Objective　To study the indication, contraindication and short-term clinical effects of stereotactic radiotherapy (X-knife) in lung cancer. Methods　A total of 56 cases (Treatment group) of lung cancer were treated with X-knife composing of the modern 3-D treatment planning system and the stereotactic body frame, and SL-75 accelerator. And another 53 cases only treated with routine external radiation served as the control. In treatment group, 42 cases received 50 Gy radiation, at 2 Gy per day, 5 d per week in the upper mediastinum, hilus of lung, below the primary focus first and then underwent X-knife for the primary focuses, and other 14 recurrent and metastatic cases received irradiation of 8-12 Gy per day, 5 d by X-knife alone. Results　The actually reexamination rate for 1, 3, 6, and 12 months after X-knife treatment in 78 focuses from 56 cases was 79.5% (62/78), 89.8% (70/78), 87.2% (68/78) and 78.2% (61/78) respectively. The focuses reappeared in 3 months after the treatment and the growth rates of focus in 3, 6, and 12 month after the treatment were 2.6% (2/78), 5.1% and 9% (7/78) respectively. There were 2 focuses received a second treatment with X-knife because no change after the first one. Both the 1-and 2-year survival rates of 36 cases of the primary lung cancer in the treatment group were higher than that of control group (89.1%, 53.9% and 77.4%, 39.6%). Conclusion　The stereotactic radiotherapy is of applicable and prospective in the treatment of lung cancer. The indications include: ①As a boost dose in case when primary focus is less than 5 cm after the first external radiotherapy may reduce the exposure of lung tissue to x-ray and the occurrence of radiation pneumonia; ②Treatment for the recurrence after radiotherapy and operation; ③Radical treatment for the metastatic focus less than 5 cm. Attention must be paid to when multiple treatment is carried on the volume of the focus, the Karnofsky scores and the general condition when stereotactic radiotherapy combined with X-knife for lung cancer.

The effects of different physical and chemical factors on hairy root growth and flavonoids production were studied in suspension culture of Saussurea medusa hairy root in 1/2 MS medium. The results showed that the following culture conditions, nitrogen concentratiaon (involved NH4+ and NO3-), 30 mmol/L; the ratio of ammonium to nitrate, 5:25; the combination of 2% sucrose and 3% glucose; 0.5 mg/L GA3; 0.5 mg/L IBA; initial pH 5.8; light cycle, 18 h/d (3500lx); temperature, 24 degrees C; shaker revolutions per minute, 100 r/min, were favourable to hairy root growth and flavonoids production. Under the above culture conditions, up to 12.8 g/L (DW) of hairy root and 1922 mg/L of flavonoids were obtained after 21 days of culture. The content of total flavonoids in hairy root was 15%, which was about 25 times as that in the wild plantlet.
Culture Media ; Flavonoids ; biosynthesis ; Plant Roots ; growth & development ; metabolism ; Saussurea ; growth & development ; metabolism ; Temperature ; Tissue Culture Techniques

Animals ; Bone Marrow Cells ; cytology ; Brain ; cytology ; physiopathology ; Brain Injuries ; physiopathology ; therapy ; Cell Movement ; Cells, Cultured ; Cisterna Magna ; Immunohistochemistry ; Rats ; Rats, Sprague-Dawley ; Stromal Cells ; transplantation

To establish a MacELISA method for the detection of IgM antibodies against Chikungunya virus (CHIKV), we prepared virus like particle (VLP) antigens of CHIKV using the whole structural protein C-E3-E2-6K-E1 encoding gene with a baculovirus expression system in Sf9 insect cells. The VLPs were purified and used to immunize Kunming mice. Then, polyclonal antibodies were purified from the samples of ascites with a protein G HiTrap SP column and labeled with horseradish peroxidase. A MacELISA method for the detection of IgM antibodies against CHIKV was assembled with goat anti-human IgM antibody, VLP antigens and an enzyme-labeled polyclonal antibody. The results were evaluated with a serum panel containing serum samples from laboratory-confirmed CHIK, HFRS patients, healthy donors, and commercially available CHIKV IgM as a quality control. It was shown that the MacELISA had a specificity of 99% (99/100), the coefficients of variation (CoV) within a plate were <10%, and the CoV of different ELISA plates in terms of the plate variation coefficient was <15%. A comparative analysis was performed to compare the current method against a commercial CHIKV IgM antibody detection kit for IIFA-IgM. The detection limit of MacELISA was significantly lower than that of the IIFA-IgM commercial kit (P< 0.0001). Here, we demonstrate that the VLP-based MacELISA is a promising tool for the early diagnosis and epidemiological investigation of CHIKV infection, with a high level of sensitivity and specificity for the detection of IgM antibodies against CHIKV.
Animals ; Antibodies, Viral ; blood ; Chikungunya Fever ; blood ; diagnosis ; virology ; Chikungunya virus ; immunology ; isolation & purification ; Enzyme-Linked Immunosorbent Assay ; methods ; Humans ; Immunoglobulin M ; blood ; Mice

OBJECTIVETo evaluate the feasibility and utility of an ex vivo sentinel lymph node (SLN) identification and ultrastaging for colorectal cancer (CRC).

METHODSCRC patients undergoing resection of a primary colorectal cancer were considered for inclusion. Following resection, SLN identification was performed. The SLN was dissected from the mesentery and submitted separately for pathologic analysis. All lymph nodes were stained with HE. Blue lymph nodes, when negative by routine HE staining, were further analyzed.

RESULTSA total of 62 tumors from 60 patients with colorectal cancer were studied. 95.2% (59/62) specimens was successfully identified. In these 59 specimens, a total of 1114 (18.9 per specimens) lymph nodes were examined; of these, 157 (14.9%) were designated as SLNs. The number of blue-stained lymph nodes removed ranged from 1 to 9, with a mean of 2.7 blue nodes identified. The sensitivity of a blue-stained lymph node identifying metastatic disease was 39.1%. The false-negative was 23.7%. In 4 specimens micrometastases were detected only by immunohistochemistry with cytokeratin.

CONCLUSIONSEx vivo sentinel lymph nodes mapping in colorectal cancer is feasible and can identify the SLNs with a very high success rate. Ex vivo SLN mapping improves pathologic staging of patients with CRC. The SLN evaluation should not replace attempts to harvest large number of nodes for standard processing. SLN mapping can help improving the number of nodes for pathological examination.

Adult ; Aged ; Aged, 80 and over ; Colorectal Neoplasms ; pathology ; Female ; Humans ; Immunohistochemistry ; Keratins ; analysis ; Lymph Nodes ; pathology ; Lymphatic Metastasis ; Male ; Middle Aged ; Rosaniline Dyes ; Sentinel Lymph Node Biopsy ; methods

Objective To detect the mutations of ATP2C1 gene in patients with Hailey-Hailey dis- ease (HHD).Methods PCR and direct sequencing were performed in 17 patients and 120 healthy controls to screen the mutations in the exons of ATP2C1 gene.Results Eight mutations were identified in nine probands, including three deletion mutations (nt1464-1487 del/nt1462-1485del,1523delAT,2375delTTGT),three splice site mutations (360—2A→G,1415—2A→T,2243+2T→C) and two missence mutations (C920T and G1942T).None of the above mutations was found in the controls.Conclusion Eight specific novel mutations were identified in nine probands of HHD,which could be causative factors of the disease.

OBJECTIVETo investigate whether mammalian target of rapamycin (mTOR) kinase was abnormally activated in maldeveloped balloon cells and dysmorphic neurons of focal cortical dysplasia (FCD) with refractory epilepsy.

METHODSA total of 12 archival cases of FCD typeIIwith medically intractable epilepsy treated between 2008 and 2010 were retrieved. Perilesional brain tissue was used as control specimens (n = 8). The expression of phosphorylated p-AKT (Ser473), p-mTOR (Ser2448) and p-P70S6K (Thr389) was investigated by imunocytochemistry.

RESULTSThe expression of p-AKT (Ser473), p-mTOR (Ser2448) and p-P70S6K (Thr389) was found in meldeveloped balloon cells and dysmorphic neurons of FCD. A weak stain in a small amount of pyramid neurons was also found in the control group.

CONCLUSIONAbnormal activation of mTOR in maldeveloped balloon cells and dysmorphic neurons of FCD may be a key molecular mechanism underlying the histological changes and repeated seizures.

Adolescent ; Adult ; Brain Diseases ; metabolism ; pathology ; Child, Preschool ; Epilepsy ; metabolism ; pathology ; Female ; Glial Fibrillary Acidic Protein ; metabolism ; Humans ; Immunohistochemistry ; Male ; Malformations of Cortical Development ; metabolism ; pathology ; Malformations of Cortical Development, Group I ; Nestin ; metabolism ; Neurons ; metabolism ; Phosphorylation ; Proto-Oncogene Proteins c-akt ; metabolism ; Ribosomal Protein S6 Kinases, 70-kDa ; metabolism ; TOR Serine-Threonine Kinases ; metabolism ; Young Adult

Hairy root clones of Saussurea involucrata transformed with Agrobacterium rhizogenes strains R1601, R1000, and LBA9402 were established to investigate the flavonoid production. Opine synthesis and PCR analysis confirmed the integration of the T-DNA fragment of Ri plasmid from A. rhizogenes strain R1601 into the transformed root genome. The frequency of hairy root formation from root segments, which were pre-cultured 2 days in N6 solid medium without plant growth regulators, amounted to 100% following infection with R1601 strain of A. rhizogenes. The transformed roots were kept in hormone-free N6 liquid medium in the dark at 25 degrees C, 110r/min and routinely subcultured every 20 - 24 days. One hairy root clone, which grew vigorously with lateral branches, was periodically examined for the ability to produce flavonoid. The maximum of biomass and flavonoid yield achieved 66.7 g/L (fresh weight) and 102.3mg/g dry weight after incubation 20 days. The calli were induced from the hairy root culture in the presence of 0.5mg/L IBA and intact plantlets were regenerated from these calli. The regeneration plantlets from hairy roots, in which the flavonoid content were 53% in that of untransformed plants, weren't different in growth and morphology of the untransformed plantlets. Therefore plant regeneration from hairy roots may be also a means for producing transformed S. involucrata plants. Hairy root cultures of S. involucrata clearly showed higher flavonoid contents compared to the wild plant or the regeneration seedlings. As the wild S. involucrata grows only in special regions with peculiar climate, and cultivation of this species in a normal climate has been unsuccessful so far. The success in obtaining a method for high production of flavonoid might very well be one of the solutions for this problem in the future.
Culture Techniques ; Flavonoids ; biosynthesis ; Plant Roots ; growth & development ; Rhizobium ; physiology ; Saussurea ; growth & development

To study the effect of micro (mi)RNA on cellular proliferation induced by hepatitis B x protein, HBx, in human liver cells and to investigate the underlying molecular mechanism of this cancer-related effect. The human L02 hepatocyte cell line was stably transfected with HBx (L02/HBx) or an HBx mutant (L02/HBx-d382) that induces higher levels of cellular proliferation. The differential miRNA expression profiles were determined by microarray analysis and confirmed by real-time PCR. Two miRNAs, miR-338-3p and miR-551b, that were found to be significantly down-regulated in the L02/HBx-d382 cells were selected for further study and transfected individually into cells using the lipofectamine procedure. The cell survival rate was analyzed by MTT assay, and cell cycles were assessed by flow cytometry. Expressions of cyclinD1, cyclinG1, and E2F1 were assessed by real-time PCR and Western blotting. Compared with the microarray miRNA profile of L02/pcDNA3.0 cells, six miRNAs were up-regulated and five miRNAs were down-regulated in the L02/HBx-d382 cells, while four miRNAs were up-regulated and 12 were down-regulated in the L02/HBx cells. The microarray results were consistent with real-time PCR results. Transfection of miR-338-3p and miR-551b significantly inhibited the cell survival rates (P less than 0.001) and induced G0/G1 phase cycle arrest. According to MTT results: for L02/HBx-d382 cells, compared with lipofectamine or non-transfected (NC) controls, the t value of miR-338-3p was 10.402, 9.133 and the t value of miR-551b was 8.763, 7.403; for L02/HBx cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 9.105, 8.074 and the t value of miR-551b was 7.673, 7.52. According to flow cytometry results: for L02/HBx-d382 cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 12.173, 11.107 and the t value of miR-551b was 15.364, 13.377; for L02/HBx cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 15.416, 13.378, and the t value of miR-551b was 13.276, 13.109. The protein levels of cyclinD1, cyclinG1, and E2F1 were significantly reduced by both miR-338-3p and miR-551b ( P less than 0.001). For L02/HBx-d382 cells, compared with lipofectamine or NC controls: E2F1 had t = 11.132, 10.031 and 12.017, 10.973, respectively; cyclinD1 had t = 15.654, 15.013 and 15.447, 14.733, respectively; cyclinG1 had t = 8.017, 7.661 and 7.402, 7.417, respectively. For L02/HBx cells, compared with lipofectamine or NC controls: E2F1 had t = 14.244, 13.331 and 15.022, 14.468, respectively; cyclinD1 had t = 8.695, 8.137 and 7.877, 7.503, respectively; cyclinG1 had t = 7.73, 7.471 and 7.596, 7.41, respectively. In contrast, the mRNA levels for E2F1, cyclinD1, and cylcinG1 showed no significant differences between the miRNA transfected cells and controls. Wild-type HBx and the high proliferation-inducing mutant HBx can influence the miRNA expression profile of L02 cells. HBx down-regulates miR-338-3p and miR-551b in L02 cells, and the high proliferation-inducing mutant has a more robust effect. The mechanism of miR-338-3p- or miR-551b-mediated cell growth inhibition appears to be related to the direct modulation of cyclinD1, cyclinG1, and E2F1.
Blotting, Western ; Carcinoma, Hepatocellular ; genetics ; metabolism ; pathology ; Cell Cycle ; Cell Line ; Cell Proliferation ; Cyclins ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Genes, Viral ; Hepatitis B virus ; genetics ; metabolism ; Hepatocytes ; metabolism ; pathology ; Humans ; Liver Neoplasms ; genetics ; metabolism ; pathology ; MicroRNAs ; genetics ; metabolism ; Mutation ; Oligonucleotide Array Sequence Analysis ; RNA, Messenger ; genetics ; Real-Time Polymerase Chain Reaction ; Trans-Activators ; genetics ; metabolism ; Transfection

The present study investigated the influence of the methyl jasmonate and salicylic acid elicitors on the formation of phenylethanoid glycosides (PeG) in the suspension cultures of Cistanche deserticola. The results showed that methyl jasmonate and salicylic acid enhanced greatly the accumulation of PeG and echinacoside (Echin), but their optimum elicitation dosage and addition time were different. The yields of PeG and Echin were significantly increased in the presence of 5 micromol/L methyl jasmonate on day 14 (up to 2.59-fold and 3.82-fold, respectively), whereas treated with 50 micromol/L salicylic acid on day 28, the maximum content of them were, respectively, 2.71 and 3.16-fold higher than the untreated cell cultures.
Acetates ; pharmacology ; Cell Culture Techniques ; Cistanche ; drug effects ; metabolism ; Culture Media ; Cyclopentanes ; pharmacology ; Glycosides ; biosynthesis ; Oxylipins ; pharmacology ; Phenylethyl Alcohol ; metabolism ; Salicylic Acid ; pharmacology