Colonic Neoplasms ; genetics ; pathology ; CpG Islands ; genetics ; DNA Methylation ; Humans ; Polymerase Chain Reaction ; methods ; Sequence Analysis, DNA ; methods

Objective To investigate the immunological pathogenesis of Kawasaki disease( KD)through examination of changes in the expression of suppressors of cytokine signaling 1(SOCS1)and SOCS3,helper T cells and CD4 + CD25 + regulatory T cells(CD4 + CD25 + Treg)in peripheral blood from children with acute KD. Methods Six-teen children[10 boys,6 girls,aged 1 - 2 years old,averaged(1. 6 ± 0. 3)years old]in the acute phase of KD(KD group),16 children[9 boys,7 girls,aged 1 - 3 years old,averaged(1. 5 ± 1. 1)years old]with pneumonia(pneumo-nia group)and 8 normal children[5 boys,3 girls,aged 1 - 5 years old,averaged(2. 0 ± 1. 1)years old]of the same age(normal control group)from the Affiliated Hospital of Qingdao University who were admitted from October 2012 to March 2013 were recruited. The mRNA levels of SOCS1 and SOCS3 in the T cells from peripheral blood were examined by way of reverse transcription - polymerase chain reaction(RT - PCR). Interferon - γ( IFN - γ),interleukin - 4 (IL - 4)and CD4 + CD25 + Treg were quantified by means of fluorescence activated cell sorting(FACS). Results The expressions of SOCS1 and SOCS3,the percentage of IL - 4 T cells observed in the peripheral blood of the pneumonia group were similar to the normal control group(P ﹥ 0. 05),but significantly decreased in the percentage of INF - γ and the level of CD4 + CD25 + Treg(t = 3. 71,12. 81,all P ﹤ 0. 05). Compared to the normal control group and the pneumo-nia group,the expressions of SOCS1 and SOCS3,the percentage of INF - γ and IL - 4 T cells decreased significantly in the peripheral blood of the KD group(t = 2. 27,4. 48,17. 64,2. 73,2. 74,1. 25,2. 36,2. 59,all P ﹤0. 05 ). On the other hand,the level of CD4 + CD25 + Treg in the peripheral blood of the KD group was markedly lower than that in the normal control group(t =7. 70,P ﹤0. 05),but similar to the pneumonia group(P ﹥0. 05). Conclusions The function of helper T cells is inhibited in acute KD. The CD4 + CD25 + Treg may be involved in the immunological pathogenesis of KD.

Objective: To obtain the transcriptome sequence database and differentially expressed genes of Asarum sieboldii, and to identify the genes related to the biosynthesis of methyleugenol, the main chemical compound in this species. Methods: Roots and leaves of the plants were chosen as experiment materials. The transcriptome sequence database was constructed by applying an Illumina Hiseq 4000 Sequencing Platform. Unigenes were assembled by BLAST similarity searches and annotated with GO and KEGG orthologs identifiers. Moreover, differentially expressed genes were analyzed. Results: 12.25 Gb database was obtained, among which 129 003 unigenes were annotated to be involved in 52 GO-terms and 363 metabolic pathways. After analysis, 439 differentially expressed genes were observed, the up-regulated genes account for 38.3% and the down-regulated genes account for 61.7%. In addition, 136 unigenes involved in phenylpropanoid biosynthesis in A. sieboldii, and 44 unigenes that were associated with biosynthesis of methyleugenol were identified. Conclusion: Unigenes explored in this study will significantly contribute to genome-wide research and analysis of this species.

With the application of monoclonal antibody technology more and more widely, its production technology is becoming more and more perfect. Small molecule monoclonal antibody technology is becoming a hot research topic for people. The application of traditional Chinese medicine small molecule monoclonal antibody technology has been more and more widely, the technology for effective Chinese medicine component knockout provide strong technical support. The preparation of monoclonal antibodies and small molecule knockout technology are reviewed in this paper. The preparation of several steps, such as: in the process of preparation of antigen, hapten carrier coupling, coupling ratio determination and identification of artificial antigen and establishment of animal immunization and hybridoma cell lines of monoclonal antibody, the large-scale preparation; small molecule monoclonal antibody on Immune in affinity chromatography column method is discussed in detail. The author believes that this technology will make the traditional Chinese medicine research on a higher level, and improve the level of internationalization of Chinese medicine research.
Animals ; Antibodies, Monoclonal ; chemistry ; genetics ; immunology ; Humans ; Hybridomas ; metabolism ; Immunologic Techniques ; methods ; trends

Background To optimize the culture method of human retinal microvascular endothelial cells is very important for the study of retinal angiogenesis disease.Human retinal microvascular endothelial cells have been successfully cultured in previous studies,but further improvement of the culture method to harvest higher yields and purity cells is still needed.Objective This study was to design a modified method to isolate and purify human retinal microvascular endothelial cells much easily and quickly,and to compare the expression of specific markers of vascular endothelial cells,factor Ⅷ and CD31/CD34 in the cells.Methods The use of human donor eyeballs was approved by the Ethic Commission of Zhongshan Ophthalmic Center of Sun Yat-sen University.The retina tissue from healthy donor was isolated and digested by the two-step digestion method with 2％ trypsin and 0.133％ collagenase Ⅳ.Human retinal microvascular endothelial cells were collected and plated in 60 mm dishes coated by 0.1％ fibronectin and cultured in endothelial cell-specialized medium supplemented with 10％ fetal bovine serum,0.3 mg/L β-endothelial cell growth factor (ECGF) and 100 ng/L sodium heparin.During the culturing,the growth situation of the cells was monitored by morphological observation,and immunohistochemical staining was performed to probe vascular endothelial cell-specific membrane protein CD31,CD34 and factor Ⅷ for identification of the cell purity.Results Human retinal microvascular endothelial cells were isolated successfully from the retina by the twostep digestion method.The primary cultured cells adhered to well 72 hours later and achieved confluence with the typical cobblestone appearance 9 to 10 days after cultured.The cells exhibited the blue nuclei and reddish cytoplasm by regular haematoxylin and eosin stain and showed a strong positive response for CD31,CD34 and factor Ⅷ by immunohistochemistry.The positive dye of CD31 and CD34 was lower than Ⅷ factor in both endothelial cells.Conclusions Modified culture method of human retinal microvascular endothelial cells can improve cell culture result and purify target cells.

In this paper, five different methods were carried out for DNA extraction directly from soil. The result shows that all five methods could generate DNA with more than 15 kb in size. They were subsequently used as templates for PCR amplification with success, using primers of the bacterial 16S rRNA gene and Shiva-1 gene encoding an antibacterial peptide. However, method 5 is more suitable for DNA extraction directly from a small amount of soil sample as it produced a good yield of DNA in high integrity with reliable reproducibility.

Selection and affinities of Cd binding peptides were assayed by phage random dodecapeptide library and affinity chromatography. Two Cd binding peptides were obtained, it was found that the affinities of Cu~ 2+ ,Co~ 2+ ,Zn~ 2+ ,Ni~ 2+ for Cd binding peptides were higher than that of Cd~ 2+ and Cr~ 2+ after detection of the amplified Cd binding peptides displayed phages to different heavy metal-charged resins; the detoxification of E.coli to Ni~ 2+ and Cd~ 2+ was enhanced when infected by Cd binding peptide displayed phages as compared with those of the control. The interaction of Cd binding peptide displayed phages with heavy metals charged resins was also observed under microscope. The work would be of great value and consequences for the study of interaction between heavy metals and proteins(peptides), as well as thedetoxification and bioremediation of heavy metals.

Background and purpose:Antracycline combined with paclitaxel is more widely applied in breast cancer as neoadjuvant chemotherapy.There are differences in applications of different paclita~els.In this research,the efficacy and toxicity of neoadjuvant chemotherapy with ET,ED regimen were compared for the patients with stageⅢbreast cancer.Methods:64 cases of stageⅢbreast cancer patients were divided in two groups.Before surgery,one group had received ET(EPI ivgtt 60 mg/m~2 d_1?21,PTX ivgtt 175 mg/m~2 d_2?21),the other group had received ED(EPI ivgtt 60 mg/m~2 d_2?21,DOC ivgtt 75 mg/m~2 d_2?21)neoadjuvant chemotherapy for three weeks. Curative effect and side effects were evaluated after 2-4 cycles.Results:Total effective rate was 87.5%.Effective rate in ED group was 92.9%,and effective rate in ET group was 83.5%.There was no significant difference(P=0.253).In pCR cases,8 cases in ED group achieved pathologically complete response compared to 3 cases in ET group(P=0.033). The number of patients in ED group(24 cases)hadⅣ-Ⅴgrade pathology evaluation after chemotherapy,it was higher than that in ET patients(21 cases).There was a significant difference(P=0.017).In both groups side effects including hair loss,nausea and vomiting,liver dysfunction were similar.Incidence rate of peripheral neurotoxicity in ET group was higher than that in ED group(P=0.002).Incidence rates of leukopenia,skin rash and phlebitis in ED group were higher than that in ET group.There was a significant difference between two groups in the leukopenia(P=0.034). Conclusions:For the patients with stageⅢbreast cancer patients,both two regimens could achieve better curative effect.ET and ED regimen have similar effect.But in ED regimen,the number ofpCR cases was obviously higher than in ET group.In both groups side effects were similar.There were significant differences in terms of leukopenia and peripheral neurotoxicity,but the side effects could be tolerated.

Aim To monitor the whole blood trough concentration of cyclosporine A (CsA) in renaltransplant recipients reciving triple therapy with mycophenolate mofetil, cyclosporine andprednisone and to establish an optimal therapeutic window of CsA. Methods Sampleswere measured by specific fluorescence polarization immunoassay. According to the timeafter operation and different therapy plan, the whole blood trough concentration of CsA ineach group was compared with that in control group.Results The optimal therapeuticwindow of CsA with MMF plan was 150~300 ?g? L-1 (less than one month after op-eration), 120~260?g?L-1 (1~

AIM To establish an HPLC method for the simultaneous content determination of four constituents in Kesuting Capsules (a fast cough suppressant,containing Ephedrae Herba,Papaveris Pericarpium,Platycodonis Radix,etc.).METHODS The analysis of trichloromethane-strong ammonia extract of Kesuting Capsules was performed on a 35 ℃ thermostatic Welch Ultimate(◎) XB-C18 column (4.6 mm ×250 mm,5 μm),with the mobile phase comprising of acetonitrile-0.01 mol/L potassium dihydrogen phosphate buffer flowing at 1.0 mL/min in an isocratic elution manner,and the detection wavelength was set at 210 nm.RESULTS Morphine,ephedrine hydrochloride,pseudoephedrine hydrochloride and codeine phosphate showed good linear relationships within the ranges of 8.054-67.12 μg/mL (r =0.999 5),22.31-185.9 μg/mL (r =0.999 9),21.26-177.2 μg/mL (r =0.999 7) and 1.212-10.09 μg/mL (r =0.999 7),whose average recoveries (n =9) were 100.9％ (RSD =2.0％),101.4％ (RSD =3.6％),105.3％ (RSD =1.2％) and 106.2％ (RSD =1.2％),respectively.CONCLUSION This simple method can be used for the rapid quality control of Kesuting Capsules.