T-cell lymphopenia is associated with an increased infecting risk in children after cardiopulmonary bypass.

BACKGROUND: children who undergo CPB operations are at an elevated risk of infection due to immunosuppression. This study aims to investigate the association between lymphopenia following CPB and early postoperative infection in children. METHODS: A retrospective analysis including 41 children under 2 years old underwent CPB. Among them, 9 subjects had an early postoperative infection, and 32 subjects were period-matched without infection. Inflammatory cytokines, serum CRP and PCT values were measured in plasma, additionally, circulating total leucocyte and lymphocyte subpopulations were counted. RESULTS: Infected subjects exhibited significantly higher levels of inflammatory cytokines, including IL-6, IL-8, IL-10, IL-1ß and TNF-α, than non-infected subjects after CPB. Additionally, lower absolute number of lymphocyte and their subpopulations CD3+ T cells, CD4+ T-helper cells and CD8+cytotoxic T-cells, were observed in infected subjects. The impairment of T-cells Immune was found to be associated with higher levels of inflammatory cytokines IL-10. The ROC demonstrated that the absolute number of CD3+ T-cells <1934/ul, CD4+ T helper cells <1203/ul and CD8+cytotoxic T-cells <327/ul were associated with early postoperative infection. CONCLUSION: Higher levels of inflammatory cytokines resulted in T-cells lymphopenia after CPB, which significantly increasing the risk of postoperative infection in infants and young children. IMPACT: Infection complications after cardiopulmonary bypass (CPB) in pediatric CHD patients are serious issues, identifing the infection from after CPB remains a challenging. CPB can release numerous inflammatory cytokines associated with T cells lymphopenia, which increases the risk of postoperative infection after surgery. Monitoring T cells lymphopenia maybe more beneficial to predict early postoperative infection than C-reactive protein and procalcitonin.

Fetal pulmonary valve stenosis or atresia with intact ventricular septum: Predictors of need for neonatal intervention.

OBJECTIVES: To compare the fetal echocardiographic measurements and neonatal outcome of fetuses with diagnosis of critical pulmonary stenosis (CPS/IVS) and pulmonary atresia with intact ventricular septum (PA/IVS) to identify the predictors of neonatal ductus dependence and the need for neonatal intervention. METHODS: Forty-four fetuses with a diagnosis of membranous PA/IVS or CPS/IVS referred to Shanghai Xinhua Hospital Affiliated to Shanghai Jiaotong University between June 2009 and November 2014 were respectively analyzed. We analyzed their fetal and pediatric echocardiographic features and clinical features after birth. RESULTS: Among the 29 infants in the final analysis, 19 were ductal dependent and were diagnosed as CPS/IVS or PA/IVS after birth. Ten patients with fetal CSP/IVS were ductal independent and were diagnosed as pulmonary stenosis after birth. In midtrimester, significance was only detected in pulmonary valve (PV) regurgitation between 2 groups (P = .009). The fetuses with PV regurgitation in midtrimester were more likely to be ductal independent (odds ratio = 6.67; P = .010) than those with absence of PV regurgitation in the midtrimester scan. In late trimester, the infants in ductus-independent group had better fetal right ventricular development than those in ductus dependent group. The right ventricle over left ventricle length ratio in last scan over 0.86 had the best capability in predicting ductus independence during neonatal period with help of receiver-operating characteristic curve and the logistic regression analysis. CONCLUSION: Among fetuses with pulmonary valve stenosis and right ventricle dysplasia at the time of first fetal echo in midtrimester, use of pulmonary valve regurgitation can identify who will be ductal dependent at births. In late trimester, right ventricle over left ventricle length ratio less than 0.86 had the best capability of predicting neonatal ductal dependence. Fetuses with reversed ductus arteriosus flow direction in late trimester were more likely to be ductus dependent after birth.

Simple-MSSM: a simple and efficient method for simultaneous multi-site saturation mutagenesis.

OBJECTIVE: To develop a practically simple and robust multi-site saturation mutagenesis (MSSM) method that enables simultaneously recombination of amino acid positions for focused mutant library generation. RESULTS: A general restriction enzyme-free and ligase-free MSSM method (Simple-MSSM) based on prolonged overlap extension PCR (POE-PCR) and Simple Cloning techniques. As a proof of principle of Simple-MSSM, the gene of eGFP (enhanced green fluorescent protein) was used as a template gene for simultaneous mutagenesis of five codons. Forty-eight randomly selected clones were sequenced. Sequencing revealed that all the 48 clones showed at least one mutant codon (mutation efficiency = 100%), and 46 out of the 48 clones had mutations at all the five codons. The obtained diversities at these five codons are 27, 24, 26, 26 and 22, respectively, which correspond to 84, 75, 81, 81, 69% of the theoretical diversity offered by NNK-degeneration (32 codons; NNK, K = T or G). CONCLUSION: The enzyme-free Simple-MSSM method can simultaneously and efficiently saturate five codons within one day, and therefore avoid missing interactions between residues in interacting amino acid networks.

Three new flavonoids from the active extract of Fallopia convolvulus.

Five solvent extracts (ethanol, petroleum ether, EtOAC, n-butanol, and water) from Fallopia convolvulus (L.) Love were separated and their inhibitory effects on nitric oxide production in lipopolysaccharide-activated macrophages were evaluated. Three new flavonoids, falloconvolin A (1), falloconvolin B (2), and quercetin-3-O-(2-E-sinapoxyl)-glucopyranoside (3), together with 17 known phenolic compounds, were isolated from the active EtOAC extract, and their structures were elucidated on the basis of spectroscopic analysis and literature data.

Potential Biochemical Mechanisms of Brain Injury in Diabetes Mellitus.

The goal of this review was to summarize current biochemical mechanisms of and risk factors for diabetic brain injury. We mainly summarized mechanisms published in the past three years and focused on diabetes induced cognitive impairment, diabetes-linked Alzheimer's disease, and diabetic stroke. We think there is a need to conduct further studies with increased sample sizes and prolonged period of follow-ups to clarify the effect of DM on brain dysfunction. Additionally, we also think that enhancing experimental reproducibility using animal models in conjunction with application of advanced devices should be considered when new experiments are designed. It is expected that further investigation of the underlying mechanisms of diabetic cognitive impairment will provide novel insights into therapeutic approaches for ameliorating diabetes-associated injury in the brain.

Distribution of cfr in Staphylococcus spp. and Escherichia coli Strains from Pig Farms in China and Characterization of a Novel cfr-Carrying F43:A-:B- Plasmid.

The multi-resistance gene cfr is widely distributed among various gram-positive and gram-negative species in livestock in China. To better understand the epidemiology of cfr among Staphylococcus spp. and E. coli isolates, 254 Staphylococcus spp. and 398 E. coli strains collected from six swine farms in China were subjected to prevalence and genetic analysis. Forty (15.7%) Staphylococcus spp. isolates, including 38 Staphylococcus sciuri strains, one Staphylococcus chromogenes strain, and one Staphylococcus lentus strain, and two (0.5%) E. coli isolates were found to contain the cfr gene. Most of the 38 S. sciuri strains were clonally unrelated; however, clonal dissemination of cfr-positive S. sciuri was detected at the same farm. In eight randomly selected cfr-positive staphylococci, a cfr-harboring module (IS21-558-cfr-ΔtnpB) was detected in six S. sciuri isolates; cfr was bracketed by two copies of ISEnfa4 or IS256 in the remaining two S. sciuri isolates. In the two E. coli isolates, EP25 and EP28, cfr was flanked by two IS26 elements in the same or opposite orientation, respectively. Complete sequence analysis of the novel F43:A-:B- plasmid pHNEP28 revealed that it contains two multi-resistance regions: cfr together with floR, qnrS1 interspersed with IS26, ΔISCR2 and ISKpn19, and blaTEM-1 together with tet(M) interspersed with IS26, ISApl1, ΔTn2, and ΔIS1B. The coexistence of cfr with other resistance genes on a conjugative plasmid may contribute to the dissemination of these genes by co-selection. Thus, rational drug use and continued surveillance of cfr in swine farms are warranted.

[The effects of calmodulin on the lipid-binding activity of CaM-binding protein-10 and maize non-specific lipid transfer protein].

Fluorescence-marked lipid binding experiment shows that CaMBP-10 has typical lipid binding feature of non-specific lipid transfer protein (nsLTP). We have compared the effects of calmodulin (CaM) on the lipid-binding activity of CaMBP-10 and maize nsLTP. Different influences were found in the presence of either Ca(2+) or EGTA. W-7 and TFP could abolish the influence of CaM. Therefore, it is suggested that CaM could interact specifically with both CaMBP-10 and maize nsLTP. Probably, there are different CaM regulatory mechanisms between CaMBP-10 and maize nsLTP.

[Cloning and expression of cDNA for maize nonspecific lipid transfer protein as well as calmodulin-binding activity analysis of the expression product].

Maize nonspecific lipid transfer protein (Zm-nsLTP) was cloned and expressed to investigate its CaM-binding activity. The cDNA of Zm-nsLTP was amplified using RT-PCR (Fig.1), and then inserted into the vector pET32a(+). The recombinant vector pET-Zm-nsLTP was expressed in E. coli BL21(DE3)trxB(-). Results of CaM-gel overlay assays (Fig.2) and CaM-sepharose pull-down experiments (Fig.3) indicated that recombinant Zm-nsLTP was bound to CaM in a Ca(2+)-independent manner, which is in accordance with the way that CaMBP-10 and Arabidopsis non-specific lipid transfer protein-1 (At-nsLTP1) are bound to CaM. The CaM-binding domain in Zm-nsLTP was mapped to the region of 47-60 amino acids (Fig.3), and online sequence analysis using Predict Protein program predicted that it has a BAA structure (Fig.4,5).

Bioconversion of isoeugenol into vanillin by crude enzyme extracted from soybean.

Crude enzyme extracted from soybean was used to convert isoeugenol into vanillin. The effects of several factors on the bioconversion were studied. Conversion was affected by the amount of substrate and was also improved by the addition of absorbents, among which powdered activated carbon was the best. The effect of H2O2 concentration on the conversion was also studied. The optimum concentration of H2O2 was 1% (v/v). With 10 g/L of powdered activated carbon and 0.1% H2O2 added, vanillin reached a maximum concentration of 2.46 g/L after 36 h, corresponding to a molar yield of 13.3%.

Tumor-derived transforming growth factor-ß is critical for tumor progression and evasion from immune surveillance.

Tumors have evolved numerous mechanisms by which they can escape from immune surveillance. One of these is to produce immunosuppressive cytokines. Transforming growth factor-ß(TGF-ß) is a pleiotropic cytokine with a crucial function in mediating immune suppression, especially in the tumor microenvironment. TGF-ß produced by T cells has been demonstrated as an important factor for suppressing antitumor immune responses, but the role of tumor-derived TGF-ß in this process is poorly understood. In this study, we demonstrated that knockdown of tumor-derived TGF-ß using shRNA resulted in dramatically reduced tumor size, slowing tumor formation, prolonging survival rate of tumor-bearing mice and inhibiting metastasis. We revealed possible underlying mechanisms as reducing the number of myeloid-derived suppressor cells (MDSC) and CD4+Foxp3+ Treg cells, and consequently enhanced IFN-γ production by CTLs. Knockdown of tumor-derived TGF-ß also significantly reduced the conversion of naive CD4+ T cells into Treg cells in vitro. Finally, we found that knockdown of TGF-ß suppressed cell migration, but did not change the proliferation and apoptosis of tumor cells in vitro. In summary, our study provided evidence that tumor-derived TGF-ß is a critical factor for tumor progression and evasion of immune surveillance, and blocking tumor-derived TGF-ß may serve as a potential therapeutic approach for cancer.

[Identification of genes regulating alveolar morphogenesis by supression subtractive hybridization].

Alveoli are the key functional units of the lungs where gas exchange takes place, yet little is known about the mechanisms of their formation. To identify the genes that regulate alveolar morphogenesis, we used suppression subtractive hybridization to isolate genes that differentially expressed during the formation of saccules and the remodeling of saccules into alveoli in the mouse. After differential screening, we got 118 candidate genes from two libraries. These genes associate with many aspects of life including proliferation, differentiation, regulation and so on. For example: Transient receptor protein 4(TRPC4) may associate with formation or remodeling of pulmonary vessels by increasing permeability of endothelia cells. Lectin, (galactose bindingl, Lgalsl) may play a significant role in stimulating smooth muscle growth in developing alveolar wall vessels and the development of pulmonary capillaries. Interesting, a putative neuronalprotein, P311 was found highly expressed in both the first and the secondary septation stages. Taking together SSH is a powerful method to identify differential expressing genes.

Biotransformation of isoeugenol to vanillin by a novel strain of Bacillus fusiformis.

A novel strain of Bacillus fusiformis, producing high amounts of vanillin from isoeugenol, was isolated from soil. Using 60% (v/v) isoeugenol as substrate and solvent and at pH 4.0, 37 degrees C and 180 rpm, vanillin was produced at 32.5 g l(-1) over 72 h. The unused isoeugenol was reusable.

[Embryonic lung cell allografts under the renal capsule--a new model for the study of lung development].

Lung is a critical respiratory organ of living body. The regulation of lung morphogenesis, especially the epithelial development in the late fetal stages, is not completely understood due to lack of efficiency strategies. In this study we showed that the growth of dissociated embryonic lung cells implanted underneath the kidney capsules of syngeneic hosts followed closely lung development in utero. The epithelium developed extensively and appeared to go through pseudoglandular stage, canallicular stage, and saccular stage at a pace similar to normal lung development. At the same time, the capillary-like vascular developed also. The greatest advantage of this model was that the dissociated single cells uptake antisence oligo at a high level, and introduced phenotype after grafting. Embryonic lung cell renal capsule graft is thus an excellent model for the study of lung morphogenesis.