Objective To investigate the morphological changes of bone marrow megakaryocytes in patients with bacterial and fungal infection.Methods Totally 76 patients with microorganism infection from the Second Affiliated Hospital,Zhejiang University School of Medicine from January 2008 to August 2009 were enrolled,including 56 bacteria infected patients and 20 fungal infected patients.All patients received bone marrow examinations,and were positive in microorganism culture.Thirty subjects without infection,hematological disease and other severe diseases were randomly selected as controls.The number and function of megakaryocytes were examined retrospectively, and the size, nuclear lobulation, and vacuolar degeneration of megakaryocytes were quantitative analyzed and compared among the groups.Results The size,nuclear lobulation,vacuolar degeneration,and Yat nuclear of megakaryocytes in bacterial infected group were 2.20 ±0.21,2.11 ±0.23,0.51 ±0.11 and 0.74 ±0.11 respectively,those in fungal infected group were 2.21 ±0.16,2.10 ±0.19,0.52 ±0.10 and 0.79 ±0.10 respectively;while those in control group were 1.40 ±0.10,1.36 ±0.12,0.28 ±0.06 and 0.54 ±0.09 respectively.The differences between bacterial infected group and control were of statistical significance(t values were 14.52,12.19,9.33 and 6.61 respectively,P ＜ 0.05),and the differences between fungal infected group and control were of statistical significance(t values were 16.27,12.34,7.85 and 6.49 respectively,P ＜ 0.05).The size,nuclear lobulation,and vacuoles of megakaryocytes in gram-negative(G-)bacteria group were 2.29 ±0.20,2.22 ±0.26 and 0.57 ±0.10,while those in the gram-positive(G+)bacteria group were 2.13 ±0.20,2.04 ±0.18 and 0.46 ±0.09,and the differences were also significant(t values were 2.07,3.03and 3.56 respectively,P ＜ 0.05).The production of platelet by megakaryocytes in bacterial infected group,in fungal infected and the control were 31.4 ±7.6,32.4 ±6.4 and 41.3 ±5.5,and the differences between bacterial infected group and control,fungal infected group and control were significant(t values were 4.78and 3.98 respectively,P ＜ 0.05).The production of platelet in G-bacteria group was 28.0 ± 6.7,while that in G + bacteria group was 34.4 ± 7.2,and the difference was also of statistical significance(t = 2.41,P ＜0.05). Conclusion Bacterial infected patients have increased megakaryocytes cell body,nuclear lobulation,obvious vacuolar degeneration,Yat nuclear and decreased platelet production function,which are more significant in G- bacteria infected group.

Four salts of ticagrelor, ticagrelor-3,5-dinitrobenzoic acid, ticagrelor-pyrazinamide, ticagrelor-D-proline and ticagrelor-L-proline were prepared by solvent suspension and liquid-assisted grinding to improve the solubility of ticagrelor. The compounds were characterized by powder X-ray diffraction, Fourier transform infrared spectroscopy, differential scanning calorimetry, nuclear magnetic resonance spectroscopy, elemental analysis, and the intermolecular salt-bonding forces were analyzed. The equilibrium solubility of salts and pure drug in hydrochloride buffer pH 1.2 and phosphate buffer pH 6.8 were measured by high-performance liquid chromatography. Ticagrelor was salted with 3,5-dinitrobenzoic acid, pyrazinamide, D-proline, L-proline all in a stoichiometric ratio of 1∶1; with the exception of ticagrelor-D-proline, the solubility of the other three salts provided significantly improved solubility in hydrochloride buffer pH 1.2, and the equilibrium solubility of ticagrelor-3,5-dinitrobenzoic acid was increased by approximately 1.7 folds as compared to pure drug. Salt-forming technology is convenient and can improve the solubility of ticagrelor.

In plant, evocation of secondary metabolism is associated with complex biochemical and molecular events that are regulated by developmental and environmental factors. In order to get more information about Taxol biosynthesis, comparison of mRNA populations from Taxus chinensis cells during Taxol-synthesis phase and those during non-Taxol-synthesis phase were performed by mRNA differential display. The results suggested that genes specifically expressed in the Taxol-synthesis phase might be involved in Taxol biosynthesis.
Gene Expression Regulation, Developmental ; Gene Expression Regulation, Plant ; Paclitaxel ; biosynthesis ; RNA, Messenger ; genetics ; metabolism ; Taxus ; genetics ; growth & development ; metabolism ; Time Factors

BACKGROUNDSome studies indicated that cases of idiopathic oculomotor nerve palsy can be explained by vascular compression of the oculomotor nerve. Vascular contact with or compression to the cisternal segment of the oculomotor nerve has been reported frequently in asymptomatic individuals. In this study, we retrospectively analyzed the relationship between the oculomotor nerve's cisternal segment and adjacent arteries in asymptomatic patients and the prevalence of this occurrence via magnetic resonance imaging (MRI).

METHODMRI of bilateral oculomotor nerves in 93 asymptomatic patients were reviewed. The oculomotor nerve-artery relationship was evaluated and classified from levels 1 to 3, representing the degrees of contact on oblique transverse and oblique sagittal reconstructed MRI. Prevalence of the nerve-artery relationship at each level was described. The correlation between the nerve-vessel relationship (levels) and the age was analyzed by Spearman's rank correlation analysis.

RESULTSCisternal segment of the oculomotor nerve did not have contact with any artery (level 1) in 27.4% (51/186) nerves. One hundred nerves made contact with at least one artery (level 2), but their shapes or configurations were not changed; 35 nerves (18.8%) were displaced or distorted due to artery compression (level 3). The posterior cerebral artery had the greatest incidence of making contact with or compressing the cisternal segment of the oculomotor nerve (58.1%). No significant correlation between nerve-vessel relationship (levels) and the age was found in this study.

CONCLUSIONSWhether oculomotor nerve contact with or compression by one or more arteries is of high prevalence in asymptomatic individuals as evidenced by MRI examination. There is no correlation with individual age. Discretion should be used when making an etiological diagnosis of vascular compression for patients with oculomotor nerve palsy. Further investigation of other causes is warranted.

Adolescent ; Adult ; Age Factors ; Aged ; Child ; Child, Preschool ; Female ; Humans ; Magnetic Resonance Imaging ; methods ; Male ; Middle Aged ; Nerve Compression Syndromes ; complications ; pathology ; Oculomotor Nerve ; pathology ; Oculomotor Nerve Diseases ; etiology ; pathology ; Young Adult

BACKGROUNDAn early identification of the composition of arterial thrombus may have diagnostic, therapeutic, and prognostic implications. The variation of magnetic resonance (MR) signal intensity between white and red thrombi, especially in the susceptibility sensitive MR sequence, remains unknown. Our research was to evaluate the feasibility of MRI in differentiating of white and red thrombi with a phantom study.

METHODSA total of 12 red and 12 white thrombi were prepared with the venous blood. Examination of the phantom was completed using a 3.0T MR unit, including fluid attenuated inversion recovery (FLAIR) T1, T2-weighted imaging (T2WI), FLAIR T2, T2 gradient echo (T2 GRE) imaging, and susceptibility weighted angiography sequences (SWAN). MR signal intensity patterns of the thrombi were objectively classified as hyperintensity, isointensity and hypointensity, compared with the background agar. The volume of thrombus was calculated and correlated with its signal intensity.

RESULTSFor white thrombi, 11/12 clots showed hyperintensity and 1/12 showed isointensity in FLAIR T1 images. In T2WI, 6/12 clots showed hyperintensity, 3/12 isointensity, and 3/12 hypointensity. In FLAIR T2, 8/12 clots showed hyperintensity and 4/12 showed isointensity. In T2 GRE, 3/12 clots showed hyperintensity and the remaining 9/12 clots showed isointensity. In SWAN, 5/12 clots demonstrated hyperintensity and 7/12 isointensity. For the red thrombus, 12/12 clots demonstrated hyperintensity in FLAIR T1, T2WI, and FLAIR T2 sequences. In T2 GRE and SWAN sequences, 3/12 clots displayed hypointensity and the remaining 9/12 clots showed slight hyperintensity. Thrombi with hypointensity displayed in T2 GRE and SWAN sequences were significantly larger than those with hyperintensity.

CONCLUSIONSDifferentiation of white and red thrombi with conventional MR sequence is unreliable, because both kinds of thrombi do not possess unique signal intensity features in these sequences. Red thrombus may or may not show hypointensity in the susceptibility sensitive MR sequences, depending on its size and time course.

Humans ; Magnetic Resonance Imaging ; methods ; Phantoms, Imaging ; Thrombosis ; diagnosis ; pathology

We identified a novel gene ST13 from a subtractive cDNA library of normal intestinal mucosa in 1993, more studies showed that ST13 was a co-chaperone of Hsp70s. Recently we detected the ST13 gene expression in tumor tissue and adjacent normal tissue of the same colorectal cancer patient and investigated if the ST13 gene expression might have any prognostic value. Analysis was performed at molecular level by reverse transcription-PCR using real-time detection method. We measured two genes simultaneously, ST13 as the target gene and glyceraldehydes-3-phosphate dehydrogenase as a reference gene, in primary colorectal tumor specimens and tumor-adjacent normal mucosa specimens from 50 colorectal cancer patients. The expression levels of the ST13 gene were significantly decreased in primary tumors compared with adjacent mucosa (P<0.05). But there were no significant differences in the expression of ST13 as compared with different Dukes' stage, tumor differentiation grade, invasion depth, lymph node metastasis and disease-specific survival.
Biomarkers, Tumor ; metabolism ; Carrier Proteins ; metabolism ; China ; epidemiology ; Colorectal Neoplasms ; diagnosis ; metabolism ; mortality ; Disease-Free Survival ; Female ; Gene Expression Profiling ; Humans ; Male ; Prevalence ; Prognosis ; Risk Assessment ; methods ; Risk Factors ; Survival Analysis ; Survival Rate ; Tumor Suppressor Proteins ; metabolism

Australia ; Environmental Monitoring ; Humans ; Occupational Health ; Safety Management ; methods ; organization & administration

Design space approach is applied in this study to enhance the robustness of first ethanol precipitation process of Codonopsis Radix (Dangshen) by optimizing parameters. Total flavonoid recovery, dry matter removal, and pigment removal were defined as the process critical quality attributes (CQAs). Plackett-Burman designed experiments were carried out to find the critical process parameters (CPPs). Dry matter content of concentrated extract (DMCE), mass ratio of ethanol to concentrated extract (E/C ratio) and concentration of ethanol (CEA) were identified as the CPPs. Box-Behnken designed experiments were performed to establish the quantitative models between CPPs and CQAs. Probability based design space was obtained and verified using Monte-Carlo simulation method. According to the verification results, the robustness of first ethanol precipitation process of Dangshen can be guaranteed by operating within the design space parameters. Recommended normal operation space are as follows: dry matter content of concentrated extract of 45.0% - 48.0%, E/C ratio of 2.48-2.80 g x g(-1), and the concentration of ethanol of 92.0% - 92.7%.
Chemical Precipitation ; Chemistry, Pharmaceutical ; methods ; Codonopsis ; chemistry ; Drugs, Chinese Herbal ; chemistry ; isolation & purification

BACKGROUNDCytosine deaminase (CD) converts 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU) in CD/5-FC gene therapy, 5-FU will be mostly converted into nontoxic beta-alanine without uracil phosphoribosyltransferase (UPRT). UPRT catalyzes the conversion of 5-FU to 5-fluorouridine monophosphate, which directly kills CD::UPRT-expressing cells and surrounding cells via the bystander effect. But the pharmacokinetics and the bystander effect of CD::UPRT/5-FC has not been verified in vivo and in vitro. Before the CD::UPRT/5-FC bi-gene therapy system is used in clinical trial, it is essential to monitor the transgene expression and function in vivo. Thus, we developed a preclinical tumor model to investigate the feasibility of using (19)F-magnetic resonance spectroscopy ((19)F-MRS) and optical imaging to measure non-invasive CD and UPRT expression and its bystander effect.

METHODSC6 and C6-CD::UPRT cells were cultured with 5-FC. The medium, cells and their mixture were analyzed by (19)F-MRS. Rats with intracranial xenografted encephalic C6-CD::UPRT glioma were injected intraperitoneally with 5-FC and their (19)F-MRS spectra recorded. Then the pharmacokinetics of 5-FC was proved. Mixtures of C6 and C6-CD::UPRT cells at different ratios were cultured with 5-FC and the cytotoxic efficacy and survival rate of cells recorded. To determine the mechanism of the bystander effect, the culture media from cell comprising 25% and 75% C6-CD::UPRT cells were examined by (19)F-MRS. A comparative study of mean was performed using analysis of variance (ANOVA).

RESULTS(19)F-MRS on samples from C6-CD::UPRT cells cultured with 5-FC showed three broad resonance signals corresponding to 5-FC, 5-FU and fluorinated nucleotides (F-Nuctd). For the C6 mixture, only the 5-FC peak was detected. In vivo serial (19)F-MRS spectra showed a strong 5-FC peak and a weak 5-FU peak at 20 minutes after 5-FC injection. The 5-FU concentration reached a maximum at about 50 minutes. The F-Nuctd signal appeared after about 1 hour, reached a maximum at around 160 minutes, and was detectable for several hours. At a 10% ratio of C6-CD::UPRT cells, the survival rate was (79.55 +/- 0.88)% (P < 0.01). As the C6-CD::UPRT ratio increased, the survival rate of the cells decreased. (19)F-MRS showed that the signals for 5-FU and F-Nuctd in the culture medium increased as the ratio of C6-CD::UPRT in the mixture increased.

CONCLUSIONS(19)F-MRS studies indicated that C6-CD::UPRT cells could effectively express CD and UPRT enzymes. The CD::UPRT/5-FC system showed an obvious bystander effect. This study demonstrated that CD::UPRT/5-FC gene therapy is suitable for 5-FC to F-Nuctd metabolism; and (19)F-MRS can monitor transferred CD::UPRT gene expression and catalysis of substrates noninvasively, dynamically and quantitatively.

Animals ; Antimetabolites ; pharmacokinetics ; therapeutic use ; Cell Line ; Cytosine Deaminase ; genetics ; physiology ; Flucytosine ; pharmacokinetics ; therapeutic use ; Genetic Therapy ; methods ; Glioma ; drug therapy ; therapy ; Humans ; Magnetic Resonance Imaging ; Male ; Pentosyltransferases ; genetics ; physiology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo cooperate with the study of HBV vector, hygromycin-resistant packaging cell line was developed that allows encapsidation of plasmids into HBV particles.

METHODSFree of packaging signal, HBV genome was inserted into plasmid pMEP4, which expresses the HBV structural proteins including core, pol and preS/S proteins. HepG2 cell lines were employed to transfect with the construct. Hygromycin selection was done at a concentration of 150 micrograms/ml in the culture medium. The hygromycin-resistant clones with the best expressions of HBsAg and HBcAg were theoretically considered as packaging cell line and propagated under the same conditions. It was infected with recombinant retrovirus vector and hen selected with G418 and hygromycin in the culture medium. The existence of recombinant HBV virion in the culture medium was examined by PCR.

RESULTSHygromycin-resistant HBV packaging cell line was generated, which harbored an HBV mutant whose packaging signal had been deleted. Expressions of HBsAg and HBcAg were detectable. Infected with recombinant retrovirus pRV-CP, the hygromycin-resistant packaging cell line was found to secrete mutant HBV particles and no wild-type HBV was detectable in the culture medium.

CONCLUSIONAfter the packaging signal was deleted and transfected into HepG2 cell lines, the partial HBV genome lost its ability to form wild-type HBV, but conserves cis-action providing structural proteins for the packaging of the replication-defective HBV.

Cell Line ; Drug Resistance, Viral ; Genetic Vectors ; Genome, Viral ; Hepatitis B virus ; drug effects ; genetics ; Humans ; Hygromycin B ; pharmacology ; Mutation ; Plasmids ; Retroviridae ; genetics ; Transfection ; Virus Assembly