induction of TNF-αexpression, and M2 macrophage-mediated later recovery was due to enhancing TGF-β1 levels.CONCLUSION:The polarization of M1 and M2 macrophages is involved in the processes of UUO and IRI .M1 macrophages play a key role in early injury , and M2 macrophages contribute to the late stage of fibrotic repair .The polari-zation of macrophages during renal injury and repair provides a guiding significance for the clinical treatment .

Objective To explore the siRNA as a new antiviral therapy,evaluate the inhibition effect of siRNA based on vector on the HBV of HepG2.2.15 cell,and observe the side effect and toxicity of siRNA vector on cells and the off-target effect of siRNA.Methods Three pairs of siRNA duplexes targeting HBV C gene were designed as double strands,and the duplex were annealed and ligated into the p-Silencer-Cmv 4.1-hygro vector.The ligation products were used to transform JM109 cells.The clones with shRNA were obtained,and the vectors were purified.After the initial identification of the vector with agarose gel and the size of the inserted sequence got examined by native polyacrylamide gel electrophoresis,furthermore the sequencing was further carried out.The recombinant plasmids were purified with ultrapure Midipreps DNA Purification System.Then HepG2.2.15 cells were transfected with the plasmid mixed with siPort XP-1.The expression of HBsAg and HBeAg were detected by immunofluorescence and Western blot,and the HBV RNA was investigated by RT-PCR.Furthermore the real-time quantitive PCR was carried out to detect the changes of HBV DNA.In order to evaluate the toxicity of the shRNA,MTT was used to examine the growth rate and curve of cells.The ELISA was performed to detect the changes of interferon-? (IFN-?).Results The Western blot showed that the HBsAg and HBeAg protein were suppressed with (81.15?0.69)%,(88.12?0.92)% respectively by vector p-C2 on the third day of post-transfection.It had the similar result indicated by immunofluorescence.And the RT-PCR showed that the specific siRNA targeting HBV C gene could markedly suppress the expression of HBV mRNA and the HBV C gene mRNA was inhibited with 96.9%.The real-time quantitive PCR showed that the specific functional siRNA could markedly suppress HBV DNA copy with two orders of magnitude,while the siRNA vector had no effect on the growth of cell showed by MTT detection.Compared with the non-transfected group and p-NC group,the IFN-? level was almost the same with siRNA p-C1,p-C2,p-C3 groups.Conclusions The siRNA based on the expression vector can suppress the expression and replication of HBV in HepG2.2.15 cell.The inhibition effect was specific and had a certain dependency on siRNA concentration.No toxicity effect was found in the study.And the drug resistance wouldn′t happen because the silence was based on the split of gene.

Aim To reveal the aetion mechanism of Trillium tschonoskii Maxim (TTM) in the treatment of myoeardial ischemia ( MI) by using network pharma¬cology combined with molecular docking.Methods Compounds of TTM were detected and fished out from TCMID, TCM@TAIWAN , BATMAN-TCM database, and the literature data from PubMed , CNK1, and WAN- FANGD database.PharmMapper database was used to find the targets related to compounds, and DISGeNET, GeneCards, DrugBank and OMIM databases were used to find the targets related to Ml.The predictive targets of TTM in the treatment of Ml were obtained.Cytosca- ope 3.1.2 Software and String database were used to build compound-target network and PP1 network.Gene ontology ( GO ) functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes ( KEGG ) pathway enrichment analysis were performed by utili¬zing the CludterProfiler Software package of RStudio software.The molecular docking was used for verifying the results of network analysis.Results The 10 active compounds of TTM were screened , and 13 core targets of Ml were predicted, such as ALB, EGFR, MAPK1 , CASP3,ESR1 ,etc.A total of 28 Ml-related signaling pathways were fished out.The results of molecular docking showed that the core active ingredients had good binding activity with the key targets.Conclusion TTM may play a role in the treatment of Ml through regulating multiple ingredients, multiple pathways, and multiple targets.

OBJECTIVE: To analyze the intellectual landscape and emerging research trends of Chinese medicine (CM) in the management of pediatric asthma through a scientometric study. METHODS: Publications related to CM in the management of pediatric asthma were retrieved from the Web of Science Core Collection using relevant keywords. A scientometric study was performed using CiteSpace and VOSviewer. RESULTS: A total of 1,673 original articles and reviews from 1991 to 2019 were included in the analysis. The amount of annual publications had a gradual increase with time. USA was the major contributor both in country and institution analyses. Based on the co-citation, the published journals were grouped into 4 clusters. Keyword analysis indicated that the main hotspots were: (1) comprehensive management; (2) risk factors, mechanism, and prevalence; (3) prevention and treatment; (4) inflammation; and (5) environmental research. Lastly, we predicted that three emerging trends were quality of life promotion, immune response, and combination therapy. CONCLUSIONS CM research in the management of pediatric asthma will maintain the current trend of steady growth. This scientometric analysis may help scientists to identify the areas of interests and future directions in the field.
Asthma/drug therapy* ; Bibliometrics ; Child ; Humans ; Medicine, Chinese Traditional ; Publications ; Quality of Life