Objective: To establish an animal model of primary cutaneous cryptococcosis and to study the role of immune suppression in Cryptococcus neoformans (C. neoformans) infection. Methods: Twenty-four BALB/c mice were equally randomized into 2 groups: immunocompetent group and immunocompromised group. C. neoformans isolate, B3501, was intradermaly inoculated at a concentration of 5 × 107/ml into all mice. Mycology and histopathology examinations of the skin were done to confirm the infection. The skin damage courses were recorded and compared in 2 groups. Results: Different manifestations of skin infection, such as nodule, papule, ulcer and lesions mimicking molluscum contagiosum appeared 2-9 d after inoculation, with a mean of (3.42±1.17) d in immunocompromised group and (4.25±1.42) d in immunocompetent group. The lesions became self-cured 22-44 d after their onset, with a mean of (36.75±4.20) d in immunocompromised group and (29.00±4.75) d in immunocompetent group (P<0.05). Cryptococcal infection was confirmed by mycology culture and histopathology examination. Conclusion: Intradermal inoculation with C. neoformans is feasible for establishing a cutaneous cryptococcosis model in BALB/c mice. Immnunosupressent state may not be a key factor for primary cutaneous cryptococcosis.

BACKGROUND:About 10%-49%of maintenance hemodialysis patients are accompanied by refractory hypertension and common drug therapy is ineffective. Continuous renal replacement therapy could clear the middle and giant molecule toxins in the blood plasma of patients, it is theoretical y treat refractory hypertension through reducing the toxin levels.
OBJECTIVE:To observe the effect of continuous renal replacement therapy on maintenance hemodialysis patients with refractory hypertension.
METHODS:A total of 45 maintenance hemodialysis patients with refractory hypertension were randomly divided into two groups, hemodialysis group (n=22) and continuous blood purification group (n=23). Hemodialysis group underwent conventional hemodialysis treatment. Continuous blood purification group underwent conventional hemodialysis therapy, and treatment of continuous blood purification weekly.
RESULTS AND CONCLUSION:After 3 months of treatment, the renin activity, endothelin, angiotensin II, C-reactive protein, interleukin-6, and tumor necrosis factor-αlevels in continuous blood purification group were significantly lower than the levels before the experiment, while 24-hour mean blood pressure levels were significantly decreased (P<0.05). In hemodialysis group, renin activity, endothelin, angiotensin II, C-reactive
protein, interleukin-6, and tumor necrosis factor-αlevels and 24-hour mean blood pressure did not change (P>0.05). After the test, continuous blood purification group showed significantly lower levels of the above index than the hemodialysis group (P<0.05). Continuous renal replacement therapy plus hemodialysis significantly reduce the blood pressure in maintenance hemodialysis patients with refractory hypertension, the potential mechanisms are mediated by lowering the middle and giant molecule toxin as wel as inflammatory factor in plasma.

Given the vital role of vasculature in solid tumors, the potential of vascular disrupting therapy in the treatment of triple-negative breast cancer (TNBC) is promising. In this study, we prepared the acid-sensitive liposome PPD/CA4P/Lip-Rap loaded with the vascular disrupting agent CA4P and the anti-angiogenic drug rapamycin (Rap) to explore the potential of the vascular disrupting strategy in TNBC. PPD/CA4P/Lip-Rap was characterized by ¹H NMR, dynamic light scattering, and transmission electron microscopy. Its drug loading and acid sensitivity were determined. The particle size of PPD/CA4P/Lip-Rap is 161.53 ± 1.89 nm, the zeta potential is -20.03 ± 0.9 mV and it demonstrated good drug release on acidic sensitivity responses. CCK-8 experiments proved that Rap can enhance the ability of CA4P to destroy tumor vascular endothelial cells. Rap can kill marginal residual tumor cells, suppress tumor recurrence. Nanocarriers can further enhance the therapeutic effect. Western blot (WB) showed that Rap decreased the expression of hypoxia-inducible factor-1α (HIF-1α) via the mTOR/p70S6K and mTOR/4E-BP1 pathways. Thus, tumor hypoxia activation and angiogenesis were inhibited. PPD/CA4P/Lip-Rap can effectively destroy tumor vessels, inhibit tumor angiogenesis and recurrence, and provide a new strategy for the treatment of TNBC by targeting disruption of tumor vessels.

Codon usage bias is an important characteristic of genetic information transfer in organisms. Analysis of codon usage bias of different species is important for understanding the rules on genetic information transfer. The previous method for analysis of codon usage bias is mainly based on genomic data. However, this method is greatly limited, because the genome sequences of higher organisms are still not available up to now. In this study, we found that we could obtain the same optimal codons of Ganoderma lucidum (Curtis: Fr.) P. Karst based on its whole genomic data or large-scale transcriptomic data from its liquid-cultured hyphae, primordium and fruiting body, separately. This result indicated the feasibility to understand the codon usage bias based on the large-scale transcriptomic data. By calculating the proportion of rare codons of Escherichia coli and Saccharomyces cerevisiae in 26 terpene synthases (TS) of G. lucidum, we found that the rare codons of S. cerevisiae have a higher proportion in TS genes, while the rare codons of E. coli have relatively lower, suggesting that the TS genes of G. lucidum are possibly more difficult to be expressed in S. cerevisiae than in E. coli. Chemical synthesis of TS genes according to the yeast optimal codons will be an effective way to solve the problem on the mismatch of gene codon bias between the foreign genes and the host strain.
Codon ; Escherichia coli ; Genome, Fungal ; Reishi ; genetics ; Saccharomyces cerevisiae ; Transcriptome

BACKGROUND:Stem cels are induced to differentiate into endothelial-like cels that can be used for the treatment of diabetic lower extremity vascular disease. However, it is unclear whether these endothelial-like cels can completely replace endothelial cels to improve vascular disease and what are the differences between endothelial-like cels and endothelial cels. OBJECTIVE:To explore the differences and similarities between endothelial-like cels and human umbilical vein endothelial cels in the aspects of morphology, function, and viability. METHODS:Umbilical cord mesenchymal stem cels and umbilical vein endothelial cels were isolated, cultured and identified using flow cytometry and immunohistochemical method. Isolated umbilical cord mesenchymal stem cels were induced in DMEM-LG/F12 containing 10 μg/L vascular endothelial growth factor, 10 μg/L basic
fibroblast growth factor and 2% fetal bovine serum to differentiate into endothelial-like cels folowed by immunohistochemical identification. To compare endothelial-like cels with human umbilical vein endothelial cels, cel migration detection, active substance measurement and three-dimensional angiogenesis test were performed. RESULTS AND CONCLUSION:Isolated umbilical cord mesenchymal stem cels strongly expressed the surface markers of mesenchymal stem cels, and human umbilical vein endothelial cels strongly expressed CD31 and VWF. After induction, the umbilical cord mesenchymal stem cels were identified to highly express CD31 and VWF. Through cel migration, active substance and three-dimensional angiogenesis tests, endothelial-like cels were similar to endothelial cels in the function and activity, and superior to endothelial cels. Cite this article:Hao XJ, Hao HY, Zhu MJ, Yuan Z, Li WW, Chen J, Zhu LY. Endothelial-like cels versus human umbilical vein endothelial cels. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):83-88.

To study the expression of tankyrase (a positive regulator of telomerase activity) in malignant hematopoietic cells and its relation with telomerase activity, the method of realtime quantitative PCR with fluorescence probe hybridization were used to measure expression of tankyrase and hTERT in myeloid leukemia cell lines K562, HL-60, U937, NB4, THP-1, HEL, Dami and T lymphocytic leukemia cell lines 6T-CEM, Jurkat and B-cell lymphoma cell line Raji. CD3(+), CD19(+) and CD33(+) cells separated from normal human mobilized peripheral blood by immunomagnetic bead system and 10 mononuclear cell samples separated from bone marrow of normal individuals were served as normal controls. The results indicated that the expression of tankyrase in malignant hematopoietic cell lines was significantly higher than that in normal controls (U = 19, P < 0.01). Its expression in myeloid leukemia cell lines is higher than in normal CD33(+) cells, the expression in T lymphocytic leukemia and B-cell lymphoma cell lines is higher than in CD3(+) and CD19(+) cells respectively. Its expression in myeloid malignant hematopoietic cell lines is significantly lower than in lymphocytic ones (0.0032 +/- 0.0010 vs. 0.012 +/- 0.0016, F = 23, P < 0.01). The expression of tankyrase correlated positively with hTERT (Spearman correlation coefficient is 0.395, P < 0.05). It is concluded that tankyrase is overexpressed in malignant hematopoietic cell lines, that may be one of the causes of high-produced telomerase activity in malignant hematopoietic diseases.
DNA-Binding Proteins ; HL-60 Cells ; Humans ; K562 Cells ; Leukemia ; enzymology ; Polymerase Chain Reaction ; Tankyrases ; genetics ; Telomerase ; genetics ; U937 Cells

OBJECTIVETo detect mutations of human telomeric repeat binding factor 1 (TERF1) gene in 11 malignant hematopoietic cell lines, which have positive telomerase activity, and evaluate the significance of the mutations.

METHODSGenome structure of TERF1 was predicted by using biology information program, and verified by PCR and sequencing. Telomerase activity was detected by telomeric repeat amplification (TRAP)-ELISA. PCR and sequencing were used to detect mutation of each exon of TERF1 in 11 cell lines, including myelogenous leukemia cell lines K562, HL-60, U-937, NB4, THP-1, HEL and Dami; lymphoblastic leukemia cell lines 6T-CEM, Jurkat and Raji and MDS-RAEB cell line MUTZ-1. Five DNA samples from healthy volunteers were detected as normal controls.

RESULTSTERF1 gene has 10 exons and spans 38.6 kb. All the 11 cell lines showed positive telomerase activity. No mutation was found in all exons of TERF1 in the 11 cell lines. However, 4 variants were found in intron1, 2 and 8 near exon1, exon2 and exon9, respectively. The variants in MUTZ-1 was different from those in leukemia cell lines; but no difference was found between the variants in myelogenous and lymphoblastic leukemia cell lines.

CONCLUSIONTERF1 mutation is probably not among the main factors of the gene dysfunction in malignant hematopoietic diseases.

Base Sequence ; Cell Line, Tumor ; DNA Mutational Analysis ; Enzyme-Linked Immunosorbent Assay ; Exons ; genetics ; HL-60 Cells ; Hematologic Neoplasms ; genetics ; metabolism ; pathology ; Humans ; Jurkat Cells ; K562 Cells ; Mutation ; Polymerase Chain Reaction ; Telomerase ; metabolism ; Telomere-Binding Proteins ; genetics ; metabolism ; U937 Cells

Objective To evaluate a normative and systematic training mode for teachers of community general practice.Methods A total of 16 general practice teachers,who came from Fenglin community health service centre in Shanghai,participated three-stage training mode during March 2011 to February 2012.The clinical competence evaluation and questionnaire survey were conducted before and after training.Fifty trainees trained by the teachers also received clinical competence evaluation.Results After the three-stage training,the test score in medical knowledge,physical examination and clinical skills of the teachers did not change significantly (P =0.794,0.674 and 0.326).The self assessment questionnaire survey of general practice teachers indicated a significant increase,especially in practice capability (t =-2.840,P =0.015) and overall quality (t =-3.017,P =0.011).After training by the teachers,the medical knowledge (t =-9.200,P =0.000),physical examination (t =-9.947,P =0.000) and clinical skills (t =-14.828,P =0.000) of 50 trainees increased markedly.Conclusions Differed from conventional training courses,the three-stage training enhances teaching ability and overall quality of community general practice teachers,and provides a effective training model.

The research on clinical efficacy evaluation of traditional Chinese medicine (TCM) is a hot and difficult issue in the field of TCM. However, the overall level of current related research also needs to be improved. There are still a lot of academic controversies in the field of TCM clinical efficacy evaluation. The author believed that before conducting research, it is necessary to sort out core issues of TCM clinical efficacy evaluation, which in-clude the concept of TCM clinical efficacy and its evaluation. In recent years, Chinese scholars have conducted a series of studies on these issues and have some achievements. To think and answer these questions will play an important role for the ultimate resolution of problem of TCM clinical efficacy evaluation.

Objective To investigate the sequences of the 3' end of genomes from enterovirus 71 isolated from pediatric patients with different symptoms in Beijing during 2008-2009.Methods Clinical specimens were collected from pediatric patients suffering from hand,foot and mouth disease(HFMD)and/or with neurological complications who visited the affiliated Children's Hospital during the epidemic seasons of 2008 and 2009 in Beijing.The samples were inoculated into the Vero cell line,and the virus isolates were further identified by nested reverse transcription-nested PCR(RT-nPCR)assay using universal enterovirus primers,type specific primers for EV71 and CA16.The 3' end of genomes(including 3D and 3' UTR regions)from 10 EV71 strains derived from various clinical presentations were amplified and sequenced.Results Analysis of the nucleotide sequences of the amplified fragments showed that the 3' end of genomes of 10 EV71 isolates include the 3D region of 1386 nucleotides(nt)encoding the 3D polymerase(3Dpol,462 amino acids),the terminator codon TGA and 3' UTR of 81 nt.Nucleotide sequence identities among 3D regions from these 10 EV71 isolates were in the range of 95.8%-99.6%,while the nucleotide sequence identities for 3' UTR were 96.3%-100%.The majority of nucleotide changes were located at the third codon positions which caused silent mutations,thus the amino acid sequence changes of 3Dpol among those 10 EV71isolates were scanty.The residues 140 and 263 which were R and 1 were substituted by K and V,respectively in 3 of 4 neurovirulent strains,whereas only 1 of 6 strains from mild cases had these 2 amino acid changes.The sequences of the 3D and 3' UTR regions of 10 EV71 isolates were compared to the representative strains of known genotypes from the GenBank.The nucleotide and amino acid sequences of 10 EV71 isolates in the 3D region exhibited highest homology to the subgenotype C4 of EV71(92.7%-94.2%and 96.8%-97.6%.respectively).However,3' UTR of 10 EV71 isolates shared the highest nucleotide identity with CA16/G10(88.9%-91.4%).The phylogenetic analysis based on the 3D regions demonstrated that 10 EV71 isolates had the closest genetic relationship with the representative isolate of CA sub-genotype of EV71 and shared more homology with CA16/G10 than other known genotypes of EV71.Conclusion Genetic analysis of the 3' end of genomes from 10 EV71 strains indicated that the 3' end of genome may play a role in the evolution of EV71.