Suppression of lysosome metabolism-meditated GARP/TGF-ß1 complexes specifically depletes regulatory T cells to inhibit breast cancer metastasis.

Regulatory T cells (Tregs) prevent autoimmunity and contribute to cancer progression. They exert contact-dependent inhibition of immune cells through the production of active transforming growth factor-ß1 (TGF-ß1). However, the absence of a specific surface marker makes inhibiting the production of active TGF-ß1 to specifically deplete human Tregs but not other cell types a challenge. TGF-ß1 in an inactive form binds to Tregs membrane protein Glycoprotein A Repetitions Predominant (GARP) and then activates it via an unknown mechanism. Here, we demonstrated that tumour necrosis factor receptor-associated factor 3 interacting protein 3 (TRAF3IP3) in the Treg lysosome is involved in this activation mechanism. Using a novel naphthalenelactam-platinum-based anticancer drug (NPt), we developed a new synergistic effect by suppressing ATP-binding cassette subfamily B member 9 (ABCB9) and TRAF3IP3-mediated divergent lysosomal metabolic programs in tumors and human Tregs to block the production of active GARP/TGF-ß1 for remodeling the tumor microenvironment. Mechanistically, NPt is stored in Treg lysosome to inhibit TRAF3IP3-meditated GARP/TGF-ß1 complex activation to specifically deplete Tregs. In addition, by promoting the expression of ABCB9 in lysosome membrane, NPt inhibits SARA/p-SMAD2/3 through CHRD-induced TGF-ß1 signaling pathway. In addition to expose a previously undefined divergent lysosomal metabolic program-meditated GARP/TGF-ß1 complex blockade by exploring the inherent metabolic plasticity, NPt may serve as a therapeutic tool to boost unrecognized Treg-based immune responses to infection or cancer via a mechanism distinct from traditional platinum drugs and currently available immune-modulatory antibodies.

2-Phenylquinoline-polyamine conjugate (QPC): Interaction with bovine serum albumin (BSA).

A novel 2-phenylquinoline-polyamine conjugate (QPC) was synthesized and characterized, its interaction with bovine serum albumin (BSA) was evaluated using UV-Vis, fluorescence and circular dichroism (CD) spectroscopy. The results showed that QPC caused a whole train of spectral variation, including enhancement of UV-vis absorption and reduction of fluorescence (FL), indicating QPC-BSA complex formed. FL results showed that the type of FL quenching waslarge static quenching, which was also accompanied with a process of dynamic quenching. Binding constants, thermodynamic parameters and docking results showed that the interaction between QPC and BSA was basically a Van der Waals, hydrogen bond and hydrophobic interaction. Synchronous and 3D-FL analysis revealed that QPC resulted in unapparent conformational alteration of BSA. The docking study suggested QPC was situated at the binding sites II of BSA, and 2-phenylquinoline moiety contributed to the hydrophobic interaction. The results of molecular dynamics revealed QPC altered the conformation of BSA, which showed that the inconsistency between experimental data and theoretical calculation results may be due to the instability of the compound.

Genetic diversity patterns of Haemonchus contortus isolated from sheep and goats in Bangladesh.

Haemonchus contortus is the most prevalent parasitic nematode among the Trichostrongylids causing severe health hazards leading to production losses in small ruminants around the world. This study was conducted to explore genetic variation within and among H. contortus populations from seven topographic zones of Bangladesh in small ruminants using second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA and the mitochondrial nicotinamide dehydrogenase subunit 4 (nad4) genes. To do this, a total of 95 adult H. contortus were collected from abomasa of slaughtered sheep and goats from seven different geographic zones of Bangladesh. After the extraction of DNA, ITS-2 of nuclear ribosomal DNA and partial region of the mitochondrial nad4 genes were amplified and sequenced for 95 and 85 worms, respectively. After editing and alignment, sequences were employed for analysis to determine sequence variation, genetic diversity and population genetic structure. Genetic analysis defined 19 distinct ITS-2 genotypes and 77 unique nad4 haplotypes among the H. contortus isolates. The nucleotide diversities were 0.0098 and 0.025 for ITS-2 and nad4 gene, respectively. Phylogenetic analysis (neighbor joining, maximum likelihood and maximum parsimony) of haplotypes indicated the existence of two populations without marked specification of host and locations within H. contortus populations in Bangladesh. By population genetic analysis, 93.67% of genetic variance was partitioned within the population. Very low genetic differentiation but high gene flow was observed among different populations of H. contortus in Bangladesh. This is the first study on genetic variability of H. contortus isolates of small ruminants in Bangladesh. Our study could be the basis for further molecular epidemiological studies, using more discriminative markers and tracing possible changes in the population structure of H. contortus.

A plate method for rapid screening of Ketogulonicigenium vulgare mutants for enhanced 2-keto-l-gulonic acid production

Abstract A new plate method was developed for rapid screening of Ketogulonicigenium vulgare mutants overproducing 2-keto-l-gulonic acid (2-KLG). The screening methodology took the advantage of the acidity caused by 2-KLG, which changes the color of bromothymol blue (pH indicator) from blue to yellow. Using the proposed method, a mutant, K. vulgare 65, was selected from 20,000 colonies produced by a strain subjected to spaceflight mutagenesis. When co-cultured with Bacillus megaterium 2980 in 20-L fermenters, K. vulgare 65 showed a high conversion rate (94.45%) of l-sorbose to 2-KLG. In contrast to the traditional screening method, this one significantly improved the frequency of obtaining positive mutants. The proposed plate screening method is cost-effective and easy to run and is thus useful for the isolation and screening of K. vulgare mutants overproducing 2-KLG.

A plate method for rapid screening of Ketogulonicigenium vulgare mutants for enhanced 2-keto-l-gulonic acid production.

A new plate method was developed for rapid screening of Ketogulonicigenium vulgare mutants overproducing 2-keto-l-gulonic acid (2-KLG). The screening methodology took the advantage of the acidity caused by 2-KLG, which changes the color of bromothymol blue (pH indicator) from blue to yellow. Using the proposed method, a mutant, K. vulgare 65, was selected from 20,000 colonies produced by a strain subjected to spaceflight mutagenesis. When co-cultured with Bacillus megaterium 2980 in 20-L fermenters, K. vulgare 65 showed a high conversion rate (94.45%) of l-sorbose to 2-KLG. In contrast to the traditional screening method, this one significantly improved the frequency of obtaining positive mutants. The proposed plate screening method is cost-effective and easy to run and is thus useful for the isolation and screening of K. vulgare mutants overproducing 2-KLG.

Honokiol protects rat hearts against myocardial ischemia reperfusion injury by reducing oxidative stress and inflammation.

Honokiol, a potent radical scavenger, has been demonstrated to ameliorate cerebral infarction following ischemia/reperfusion (I/R) injury. However, its effects on myocardial I/R injury remain unclear. The present study aimed to examine the effects of honokiol on myocardial I/R injury and to investigate its potential cardioprotective mechanisms. Sprague-Dawley rats were pretreated with honokiol and exposed to a 30-min myocardial ischemia followed by 2-h coronary reperfusion. Myocardial I/R-induced infarct size and biochemical and histological changes were compared. The expression of nuclear factor κB(NF-κB; p65) was assessed by western blotting. Pretreatment with honokiol significantly reduced infarct size, and serum creatine kinase (CK) and lactate dehydrogenase (LDH) release compared with those in the I/R group following a 2-h reperfusion. The malondialdehyde (MDA) level, myeloperoxidase (MPO) activity, concentrations of tumor necrosis factor (TNF)-α and interleukin (IL)-6 and expression level of NF-κB were all reduced by honokiol pretreatment, while honokiol inhibited the decreases in superoxide dismutase (SOD) and catalase (CAT) activities. In addition, less neutrophil infiltration and histopathological damage in the myocardium were observed in the honokiol-pretreated group. These findings indicate that honokiol pretreatment diminished myocardial I/R injury through attenuation of oxidative stress and inflammation.

Correlations between the ages of Alnus host species and the genetic diversity of associated endosymbiotic Frankia strains from nodules.

Nodule samples were collected from four alder species: Alnus nepalensis, A. sibirica, A. tinctoria and A. mandshurica growing in different environments on Gaoligong Mountains, Yunnan Province of Southwest China and on Changbai Mountains, Jilin Province of Northeast China. PCR-RFLP analysis of the IGS between nifD and nifK genes was directly applied to uncultured Frankia strains in the nodules. A total of 21 restriction patterns were obtained. The Frankia population in the nodules of A. nepalensis had the highest genetic diversity among all four Frankia populations; by contrast, the population in the nodules of A. mandshurica had the lowest degree of divergence; the ones in the nodules of A. sibirica and A. tinctoria were intermediate. A dendrogram, which was constructed based on the genetic distance between the restriction patterns, indicated that Frankia strains from A. sibirica and A. tinctoria had a close genetic relationship. Frankia strains from A. nepalensis might be the ancestor of Frankia strains infecting other Alnus species. From these results and the inference of the ages of Alnus host species, it is deduced that there was a co-evolution between Alnus and its microsymbiont Frankia in China.

Isolation and characterization of polycyclic aromatic hydrocarbons-degrading Sphingomonas sp. strain ZL5.

A bacterial strain ZL5, capable of growing on phenanthrene as a sole carbon and energy source but not naphthalene, was isolated by selective enrichment from crude-oil-contaminated soil of Liaohe Oil Field in China. The isolate was identified as a Sphingomonas sp. strain on the basis of 16S ribosomal DNA analysis. Strain ZL5 grown on phenanthrene exhibited catechol 2,3-dioxygenase (C23O) activity but no catechol 1,2-dioxygenase, gentisate 1,2-dioxygenase, protocatechuate 3,4-dioxygenase and protocatechuate 4,5-dioxygenase activities. This suggests that the mode of cleavage of phenanthrene by strain ZL5 could be meta via the intermediate catechol, which is different from the protocatechuate way of other two bacteria, Alcaligenes faecelis AFK2 and Nocardioides sp. strain KP7, also capable of growing on phenanthrene but not naphthalene. A resident plasmid (approximately 60 kb in size), designated as pZL, was detected from strain ZL5. Curing the plasmid with mitomycin C and transferring the plasmid to E. coli revealed that pZL was responsible for polycyclic aromatic hydrocarbons degradation. The C23O gene located on plasmid pZL was cloned and overexpressed in E. coli JM109(DE3). The ring-fission activity of the purified C23O from the recombinant E. coli on dihydroxylated aromatics was in order of catechol > 4-methylcatechol > 3-methylcatechol > 4-chlorocatechol >> 3,4-dihydroxyphenanthrene > 3-chlorocatechol.

[Application of Vc fermentation waste residue on prevention and control of plant diseases in protective ground].

The prevention and control of tomato plant diseases were conducted in protective ground using Vc fermentation waste residue treated by enzymolysis and ultrasonic wave. The results showed that the seedlings planted for 3 weeks on the protective ground soil continuously cropped tomato plant for 9 years and fertilized 75, 150 and 300 kg.hm-2 grew well. Their biomass were increased by 123%, 164% and 182%, and the disease incidence rates were decreased by 59%, 78% and 85%, respectively. Under application of 300 kg.hm-2 Vc fermentation waste residue, the products of tomato grown for 10 weeks on the soil continuously cropped tomato plant for 9, 6 and 2 years were increased by 60%, 43% and 14%, respectively, and the disease incidence rates were all decreased by 50%.

[Effect of natural stress on genetic diversity of Frankia in Alnus nodules].

The genetic diversity of Frankia in the nodules of Alnus nepalensis growing in different habitats of Jizu and Wuliang Mountains in Yunnan Province was studied by rep-PCR technique. The results indicated that the Frankia had an abundant genetic diversity, which was varied with different zones, altitudes and slopes. More genotypes of Alnus-infective Frankia were found at Jizu than at Wuliang Mountain. At the bottom of Jizu Mountain (2,300 m), the Shannon-Weiner index was 0.90, and increased with raising altitude, up to 1.33 at the top (2,700 m). The genetic diversity of Frankia was more abundant along the southern slope than the northern slope. It was concluded that the diversity index was positively related to natural stresses which were one of the important factors producing and maintaining the genetic diversity of Frankia.

[Isolation and characteristics of PAHs-degrading strains].

The predominant indigenous microbial population and the strain ZL5 PAHS-degrading were obtained by selective enrichment culture from Liaohe viscous oil-contaminated field. The sequence analysis of 16s rDNA showed that strain ZL5 belonged to Sphingomonas sp. The microbial population and the strain could degrade phenanthrene well. 95.28% and 69.24% of phenanthrene in the medium were degraded in 120 hrs. However, the degradation of pyrene by the microbial population or by the strain was low. The degradation rate of phenanthrene or pyrene by the microbial population or the strain was greatly increased while glucose was used as an added substrate. The effect was closely related to the amount of glucose. If the concentration were too high, glucose would show inhibition effect on the degradation of PAHs. Therefore, it was necessary to control the amount of glucose while using glucose as an added substrate to increase degradation of phenanthrene or pyrene.

[Semi-nested LP-PCR-SSCP identification of arbuscular endomycorrhizal fungi diversity of Alnus at Changbai Mountain].

The diversity of arbuscular endomycorrhizal fungi colonized on three species of Alnus at Changbai Mountain was identified based on coupling the sensitivity of semi-nested LP-PCR and the specificity afforded by Single Strand Conformation Polymorphism analysis. It is suggested that altitudes and species of Alnus were not the important factors deciding the distribution of AMF in the grade of family, and there was at least one species of Glomaceae, Glomus intraradix, dominated in Alnus at Changbai Mountain.

[Diversity of the microorganisms degrading aromatic hydrocarbons].

Aromatic hydrocarbons are biological xenobiotics. Indigenous microorganism groups turn from being unfamiliar with into active response to environmental changes by their adaptability to environment. There are changes in their genetic background, and then, the biodiversity turns into being. This paper reviewed the microorganism groups resources, biocharacters, genetic background, evolvement and their adaptability to environment; described the genetic information, expression and regulatory for some species in detail; and pointed out that the degradation of aromatic hydrocarbons depended mainly on new microorganisms formed by genetic engineering and on their highly efficient metabolic regulatory.

[Microbiological eco-regulation in Vc two-step fermentation].

The growth and interaction of Gluconobacter axydans and Bacillus megaterium in Vc two step fermentation have been studied. The results showed that there is a kind of coordinate symbiosis between G. axydans and B. megaterium during the fermentation, which could promoted 2-Keto-L-gulonic acid (2KGA) synthesis. The efficiency on the synbiosis would vary as the conditions of fermentation were different and were improved by the environmental factors.