Humans ; Infant ; Lymphohistiocytosis, Hemophagocytic ; complications ; Male ; Mucocutaneous Lymph Node Syndrome ; complications

Linoleic acid (C18∶2n-6) and ?-linolenic acid (C18∶3n-3) are found widely in fungi, plants and some lower animals. However, they can not be synthesized in mammals due to lack of △12 and ?-3 fatty acid desaturases. To enable endogenous production of essential fatty acids in mammalian cells, here the stable expression of a Caenorhabditis elegans gene FAT-2 encoding △12 fatty acid desaturase in CHO cells was reported. First, the FAT-2 coding sequence was cloned by RT-PCR. To facilitate high level synthesis of heterogeneous protein, the codon usage of the fatty acid desaturase genes was optimized according to the codon preference of mouse by site-directed mutagenesis, 2 synonymous mutations were introduced into FAT-2 gene by overlapping PCR. The codon-modified gene was finally fused to pBudCE4.1 vector (Invitrogen) under the control of CMV promoter. The expression vector pBudCE-FAT2 was linearized with NheⅠ, and then transfected CHO cells, the cells were under Zeocin selection for nine days and then propagated, then the transfected cells were harvested. The genome and total RNA were isolated for PCR and Norhern blot ananlysis. The results revealed that FAT-2 gene has been integrated into the genome of CHO cells and expressed properly. Fatty acids of total cellular lipids were analyzed by gas chromatography. The results indicate that the expression and function of △-12 fatty acid desaturase resulted in accumulation of linoleic acid. The levels of linoleic acid in transgenic cells were 2.4-fold higher than those in wild-type cells. The moderate linoleic acid in CHO cells was derived from cell culture media uptaken by cell membrane. The results demonstrate that a heterogenous desaturase gene can function well in mammalian cells and prove that transgenic approach is an efficient strategy for changing fatty acid composition of mammals.

Objective To investigate jugular vein morphological changes with three dimension phase contrast magnetic resonance venography(3D PC MRV),and to explore hemodynamic features using MR phase contrast cine (MR PC cine).Methods Sixty-five healthy volunteers performed 3D PC MRV and MR PC-cine sannings.MRV ranged from torcular herophili to brachiocephalic veins, and the raw data of PC-cine was acquired at cervical 2-3(C2-C3)level perpendicular to the Jugular veins(JVs)with the maximum encoding velocity of 50 cm/sec.Jugular vein showing absent or tip shape(cross-sectional area less than 12.5 mm2 )was considered abnormal,and flat,crescent,oval,round shapes were considered normal.Data of PC-cine was processed by computer to evaluate the hemodynamic features.Results Nine (13.85%)of 65 cases were abnormal that unilateral jugular vein showing needle-pointed narrow or absent,and 8 cases on the left,and one case on the right;Weak correlation was found between jugular veins pattern and the age.The right sided values in volunteers were higher than that of the left side.Conclusion The morphology and hemodynamics of jugular veins in volunteers showed significant difference between sides,and weak correlation is found between the morphology and aging.

Objective To evaluate the deep brain venous blood oxygen content changes in patients with multiple sclerosis(MS) using susceptibility weighted imaging (SWI), and to explore the ability of SWI in reflecting the clinical condition. Methods Forty-four MS patients were prospectively enrolled in the study. All the clinical-proved patients meeting the McDonald standards (2005 revised) underwent conventional MRI, SWI, and 12 cases of them underwent MRI review from 12 to 16 months interval. all the patients' clinical condition were quantified according to the expanded disability status scale(EDSS). The score was 0.5—6.5. Sixty-five age- and gender- matched healthy volunteers underwent conventional MRI and SWI. The blood oxygen content of the deep brain venous were estimated by the veins phase value, and differential phase values of blood vessels and surrounding tissues (Δφ) were processed with SPIN software. The blood vessels consist of bilateral BV, SMCV, ICV, STV and FMV, PMV, OMV. The difference of Δφvalue in different veins between MS patients and the controls was compared using independent sample t-test, and the Δφ value comparison of MS patients in different time were performed by using paired t test; The correlation ofΔφvalue between MS and EDSS was analyzed using Spearman correlation. Results TheΔφvalue of BV, SMCV, ICV, STV were 856.6 ± 246.4, 600.6 ± 155.2, 965.9 ± 205.4, 844.2 ± 149.7 in MS, and 767.6±145.1, 536.2±123.5, 892.8±156.3, 783.1±148.5 in controls, respectively. TheΔφvalue was higher in MS patients than the controls (t=2.157, 2.303, 2.005, 2.103,P<0.05). The twelve patients'Δφvalues of BV, ICV,STV were 729.4±275.1, 906.1±219.2, 737.2±159.1 in the first time, and 923.2±211.6, 1017.3±211.1, 919.3 ± 165.9 in the second time, and all the values increased in the review of the interval of 12 to 16 months (t=-3.092,-6.420,-3.972,P<0.05). The phase value of PMV and OMV had significant positive correlation with EDSS scores(r=0.638, 0.642,P<0.01). Conclusions The state of hypoxia of the brain parenchyma appears in MS patients, and hypoxia may become worse with the extension of course. The extent of hypoxia can reflect the disability of the patients.

Objective To investigate the current status of secondary prevention of cerebral infarction in Longquanyi District of Chengdu. Methods Prospective Investigation on the secondary prevention was conducted in 94 pa-tients with cerebral infarction at 3, 12 and 24 months following stroke. Results The percentages of patients receiving anti-platelet therapy, antihypertensive treatment, antidiabetic drug treatment and statin therapy were 96.8%, 92.1%, 91.7%and 63.8%at hospital discharge, 79.8%, 88.9%, 91.7%and 55.3%at 3 months, 76.1%, 75.0%and 41.4%, 63.8%at 12 months and 33.0%, 71.4%;58.3%and 22.3%at 24 months, respectively. None of stroke patients with atrial fibrillation re-ceived anticoagulation therapy. Conclusion There is a large gap between the current practice of secondary prevention and guideline in patients with cerebral infarction in Longquanyi District of Chengdu.

Objective To establish a rabbit model of restenosis and analyze the expressions of VEGFmRNA and TGF-?_1mRNA during the intimal proliferation.We also explored the relationship between VEGFmRNA,TGF-?_1mRNA and restenosis.Methods 40 healthy male New Zealand white rabbits were evenly divided into three injury groups and one control group.Right carotid arteries were injured with PCI balloon in the injury groups.10 rabbits of each injury group were sacrificed on weeks 1,2 and 4 after the injury.VEGFmRNA and TGF-?_1mRNA were examined by in situ hybridization.All the samples were analyzed using a computerized imaging analysis system.Results In the injury groups,neointimal areas were significantly larger than those in control group(P

OBJECTIVEThis study examined the levels of exhaled nitric oxide (eNO) and peripheral blood eosinophils (EOS) as well as the correlation between the two markers in children with bronchial asthma (AS),AS complicated by allergic rhinitis (AS/AR) and chronic cough variant asthma (CVA), in order to explore the value of eNOS detection in children with AS.

METHODSThe eNO level was measured using light-emitting electrochemical photometry in 12 children with AS, 29 children with AS/AR and 10 children with CVA. Peripheral blood EOS was counted by blood cell counter (Coulter JT). Forced expiratory volume in one second (FEV1) was assessed by lung function measurement. Thirty children without atopic disease and acute respiratory infection as well as without a family history of atopic diseasea served as the control group.

RESULTSThe levels of eNO and blood EOS in the AS, the AS/AR and the CVA groups were significantly higher than those in the control group (p<0.01). The AS/AR group showed increased levels of eNO (50.3 + or - 6.7 ppb) and EOS (5.9 + or -4.2 x 109 ) compared with the AS (30.5 + or - 8.8 ppb and 4.2 + or - 3.2 x 109 respectively) and the CVA groups (26.0 + or - 3.2 ppb and 3.7 + or - 6.9 x 109 respectively) (p<0.05). There were no significant differences in eNO and EOS levels between the AS and the CVA groups. The eNO level was positively correlated with the EOS level (r=0.51, p<0.05), but not with FEV1 (r=0.144, p>0.05) in the AS group.

CONCLUSIONSNO is highly expressed in children with symptoms of atopy and can reflect the levels of eosinophilic airway inflammation in children with AS.

Adolescent ; Asthma ; blood ; physiopathology ; Breath Tests ; Child ; Child, Preschool ; Eosinophils ; physiology ; Female ; Forced Expiratory Volume ; Humans ; Male ; Nitric Oxide ; metabolism

OBJECTIVETo acquire the genes of light chain variable region (VL) and heavy chain variable region (VH) of a novel clone ZCH-7-2F9(2F9) of anti-hCD14 for construction of anti-hCD14 single chain antibody(ScFv).

METHODSFrom the mouse hybridoma cell line 2F9 and its fusion partner murine myeloma cell line NS-1, total RNA was prepared. The VL and VH genes were amplified by RT-PCR with family specific primer pairs, respectively. The PCR products were cloned into pGEM(sound recording copyright sign)-T Easy vectors, then transfected into DH5alpha and the positive recombinants were identified and purified. After sequencing with automatic DNA sequencer the sequences were analyzed online.

RESULTSVL gene of the new clone of CD14 monoclonal antibody (McAb) 2F9 consisted of 321 bps encoding a peptide of 107 amino acid residues, and VH gene of the 2F9 antibody contained 360 bps encoding a peptide of 120 amino acid residues. According to IMMUNOGENETICS online analysis by IMGT/V-QUEST, the VL and VH genes belonged to mouse IGKV and mouse IGHV subgroups, respectively. On the position 23/88 of the light chain and 22/96 of the heavy chain genes there were cysteines, which play the key role in forming disulfo-bond in each chain. Both VL and VH chains had definitely 4 frame regions (FR) and 3 complementary determinant regions (CDR).

CONCLUSIONThe treatment of CML consisting of myeloablative Allo-SCT combined with Gleevec before and after transplantation is an effective and safe method for CML.

Amino Acid Sequence ; Animals ; Antibodies, Monoclonal ; genetics ; immunology ; Base Sequence ; Cloning, Molecular ; Humans ; Immunoglobulin Heavy Chains ; genetics ; Immunoglobulin Light Chains ; genetics ; Immunoglobulin Variable Region ; genetics ; Lipopolysaccharide Receptors ; immunology ; Mice ; Mice, Inbred BALB C ; Molecular Sequence Data ; Polymerase Chain Reaction ; Sequence Analysis, DNA

OBJECTIVETo investigate the T2 values and knee thickness in healthy young adults using 3.0 T magnetic resonance imaging(MRI) .

METHODSTotally 40 volunteers(18-30 years old) with body mass index between 18.5-24.0 kg/m(2) were divided into two groups(22 men and 18 women) according to their gender. Also in addition, each group was divided into two subgroups(right knee and left knee) . The T2 values and the thickness of the areas on the medial condyle of femur, the lateral condyle of femur, the medial tibial plateau, the lateral tibial plateau, and the patella of the knee cartilage were measured.

RESULTSThe T2 values and the thickness of the right and left knee cartilages showed no significant differences between men and women (P>0.05) . Also, the T2 values in the five parts of the knee cartilage also were not significantly different between men and women (P>0.05) . However, the thickness of the 5 parts of the knee cartilage significantly differed between men and women(P<0.05) .

CONCLUSIONSThe thickness of the knee cartilage may different between male and female young adults. The T2 values of the cartilage may be not affected by the gender.

Adolescent ; Adult ; Cartilage, Articular ; anatomy & histology ; Female ; Humans ; Knee Joint ; anatomy & histology ; Magnetic Resonance Imaging ; Male ; Sex Factors ; Young Adult

OBJECTIVEAcute monocytic leukemia (AML)-M5 is the common type of acute myeloid leukemias in children. Studies have shown that there are abundant lipopolysaccharide (LPS) receptor (designated as CD14) molecules on the cell membrane of M5 cells and they play an important role in the diagnosis of M5, since they can be recognized and bound by mouse-anti-human CD14 monoclonal antibody (McAb). However, mouse-originated antibodies are largely not suitable for clinical application due to the severe side effects, thus "humanized antibody" is desired. As the first step for developing humanized antibody, the construction and expression of single chain antibody (scFv) with functional protein are necessary. The present study aimed to express ZCH-7-2F9 ScFv (scFv(2F9)) in eukaryotic cells and obtain the scFv(2F9) protein with a high biological activity.

METHODSFour primers were synthesized to construct the eukaryotic expressional vector, which included SfiI and EcoRI enzyme cleaving site, 6 x His and stop code TGA sequences. scFv(2F9) gene was amplified through splicing by overlap extension (SOE) using the high fidelity Taq polymerase. Positive recombinants (pSectag2A/scFv(2F9)) were identified through enzyme cleaving and sequenced before expression and were transformed into Chinese hamster ovary (CHO) cells for expression. Western-Blot and flow cytometry (FCM) were carried out to determine the relative molecular mass (Mr) and binding activity of scFv(2F9).

RESULTSThe cloned scFv(2F9) gene was identified to be functional by sequencing and expressing. The interested protein could be detected in the culture supernatant of transformed CHO cells with an Mr of 31 000. The blocking test showed that the positive cell percentages, the mean fluorescence intensity (MFI) and the peak of channel (peak Ch) were reduced by 90.02%, 63.30% and 63.38%, respectively after blocking with scFv(2F9), while those were 4.55%, 10.09% and 5.02% after blockage using the supernatant from the CHO cells transfected with blanked vector pSectag2A.

CONCLUSIONSThe scFv(2F9) against human CD14 antigen was successfully expressed in eukaryotic cells and showed a high biological activity, which may be useful for the further studies on its humanized antibodies.

Amino Acid Sequence ; Animals ; Antibodies, Monoclonal ; genetics ; isolation & purification ; Base Sequence ; CHO Cells ; Child ; Cloning, Molecular ; Cricetinae ; Cricetulus ; Humans ; Lipopolysaccharide Receptors ; immunology ; Molecular Sequence Data ; Single-Chain Antibodies ; genetics ; isolation & purification