Objective To develop a matrix metalloproteinase(MMP)-responsive hyaluronic acid(HA)-based controlled-release material for brain-derived neurotrophic factor(BDNF)to provide a novel therapeutic strategy for intervention and repair of traumatic brain injury(TBI).Methods HA was modified with amination,followed by condensation with Suflo-SMCC carboxyl group to form amide,and then linked with glutathione(GSH)to synthesize HA-GSH.The recombinant glutathione S-transferase(GST)-tissue inhibitor of metalloproteinase(TIMP)-BDNF(GST-TIMP-BDNF)expression plasmid was constructed using molecular cloning technique with double enzyme digestion by Bam H Ⅰ and Eco R Ⅰ.The recombinant GST-TIMP-BDNF protein was expressed in the Escherichia coli prokaryotic expression system,and purified by ion exchange chromatography,confirmed by Western blotting.MMP diluents were supplemented with PBS,MMP inhibitor marimastat,and varing concentrations(0.4,0.6,0.8 mg/ml)of GST-TIMP-BDNF or GST-BDNF.MMP-2 activity was analyzed using an MMP activity detection kit to evaluate the inhibitory effect of the recombinant protein on MMP.Primary rat neurons were extracted and cultured to establish an iron death model induced by RSL3.The effect of recombinant protein GST-TIMP-BDNF on neuronal injury was detected by immunofluorescence staining.Results MRI hydrogen spectrum identification confirmed the successful synthesis of HA-GSH.Western blotting results showed the successful expression of the recombinant protein GST-TIMP-BDNF containing the GST tag using the E.coli prokaryotic expression system.MMP activity detection results indicated that the recombinant protein GST-TIMP-BDNF had a superior inhibitory effect on MMP-2 activity compared to GST-BDNF(P<0.05).Immunofluorescence staining results showed a significant increase in fluorescence intensity in rat neurons treated with GST-TIMP-BDNF after RSL3 induction(P<0.05).Conclusion A MMP-responsive HA-based BDNF controlled-release material has been successfully developed,exhibiting a protective effect on neuron damage.

【Objective】To investigate the effect of dexmedetomidine on the phagocytosis of macrophages.【Methods】 RAW264.7 cells were divided into control group，DEX group，and BRL44408 + DEX group. Expression of α2A adrenergic receptor，p-Akt and Akt were detected by Western Blotting；Phagocytosis Assay Kit（IgG PE）was used to measure the phagocytosis of macrophages. 【Results】 α2A adrenergic receptor was detected in RAW264.7 cells；Dexmedetomidine could enhance the phagocytosis of macrophages（P < 0.001），and BRL44408 reversed the enhancement of phagocytic ability of macrophages（P < 0.001）；Dexmedetomidine upregulated the expression of Akt in RAW264.7 cells，while the use of BRL44408 inhibited the activation of the Akt pathway（P < 0.01）.【Conclusion】Dexmedetomidine could enhance phagocytosis of RAW264.7 by activating the Akt pathway through the α2A adrenergic receptor.

In this study, RACE technology was employed to isolate the full length cDNA of DoHT1 in Dendrobium officinale, followed by bioinformatics analysis of the sequence characteristics. And the expression pattern of the gene was also analyzed by quantitative PCR. The full length cDNA of DoHT1 was 1 586 bp in length, containing a 1 536 bp ORF, which encoded a 511-aa protein with molecular weight of 56.18 kD and isoelectric point of 9.08. The deduced DoHT1 protein had the major facilitator superfamily conserved domain (22-483), SUGAR₋TRANSPORT₋1 (139-164), and SUGAR₋TRANSPORT₋2 (338-355), typical for sugar transporter; DoHT1, without a single peptide had 11 transmembrane regions, and was predicted to locate in the plasma membrane; DoHT1 had high identities (54.7%-80.7%) with HTs proteins from various plants. DoHT1 belonged to the MST (monosaccharide transporter) group of the evolutionary tree, and was closely related to the Phalaenopsis equestris. DoHT1 was differentially expressed in the three included organs. The transcripts were significantly the most abundant in the leaves with 19.36 fold than roots, then 1.82 fold in the stems than the roots. The identification and molecular characterization of the full length DoHT1 will be essential for further function study of the gene during the regulation of sugar metabolism of D. officinale.

The calcineurin B-like protein (CBL)-interacting protein kinase (CIPK) plays a vital role in the growth, development, and stresses adaptation in plants by interaction with the calcium signaling. In this study, four full length cDNAs of CIPKs genes, namely DoCIPK1, DoCIPK2, DoCIPK3 and DoCIPK4 (GenBank accession No. KT957557, KT957558, KT957559 and KT957560, respectively) were cloned from the rear and medicinal plant, Dendrobium officinale, by rapid amplification of cDNA ends (RACE) for the first time. The corresponding encoded proteins, consisting of 473, 449, 451 and 440 amino acids (aa), respectively, with a molecular weight of 53.50, 50.93, 51.50 and 50.16 kDa, and an isoelectric point (pI) of 7.99, 9.25, 8.81 and 9.11, respectively, shared 70%-90%, 69%-80%, 78%-93%, and 66%-82% identities CIPKs with various plants. Each deduced protein contained a conserved protein kinase domain (respectively at 21 -275, 14-268, 16-271 and 12-266 aa position), a CIPKs family characteristic NAF/FISL domain (respectively at 335-391, 313-370, 310-369 and 305-362 aa position) and some functional motifs. The four DoCIPK proteins, without signal peptide or transmembrane region, were located in the plasma membrane and endoplasmic reticulum at the subcellular level. The three dimensional structure of the proteins were similar to that of Arabidopsis AtCIPK24. DoCIPK1 and DoCIPK3 were respectively clustered in the group E and A of the Arabidopsis and rice CIPK evolutionary tree, while DoCIPK2 and DoCIPK4 belonged to group C. The relative expression of DoCIPK1 showed no significant difference in the leaves and stems, and its transcripts in the roots was 0.35 fold over that in the leaves. The abundance of DoCIPK3 transcripts in the stems and the roots were 3.36 fold and 3.47 fold higher, respectively, than those in the leaves. DoCIPK2 exhibited similar expression pattern to DoCIPK4. Their relative expression in the leaves and the stems had no apparent difference, and the transcript levels were higher in the roots than that in the leaves, with 2.08 fold and 7.86 fold, respectively. Cloning, bioinformatics analyses, and expression patterns of the four DoCIPK genes provide a basis for functional elucidation of these genes further during the physiological responses in D. officinale.

Anthraquinones are not only the main active constituents but also the index components for the quality control of Rhei Radix et Rhizoma. To study the anthraquinone biosynthesis, Rheum palmatum L. seedlings were subjected to a high-throughput transcriptomic sequencing analysis by Illumina HiSeq^TM 2000 150PE. The Illumina sequencing generated a total of 11.04 G clean data resulting in 736 309 74 clean reads, deposited in the sequence read archive (SRA accession SRP160030). Trinity do novo assembly yielded 93 646 unigenes, with an average of 1 108 nt. Functional annotation revealed that all unigenes were successfully annotated in the NR, NT, Swiss-port, PFAM, and KOG databases. GO enrichments showed that 57 subgroups were involved in biological process, cellular component, and molecular function. KEGG analysis indicated that 1 107 unigenes were implicated in 19 standard secondary metabolic pathways. 172 unigenes were analyzed to encode 28 key enzymes during the MVA, MEP, shikimic acid, and polyketide pathways related to anthraquinone biosynthesis. 125 CYP450 and 73 UGTs unigenes were related the modification of secondary metabolites in R. palmatum L. Furthermore, seven unigenes with full length cDNAs were successfully verified by RT-PCR and sequencing analyses. Then, MISA prediction produced a number of 18 885 simple sequence repeats (SSRs). Herein, the transcriptomic gene expression profiles of R. palmatum L. and candidate genes during the anthraquinone biosynthesis pathway were obtained for the first time. The results provided basic information for subsequent gene function characterization, secondary metabolic pathway analysis, and anthraquinone biosynthesis and regulation elucidation in R. palmatum L.

OBJECTIVETo study the effect of exogenous Ca2+ on photosynthetic parameters of Pinellia ternate and accumulations of active components under high temperature stress.

METHODThe pigment contents of P. ternata leaves, photosynthesis parameters and chlorophyll fluorescence parameters of P. ternata leaves, the contents of guanosine, adenosine and polysaccharide in P. ternata tubers were measured based on different concentrations of exogenous Ca2+ in heat stress when the plant height of P. ternata was around 10 cm.

RESULTThe contents of total chlorophyll and ratio of chlorophyll a/b were relatively higher by spaying Ca2+. Compared with the control, spaying 6 mmol x L(-1) Ca2+ significantly enhanced the net photosynthetic rate (Pn), transpiration (Tr) and stomatal limitation (L8), but reduced intercellular CO2 concentration (C) in P. ternata leaves. With the increase of Ca2+ concentration, maximal PS II efficiency (Fv/Fm), actual photosynthetic efficiency (Yield) and photochemical quenching coefficient (qP) initially increased and then decreased, however, minimal fluorescence (Fo) and non-photochemical quenching coefficient (NPQ) went down first and then went up. The contents of guanosine and polysaccharide and dry weight of P. ternata tubers showed a tendency of increase after decrease, and the content of adenosine increased with the increase of Ca2+ concentration. The content of guanosine and polysaccharide in P. ternata tubers and its dry weight reached maximum when spaying 6 mmol x L(-1) Ca2+.

CONCLUSIONWith the treatment of calcium ion, the inhibition of photosynthesis and the damage of PS II system were relieved in heat stress, which increased the production of P. ternata tubers.

Breeding ; Calcium ; pharmacology ; Chlorophyll ; metabolism ; Dose-Response Relationship, Drug ; Heat-Shock Response ; drug effects ; Organ Size ; drug effects ; Photosynthesis ; drug effects ; Pinellia ; drug effects ; growth & development ; metabolism ; physiology ; Plant Leaves ; drug effects ; growth & development ; metabolism

Objective To examine the cognitive function of the childhood schizophrenia and healthy children,and to explore the characterized impairment of cognitive function in childhood schizophrenia.Methods Fifty untreated children with first-episode schizophrenia(schizophrenia group) and 50 healthy children(control group) were enrolled in the study.Chinese Wechsler intelligence scale for children (C-WISC),Wiscosin Cards Sorting Test (WCST),Stroop test and Trail making test A and B(TMT-A/B) were used to measure the cognitive function.The Clinical features were assessed with the scale for positive and negative syndrome(PANSS).Results Full intelligence quotient (FIQ),verbal intelligence quotient (VIQ) and performance intelligence quotient (PIQ) in schizophrenia group were significantly lower than those in the control group(P ＜ 0.05),but the change between VIQ and PIQ in schizophrenia group was higher than that in the control group(P =0.023).The complete categories in schizophrenia group were significantly lower than that in the control group(P ＜ 0.05).The total number of errors and the continuous errors in the schizophrenia group were significantly higher than those in the control group(P ＜0.05).No significant difference of the random errors was observed between the schizophrenia group and the control group.The times to complete word A,color B,double-word C and double-color D in the control group were significantly less than those in the schizophrenia group(P ＜ 0.05).The reaction time of Trail Making Test A/B and the error number of B in the schizophrenia group were significantly higher than those in the control group(P ＜ 0.05).No significant difference of the A error numbers were observed between the 2 groups (P ＞ 0.05).There were significant negative relationships between the negative syndrome and VIQ,PIQ(P ＜0.05) ;in contrast,there was significantly positive relationship between the negative syndrome and the A error numbers (P ＜ 0.05).There was significantly negative relationship between the total score of PANSS and completed categories (P ＜ 0.05).Multiple regression analysis showed that:there was a significant correlation between the damage of cognitive function in children and negative symptoms,PANSS total score,gender,age,years of education and place of residence (all P ＜ 0.05).Conclusions Children with first-episode schizophrenia have multiple cognitive deficits in speed of processing working memory,spatial memory,attention and executive function.Some of the cognitive impairments have certain relationships with negative syndrome.

BACKGROUNDThe aim of this study was to investigate the potential relationship between the dynamic expression of Toll-like receptor 2 and 4 (TLR2/4) in peripheral blood mononuclear cells as well as changes in serum concentration of inflammatory factors and acute lung injury (ALI) in patients after orthotopic liver transplantation (OLT).

METHODSThe peripheral blood samples of 27 patients (23 men and 4 women with ASA III to IV) who received OLT were collected for measurement of TLR2/4 at T1 (after induction of anesthesia), T2 (25 minutes after anhepatic phase), T3 (3 hours after graft reperfusion) and T4 (24 hours after graft reperfusion). The expression of TLR2/4 in mononuclear cells was measured by flow cytometry. The serum concentrations of tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-8 were measured by enzyme-linked immunosorbent assay (ELISA). Twenty-seven patients were assigned to ALI group (n = 9) and non-ALI group (n = 18) according to the diagnostic criteria of ALI. The expression of TLR2/4 in the ALI group or non-ALI group was analyzed.

RESULTSCompared to the non-ALI group, the volumes of blood loss, ascites, total output and transfused red blood cells were higher in the ALI group, and the anhepatic phase lasted longer (P < 0.05, P < 0.01). The expression of TLR2/4 in mononuclear cells increased significantly at T3 and T4, and serum concentrations of TNF-alpha, IL-1beta and IL-8 increased significantly too. There was no significant difference in Child-Turcotte-Pugh (CTP) scores between the ALI group and non-ALI group (P > 0.05). The expression of TLR2/4 in mononuclear cells increased significantly at T3 and T4 in the ALI group (P < 0.05, P < 0.01). A positive correlation was noted between the expression of TLR4 in mononuclear cells and the serum concentrations of TNF-alpha, IL-1beta (P = 0.041, P = 0.046) in the ALI group. In the non-ALI group, statistical results showed that the expression level of TLR2/4 in mononuclear cells was not significantly different during the peri-operative period of OLT (besides TLR4 expression at T4). Compared to the non-ALI group, the increasing amplitude of TLR2/4 expression in mononuclear cells was more significant in the ALI group. The patients whose TLR2/4 expression in mononuclear cells exceeded that at T1 by one time were more likely to suffer from ALI (P = 0.013), with a relative risk of 16.

CONCLUSIONThe expression level of TLR2/4 in mononuclear cells increases significantly in the peri-operative period of OLT, and it may be a high risk factor for occurrence of postoperative ALI.

Acute Lung Injury ; etiology ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Female ; Flow Cytometry ; Humans ; Interleukin-1beta ; metabolism ; Interleukin-8 ; metabolism ; Leukocytes, Mononuclear ; metabolism ; Liver Transplantation ; adverse effects ; Male ; Postoperative Period ; Toll-Like Receptor 2 ; metabolism ; Toll-Like Receptor 4 ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo observe the effects of astragalus membranacaus injection on the activity of the intestinal mucosal mast cells (IMMC) and inflammatory response after hemorrahagic shock-reperfusion in rats.

METHODThirty-two Wistar rats were randomly divided into four groups: normal group, model group, low dosage group, (treated with astragalus membranacaus 10 g kg(-1)) and high dosage group (treated with astragalus membranacaus 20 g kg(-1)). Models of hemorrhage shock for 60 minutes and reperfusion for 90 minutes were created. The animals were administrated 3 mL therapeutic solution before reperfusion. At the end of study, intestinal pathology, ultrastructure of IMMC, and expression of tryptase were observed. The levels of MDA, TNF-a, histamine, and SOD activity of intestinal were detected, and the number of IMMC was counted.

RESULTThe degranulation of IMMC was seen in model group and was attenuated by astragalus membranacaus treatment. Chiu's score of model group was higher than that of the other groups. Astragalus membranacaus could attenuate the up-regulation of the Chiu' s score, the levels of MDA and TNF-alpha, expression of tryptase, and the down-regulation of SOD activity and histamine concentration. The Chiu's score and MDA content were negatively, while SOD activity was positively correlated to the histamine concentration respectively in the four groups.

CONCLUSIONAstragalus membranacaus can reduce small intestine mucosal damage by inhibiting the activity of IMMC after hemorrhage shock reperfusion.

Animals ; Astragalus membranaceus ; chemistry ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Female ; Injections, Intravenous ; Intestinal Mucosa ; metabolism ; pathology ; Intestine, Small ; metabolism ; Male ; Malondialdehyde ; metabolism ; Mast Cells ; drug effects ; metabolism ; ultrastructure ; Random Allocation ; Rats ; Rats, Wistar ; Reperfusion Injury ; metabolism ; pathology ; Shock, Hemorrhagic ; metabolism ; pathology ; Tryptases ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo summarize the experiences of cardiopulmonary bypass (CPB) techniques in 15 patients of orthotopic heart transplantation.

METHODSAll patients received mild hypothermia and high flow rate perfusion. Effective strategies were taken to protect myocardium, lung, kidney, and blood conservation. The donor hearts were arrested with aorto perfusion using 1 000 ml St. Thomas solution at 4 degrees C, perfused with 1 000 ml University of Wiscosin (UW) solution or Histidin-Tryptophan-Ketoglutarat (HTK) solution at 4 degrees C, and then preserved in ice saline. Ice mud was covered on the donor heart during anastomosis. Low potassium cardioplegia solution was perfused before the unclamping of aorta.

RESULTSThe CPB time was (165.2 +/- 22. 8) min, the warm ischemia time was ( 7. 7 +/- 1. 7) min, and the cold ischemia time was ( 142. 4 +/- 11. 5) min. Heart beats was automatically recovered in 10 patients, and was recovered upon defibrillation in 5 patients. Left ventricular ejection fractions were (64. 1 +/- 4. 6) % after one month. All patients survived.

CONCLUSIONProper CPB management and effective donor heart protection are essential to guarantee the success of heart transplantation.

Cardiopulmonary Bypass ; Heart Transplantation ; methods ; Humans