The influence of a TNF gene polymorphism on the severity of rheumatoid arthritis in the Brazilian Amazon

PurposeThe aim of this study was to investigate the influence of the TNF -308 G/A polymorphism in the promoter region of the tumor necrosis factor-α gene on the susceptibility and severity of rheumatoid arthritis (RA) in individuals from the Brazilian Amazon.MethodsA total of 323 individuals—192 healthy controls without arthritis and 131 individuals suffering from arthritis—were genotyped for this polymorphism using a methodology based on PCR-RFLP.ResultsThe frequency of the A allele (TNF2) in rheumatoid arthritis sufferers was not significantly higher than in the controls (p = 0.926; OR = 0.97; confidence interval 0.54–1.76). However, using a logistic regression model, when the patients were stratified according to whether the manifestations were preponderantly articular or systemic, there was a strong association between the TNF2 allele and systemic arthritis (p = 0.001; OR = 5.89; confidence interval = 1.98–17.5) as well as the use of anti-TNF immunotherapy (p = 0.023; OR = 1.10; confidence interval = 1.00–1.14). The main factors that were found to influence the risk of extra-articular disease were age greater than or equal to 60 years (p = 0.008; OR = 4.06; confidence interval = 1.45–11.38), disease duration greater than 10 years (p = 0.031; OR = 3.10; confidence interval = 1.11–8.63) and positive rheumatoid factor (p = 0.035; OR = 2.07; confidence interval = 1.05–4.09).ConclusionsThese results suggest that the TNF2 allele is associated with the more serious forms of the disease in individuals from the Brazilian Amazon but not with a risk for developing RA.     

Synthesis and evaluation of benzo[b]thiophene derivatives as inhibitors of alkaline phosphatases

Presence of basic calcium phosphate in knee joints of osteoarthritis patients could be prevented by inhibiting tissue non-specific alkaline phosphatase (TNAP) activity. Levamisole or the L stereoisomer of tetramisole (a known TNAP inhibitor) has been used as a treatment for curing rheumatoid arthritis but its therapeutical use is limited due to side effects. We report the synthesis and the TNAP inhibition property of benzo[b]thiophene derivatives, among which benzothiopheno-tetramisole and benzothiopheno-2,3-dehydrotetramisole, which could be involved in a drug therapy for osteoarthritis. Two water soluble racemic benzothiopheno-tetramisole and -2,3-dehydrotetramisole with apparent inhibition constants K_i = 85 ± 6 μM and 135 ± 3 μM (n = 3) comparable to that of enantiomeric levamisole 93 ± 4 μM were found. Several novel derivatives showed more pronounced inhibition properties towards intestinal alkaline phosphatase than TNAP.     

Molecular distinction of C × R hybrid (Coffea congensis × Coffea canephora) from morphologically resembling male parent using rbcL and matK gene sequences

Interspecific C × R hybrid (Coffea congensis × Coffea canephora) in India is cultivated as mixed population with male parent C. canephora as this species is an efficient pollen donor for enhanced yield. But distinction of C × R hybrid from C. canephora in old plantation is difficult due to varying plant sizes of C × R hybrid and often resembles with C. canephora. C × R hybrid cultivated under different agroclimatic conditions show distinct vegetative growth pattern with varying yields. Thus development of DNA marker for identification of C × R hybrid is important for clonal propagation and seed preparation from selective individuals. In this study, two DNA bar coding loci of chloroplast genome (rbcL and matK) of parents, F1 hybrid and its back cross progeny were partially sequenced to identify SNPs as DNA marker for distinction of C × R hybrid from C. canephora. Seven SNPs in the matK gene sequence and three nucleotides in the rbcL gene sequence were identified as DNA markers for the genetic identity of C. congensis. These SNPs were found in F1 and advanced progenies of C × R hybrid due to maternal inheritance. Large number of samples of C × R hybrids with varying morphological features revealed no polymorphism among C × R hybrid and C. congensis. Thus, the SNPs in C. congensis can be used as DNA markers for precise identification of C × R hybrid for production of clones besides tagging the chloroplast inheritance in advanced progenies.     

Crucial factors affecting the nitrile hydratase production of Mesorhizobium sp. F28

Mesorhizobium sp. F28 contains cobalt-NHase, which effectively converts acrylonitrile into acrylamide. When urea was added to the culture medium, the NHase activity was 62.3 U ml^?1 (R2A–R2A/urea) after 22.5 h of cultivation, which was similar to that in the medium without addition (R2A–R2A, 70.0 U ml^?1). The relative activity of the purified NHase was 100%, 92%, 94%, and 92% in the medium containing, respectively, 0 mM, 2 mM, 5 mM, and 10 mM of urea. Urea had no significant effect on the purified NHase activity of Mesorhizobium sp. F28. This research did not observe the NHase production by Mesorhizobium sp. F28 when acrylonitrile was supplemented in the culture medium except that cobalt ions existed. The highest enzyme activity was 328.5 U ml^?1 as cobalt ions were added in the pre-culture and culture medium after 22.5 h of cultivation (R2A/Co-R2A/Co); compared to media without cobalt ions (R2A–R2A, 22.5 h, 70.5 U ml^?1) this is an almost five-fold enhancement. It can be concluded that culture media containing cobalt ions was beneficial for the formation of active NHase of Mesorhizobium sp. F28.     

Impact of rock materials and biofertilizations on P and K availability for maize (Zea Maize) under calcareous soil conditions

The present work evaluated the synergistic effects of soil fertilization with rock P and K materials and co-inoculation with P and K-dissolving bacteria [PDB (Bacillus megaterium var. phosphaticum) and KDB (Bacillus mucilaginosus and B. subtilis)] on the improvement of P and K uptake, P and K availability and growth of maize plant grown under limited P and K soil conditions (calcareous soil). The experiment was establishment with eight treatments: without rock P and K materials or bacteria inoculation (control), rock P (RP), rock K (RK), RP + PDB, RK + KDB and R(P + K)+(P + K)DB. Under the same conditions of this study, co-inoculation of PDB and KDB in conjunction with direct application of rock P and K materials (R(P + K)) into the soil increased P and K availability and uptake, and the plant growth (shoot and root growth) of maize plants grown on P and K limited soils.     

Decreased serum level of IL-7 in patients with active Graves’ disease

BackgroundGraves’ disease (GD) is a common autoimmune disease which is one of the major causes of hyperthyroidism. Interleukin 7 (IL-7) has been recently reported to play an important role in various autoimmune diseases, but its role in the pathogenesis of GD has not been assessed. The aim of this study was to evaluate the levels of IL-7 and the soluble form of its receptor (sIL-7R) in the serum of GD patients, and to identify their association with disease activity.MethodsA total of 37 GD patients were enrolled into the experimental group and 16 individuals into the control group. All patients were further classified into three subgroups: a GD-active group (hyperthyroidism and TRAb (thyroid stimulating hormone receptor antibody) >7.5 U/L) (N = 15), a GD-inactive group (euthyreosis and TRAb < 1 U/L) (N = 8), and other GD patients (euthyreosis and TRAb > 1 U/L) (N = 14). Concentrations of IL-7 and sIL-7R were assayed with ELISA. Additionally, the relationship between IL-7 and sIL-7R serum concentrations with disease activity (free triiodothyronine [FT3], free thyroxine [FT4], thyroid stimulating hormone [TSH] and TRAb) was also analyzed.ResultsThe serum concentrations of IL-7 in GD-active patients were significantly lower than those of the control group as well as the GD-inactive and GD-other groups. The serum level of IL-7 in GD patients negatively correlated with FT4 and TRAb concentrations. Moreover, no significant difference was observed in the serum level of sIL-7R in GD patients compared to the control group.ConclusionsThese observations suggest that IL-7 may play a role in the pathogenesis of GD and may be associated with its clinical activity. To this end, the serum level of IL-7 could be an additional diagnostic biomarker predictive of the disease and could be particularly valuable for TRAb-negative GD patients.     

Bioprocess strategies for mass multiplication of and metabolite synthesis by plant growth promoting pseudomonads for agronomical applications

The exponential substrate feeding (open-loop) and automated feedback substrate feeding (closed loop) strategies were developed to obtain high cell densities of fluorescent pseudomonad strains R62 and R81 and enhanced production of antifungal compound 2,4-diacetylphloroglucinol (DAPG) from glycerol as a sole carbon source. The exponential feeding strategy resulted in increased glycerol accumulation during the fed-batch cultivation when the predetermined specific growth rate (μ) was set at 0.10 or 0.20 h^?1 (<μ_m = 0.29 h^?1). Automated feeding strategies using dissolved oxygen (DO) or pH as feedback signals resulted in minimal to zero accumulation of glycerol for both the strains. In case of DO-based feeding strategy, biomass productivity of 0.24 g/(L h) and 0.29 g/(L h) was obtained for R62 and R81, respectively. Using pH-based feeding strategy, biomass productivity could be increased to a maximum of 0.51 and 0.54 g/(L h), for the strains R62 and R81, respectively, whereas the DAPG concentration was enhanced to 298 mg/L for R62 and 342 mg/L for R81 strains. These yields of DAPG are thus far the highest reported from GRAS organisms.     

Genome size variation and polyploidy in the resurrection plant genus Ramonda: Cytogeography of living fossils

The genus Ramonda includes three preglacial paleoendemic species surviving as the rare resurrection angiosperms of the Northern hemisphere in refugia habitats in the Balkan (Ramonda nathaliae and Ramonda serbica) and Iberian Peninsulas (Ramonda myconi). This study focuses on: assessing genome size and base composition, determining chromosome number and ploidy level in several populations, evaluating inter- and intra-specific variations in DNA content and chromosome number as well as looking for the possible hybridization in the sympatric zones of Balkan species. R. nathaliae and R. myconi are diploid species (2n = 2x = 48) while R. serbica is hexaploid (2n = 6x = 144). The mean 2C DNA values ranged from 2.30 pg for R. nathaliae to 2.59 pg for R. myconi compared to 7.91 pg for R. serbica. The base composition for R. nathaliae was 42.1% GC, for R. myconi 39.9% and for R. serbica 41.2%. In one population of R. serbica the DNA content ranged from 2C = 7.65 to 11.82 pg, revealing different ploidy levels among its individuals. In sympatric populations genome size was intermediary (～5 pg) between the diploid and hexaploid classes which indicates the hybridization ability between R. serbica and R. nathaliae. It appears that polyploidization is the major evolutionary mechanism in the genus Ramonda.     

The site-directed mutation I(L177)H in Rhodobacter sphaeroides reaction center affects coordination of PA and BB bacteriochlorophylls

To explore the influence of the I(L177)H single mutation on the properties of the nearest bacteriochlorophylls (BChls), three reaction centers (RCs) bearing double mutations were constructed in the photosynthetic purple bacterium Rhodobacter sphaeroides, and their properties and pigment content were compared with those of the correspondent single mutant RCs. Each pair of the mutations comprised the amino acid substitution I(L177)H and another mutation altering histidine ligand of BChl P_A or BChl B_B. Contrary to expectations, the double mutation I(L177)H + H(L173)L does not bring about a heterodimer RC but causes a 46 nm blue shift of the long-wavelength P absorbance band. The histidine L177 or a water molecule were suggested as putative ligands for P_A in the RC I(L177)H + H(L173)L although this would imply a reorientation of the His backbone and additional rearrangements in the primary donor environment or even a repositioning of the BChl dimer. The crystal structure of the mutant I(L177)H reaction center determined to a resolution of 2.9 Å shows changes at the interface region between the BChl P_A and the monomeric BChl B_B. Spectral and pigment analysis provided evidence for β-coordination of the BChl B_B in the double mutant RC I(L177)H + H(M182)L and for its hexacoordination in the mutant reaction center I(L177)H. Computer modeling suggests involvement of two water molecules in the β-coordination of the BChl B_B. Possible structural consequences of the L177 mutation affecting the coordination of the two BChls P_A and B_B are discussed. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.     

Serum evaluation of soluble interferon-α/β receptor and high-sensitivity C-reactive protein for diagnosis of the patients with gastrointestinal and hepatobiliary-pancreatic cancer

Serum soluble interferon-α/β receptor (sIFN-α/βR) and high-sensitivity C-reactive protein (hs-CRP) levels were evaluated in the patients with gastrointestinal and hepatobiliary-pancreatic cancer. We compared the sensitivity and specificity of serum sIFN-α/βR with that of serum hs-CRP and evaluated the two diagnostic parameters in combination. Serum sIFN-α/βR levels were measured in 92 patients and 25 healthy individuals by enzyme-linked immunosorbent assay. The diagnoses were 37 cases of hepatocellular carcinoma, 17 cases of pancreatic cancer, 15 cases of colon cancer, 13 cases of biliary tract cancer, and 10 cases of gastric cancer. Serum levels of sIFN-α/βR and hs-CRP were significantly higher in the patients than in healthy individuals (p < 0.05). The optimal cut-off values of sIFN-α/βR and hs-CRP were 3600 pg/ml and 0.5 μg/ml, respectively. The sensitivity and specificity for these thresholds were 94.6% and 88.0%, whereas positive predictive and negative predictive values were 96.7% and 81.5%. These results suggest that a combination of serum sIFN-α/βR and hs-CRP thresholds may be more reliable diagnostic parameter for gastrointestinal and hepatobiliary-pancreatic cancer.     

Cardioprotective effects of lipoic acid,quercetin and resveratrol on oxidative stress related to thyroid hormone alterations in long-term obesity

This study investigated possible mechanisms for cardioprotective effects of lipoic acid (LA), quercetin (Q) and resveratrol (R) on oxidative stress related to thyroid hormone alterations in long-term obesity. Female C57BL/6 mice were fed on high-fat diet (HFD), HFD + LA, HFD + R, HFD + Q and normal diet for 26 weeks. Body weight, blood pressure, thyroid hormones, oxidative stress markers, angiotensin converting enzyme (ACE), nitric oxide synthase (NOS) and ion pump activities were measured, and expression of cardiac genes was analyzed by real-time polymerase chain reaction. HFD induced marked increase (P < .05) in body weight, blood pressure and oxidative stress, while plasma triidothyronine levels reduced. ACE activity increased (P < .05) in HFD mice (0.69 ± 0.225 U/mg protein) compared with controls (0.28 ± 0.114 U/mg protein), HFD + LA (0.231 ± 0.02 U/mg protein) and HFD + Q (0.182 ± 0.096 U/mg protein) at 26 weeks. Moreover, Na⁺/K⁺-ATPase and Ca^2 +-ATPase activities increased in HFD mice whereas NOS reduced. A 1.5-fold increase in TRα1 and reduction in expression of the deiodinase iodothyronine DIO1, threonine protein kinase and NOS3 as well as up-regulation of AT1α, ACE, ATP1B1, GSK3β and Cja1 genes also occurred in HFD mice. Conversely, LA, Q and R inhibited weight gain; reduced TRα1 expression as well as increased DIO1; reduced ACE activity and AT1α, ATP1B1 and Cja1 gene expression as well as inhibited GSK3β; increased total antioxidant capacity, GSH and catalase activity; and reduced blood pressure. In conclusion, LA, resveratrol and quercetin supplementation reduces obesity thereby restoring plasma thyroid hormone levels and attenuating oxidative stress in the heart and thus may have therapeutic potential in heart diseases.     

Improved PCR performance and fidelity of double mutant Neq A523R/N540R DNA polymerase

We previously reported that Neq A523R DNA polymerase is more efficient in PCR than wild-type Neq DNA polymerase, and amplifies products more rapidly. Neq A523R DNA polymerase also amplifies templates more rapidly than Pfu DNA polymerase, but has a lower fidelity than Pfu DNA polymerase. To improve product yield and the fidelity of amplification simultaneously, we constructed and characterized the double mutant Neq A523R/N540R. The yield of PCR products was greater for Neq A523R/N540R DNA polymerase than wild-type and other mutant DNA polymerases, and the Neq double mutant catalyzed amplification of a 12-kb PCR product from a lambda template with an extension time of 3 min. The PCR error rate of Neq A523R/N540R DNA polymerase (6.3 × 10⁻⁵) was roughly similar to that of Pfu DNA polymerase (4.8 × 10⁻⁵), but much lower than those of wild-type Neq DNA polymerase (57.2 × 10⁻⁵), Neq A523R DNA polymerase (13.1 × 10⁻⁵), and Neq N540R DNA polymerase (37.7 × 10⁻⁵). These results indicated that A523R and N540R mutations of Neq DNA polymerase had synergistic effects on its fidelity.     

Enhanced acylation activity of esterase BioH from Escherichia coli by directed evolution towards improved hydrolysis activity

Esterase BioH is a critical enzyme for Biotin synthesis in Escherichia coli, which has been previously found to be active in the acylation of secondary alcohols and amines. Directed evolution towards improved acylation activity requires a high-throughput screening method. The aim of this study is to explore whether the acylation activity of BioH can be improved by directed evolution of its hydrolysis activity. A colorimetric method based on p-nitrophenyl butyrate hydrolysis was adopted in the high-throughput determination of hydrolysis activity. The wild-type BioH showed a hydrolysis activity of 18 U/mg, and the specific activities for α-phenylethanol and α-phenylethylamine acylation were 12.8 U/mg and 3.5 U/mg, respectively. After two rounds of directed evolution, seven mutants with improved hydrolysis activity were obtained, among which, K213E, Q70L/M170T and M197L/K213E also showed improvement in acylation activity. To further improve the acylation activity, site mutations were generated in different combinations at the four hot spots Q70L, M170T, M197L and K213E. Among the resulting mutants, Q70L/M197L/K213E showed the highest activity in α-phenylethylamine acylation with a 120% improvement, while Q70L/K213E had the highest α-phenylethanol acylation activity, which was improved by 70%. The results demonstrated that directed evolution of the hydrolysis activity might be a possible approach to improve the acylation activity of the esterase BioH.     

Promoter polymorphism (−590, T/C) of interleukin 4 (IL4) gene is associated with rheumatoid arthritis: An updated meta-analysis

Rheumatoid arthritis (RA) is a chronic disease. It causes chronic inflammation of the joint. Recent studies suggested that interleukin 4 (IL4) contributes to susceptibility and severity of rheumatoid arthritis (RA). Especially, it was reported that promoter polymorphism (−590, T/C) of IL4 gene has been associated with susceptibility of RA. The aim of present study was to investigate whether the promoter polymorphism (−590, T/C) of IL4 gene is associated with the susceptibility of RA using meta-analysis. And in order to perform meta-analysis, comprehensive meta analysis program was used. Genetic models (co-dominant, dominant, recessive, and allele) were used to determine odds ratios (ORs), 95% confidence intervals (CIs), and P values. Nine case-control studies with case and control design were included in this meta-analysis. Overall, meta-analysis revealed a strong association with susceptibility of RA [OR = 1.303, 95% CI = 1.093–1.554, P = 0.003 in allele model (C vs. T); OR = 1.247, 95% CI = 1.054–1.474, P = 0.010 in dominant model (CC vs. CT + TT); OR = 2.148, 95% CI = 1.263–3.651, P = 0.005 in recessive model (CC + CT vs. TT)]. Our data demonstrated that promoter polymorphism (−590, T/C) of IL4 gene may be contributed to susceptibility of RA. However, more studies with a larger sample size are needed to provide more precise evidence.     

Differing patterns of peripheral blood leukocyte telomere length in rheumatologic diseases

Telomeres progressively shorten with repeated somatic tissue cell division, their length being an indicator of cellular ageing. Telomeric dysfunction may be implicated in a variety of diseases. We measured mean telomere length in peripheral blood leukocytes (PBL) from patients with various rheumatologic diseases. Mean PBL telomere length was measured using real-time quantitative polymerase chain reaction (Q-PCR) assay in a control population (n = 130; age range: 3–94 years) and in subjects diagnosed with rheumatoid arthritis (RA; n = 86; age range: 31–82 years), psoriatic arthritis (PA; n = 56; age range: 26–79 years) and ankylosing spondylitis (AS; n = 59; age range: 21–75 years). These diseases are associated with chronic systemic inflammatory activity. Telomere length was also quantified in subjects with osteoarthritis (OA; n = 34; age range: 43–82 years) and osteoporosis (OP; n = 35; age range: 59–95 years), diseases without a chronic systemic inflammatory component. Telomere length in OA showed no differences from age-matched controls (p = 0.234), but was significantly shorter in OP (p = 0.001). Telomere length was significantly longer than controls in RA (p = 0.015), PA (p < 0.001) and AS (p < 0.001). Different patterns in telomere length from PBL are evidenced in rheumatologic pathologies, possibly dependent on the presence or absence of chronic systemic inflammation.     

Development of Halomonas TD01 as a host for open production of chemicals

Genetic engineering of Halomonas spp. was seldom reported due to the difficulty of genetic manipulation and lack of molecular biology tools. Halomonas TD01 can grow in a continuous and unsterile process without other microbial contaminations. It can be therefore exploited for economic production of chemicals. Here, Halomonas TD01 was metabolically engineered using the gene knockout procedure based on markerless gene replacement stimulated by double-strand breaks in the chromosome. When gene encoding 2-methylcitrate synthase in Halomonas TD01 was deleted, the conversion efficiency of propionic acid to 3-hydroxyvalerate (3HV) monomer fraction in random PHBV copolymers of 3-hydroxybutyrate (3HB) and 3HV was increased from around 10% to almost 100%, as a result, cells were grown to accumulate 70% PHBV in dry weight (CDW) consisting of 12 mol% 3HV from 0.5 g/L propionic acid in glucose mineral medium. Furthermore, successful deletions on three PHA depolymerases eliminate the possible influence of PHA depolymerases on PHA degradation in the complicated industrial fermentation process even though significant enhanced PHA content was not observed. In two 500 L pilot-scale fermentor studies lasting 70 h, the above engineered Halomonas TD01 grew to 112 g/L CDW containing 70 wt% P3HB, and to 80 g/L CDW with 70 wt% P(3HB-co-8 mol% 3HV) in the presence of propionic acid. The cells grown in shake flasks even accumulated close to 92% PHB in CDW with a significant increase of glucose to PHB conversion efficiency from around 30% to 42% after 48 h cultivation when pyridine nucleotide transhydrogenase was overexpressed. Halomonas TD01 was also engineered for producing a PHA regulatory protein PhaR which is a robust biosurfactant.     

Stereoselective biocatalytic hydride transfer to substituted acetophenones by the yeast Metschnikowia koreensis

Freely suspended and variously entrapped viable cells of the yeast Metschnikowia koreensis were examined for the asymmetric reduction of prochiral acetophenone. A ketone substrate at 25 mM can be converted (92%) to the corresponding alcohol within 3 h using freely suspended cells [46 mg/mL dry cell weight (DCW)] at pH 9 (Tris buffer, 50 mM), 25 °C, in an agitated reactor (200 rpm). The reaction displayed an excellent stereoselectivity of >99%. Supplementation of the reaction mixture with glucose (20 g/L) greatly enhanced the rate of the bioreduction reaction likely because of improved cofactor recycling in the cells. The cells could successfully reduce various acetophenones substituted with electron withdrawing groups on the phenyl ring, particularly at the para-position compared to ortho- or meta-substituted acetophenones. The ketone reductase of M. koreensis showed Prelog-selectivity as the reaction exclusively yielded (S)-alcohols. The thermostability and the substrate tolerance of the yeast were improved by immobilization in calcium alginate beads. Immobilization reduced the effectiveness factor only slightly.     

Breast cancer risk and common single nucleotide polymorphisms in homologous recombination DNA repair pathway genes XRCC2, XRCC3, NBS1 and RAD51

The possible role for DNA repair deficiencies in cancer development, namely in breast cancer has been the subject of increasing interest since it has been reported that breast cancer patients might be deficient in the repair of DNA damage. Exposure to ionizing radiation has been pointed out as a risk factor for breast cancer, and the type of DNA lesions induced by this carcinogen can be repaired by homologous recombination DNA repair (HRR) pathway. To evaluate the potential modifying role of some single nucleotide polymorphisms (SNP) in HRR involved genes on the individual susceptibility to breast cancer we carried out a hospital based case–control study in a Caucasian Portuguese population (289 histological confirmed breast cancer patients and 548 control individuals). We genotyped 4 SNPs in 4 different HRR pathway genes, XRCC2 (Ex3 + 442G > A, R188H, rs3218536), XRCC3 (Ex8-5C > T, T241M, rs861539), NBS1 (Ex5-32C > G, E185Q, rs1805794) and RAD51 5′UTR (Ex1-59G > T, rs1801321), tagging 41 SNPs in these genes. The frequency of the different polymorphisms in the Portuguese control population is similar to the ones reported for other Caucasian populations, and the deviation of the Hardy–Weinberg equilibrium was only observed for the XRCC2 (Ex3 + 442G > A, R188H, rs3218536) polymorphism in the control population. The results obtained, after logistic regression analysis, did not reveal a major role of these polymorphisms on breast cancer susceptibility. However, when the population was stratified according to breast feeding (women that breast fed and women that never breast fed) it is observed, in women that never breast fed, that the heterozygous individuals for the XRCC2 (Ex3 + 442G > A, R188H, rs3218536) polymorphism have a decreased risk for breast cancer [adjusted OR = 0.45; 95% CI = 0.22–0.92] (P = 0.03). Additionally, after stratification according to menopausal status, our results suggest that post-menopausal women carrying at least one variant allele for the XRCC3 (Ex8-5C > T, T241M, rs861539) polymorphism have a lower risk for breast cancer [adjusted OR = 0.67; 95% CI, 0.47–0.94] (P = 0.03). Most of the studies suggest that breastfeeding may be responsible for 2/3 of the estimate reduction of breast cancer. The longer the duration of breastfeeding the lower the potential risk associated with breast cancer. Therefore, in our study the potential protective role of the variant allele of XRCC2 (Ex3 + 442G > A, R188H, rs3218536), in never breast fed women, might be related with a more efficient DNA repair activity.     

Metabolic engineering of Ralstonia eutropha for the biosynthesis of 2-hydroxyacid-containing polyhydroxyalkanoates

Polyhydroxyalkanoates (PHAs) are bio-based and biodegradable polyesters synthesized by numerous microorganisms. PHAs containing 2-hydroxyacids as monomer units have attracted much attention, but their production has not been efficient. Here, we metabolically engineered Ralstonia eutropha strains for the in vivo synthesis of PHAs containing 2-hydroxyacids as monomers. This was accomplished by replacing the R. eutropha phaC gene in the chromosome with either the R. eutropha phaC S506G A510K gene, which contains two point mutations, or the Pseudomonas sp. MBEL 6–19 phaC1437 gene. In addition, the R. eutropha phaAB genes in the chromosome were replaced with the Clostridium propionicum pct540 gene. All of the engineered R. eutropha strains produced PHAs containing 2-hydroxyacid monomers, including lactate and 2-hydroxybutyrate (2HB), along with 3-hydroxybutyrate (3HB) and/or 3-hydroxyvalerate (3HV), when they were cultured in nitrogen-free medium containing 5 g/L lactate or 4 g/L 2HB and 20 g/L glucose as carbon sources. Expression of the Escherichia coli ldhA gene in engineered R. eutropha strains allowed production of poly(3-hydroxybutyrate-co-lactate) [P(3HB-co-LA)] from glucose as the sole carbon source. This is the first report on the production of 2-hydroxyacid-containing PHAs by metabolically engineered R. eutropha.